WHIP2015 book - Marine Biological Laboratory
WOODS HOLE IMMUNOPARASITOLOGY CONFERENCE 2015 Sunday APRIL 19 – Wednesday APRIL 22 Meeting Co-‐organizers: JUDITH ALLEN School of Biological Sciences University of Edinburgh, UK NICOLA HARRIS Ecole Polytechnique Fédérale de Lausanne Lausanne, Switzerland ANDREA GRAHAM Princeton University Princeton, New Jersey, USA Organizational support provided by: Marine Biological Laboratory, Woods Hole, MA FINANCIAL SUPPORT PROVIDED BY: Global Health Institute, EPFL SUNDAY APRIL 19, 2015 (following Immune selection workshop*) 16:30 Opening Remarks – Judi Allen, Nicola Harris, Andrea Graham 16:40-‐-‐-‐17:40 Ecology of the Immune System The immunologist’s view – Chris Hunter The ecologist’s view – Nicole Mideo CO-‐INFECTION – CHAIRs Andrea Graham & Constance Finney 17:40 Matteo Rossi -‐-‐-‐ Type I interferons induced by viral co-‐infection worsen murine leishmaniasis caused by L. guyanensis 17:55 Melanie Clerc – Co-‐infection and immunity affect parasite burden in the wild; insights from a wild wood mouse system 18:10 Dinner Swope 20:00 Social Hour MONDAY APRIL 20, 2015 LEISHMANIA -‐-‐-‐ CHAIRs Phil Scott & Michael Grigg 8:30 Susmita Ghosh -‐-‐-‐ Leishmania and/or Host: Who drives the Immune Response during Indian Leishmaniasis? 8:45 Benjamin Hurrell -‐-‐-‐ Rapid sequestration of Leishmania mexicana by neutrophils results in the development of chronic lesion 9:00 Clara Gimblet -‐-‐-‐ Leishmania major infection induces transmissible alterations in the skin microbiome 9:15 Stephen Christensen, RNA sequencing to reveal host and parasite transcriptional responses during human cutaneous leishmaniasis 9:30 Patrick Kelly -‐-‐-‐ Alterations of the gut microbiota of the vector Lutzomyia longipalpis over time due to alterations in diet and Leishmania infection 9:45 POSTER PITCHES 10:30 – 11:00 COFFEE BREAK 11:00 -‐-‐-‐ 12:00 KEYNOTE – GWEN RANDOLPH “Tissue resident macrophage diversity: What is the point?” 12:15 – 13:30 LUNCH SWOPE MYELOID CELLS – CHAIRs David Mosser & P’ng Loke 13:30 Michael Gonzalez -‐-‐-‐ Conversion of monocyte derived M2 macrophages into a tissue resident macrophage phenotype 13:45 Stanley Huang -‐-‐-‐ The mTORC2-‐IRF4 Axis Regulates Alternatively Activated Macrophages 14:00 Keke Fairfax -‐-‐-‐ Schistosoma Mansoni infection induces anti-‐atherogenic transcriptional changes in hepatic macrophages 14:15 Mark Viney -‐-‐-‐ The Immune Lives of Wild Mice 14:30 Stephen Redpath -‐-‐-‐ Type 2 cytokine signalling alters dendritic cell phenotype and function 14:45 Lanny Gov -‐-‐-‐ Toxoplasma gondii preferentially invades human monocytes and selectively activates host signaling pathways 15:00 – 15:30 COFFEE BREAK MALARIA – CHAIRs Mary Stevenson & Tracey Lamb 15:45 Seth Amanfo -‐-‐-‐ Multiple Plasmodium Species Co-‐-‐-‐infections In African Populations 16:00 Chester Joyner, Dysfunctional neutrophils are a feature of Plasmodium cynmologi infection in rhesus monkeys 16:15 -‐-‐-‐ 17:00 POSTER PITCHES 18:00 Dinner Swope 20:00 Poster Session & Social Hour TUESDAY APRIL 21, 2015 HELMINTHS – CHAIRs Richard Grencis & Nicola Harris 8:30 Romain Garnier -‐-‐-‐ Integrating immune mechanisms in model of sheep gastro-‐-‐-‐intestinal nematodes 8:45 Lalit Kumar Dubey – Mesenteric lymph node organization during helminth infection 9:00 Laura Campbell -‐-‐-‐ Mucus and immunity to Trichinella spiralis: A systemic affair. 9:15 Christian Owusu, Temporal transcriptional profiling of livers from Schistosoma mansoni-‐infected mice 9:30 Sarah Budischak, Experimental and longitudinal evidence for the fitness costs of helminth infection in African buffalo HOST DEFENSE – Paul Kaye & Tajie Harris 9:45 Edmund Legrand -‐-‐-‐ Non-‐-‐-‐specific stressors: under-‐-‐-‐appreciated innate defenses against pathogens 10:00 Dionne Robinson -‐-‐-‐ Biological Sex is a Major Determinant of the Immune Response to Toxoplasma gondii 10:15 Paul Capewell -‐-‐-‐ Genetic Determinants of Trypanotolerance in Humans 10:30 – 11:00 COFFE BREAK 11:00 – KEYNOTE JOHN MANSFIELD – “New Paradigms of Host Resistance to African Trypanosomiasis” 12:15 – 13:30 LUNCH SWOPE LYMPHOCYTES – CHAIRs Chris Hunter & Georgia Perona Wright 13:30 Jennifer Cnops -‐-‐-‐ NK, NKT and CD8-‐derived IFNγ drives myeloid cell activation and erythrophagocytosis, resulting in Trypanosomosis-‐associated acute anemia 13:45 Sasha Silva -‐-‐-‐ Leishmania donovani activates B cells and induces cytokine expression by triggering endosomal TLRs 14:00 Gretchen Pritchard – a role for T-‐bet in co-‐ordinating T cell activation 14:15 Lucy Jones -‐-‐-‐ Fat Associated Lymphoid Clusters are IL-‐33R Dependent Sites of B Cell Proliferation and Local IgM Production 14:30 Anna Sanecka -‐-‐-‐ The formation and maintenance of brain resident memory CD8 T cells is dependent on the affinity of TCR-‐MHC interaction 14:45 Kyle Burrows -‐-‐-‐ The transcriptional repressor Hic1 regulates intestinal immunity 15:00 – 15:30 COFFEE BREAK IMMUNOPATHOLOGY – CHAIRs -‐-‐-‐ Nicola Harris & Herbert De’Broski 15:30 Danielle Worth – A role for dectin in the CNS during chronic Toxoplasma gondii infection 15:45 Barbara Capuccini – Role of microglia during cerebral malaria 16:00 Elise Burger -‐-‐-‐ The Paneth-‐-‐-‐cell specific role of autophagy in Toxoplasma gondii-‐-‐-‐induced intestinal inflammation 16:15 Li Yin Hung -‐-‐-‐ Trefoil factor 2 mediates macrophage-‐-‐-‐dependent lung repair following hookworm infection 16:30 Oliver Preham – Immunopathogenesis of visceral leishmaniasis 16:45 Caroline Junqueira-‐-‐-‐ A highly attenuated clone of Trypanosoma cruzi lacks the SAPA domain of trans-‐-‐-‐sialidase protein 18:00 Dinner Swope 20:00 Poster Session & Social Hour (Bar & DJ) WEDNESDAY APRIL 22, 2015 0700-‐-‐-‐0830 BREAKFAST – Swope [0830-‐-‐-‐1130 *Immune selection workshop continues from Sunday] DEPARTURE: Safe travels everyone!! Thank you to: NIH NSF Global Health Institute, EPFL University of Pennsylvannia, Dept of Pathobiology Parasite Immunology * Immune selection workshop is scheduled for 14-‐-‐-‐16.30 Sunday and 08.30-‐-‐-‐11.30 Wednesday Participation of WHIP conference-‐-‐-‐goers very welcome! Please contact Andrea for details and schedule: algraham@princeton.edu TALK ABSTRACT LIST Type I interferons induced by viral co-‐infection worsen murine leishmaniasis caused by L. guyanensis Rossi, Matteo (Department of Biochemistry, University of Lausanne, Lausanne, CHE); Castiglioni, Patrik ; Hartley, Mary-‐Anne; Eren, Remzi Onur; Prével, Florence; Utzschneider, Daniel T. (Swiss Vaccine Research Institute, Epalinges, Switzerland); Zehn, Dietmar (Swiss Vaccine Research Institute, Epalinges, Switzerland); Beverley, Stephen M.; Ronet, Catherine; Fasel, Nicolas Co-‐infection and immunity affect parasite burden in the wild: Insights from a wild wood mouse system Clerc, Melanie (University of Edinburgh, UK), Devevey, Godefroy; Pedersen, Amy B Leishmania and/or Host: Who drives the Immune Response during Indian Leishmaniasis? Ghosh, Susmita (Institute of Post Graduate Medical Education and Research, Kolkata, IND); Dey, Badal; Nath, Sankha; Roy, Susmita; Mukherjee, Shibabrata; Mukherjee, Souvik; Chatterjee, Mitali Rapid sequestration of Leishmania mexicana by neutrophils results in the development of chronic lesion Hurrell, Benjamin (University of Lausanne, Biochemistry Department, Epalinges, CHE); Schuster, Steffen; Grün, Eva; Williams, Roderick; Held, Werner; Malissen, Bernard; Malissen, Marie; Yousefi, Shida; Simon, Hans-‐Uwe, Muller, Andreas; Tacchini-‐Cottier, Fabienne Leishmania major infection induces transmissible alterations in the skin microbiome Gimblet, Ciara (University of Pennsylvania, Philadelphia, PA, USA); Loesche,; Grice, Elizabeth; Scott, Phillip RNA sequencing to reveal host and parasite transcriptional responses during human cutaneous leishmaniasis Christensen, Stephen M. (University of Maryland, College Park, MD, USA); Dillon, Laura; Novais, Fernanda; Carvalho, Lucas; Passos, Sarah; Beiting, Daniel; Carvalho, Edgar; El-‐Sayed, Najib; Scott, Phillip; Mosser, David Alterations of the gut microbiota of the vector Lutzomyia longipalpis over time due to diet and Leishmania infection Kelly, Patrick H. (Department of Microbiology, University of Iowa, Iowa City, IA 52246, USA, Iowa City, IA, USA); Bahr, Sarah; Kamhawi, Shaden; Petrosino, Joseph; Valenzuela, Jesus; Wilson, Mary Conversion of monocyte derived M2 macrophages into a tissue resident macrophage phenotype Gonzalez, Michael (New York University School of Medicine, USA); Girgis, Natasha; Gundra, Uma Mahesh; Tang, Mei San; Gan, Wenbao; Loke, Png The mTORC2-‐IRF4 Axis Regulates Alternatively Activated Macrophages Huang, Stanley (Washington University School of Medicine, USA); Smith, Amber; O’Neill, Christina; Schilling, Joel; Pearce, Edward Schistosoma Mansoni infection induces anti-‐atherogenic transcriptional changes in hepatic macrophages Fairfax, Keke (Purdue University, West Lafayette, IN, USA); Elvington, Andrew (Washington University in St. Louis, USA); Pearce, Edward; Randolph, Gwendalyn The Immune Lives of Wild Mice Viney, Mark (University of Bristol, Bristol, GBR); Abolins, Steve (University of Bristol, USA); King, Liz (London School of Hygiene and Tropical Medicine, UK); Hafalla, Julius (London School of Hygiene and Tropical Medicine, UK); Riley, Eleanor (London School of Hygiene and Tropical Medicine, UK) Type 2 cytokine signalling alters dendritic cell phenotype and function Redpath, Stephen A (University of British Columbia, Vancouver, BC, CAN); Reynolds, Lisa; Heieis, Graham; Fonseca, Nicolette; Rahanjam, Negin; Perona-‐Wright, Georgia Toxoplasma gondii preferentially invades human monocytes and selectively activates host signaling pathways Gov, Lanny (University of California, Irvine, Irvine, CA); Ueno, Norikiyo; Wang, Kehui; Lodoen, Melissa Multiple Plasmodium Species Co-‐infections In African Populations Amanfo, Seth A. (The University of Edinburgh, Edinburgh, GBR); Midzi, Nicholas (The University of Zimbabwe); Mduluza, Takafira (The University of Zimbabwe); Theander, Thor G. (National University Hospital (Rigshospitalet), Copenhagen); Elhassan, Ibrahim; Arnot, David E.; Mutapi, Francisca; Cavanagh, David Dysfunctional neutrophils are a feature of Plasmodium cynomolgi infection in rhesus macaques Joyner, Chester (Emory University, USA); Lamb, Tracey; Ibegbu, Chris (Yerkes National Primate Research Center); Tirouvanziam, Rabindra; Galinski, Mary; MaHPIC, Consortium (USA) Integrating immune mechanisms in model of sheep gastro-‐intestinal nematodes. Garnier, Romain (Princeton University, Princeton, NJ, USA) MESENTERIC LYMPH NODE ORGANIZATION DURING HELMINTH INFECTION Dubey, Lalit Kumar (Ecole Polytechnique Fédérale de Lausanne, Lausanne, CHE); Mosconi, Ilaria; Burri, Olivier; Seitz, Arne; Luther, Sanjiv; Harris, Nicola Mucus and immunity to Trichinella spiralis: A systemic affair. Campbell, Laura (University of Manchester, Manchester, GBR); Bancroft, Allison; Whittingham-‐Dowd, Jayde; Gallagher, Amanda; Roberts, Ian; Thornton, David, Grencis, Richard Temporal transcriptional profiling of livers from Schistosoma mansoni-‐infected mice Owusu, Christian K. (Wellcome Trust Sanger Institute, Hinxton, GBR); Phythian-‐Adams, Alexander (University of Manchester); MacDonald, Andrew (University of Manchester); Berriman, Matt (Wellcome Trust Sanger Institute) Experimental and longitudinal evidence for the fitness costs of helminth infection in African buffalo Budischak, Sarah A. (Princeton University, Athens, GA, USA); Jolles, Anna (Oregon State University, USA); Ezenwa, Vanessa (University of Georgia, USA) Non-‐specific stressors: under-‐appreciated innate defenses against pathogens LeGrand, Edmund K. (College of Veterinary Medicine, University of Tennessee, Knoxville, TN, USA) Biological Sex is a Major Determinant of the Immune Response to Toxoplasma gondii Robinson, Dionne P. (National Institute of Allergy and Infectious Disease, USA); Gregg, Beth; Sikorski, Patricia; Grigg, Michael E. Genetic Determinants of Trypanotolerance in Humans Capewell, Paul (University of Glasgow, UK, Glasgow, GBR); Ilboudo, Hamidou (Centre International de Recherche Développement, Burkina Faso); Camara, Mamady (Programme National de Lutte contre la Trypanosomiase Humaine Africaine, Guinea); Garside, Paul ; Bucheton, Bruno (Institut de Recherche pour le Développement, France); Jamonneau, Vincent ; MacLeod, Annette NK-‐, NKT-‐ and CD8-‐derived IFN? drives myeloid cell activation and erythrophagocytosis, resulting in Trypanosomosis-‐associated acute anemia Cnops, Jennifer (Vrije Universiteit Brussel, brussel, BEL); De Trez, Carl; STIJLEMANS, Benoit; Keirsse, JIRI; Kauffmann, FLorence; Barkhuizen, Mark; Keathon, Roanne; Brombacher, Frank; Magez, Stefan Leishmania donovani activates B cells and induces cytokine expression by triggering endosomal TLRs Silva, Sasha (INRS-‐ Institut Armand Frappier, Laval, PQ, CAN) A role for T-‐bet in co-‐ordinating T cell activation Pritchard, Gretchen (University of Pennsylvania, Philadelphia, PA, USA); Konradt, Christoph; Roy, Nathan (University of Pennsylvania, USA); Dupont, Christopher; Burkhardt, Janis; Hunter, Christopher Fat Associated Lymphoid Clusters are IL-‐33R Dependent Sites of B Cell Proliferation and Local IgM Production. Jones, Lucy H. (University of Edinburgh, Edinburgh, GBR); Duncan, Sheelagh; Magalhaes, Marlène; McKenzie, Andrew; Maizels, Richard; McSorely, Henry; Allen, Judith; Bénézech, Cécile The formation and maintenance of brain resident memory CD8 T cells is dependent on the affinity of TCR-‐MHC interaction Sanecka, Anna (The Francis Crick Institute, Mill Hill Laboratory, London, GBR); Yoshida,; Frickel, Eva-‐ Maria The transcriptional repressor Hic1 regulates intestinal immunity Burrows, Kyle (University of British Columbia, Vancouver, BC, CAN); Antignano, Frann; Chenery, Alistair; Underhill, Michael; Zaph, Colby (Monash University) A role for dectin-‐1 in the CNS during chronic Toxoplasma gondii infection Worth, Danielle (University of California, Riverside, Riverside, CA, USA); Nance, J. Philip; Jang, Jessica; David, Clement; Nair, Meera; Wilson, Emma Role of microglia during cerebral malaria Capuccini, Barbara (The Francis Crick Institute, Mill Hill Laboratory, London, GBR); Lin, Jingwen; Sodenkamp, Jan; Keller-‐Mclauglin; Kushinga, Garikai; Langhorne, Jean The Paneth-‐cell specific role of autophagy in Toxoplasma gondii-‐induced intestinal inflammation Burger, Elise (UT Southwestern, USA); Yarovinsky, Felix Immunopathogenesis of visceral leishmaniasis -‐ New insights into infection-‐induced haematological disorders Preham, Olivier (University of York, York, GBR); Pinto, Ana; Brown, Najmeeyah; Chan, Samantha; Kaye, Paul Trefoil factor 2 mediates macrophage-‐dependent lung repair following hookworm infection Hung, Li-‐Yin (UCSF, San Francisco, CA, USA); Sen, Debasish; Oniskey, Taylor; Nieves, Wildaliz; Kim, Charles; Krummel, Matthew; Herbert, De'Broski A highly attenuated clone of Trypanosoma cruzi lacks the SAPA domain of trans-‐sialidase protein Junqueira, Caroline (Centro de Pesquisas Rene Rachou/ FIOCRUZ, Belo Horizonte, BRA); Neto, Rondon (ICB/ UFMG); Zuccherato, Luciana (ICB/ UFMG); Bartholomeu, Daniella (ICB/ UFMG); Vasconcelos, Ana Tereza (Laboratório Nacional de Computação); El-‐Sayad, Najib (University of Maryland); Burleigh, Barbara (Harvard University); Teixeira, Santuza (ICB/ UFMG); Gazzinelli, Ricardo (CpQRR/ FIOCRUZ) TALK ABSTRACTS (in order of presentation) Type I interferons induced by viral co-‐infection worsen murine leishmaniasis caused by L. guyanensis Rossi, Matteo (Department of Biochemistry, University of Lausanne, Lausanne, CHE); Castiglioni, Patrik ; Hartley, Mary-‐Anne; Eren, Remzi Onur; Prével, Florence; Utzschneider, Daniel T. (Swiss Vaccine Research Institute, Epalinges, Switzerland); Zehn, Dietmar (Swiss Vaccine Research Institute, Epalinges, Switzerland); Beverley, Stephen M.; Ronet, Catherine; Fasel, Nicolas Leishmania parasites are the causative agents of leishmaniasis: the second biggest protozoan parasitic infection after malaria, which affects over 12 million people in 98 countries worldwide. Leishmaniasis has various outcomes, depending on the species of the infecting parasite and the immunocompetence of its infected host. Symptoms range from a simple cutaneous, ulcerating lesion at the inoculation site to a potentially fatal visceralizing disease, where the parasite disseminates to internal organs. Infection with Leishmania guyanensis mainly results in simple cutaneous leishmaniasis, however 5-‐10% of patients develop disseminated or mucocutaneous lesions, which are refractory to current therapies. While the mechanism of this pathology is unknown, we have recently identified a virulence factor in L. guyanensis parasites in the form of a cytoplasmic virus, named Leishmania RNA virus (LRV). In this study, we investigated a possible role for type I interferons (IFN) in LRV-‐mediated virulence in a murine model of cutaneous leishmaniasis. We found that macrophages produce significantly higher levels of IFN-‐β at early time points following infection with LRVpositive L. guyanensis parasites in comparison to LRV negative L. guyanensis parasites. Interestingly, treating mice with IFN-‐b increases both lesion size and parasite burden, indicating that IFN-‐β is sufficient and essential to worsen the outcome of murine leishmaniasis. Further, we show that LCMV co-‐infection (notorious for its potent induction of type I IFN) can reproduce LRVpositive parasite phenotype in LRVnegative infected mice. These findings highlight the detrimental role of IFN-‐β during L. guyanensis infection and reveal the potential clinical significance of exogenous viral co-‐infection in disseminated or mucocutaneous leishmaniasis. Co-‐infection and immunity affect parasite burden in the wild: Insights from a wild wood mouse system Clerc, Melanie (University of Edinburgh, UK), Devevey, Godefroy; Pedersen, Amy B In nature, hosts are frequently co-‐infected with multiple parasites, which can interact directly or indirectly via resource competition or the host immune system. These interactions can determine disease severity, the release of infective stages, the parasites’ response to control strategies and, ultimately, the epidemiology of each parasite species. However, most studies addressing within-‐host parasite interactions use lab-‐based systems, which struggle to reflect the dynamics of natural systems. Wild wood mice (Apodemus sylvaticus), harbour a diverse community of >30 parasite species, are commonly co-‐infected and can be monitored longitudinally. In a wild population, we used anthelmintic treatment to reduce helminth burdens (dominated by Heligmosomoides polygyrus). Importantly, we found a 15-‐fold increase in burden of co-‐infecting intestinal coccidian parasites, specifically Eimeria hungaryensis. To better understand this antagonistic within-‐host interaction, we collected host and parasite demographic and immunological data in wild wood mice. We found a significant association between immune markers and worm infection. Interestingly, high total faecal IgA was negatively associated with infection risk, while high serum H. polygyrus-‐specific IgG1 was associated with reduced H. polygyrus burden. While these markers suggest a protective response, we found that concurrent Eimeria infection is associated with a reduction in total levels of both antibodies. This suggests that Eimeria infection may reduce the host’s response to H. polygyrus via down-‐regulation/interference with its antibody –dependent immune response. These findings highlight how the host’s immune system may be mediating this antagonistic interaction and stress the need for understanding the immune profiles of wild animals to predict disease outcomes. Leishmania and/or Host: Who drives the Immune Response during Indian Leishmaniasis? Ghosh, Susmita (Institute of Post Graduate Medical Education and Research, Kolkata, IND); Dey, Badal; Nath, Sankha; Roy, Susmita; Mukherjee, Shibabrata; Mukherjee, Souvik; Chatterjee, Mitali Visceral Leishmaniasis (VL) and Post Kala-‐azar dermal Leishmaniasis (PKDL) caused by Leishmania donovani are the two prevalent forms in South Asia. VL is associated with pancytopenia and severe immune suppression whereas a Th2 biased immune alteration occurs during PKDL. However, clinical studies regarding factors responsible for these varying immune responses remain ill defined, this study aimed to deliver a comprehensive picture from the point of view of pathogen and host. All clinically diagnosed patients were confirmed as VL/PKDL by PCR, rK39 strip test and ELISA; parasite load was measured by real time PCR. The circulatory markers were detected by real-‐time PCR or ELISA or by Multiplex system. Specific regions of the patient’s genome were sequenced by Sanger’s method to detect functional variants of the gene coding for Toll Like Receptors. The parasite load of patients with VL was significantly higher than patients with PKDL; however, the number of parasites did not correlate with humoral markers indicating that immune changes in patients are more likely host specific and not parasite driven. In an in vivo hamster model of VL, IFNγ expression was higher and positively correlated with parasite load. A higher proportion of the Eastern Indian population including patients with VL/PKDL demonstrated a Gly-‐Ile haplotype of TLR4, indicative of hyporesponsiveness to Lipopolysaccharide; additionally, VL patients had a significantly higher proportion of this haplotype than PKDL patients. The presence of this significantly higher proportion of the Gly-‐Ile haplotype in VL patients than in PKDL might account for the immunosuppressive nature of patients with VL, not for PKDL. Rapid sequestration of Leishmania mexicana by neutrophils results in the development of chronic lesion Hurrell, Benjamin (University of Lausanne, Biochemistry Department, Epalinges, CHE); Schuster, Steffen; Grün, Eva; Williams, Roderick; Held, Werner; Malissen, Bernard; Malissen, Marie; Yousefi, Shida; Simon, Hans-‐Uwe, Muller, Andreas; Tacchini-‐Cottier, Fabienne The protozoan Leishmania mexicana parasite causes chronic non-‐healing cutaneous lesions in humans and mice with poor parasite control. The mechanisms preventing the development of a protective immune response against this parasite are unclear. Here we provide data demonstrating that parasite sequestration by neutrophils is responsible for disease progression in mice. Within hours of infection L. mexicana induced the local recruitment of neutrophils, which ingested parasites and formed extracellular traps, without markedly impairing parasite survival. We further showed that the L. mexicana-‐induced recruitment of neutrophils impaired the early recruitment of dendritic cells at the site of infection as observed by intravital 2-‐ photon microscopy and flow cytometry analysis. Indeed, infection of neutropenic Genista mice and of mice depleted of neutrophils at the onset of infection demonstrated a prominent role for neutrophils in this process. Furthermore, an increase in monocyte-‐ derived dendritic cells was also observed in draining lymph nodes of neutropenic mice, correlating with subsequent increased frequency of IFNγ-‐secreting T helper cells, and better parasite control leading ultimately to complete healing of the lesion. Altogether, these findings show that L. mexicana exploits neutrophils to block the induction of a protective immune response and impairs the control of lesion development. Our data thus demonstrate an unanticipated negative role for these innate immune cells in host defense, suggesting that in certain forms of cutaneous leishmaniasis, regulating neutrophil recruitment could be a strategy to promote lesion healing. Leishmania major infection induces transmissible alterations in the skin microbiome Gimblet, Ciara (University of Pennsylvania, Philadelphia, PA, USA); Loesche,; Grice, Elizabeth; Scott, Phillip Cutaneous leishmaniasis is a disease characterized by ulcerating skin lesions, the resolution of which requires an effective, but regulated, immune response that limits parasite growth without causing permanent tissue damage. Studies have shown that skin commensals enhance the immune response to L. major, but how the microbiome changes during infection has not been characterized. Using analysis of the 16S ribosomal RNA gene, we found that infection with L. major causes a loss in bacterial diversity during the peak of infection, resulting in a dominance of the genus Staphylococcus. However once the lesions resolved, bacterial diversity returned to pre-‐infection levels. Alternatively, when mice developed more severely ulcerated lesions, the proportion of Streptococcus significantly increased on the lesions, demonstrating that L. major induced alterations in the skin microbiome can vary depending on the severity of disease. Strikingly, we observed similar changes in the microbiome on non-‐inflamed skin distant from the infection site, as well as in skin from cohoused naïve mice. These results indicate that the altered skin microbiome present at the site of infection is transmissible to uninfected skin despite having different immune responses at these sites. Current studies are directed at defining what factors mediate the changes in the skin microbiome during L. major infection and how this transmissible microbiome influences disease RNA sequencing to reveal host and parasite transcriptional responses during human cutaneous leishmaniasis Christensen, Stephen M. (University of Maryland, College Park, MD, USA); Dillon, Laura; Novais, Fernanda; Carvalho, Lucas; Passos, Sarah; Beiting, Daniel; Carvalho, Edgar; El-‐Sayed, Najib; Scott, Phillip; Mosser, David RNA sequencing using the Illumina platform was performed on biopsies from 13 patients with cutaneous leishmaniasis, and compared to 4 normal uninfected endemic controls. The analysis included biopsies from 5 L. braziliensis infected patients with early cutaneous leishmaniasis and 8 with late cutaneous leishmaniasis. High coverage sequencing allowed the sensitive detection of parasite transcripts and revealed that 2 of the early lesions and 2 of the late lesions had readily detectable parasite transcripts (greater than 300,000 counts) which accounted for almost 1% of total reads. Two of the early and 5 of the late lesions had no detectable parasite transcripts. This analysis allowed us to compare host responses in lesions where parasites were presumable being cleared (parasite transcript-‐negative) to lesions in which parasites were persisting or even multiplying. Lesions that contained parasite “detectable-‐positive transcripts” expressed substantial differences in host response transcripts relative to “detectable-‐negative” biopsies. The primary host transcripts that were produced in detectable-‐ positive lesions were overwhelmingly derived from B cells, with greater than half of the top 100 differentially expressed genes belonging to the immunoglobulin family. These data suggest that B cells and their products make a substantial contribution to the outcome of this disease. Studies from 12 more patients and 6 normal controls are currently being sequenced to expand this analysis. Alterations of the gut microbiota of the vector Lutzomyia longipalpis over time due to diet and Leishmania infection Kelly, Patrick H. (Department of Microbiology, University of Iowa, Iowa City, IA 52246, USA, Iowa City, IA, USA); Bahr, Sarah; Kamhawi, Shaden; Petrosino, Joseph; Valenzuela, Jesus; Wilson, Mary Leishmaniasis is a neglected tropical disease that affects one million people annually. Development of a successful preventive vaccine has been elusive. Therefore additional methods of sand fly vector transmission could be beneficial. Leishmania spp. parasites develop to an infectious form solely in the lumen of the female sand fly midgut, prior to transmission via blood meal. The infectious metacyclic is inoculated into host dermis, along with other components of the gut. This study is based on the dual hypotheses that: (1) The microorganisms resident in the gut of sand flies, collectively known as the microbiome, is a dynamic environment over time pending the sand fly diet, and (2) the microbiome may influence the development of Leishmania to an infectious form. In this study, we identified alterations in the gut microbiome over time in the female vector Lutzomyia longipalpis after infection with Leishmania infantum amastigotes. Control sand flies were fed blood only or sucrose at baseline, and flies fed with sterile or infected blood were shifted to sucrose meals after initial blood feeding. 16S rRNA Illumina sequencing was performed, and data were analyzed with the QIIME bioinformatic pipeline. The results indicated blood-‐fed sand flies experienced a dramatic down-‐shift in microbial diversity which recovered over time, becoming similar to sucrose controls. However, in sand flies fed Leishmania infected blood meals, the microbiota did not return to basal diversity levels. Loss/acquisition of genera was associated with maturing infection. These data provided the basis for metagenomic analyses. KEGG ortholog analysis revealed bacterial heme and iron(III) transport and phosphate transport pathways were represented at a decreased relative frequency in infected versus uninfected blood fed flies. These findings indicate that the sand fly vector microbiome is affected by Leishmania infection. They also raise hypotheses that parasite-‐induced perturbation of the microbiota and consequent nutrient availability may influence parasite fitness and transmission to hosts Conversion of monocyte derived M2 macrophages into a tissue resident macrophage phenotype Gonzalez, Michael (New York University School of Medicine, USA); Girgis, Natasha; Gundra, Uma Mahesh; Tang, Mei San; Gan, Wenbao; Loke, Png Alternatively activated M2 macrophages accumulate during inflammation via recruitment of inflammatory monocytes, or through the proliferation of tissue resident macrophages. Monocyte-‐ derived and tissue-‐resident M2 macrophages are phenotypically and functionally distinct, with UCP1 expression being a unique feature of tissue-‐resident cells. We now show that UCP1 expression and proliferating cells accumulate in the periphery of mature granulomas during S. mansoni infection. Genetic fate mapping of macrophages derived from CX3CR1(+) monocytes indicate that UCP1-‐ expressing cells are derived from monocyte precursors. Conversion to a tissue-‐resident phenotype occurs independently of Stat6, although Stat6 regulates macrophage proliferation both intrinsically and extrinsically. Vitamin A deficiency (VAD) leads to dysregulation of the balance between monocyte-‐ derived and tissue-‐resident M2 macrophages. Liver granulomas of S. mansoni infected VAD mice no longer express UCP1 and GATA6, indicating a failure to adopt a tissue-‐resident phenotype, and are associated with increased mortality. Retinoic acid treatment of S. mansoni infected VAD mice reverses some of the defects. Hence, retinoic acid may regulate the conversion of monocyte-‐derived inflammatory M2 macrophages into a tissue-‐resident phenotype. The mTORC2-‐IRF4 Axis Regulates Alternatively Activated Macrophages Huang, Stanley (Washington University School of Medicine, USA); Smith, Amber; O’Neill, Christina; Schilling, Joel; Pearce, Edward The mammalian target of rapamycin complexes, mTORC1 and mTORC2 have emerged as important regulators of environmental cues for development of immune cells. mTORC1 is highly responsive to changes in cellular glucose and amino acid levels. The constitutive activation of mTORC1 has been shown to negatively regulate alternative (M2) macrophage activation, but the role of mTORC2 in macrophage activation is still unknown. Here we report that macrophages lacking Rictor, a critical adaptor protein required for mTORC2 activity in macrophages fail to become fully M2 activated during Heligmosomoides polygyrus infection. Moreover, IL-‐4 stimulated protective immunity to this parasite is significantly diminished in LysMcre x Rictorfl/fl mice. We noted that IRF4 expression and glycolytic/ mitochondrial metabolism were markedly decreased in Rictor-‐deficient M2 macrophages. IRF4 deletion in macrophages affected neither mTORC2 activity nor the phosphorylation of Stat6 in response to IL-‐4, but led to a significant reduction in glucose metabolism and M2 activation. Taken together, our finding suggests that signaling through mTORC2-‐IRF4 is critical for IL-‐4 induced changes in metabolism and M2 activation. Schistosoma Mansoni infection induces anti-‐atherogenic transcriptional changes in hepatic macrophages Fairfax, Keke (Purdue University, West Lafayette, IN, USA); Elvington, Andrew (Washington University in St. Louis, USA); Pearce, Edward; Randolph, Gwendalyn Hepatic macrophages play an essential role in the granulomatous response to infection with the parasitic helminth Schistosoma Mansoni, but the transcriptional changes that underlie this participation are poorly understood. To explore this, we sorted the two previously recognized hepatic macrophage populations (perivascular and Kupffer) from naïve and S. mansoni infected mice and performed micro-‐ array analysis as part of the Immunological Genome Project. Consistent with the pattern of great diversity identified in other organ macrophages, the two hepatic macrophage populations displayed signatures distinct from all other macrophages, with the two populations exhibiting remarkable differences between them. However, this diversity was greatly reduced following infection with S. mansoni, and in fact, many of the transcripts identified as uniquely perivascular or Kupffer cell specific were lost following infection, raising the possibility that both populations may be replenished by monocytes following infection. Our analysis showed a profound alteration in phospholipid and cholesterol metabolic pathways, including prostaglandin signaling, in addition to alterations in M2 markers. These changes suggested a possible mechanism for the previously reported atheroprotective effects of S. mansoni infection. Indeed we find that ApoE null mice fed a high fat diet in combination with S. mansoni infection have reduced body mass in addition to reduced plaque area as compared to control mice. The Immune Lives of Wild Mice Viney, Mark (University of Bristol, Bristol, GBR); Abolins, Steve (University of Bristol, USA); King, Liz (London School of Hygiene and Tropical Medicine, UK); Hafalla, Julius (London School of Hygiene and Tropical Medicine, UK); Riley, Eleanor (London School of Hygiene and Tropical Medicine, UK) The immune responses made by laboratory mice, Mus musculus domesticus, are very well known, but those of their wild relatives are not. We have characterised the immune status of c.500 wild mice; specifically, the total serum immunoglobulin (Ig) concentration, the faecal IgA concentration, a FACS characterisation of splenocytes, and the cytokine response of in vitro-‐stimulated splenocytes. In parallel we have characterised the metazoan parasite fauna of these animals, together with evidence of their viral infections. We find that these wild mice, compared with laboratory-‐bred mice, have (i) substantially higher total IgG and IgE concentrations and higher concentrations of the acute phase proteins haptoglobin and serum amyloid P; (ii) significantly lower numbers of splenic T and B lymphocytes and NK cells, but substantially higher levels of lymphocyte activation and differentiation (i.e. a higher ratio of memory or effector cells to naïve cells; increased expression of CD69, MHC Class II, KLRG1 and higher IFN-‐γ production in response to CD3/CD28 cross-‐linking); (iii) broadly similar cytokine responses to a range of innate immune ligands although with a tendency for lower levels of secretion of IL-‐1β, IL-‐12 and IL-‐13. These observations are consistent with continual, high-‐level antigen exposure among wild mice but we find little correlation between current infection status and measures of immune function. Type 2 cytokine signalling alters dendritic cell phenotype and function Redpath, Stephen A (University of British Columbia, Vancouver, BC, CAN); Reynolds, Lisa; Heieis, Graham; Fonseca, Nicolette; Rahanjam, Negin; Perona-‐Wright, Georgia The immune responses generated by chronic infections can compromise the host’s ability to respond to unrelated infections. It has already been shown that the type 2 cytokines elicited by helminth infection influence naïve T cells and inhibit their ability to mount a Th1 response during subsequent antigen challenge. Antigen presenting cells such as dendritic cells (DC) are also exposed to type 2 cytokines during helminth infection. We therefore hypothesized that exposure to type 2 cytokine signals during infection would alter DC function, skewing them towards a phenotype that would favour Th2 differentiation. To test this, we used the nematode Heligmosomoides polygyrus to set up a strong Th2 environment in the small intestine and gut-‐associated lymphoid tissue (GALT). H. polygyrus infection caused marked changes in DC phenotype in the GALT, imposing an alternatively activated phenotype that included increased CD40, IL-‐6 and RELM-‐a expression. These changes were dependent on IL-‐13 signalling, and enabled the DC to drive Th2 differentiation in naive CD4+ T cells. Our preliminary data suggest that, during infection, the principal source of IL-‐13 is type 2 innate lymphoid cells. Together our data indicate that IL-‐13 signalling in helminth infection imprints gut-‐associated DC with a phenotypic and functional bias that preferentially primes Th2 immune responses. This Th2 bias could impair the development of protective immunity to a subsequent infection with a different pathogen. Given the prevalance of helminth infections and co-‐infections, further understanding of the mechanisms that dictate appropriate immune polarisation are essential for the successful design of treatment and vaccination programs. Toxoplasma gondii preferentially invades human monocytes and selectively activates host signaling pathways Gov, Lanny (University of California, Irvine, Irvine, CA); Ueno, Norikiyo; Wang, Kehui; Lodoen, Melissa Toxoplasma gondii is a parasite that infects one third of the human population and can cause life-‐ threatening disease in the developing fetus and in immunocompromised individuals. Transmission to humans typically occurs via accidental ingestion of parasite cysts. During acute infection, a major avenue by which T. gondii disseminates in the body is the bloodstream. To better understand the interaction of blood leukocytes with T. gondii, we infected human peripheral blood mononuclear cells (PBMC) with T. gondii and examined the association of the parasite with each immune cell population. Strikingly, even though monocytes were a minority of the PBMC population, monocytes preferentially harbored T. gondii. Among purified monocytes, T. gondii associated with all three monocyte subsets (classical, resident, and intermediate). Similarly, during infection of mice, T. gondii was predominantly associated with CD11b+ cells in the peripheral blood. Monocytes are phagocytes, and the parasite has two potential fates when interacting with a phagocytic cell: T. gondii can directly invade a monocyte and establish a parasitophorous vacuole, leading to parasite survival, or the parasite can be phagocytosed, which typically leads to its destruction. In a detailed kinetic analysis, ~60% of intracellular parasites had invaded the monocytes based on GRA7 staining whereas ~40% of intracellular parasites had been phagocytosed and were positive for the phagolysosomal marker LAMP1. We examined the activation of the signaling pathways in response to invasion or phagocytosis and found selective phosphorylation of MAPK proteins. Collectively, this work provides novel insight into the interactions of human leukocytes with a parasitic pathogen of global importance. Multiple Plasmodium Species Co-‐infections In African Populations Amanfo, Seth A. (The University of Edinburgh, Edinburgh, GBR); Midzi, Nicholas (The University of Zimbabwe); Mduluza, Takafira (The University of Zimbabwe); Theander, Thor G. (National University Hospital (Rigshospitalet), Copenhagen); Elhassan, Ibrahim; Arnot, David E.; Mutapi, Francisca; Cavanagh, David In malaria endemic countries individuals may harbour multiple Plasmodium species. However, the epidemiology of multiple Plasmodium species co-‐infections in many African populations has not been well characterised owing to the several limitations associated with malaria diagnosis by blood film microscopy. We have used merozoite surface protein 1 (MSP-‐119) from the four major human Plasmodium parasites and a panel of other antigens derived from MSPs of P. falciparum to determine the antibody responses to multiple P. falciparum serotypes and other Plasmodium species co-‐infections in both cross-‐sectional Zimbabwean cohort and a 10-‐year longitudinal sampling of individuals from Daraweesh, Sudan. Preliminary analysis shows that anti-‐MSP-‐119 antibodies are highly species specific. In both studies, IgG responses to MSP-‐119 of P. falciparum (PfMSP-‐119) were the most frequently observed followed by responses to P. malariae and P. ovale, with responses to P. vivax MSP-‐119 rarely detected. Anti-‐MSP-‐119 responses to multiple Plasmodium species were more frequently observed in these population. Interestingly, almost all sera positive for P. malariae, P. ovale and/or P. vivax MSP-‐119 were also positive for PfMSP-‐119, indicating that co-‐infections are a common occurrence, and that mono infections with the non-‐falciparum species are rare. In the Daraweesh study, very few individuals with cases of documented clinical malaria had no detectable Ab response to any of the four parasites’ MSP-‐ 119. Some individuals however, showed a selective responsiveness to MSP-‐119, having no Ab response in earlier malaria infections, but mounting a strong Ab response to P. falciparum MSP-‐119 in later episodes. Serum samples with antibody reactivities to PfMSP-‐119 often also showed reactivity to PfMSP-‐2A and PfMSP-‐2B derived antigens, but the patterns of reactivity with these polymorphic antigens varied markedly between individuals. This study reinforces the importance of diagnosing not only the presence of Plasmodium parasites, but also the species of Plasmodium infecting a patient for optimal treatment of malaria co-‐infections. Dysfunctional neutrophils are a feature of Plasmodium cynomolgi infection in rhesus macaques Joyner, Chester (Emory University, USA); Lamb, Tracey; Ibegbu, Chris (Yerkes National Primate Research Center); Tirouvanziam, Rabindra; Galinski, Mary; MaHPIC, Consortium (USA) Plasmodium vivax is the most widespread human malaria parasite and accounts for a significant portion of malaria worldwide. The innate immune system is likely to play an important role in P. vivax infection, but studies to-‐date have been limited. Neutrophils are the most abundant leukocytes in the blood and serve as a primary defense against invading microorganisms through phagocytosis and the production and release of inflammatory mediators that amplify immune responses. When excessive, however, these responses can lead to immunopathology. The role of neutrophils during acute P. vivax infection has not been extensively characterized. Here, we have utilized the rhesus macaque -‐ Plasmodium cynomolgi model system of P. vivax infection to determine how neutrophils respond during infection. We first developed novel flow cytometry-‐based assays to monitor the activation and functional status of rhesus neutrophils using small volumes of blood. Five rhesus were then infected with sporozoites, clinically monitored, and their neutrophil function assessed alongside other cellular subsets during acute infection. Neutrophils became significantly activated based on expression of surface markers such as CD66 during acute infection, demonstrating a role for this cellular subset during infection. Additionally, neutrophil caspase-‐1 activity, measured by the cell-‐permeant, fluorescent substrate FLICA, increased significantly and correlated with an increase in IL-‐1β in the plasma. However, the phagocytic ability of neutrophils was impaired at this point during infection. Collectively, these results indicate that neutrophils become dysfunctional during malaria and lose their phagocytic ability but continue to produce pro-‐inflammatory cytokines, which could contribute significantly to malaria pathogenesis. Furthermore, the diminished neutrophil phagocytic ability could provide mechanistic insight into secondary bacterial infections that are common during this disease. Integrating immune mechanisms in model of sheep gastro-‐intestinal nematodes. Garnier, Romain (Princeton University, Princeton, NJ, USA) Gastrointestinal nematodes are a major source of disease not only in humans but also in livestock. In small ruminants, several species have the potential to cause substantial (economic) losses and or even death. Because of this importance, a large number of theoretical models have been developed ranging from simple phenomenological models to very detailed multi-‐stage models. However, there is a distinct lack of relatively simple models that include specific characteristic of the immune response to gastro-‐ intestinal nematodes. Here I will present a phenomenological model that has been modified to integrate empiric knowledge of the immune response elicited by Teladorsagia circumcincta in sheep. In this system, immunoglobulins A and E are thought to be the most important immune effectors and to act on larval stages of the nematode. This is modelled by considering that immunity accumulates with host exposure to larval parasites and subsequently reduces establishment, larval maturation and adult fecundity. The model is parameterized using published data of domestic sheep trickle infection experiments. I will show that this limited set of immuno-‐epidemiological assumptions is sufficient to describe both the adult worm burdens and the fecal egg counts under different infection regimes. Such models could benefit from the increasing availability of immune markers. A better ability to ground-‐ truth the immune component of the model with data would be crucial if these models are to be used to predict the efficacy of different treatments or of vaccines that are currently being developed. MESENTERIC LYMPH NODE ORGANIZATION DURING HELMINTH INFECTION Dubey, Lalit Kumar (Ecole Polytechnique Fédérale de Lausanne, Lausanne, CHE); Mosconi, Ilaria; Burri, Olivier; Seitz, Arne; Luther, Sanjiv; Harris, Nicola The lymph node stroma acts to form conduits, to provide a scaffold and can additionally regulate CD8+ T cell immune responses. We investigated the organizational and functional roles of the stroma following type 2 immunity raised against the intestinal helminth Heligmosomoides polygyrus bakeri (Hp). Visualization of the mesenteric lymph node revealed dramatic changes in cellularity and organization with an extensive expansion of the T cell zone stroma i.e. FRCs. Stromal cell re-‐organization and expansion was driven by LTβR signaling and required IL-‐4Rα which acted to promote LTαβ expression by lymphocytes. Strikingly, during the course of infection the FRCs start aggregating around the newly formed B cell/germinal centers indicating a novel role of T cell stromal cells in B cell organization. In conclusion, our study provides further evidence to understand the mesenteric lymph node biology during infection with a Th2-‐polarizing pathogen. Our data showed that stromal cells are early sensors of infection, which can adapt structurally and functionally with the help of hematopoietic cells and all of these effects were dependent on IL-‐4Rα signaling. Mucus and immunity to Trichinella spiralis: A systemic affair. Campbell, Laura (University of Manchester, Manchester, GBR); Bancroft, Allison; Whittingham-‐Dowd, Jayde; Gallagher, Amanda; Roberts, Ian; Thornton, David, Grencis, Richard Trichinella spiralis induces a strong Type 2 cytokine response to infection, which drives expulsion of the parasite from the intestine by generating a broad inflammatory response that includes intestinal mast cell and goblet cell hyperplasia. The present work focuses on the mucosal goblet cell response both within the intestine and in the lung as an example of a non-‐parasitised epithelium to explore the broader systemic mucin response to infection. C57BL/6 mice were infected with T. spiralis and goblet cell responses/mucin analyses were investigated in the small intestine and lungs. The data demonstrates that in addition to a profound goblet cell hyperplasia in the small intestine, T. spiralis induces a strong goblet cell hyperplasia within the lungs, initiated before newborn larvae have entered the circulatory system (characterized by elevation of Muc5b and Muc5ac). The Type 2 cytokine, interleukin 13 is known to promote goblet cell hyperplasia in the intestine, and utilisation of a neutralising IL-‐13 antibody confirms this also drives the systemic mucin response with the lung. Both mucins were also found to be up-‐regulated in the lungs of infected immunodeficient animals (SCID mice) and even after treatment to remove innate lymphoid cells, known producers of IL-‐13. These results suggest that Trichinella drives a systemic mucosal response characterised by an IL-‐13 driven production of Muc5ac producing goblet cells at sites distinct from the infection site. This response can occur in the absence of adaptive immunity and does not appear to depend on innate lymphoid cells. Temporal transcriptional profiling of livers from Schistosoma mansoni-‐infected mice Owusu, Christian K. (Wellcome Trust Sanger Institute, Hinxton, GBR); Phythian-‐Adams, Alexander (University of Manchester); MacDonald, Andrew (University of Manchester); Berriman, Matt (Wellcome Trust Sanger Institute) The immune response to Schistosoma mansoni infection has been well-‐characterised in experimental mouse models. Early in the infection, migrating larvae trigger a mixed type 1 (Th1) and 2 (Th2) inflammatory response. This changes to a dominant Th2 response after mated worm pairs begin producing eggs around week 5 post infection. The Th2 response peaks around week 8, after which the response gradually weakens, corresponding to the chronic phase of the disease. While much is known about the major effector cells involved in immunopathological changes in the liver, we are still largely ignorant of the exact molecular triggers leading to the observed pathology. I will present a transcriptome data profiling changes in gene expression in the livers of infected mice across 6 time points. The time points were chosen to reflect the progression of the immune response, beginning prior to the onset of egg deposition, and extending all the way through to the chronic stage of the disease. We identify putative mRNA targets of differentially expressed miRNAs through the integrated analysis of miRNA and mRNA expression profiles. In addition, we show evidence of changes in expression of receptors and ligands involved in antigen presentation and T cell activation. These data provide clues regarding possible mechanisms underlying T cell hyporesponsiveness seen during the later time points. We also show temporal changes in the expression of genes involved in extracellular matrix organisation and corresponding to changes in liver tissue architecture as the granulomatous response becomes fibrotic. Finally, cluster analysis reveals several genes which share similar patterns of expression. In many clusters, genes grouped together either act in the same pathway or identify effector cell types showing coordinated recruitment to the liver across the different time points. Experimental and longitudinal evidence for the fitness costs of helminth infection in African buffalo Budischak, Sarah A. (Princeton University, Athens, GA, USA); Jolles, Anna (Oregon State University, USA); Ezenwa, Vanessa (University of Georgia, USA) The fitness costs of parasite infection are poorly described for wild hosts where coinfection with multiple sub-‐lethal parasites is common. To explore links between helminth infection and components of host physiology and fitness, we conducted a 4-‐year deworming experiment and longitudinal study of the gastrointestinal helminths of African buffalo (Syncerus caffer). We combined experimental, longitudinal, and species-‐specific approaches to prove strong inference into the fitness costs of helminth infection. Both experimental and longitudinal evidence suggest that helminths pose significant fitness costs, but these costs may be species-‐specific. Several condition fitness-‐metrics were negatively associated with Haemonchus abundance, but surprisingly, others were positively associated with Cooperia abundance. Again, species-‐specific differences emerged with declines in Haemonchus observed when hosts elevated eosinophil levels and a negative relationship between Haemonchus abundance and host nutrition. Conversely, Cooperia abundance was not associated with host eosinophilia or nutrition. The positive effects of experimental removal on buffalo condition and survival were not evident when total helminth abundance of control hosts was examined. This discrepancy suggests that helminths pose significant fitness costs to their hosts, but having a naturally low helminth load also impacts host fitness. In combination, our data suggest that adopting a tolerance strategy to low-‐virulence parasites may be more beneficial for host fitness than a resistance strategy. Overall, the effects of helminth infection on host fitness may be cryptic and indirect, yet have substantial consequences for the dynamics of host populations. Non-‐specific stressors: under-‐appreciated innate defenses against pathogens LeGrand, Edmund K. (College of Veterinary Medicine, University of Tennessee, Knoxville, TN, USA) Ideally host defenses should harm pathogens without causing self-‐harm. Nevertheless, even completely non-‐specific stressors which harm host and pathogens alike, such as reactive molecular species, heat, nutrient and oxygen restriction, and acidity, have utility for host defense. This utility is based on differences in vulnerability between pathogens and host and on degree of stress localization. The universal vulnerability to stress associated with growth and replication can be used by the host to preferentially harm rapidly replicating pathogens. Infected foci experience intense local non-‐specific stress, affecting pathogens, host tissue, and immune cells, though pathogens must replicate there while infiltrating immune cells are expendable. Regionally, linkage of coagulation and inflammation reduces blood flow around infected sites, adding to the non-‐specific stress. Systemic stressors of the acute-‐phase response, e.g., fever and nutrient restriction, support the more intense regional and local stress, again disproportionally harming the pathogens. We used a simple agent-‐based model of a locally infected host to explore the efficacy of host defenses consisting of completely non-‐specific stress in controlling rapidly replicating pathogens. In our model local, regional, and systemic stress act synergistically in eliminating pathogens; the key premise being that pathogens use their resources for more rapid replication than host cells, thus having fewer reserves to withstand stress. Since non-‐specific stress has a multitude of targets and since defenses against stress are so ancient, evolution of new anti-‐stress defenses by pathogens seems unlikely, in contrast to evolving defenses against highly specific therapies. Biological Sex is a Major Determinant of the Immune Response to Toxoplasma gondii Robinson, Dionne P. (National Institute of Allergy and Infectious Disease, USA); Gregg, Beth; Sikorski, Patricia; Grigg, Michael E. Females across numerous species respond immunologically to infectious agents differently than males. Females typically mount more robust immune responses, and for the majority of murine protozoal infections, are better protected from severe disease compared with males. However, this is not the case during Toxoplasma gondii infection. Excessive immune responsiveness in females (e.g., increased IFN-‐γ, TNF-‐α) coupled with decreased immune regulation (e.g., regulatory T cell (Tregs)) leads to the development of immunopathology. Using male and female C57BL/6J mice inoculated with Toxoplasma (Type II), we observe greater mortality in females than males following peroral, but not intraperitoneal (i.p.) inoculation that is not a result of differential parasite replication but is exacerbated by small intestine microbial dysbiosis. To investigate the relative and contributing roles of differential dysbiosis, inflammation and endocrine responses in the more susceptible females, small intestine contents had greater γ-‐proteobacteria outgrowth in females compared with males when assessed by RT-‐PCR. To determine whether endocrine function is influencing inflammation in Toxoplasma-‐infected mice, uterine horns and seminal vesicles (i.e., hormone-‐responsive tissues) were weighed. Males exhibited no change in seminal vesicle mass following Toxoplasma infection, and possessed greater serum concentrations of the regulatory cytokines, IL-‐5, IL-‐13, and GM-‐CSF. Conversely, females inoculated perorally, but not i.p., experienced a significant loss in uterine horn mass, indicative of reduced estradiol concentrations, and had higher serum concentrations of IFN-‐γ, which is associated with immunopathology. Interestingly, males experienced a greater Treg collapse in the small intestine than females, despite being better protected against toxoplasmosis. Collectively, our data suggest the existence of a ‘male-‐typic’ regulatory profile that modulates inflammation and microbial dysbiosis. Genetic Determinants of Trypanotolerance in Humans Capewell, Paul (University of Glasgow, UK, Glasgow, GBR); Ilboudo, Hamidou (Centre International de Recherche Développement, Burkina Faso); Camara, Mamady (Programme National de Lutte contre la Trypanosomiase Humaine Africaine, Guinea); Garside, Paul ; Bucheton, Bruno (Institut de Recherche pour le Développement, France); Jamonneau, Vincent ; MacLeod, Annette Trypanosoma brucei gambiense causes 97% of human African trypanosomiasis (HAT) cases and was believed to be invariably fatal. The recent discovery of asymptomatic individuals demonstrates that this is not the case. Asymptomatic (or trypanotolerant) individuals are positive for trypanosomes using antibody and PCR assays, although no symptoms are presented nor are parasites found within the vasculature. This asymptomatic period can be protracted – one recent case persisted for 29 years before immunosuppressive treatment for an unrelated condition caused symptoms to emerge, highlighting involvement of immunological components in controlling the parasite. The trypanosomes infecting asymptomatic individuals have been characterised and do not differ from those in symptomatic patients, indicating asymptomatic individuals directly moderate infection. To identify host immune factors that may be associated with the asymptomatic phenotype, we analysed the transcriptomes of peripheral blood mononuclear cells (PBMCs) from sympatric uninfected, symptomatic and asymptomatic T. b. gambiense patients from a HAT focus in Guinea. A comprehensive chemokine and cytokine analysis was also performed on matched samples to support inferences from the PBMC transcriptomics. Previous studies on human trypanosome infections focused on cytokine levels at various stages of disease, only examining patients with symptomatic disease and thus not assessing the protective response. Our study identified several transcriptional and cytokine/chemokine differences that associate with the asymptomatic phenotype and may be directly involved in controlling T. b. gambiense infection. These factors may also prove useful for biomarker development and diagnostic measures to identify asymptomatic T. b. gambiense carriers. NK-‐, NKT-‐ and CD8-‐derived IFN? drives myeloid cell activation and erythrophagocytosis, resulting in Trypanosomosis-‐associated acute anemia Cnops, Jennifer (Vrije Universiteit Brussel, brussel, BEL); De Trez, Carl; STIJLEMANS, Benoit; Keirsse, JIRI; Kauffmann, FLorence; Barkhuizen, Mark; Keathon, Roanne; Brombacher, Frank; Magez, Stefan African trypanosomes are the causative agents of Human African Trypanosomosis (HAT/Sleeping Sickness) and Animal African Trypanosomosis (AAT/Nagana). A common hallmark of African trypanosome infections is inflammation. In murine trypanosomosis, the onset of inflammation occurs rapidly after infection and is manifested by an influx of myeloid cells in both liver and spleen, accompanied by a burst of serum pro-‐inflammatory cytokines. Within 48 hours after reaching peak parasitemia, acute anemia develops and the percentage of red blood cells drops by 50%. Using a newly developed in vivo erythrophagocytosis assay, we demonstrate that activated cells of the myeloid phagocytic system display enhanced erythrophagocytosis causing acute anemia. Results indicate that IFNγ plays a crucial role in the recruitment and activation of erythrophagocytic myeloid cells, as mice lacking the IFNγ receptor were partially protected against trypanosomosis-‐associated inflammation and acute anemia. NK and NKT cells were the earliest source of IFNγ during T. b. brucei infection. Later in infection, CD8+ and to a lesser extent CD4+ T cells become the main IFNγ producers. Cell depletion and transfer experiments indicated that during infection the absence of NK, NKT and CD8+ T cells, but not CD4+ T cells, resulted in a reduced anemic phenotype similar to trypanosome infected IFNγR-‐/-‐ mice. Collectively, this study shows that NK, NKT and CD8+ T cell-‐derived IFNγ is a critical mediator in trypanosomosis-‐associated pathology, driving enhanced erythrophagocytosis by myeloid phagocytic cells and the induction of acute inflammation-‐associated anemia Leishmania donovani activates B cells and induces cytokine expression by triggering endosomal TLRs Silva, Sasha (INRS-‐ Institut Armand Frappier, Laval, PQ, CAN) Leishmania donovani, a causative agent of Visceral Leishmaniasis (VL), is known to induce polyclonal B cell activation and hypergammaglobulinemia. B cell activation by the parasite is detrimental to infection; indeed B-‐cell deficient mice are highly resistant to VL. Several publications have now demonstrated that B cells can regulate adaptive T cell responses by various antibody independent mechanisms, such as cytokine production. B cells can both enhance or suppress CD4+ T cell responses, depending on the disease model. We have recently shown that B cells and marginal zone B cells (MZB) suppress protective T cell responses during experimental VL. This suppression is partly mediated by IL-‐10 produced by MZB. Here, we characterize the interaction between L. donovani amastigotes and splenic B cell subsets, and investigate activation pathways triggered by the parasite. We show that upon exposure to L. donovani, B cells form clusters retaining one or more parasites on their surface. This close interaction induces the expression of various proinflammatory cytokines, type I interferons, and IL-‐10. Interestingly, cytokines expression was completely abrogated in B cells from UNC93b1-‐deficient mice, suggesting that endosomal TLRs are triggered by the parasite and required for B cell activation. Taken together, we demonstrate that endosomal TLR activation is one of the main mechanism by which L. donovani activates B cells. Further studies are granted in order to understand the exact role of the various B cell-‐ derived cytokines in regulating T cell responses and suppressing anti-‐leishmanial immunity. A role for T-‐bet in co-‐ordinating T cell activation Pritchard, Gretchen (University of Pennsylvania, Philadelphia, PA, USA); Konradt, Christoph; Roy, Nathan (University of Pennsylvania, USA); Dupont, Christopher; Burkhardt, Janis; Hunter, Christopher The T-‐box transcription factor T-‐bet is perhaps most prominently known as a master regulator of Th1 differentiation and IFNγ production. Recent work from our laboratory has identified a novel role for this transcription factor in coordinating the effector T cell responses necessary to control the intracellular parasite Toxoplasma gondii in peripheral tissues, and suggests that T-‐bet may function in a diverse array of immunological processes. The chemokine CCL3 and the integrin CD11a are both involved in cell:cell interactions during T cell priming and activation. Here, we show that T-‐bet expression correlates with that of CCL3 and that T-‐bet is necessary for its production. In addition, T-‐bet is required for optimal upregulation of CD11a and ICAM-‐binding, an observation that suggests that T-‐bet is required for optimal cell:cell interactions during T cell activation. Support for this idea is provided by studies which showed that in mice challenged with T. gondii, the absence of T-‐bet or CCL3, or CD11a blockade, results in similar reduction in T cell responses. Thus, these findings highlight the importance of these proteins for in vivo T cell responses and suggest a role for T-‐bet in the coordination of the very early events in T cell activation that are necessary for optimal CD8+ T cell responses. Fat Associated Lymphoid Clusters are IL-‐33R Dependent Sites of B Cell Proliferation and Local IgM Production. Jones, Lucy H. (University of Edinburgh, Edinburgh, GBR); Duncan, Sheelagh; Magalhaes, Marlène; McKenzie, Andrew; Maizels, Richard; McSorely, Henry; Allen, Judith; Bénézech, Cécile The peritoneal and pleural cavities constitute a reservoir of B cells that produce natural antibodies vital for the early control of infection. In contrast to the peritoneal cavity, very little is known about the structures sustaining B cells in the pleural cavity. Using a mouse model of filarial nematode infection (Litomosoides sigmodontis), we showed that the Fat Associated Lymphoid Clusters (FALCs) found in the pericardial and mediastinal fat deposits of the pleural cavity supported the proliferation of B1 and B2 cells that differentiated rapidly into IgM producing plasma cells. These nematode induced clusters supported local secretion of IgM directly into the pleural cavity. Moreover, IL-‐33R was crucial for the local proliferation of all pleural B cell subsets and FALC B-‐cell IgM secretion. Finally, we demonstrated that FALCs themselves are a local source of IL-‐33 within the pleural cavity. The formation and maintenance of brain resident memory CD8 T cells is dependent on the affinity of TCR-‐MHC interaction Sanecka, Anna (The Francis Crick Institute, Mill Hill Laboratory, London, GBR); Yoshida,; Frickel, Eva-‐ Maria The protozoan parasite Toxoplasma gondii is the most common parasitic infection in man. In immunocompetent hosts the acute phase of infection is generally asymptomatic and proceeds to the chronic phase, which is incurable and defined by tissue cyst formation preferably in the brain. The parasite poses a serious health threat to immunocompromised individuals. IFNγ and CD8 T cells are essential for the control of chronic Toxoplasma infection. We are defining how affinity of the TCR-‐MHC interaction shapes the phenotype of memory CD8 T cells in the chronically Toxoplasma-‐infected brain. As a unique tool we are using CD8 T cells from three lines of transnuclear mice that harbour, in their endogenous loci, different T cell receptors specific for the same Toxoplasma antigenic epitope Rop7. The three transnuclear CD8 T cell clones span a wide range of affinities to MHCI-‐Rop7. Our data show that these three CD8 T cell clones have a distinct and fixed hierarchy in effector function such as proliferation capacity, trafficking, maintenance and memory formation. In particular, the clone of lowest affinity does not home to the brain. The two higher affinity T cell clones show differences in establishing resident memory populations (CD103+) in the brain and vary in their time of persistence in the host during chronic infection. We are currently transcriptionally profiling naïve, activated and memory subpopulations of Rop7-‐specific CD8 T cells to better understand signalling pathways leading to observed phenotypic differences. This study uses Balb/c mice that are resistant to chronic Toxoplasma infection advancing the knowledge of control of Toxoplasma by CD8 T cells specifically and resident memory CD8 T cells in the brain in general. The transcriptional repressor Hic1 regulates intestinal immunity Burrows, Kyle (University of British Columbia, Vancouver, BC, CAN); Antignano, Frann; Chenery, Alistair; Underhill, Michael; Zaph, Colby (Monash University) The BTB/POZ family of transcriptional repressors, which includes Plzf, Thpok, and Bcl6, plays a critical role in the development of both the innate and adaptive immune systems. Hypermethylated in cancer 1 (Hic1) is a member of this family and has been identified as a target gene of retinoic acid (RA) signaling. RA, a vitamin A metabolite, has also been shown to be critically required for intestinal immunity. RA is required to license lymphocytes for intestinal homing, maintain innate lymphoid cell (ILC) homeostasis, as well as to promote CD4+ T helper (TH) cell activation. Here we demonstrate that Hic1 is critically required for intestinal ILC and TH cell function. In vivo, RA is highly enriched in the gut, concurrently; Hic1 expression within the hematopoietic compartment is restricted to the small intestine. Strikingly, mice fed a vitamin A deficient diet exhibit notably diminished expression of Hic1 in intestinal immune cells, demonstrating Hic1’s dependence on RA signaling. Mice with a specific deletion of Hic1 in hematopoietic cells (Hic1Vav mice), demonstrate a significantly altered intestinal immune homeostasis with reduced frequencies of ILCs and T cells at steady state. Following infection with Citrobacter rodentium, Hic1Vav mice displayed exacerbated pathology, associated with impaired Rorγt+ ILC responses. Intriguingly, Hic1Vav mice were also unable to develop protective TH2 immunity in response to Trichuris muris infection; conversely Hic1Vav mice developed a TH1 response and maintained a chronic parasite burden. Taken together, these results establish a hematopoietic-‐intrinsic role for Hic1 in maintaining intestinal immune homeostasis as well as being critically required for protection from intestinal helminth and bacterial infections. A role for dectin-‐1 in the CNS during chronic Toxoplasma gondii infection Worth, Danielle (University of California, Riverside, Riverside, CA, USA); Nance, J. Philip; Jang, Jessica; David, Clement; Nair, Meera; Wilson, Emma The latent stage of T.gondii infection is characterized by the formation of tissue cysts within the CNS. These tissue cysts are formed by manipulation of the parasitophorous vacuole, and chitin is a structural component of the cyst wall. Our lab has shown that Toxoplasma cysts induce an M2 phenotype in macrophages in a contact-‐dependent manner. Upon contact, these macrophages actively secrete the mammalian chitinase, AMCase, to break down the cyst wall. We have previously demonstrated that AMCase production by macrophages is essential for control of cyst burden in the brain during T. gondii infection. The molecular interactions involved in macrophage-‐cyst recognition, and the signaling pathway for AMCase production are as yet unknown. Although chitin recognition has been poorly defined, studies suggest dectin-‐1 may be a receptor. Here, we present data on dectin-‐1 during T. gondii infection. Dectin-‐ 1 is upregulated in the brain in response to infection and is expressed on arginase-‐ 1+, M2 macrophages. Consistent with its expression on M2 macrophages, dectin-‐1 upregulation is STAT6 dependent, and dectin-‐1 is not required to control parasite burden or induce an inflammatory immune response in vivo. Contrary to our hypothesis, dectin-‐1 deficient mice do not exhibit differences in cyst numbers in the brain or AMCase activity ex vivo. Thus, this data suggests that dectin-‐1 plays no significant role during Toxoplasma infection. Role of microglia during cerebral malaria Capuccini, Barbara (The Francis Crick Institute, Mill Hill Laboratory, London, GBR); Lin, Jingwen; Sodenkamp, Jan; Keller-‐Mclauglin; Kushinga, Garikai; Langhorne, Jean Cerebral malaria (CM) is a major life-‐threatening complication of Plasmodium falciparum infection. The pathogenic mechanisms underlying the occurrence of cerebral lesions are still not completely understood but, clearly, cerebral complications may result from concomitant microvessel obstruction and inflammation. Microglia represent the resident macrophages of the central nervous system. These cells exhibit a resting phenotype in a healthy brain, but following challenge or injury they can rapidly convert to an activated state. Up to now, there is little information on the role of microglial cells and their contribution to CM. Therefore, there is a need to unravel the mechanisms leading to microglial activation and whether this response influences the morbidity and mortality during CM. The major goal of this project is to define the role of these peculiar cells during the inflammatory response causing the fatal outcome of CM. To understand these processes better, we exploit the murine model of experimental cerebral malaria (ECM), in which C57BL/6 mice have been infected with Plasmodium berghei ANKA. We studied the gene expression profile of microglia cells from mice that showed ECM symptoms compared to the same cells from non-‐infected mice. Network analysis identified type I IFNs as possible regulatory molecules. Type I IFNs could not be detected in microglia themselves. We hypothesise that signalling through the IFNAB Receptor expressed on microglia results in up-‐ regulation of co-‐stimulatory molecules and chemokine production, and may regulate recruitment of other immune cells, thus contributing to the inflammatory response associated with ECM. The Paneth-‐cell specific role of autophagy in Toxoplasma gondii-‐induced intestinal inflammation Burger, Elise (UT Southwestern, USA); Yarovinsky, Felix The protozoan parasite Toxoplasma gondii triggers severe immunopathology characterized by intestinal inflammation, a loss of Paneth cells and change in the microbiota. Paneth cells are the predominant intestinal secretory epithelial cells which reside at the base of the crypt and release antimicrobial peptides to regulate the intestinal microbiota. IFN-‐gamma triggered by T. gondii infection is a major inducer of Paneth cell death. However, mechanisms of T. gondii-‐induced IFN-‐gamma dependent Paneth cell death are currently unknown. We have observed that under steady state conditions, Paneth cells have the highest level of autophagy when compared to other intestinal cells. To investigate if autophagy plays a role in Paneth cell loss during T. gondii infection, we investigated autophagy reporter mice as well as the animals that are deficient in the critical autophagy component Atg5. We have found that upon challenge with T. gondii, the autophagy-‐deficient mice display enhanced susceptibility to intestinal inflammation characterized by complete destruction of the intestinal crypt. Our results revealed that contrary to our initial hypothesis, autophagy plays a protective role in regulation of Paneth cell functions during acute responses to the parasite. We are currently further investigating the protective mechanism of autophagy in Paneth cells during IFN-‐gamma-‐mediated intestinal inflammation. Once the mechanism is established, we aim to develop approaches for the prevention of T. gondii triggered Paneth cell death, intestinal dysbiosis and pathology in vivo. Immunopathogenesis of visceral leishmaniasis -‐ New insights into infection-‐induced haematological disorders Preham, Olivier (University of York, York, GBR); Pinto, Ana; Brown, Najmeeyah; Chan, Samantha; Kaye, Paul Human visceral leishmaniasis (VL) is a major neglected disease of poverty. We have used a mouse model to understand why patients with this disease present haematological disorders such as anemia and thrombocytopenia. We have shown that medullary erythropoiesis is significantly reduced in B6 mice chronically infected with Leishmania donovani. Depletion of erythroblasts from the bone marrow was associated with a down-‐regulation of the chemokine (C-‐X-‐C motif) ligand 12, (CXCL12) a key cytokine for the retention of hematopoietic progenitors in the bone marrow, as well as an up-‐regulation G-‐CSF, a potent inhibitor of medullary CXCL12 production. Loss of bone marrow function also correlated with a disruption of the splenic architecture due to extramedullary haematopoiesis and the resulting splenomegaly, a common feature of human disease. Observed changes in erythropoiesis were T-‐cell dependent, as erythropoiesis was unaltered in immune-‐deficient RAG2 KO mice and adoptive transfer of purified CD4 T cells was sufficient to induce these changes in erythropoiesis in infected RAG2 KO mice. To examine a causal relationship between splenomegaly and anemia, we examined infection in splenectomized mice. Splenectomized B6 mice infected with L. donovani were equally deficient for medullary erythropoiesis as sham-‐operated controls. However, these mice had normal platelet counts, suggesting that different mechanisms underlie thrombocytopenia and anemia and that the spleen is involved, directly or indirectly in regulating platelet count during infection. In summary, our data provide new understanding of the links between anaemia, thrombocytopenia and splenomegaly, major symptoms affecting patients with visceral leishmaniasis. Our research may lead to new treatments aimed at restoring bone marrow function, as a novel and alternative approach to directly targeting the parasite itself Trefoil factor 2 mediates macrophage-‐dependent lung repair following hookworm infection Hung, Li-‐Yin (UCSF, San Francisco, CA, USA); Sen, Debasish; Oniskey, Taylor; Nieves, Wildaliz; Kim, Charles; Krummel, Matthew; Herbert, De'Broski Coordination between epithelia and myeloid subsets is important for tissue repair, particularly in the context of hookworm-‐induced lung damage, but mechanistic evidence for macrophage-‐driven lung repair is lacking. Our study demonstrates that pulmonary macrophages accelerate tissue regeneration through a pathway independent of Type 2 cytokines (interleukin 4/13), and T, B or ILC populations. Macrophages rely upon Trefoil factor 2 (TFF2), a mucosal cytokine considered solely epithelial-‐derived, to repair lungs damaged by Nippostrongylus brasiliensis. Fluorescent reporter mice reveal that alveolar macrophages produce TFF2 under baseline and damage-‐induced conditions. CD11c-‐specific Tff2 deletion impairs tissue regeneration following hemorrhagic lung injury. Macrophage-‐epithelia co-‐culture experiments indicate that macrophage-‐derived TFF2 directly accelerates epithelial proliferation, trans-‐ epithelial resistance, and barrier function. Combined, this demonstrates TFF2 as a regenerative cytokine released from macrophages to facilitate repair of infectious or non-‐infectious lung damage. A highly attenuated clone of Trypanosoma cruzi lacks the SAPA domain of trans-‐sialidase protein Junqueira, Caroline (Centro de Pesquisas Rene Rachou/ FIOCRUZ, Belo Horizonte, BRA); Neto, Rondon (ICB/ UFMG); Zuccherato, Luciana (ICB/ UFMG); Bartholomeu, Daniella (ICB/ UFMG); Vasconcelos, Ana Tereza (Laboratório Nacional de Computação); El-‐Sayad, Najib (University of Maryland); Burleigh, Barbara (Harvard University); Teixeira, Santuza (ICB/ UFMG); Gazzinelli, Ricardo (CpQRR/ FIOCRUZ) CL-‐14 is a highly attenuated clone of T. cruzi that was isolated from the CL strain, as well as its parental clone CL Brener. CL-‐14 was characterized as a non-‐pathogenic T. cruzi, which is not able to induce parasitemia and tissue parasitism. This clone was originally described as a vaccine clone, since it leads to protection against a challenge with the virulent clone CL Brener. Recent studies from our research group used the attenuated clone as a vaccine delivery vector expressing the cancer antigen NY-‐ESO-‐1 to be employed as an antitumor vaccine. Regarding the promising results obtained with the vaccine vector, we decided to search for the molecular basis by which the CL-‐14 clone is less pathogenic than the parental clone CL Brener and prove its biosafety for vaccination protocols. Aiming to achieve this goal was fully sequenced the genome of CL-‐14, as well as its transcriptome. Based on the comparative analysis of the multigenic families belonging to the T. cruzi parasite, we found that the CL-‐14, when compared with the CL Brener, lacks the SAPA domain of the trans-‐sialidase protein. The data was confirmed by molecular biology and immunodetection assays. This protein is known to play a key role in invasion process, even as one of main immunodominant antigens in T. cruzi, leading to immunomodulation and parasite escape. The absence of SAPA domain can be correlated with the attenuated phenotype of this clone and it is now being evaluated by the reconstitution of TS SAPA into the CL-‐14. Financial Support: INCTV, CNPq, Fapemig POSTERS Immune Regulation 1. IFN-‐(g) prevents adenosine receptor (A2bR) upregulation to sustain the macrophage activation response and promote the killing of L. major Hamidzadeh, Kajal (University of Maryland, College Park, MD, USA); Cohen, Heather; Mosser, David We recently demonstrated that TLR-‐stimulated macrophages modulate their activation state by increasing the synthesis and secretion of ATP. This endogenous production of ATP is paradoxically immunosuppressive due to its rapid catabolism into adenosine by CD39 and CD73 on the macrophage surface. Macrophages lacking this purinergic regulatory mechanism are unable to transition to a regulatory state and consequently continue to produce inflammatory cytokines. The priming of macrophages with IFN-‐γ prior to TLR stimulation results in enhanced and prolonged inflammatory cytokine production. Here, we demonstrate that following TLR stimulation, macrophages up regulate the adenosine 2b receptor (A2bR) to enhance their sensitivity to immunosuppressive extracellular adenosine. This up regulation of A2bR leads to the induction of a regulatory macrophage phenotype and the down regulation of inflammation. IFN-‐γ priming of macrophages selectively prevents the induction of the A2bR in macrophages to mitigate sensitivity to adenosine and prevent this regulatory transition. IFN-‐γ-‐mediated A2bR blockade leads to a prolonged production of TNFa and IL-‐12 in response to TLR ligation. The pharmacological inhibition or the genetic deletion of the A2bR results in a hyper-‐ inflammatory response to TLR ligation, similar to IFN-‐γ treatment of macrophages. Conversely, the overexpression of A2bR on macrophages blunts the IFN-‐γ effects and promotes the development of regulatory macrophages. Thus, we propose a novel mechanism whereby IFN-‐γ contributes to host defense, by desensitizing macrophages to the immunoregulatory effects of adenosine. This mechanism overcomes the transient nature of TLR activation, and prolongs the anti-‐microbial state of the classically activated macrophages to allow them to kill intracellular pathogens. 2. Phlebotomus duboscqi salivary proteins modulate human Langerhans cell and monocyte function Huda, Naureen (National Institutes of Health, Bethesda, MD, USA); Sangare, Moussa Brema (Mali International Center for Excellence in Research); Abdeladhim, Maha; Doumbia, Seydou; Coulibaly, Yaya Ibrahim; Nutman, Thomas B.; Valenzuela, Jesus; Oliveira, Luiz; Kamhawi, Shaden; Semnani, Roshanak Tolouei Leishmaniasis is a vector-‐borne disease transmitted by bites of infected phlebotomine sand flies. Transmission occurs during bloodfeeding when female sand flies inject both saliva and parasites into the host. Previous studies in animal models showed that sand fly saliva enhances infectivity of Leishmania parasites in naïve hosts, while saliva-‐specific adaptive immune responses protect against leishmaniasis. However, little is known about the innate immune response to saliva in humans. Here we investigated the effects of sand fly salivary glands (SG) of Phlebotomus duboscqi (PD) on antigen-‐presenting cells, particularly monocytes (Mφ) and Langerhans cells (LCs), from saliva-‐unexposed healthy volunteers. Mφ were isolated and cultured in vitro either in media alone or with GM-‐CSF, IL-‐4, and TGF-‐β to generate LCs. Both cell types were stimulated with SG, lipopolysaccharide (LPS), Brugia malayi microfilariae (mf) as an unrelated helminth parasite, or Leishmania antigen. After 30 minutes to 48 hours incubation, the mRNA expression of genes involved in Type 1, Type 2, and regulatory responses was measured, as were cytokines (IL-‐6, IL-‐10) and chemokines (CCL2, CCL4, CCL22, CXCL10) associated with these responses. While stimulation with LPS, mf, or Leishmania antigen largely upregulated the cytokine production in Mφ, SG generally had no effect on cytokine production. Interestingly, exposure of Mφ to SG resulted in significant downregulation of a Type 2-‐associated chemokine CCL22 (4 hours, p<0.05) and upregulation of a Type 1-‐associated chemokine CCL4 (24 hours, p<0.05). Induction of CCL4 by SG may be partially responsible for the persistent neutrophilic inflammation observed following sand fly bites. Further, in LCs while mRNA expression was not altered, the production of Type 1-‐associated chemokines CXCL10 and CCL2 (p=0.015) was significantly diminished after 48 hours. These results suggest that sand fly saliva may have a differential effect on human LCs and Mφ, primarily modulating chemokine production and cellular recruitment to the bite site. 3 The phenotype and behavior of regulatory T cells in the CNS during chronic T. gondii infection O'Brien, Carleigh (University of Virginia, USA); Hayes, Nik; Harris, Tajie Regulatory T cells (Tregs) have potent suppressive capacity capable of controlling effector T cell responses and limiting immune-‐mediated pathology. While the importance of Tregs during the immune response to multiple infections has been demonstrated, the phenotype and specific mechanisms of suppression in the inflamed CNS during infection have not been completely elucidated. During chronic infection with Toxoplasma gondii, effector T cells are required to limit parasite replication in the CNS. Tregs are also found in the CNS during this chronic infection, primarily in the perivascular spaces and meninges, and are predicted to limit excessive immunopathology. To begin understanding the role of these Tregs and their mechanisms of immune suppression in the CNS, we have used several techniques to analyze their phenotype. We have also been able to isolate Tregs from the meninges of both naïve and infected mice and analyze them by flow cytometry. Tregs isolated from the CNS of infected mice express IL-‐10 and have an activated, Th1-‐like phenotype, shown by expression of the transcription factor T-‐bet and CXCR3 and upregulation of the markers CD44, ICOS, CD103, CD11a, and ICAM. Using MHC Class II tetramers specific for T. gondii, we find that Tregs in the CNS are not specific for this T. gondii antigen. Using intravital multiphoton imaging, we are able to analyze their behavior. These Tregs form long-‐lasting contacts with CD11c+ APCs in the meninges, yet remain mobile. Overall, understanding the phenotype of CNS-‐resident Tregs, and how this differs from other peripheral and lymphoid tissues, is an important first step to understanding their role and specific mechanisms of immune suppression during chronic CNS infection. 4 The B Cell Adaptor Molecule Bam32 is Critically Important for Optimal Antibody Response and Resistance to Trypanosoma congolense Infection in Mice Onyilagha, Chukwunonso (University of Manitoba, Winnipeg, MN, CAN); Jia, Ping; Jayachandran, Nipun, Hou, Sen; Okwor, Ifeoma; Kuriakose, Shiby; Marshall, Aaron; Uzonna, Jude Background: Bam32, a 32 kDa adaptor molecule, plays important role in B cell receptor signalling, T cell receptor signalling and antibody affinity maturation in germinal centres. Since antibodies against trypanosome variant surface glycoproteins (VSG) are critically important for control of parasitemia, we hypothesized that Bam32 deficient (Bam32-‐/-‐) mice would be susceptible to T. congolense infection. Methodology/Principal Findings: We found that T. congolense-‐infected Bam32-‐/-‐ mice successfully control the first wave of parasitemia but then fail to control subsequent waves and ultimately succumb to their infection unlike wild type (WT) C57BL6 mice which are relatively resistant. Although infected Bam32-‐/-‐ mice had significantly higher hepatomegaly and splenomegaly, their serum AST and ALT levels were not different, suggesting that increased liver pathology may not be responsible for the increased susceptibility of Bam32-‐/-‐ mice to T. congolense. Using direct ex vivo flow cytometry and ELISA, we show that CD4+ T cells from infected Bam32-‐/-‐ mice produced significantly increased amounts of disease-‐ exacerbating proinflammatory cytokines (including IFN-‐g, TNF-‐a and IL-‐6). However, the percentages of regulatory T cells and IL-‐10-‐producing CD4+ cells were similar in infected WT and Bam32-‐/-‐ mice. While serum levels of parasite-‐specific IgM antibodies were normal, the levels of parasite-‐specific IgG, (particularly IgG1 and IgG2a) were significantly lower in Bam32-‐/-‐ mice throughout infection. This was associated with impaired germinal centre response in Bam32-‐/-‐ mice despite increased numbers of T follicular helper (Tfh) cells. Adoptive transfer studies indicate that intrinsic B cell defect was responsible for the enhanced susceptibility of Bam32-‐/-‐ mice to T. congolense infection. Conclusions/Significance: Collectively, our data show that Bam32 is important for optimal anti-‐trypanosome IgG antibody response and suppression of disease-‐promoting proinflammatory cytokines and its deficiency leads to inability to control T. congolense infection in mice. 5 Adaptive and innate immune regulation of allergic airway inflammation by the intestinal helminth Trichuris muris Chenery, Alistair (University of British Columbia, Vancouver, BC, CAN); Antignano, Frann; Burrows, Kyle; Scheer, Sebastian; Perona-‐Wright, Georgia; Zaph, Colby (Monash University) Background: Allergic diseases such as asthma are becoming increasingly prevalent globally. Environmental factors are known to play a major role in determining allergic disease risk. Intestinal helminths are well-‐known to immunoregulate their hosts and there is evidence that they can directly protect against allergic airway inflammation (AAI). However, mucosal immune crosstalk between the intestine and the lungs is a poorly-‐defined concept. The objective of this study is to examine the gut-‐lung immunological crosstalk during an intestinally-‐restricted helminth infection in the context of lung disease. We hypothesized that Th1-‐mediated intestinal helminth infections would protect against Th2-‐ dependent AAI through this gut-‐lung immune axis. Methods: Mice were chronically infected with Trichuris muris and then challenged intranasally with protease allergens to induce AAI. Lung disease was assessed by bronchoalveolar lavage fluid analysis by flow cytometry. Lung QRT-‐PCR, ELISA, and histopathology were also performed. Results: T. muris-‐infected mice were protected from AAI. Infection significantly blocked airway cell infiltration, eosinophilia, and mucus hyperproduction. Infection also caused a mixed immune response in the lung characterized by increased Th1 IFN-‐γ and myeloid IL-‐10 that blocked Th2-‐dependent AAI. T. muris exerted effects on both innate cell and adaptive CD4+ T cell populations indicating that there are multiple immune pathways involved in which T. muris can protect against allergic lung disease. Conclusions: An intestinally-‐restricted helminth can exert protective immune effects that can suppress the development of AAI in the lung. An intestinal Th1 response regulates AAI through convergent innate and adaptive immunomodulation. These results are relevant for both studying host-‐parasite relationships as well as for defining novel immune pathways that may have therapeutic potential for the treatment of AAI. 6 Identification and characterization of larval Trichuris suis secretory proteins Balster, Katja (Institute of Immunology, Department of Veterinary Medicine, Freie Universität Berlin, USA); Ebner, Friederike, Hartmann, Susanne The intestinal nematode Trichuris suis (T. suis) naturally infects the caecum and colon of pigs. Due to the immunomodulatory potential the administration of T. suis ova (TSO) was used as a novel therapy for immune mediated diseases in clinical trials. Previous studies of our group demonstrate the immunomodulatory potential of T. suis larval excretory/secretory (E/S) products in murine in vitro assays. Among others, dendritic cells pre-‐treated with T. suis E/S showed diminished upregulation of MHC class II and the co-‐stimulatory molecule CD86. Furthermore, we demonstrated a massive immunomodulatory potential of T. suis E/S products in vivo in a murine model of allergic hyperractivity. In this study we aim to identify single immunomodulatory molecules in T. suis larval E/S in order to develop a novel therapy based on the immunomodulatory capacity of the T. suis E/S products. Via mass spectrometry we identified ten signal-‐peptide-‐containing proteins of T. suis larval E/S. Six of the identified E/S proteins showed homologies to known proteins like chitinase, venom allergen-‐like protein or trypsin. Subsequently, these proteins were cloned, eukaryotically expressed in Leishmania and purified via affinity chromatography. Analyses of the immunmodulatory function of the recombinant expressed proteins are ongoing. Hence, T. suis larval E/S proteins exhibit significant immunomodulatory potential and ten novel proteins in the E/S products have been identified. These proteins were eukaryotically expressed and will now be characterized for their immunomodulatory potential. Innate Immune Cells 7 The prostaglandin D2 receptor CRTH2 regulates accumulation of group 2 innate lymphoid cells in the inflamed lung Tait Wojno, Elia (Cornell University, Ithaca, NY, USA) Group 2 innate lymphoid cells (ILC2s) promote type 2 immune responses following helminth infection and during allergic inflammation, but the pathways that control ILC2 migration into inflamed tissues remain poorly understood. Here, we show that the prostaglandin D2 (PGD2) receptor CRTH2 (chemoattractant receptor-‐homologous molecule expressed on Th2 cells) regulates ILC2 accumulation in the lung in vivo. The frequency of ILC2s that expressed CRTH2 was significantly higher in healthy human and murine peripheral blood than in the lung, suggesting that regulation of CRTH2 expression is associated with ILC2 accumulation in the lung. Consistent with this, murine ILC2s accumulated in the lung in response to PGD2. Further, using a model of pulmonary inflammation induced following infection with the hookworm Nippostrongylus brasiliensis, CRTH2-‐deficient mice exhibited reduced ILC2 accumulation and type 2 inflammation in the lung compared to wild-‐type mice. Critically, adoptive transfer of CRTH2-‐sufficient ILC2s restored hookworm-‐induced type 2 pulmonary inflammation and pathology in CRTH2-‐deficient mice. Together, these data suggest that the PGD2-‐CRTH2 pathway regulates ILC2 accumulation and type 2 inflammation in the lung in vivo. 8 Vitamin-‐A regulates eosinophil tissue accumulation during homeostasis and helminth-‐induced inflammation. Sutherland, Duncan (Ecole Polytechnique Fédérale de Lausanne, USA); Esser-‐von Bieren, Julia; Kulagin, Manuel; Fagarasan, Sidonia (RIKEN); Harris, Nicola Eosinophils are tissue dwelling cells with multiple roles in homeostasis and disease. They are produced in the bone marrow and accumulate in tissues in response to type 2 inflammation. However they also accumulate in the upper small intestine during homeostasis, prompting us to investigate their interaction with dietary metabolites. We observed that chronic vitamin-‐A deficiency (VAD) or pharmacological blockade of RA receptor (RAR) signaling abolishes homeostatic eosinophil accumulation in the small intestine and adipose tissue, and abrogated chemokine-‐receptor-‐3 (CCR3) expression by eosinophils in the bone marrow. Addition of increasing concentrations of the Vitamin A metabolite, retinoic acid (RA), to bone marrow-‐derived eosinophils in vitro resulted in a distinct acceleration of eosinophil maturation kinetics and the activation of distinct gene clusters. Helminth-‐induced eosinophilia also required RAR signaling and these cells functioned to promote the removal of trapped helminth larvae through a process involving the expression of oxidative enzymes. These data reveal a novel role for dietary vitamin A in regulating the homeostatic and pathogen-‐induced tissue accumulation of eosinophils. 9 Transcriptional Profiling of In Vitro Activated Primary Mouse Astrocytes Dawson, Lucas A. (University of Pennsylvania, Philadelphia, PA, USA); Hidano, Shinya; Beiting, Daniel; Randall, Louise (University of Melbourne); Hunter, Christopher Astrocytes are the predominant subtype of glial cell in the brain where they provide support and protection for neurons, as well as maintain the blood brain barrier. There are a limited number of bacterial or eukaryotic pathogens that can replicate in these cells and Toxoplasma gondii is perhaps the most common clinically relevant organism that interacts with these glia. In murine models of Toxoplasmic encephalitis astrocyte activation and proliferation are prominent, and astrocytes produce chemokines that can influence the recruitment of T cells and dendritic cells as well as microglial cell activation. In an attempt to identify the transcriptional changes of activated astrocytes we utilized a microarray approach to compare the impact of interferon gamma, interferon alpha, and T. gondii infection on mouse primary astrocytes. Analysis of this data revealed subsets of differentially expressed genes specific to either interferon gamma or interferon alpha, as well as a subset of differentially expressed genes regulated by both interferons. In contrast, far fewer genes were differentially regulated during T. gondii infection with almost no overlap to those genes regulated by interferon stimulation. Gene set enrichment analysis reveals that both interferons induce canonical pathways of immune responses, while T. gondii infection induces DNA synthesis pathways. Future studies will endeavor to determine if the genes identified in this study are differentially regulated during chronic T. gondii infection, and their roles in astrocyte function. 10 Eosinophil-‐derived IL-‐4 promotes nematode growth in an innate context Huang, Lu (Cornell University, Ithaca, NY, USA); Beiting, Daniel (University of Pennsylvania, USA); Gebreselassie, Nebiat; Gagliardo, Lucille; Ruyechan, Maura; Lee, Nancy; Lee, James; Appleton, Judith It has become increasingly clear in recent years that the function for eosinophils in health and disease is highly varied. Recent studies have revealed novel functions for eosinophils in metabolism and tissue regeneration. Such mechanisms have strong potential to be relevant in worm infections in which parasites rely on host metabolism for nutrients that support worm growth or reproduction. The aim of this study was to investigate the mechanism underlying the observation that eosinophils promote growth of Trichinella spiralis larvae in skeletal muscle. Our results indicate that normal larval growth requires IL-‐4 and STAT6 signaling that are intrinsic to eosinophils. Support of growth by eosinophil-‐ derived IL-‐4 occurs independently of adaptive immunity. Host gene expression in skeletal muscle was compatible with a regenerative response to invasion by newborn larvae and a shift in the source of energy in infected tissue. Furthermore, the presence of eosinophils suppressed local inflammation while also influencing nutrient homeostasis in muscle. Redistribution of glucose transporter 4 (GLUT4) and phosphorylation of Akt were observed in nurse cells, consistent with enhancement of glucose uptake and storage of glycogen by larvae that is known to occur. The results suggest that eosinophil-‐derived IL-‐ 4 may promote larval growth by altering nutrient metabolism in infected muscle. Our findings document a novel interaction between worms and host, in which worms have evolved a strategy to co-‐opt an innate host cell response in a way that facilitates their growth. Host Protection 11 Immunization study using a combination of B-‐ and T-‐cell epitopes in the a1,3-‐galactosy-‐ ltransferase-‐knockout murine model of Chagas disease Portillo, Susana (The University of Texas at El Paso, USA); Schocker; Bottazzi, Maria E. (Baylor College of Medicine National School of Tropical Medicine, USA); Jones, Kathryn M.; Zhan, Bin; Hammond, Molly; Hotez, Peter J.; Michael, Katja Chagas disease affects 8-‐10 million in Latin America, resulting in approximately 10,000-‐12,000 deaths every year. Lately, it has also become a public health concern for non-‐endemic countries, like the United States, Spain, Japan and Australia. In the U.S., it is one of the top five neglected parasitic infections with an estimated 300,000 people chronically infected, asymptomatic individuals. The available drugs are partially effective and toxic, and there are no current vaccines to prevent or treat this disease. In this study, we evaluated a CD8 T cell epitope (i.e., recombinant protein Tc24, recTc24) and a B cell glycan epitope (i.e., synthetic neoglycoprotein Galα(1,3)Galβ(1,4)LacNAc-‐BSA, Galα3LN-‐BSA), alone or combined, as potential experimental vaccine candidates, using an α1,3-‐galactosyltransferase gene-‐ knockout (α1,3GalT-‐KO) mice. This murine model lacks terminal Galα1,3Gal epitopes, which are not expressed in humans and Old World primates, but abundantly expressed on all other mammals and T. cruzi infective trypomastigote stage. Our preliminary results demonstrated a very strong antigen-‐specific antibody response when animals were immunized with either antigen alone or combined. Cytokine and immunoglobulin profiling, trypanolytic assay, and survival experiments are underway and will be discussed. We anticipate that the combination of a CD8 T cell epitope together with a B cell epitope will result in a more effective immunological response and protection against experimental T. cruzi infection. 12 Vaccine-‐Induced T cell Responses to Toxoplasma gondii are Primed in the Omentum Christian, David (University of Pennsylvania, USA); Harms Pritchard, Gretchen; Wang, Qiuling (Washington University in St. Louis, USA); Koshy, Anita (University of Arizona, USA); Park, Sunmin, Virgin IV, Herbert; Sibley, L. David; Hunter, Christopher Long-‐term resistance to infection with the intracellular parasite T. gondii is dependent on CD8+ T cells. While vaccine strategies to generate this protective T cell response have been elusive, immunization with the replication-‐deficient strain of T. gondii (cpsII strain) has been shown to drive a parasite-‐specific CD8+ T cell response that is dependent on CD4+ T cells. Previous work has demonstrated that cpsII T. gondii must invade host cells rather than be phagocytosed to prime both CD4+ and CD8+ T cells. The focus of this work is to elucidate what APCs drive this T cell priming and what molecular mechanisms are responsible for processing and presenting parasite antigens. By combining a fluorescent, Cre-‐secreting cpsII strain of T. gondii with the Ai6 floxed reporter mouse strain, cells infected and/or injected by the parasite were tracked in different tissues in vivo. After intraperitoneal injection, the cells that primarily interacted with the parasite were CD11b+ DCs and large peritoneal macrophages that were found later in the omentum, suggesting that the omentum could serve as the primary site for T cell priming. Indeed, tracking of initial T cell priming events by CD69 expression revealed that T cells begin to upregulate CD69 in the peritoneum and omentum but not the draining lymph nodes or spleen by 2 days after immunization. In mice deficient in DC populations (BATF3 and FLT3L KO mice), were defective in T cell CD69 expression at these early time points, suggesting that DCs are critical for T cell priming. Further work will examine the role of autophagy as a mechanism for presentation of parasite antigens by different APC populations. 13 Production and testing of the Lutzomyia intermedia sand fly salivary recombinant proteins as markers of risk to cutaneous leishmaniasis disease Carvalho, Augusto M P (NIH, Rocvkville, MD, USA); Valenzuela, JG; Costa-‐Guimaraes, A; de Oliveira, CI; Oliveira, F Currently there are no human vaccines against leishmaniasis. In rodents, cellular immunity to salivary proteins of sand fly vectors is associated to protection against leishmaniasis making them worthy targets for further exploration as vaccines. Anti-‐saliva human responses from residents of cutaneous leishmaniasis (CL) endemic areas have shown that sand fly salivary proteins are immunogenic, eliciting specific-‐anti-‐saliva antibodies and cellular immune responses. Importantly, the presence of anti-‐sand fly saliva antibodies correlates with an two-‐fold increase risk of CL. Stimulation of PBMC from endemic area individuals with whole sand fly saliva induces a mixed type of response with participation of both Th1 and Th2 cytokines. No data on the correlation of anti-‐saliva cellular immune responses and CL disease outcome is available to date. Due to the complexity of antigens present in the whole sand fly saliva, interpretation of the meaning of the anti-‐saliva cellular immune responses to CL disease would become cumbersome. Here, we propose to generate the most abundant recombinant proteins from the sand fly vector Lutzomyia intermedia and test them for the ability to induce either a Th1 or Th2 immune response. We will also screen the sand fly salivary recombinant proteins against the sera of the endemic area individuals for markers of exposure and or risk of CL disease. Moreover, by studying CL patients versus matched health controls from the same endemic area we hope to determine if immune responses to particular sand fly salivary proteins that can be correlated to protection or enhancement of CL disease outcome. 14 IL-‐1a is required for control of the Trypanosoma cruzi infection SILVA, GRACE (University of Massachusetts, Worcester); Ketelut-‐Carneiro, Natália (University of São Paulo); Zamboni, Dario; Gazzinelli, Ricardo (University of Minas Gerais); Golenbock, Douglas; Silva, João Santana Trypanosoma cruzi is the parasite causative of Chagas disease that causes robust immune responses in immunocompetent hosts, resulting in the recruitment of cells to the heart. Recently, we demonstrated that IL-‐1R is required to the control of T. cruzi infection. Notably, both IL-‐1α and IL-‐1β signal through IL-‐1 receptor, then our aim is understand the mechanisms that regulate the processing of IL-‐1α and it role during experimental T. cruzi infection. Firstly, we verify whether this parasite is able to trigger IL-‐1α in the bone marrow macrophages (BMMs) and in the heart at different times after infection. Interesting, we observed that only live T. cruzi, in the presence of IFN-‐γ, triggers robust amount of IL-‐1α by BMMs at 48 hs pi. Additionally, we found a significant amount of IL-‐1α in the heart from infected mice as compared with not infected animals. To test the involvement of IL-‐1α in the Killing of Trypomastigotes we infected WT, IL-‐1R-‐/-‐ and IL-‐1α-‐/-‐ BMMs primed or not with 10 ng/mL of IFN-‐γ to quantify the growth parasites and Nitric oxide (NO) production. Strikingly, IL-‐1R-‐/-‐ and IL-‐1α-‐/-‐ BMMs, primed with IFN-‐γ, failed to eliminate efficiently the T. cruzi and to produce NO when compared with WT cells. To analyze the resistance of these animals to T. cruzi infection we infected WT, IL-‐1R-‐/-‐ and IL-‐1α-‐/-‐ mice with 103 Trypomastigotes to determine the parasitemia and mortality. We observed that IL-‐1R-‐/-‐ and IL-‐1α-‐/-‐ presented high amount of parasites and succumbed infection precocity, while the WT mice were resistant. Altogether these results reveal, by the first time, that IL-‐1α is essential to control the T. cruzi infection in vitro and in vivo. Financial Support: FAPESP 15 A Newly Identified Role for Interleukin-‐22 in Bladder Immunity Honeycutt, Jared (Stanford University, Stanford, CA, USA); Hammam, Olfat (Theodor Bilharz Institute, Cairo, Egypt); Hsieh, Yi-‐Ju (Biomedical Research Institute, USA); Hsieh, Michael H. (Biomedical Research Institute; George Washington University; Children's National, USA) In contrast to the gut and lung, it is relatively unclear what inflammatory pathways in the urinary tract contribute to host protection and pathology during parasitic or bacterial infection. Recent reports identify interleukin-‐22 (IL-‐22) as a key cytokine supporting epithelial immunity; additionally, its improper regulation may contribute to pathologies such as fibrosis and cancer. We hypothesized that IL-‐22 may have an important role in bladder immunity during Schistosoma haematobium and E.coli infections. In wild-‐type (wt) mice, IL-‐22 receptor α1 expression was detectable in the urothelium by immune-‐ fluorescence and qPCR. Levels of IL-‐22 and its soluble binding protein, IL-‐22BP, were increased in the bladders of wt mice after bladder wall injection of S. haematobium eggs. IL-‐22-‐null (KO) and wild type (wt) female mice underwent bladder wall injection with S. haematobium eggs. Comparing whole bladder RNA by microarray, IL-‐22-‐KO mice showed differential expression of genes related to glutathione transferase activity, transferase activity, and epithelial cell development. These changes in the bladder suggest that, as in the gut, IL22 is important for replenishing the epithelial lining during infection-‐related injury. IL-‐22-‐KO and wt littermate mice were transurethrally infected with E.coli UTI89. IL-‐22-‐KO mice had lower bacterial counts in their urine, bladder, and kidneys. Giving stabilized IL-‐22 cytokine (IL-‐22-‐Fc) to UTI89-‐infected mice led to higher kidney bacterial counts and increased morbidity. Our data suggest that IL-‐22 is indeed important in urinary tract immunity and may interfere with clearance of bacteria from the urinary tract, potentially through its role in maintenance of mature urothelium. Malaria Immunobiology 16 CD4+ T Cell Response Correlates with Naturally Acquired Antibodies against Plasmodium vivax Tryptophan-‐Rich Antigens Zeeshan, Mohammad (All India Institute of Medical Sciences,New Delhi); Tyagi, Kriti; Sharma, Yagya D Tryptophan-‐rich proteins play important biological functions for the Plasmodium parasite. Plasmodium vivax contains remarkably larger number of such proteins belonging to the “Pv-‐fam-‐a” family that need to be characterized. Earlier, we reported the presence of memory T cells and naturally acquired antibodies against 15 of these proteins in P. vivax malaria-‐exposed individuals (M. Zeeshan et al., J. Infect. Dis. 207:175–185, 2013). Here, we sought to characterize and ascertain the cross talk between effector responses of T and B cells in malarial patients against all Pv-‐fam-‐a family proteins. Therefore, we expressed the remaining 21 of these proteins in E. coli and studied the humoral and cellular immune responses based on the same parameters used in our previous study. Naturally acquired IgG antibodies were detected against all 21 antigens in P. vivax patient sera (37.7 to 94.4% seropositivity). These antigens were able to activate the lymphocytes of P. vivax-‐exposed individuals, and the activated CD4 T lymphocytes produced higher levels of Th1 (interleukin-‐2 [IL-‐2] and gamma interferon [IFN-‐]) and Th2 (IL-‐4 and IL-‐10) cytokines than the healthy controls, but the response was Th2 biased. The combined results of present and previous studies seem to suggest a striking link between induction of the CD4 T cell response and naturally acquired antibodies against all 36 proteins of the Pv-‐fam-‐a family, the majority of them having conserved sequences in the parasite population. 17 Watching the clock: the story of circadian rhythms in malaria parasites Prior, Kimberley (University of Edinburgh, Edinburgh, GBR); O'Donnell,Aidan; Savill, Nicholas; Reece, Sarah The evolution and ecology of infectious diseases is a growing and dynamic field of research. A novel area within this field involves integrating chronobiology, immunology, parasitology, and evolutionary theory with mathematical models to investigate the circadian rhythms of malaria parasites. These parasites live within red blood cells of the mammalian host. Recent work indicates that mismatching the biological (developmental) rhythms of parasite and host, via a means of jet lagging parasites, results in a loss of parasite fitness. This highlights the importance of synchrony and timing of parasite developmental rhythms with its host. Major questions arising from this finding are “to what extent are parasites and hosts in control of synchrony and timing of parasite developmental rhythms” and “what roles do parasite developmental rhythms play in strategies for parasite offence and host defence”? To start to answer these questions a three day time series was performed to measure parasite and host factors every three hours, then twice per day sampling until the end of the infection. Integrating this large dataset with mathematical models it is possible to 1) characterise parasite development in terms of timing and synchrony 2) assess genetic variation in parasite rhythms and 3) ask whether rhythms co-‐vary with variation in the in-‐host environment. A better understanding of parasite and host dynamics during malaria infections means one can start to examine whether hosts or parasites are in charge, identify the factors responsible for rhythms and ultimately what the costs and benefits are to parasite and host. 18 The role of interleukin-‐22 in a mouse model of malaria infection Sodenkamp, Jan; Mastelic, Beatris; Keller-‐Mclaughlin, Sarah; Ahlfors, Helena; Stockinger, Brigitta; Langhorne, Jean; Kouassi, Victor M. (MRC National Institute for Medical Research, Mill Hill, GBR) Interleukin-‐22 (IL-‐22) is known to have a role in protecting tissues from the damaging effects of inflammation and protecting their innate immunity. On infecting C57BL/6 mice deficient in this cytokine with Plasmodium chabaudi chabaudi A.S, increased mortality was observed after the peak of the acute blood-‐stage infection despite no difference in parasitaemia. This is associated with cachexia and increases in alanine transaminase (ALT) and aspartate transaminase (AST) in the plasma, markers of liver damage. There were no obvious differences in histopathology in these organs suggesting subtle effects of the lack of IL-‐22. Il22-‐/-‐ mice infected via mosquito bite showed a less virulent infection; however, plasma ALT was still significantly elevated. Previously, we have reported a small increase in the numbers of CD8+ T cells producing IL-‐22 in the liver in blood-‐stage P. chabaudi infection. Here we have used an IL-‐ 22crexRosa26eYFP fate reporter mouse to detect any cell that has produced IL-‐22 at any time during infection. Although only very few eYFP+ cells were found in the spleen and liver, an increase of eYFP+ lymphocytes in the lungs was detected at day 5 of the infection. This was found to mostly consist of γδ T cells. Increases in eYFP+ gd T cells were also observed in the lamina propria of the small and large intestine at day 5 of infection. At day 10, eYFP+ lymphocytes devoid of T-‐cell markers were more dominant in this population, strongly suggesting involvement of innate lymphoid cells. Both the lung and gut epithelia are known to be vulnerable during malaria infection, and further investigations aim to examine whether IL-‐ 22 may act to mitigate this damage. 19 Low-‐affinity T cells are a feature of the anti-‐malarial immune response King, Thayer (Emory University, USA); Neeld, Dennia; Evavold, Brian; Lamb, Tracey According to the classical understanding of T cell expansion, T cells with high-‐affinity T cell receptors (TCRs) for their cognate peptide:MHC will selectively expand and predominate following infection of a host. However, recent research has shown that low-‐affinity T cells also expand during polyclonal T cell responses in the contexts of both autoimmunity and infection. It is currently unknown how TCR affinity impacts the anti-‐malarial immune response. We have used a malaria antigen-‐specific tetramer containing the glideosome-‐associated protein 50 (GAP50) epitope to detect high-‐affinity antigen-‐specific CD8 T cells in the spleens of Plasmodium berghei ANKA infected C57BL/6J mice. However, since tetramers are believed to primarily detect high-‐affinity T cells, we sought to utilize a more sensitive method that could detect T cells of varying affinities. Using a novel 2D micropipette adhesion frequency assay with GAP50 monomers, we found that low-‐affinity splenic CD8 T cells proliferate in response to blood-‐stage P. berghei ANKA infection. Interestingly, we did not detect T cells with intermediate affinity in this polyclonal CD8 T cell population. At day 6 post-‐infection, both low-‐affinity and high-‐affinity GAP50-‐specific T cells migrated to the brain in response to sequestered P. berghei ANKA infected red blood cells. Both expansion and accumulation of these low-‐affinity T cells in the brain occurred in an interferon gamma (IFN-‐γ)-‐independent manner. Our data demonstrates that low-‐affinity T cells can be a feature of the anti-‐Plasmodium blood stage response and the potential implications of this finding are discussed. Toxoplasma Immunobiology 20 Immune response induced by vaccination with non-‐replicative type I strain of Toxoplasma gondii (CPS-‐1) induces more tetramer specific CD8 T cells with central memory phenotype. Dominguez, Mariana (The George Washington University, USA); Hwang, Sujin; Nixon, Douglas; Moretto, Magali Role of CD8 T cells in immunoprotection against Toxoplasma gondii infection is well established. However, in an encephalitis model, CD8 immunity is functionally impaired (immune exhaustion), leading to reactivation of latent infection. Studies from our laboratory have reported that functional defect during chronic infection is primarily exhibited by memory subset of CD8 T cells. Thus restoration of dysfunctional CD8 response may be critical for maintaining the chronicity and preventing the reactivation of latent infection. As a first step we evaluated and compared the antigen-‐specific CD8 T cell response induced by vaccination with the non-‐replicative type I strain of Toxoplasma gondii (which efficiently controls per-‐oral infection with type II strain) to the one induced during chronic infection. While type II strain induced a stronger SLEC (short lived effector CD8 cell) response as compared to vaccine strain in all the tissues tested, MPEC (memory precursor effector cell) response with both the strains was were similar. However, interestingly the frequency of central memory cells subset (CD127+CD62Lhigh) in the vaccinated mice was significantly higher as compared to those receiving type II infection. These findings suggest that the development of inadequate central memory CD8 response may be an important defect responsible for reactivation of infection in mice susceptible to toxoplasmic encephalitis. 21 CCR7+ dendritic cells are sufficient to recover a deficient population of KLRG1+ T cells in the brain of chronically infected CCR7-‐/-‐ mice Landrith, Tyler (University of California, Riverside, Loma Linda, CA, USA); Noor, Shahani (University of New Mexico, USA); Wilson, Emma H. Toxoplasma gondii is a protozoan parasite that establishes a chronic infection in the brain. CCR7-‐/-‐ mice are susceptible to challenge with normally avirulent strains of T. gondii, but if the mice are treated with sulfadiazine, a chronic infection is established. At this stage, equivalent numbers and proportions of parasite-‐specific T cells are seen in the brain of CCR7-‐/-‐ and wild type mice suggesting normal priming is possible in the absence of CCR7. However, CCR7-‐/-‐ mice fail to mount a protective response to rechallenge with a virulent strain of parasite in contrast to their wild type counterparts, implying deficient effector mechanisms despite equivalent priming. Indeed, a profound defect is observed in the number and proportion of T cells expressing the effector marker KLRG1 in the brain of CCR7-‐/-‐ mice. Here we present data indicating that this defective effector population is restored to wild type levels when CCR7+ dendritic cells are transferred to CCR7-‐/-‐ recipients at the chronic stage. This data suggests a novel role for CCR7 during chronic T. gondii infection in guiding efficient dendritic cells interactions required to maintain effector populations of T cells in the brain. 22 Genetic approaches to identify Toxoplasma effector proteins that alter inflammasome activation Gregg, Beth (NIH/NIAID, Bethesda, MD, USA); Khan, Asis; Kennard, Andrea; Grigg, Michael E. The ability of protozoan pathogens to subvert host immune responses and establish chronic transmissible infections is a fascinating dynamic in the ecology and evolution of infectious disease processes. Using Toxoplasma gondii as a model organism, parasite effector proteins that actively subvert host innate immune responses were identified using both population and forward genetic screens. Progeny from a collection of T. gondii crosses were screened using bone-‐marrow derived macrophages for their ability to differentially modulate activation of the host inflammasome: a two-‐ signal process that regulates parasite proliferation. Signal one involves TLR and NFκB signaling, resulting in the upregulation of pro-‐Caspase-‐1, Nlrp’s, and immature IL-‐1β and IL-‐18. Signal two causes the oligomerization of NLRP1 or 3 resulting in the cleavage of pro-‐Caspase-‐1 and the secretion of mature IL-‐ 1β and IL-‐18. This study identified significant strain-‐specific differences in the upregulation of IL-‐1β. Previous work identified GRA15 as the critical parasite factor mediating IL-‐1β upregulation; however, QTL mapping of T. gondii cross-‐progeny identified at least three parasite loci regulating IL-‐1β, indicating that this is a multi-‐locus phenotype that results in strain-‐specific immune modulation. Characterization of these as yet unidentified polymorphic effector molecules should provide considerable insight into their impact on host immune selection, parasite niche localization, and transmissibility. 23 The IL-‐27 subunit p28 inhibits the germinal center B cell response to Toxoplasma gondii DeLong, Jonathan H. (University of Pennsylvania, Philadelphia, PA); Konradt, Christoph; Tait Wojno, Elia D. (Cornell University, USA); Glatman Zaretsky, Arielle (NIH); Hunter, Christopher A. Interleukin (IL)-‐27 is a heterodimeric cytokine composed of the proteins EBI-‐3 and IL-‐27p28. It is critical for controlling the immune response to several pathogens including Toxoplasma gondii. The IL-‐27p28 subunit can be secreted independently of EBI-‐3. Our group previously found that IL-‐27p28 alone blocks signaling through gp130, a component of the receptors for IL-‐6 and IL-‐27, among other cytokines. Through development of a transgenic mouse (p28Tg mouse) that overexpresses IL-‐27p28 in T cells and B cells, we found that IL-‐27p28 inhibits the development of germinal centers, GC B cells, and class-‐ switched antibody in response to NP-‐Ova immunization. The goals of the present study are to investigate the mechanism behind these deficiencies and to determine the role of IL-‐27p28 in the immune response to Toxoplasma gondii. Resistance to T. gondii depends on cell-‐mediated immunity and humoral responses. p28Tg mice are highly susceptible to toxoplasmosis, developing severe toxoplasmic encephalitis. Further analysis revealed that infection-‐induced expansion of follicular helper T cells is not reduced in p28Tg mice, but germinal center B cell numbers are deficient. In addition, serum titers of T. gondii-‐specific IgM, IgG and IgG2c are deficient in p28Tg mice. It is therefore possible that IL-‐27p28 limits the GC B cell response through directly inhibiting IL-‐6 signaling on B cells. As IL-‐27 has been shown to promote IL-‐21 production, it is also possible that by inhibiting IL-‐27 signaling, IL-‐27p28 limits IL-‐21 production by T cells. 24 The Role of Epigenetics in Immunity to T. gondii in Mice Zwicker, Christian (Institute of Specific Prophylaxis and Tropical Medicine, Medical University of Vienna, Vienna, Austria, Vienna, AUT); Hassler, Melanie R. (Clinical Institute of Pathology, Medical University of Vienna, Vienna, Austria); Joachim, Anja (Institute of Parasitology, University of Veterinary Medicine, Vienna, Austria); Egger, Gerda; Wiedermann, Ursula; Schabussova, Irma Infection with Toxoplasma gondii, a protozoan, apicomplexan parasite is highly prevalent worldwide evident by around 30% seropositivity among the human population. In support of the "Hygiene Hypothesis" several epidemiological studies have demonstrated an inverse relationship between microbial/parasitic infections, such as toxoplasmosis, and the development of allergic diseases. Recently, we have shown that chronic exposure to T. gondii induced a robust Th1 immune response associated with reduced development of a pro-‐allergic Th2 phenotype in a mouse model of birch pollen allergy. In this study we aim to explore whether epigenetic mechanisms play a role during the infection with T. gondii and how such mechanisms might be involved in the immunomodulatory capacities of this parasite. Mice were orally infected with T. gondii oocysts and mRNA expression and chromatin remodeling in CD4+ T cells was analyzed in spleen and mesenteric lymph nodes (MLNs). After 8 days of infection we detected a strongly upregulated expression of Th1 cytokines on mRNA level as well as increased histone markers related to active genes at Th1 loci in CD4+ T cells of mice having received T. gondii. To further investigate whether epigenetic mechanisms are involved in the development of immune responses to T. gondii, we will characterize the role of histone deacetylases (HDACs) during acute and chronic infection. For this purpose, we will infect T cell-‐specific HDAC1 knock-‐out mice (LckCre deleter strain) with T. gondii and analyze the immune response to parasite and bystander antigens, such as ovalbumin or the major birch pollen allergen Bet v 1. First results of epigenetic studies show that infection with T. gondii in mice induced histone modifications at Th1-‐associated genes in CD4+ T cells. 25 RabGDIa is a negative regulator of interferon-‐gamma-‐inducible GTPase-‐dependent cell-‐ autonomous immunity to Toxoplasma gondii Yamamoto, Masahiro (Research Institute for Microbial Disases, Osaka University, Suita, JPN); Ohshima, Jun; Sasai, Miwa; Ma, Jisu ; Lee, Youngae (RIMD, Osaka University, USA); Bando, Hironori Interferon-‐gamma (IFN-‐γ)-‐inducible GTPases such as p47 immunity-‐related GTPases (IRGs) and p65 guanylate-‐binding proteins (GBPs) are recruited to pathogen containing vacuoles, which is important for cell-‐autonomous immunity against vacuolar pathogens. Although the positive regulation for the proper recruitment of IRGs and GBPs to the vacuoles is being elucidated, the suppressive mechanism is unclear. Here, we show that RabGDIα, originally identified as a Rab small GTPase inhibitor, is a negative regulator of IFN-‐γ-‐inducible GTPases in cell-‐autonomous immunity to the intracellular pathogen Toxoplasma gondii. Overexpression of RabGDIα, but not of RabGDIβ, impaired IFN-‐γ-‐dependent reduction of T. gondii numbers. Conversely, RabGDIα deletion in macrophages and fibroblasts enhanced the IFN-‐γ-‐ induced clearance of T. gondii. Furthermore, RabGDIα-‐deficient mice were more resistant to infection by T. gondii than control mice. Among members of IRGs and GBPs important for the parasite clearance, Irga6 and Gbp2 alone were more frequently recruited to T. gondii-‐forming parasitophorous vacuoles in RabGDIα-‐deficient cells. Notably, Gbp2 positively controlled Irga6 recruitment that was inhibited by direct and specific interactions of RabGDIα with Gbp2 through the lipid binding pocket. Taken together, our results suggest that RabGDIα inhibits host defense against T. gondii by negatively regulating the Gbp2-‐Irga6 axis of IFN-‐γ-‐dependent cell-‐autonomous innate immunity. 26 SPARC promotes efficient T cell migration during CNS infection McGovern, Kathryn E. (UC Riverside, Riverside, CA, USA); Nance, J. Philip; David, Clement; Worth, Danielle; Noor, Shahani; Wilson, Emma H. Infection with the protozoan parasite Toxoplasma gondii results in a life-‐long infection in the brain. Control of parasite replication requires the continuous presence of IFN-‐γ producing T cells to keep T. gondii in its slowly replicating, cyst form. A system of reticular fibers, detectable by two-‐photon microscopy, is produced upon infection and is associated with T cell migration within the brain parenchyma. These fibers are present areas associated with immune cell infiltration. Data generated in our laboratory suggests that the extracellular matrix-‐remodeling molecule SPARC (secreted protein, acidic, rich in cysteine) is necessary for successful formation of the network. Consequences of an absence of SPARC include a reduced volume of the reticular fibers found in the brain, an increase in parasite burden and increased accumulation of cells in the perivascular space. In addition, SPARC null mice accumulate or retain fewer antigen specific cells in the inflamed brain. Thus we show a new role for SPARC, providing protective immunity by regulating T cell migration in the CNS. 27 T-‐bet-‐independent IFN-‐gamma responses are sufficient for induction of Toxoplasma gondii mediated intestinal immunopathology Lopez-‐Yglesias, Americo (UT Southwestern, Dallas, TX); Burger, Elise; Sturge, Carolyn; Yarovinsky, Felix Toxoplasma gondii is a protozoan parasite recognized by innate receptors, TLR11 and TLR12, leading to MyD88-‐dependent IL-‐12 production. Subsequently, IL-‐12 production results in a robust CD4+ Th1 IFN-‐ gamma response that is essential for host resistance. In an oral T. gondii infection model, we have previously demonstrated that the CD4+ T cell IFN-‐gamma response leads to intestinal immunopathology characterized by elimination of Paneth cells, loss of antimicrobial peptides, and significant expansion of Escherichia coli in the small intestine (Raetz et. al., 2012). The transcription factor T-‐bet, encoded by Tbx21, is well described as being essential for CD4+ T cell Th1 development and IFN-‐gamma responses. T-‐bet is also indispensable for the development of type I innate lymphoid cells (ILC1) that play a major role in IFN-‐gamma mediated host defense and immunopathology during mucosal responses to T. gondii (Klose et al., 2014). Therefore, we set out to determine the relative contribution of CD4+ T cells and ILC1s in T. gondii-‐triggered intestinal immunopathology. We established that T-‐bet-‐deficient mice orally infected with T. gondii exhibit strong CD4+ T cell IFN-‐gamma responses in both the draining lymph node and spleen. Tbx21 KO mice also show significant loss of Paneth cells and antimicrobial peptides, resulting in expansion of E. coli. Additionally, adoptive transfers in mice deficient in T cells, but not ILC1s, revealed that the CD4+ T cells and not the ILC1s are responsible for IFN-‐gamma responses to T. gondii and subsequent parasite induced intestinal damage. Overall, we established that the transcription factor T-‐bet, while required for ILC1 development, is dispensable for parasite-‐induced CD4+ Th1 responses and IFN-‐gamma-‐dependent intestinal immunopathology. Leishmania Immunobiology 28 Sand fly salivary proteins regulate Langerhans cells immune response to Leishmania parasites Guimaraes B Costa, Anderson (NIAID/NIH, Rockville, MD, USA); Carvalho, Augusto; Meneses, Claudio; Clausen, Bjorn (University Medical Center, Netherlands); Von Stebut, Esther (Johannes-‐Gutenberg University); Valenzuela, Jesus; Oliveira, Fabiano Leishmania infection begins when an infected sand fly bites a host inoculating parasites along with its saliva into the skin. Murine skin contains three Langerin+ cells populations: epidermal Langerhans cells (LCs) and two Langerin+ dermal DCs. Using knock-‐in mice (Ko-‐LC) expressing the diphtheria toxin (DT) receptor under the control of the Langerin promoter, it was shown that depletion of LCs correlates with protection against Leishmania infection. Here, we evaluate the role of LCs when parasites are delivered along with vector saliva, mimicking what happens in nature. We depleted LCs by DT treatment of C57BL/6 mice and infected the mice ear pinna with 1000 Leishmania major metacyclics with (Lm-‐SGH) or without (Lm) 0.5 pair of salivary gland homogenate from Phlebotomus duboscqi. As expected, depletion of LCs (Ko-‐LC) protected animals infected with Lm from developing the disease compared to WT. Counter-‐intuitively, Ko-‐LC mice when infected with Lm-‐SGH displayed a much more severe outcome of infection compared to WT. At the third week post infection with Lm-‐SGH, Ko-‐LCs mice present larger lesion thickness compared to all other groups. Parasite burden was higher (2.5 fold) in Ko-‐LC Lm-‐SGH group correlating with the observed increased lesion size. Nine weeks post-‐infection, Ko-‐LCs Lm-‐SGH still had unhealed larger lesions when compared to other groups. Flow cytometric analysis of the infected ear cells revealed that Ko-‐LC Lm-‐SGH mice had approximately seven times higher frequency of CD4+Foxp3+ T regulatory cells than Ko-‐LC Lm group. Taken together this data indicates that in absence of LCs sand fly saliva potentiates Leishmania infection by induction of CD4+Foxp3+ T regulatory cells. Our next step is to confirm the role of LCs when parasites are delivered by the natural route of transmission, the bites of an infected sand fly. 29 a-‐gal-‐trissacaride as a Vaccine against Cutaneous Leishmaniasis Iniguez, Eva (Department of Biological Sciences. The Border Biomedical Research Center. University of Texas at El Paso. El Paso TX); Portillo, Susana; Almeida, Igor; Maldonado, Rosa Cutaneous leishmaniasis caused by the protozoan parasite Leishmania major affects millions of people worldwide. Most available drugs are expensive; require long treatment regimens and due to the development of resistant parasite strains are becoming more ineffective. In addition, no preventive or therapeutic human vaccine is currently available. For this reasons, there is a crucial need of a protective vaccine against this disease. Except for Old World primates and humans, mammals express α-‐gal epitopes and, as a consequence, do not produce anti-‐Gal antibodies. In addition, we know that α-‐gal is abundant in the surfaces of Leishmania parasites. Therefore, using these specific α-‐gal epitopes would allow us to target α-‐Gal residues that are not expressed in human cells, but highly abundant in the surface of the parasite. Based in all this evidence, we evaluate if the immune response to Galα 1-‐3Galβ1-‐ 4GlcNAc-‐R (α-‐gal-‐trissacaride) protects a against the L. major infection in a murine model. α-‐1,3-‐ Galactosyltransferase gene knockout (α1,3GalT-‐KO) mice were immunized with 10 μg of α-‐gal-‐ trissacaride and control group with BSA; and levels of antibody titers (anti-‐α-‐gal) were determined after immunizations by Chemiluminescent enzyme-‐linked immunosorbent assay (CL-‐ELISA). High anti-‐α-‐Gal levels were observed in the α-‐gal-‐trissacaride immunized group in comparison with control group. In order to validate if α-‐gal-‐trissacaride could protect against infection with L. major, mice were challenge with 106 L. major metacyclic promastigotes. A two-‐fold reduction was observed in the footpad lesions of mice immunized with α-‐gal-‐trissacaride, in comparison with control group. All together, these results suggest that α-‐gal-‐trissacaride produced specific antibodies with a protective immunity against L. major infection. Future experiments with the ideal murine model for CL (BALB/c α1,3GalT-‐KO mice) are planned. 30 ATP metabolizing activity of Leishmania donovani and its implications in establishment of infection Arora, Kashika (Motilal Nehru National Institute of Technology, Allahabad, IND); Verma, Pankaj; Saini, Sheetal; Kureel, Amit K. ; Rai, Ambak K. Even with the advancement in our understanding about leishmaniasis, it is still affecting large number of people every year in the world including Indian subcontinent. Various drugs have been introduced worldwide till date but still the disease remains to be eradicated. The need to look for new drug stepped up various approaches to analyze the metabolic pathways of the causative organism L. donovani. One of the premium survival routes is inability of the parasite to synthesize purines de novo due to which it is dependent upon its host. This calls for a transport system to make the path for purines and allow their entry inside the cell. But the purines must be available in the extracellular environment, which are actually supplemented to the parasite by the human host in the form of ATP. The ATP is actually broken down to adenosine by the ecto-‐nucleotidase, an enzyme present on the surface of the parasite. The released adenosine can then be taken up via transporters and the parasitic cell can continue with its metabolism. Although the purine transporters have been the drug targets but much about their active sites remains to be explored. Our findings suggest discordance in ATP metabolizing activity of the parasite and its modulation by extracellular concentration of inorganic phosphate (Pi). Inhibiting ATP metabolism of L. donovani could be a therapeutic advancement and may offer a better alternative to current therapies. 31 Chemokines, targetable biomarkers of the outcome of treatment of cutaneous leishmaniasis caused by Leishmania Viannia? Vargas, Alejandro A. (CIDEIM-‐Colombia, Cali, COL); Barrera, María ; Dillon, Laura (University of Maryland); Navas, Adriana; El-‐ Sayed, Najib (University of Maryland); Gómez, María Antimonial drugs remain the first-‐line treatment for cutaneous leishmaniasis (CL) caused by Leishmania species, endemic in South America. The increasing rates of therapeutic failure and the toxicity of antimonials threaten its usefulness. Although Leishmania drug susceptibility has been considered a determinant of therapeutic responsiveness, treatment failure can occur during infections with drug-‐ susceptible parasites. Being Leishmania an obligate intracellular pathogen, we sought to explore the role of the human host cells in the outcome of antimonial drug treatment during infections with drug susceptible L. panamensis. Using high-‐throughput, high resolution RNA Sequencing (RNA-‐Seq), we profiled the gene expression signature of L. panamensis infected and drug-‐treated primary human macrophages from patients with successful (n=2) and failed (n=2) therapeutic outcomes after standard-‐ of-‐care treatment with meglumine antimoniate (MA). Results revealed five times more transcripts being modulated in macrophages from responders (631 vs. 128 in non-‐responders). Gene ontology analyses showed that biological processes predominantly related to immunological functions were significantly up-‐regulated and enriched in macrophages from responders, compared to oxidative-‐stress response genes enriched in host cells from non-‐responders. Chemotactic chemokine genes (ccl8,cxcl3) were found up-‐regulated in macrophages from patients who responded to treatment. To corroborate this, primary macrophages from CL patients who cured (n=9) or failed (n=5) treatment were analyzed. L. panamensis infection induced expression of cxcl3 and ccl8, and that of ccl2 and cxcl5 in macrophages from both patients groups. Notably, infection followed by drug exposure (MA: 32µg/ml) markedly repressed expression of all four chemokine genes in macrophages from non-‐responders. These exploratory analyses suggest that early drug and parasite-‐mediated modulation of the chemokine network could be determinant to the outcome of treatment of CL. 32 Cutaneous infection with Leishmania major mediates heterologous protection against visceral infection with Leishmania infantum ROMANO, Audrey (NIH/LPD, Bethesda, MD, USA); Doria, Nicole; Mendez, Jonatan; Sacks, David; Peters, Nathan The leishmaniases are a spectrum of cutaneous, mucocutaneous, and visceral diseases caused by the eukarytotic parasite Leishmania. No vaccine is currently available, however, deliberate needle inoculation of infectious L. major (L.m) into the skin without the disease exacerbating factors co-‐egested during natural sand fly transmission, termed leishmanization, produces a self-‐limiting cutaneous lesion that provides complete homologous protection against subsequent sand fly transmitted cutaneous disease. The most beneficial application of leishmanization would be to vaccinate against strains, such as L. donovani or L. infantum (L.i), that cause lethal visceral leishmaniasis (VL). We show that mice with a healed primary L.m. infection in the skin significantly reduced parasite numbers in the ear, draining lymph node, spleen and liver versus control animals upon i.d. challenge with L.i., and achieved sterilizing immunity in the viscera at 6 weeks post-‐challenge. Protection correlated with an early and sustained recruitment of IFN-‐g+CD4+ in the skin and viscera. In vivo staining with TCRβ-‐fluorescent antibody allows the distinction between parenchymal cells and cells localized in the vasculature of the liver and the red pulp of the spleen. By using this technique we demonstrated that primary infection with L.m. in the skin maintained a high number of IFN-‐g+CD4+-‐cells that were trapped in the vasculature of the liver and spleen, but also maintained a small number within the tissue parenchyma. L.i. challenge stimulated the rapid recruitment and/or retention of IFN-‐g+CD4+ cells in visceral tissue. This is the first demonstration that the protective immunity generated by chronic infection with a cutaneous strain of Leishmania can cross-‐protect against i.d. challenge with a visceral strain. Employing a hamster model of VL that mimics human disease will help us verify if leishmanization can protect against fatal visceral disease following natural sand fly transmission. 33 Activation of human keratinocytes by Leishmania spp.: divergent effects of Leishmania infantum versus Leishmania major Scorza, Breanna M. (University of Iowa, Iowa City, IA, USA); Wacker, Mark; Messingham, Kelly; Fairley, Janet; Wilson, Mary Diverse clinical manifestations of leishmaniasis are caused by different infecting Leishmania species. Leishmania major (Lm) cause localized cutaneous lesions at the inoculation site, whereas visceralizing L. infantum (Li) cause potentially fatal disseminated disease. Early local inflammatory responses to Leishmania at the site of skin infection influence the adaptive response and disease outcome, but surprisingly little is known about the role of keratinocytes, the major immunomodulatory cell type of the epidermis. We hypothesized Leishmania induce keratinocytes to produce cytokines and chemokines that influence the subsequent immune response. Using primary human keratinocytes or a human keratinocyte cell line, gene expression upon exposure to Leishmania was investigated using RT-‐qPCR. Incubation of Li with human keratinocytes caused a significant increase in the transcripts encoding pro-‐ inflammatory cytokines il6, il8, and tnfa. However, keratinocytes exposed to a series of distinct Lm isolates did not increase production of cytokine mRNA, suggesting a species-‐specific inflammatory response. Western blotting confirmed NFkB-‐p65 activation following Li, but not Lm, exposure. Furthermore, Li-‐derived exosomes also induced il6 and il8 expression by keratinocytes, whereas Lm-‐ or non-‐specific exosomes did not. Based on the hypothesis that keratinocyte cross-‐talk with immune cells may contribute to disease outcome, Li-‐infected human monocytes were co-‐cultured with Li-‐exposed or control keratinocytes across a semi-‐permeable membrane. Monocytes co-‐cultured with Li-‐exposed keratinocytes were significantly better able to control intracellular parasite replication than infected monocytes alone. This control was mediated through a soluble, or exosome-‐packaged, factor. In contrast, co-‐culture with Lm-‐exposed keratinocytes did not affect monocyte intracellular Lm infection. These observations suggest activation of human keratinocytes by Li, but not by Lm promastigotes, leads to release of cytokines that promote a local inflammatory environment and macrophage leishmanicidal activity. These observations may be relevant to the propensity of Lm, but not Li, to cause a chronic skin lesion at the site of inoculation. 34 Leishmania RNA virus 1 induced microRNA lead to parasite persistence by promoting host survival Eren, Onur (University of Lausanne); Reverte Royo, Marta; Hartley, Mary-‐Anne; Rossi, Matteo; Ronet, Catherine; Fasel, Nicolas Metastatic forms of leishmaniasis are almost exclusively caused by infections with species of the Leishmania Viannia subgenus. Previously, we showed that a naturally occuring cytoplasmic virus within L. (Viannia) guyanensis (L.g) was able to worsen disease outcome in mice. Leishmania RNA virus (LRV) induced a destructive anti-‐viral immune response via Toll-‐like receptor 3 (TLR3), which worsened lesional swelling and increased parasite burden. The underlying molecular mechanisms responsible for LRV-‐dependent parasite persistence are not fully described. Here, we determined the microRNAome profile of murine macrophages infected with isogenic L.g strains either containing or depleted of LRV. We found that miR-‐155 was the only microRNA among 1200 unique murine miRNAs upregulated in the presence of LRV and further demonstrated this induction to be dependent on LRV-‐TLR3 interaction. Finally, we found that miR-‐155 indirectly activates PI3K/Akt pathway, which prolongs macrophage survival and parasite persistence. Our data supports a growing body of evidence that indicates TLR ligation as a cell survival stimulus and reveals a mechanism of action by which the anti-‐viral immune response contributes to parasite virulence at a microRNA level. 35 Understanding glyoxalase pathway of L. donovani for development of alternate treatment option Tiwari, Praachi (Motilal Nehru National Institute of Technology, Allahabad, IND); Saini, Sheetal; Chiranjivi, Adarsh K; Rai, Ambak K. Visceral leishmaniasis is a tropical disease that has posed a major problem in many tropical and temperate countries including Indian subcontinent. The current treatment options are not cost-‐effective and exhibit one or other side effects. Moreover, these drugs require the help from immune system of the patient which is already suppressed. Occurrence of disease at a slow but steady rate indicates the persistence of parasite as well as a potential threat of an outbreak sooner or later if not properly taken care. Therefore, there is a need to focus on the development of new drugs which is independent of the immune responses ongoing within the patients. Similar to their mammalian counterpart, L. donovani neutralizes methylglyoxal generated in glycolysis pathway. However, it differs in one or more way(s) from the humans. It is known that methylglyoxal is a toxic ubiquitous product of cellular metabolism and eliminated either through glyoxalase pathways and/or ejected across the cell membrane. This toxic metabolic by-‐product is known to be detoxified predominantly by the glyoxalase pathway. Our approaches leading to accumulation of methylglyoxal in the parasite may result in its death due to toxicity. Besides targeting the glyoxalase pathway, we intend to understand the mechanism of methylglyoxal release across the cell membrane and possible ways to inhibit its exclusion from parasite. Targeting this pathway may lead to decipher ways for accumulation Methylglyoxal in cells and can further help in identifying possible drug targets. 36 Combinatorial proteomics and cellular immunology identify conserved, immunodominant and cross species protective Leishmania antigen and the responding CD4+ T cells at clonal level Mou, Zhirong (University of Manitoba, Winnipeg, CAN); Liu, Jintao; Liu, Dong; Khadem, Forough; Okwor, Ifeoma; Jia, Ping; Uzonna, Jude There is currently no clinically effective vaccine against leishmaniasis due to poor understanding of the antigens that elicit dominant and protective T cell immunity. Using cellular immunology and proteomics, we identified a dominant naturally processed peptide (Leishmania major PEPCK335-‐351) derived from Leishmania glycosomal phosphoenolpyruvate carboxykinase (PEPCK). PEPCK is highly conserved in all pathogenic Leishmania, expressed in glycosomes of promastigotes and amastigotes and elicits strong CD4+ T cell responses (proliferation and polyfunctional cytokine production) in infected mice and human patients. L. major I-‐Ab-‐PEPCK335-‐351 tetramer identified for the first time protective Leishmania-‐ specific CD4+ T cell responses at clonal level, which comprised ~ 17% of responding CD4+ T cells at the peak of L. major infection. PEPCK335-‐351-‐specific CD4+ T cells undergo expansion, effector activities (cytokine production), contraction and stable maintenance following lesion resolution. Vaccination with PEPCK peptide, DNA or recombinant protein induced strong cross-‐species protection in both the resistant and susceptible mice. 37 Leishmania donovani L82 strain: a new mouse reference strain with different properties compared to LV9? Kauffmann, Florence (Vrije Universiteit Brussel, Elsene, BEL); Cnops, Jennifer; Maes, Louis (University of Antwerpen, USA); Dujardin, Jean-‐Claude; Magez, Stefan; De Trez, Carl Leishmania donovani, a sandfly-‐transmitted intracellular protozoan parasite, is the main causative agent of visceral leishmaniasis (VL). VL is characterized by hepatosplenomegaly, fever and weight loss and is fatal if left untreated. The current knowledge on the immunopathology of L. donovani infections is largely based on infections with the laboratory strain LV9. Hence, we decided to analyze the in vivo behavior and characteristics of another L. donovani strain, L82, in different models. Like with LV9, we observed that the infection of L82 in Syrian hamsters, a reference model of human VL, also lead to weight loss, hepatosplenomegaly and is fatal. Using the mouse model, we analyzed the contribution of two key effector players in the control of LV9 infection in vivo, TNF and IL-‐10, which play a protective and detrimental role, respectively. Firstly, similarly to LV9 infection, TNF-‐deficient mice infected with L82 also succumbed from infection by 8 weeks post-‐infection. However, surprisingly and in contrast to LV9 infection, this susceptible phenotype did not seem to be associated to uncontrolled liver parasitemia. Secondly, using L82-‐infected IL-‐10 reporter mice, we demonstrated that spleen and liver CD4+ T cells are the main producers of IL-‐10, as is also shown following LV9 infection. Hence, infection of IL-‐10-‐deficient mice with L82 matched with results obtained in the LV9 model showing a negative role of IL-‐10 in parasitemia control. However, in contrast to LV9 model, preliminary results suggest that B cell-‐derived IL-‐10 does not hamper parasite control. Together, these results suggest that LV9 and L82 strains of L. donovani induce different immunological effector mechanisms depending on their molecular environment. 38 The Role of Vascular Remodeling in the Immunopathology of Leishmania Infection Weinkopff, Tiffany (University of Pennsylvania, Philadelphia, PA, USA); Konradt, Christoph; Christian, David Cutaneous leishmaniasis is caused by intracellular protozoan parasites, and has a wide spectrum of clinical presentations mediated in large part by an exaggerated inflammatory response. Since vascular remodeling contributes to the magnitude of the inflammatory response, we hypothesized that manipulation of the cellular and molecular mediators promoting vascular remodeling might provide a novel approach to limit pathology in leishmaniasis. To address this, we first characterized the vascular remodeling that occurs in mice infected with Leishmania major. We found dramatic changes in vessel morphology, structure, and number within leishmanial lesions, and using intravital imaging we visualized a significant increase in vascular permeability that is a consequence of vascular remodeling in leishmaniasis. At the peak of infection, VEGF-‐A and VEGFR-‐2 expression were upregulated and correspondingly endothelial cells were proliferating at the site of infection. To determine if these VEGF-‐ mediated responses contributed to the magnitude of the pathology in the disease, we treated L. major infected mice with neutralizing antibodies directed against VEGFR-‐2. This treatment led to a decrease in the pathology seen in L. major infected mice compared with control animals. Taken together these data suggest that VEGF-‐A-‐mediated vascular remodeling occurs in leishmaniasis and that blockade of this remodeling can lessen the magnitude of the immunopathology associated with this disease. 39 Jak3 inhibition controls immunopathology induced by CD8+ T cells in Leishmania braziliensis infection Novais, Fernanda O. (University of Pennsylvania, School of veterinary medicine, Philadelphia, PA, USA); Robertson, Tanner; Scott, Phillip The host immune response plays a critical role not only in protection from human leishmaniasis, but also in promoting disease severity. Using experimental models of infection and skin samples from infected patients, we found that cytotoxicity induced by CD8+ T cells is a major mechanism for immunopathology in cutaneous leishmaniasis caused by L. braziliensis. However, the mechanisms that lead to a cytolytic profile in CD8+ T cells in the skin remain unclear. We hypothesized that the cytokine environment in the skin promotes a cytolytic profile in CD8+ T cells and enhance pathology. We found that IL-‐15 was induced in lesions of mice and humans infected with L. braziliensis and in addition IL-‐15 was required for the induction of cytotoxic markers in CD8+ T cells in vitro and in vivo. Importantly, using mouse models of pathology dependent on CD8+ T cells we show that blockage of IL-‐15 signaling through Jak3 inhibition controls immunopathology induced by CD8+ T cells without affecting protective responses. In conclusion, we propose that Jak3 signaling in CD8+ T cells is detrimental to the host and targeting this pathway should be considered for immunotherapy in patients infected with L. braziliensis. 40 A PHASE I STUDY TO ASSESS THE SAFETY AND IMMUNOGENICITY OF A CANDIDATE LEISHMANIA VACCINE CHAD63-‐KH Osman, Mohamed (University of York, USA); Wiggins, Rebbbeca ; Aebischer, Toni (Robert Kock, Germany); Smith, Deborah; Lacey, Charles; Kaye, Paul Visceral leishmaniasis (VL) causes an estimated 40,000 deaths annually. With the paucity of new treatment options and concerns over drug resistance, the development of new preventative and/or therapeutic measures is a major international research priority. Here, we report on progress in developing a novel therapeutic vaccine for VL and PKDL, targeting the induction / re-‐activation of CD8+ T cells. We combined a number of innovative features, to develop a polyprotein vaccine comprising two Leishmania antigens (HASPB and KMP11) vectored in a manner designed to elicit high levels of CD8+ T cell effector function (along with strong CD4+ T cell responses). HASPB is known to feature stretches of 11-‐14 amino acid-‐long repeats and to minimise any impact of isolate-‐specific variation in the repeat regions, we designed a synthetic Haspb gene comprising the conserved N and C termini flanking 10 repeats from the 17 identified to date. Repeats were arranged to preserve their native order as observed in multiple field isolates, to conserve their natural re-‐iteration, and to maintain natural protein length. The unique Haspb gene was co-‐translationally expressed with the Kmp11 gene through the use of a viral 2A sequence, allowing use of a single vector for vaccine delivery. HuAd5-‐KH was previously been reported to have efficacy as a stand-‐alone therapy in mice infected with L. donovani (Maroof et. al. JID 2012. 205:853-‐63). For clinical studies, we manufactured ChAd63-‐KH replacing HuAd5 with a simian adenovirus with an established safety and immunogenicity profile in man.. Here, we will update on our progress taking this vaccine into Phase 1 first-‐in-‐man clinical trial, to determine the safety and immunogenicity of AdCh63-‐KH in healthy adults. 41 Leishmania donovani infection impacts on early events in hematopoiesis Pinto, Ana Isabel (University of York, York, GBR); Preham, Olivier; Brown, Najmeeyah; Dalton, Jane E.; Beattie, Lynette (University of Queensland); Doehl, Johannes; Kaye, Paul M Visceral leishmaniasis (VL) in humans and animal models is associated with significant changes in haematological function, including anaemia and leukopenia, wherein the bone marrow (BM) represents a site of parasite persistence. However, the mechanisms underlying hematologic deregulation are largely unknown. In B6 mice, using a panel of stem cell markers, murine haematopoietic stem cells and precursors (HSCPs) were characterized in the BM during the course of L.donovani-‐infection. Chronically infected mice showed a significant increase in intermediary non-‐committed multipotent haematopoietic precursors (Lineage-‐ Sca1+ cKit+ (LSK), CD150+ CD48+) without reciprocal increases either in lineage-‐committed precursors or mature cells in circulation. In the steady state, the majority of HSCs (LSK CD150+ CD48-‐) are found in a quiescent state, representing the cells with the highest degree of reconstitution potential. Conversely, during chronic infection, most entered the cell-‐cycle which correlated with a reduced potential to engraft into syngeneic recipients. The loss of quiescent HSCs was not observed in RAG2-‐/-‐ mice, but adoptive transfer of CD4+ T cells restored this phenotype. Compared to naive mice, CD4+ T cells producing high levels IFN-‐γ were found in high numbers in the BM of infected mice. To test the hypothesis that IFN-‐γ signalling in HSCPs was involved in BM dysregulation, we generated chimeras by the transfer of B6.Ifngr2-‐/-‐ BM cells into B6 recipients. Loss of intrinsic IFN-‐γ signalling led to a partial recovery in the quiescent HSCs reservoir. Our data suggests that CD4+ T cells, partially operating via IFN-‐ γ, regulate the capacity for self-‐renewal of HSCs in L.donovani-‐infected mice. These changes in haematological function should be considered in the design of new therapeutic approaches. 42 Skin Resident Memory CD4+ T Cells Enhance Protection Against Leishmania major Infection Glennie, Nelson (University of Pennsylvania, Philadelphia, PA, USA); Yeramilli, Venkata; Beiting, Daniel; Volk, Susan; Weaver, Casey; Scott, Phillip Leishmania-‐infected patients become refractory to reinfection following disease resolution, and following a primary infection with Leishmania major C57BL/6 mice are highly resistant to reinfection, yet effective immune protection has not been achieved by vaccines. While circulating Leishmania-‐specific T cells are known to play a critical role in immunity, the role of memory T cells present in peripheral tissues has not been explored. We have identified a population of Leishmania-‐specific memory CD4+ T cells present in the skin of mice that have resolved a primary infection with L. major. These cells are present in skin distant from the primary infection, and produce IFNγ in response to L. major stimulation. Using skin grafts we show that they are resident in the skin, rather than recirculating. Following Leishmania challenge, these cells enhance the early recruitment of circulating T cells to the skin in a CXCR3 dependent manner, resulting in better control of the parasites. Our findings indicate that protective immunity to Leishmania, and thus the success of a vaccine, may depend on generating both circulating and skin-‐resident memory T cells. 43 Inflammation mediated induction of HIF-‐1a limits effector T cell responses by impairing dendritic cell function during acute Leishmania infection Hamami, Akil (INRS -‐ Institut Armand-‐Frappier, Montreal, PQ, CAN) Inflammation, although responsible for controlling infection, is often associated with the pathogenesis of chronic diseases. Leishmania donovani, the causative agent of visceral leishmaniasis, quickly induces a strong inflammatory response on the onset of infection. Inflamed tissues are typically characterized by low levels of oxygen, a microenvironment that triggers the hypoxia-‐inducible transcription factor (HIF-‐ 1α). HIF-‐1α plays an integral role in the metabolic changes that drive cellular adaptation to low oxygen availability. Indeed HIF-‐1α induces a number of aspects of host immune function, but can also lead to immunesuppression. HIF-‐1α expression and stabilization in immune cells can be triggered by hypoxia; nevertheless, other factors associated with pathological stress such as inflammation, cancer, and infectious microorganisms can also induce this transcription factor. For instance, Leishmania promastigotes were recently shown to stabilize HIF-‐1α in in vitro infected macrophages. Here, we investigate the role of HIF-‐1α in dendritic cell (DC) function and its effect in regulating CD8 T cell responses in L. donovani infected mice. Our data demonstrate that mice with a targeted deletion of HIF-‐ 1α in CD11c+ cells showed a higher frequency of short-‐lived effector cells (SLEC), enhanced CD8+ T cell expansion, and sustained IL-‐12 expression in splenic DCs. Ablation of HIF-‐1α in CD11c+ cells also resulted in a significantly lower parasite burden. Interestingly, HIF-‐1α expression in CD11chi splenic DCs was induced in an IRF-‐5-‐dependent manner, suggesting that the inflammatory response limits antigen specific CD8+ T cell expansion. Taken together, these findings suggest that Leishmania induces inflammation to target the HIF-‐1α pathway, evade the adaptive immune response, and establish chronic infection. Helminth Immunobiology 44 What do natural infection models tell us about host-‐parasite interactions? Luzzi, M; Bousquet, M; Szabo, Edina; Finney, Constance (University of Calgary,) Sterile immunity, which continues even after the causative agent disappears, is rarely generated against helminths. Infected hosts must balance protective anti-‐parasite responses with infection-‐limiting immune-‐mediated pathology. Since most worm infections are not lethal, chronic infections are tolerated by hosts to limit pathology, leading to two common features: 1. The polarization of immune responses towards a Th2 phenotype. 2. The immunosuppression of worm-‐specific and general inflammatory responses. Helminths facilitate many secondary infections by skewing immune responses towards this phenotype and therefore providing an immune environment that is beneficial to the establishment of other pathogens. However, this is not true in all contexts, and is strongly dependent on the exact parameters of the study (e.g. helminth dose and timing of infections). To discover the mechanisms that underlie these contradictory results, we are developing a natural model of helminth infection using Heligmosomoides polygyrus. This worm is a natural gastrointestinal parasite of wild mice. For H. poly-‐ gyrus, most studies employ a single bolus infection, with doses that are far from physiological or repre-‐ sentative of field conditions We tested whether changes in infection dynamics could be observed using two different doses (100 vs. 200 worms) and infection types (bolus vs. trickle infection models) in the same susceptible strain of mouse (C57BL/6). Gross gut pathology and cell recruitment to the spleen and mesenteric lymph nodes differed between groups, as did cell markers associated with an immune-‐ suppressive phenotype such FoxP3 and surface TGFbeta. These preliminary findings indicate that the nature of infection models strongly impacts disease dynamics, and may help explain why observations made in model systems do not always translate well to what is observed in a natural setting. 45 Th2/1 HYBRID T HELPER CELLS: A REGULATORY POPULATION IN NEMATODE INFECTED PATIENTS? Bock, Cristin Nadine (Institute of Immunology, Freie Universität Berlin, Berlin, DEU); Rausch, Sebastian; Breloer, Minka (Bernhard Nocht Institute for Tropical Medicine, Hamburg); Babu, Subash (National Institute of Research in Tuberculosis, Chennai); Hartmann, Susanne (Institute of Immunology, Freie Universität Berlin) Protective immunity against nematodes depends on the development of efficient Th2 responses, however excessive immune reactivity needs to be controlled to prevent immunopathology. Recently we showed in a mouse model for human hookworm infections that a large fraction of T helper cells differentiates into a bifunctional Th2/Th1 hybrid phenotype in infected mice following the kinetic seen for the differentiation of classical Th2 cells. The hybrid phenotype co-‐expressed the lineage-‐defining transcription factors GATA-‐3 and T-‐bet and co-‐produced Th2 and Th1 cytokines at intermediate (int) levels. Importantly, the hybrid phenotype caused less immunopathology in vivo than the respective classical Th2 subset. Therefore excessive inflammation during chronic nematode infections may be prevented by the self-‐limitation of T helper cells based on the acquisition of a stable hybrid phenotype, able for a balanced and moderate expression of two opposing T helper programs. We hypothesized that Th2/Th1 hybrid cells also occur in humans and are associated with the outcome of nematode infections. In order to characterize the phenotype of human Th2 cells the expression of GATA-‐3, T-‐bet and the associated cytokines IL-‐4, IL-‐13, IL-‐5 and IFN-‐γ was surveyed in peripheral blood CD4+ T cells from patients infected with the intestinal threadworm Strongyloides stercoralis or the tissue filaria W. bancrofti in endemic regions in South India. Indeed a substantial fraction of CD4+ T cells co-‐expressed IFN-‐γ and IL-‐4/IL-‐13. Remarkably, unlike murine GATA3intIFN-‐γint Th2/1 hybrids detected in H. polygyrus and S. ratti infections, human Th2/1 hybrids expressed very low levels of GATA-‐3 and high levels of IFN-‐γ. Furthermore, Th2/1 hybrids were restricted to the IL-‐4/13 producing population, while IL-‐5+ Th2 cells uniformly expressed high levels of GATA-‐3 and no IFN-‐γ. Further studies assessing the differential programming of murine and human Th2/Th1 hybrids as well as their contribution to the outcome of nematode infections are under way. 46 Ascaris lumbricoides population dynamics and co-‐infection in rural western Kenya Easton, Alice V (NIH, USA); Njenga, Sammy (Kenya Medical Research Institute); Nutman, Thomas B (NIH, USA); Anderson, Roy M (Imperial College London) To examine how helminth population biology may inform deworming efforts for soil transmitted helminths (STH), we intensively screened 1671 people in four villages for helminth infections by stool egg-‐counts, treated everyone with albendazole, and collected Ascaris lumbricoides adult worms expelled post-‐treatment. Three months post-‐treatment, we re-‐screened and re-‐treated 1225 people, and re-‐ collected expelled worms. A. lumbricoides prevalence by egg-‐positivity was 8% at baseline and 5% at follow-‐up. Worms were recovered from only half of people who were egg-‐positive, but from some egg-‐ negative individuals as well. These worms were highly aggregated. A. lumbricoides were expelled over 13 days, and 75% were female. There was a strong correlation (r=.64; p<0.0001) between adult worms and intensity of infection (egg counts). A nonparametric correlation matrix between A. lumbricoides, Necator americanus, Giardia lamblia and Entamoeba histolytica for 332 samples showed a negative correlation between N. americanus and G. lamblia infection intensity (r=-‐0.11; p<0.05). Spatial data collected on infection and reinfection, host predisposition and worm persistence, and worm genetic variability among villages/individuals should provide a framework underlying A. lumbricoides population biology that will have implications for control efforts. Ongoing molecular work to quantify the presence of major STH and other intestinal pathogen DNA in stool from study participants, and sequencing of individual worms collected, will add depth to these analyses that cannot be obtained with traditional field-‐based methods. Starting with a backbone proteome for A. lumbricoides, we are sequencing a set of the worms collected, to look for structuring at the household and regional levels, and examine differences between worms collected at different time-‐points. We will explore evolutionary explanations for temporal changes in A. lumbricoides population structure, genetic diversity and markers for resistance, and the host’s microbiome. 47 Bladder Urothelial Cell Cycle Responses to Schistosoma haematobium Infection Are Modulated by IL-‐4 Receptor-‐alpha Fu, Chi-‐Ling; Hsieh, Michael (USA) The bladder urothelium, a normally quiescent epithelial organ, undergoes dynamic changes during Schistosoma haematobium infection (urogenital schistosomiasis). These alterations include hyperplasia, ulceration, and carcinogenesis, and likely involve shifts in urothelial cell cycle status. Defining the pathways underpinning these urothelial responses will contribute to a deeper understanding of how S. haematobium egg-‐induced expulsion, hematuria, and bladder cancer develop in humans. Mammals mount an IL-‐4 and -‐13-‐associated type 2 immune response in response to many different helminth infections, including S. haematobium infection. These cytokines share the IL-‐4 receptor-‐α subunit (IL4R) as a cognate receptor subunit. To determine whether IL-‐4 receptor-‐α signaling regulates urothelial cell cycle alterations in urogenital schistosomiasis, we injected S. haematobium eggs or control vehicle into the bladder walls of wild type, IL4R-‐deficient, and myeloid-‐associated IL4R-‐deficient mice. Three weeks later, mice were sacrificed and their bladder urothelia prepared as single cell suspensions. These suspensions were stained with DAPI and antibodies to CD45 and EpCAM. CD45-‐EpCAM+ urothelial cells were gated and their DAPI staining analyzed to assess cell cycle status. Relative to vehicle controls, wild type mice injected with eggs demonstrated increased proportions of urothelial cells in S phase and decreased proportions in G2/M phase. Although egg-‐injected, macrophage-‐associated IL4R-‐deficient mice featured similar urothelial responses as their egg-‐injected wild type counterparts, egg-‐injected conventional IL4R-‐deficient mice exhibited fewer urothelial cells in S phase. Thus, IL4R signaling through non-‐myeloid cells may affect urothelial cell cycle status; this effect may target DNA synthesis, a crucial process in carcinogenesis. Ongoing work is determining which IL4R-‐expressing cells are crucial to these phenomena, and how these cells mediate alterations in the cell cycle status of the bladder urothelium. 48 A central role for Type I IFN in the induction of Th2 responses by dendritic cells Webb, Lauren (University of Manchester, Manchester, GBR); Lundie, Rachel; Borger, Jessica; Cartwright, Adam; Cook, Peter; Brown, Sheila; Jones, Lucy; Phythian-‐Adams, Alex; Davis, Daniel; MacDonald, Andrew Although dendritic cells (DCs) are critical for induction of Th2 immunity against helminths or allergens, relatively little is known about how they become activated and function in response to Th2-‐polarizing antigens. We have discovered a previously unrecognized role for Type I IFN (IFN-‐I) in the optimal activation and function of DCs following exposure to strongly Th2-‐polarizing antigens from the parasitic helminth Schistosoma mansoni. To date, IFN-‐I has primarily been associated with anti-‐viral immunity, and its role in Th2 settings is currently unclear. DCs lacking the IFN-‐I receptor displayed a dramatically impaired ability to induce Th2 cytokines in vivo, but unimpaired ability to support CD4 T cell polarization in vitro. Further, Th2-‐promoting DCs depended on IFN-‐I signaling for optimal activation, efficient migration to the draining LN, and effective localization within the T cell zone. Together, our data suggest a key role for IFN-‐I to enable Th2 induction by DCs against helminths in vivo. Future work will address the wider role of IFN-‐I in Th2 inflammation, including against other Th2-‐inducing antigens and during patent helminth infection in vivo. 49 Identification of genetic markers linked to susceptibility for filaria-‐induced immunopathology Venugopal, Gopinath (Institute of Immunology, Freie Universität, Berlin, Berlin, DEU); O'Regan, Noëlle Louise; Steinfelder, Svenja; Hartmann, Susanne Lymphatic Filariasis (LF) is an infection affecting more than 120 million people worldwide, which is caused by three nematodes namely Wuchereria bancrofti, Brugia malayi and Brugia timori. The most common manifestations of lymphatic filariasis are asymptomatic infection, characterized by the presence of circulating microfilaria, and chronic infection without microfilaremia, which is associated with severe pathology in the form of lymphedema, hydrocele or elephantiasis. T cells play a major role in immune responses to filarial infections, whereby a significant enhancement of Th1/Th17 cells is observed in chronic pathology, whereas in asymptomatic infection, a general hypo-‐responsiveness develops. Infection with filarial parasites induces alternatively activated macrophages which are involved in immunoregulation and display a characteristic phenotype also found in monocytes from filaria-‐infected individuals. Here, we demonstrate that monocytes from non-‐endemic donors are targeted in vitro by lysate of Brugia malayi microfilaria (Mf), resulting in a regulatory phenotype and up-‐ regulation of IL-‐10, IL-‐6, IL-‐8 and PD-‐L1. Specific worm molecules (e.g. filarial cystatins) have been described and were shown to powerfully suppress unrelated inflammatory responses by addressing host receptors and exploiting signalling pathways related to innate immune responses, such as expression of IL-‐10 and PD-‐L1 in monocytes and macrophages. Therefore, genetic polymorphisms of these receptors and signalling molecules could account for the distribution of clinical manifestations in filarial-‐infected individuals. Co-‐Infection Immunobiology 50 Mechanisms of defense in a Drosophila defensive symbiosis Hamilton, Phineas (University of Victoria, Victoria, BC, CAN); Peng, Fangni; Boulanger, Martin; Perlman, Steve It has become clear that interactions between multiple parasites within a host can be critical in determining disease outcomes. Perhaps the most dramatic way in which infectious agents interact is in defensive symbiosis – wherein one infectious agent defends the host against another. Such defense is now known to be common in heritable symbioses, where symbiont transmission depends on host fitness. The fly Drosophila neotestacea, for instance, is infected by a heritable bacterium − Spiroplasma − that protects it against a virulent and common nematode parasite − Howardula aoronymphium − and positive selection on this defensive trait has caused the recent spread of Spiroplasma across North America. To understand the mechanism underlying Spiroplasma-‐mediated defense, we conducted a series of experiments that included comprehensive gene expression profiling of infected flies, uncovering evidence of a Spiroplasma-‐encoded toxin − a putative ribosome inactivating protein (RIP) − up-‐regulated by Spiroplasma in response to nematode challenge. Subsequent experiments confirmed that Howardula suffers attack characteristic of an RIP in vivo, but only in the presence of Spiroplasma, while heterologous expression of the toxin confirmed its activity against nematodes in vitro. Our results demonstrate the participation of this toxin in Spiroplasma-‐mediated host defense, providing one of the first examples of the mechanism underlying a heritable defensive symbiosis. Attack of invaders with specific toxins can thus be an effective means of protecting the host when co-‐infecting parasites or symbionts clash in their transmission requirements. Other 51 Chemiluminescent-‐ELISA for the diagnosis of Trypanosoma cruzi in Mexico Orozco, Jose A. (University of Texas at El Paso, El Paso, TX); Manriquez-‐Roman, Claudia; Flores, Marco; Avitia, Alfonso (Centro Estatal de Transfusión Sanguínea de Chihuahua, Mexico); Duque, Jorge; Maldonado, Rosa A. Chagas disease is zoonosis caused by the protozoa Trypanosoma cruzi, affects around 10 million individuals. Recently, few acute cases of T. cruzi infection in the United States have been reported following blood transfusion or bone marrow transplantation. Conventional serological tests, along with enzyme-‐linked immunosorbent assay (ELISA) used by blood banks are sensitive enough to detected the presence of the parasite, giving inconclusive, false-‐positive or false-‐negative results. Currently there is no specific serological test that can diagnose the infection of this protozoan in the border region, between the United States and Mexico. In this study we developed a highly specific chemiluminescent-‐ ELISA (CL-‐ELISA) using an epimastigote antigen mix from T. cruzi strains endemic to South America (Dm28c, Y-‐strain and CL-‐Brener) and southern United States (Houston and Texas-‐Tulane). In this CL-‐ ELISA, 319 human sera from Chihuahua blood bank were tested. Previously, these sera were tested utilizing three different methodologies which combined proved to be uncertain for T. cruzi infection. Our results demonstrate this CL-‐ELISA is effective and specific to detect T. cruzi infection compared to other serological tests. This assay could be used in blood banks to test blood units and reduce T. cruzi infection among the population of South America, Central America and the United States. 52 Using bacterial infection in the sand fly gut to control leishmaniasis Oliveira, Fabiano (NIH/LMVR, Rockville, MD, USA); Meneses, Claudio; Guimaraes-‐costa, A; Carvalho, A M; Jacobs-‐Lorena, M (John Hopkins); Valenzuela, J. G. Leishmaniasis is a vector-‐borne neglected tropical disease that affects the poorest of communities and comes only second to malaria and fourth among tropical parasitic diseases in mortality and morbidity, respectively. Here, we tested a control strategy that exploits the use of an engineered bacteria that secrets a defensin-‐like peptide (Scorpine). We tested the effect of the engineered Pantoea agglomerans, a common symbiotic bacterium in many insects including sand flies, on the development of Leishmania parasites inside the sand fly gut. Our data indicates that Pantoea agglomerans was able to significantly reduce Leishmania parasite burden inside sand fly midguts. Importantly, scopine secreting-‐ P. agglomerans impacted colonization by the Leishmania parasites of the anterior portion of the sand fly gut, a key step in parasite transmission to mammalian hosts. Future experiments are targeted to test if scorpine secreting-‐P. agglomerans would efficiently block transmission in animal models of leishmaniasis, including reservoirs of the disease. 53 Development of a Monoclonal Antibody Capture ELISA for Detection of Parasite Antigens in Taenia Solium Cysticercosis Carbajal, Edwin A. (National Institute of Allergy and Infectious Diseases, Washington, DC, USA); Paredes, Adriana (Cayetano Heredia University); Marzal, Miguel; Guerra-‐Giraldez, Cristina; Kwan, Jennifer; Garcia, Hector Hugo; Nash, Theodore Mahanty, Siddhartha Neurocysticercosis (NCC) is an infection of the central nervous system by cysts of the cestode Taenia solium (Ts). We have developed a monoclonal antibody (MAb) based antigen (Ag) detection test for NCC comparable to the current B158-‐B60 Antigen-‐ELISA to be readily used in laboratories and the field. The most reactive of the MAbs produced (designated A7), an IgM isotype, was chosen. An immunofluorescence assay using affinity purified MAb to Ts cysts showed a markedly different pattern of reactivity compared to B158-‐B60, indicating unique target Ags. The assay was developed with purified A7 as the capture antibody and biotin conjugated A7 for detection, and was used for Ts Ag detection in cerebrospinal fluid from individuals with subarachnoid neurocysticercosis. The detection limit of the assay was ≥ 3 ng/ml Ts Ags. The A7-‐based assay was highly sensitive (94%; 95% CI: 70-‐99%) and specific (100%; 95% CI: 66-‐100%). Receiver operating characteristic (ROC) curve analysis determined an optimal cutoff for the assay (5 ng/ml) using CSF samples clinically defined as NCC or non-‐NCC. We studied the evolution of Ag levels at presentation and after treatment with anthelmintics in nine patients with NCC. Seven of nine (78%) individuals with pre-‐ and post-‐ treatment samples had Ag detectable by the A7 capture ELISA, and all nine (100%) experienced a decline in levels to undetectable after completion of treatment. The A7 ELISA correlated significantly with the B158-‐B60 assay (Spearman test; ρ=0.61, p<0.002). This assay will likely be useful in verifying persons with active infections, determining if ongoing treatment is effective and the time cysticidal treatment can be stopped, and potentially, diagnosing regrowth and recurrence of infection. 54 Giardia duodenalis: Characterization of a virulent canine strain with zoonotic potencial through biological and proteomics tools COELHO, CAMILA (GEORGETOWN UNIVERSITY, WASHINGTON, DC, USA); Pereira, Rosiane (Rene Rachou -‐ Fiocruz-‐MG, Brazil); Silva, Ana Carolina (UFMG); Silva e Silva, Daniel (Department of Genetics, UFMG); Nascimento, Frederico (UFMG); Serufo, Angela (UFMG); Chaperouge, Alex (Oswaldo Cruz Institute, Fiocruz, RJ); Perales, Jonas (Oswaldo Cruz Institute, Fiocruz, RJ); Silva, Lorena (Department of Microbiology, UFMG); Abrãao, Jonatas (Department of Microbiology, UFMG); Costa, Adriana (UFMG); Fernandes, Ana Paula (UFMG) Giardiasis is a neglected disease that affects more than 200,000 people in the world. Symptoms include abdominal pain, diarrhea and malabsorption syndrome. Giardiasis is a zoonotic disease and its transmission may occur from domestic animals to humans. Canine strains are less characterized than human isolates of G. duodenalis. In this study, a novel strain (BHFC1), isolated from a symptomatic dog, was characterized through growth aspects, infectivity in Swiss mice, phylogenetic and ultra-‐structural analysis, encystment profile, reactivity with canine and human sera, proteomic map and presence of viral genome. The results revealed that BHFC1 is more infective for Swiss mice and grows faster than the human strain Portland-‐1. Electron microscopy revealed that the flange of the canine strain is larger and the process of encystment is slower or incomplete, at one time point when compared to Portland-‐1. Antibodies in human sera from patients with giardiasis, reacted more strongly with proteins of the canine than the human strain, suggesting a zoonotic potencial for BHCFC1 strain. The proteomic map of soluble and insoluble fractions led to the identification of 212 known and 28 hypothetical proteins of G. duodenalis. Some of these proteins correlated with virulence and pathogenic mechanisms. Amplification of viral genome was not detected, suggesting absence of Giardiavirus. These data provide an important source of information for future studies correlating various aspects of the biology of this parasite, such as virulence, protein interaction, metabolomics, ultrastructural characterization, among others. In addition, a well characterized G. duodenalis strain, virulent in experimental model, becomes an useful tool for investigations of antigenicity, immunogenicity, drug resistance and the development of vaccines, diagnosis and effective treatment. 55 Immunological responses to a novel anthelmintic Aroian, Raffi V. (UMASS Medical School, Worcester, Worcester, MA, USA) Cry5B is a natural Crystal protein produced by the soil-‐bacterium Bacillus thuringiensis and has recently been explored as a powerful, new anthelmintic. Anthelmintic therapy for human intestinal nematodes (soil transmitted helminths) currently almost exclusively upon the benzimidazoles, which are being increasingly recognized as sub-‐optimal. There is increasing urgency for new classes of anthlemintics. Single and double-‐dose treatments with Cry5B have excellent efficacy against hookworm (Ancylostoma ceylanicum) and Ascaris infections in laboratory hosts. Cry5B thus represents a promising new class of anthelmintics. Here, we explore immunological aspects of Cry5B treatment. In particular, given that it is a foreign bacterial protein with the potential to elicit an immunological response, here we test the hypothesis that oral treatment of animals with Cry5B might elicit an immune reaction that prevents its efficacy in subsequent applications. 56 A theoretical exploration of the evolutionary potential in a two-‐step infection processe in coevolving hosts and parasites Dybdahl, Mark (Washington State University, Pullman, WA, USA); Nuismer, Scott (University of Idaho) Traditional host-‐parasite coevolutionary models, which assume infection depends on molecular recognition and major genes, have been valuable, but we know little about coevolution of common and complex multiple step infection processes. For example, in plant and animal innate immunity, recognition steps are followed by downstream effector steps that kill recognized parasites, the outcome depending on an escalatory molecular arms race (reviewed in Dybdahl et al Am Nat 2014). We developed a coevolutionary model to predict levels of genetic diversity and local adaptation for each step of this common infection process in hosts and parasites. We found that levels of polymorphism were much higher at recognition loci than effector loci, and that levels of variation explained by host-‐ genotype by parasite-‐genotype (Gh x Gp) interactions were higher for the recognition step. Finally, we found that Gh x Gp variation contributed more to local adaptation for the recognition than the effector step. These results suggest that (1) local adaptation is more likely when fitness measures are related to recognition (infectivity) versus effectors (virulence), (2) effector loci, while functionally important, are less likely to harbor Gh x Gp variation that fuels coevolution, and (3) recognition loci are better candidates for genomic hotspots of coevolution. Cancelled Abstract: Bone marrow resident NK cells prime monocytes for regulatory function during infection Askenase, Michael (University of Pennsylvania/NIH Partnership, Bethesda, MD, USA); Han, Seong-‐Ji; Byrd, Allyson; Morais da Fonseca, Denise; Bouladoux, Nicolas; Wilhelm, Christoph; Konkel, Joanne; Hand, Timothy; Lacerda-‐Queiroz, Norinne ; Su, Xin-‐Zhuan; Trinchieri, Girogio; Grainger, John; Belkaid, Yasmine Tissue-‐infiltrating Ly6Chi monocytes play diverse roles in immunity, ranging from pathogen killing to immune regulation. A number of recent studies have demonstrated that local factors play an important role in determining the functional role of monocytes upon tissue recruitment, but the effects of systemic signals on their function remains poorly understood. We recently reported a previously unappreciated dual activity of Ly6Chi monocytes in controlling pathogen expansion while limiting immunopathology in response to commensal microbes during acute intestinal Toxoplasma gondii infection. These cells ensure host survival via production of the regulatory mediator Prostaglandin E2 (PGE2). Given the importance of a rapid and appropriate monocyte response, we hypothesized that systemic signals emanating from mucosal tissue early during infection induce these unique regulatory features in monocytes prior to tissue recruitment. To address this question, we infected mice per-‐orally Toxoplasma gondii. Early post-‐ infection when the parasite is limited to the intestine and its draining lymph, we found that Ly6Chi monocytes and their precursors in the bone marrow and blood acquired a previously undescribed MHCIIhiSca-‐1hiCX3CR1lo phenotype and produced significantly more PGE2 and IL-‐10 after stimulation than those from naïve mice. Acquisition of regulatory function by monocytes was the result of changes to the transcriptional profile of myeloid progenitors in response to IFN-‐g produced by bone marrow resident NK cells. Notably, this IFN-‐g production was not stimulated by local factors in the bone marrow, but rather by IL-‐12 produced by DC in the mucosal-‐associated lymphoid tissue (MALT). These results suggest that bone marrow resident lymphocytes may act as an alarm system responding to signals from infected distal tissues to pre-‐emptively control function of effector myeloid cells Participant List Last Name Allen First Name Judith Company Name University of Edinburgh City Edinburgh Country United Kingdom United Kingdom United States India E-Mail Address j.allen@ed.ac.uk Amanfo Seth The University of Edinburgh Edinburgh Aroian Raffi Worcester Arora Kashika Askenase Michael Balster Katja Bock Cristin Brodskyn Cláudia Budischak Sarah University of Massachusetts Medical School Motilal Nehru National Institute of Technology University of Pennsylvania/NIH Partnership Institute of Immunology, Department of Veterinary Medicine, Freie Universität Berlin Institute of Immunology, Freie Universität Berlin Fiocruz/Centro de Pesquisa Gonçalo moniz Princeton University United States Germany michael.askenase@nih.gov Berlin Germany cristin.bock@fu-berlin.de Salvador Brazil brodskyn@bahia.fiocruz.br Athens GA United States United States Canada Canada United States United Kingdom United Kingdom United Kingdom United States U.S.A. Canada United States United States United Kingdom Belgium U.S.A. Canada sabudischak@gmail.com Burger Elise UT Southwestern Dallas TX Burnett Burrows Camejo Elton Kyle Ana Institute of Parasitology, McGill University of British Columbia Cell Press Montreal Vancouver Cambridge BC MA Campbell Laura University of Manchester Manchester Capewell Paul University of Glasgow, UK Glasgow Capuccini Barbara London Carbajal Edwin Carvalho Chenery Christensen Augusto Alistair Stephen The Francis Crick Institute, Mill Hill Laboratory National Institute of Allergy and Infectious Diseases NIH University of British Columbia University of Maryland Christian David Clerc Melanie Cnops COELHO Dagenais United States Brazil Belgium ldaws@mail.med.upenn.edu United States United States Switzerland delongj@mail.med.upenn.edu United States United dybdahl@wsu.edu MA Allahabad Bethesda MD Berlin Washington DC Rocvkville Vancouver College Park MD BC MD University of Pennsylvania Philadelphia PA Edinburgh Jennifer CAMILA Maude University of Edinburgh, Institute of Evolutionary Biology Vrije Universiteit Brussel GEORGETOWN UNIVERSITY McGill University Dawson Lucas University of Pennsylvania de Oliveira De Trez Camila Carl Salvador-BA Brussels DeLong Jonathan CPqGM-FIOCRUZ VIB Department of Structural Biology, Laboratory of Cellular and Molecular Immunology University of Pennsylvania Philadelphia PA Dominguez Mariana The George Washington University Washington DC Dubey Lalit Kumar Lausanne Dybdahl Mark Ecole Polytechnique Fédérale de Lausanne Washington State University Pullman WA Easton Alice National Institutes of Health Bethesda MD brussel WASHINGTON Ste Anne de Bellevue Philadelphia DC PQ PA s.a.amanfo@sms.ed.ac.uk raffi.aroian@umassmed.edu kashikabt14@gmail.com k.balster@fu-berlin.de elise.burger@utsouthwestern.edu elton.burnett@mail.mcgill.ca kyle@brc.ubc.ca acamejo@cell.com laura.campbell-2@manchester.ac.uk paul.capewell@glasgow.ac.uk barbaracapuccini@gmail.com noexa17@gmail.com oliveira.fabiano@me.com chenery@brc.ubc.ca schrist1@umd.edu dac2@vet.upenn.edu m.clerc@sms.ed.ac.uk jcnops@vub.ac.be camila.coelho@yahoo.com maude.dagenais@mail.mcgill.ca camila@bahia.fiocruz.br carldetrez@yahoo.fr dizcobre@yahoo.com.br lalit.dubey@epfl.ch alice.easton@nih.gov Eren Fairfax Onur Keke University of Lausanne Purdue University Finney Ganusov Constance Vitaly University of Calgary University of Tennessee Epalinges West Lafayette Calgary Knoxville Garnier Romain Princeton University Princeton Ghosh Susmita Kolkata Gimblet Ciara Institute of Post Graduate Medical Education and Research University of Pennsylvania Philadelphia PA Glennie Nelson University of Pennsylvania Philadelphia PA Gonzalez Michael NYU Medical Center New York City NY Gov Lanny University of California, Irvine Irvine CA Graham Andrea Princeton University Princeton NJ Gregg Beth NIH/NIAID Bethesda MD Greischar GRENCIS Megan RICHARD University of Toronto UNIVERSITY OF MANCHESTER Toronto MANCHESTER ON Grigg Michael LPD/NIAID/NIH Bethesda Guimaraes B Costa Hamami Hamidzadeh Anderson NIAID/NIH Rockville MD Akil Kajal INRS - Institut Armand-Frappier University of Maryland Montreal College Park PQ MD Hamilton Phineas University of Victoria Victoria BC Harris Harris Nicola Tajie EPFL University of Virginia Lausanne Charlottesville VA Herbert DeBroski UCSF San Francisco Honeycutt Jared Stanford University Stanford CA Hopke Alex The University of Maine Orono ME Hsieh Michael Children's National Health System Washington DC Huang Lu Cornell University Ithaca NY Huang Stanley Washington University School of Medicine Saint Louis MO Huda Naureen National Institutes of Health Bethesda MD Hung Li-Yin UCSF San Francisco CA Hunter Christopher University of Pennsylvania Philadelphia PA Hurrell Benjamin University of Lausanne, Biochemistry Department Epalinges IN AB TN NJ States Switzerland United States Canada United States United States India remzionur.eren@unil.ch kfairfax@purdue.edu constance.finney@ucalgary.ca vitaly.ganusov@gmail.com romaing@princeton.edu susmita_ghosh2010@yahoo.co.in United States United States United States United States United States United States Canada United Kingdom United States United States Canada United States Canada cgimblet@mail.med.upenn.edu Switzerland United States United States United States United States United States United States United States nicola.harris@epfl.ch tajieharris@virginia.edu United States United States United States Switzerland naureen.huda@nih.gov nglennie@upenn.edu michael.gonzalez@med.nyu.edu lgov@uci.edu algraham@princeton.edu beth.gregg@nih.gov megan.greischar@utoronto.ca anna.fildes@manchester.ac.uk griggm@niaid.nih.gov andersoncbg@gmail.com akil.hamami@iaf.inrs.ca khamidza@terpmail.umd.edu phin.hamilton@gmail.com debroski.herbert@ucsf.edu jaredh@stanford.edu alex_hopke@umit.maine.edu mhsieh@cnmc.org lh346@cornell.edu shuang@pathology.wustl.edu HungL@medsfgh.ucsf.edu chunter@vet.upenn.edu benjamin.hurrell@unil.ch Iniguez Eva Department of Biological Sciences. The Border Biomedical Research Center. University of Texas at El Paso. El Paso TX, 79968, USA. el paso Jones Lucy University of Edinburgh Edinburgh Joyner Chet Malaria Host-Pathogen Interaction Center/Emory Vaccine Center/Emory University Atlanta Junqueira Caroline Centro de Pesquisas Rene Rachou/ FIOCRUZ Kauffmann Florence Kaye United States eainiguez@miners.utep.edu United Kingdom United States lucy.jones@ed.ac.uk Belo Horizonte Brazil carolinejunqueira@cpqrr.fiocruz.br Vrije Universiteit Brussel Elsene Belgium florence.kauffmann@gmail.com Paul University of York York paul.kaye@york.ac.uk Kelly Patrick Department of Microbiology, University of Iowa, Iowa City, IA 52246, USA Iowa City IA United Kingdom United States King Aaron University of Michigan Ann Arbor MI kingaa@umich.edu King Thayer Emory University Atlanta GA Kouassi Victor MRC National Institute for Medical Research Mill Hill United States United States United Kingdom Kurup Sam University of Iowa Iowa City IA samar@uga.edu Lamb Tracey Emory University School of Medicine Atlanta GA Landrith Tyler University of California, Riverside Loma Linda CA United States United States United States LeGrand Edmund College of Veterinary Medicine, University of Tennessee Knoxville TN United States elegrand@utk.edu Loke Png NYU School of Medicine New York NY png.loke@nyumc.org LopezYglesias Americo UT Southwestern Dallas TX United States United States MacLeod Annette University of Glasgow Glasgow annette.macleod@glasgow.ac.uk Magez Stefan Vrije Universiteit Brussel Brussel United Kingdom Belgium Mansfield John University of Wisconsin-Madison Madison john.mansfield@wisc.edu Matuschewski Kai MPI Infection Biology Berlin United States Germany McGovern Kathryn UC Riverside Riverside United States kmcgo001@ucr.edu TX GA WI CA cjjoyne@emory.edu patrick-h-kelly@uiowa.edu thayer.king@emory.edu vkouass@nimr.mrc.ac.uk tracey.j.lamb@emory.edu tyler.landrith@gmail.com americo.lopezyglesias@utsouthwestern.edu stemagez@vub.ac.be matuschewski@mpiib-berlin.mpg.de Mideo Nicole University of Toronto Toronto ON Canada nicole.mideo@utoronto.ca Mishra Bibhuti UND School of Medicine & Health Sciences Grand Forks ND United States bibhuti.mishra@med.und.edu Mosser David University of Maryland College Park MD dmosser@umd.edu Mou Zhirong University of Manitoba Winnipeg United States Canada Novais Fernanda University of Pennsylvania, School of veterinary medicine Philadelphia PA United States novais@vet.upenn.edu O'Brien Carleigh University of Virginia Charlottesville VA co2hh@virginia.edu Oliveira Fabiano NIH/LMVR Rockville MD Onyilagha Chukwunonso University of Manitoba Winnipeg MN United States United States Canada Orozco Jose University of Texas at El Paso El Paso TX jaorozco5@miners.utep.edu Osman Mohamed University of York York Owusu Christian Wellcome Trust Sanger Institute Hinxton PeronaWright Peters Georgia University of British Columbia Vancouver BC United States United Kingdom United Kingdom Canada Nathan Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases Bethesda MD United States ncpeters@ucalgary.ca Pinto Ana Isabel University of York York ana.pinto@york.ac.uk Portillo Susana The University of Texas at El Paso EL PASO Preham Olivier University of York York Prior Kimberley University of Edinburgh Edinburgh Pritchard Gretchen University of Pennsylvania Philadelphia PA Randolph Gwendalyn Washington University School of Medicine St Louis MO United Kingdom United States United Kingdom United Kingdom United States United States Redpath Stephen University of British Columbia Vancouver BC Canada sredpath@mail.ubc.ca Regli Ivo University of Lausanne Epalinges Switzerland ivo.regli@yandex.com Robertson Tanner University of Pennsylvania Philadelphia PA rotan@mail.med.upenn.edu Robinson Dionne NIH Bethesda MD ROMANO Audrey NIH/LPD Bethesda MD Rossi Matteo Department of Biochemistry, University of Lausanne Lausanne United States United States United States Switzerland TX mouzr@yahoo.com loliveira@niaid.nih.gov chukwunonso.onyilagha@umanitoba.ca mohamed.osman@york.ac.uk co7@sanger.ac.uk gperona@mail.ubc.ca sportillo7@miners.utep.edu olivier.preham@york.ac.uk k.f.prior@sms.ed.ac.uk gharms@mail.med.upenn.edu grandolph@path.wustl.edu dionne.robinson@nih.gov romanoa2@niaid.nih.gov matteo.rossi.2@unil.ch Sanecka Anna The Francis Crick Institute, Mill Hill Laboratory London United Kingdom anna.saneck-duin@crick.ac.uk Satti Mohamed Michigan Sate University Canton MI United States United States United States Brazil msatti@msu.edu Scorza Breanna University of Iowa Iowa City IA Scott Phillip University of Pennsylvania Philadelphia PA SILVA GRACE University of Massachusetts Worcester Silva Sasha INRS- Institut Armand Frappier Laval Canada silva.sasha@iaf.inrs.ca Sorci Gabriele CNRS Dijon France gabriele.sorci@u-bourgogne.fr Stager Simona INRS - Institut Armand-Frappier Laval PQ Canada simona.stager@iaf.inrs.ca Stevenson Stumhofer Mary Jason McGill University UAMS Montreal Little Rock AR mary.stevenson@mcgill.ca jstumhofer@uams.edu Lausanne Calgary Canada United States Switzerland Canada Sutherland Szabo Duncan Edina EPFL University of Calgary TacchiniCottier Tait Wojno Fabienne University of Lausanne Epalinges Switzerland fabienne.tacchini-cottier@unil.ch Elia Cornell University Ithaca NY edt42@cornell.edu Tam Mifong McGill University UC Irvine Montreal PQ UZONNA JUDE UNIVERSITY OF MANITOBA WINNIPEG United States Canada United States Canada Van Meulder Frederik Ghent University Ledeberg Belgium frederik.vanmeulder@ugent.be Vargas Venugopal Alejandro Gopinath CIDEIM-Colombia Institute of Immunology, Freie Universität, Berlin Cali Berlin Colombia Germany davargas@cideim.org.co gopinath.v@fu-berlin.de Viney Mark University of Bristol Bristol mark.viney@bristol.ac.uk Vogel Webb Ian Lauren McGill University University of Manchester Montréal Manchester PQ Weinkopff Tiffany University of Pennsylvania Philadelphia PA Worth Danielle University of California, Riverside Riverside CA United Kingdom Canada United Kingdom United States United States Yamamoto Masahiro Research Institute for Microbial Disases, Osaka University Suita Japan myamamoto@biken.osaka-u.ac.jp Yang Scarlett University of Pennsylvania Philadelphia PA scya@upenn.edu yarovinsky felix UT Southwetsern Dallas TX Zwicker Christian Institute of Specific Prophylaxis and Tropical Medicine, Medical University of Vienna, Vienna, Austria Vienna United States United States Austria PQ breanna-scorza@uiowa.edu pscott@vet.upenn.edu gksimuno@gmail.com duncan.sutherland@epfl.ch edina.szabo@ucalgary.ca mifong.tam@mail.mcgill.ca jude.uzonna@umanitoba.ca ian.vogel@mail.mcgill.ca lauren.webb@manchester.ac.uk wtiff@vet.upenn.edu worth.danielle@gmail.com felix.yarovinsky@utsouthwestern.edu christian.zwicker@meduniwien.ac.at
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