Monitoring of Vibrio cholerae in Environmental Reservoirs in Haiti

1. Monitoring of Vibrio choleraein Environmental Reservoirs in Haiti Meer TaifurAlam Emerging Pathogens Institute University of Florida

2. Introduction • Cholera appeared in epidemic form in Haiti in 2010 for the first time in over 100 years and has continued to occur in subsequent years • As of January 2014, about 698,000 cases and more than 8,500 deaths have been reported • It is essential to understand patterns of ongoing cholera transmission and seasonality of cholera in Haiti and to assess the likelihood of future epidemics • This raises the question of whether the microorganism has established in environmental reservoirs in Haiti • If so, which environmental factors enhance the survival of the bacterium?

3. Objectives • Selection of environmental sites to monitor V. cholerae • Observation of the physical parameters of environmental reservoirs and their correlation to the occurrence of V. cholerae • Isolation of V. choleraefrom environmental reservoirs • Genetic characterization of the isolated V. cholerae

4. Sample Collection Sites • Fifteen fixed environmental sites along the transects of 3 rivers in the Gressier/ Leogane region including one independent estuarine site • MomanceRiver (4 up-river sites and one estuarine site) • GressierRiver (4 up-river sites and one estuarine site) • Tapion River (4 river sites) • Four-a-chaux (an independent estuarine site) • All sites were at least a half-mile apart, with the exception of the Christianvillebridge and spring sites, which were 0.25 miles apart • Samples were collected on a monthly basis from each site between April, 2012 and March 2013

5. Sample Collection Sites

6. Physical Parameters • Temperature • pH • Dissolved oxygen (DO) • Total dissolved solids (TDS) • Conductivity • Salinity

7. Physical Parameters Results

8. Correlation of V. choleraewith Temperature

9. Bacteriological Assays • Conventional culture and isolation of V. cholerae • Serology to confirm V. cholerae O1 • PCR assay to identify Vibrio specific genes • Aerobic plate count

10. Isolation of V. cholerae from EnvironmentApril 2012-March 2013

11. Distribution of V. choleraeO1

12. Distribution of V. choleraenon-O1/non-O139

13. PCR Analysis of V. choleraeGenes

14. PCR Analysis of V. choleraeGenes ** A shrimp isolate

15. Conclusions • Vibrio choleraeO1 and non-O1 were successfully isolated from the environment in Haiti • Isolation of non-O1 strains in environmental reservoirs suggests that conditions are appropriate for establishment of O1 strains at these same sites • Novel Vibrio choleraeO1 type biotype El Tor strain was isolated which lost its entire ctxphage • Water temperature emerged as statistically significant in a conditional logistic regression model

16. Future Goals • Development of new molecular beacon hybridization probes for the identification of V. cholerae • Perform a more sensitive colony blot assay and correlate the results with culture data and physical parameters • Develop a new enrichment technique for V. cholerae isolation • Introduce membrane fecal coliform counts (mFC) to observe correlation between mFC and V. cholerae

17. Acknowledgments Afsar Ali, PhDValery Madsen De Beau Rochars, MDJ. Glenn Morris Jr., PhD (EPI Director )Christianville FoundationHaitian laboratory personnel (UF-EPI Haiti Lab 1-Gressier)This work is supported by: NIH grant RO1 AI097405 (Drs. Morris and Ali) NIH grant RO1 AI3929 (Drs. R. Bradley Sack, John Hopkins University and Ali)DoDgrant C0654_12_UN (Dr. Ali)

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