Vibrio parahaemolyticus Levels in Atlantic Coast Shellfish

Vibrio parahaemolyticus Levels in Atlantic Coast Shellfish

ISSC, VARB & Other Vp Research Jessica L. Jones, Ph.D. FDA Gulf Coast Seafood Laboratory Dauphin Island, Alabama NY/CT Reopening Vibrio levels higher in oysters than clams from the same harvest area. SC Vibrio Levels in Summer Vibrio levels differ based on shellfish type/storage

condition. AL Resubmerging (Aquaculture) Vibrio levels return to background after 14d resubmerging following routine desiccation. Auburn Univ. Resubmerging Variation was seen amongst the trials, but Vibrio levels returned to background after 7d of resubmergence. WA Intertidal

Regardless of container, vibrio levels returned to background after one tidal cycle. NJ Intertidal Vibrio levels returned to background after the second day, following an initial increase postsorting. AL Handling Practices Vibrio levels increased

after 5h of ambient storage, especially under refrigeration compared to ice slurry. NJ Handling Practices Vibrio levels increased after 5h and 7h of shaded storage. NJ Handling Practices After 5h of ambient exposure, Vibrio levels increase, more so with mechanical refrigeration.

WA Handling Practices Variation was seen amongst trials, but 5h refrigerated and both 9h treatments showed increased Vibrio levels. CT Handling Practices After 5h ambient

exposure, Vibrio levels increased significantly. Duxbury Bay, MA total: 85/114; 21.55.1 log MPN/g tdh: 26/113; 5.01.9 log MPN/g trh: 40/113; 7.82.9 log MPN/g Katama Bay, MA total: 75/104; 23.612.6 log MPN/g tdh: 19/100; 28.251.2 log MPN/g trh: 25/98; 20.237.7 log MPN/g

Barnstable Harbor, MA total: 20/29; 100.011.0 log MPN/g tdh: 5/29; 23.619.5 log MPN/g trh: 6/21; 28.87.3 log MPN/g Massachusetts Total Vp tdh MA D

K B MA D Temp. + +

+ + + + Sal. none + none

+ none Depth - - none none pH

- Turb. Chl. trh MA D K B none none

+ + + none none none none none

none none none none none none none - none none none none none none none

none none none none none none none none none none none none none

none none none none none none none + none none + +

+ none none K B + none none Correlations with environmental parameters is dependent on specific harvest location.

In CT, bottom temperature and Vp levels show a strong correlation. WA Substrate Study 4.0 3.5 A seasonal difference was noted, but also harder substrates had higher levels of Vibrio.

3.0 2.5 Log Vp (CFU/g) 2.0 1.5 Regular Mud Substrate Type 2015

2016 Gravel WA Depuration Temperature affects purging in ASW, but not with EO water. WA Depuration Naturally contaminated oysters in ASW at 5C reduce Vp levels ~3 logs in 3 days. AL Relaying for Vibrio Reduction After an increase in Vibrio levels upon initial resubmergence, Vibrio

decreased after 7 days. WA Deep Water Purging Cold, deep water purging shows promise for reducing Vibrio levels. Johnson et al 2012 VARB

Developed in 2014 to address the need to standardize how FDA responds to external requests for assistance Does not apply to internal requests which require only minimal CFSAN resources This process is also not intended to supersede FDAs ability and willingness to respond to emergency situations and requests that arise due too such emergencies VARB Process States submit requests via their Specialist to CFSAN (Stacey Degrasse, Chair) Requests can be submitted at any time, but cut off dates for quarterly review meetings are: March 31

June 30 September 30 December 31 VARB Process (cont) Submissions ranked at quarterly meetings based on specific evaluation criteria: Programmatic impact FDA capability and capacity Requestor capability and capacity Leveraged resources of requestor Sustainability Final rankings, decision to support, along with specific feedback are provided to requestors and ISSC

F14-WA-02: Icing Study Triplicate samples collected from eight treatments Immediately ice- Initial harvest 1h ambient, then iced - 1h refrigerated 5h ambient, then iced - 5h refrigerated 9h ambient, then iced - 9h refrigerated Experiment replicated five times between 1- Aug 29 Jul F14-WA-02:Preliminary Results

ISSC - MA 2014-2016 weekly collection data Intertidial collection within 1h of exposure Stored on ice, analyzed same day or next Method: NSSP (MPN-PCR) for total, tdh, trh Samples with temp abuse in transit omitted; <3 considered negative

ISSC - WA Influence of shellfish bed substrate on rate of growth of Vibrio parahaemolyticus in oysters h 2015-2016, V. parahaemolyticus concentrations were analyzed in oysters harvested from three different substrate types rocky (gravel), muddy and a mixture of sand and mud (regular) within each growing area harvested from Washington State. Oakland Bay, Samish Bay and Hood Canal 5 Method: WA MPN-PCR for total, tdh??? ISSC - WA Influence of shellfish bed substrate on rate of growth of Vibrio parahaemolyticus in oysters h 2015-2016, V. parahaemolyticus concentrations

were analyzed in oysters harvested from three different substrate types rocky (gravel), muddy and a mixture of sand and mud (regular) within each growing area harvested from Washington State. Oakland Bay, Samish Bay and Hood Canal 5 Method: WA MPN-PCR for total, tdh??? Wa Laboratory assessment of the relation between water temperature, immersion time and exposure to bactericidal oxidation products on Vibrio parahaemolyticus levels in Pacific oysters 5 strain cocktail; BAM MPN-culture (no confirmation) Ren T, Su Y-C. 2006. Effects of electrolyzed oxidizing water on reducing Vibrio parahaemolyticus and

Vibrio vulnificus in raw oysters. J. Food Prot. 69:18291834. ISSC CT (2016) Techniques and Practices for Vibrio Reduction: Connecticut Methods: NSSP for total, tdh, trh 1) Zero (0) Hour (Baseline): Immediate post-harvest rapid cooling to internal temperature of 50F (10C) or less using ice slurry, and 2) One (1) hour from harvest to internal temperature of 50F (10C) or less using ice slurry (45 minutes on deck then into slurry for 15 minutes rapid cooling), and 3) Three (3) hours from harvest to internal temperature of 50F (10C) or less using ice slurry (two (2) hours 45 minutes on deck prior to slurry for 15 minutes), and 4) Five (5) hours from harvest to internal temperature of 50F (10C) or less using ice slurry (four (4) hours 45 minutes on deck prior to slurry for 15 minutes), and 5) NSSP standard VPCP: Five (5) hours from harvest into mechanical refrigeration at or below 45F (7.2C) and maximum of ten (10) hours to an internal temperature of

50F (10C). Diff between mechaical refrig and ice slurry w/ in 1 and 3 h Group Name 0Hr 1Hr 3Hr 5Hr 5/10 N 13 7 7

13 11 Mean 1.908 1.98 2.201 2.581 2.918 Std Dev 0.51 0.585 0.308 0.582

0.924 SEM 0.141 0.221 0.117 0.162 0.279 All Pairwise Multiple Comparison Procedures (Fisher LSD Method): Comparisons for factor: Process Study Code Comparison Diff of Means

LSD(alpha=0.050) P Diff >= LSD 5/10 vs. 0Hr 1.01 0.521 <0.001

Yes 5/10 vs. 1Hr 0.938 0.615 0.004 Yes 5/10 vs. 3Hr 0.717

0.615 0.023 Yes 5/10 vs. 5Hr 0.337 0.521 0.199

No 5Hr vs. 0Hr 0.673 0.499 0.009 Yes 5Hr vs. 1Hr 0.601

0.596 0.048 Yes 5Hr vs. 3Hr 0.38 0.596 0.206

No 3Hr vs. 0Hr 0.294 0.596 0.327 No 3Hr vs. 1Hr 0.222

0.68 0.515 Do Not Test 1Hr vs. 0Hr 0.072 0.596 0.809

Do Not Test ISSC AL (2012) Test of Effectiveness of Relaying as a Post Harvest Process for reducing levels of Vibrio vulnificus and V. parahaemolyticus in Shellstock Oysters Immediate harvest, refrigerated within one hour (white tag), unrefrigerated (green tag) until landing Methods: BAM MPN-culture (gene probe) Day Sandy Bay

Temp (C) Dauphin Island Salinity (PSU) Temp (C) Salinity (PSU) 29.1 25.3 30.2

22.6 7 29.8 26.9 30.8 27.6 14

28.2 27.5 30.2 29.0 2 28.5 21.2 27.1

27.8 7 26.8 22.3 27.4 26.8 14

22.6 23.7 21.6 22.1 August Run 2 Sept. Run parahaemolyticus V. vulnificus (+ SD) V.

(+ SD) Immediately Iced (t0) 11,240 (+ 6,562) 318 (+ 308) White Tagged (t0WT) 11,730 (+ 10,734)

1,066 (+ 1,391) Green Tagged (t0GT) 67,600 (+ 42,426) 3,761 (+ 3,785) ISSC PSI (2017) Techniques and Practices for Vibrio Reduction 2014-2016 Methods: BAM (AmTest Laboratories) for total

ISSC NJ (2015) Techniques and Practices for Vibrio Reduction Use of Shading and Rapid Cooling (ice slurry) to Control Vibrio Growth Methods: NSSP, but LC480 total, tdh, trh A N Collect 15 -20 oysters, immediately place on ice, transport, shuck, and analyze. Do during weeks of (05/18/15, 06/15/15, 06/22/15, 07/13/15, and 08/17/15). Lab processes on arrival during those weeks! Baseline/Zero Hour: Immediate rapid cooling of 15 20 harvested oysters for 10 minutes to meat temperature of 10C or 50 F or less using ice slurry! Take shell & meat temps after slurry. Place on ice after slurry. Lab processes on arrival.

O 1 hour from harvest of 15 20 oysters to meat temperature of 10C or 50 F or less using ice slurry (50 min on deck in shade then into slurry for 10 minutes rapid cooling)! Take shell and meat temps. after slurry. Place on ice after slurry. Lab processes on arrival. P 3 hours from harvest of 15 20 oysters to meat temperature of 10C or 50 F or less using ice slurry (2 hours 50 min on deck in shade then into slurry for 10 minutes rapid cooling)! Field crew/lab take shell and meat temps after slurry on P, as slurry might be done at lab! Collect and keep shaded for 2 hours and 50 minutes only; then place in slurry, and if needed, follow w/ ice. Lab processes on arrival or after slurry. Q

5 hours from harvest of 15 20 oysters to meat temperature of 10C or 50 F or less using ice slurry (4 hours 50 min on deck in shade then into slurry for 10 minutes rapid cooling)! Field crew/lab take shell and meat temps after slurry on Q, as slurry likely done at lab. Collect and keep shaded for 4 hours and 50 minutes only, then place in slurry, and if needed, follow w/ ice. Lab processes on arrival or after slurry process at lab. R NSSP standard VPCP: Shade 15 20 oysters for 5 hours from harvest then into traditional mechanical temperature control for 10 hours to an internal temperature of 10 C or 50 F. Lab takes shaded product, places in refrigerator for 10 hours of cooling and then lab takes pre-process shell and meat temperature and processes next day! Taylor Shellfish

Our methods were straight forward and simple. Identify oysters with elevated levels during vibrio season, relay them in to growing trays stocked at grow out densities and hang them below rafts in water deep enough to reach the 12 15 C range while sampling at intervals to measure the changes in Vp levels. Methods: ?? Apparent reduction across board, with all lots meeting then Canadian standards (n=5 sampling with only 1 result >100 MPN) for oysters intended for raw consumption. Some lot samples showed a low Zero hour and then suggested an increase over the first few days, but

this is likely an indication of inadequate sampling at the zero hour and/or the inconsistency in Vp loads at the zero hour. Regardless of zero-hour levels all lots showed a significant reduction by day 7. Points of contact Dr. Stacey Degrasse, VARB Chair [email protected] Dr. Jessica Jones, VARB Vice-Chair [email protected]

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