Current Perspectives on Vibrios in the Maryland Coastal Bays

Transcription

1 Food Science & Technology Program Current Perspectives on Vibrios in the Maryland Coastal Bays Salina Parveen, Ph.D. Professor Dept. of Agriculture, Food and Resource Sciences University of Maryland Eastern Shore Princess Anne, MD Food Science & Technology Program

2 Vibrio Family: Vibrionaceae Genus:Vibrio Species: 20 species, 12 pathogens 8 associated with seafood and the aquatic environment V. parahemolyticus, V. vulnificus, V. cholerae V. mimicus, V. alginolyticus, V. hollisae,v. furnissii, V. fluvialis Gram-negative non-spore forming curves rod, motile, ph (Oliver and Kaper 2007; Parveen and Tamplin 2013)

3 Vibrio parahaemolyticus & Vibrio vulnificus Gram-negative, halophilic, naturally occurring bacteria Raw shellfish (molluscs and crustaceans) are the main vehicle of transmission Most of the illnesses occur during the warmer months Seafood related illness and mortality (Oliver and Kaper 2001; CDC 2013; Parveen and Tamplin 2013)

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5 Vibrio parahaemolyticus & Vibrio vulnificus Total V. parahaemolyticus- a species specific marker thermolabile hemolysin gene (tlh + ) Pathogenic V. parahaemolyticus- pathogenicity markers two genes: thermostable direct hemolysin (tdh + ) thermostable-related hemolysin (trh + ) Total V. vulnificus - a species specific marker V. vulnificus hemolysin gene A (vvha) Clinical V. vulnificus - pathogenicity marker virulence correlated gene (vcgc) (Oliver and Kaper 2001; CDC 2013 ; Parveen and Tamplin 2013)

6 Vibrio parahaemolyticus A significant cause of bacterial seafood-borne illness U.S. CDC estimates ~34,664 Vibrio illnesses/year ~Common symptom-gastroenteritis with occasional bloody diarrhea Septicemia-individuals with underlying chronic illness (Oliver and Kaper 2001; Scallan 2011; CDC 2013; Parveen and Tamplin 2013)

7 Vibrio vulnificus Causes 95% of all seafood-related deaths Invasive and life-threatening diseases Healthy people Gastroenteritis (vomiting, diarrhea, and abdominal pain) within 16h of eating contaminated raw or undercooked seafood Immunocompromised individuals Blood stream infection [fever and chills, decreased blood pressure (septic shock) and blistering skin lesions] -50% fatal -Flesh eating bacterium Wound infection (skin breakdown and ulceration) (Oliver and Kaper 2001; Parveen and Tamplin 2013; CDC 2013)

8 Blue crabs (Callinectes sapidus) Blue Crabs and MCBs Maryland Coastal Bays 175 sq. miles watershed Commercial and Recreational blue crab fishery

9 Concerns Improperly cooked blue crabs Handling of live crabs Increase risk of Vibrio parahaemolyticus and Vibrio vulnificus infections for consumers Recent Death in Maryland

10 Occurrence of Vibrio parahaemolyticus & Vibrio vulnificus Chesapeake Bay Parveen et al., (2008) Total (tlh + ) and pathogenic (tdh + and trh + ) V. parahaemolyticus in oysters and water Jacobs et al., (2010) Ecological forecasting for V. vulnificus Dasilva et al., (2012)- Predictive modeling for V. vulnificus in post-harvest shellshock oysters Parveen et al., (2013)- Predictive modeling for V. parahaemolyticus in postharvest shellshock oysters Current study-vibrio, rapid method, metagenomics

11 Occurrence of Vibrio parahaemolyticus & Vibrio vulnificus Chesapeake Bay Vibrio spp. in blue crabs Fishbein et al., (1970) V. parahaemolyticus in processed blue crab meat Molenda et al., (1972)- First Vibrio parahaemolyticus outbreak in US, Maryland Sizemore et al., (1975) Isolated V. parahaemolyticus from blue crab

12 Data Gap No study on V. vulnificus in blue crab No study in the MCBs Chesapeake Bay study-four decades ago Since significant changes have occurred in the water quality of the Chesapeake Bay and MCBs in the past four decades

13 Objectives Determine the incidence of total and pathogenic V. parahaemolyticus, & total and clinical V. vulnificus blue crabs, water and sediments from four sites of the MCBs. Examine the correlation of V. parahaemolyticus and V. vulnificus levels in blue crabs, water and sediments with physico-chemical parameters.

14 Sampling Sites 8 Chincoteague Bay 9 Newport Bay 10 Sinepuxent Bay 13 St. Martin River Location of sampling sites in the Maryland Coastal Bays

15 Collection of Samples Sampling period February-October 2012 Water Master flex water pump YSI 6600 Sediment Petite Ponar grab Blue Crabs-Apr.-Nov. Otter trawl, crab pots Muscle and hemolymph Physico-chemical parameters

16 Methodology V. parahaemolyticus V. vulnificus Kaysner and DePaola 2004 Nordstrom et al Parveen et al Baker-Austin et al. 2010

17 Statistical Analyses One way analysis of variance (ANOVA) followed by post hoc Tukey test Vibrio spp. vs. site Vibrio spp. vs. month Spearman's Rank Correlation Coefficient (r) Vibrio spp. vs. physico-chemical parameters Log-transformation An alpha level of 0.05 Statistix 9

18 Average Surface Water Temperature, Salinity, Dissolved Oxygen and ph Feb Mar Apr May Jun Jul Aug Sep Oct Month Dissolved Oxygen Salinity Temperature ph Temp.: ⁰C; Salinity: 25-33ppt.; Dissolved oxygen: mg/l; ph:

19 C or ppt Total (tlh + ) V. parahaemolyticus in Crab, Hemolymph, Sediment and Seawater Crabs Hemolymph Sediment Seawater Temperature Salinity Log MPNg -1 or ml Feb Mar Apr May Jun Jul Aug Sep Oct 0 Positive samples=100%

20 C or ppt Pathogenic (tdh + ) V. parahaemolyticus in Crab, Hemolymph, Sediment and Seawater Crab Hemolymph Sediment Seawater Temperature Salinity Log MPNg -1 or ml Feb Mar Apr May Jun Jul Aug Sep Oct 0 Positive samples: 32% of crab; 45% of hemolymph, 28% of sediments and 39% of seawater

21 C or ppt Pathogenic (trh + ) V. parahaemolyticus in Crab, Hemolymph, Sediment and Seawater Crabs Hemolymph Sediment Seawater Temperature Salinity Log MPNg -1 or ml Feb Mar Apr May Jun Jul Aug Sep Oct Positive samples: 36% of crab; 35% of hemolymph, 25% of sediment and 8% of seawater 0

22 C or ppt Total (vvha) V. vulnificus in crab, hemolymph, sediment and seawater Crab Hemolymph Sediment Seawater Temperature Salinity Log MPNg -1 or ml Feb Mar Apr May Jun Jul Aug Sep Oct 0 Positive samples =100%

23 Clinical (vcgc type) V. vulnificus in Crab, Hemolymph, Sediment and Seawater Log MPNg -1 or ml Feb Mar Apr May Jun Jul Aug Sep Oct Month Crab Hemolymph Sediment Seawater Positive samples: 32% of crab; 25% of hemolymph, 5% of sediments and 11% of seawater

24 Conclusions V. parahaemolyticus and V. vulnificus varied throughout the year Did not show significant differences among the sites Pathogenic V. parahaemolyticus (trh + ) were not frequently present during the sampling period Variability in the levels of total and pathogenic V. parahaemolyticus, and total V. vulnificus was explained by four parameters These results highlight the potential for blue crab related vibriosis and the importance of proper handling, cooking, and care of this popular seafood before consumption

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26 Climate Variability and Microbial Dynamics in the Maryland Coastal Bays PI: P. Chigbu Sub-Project 3 Leader: S. Parveen Collaborators: J. Pitula, J. Jacobs, S. Chung, K. Kunkel Overall goal Spatial, seasonal and inter-annual variations in the occurrence of heterotrophic bacterioplankton,vibrio parahaemolyticus and V. vulnificus in water and blue crabs in relation to biotic and abiotic factors in order to improve our understanding of their dynamics in the Maryland Coastal Bays (MCBs)

27 Specific Objectives Investigate the environmental occurrence of heterotrophic bacterioplankton including V. parahaemolyticus and V. vulnificus Develop predictive models of V. parahaemolyticus and V. vulnificus Investigate the broader microbial community in blue crab and water samples using 16S rrna genes Explore the role of commensal Vibrio spp. as stressors of blue crab immunological responses, and their influence on blue crab disease

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31 High Salinity Relaying Wet overboard study (Field study) Virginia:xxx Maryland Low Salinity (xxx) High Sailinity (xxx)

32 High Salinity Relaying Chill Tank Study: VT

33 Changes of V. parahaemolyticus and V. vulnificus levels (log MPN/g) in Oysters During Relaying in Water High Salinity V. parahaemolyticus (mean±standard error) V. vulnificus (mean±standard error) Day Trial 1 Trial 2 Trial 3 Trial 4 Trial 5 Day Trial 1 Trial 2 Trial 3 Trial 4 Trial ± ± ± ± ± ± ± ± ± ± a a a a a 0 a a a a a ±0.17 ab 2.67±0.51 a 4.39±0.16 ab 4.17±0.19 a 2.41±0.14 b ±0.23 b 2.77±0.27 bc 3.04±0.09 b 3.73±0.35 b 3.29±0.18 b ±0.17 b 3.83±0.51 a 3.94±0.16 bc 2.91±0.19 b 2.27±0.14 b ±0.23 bc 3.06±0.27 ab 2.74±0.09 b 1.84±0.35 c 2.31±0.15 c ±0.17 b 2.24±0.51 a 3.58±0.16 c 2.91±0.19 b 2.17±0.14 b ±0.23 c 1.85±0.27 c 1.52±0.09 c 1.64±0.35 c 1.83±0.15 c Values are mean of three replicates (Trials 1-4 in MD) Values are mean of ten replicates (Trial 5 in MD) Means with the same letter in the same column are not statistically significant (P>0.05)

34 Acknowledgements UMES Ms. Candace Rodgers Dr. Paulinus Chigbu Dr. Jeannine Harter-Dennis Ms. Sylvia Ossai Mr. Chris Daniels Dr. Andrés G. Morales-Núñez Dr. Ligia DaSilva NOAA Dr. John Jacobs Mr. Matt Rhodes Funding NSF CREST NOAA LMRCSC USDA CBG

35 QUESTIONS