Bacterial Emerging Risks to Seafood Safety WORKSHOP "SEAFOOD AND EMERGING FOOD SAFETY ISSUES"

Transcription

1 Bacterial Emerging Risks to Seafood Safety WORKSHOP "SEAFOOD AND EMERGING FOOD SAFETY ISSUES"

2 Populations dynamics and diseases Two Models: Salmonella Vibrio V. cholerae V. parahaemolyticus Foodborne pathogen Waterborne pathogen

3 Analysing disease emergence using a combination of approaches Epidemiology and risk Remote sensing Molecular biology Oceanography & climate science

4 Pathogenic vibrios present in the marine environment V. vulnificus V. alginolyticus Gram negative bacteria common in marine and estuarine environments. V. parahaemolyticus All these species proliferate in warm (>15 C) and low salinity (<30 ppt NaCl) seawater. V. cholerae Complex life cycle: planktonic and attached to marine organisms

5 Different clinical manifestation for vibriosis 1. Wound infections. Caused by a range of different Vibrio spp. including V. vulnificus, V. alginolyticus, V. cholerae normally caused by exposure of cut, wound or abrasion to contaminated seawater. Seriousness of infection is partly determined by species in question V. vulnificus probably the worst ~25% mortality rate (Oliver 2005). 2. Gastroenteritis. Nausea, vomiting, diarrhoea - again, caused by a range of different Vibrio spp. most commonly V. vulnificus, V. parahaemolyticus and V. cholerae normally caused by consumption of raw/undercooked seafood and/or exposure to contaminated water sources. 3. Septicaemia (blood poisoning). Most serious clinical manifestation associated with vibriosis. Often fatal, depending on pathogen involved (V. vulnificus > 50% of cases). 100 Vibrio fatalities a year in the USA, mostly septicaemia-associated, numerous recent cases in Europe.

6 Vibrio parahaemolyticus Vibrio parahaemolyticus is the leading cause of bacterial seafood-associated gastroenteritis worldwide (including the USA). Leading cause of infectious diarrhoea in South East China. Causes gastroenteritis, and very rarely wound infections. Most common bacterial infection associated with the consumption of seafood. Pathogenic strains normally carry one of two established markers (hemolysin genes, tdh and trh). Very rarely strains carry both genes. In Europe all clinical cases tend to be related to tdh+ isolates. We now routinely pick up these strains in UK shellfish produce during the summer. Numbers of infections appear to be increasing in Europe several outbreaks related to warm weather conditions have been reported.

7 Vibrio model: ecological preferences

8 Vibrio model: infections and seawater temperature

9 The life cycle of pathogenic Vibrio

10 Transmission of Vibrio Environment (Seafood) Primary source infections

11 Vibrio parahaemolyticus

12 Vibrio parahaemolyticus

13 How can pathogenic vibrios move long distances? Ballast water Ships take up marine water, and transport it long distances as ballast. This is a well established mechanism for the introduction of exotic bacteria into new areas. Cholera was detected in Gulf of Mexico in early 1990 s after it was transported from an endemic region.

14 no. cases 1999 Galicia (Spain) Vibrio parahaemolyticus infections Rest of Spain Galicia Total Rest of Spain Galicia Total

15 Galicia 1999 First Large Outbreak of V. parahaemolyticus Vigo Date Cases Hospitalized Raw Oyster Source: Boletín Epidemiológico Gallego

16 Galicia Investigation Serotypes Vigo (2), Madrid, Tarragona Getafe O4:K11 O4:KUT Caracterization PFGE Vigo Tarragona Vigo Madrid Getafe

17 Vibrio parahaemolyticus 1999 Galicia (Spain)

18 Vibrio parahaemolyticus Pandemic O3:K6 clone Emergence: Indonesia, 1995 Epidemic expansion: India, 1996

19 Vibrio parahaemolyticus Calcutta, India Feb Nov-97 Antofagasta, Chile Nov--97 Countries affected by the pandemic spread between

20 Vibrio parahaemolyticus in Peru 40 Vp strains remited to the INS Austral winter July-1997 Cases Cases Pandemic 309 records seasonal pattern # strains O4:K8 O3:K6 O4:K8 Transition may sep may sep may sep may sep may sep Date may sep may sep may sep may sep may sep

21

22 Vibrio parahaemolyticus in Peru 2 months 2 months

23 Vibrio parahaemolyticus in Peru

24 El Niño El Niño is a disruption of the ocean-atmosphere system in the Tropical Pacific having important consequences for weather and climate around the globe.

25 Indonesia South America Pacific Ocean V. parahaemolyticus New Clone 2009 V. parahaemolyticus Pandemic Clone 1997 V. parahaemolyticus O4:K Cholera 1991

26

27 How can pathogenic vibrios move long distances? Ocean currents. Bacteria like vibrios are often found in close association with plankton, and can move large distances via oceanic currents. Largescale movements like El Nino is believed to have been responsible for the emergence of cholera into South America in Possible route for 2012 outbreak? Can bacteria survive the journey though? Ballast water. Ships take up marine water, and transport it long distances as ballast. This is a well established mechanism for the introduction of (IPCC fourth assessment report 2007). Vigo exotic bacteria into new areas. Cholera was detected in Gulf of Mexico in early 1990 s after it was transported from an endemic region. Both Vigo and New York are very large and busy ports. NY

28 Pacific Northwest, 1997

29 Pacific Northwest, 1997

30 Pacific Northwest, 1997

31 ALASKA 2004

32

33 Spanish outbreak, August 2012 Gastroenteritis outbreak on 18 th August 2012, in Galicia, NW Spain. A total of 58 people were interviewed, 44 affected and 14 without symptoms. A key characteristic of this outbreak is the high attack rate: 114 people attended the banquet, 100 people reported feeling unwell. Nine stool samples were collected and subjected to analysis and all the cultures were positive for Vibrio parahaemolyticus. 33

34 New York Spanish outbreak, August 2012 Serotype Virulence genes PFGE PFGE profiles O4:K12 tdh trh PCR

35 Spanish outbreak, August 2012 O Grove, Galicia, NW Spain Celcius Shellfish beds August 13th Celcius Celcius August 6th August 22nd Source: Trinanes, L., Martinez-Urtaza, J, and C.Baker-Austin. et al. (2013) unpublished data

36 New York outbreak, summer 2012 During April-June 2012, an outbreak of shellfish associated gastroenteritis was reported in the USA, focussed on produce from New York Harbour (NYH). U.S. Centers for Disease Control and Prevention (CDC) linked 28 sporadic cases to the consumption of raw oysters (57%) and clams (22%) harvested from Oyster Bay, NY between April and August, Strains were identified as V. parahaemolyticus. Rather significantly, all strains both tdh+/trh+, and highly similar to Pacific Northwest strains. Very occasionally Pacific strains have been implicated in infections elsewhere in mainland USA.

37 New York outbreak, summer 2012 New York NYH site Source: Joaquin Trinanes (2013)

38 New York Galicia V. parahaemoltycius outbreak, summer 2012 New York outbreak, summer 2012 Two V. parahaemolyticus outbreaks, 3,000 miles apart. Analysis of isolates using serology (both O4:K12), PCR, and pulse field gel electrophoresis suggest these may be the same strains. PFGE profiles PFGE profiles tdh PCR trh tdh PCR trh Gulf Stream

39 V. parahaemolyticus population MLST

40 Vibrio parahaemolyticus ST36 Pacific North West Atlantic North East North West Spain Outbreak- 2012

41 Vibrio parahaemolyticus ST36 Classical PNW clone New PNW clone North West Spain Outbreak-2012??? Atlantic North East clone

42

43 Raw sequences Process sequences (QA, adapter removal) (Trimmomatic) Genome assembly (A5 pipeline) Genome alignment (Harvest tools) Remove gaps (Trimal) Core genome Core SNPs Recombination analysis (ClonalframeML) Remove recombinant regions (cutseq) Extract SNPs in coding regions (SnpEff) Phylogenetic analysis (RAxML) Molecular clock analysis (Beast)

44 Non-recombining core genome cdc VpG vp32 We used BEAST to estimate the rates of evolution on the branches of the tree using a relaxed molecular clock, which allows rates of evolution to vary amongst the branches of the tree.

45 SPREAD: Spatial Phylogenetic Reconstruction of Evolutionary Dynamics application to analyze and visualize Bayesian phylogeographic reconstructions incorporating spatial temporal diffusion It maps phylogenies annotated with both discrete and continuous spatial information and can export high-dimensional posterior summaries to keyhole markup language (KML) for animation of the spatial diffusion through time in virtual globe software SPREAD implements Bayes factor calculation to evaluate the support for hypotheses of historical diffusion among pairs of discrete locations based on Bayesian stochastic search variable selection estimates Phylodynamics of transmission chains between hosts and/or geographical locations

46 Phylogeographical diffusion of lineages across the USA and Spain over time based on a discrete geospatial model with associated geographical coordinates Bayesian-based analysis of the spatial and temporal distribution of the different lineages of ST36. Maximum clade credibility trees from BEAST using core genome after removing recombination. Discrete location states are annotated to tree nodes and branches that indicate location changes are represented on the map. Colors of circular polygons (circles) are indicative of the number of lineages with the discrete state (location) at the given time point. Dark to light color gradients of polygons represent increasing number of lineages (WA). The color gradient (dark red - red) shows ages (older recent) of transmission lines.

47

48

49

50

51

52 How can pathogenic vibrios move long distances? Ocean currents. Bacteria like vibrios are often found in close association with plankton, and can move large distances via oceanic currents. Largescale movements like El Nino is believed to have been responsible for the emergence of cholera into South America in Possible route for 2012 outbreak? Can bacteria survive the journey though? Ballast water. Ships take up marine water, and transport it long distances as ballast. This is a well established mechanism for the introduction of (IPCC fourth assessment report 2007). Vigo exotic bacteria into new areas. Cholera was detected in Gulf of Mexico in early 1990 s after it was transported from an endemic region. Both Vigo and New York are very large and busy ports. NY Movement of shellfish. Live bivalve shellfish are transported large distances. Some produce stored inappropriately (e.g. Mixed, temperature abused etc). These shellfish commodities frequently contain pathogenic strains. Previous outbreaks of Vibrio parahaemolyticus linked to movement of live shellfish produce.

53 How can pathogenic vibrios move long distances? Ocean currents. Bacteria like vibrios are often found in close association with plankton, and can move large distances via oceanic currents. Largescale movements like El Nino is believed to have been responsible for the emergence of cholera into South America in Possible route for 2012 outbreak? Can bacteria survive the journey though? Ballast water. Ships take up marine water, and transport it long distances as ballast. This is a well established mechanism for the introduction of (IPCC fourth assessment report 2007). Vigo exotic bacteria into new areas. Cholera was detected in Gulf of Mexico in early 1990 s after it was transported from an endemic region. Both Vigo and New York are very large and busy ports. NY Movement of shellfish. Live bivalve shellfish are transported large distances. Some produce stored inappropriately (e.g. Mixed, temperature abused etc). These shellfish commodities frequently contain pathogenic strains. Previous outbreaks of Vibrio parahaemolyticus linked to movement of live shellfish produce.

54

55 Vibrio model: ecological preferences

56 Sea surface salinity in Europe Optimal vibrio growth V. cholerae (non O1). 2003, 2006 Average summer SST V. vulnificus (fatality). 1994, 1997, 2003, 2006 V. vulnificus (fatality). 2003, 2006 V. clginolyticus 2006 V. vulnificus (fatality). 1997, 2003, 2006

57 Baltic Sea published Vibrio cases All from 4 years , 2003, 1997 & 1994

58 A B C D

59 Vibrio infections in North Baltic region

60 Vibrio infections in North Baltic region Peak temperatures exceed 24C July 27 th 2014 Many shallow coastal sites are warmer, with temperatures in some isolated area over 25 degrees Celsius!?!

61 Sea Ice, near Oulu, Gulf of Bothnia, Finland, 26 th March 2009 Vibrio infections and Northern Europe, summer 2014

62 Notable recent Vibrio outbreaks in Europe Spain (2012) Circa 100 cases of V. parahaemolyticus traced to shellfish consumption (Martinez-Urtaza et al. 2013). Spain (2004) 80 cases of V. parahaemolyticus. Pandemic strain (O3:K6) identified at outbreak (Martinez-Urtaza et al. 2005). Norway (2011) Small outbreak of O3:K6 (pandemic complex) V. parahaemolyticus related to shellfish consumption. Baltic ( ) Numerous reported cases of wound infections (Vibrio spp.) in the Baltic Sea area. Almost all recreational exposure to seawater during summer months (Baker-Austin et al. 2013). Italy (2004) and France (2005) - several V. parahaemolyticus O3:K6 strains identified based on retrospective analysis of clinical cases. Sporadic V. cholerae (non O1) cases. Source: Baker-Austin, C., L. Stockley, R. Rangdale, and J. Martinez-Urtaza. Environmental occurrence and clinical impact of Vibrio vulnificus and Vibrio parahaemolyticus: a European perspective. Environmental Microbiology Reports, 2010.

63 Vibriosis - the enigma of under-reporting Invisible cases Visible cases Vibrio case correctly identified and reported data made nationally and/or internationally available to Vibrio community Data not made publically available to Vibrio community Vibrio not notifiable agent data not forwarded to national or regional epidemiology centre No formal identification of agent responsible Misdiagnosis of Vibrio illness Case not serious or self limiting and subsequently not reported. Source: Baker-Austin, C., et al. (2014) unpublished

64 vs

65 vs

66 Temperature ( o C) Developing risk-based mapping Baltic, 2006 Temperature (SST) Real-time mapping Risk algorithm Salinity (NaCl ppt) Salinity (SSS)

67

68

69

70

71

72 Conclusions Mechanisms for the dispersion of human pathogenic Vibrios: Introduction in new regions Success of settlement: once introduced, how they become endemic Climate Change and ecological conditions: expanding the range of distribution of pathogenic Vibrio The growing risk of infections in parallel to the warming of coastal areas Underreporting: Regional Government of Galicia (Xunta de Galicia) has been pioneer in the declaration of V. parahaemolyticus infections as notifiable from 1995 and this pathogen is now of urgent notification since January 2012 Improving diagnosis, testing capabilities and centralised reporting systems at European level.

73