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C I T R U S A D VA N C E D T E C H N O L O G Y P R O G R A M
QUARTERLY & FINAL REPORTS: Control of Citrus Greening, Canker & Emerging Diseases of Citrus
INSTRUCTIONS
Quarterly Report
Annual Report
Final
Required: What is the “headline” for this report (e.g. a one-sentence “newspaper headline” describing what you accomplished)
Beneficial bacteria in management of HLB
Proposal Title
Characterize the effect of application of beneficial bacteria (Microbe Program) on management of
Huanglongbing
Today’s Date
4/2/15
Sponsoring Organization (drop-down)
Citrus Research and Development Foundation
Category (drop down)
Unclassified
REPORT UPDATE (500 words; summarize your accomplishments )
The goal of the proposed study is to characterize the effect of application of beneficial bacteria (MICROBE Program) on
management of HLB. Currently, we are setting up the experiments to test different Microbe Products in management of
HLB. Assay for compatibility between isolates using antagonistic survival tests showed that all the selected beneficial
bacteria are compatible with each other. Plant growth promoting activity of six selected isolates was evaluated using the
model plant Arabidopsis grown in vitro. The results suggested that three isolates could promote plant growth.
The plant growth promoting activity of these six isolates was tested using citrus (grapefruit) seedlings in greenhouse.
Greenhouse assays suggested that a consortium of three Bacillus and relative isolates (AY16, PT6 and PT26A) may delay
the development of both HLB symptoms and pathogen population on citrus leaves after root inoculation. The potential of
the consortium to recover the tree decline from HLB infection is being evaluated in greenhouse. The growth conditions of
the three strains were optimized using a small fermenter. Three antifoam agents, A204, PPG200 or M-Oil did not affect the
growth of the three bacterial strains. The initial neutral to alkaline pH values (7.0 ~ 8.0) favor growth of the three bacteria
in LB, while acidic pH (5.0 ~ 6.0) suppress bacterial growth. The optimal cultural temperature was determined to be
around 30C with average bacterial population of 109-1010 cfu/ml after 20-hour incubation, although the bacteria may grow
slowly under room temperature (~ 23C). The shelf life of three different formulations of the bacterial culture is being
evaluated under room temperature. In a six-month time course, the bacterial populations in LB broth, OPB broth and tape
water are comparatively stable with initial and final both at ~ 108cfu/ml. Under room temperature, after a 20 month
storage, the population reduced about 10E5 to 10E6 folds.
The field trials for estimation for bacterial activity improving the health of HLB diseased trees continued as scheduled. We
completed 10 applications through soil drench in the two groves at LAKE WALES: 07/2013, 09/2013, 11/2013, 01/2014,
03/2014, 05/2014, 07/2014, 09/2014 11/2014 and 02/2015 (every 2-3 months); conducted 5 surveys of HLB disease
severity and collected leaf samples for Las population analysis using qPCR (03/2013, 09/2013, 03/2014, 09/2014 and
03/2015 (every 6 months)). The data collected suggested that after 10 applications (19 months after 1st application), the
three bacteria treatment showed a reduced HLB disease severity with lower values of standard area under the disease
progress stairs (sAUDPS) and Las bacterial populations in leaf samples as compared with the non-treated control in one
trial that contains relative younger trees with mild HLB symptoms when the experiment was initiated. We will collect data
for four trials. We will continue to conduct the field trial to analyze the cost of the treatment compared to the control effect.
PI First Name Nian
Organization Citrus Research and Education Center
PI Last Name Wang
Contract Number 608
Email nianwang@ufl.edu
Project Duration (years) 3
Phone 863-956-8828
Total Funds (current year) $133,286.00
Year of Project 2
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