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AimsThis study aims to assess the use of marine lactic acid bacteria (LAB) to reduce Vibrio parahaemolyticus levels during oyster depuration process.Methods and resultsThe inhibitory effect of 30 marine LAB strains against V. parahaemolyticus strains was evaluated by in vitro assays. A total of three positive strains (Latilactobacillus sakei SF1583, Lactococcus lactis SF1945, and Vagococcus fluvialis CD264) were selected for V. parahaemolyticus levels reduction during oyster depuration. Pacific oysters Crassostrea gigas were artificially and independently contaminated by four GFP-labelled V. parahaemolyticus strains (IFVp201, IFVp69, IFVp195, and LMG2850T) at 105 CFU ml−1 and then exposed by balneation to 106 CFU ml−1 of each LAB strains during 24 h, at 19°C. Quantification of V. parahaemolyticus in haemolymph by flow cytometry revealed variations in natural depuration of the different V. parahaemolyticus strains alone. Furthermore, the addition of LABs improved up to 1-log bacteria ml−1 the reduction of IFVp201 concentration in comparison to the control condition.ConclusionsAlthough further optimizations of procedure are needed, addition of marine LABs during oyster depuration may be an interesting strategy to reduce V. parahaemolyticus levels in Crassostrea gigas.

Vibrio parahaemolyticus infection in humans is associated with raw oyster consumption. Evaluation of V. parahaemolyticus presence in oysters is of most interest because of the economic and public health issues that it represents. To explore V. parahaemolyticus accumulation and depuration in adult Crassostrea gigas, we developed a GFP-tagged V. parahaemolyticus strain (IFVp201-gfp+), as well as a rapid and efficient quantification method in C. gigas oysters haemolymph by flow cytometry. Impact of the life-history of C. gigas on accumulation and depuration of V. parahaemolyticus IFVp201 was subsequently investigated. We found that naive oysters, i.e. grown in controlled facilities with UV treated seawater, accumulated significantly more IFVp201 than environmental oysters, i.e. grown in intertidal environment. We hypothesized that environmental oysters could have been immune primed, thus could limit V. parahaemolyticus accumulation. Meanwhile, both naive and environmental oysters had similar depuration rates.

International audience; Tetrodotoxins (TTXs) are potentially lethal paralytic toxins that have been identified in European shellfish over recent years. Risk assessment has suggested comparatively low levels (44 µg TTX-equivalent/kg) but stresses the lack of data on occurrence. Both bacteria and dinoflagellates were suggested as possible biogenic sources, either from an endogenous or exogenous origin. We thus investigated TTXs in (i) 98 shellfish samples and (ii) 122 bacterial strains, isolated from French environments. We optimized a method based on mass spectrometry, using a single extraction step followed by ultrafiltration without Solid Phase Extraction and matrix-matched calibration for both shellfish and bacterial matrix. Limits of detection and quantification were 6.3 and 12.5 µg/kg for shellfish and 5.0 and 10 µg/kg for bacterial matrix, respectively. Even though bacterial matrix resulted in signal enhancement, no TTX analog was detected in any strain. Bivalves (either Crassostrea gigas or Ruditapes philippinarum) were surveyed in six French production areas over 2.5–3 month periods (2018–2019). Concentrations of TTX ranged from ‘not detected’ to a maximum of 32 µg/kg (Bay of Brest, 17 June 2019), with events lasting 2 weeks at maximum. While these results are in line with previous studies, they provide new data of TTX occurrence and confirm that the link between bacteria, bivalves and TTX is complex.

In the marine environment, biofilm formation is an important lifestyle for microorganisms. A biofilm is comprised of cells embedded in an extracellular matrix that holds them close together and keeps the biofilm attached to the colonized surface. This predominant lifestyle and its main regulation pathway, namely quorum-sensing (QS), have been shown to induce specific bioactive metabolites. In this study, we investigated the biofilm formation by two marine bacteria belonging to the Vibrio species to discover potentially innovative bioactive compounds. We proposed a protocol to isolate biofilm extracts, to analyze their biochemical composition, and to compare them to planktonic cell extracts. Cells were grown attached to a plastic surface, extracts were prepared in water, NaOH, or in ethyl acetate and analyzed. Extracellular matrix components featured carbohydrates, proteins, lipids, and low amount of DNA. Carbohydrates appeared to be the main constituent of biofilm but also of the planktonic cell supernatant. Moreover, antimicrobial and QS-signaling activities were evidenced in extracts.

Globally, vibrios represent an important and well-established group of bacterial foodborne pathogens. The European Commission (EC) mandated the Comite de European Normalisation (CEN) to undertake work to provide validation data for 15 methods in microbiology to support EC legislation. As part of this mandated work programme, merging of ISO/TS 21872–1:2007, which specifies a horizontal method for the detection of V. parahaemolyticus and V. cholerae, and ISO/TS 21872–2:2007, a similar horizontal method for the detection of potentially pathogenic vibrios other than V. cholerae and V. parahaemolyticus) was proposed. Both parts of ISO/TS 21872 utilized classical culture-based isolation techniques coupled with biochemical confirmation steps. The work also considered simplification of the biochemical confirmation steps. In addition, because of advances in molecular based methods for identification of human pathogenic Vibrio spp. classical and real time PCR options were also included within the scope of the validation. These considerations formed the basis of a multi-laboratory validation study with the aim of improving the precision of this ISO technical specification and providing a single ISO standard method to enable detection of these important foodborne Vibrio spp.. To achieve this aim, an international validation study involving 13 laboratories from 9 countries in Europe was conducted in 2013. The results of this validation have enabled integration of the two existing technical specifications targeting the detection of the major foodborne Vibrio spp., simplification of the suite of recommended biochemical identification tests and the introduction of molecular procedures that provide both species level identification and discrimination of putatively pathogenic strains of V. parahaemolyticus by the determination of the presence of theromostable direct and direct related haemolysins. The method performance characteristics generated in this have been included in revised international standard, ISO 21872:2017, published in July 2017.

The multiannual dynamic of the cyst-forming and toxic marine dinoflagellate Alexandrium minutum was studied over a time scale of about 150 years by a paleoecological approach based on ancient DNA (aDNA) quantification and cyst revivification data obtained from two dated sediment cores of the Bay of Brest (Brittany, France). The first genetic traces of the species presence in the study area dated back to 1873 ± 6. Specific aDNA could be quantified by a newly-developed real-time PCR assay in the upper core layers, in which the germination of the species (in up to 17–19 year-old sediments) was also obtained. In both cores studied, our quantitative paleogenetic data showed a statistically significant increasing trend in the abundance of A. minutum ITS1 rDNA copies over time, corroborating three decades of local plankton data that have documented an increasing trend in the species cell abundance. By comparison, paleogenetic data of the dinoflagellate Scrippsiella donghaienis did not show a coherent trend between the cores studied, supporting the hypothesis of the existence of a species-specific dynamic of A. minutum in the study area. This work contributes to the development of paleoecological research, further showing its potential for biogeographical, ecological and evolutionary studies on marine microbes.

We describe an outbreak of seafood-associated Vibrio parahaemolyticus in Galicia, Spain in on 18th of August 2012 affecting 100 of the 114 passengers travelling on a food banquet cruise boat. Epidemiological information from 65 people was available from follow-on interviews, of which 51 cases showed symptoms of illness. The food items identified through the questionnaires as the most probable source of the infections was shrimp. This product was unique in showing a statistically significant and the highest OR with a value of 7.59 (1.52–37.71). All the nine strains isolated from stool samples were identified as V. parahaemolyticus, seven were positive for both virulence markers tdh and trh, a single strain was positive for trh only and the remaining strain tested negative for both trh and tdh. This is the largest foodborne Vibrio outbreak reported in Europe linked to domestically processed seafood. Moreover, this is the first instance of strains possessing both tdh+ and trh+ being implicated in an outbreak in Europe and that a combination of strains represent several pathogenicity groups and belonging to different genetic variants were isolated from a single outbreak. Clinical isolates were associated with a novel genetic variant of V. parahaemolyticus never detected before in Europe. Further analyses demonstrated that the outbreak isolates showed indistinguishable genetic profiles with hyper-virulent strains from the Pacific Northwest, USA, suggesting a recent transcontinental spread of these strains.

Aims: The aim is to evaluate the dynamic of Bacteroides–Prevotella and Bacillus–Streptococcus–Lactobacillus populations originating from pig manure and the persistence of pig-associated markers belonging to these groups according to temperature and oxygen. Methods and Results: River water was inoculated with pig manure and incubated under microaerophilic and aerobic conditions, at 4 and 20°C over 43 days. The diversity of bacterial populations was analysed by capillary electrophoresis-single-strand conformation polymorphism. The persistence of the pig-associated markers was measured by real-time PCR and compared with the survival of Escherichia coli and enterococci. Decay was characterized by the estimation of the time needed to produce a 1-log reduction (T90). The greatest changes were observed at 20°C under aerobic conditions, leading to a reduction in the diversity of the bacterial populations and in the concentrations of the Pig-1-Bac, Pig-2-Bac and Lactobacillus amylovorus markers with a T90 of 10·5, 8·1 and 17·2 days, respectively. Conclusions: Oxygen and temperature were found to have a combined effect on the persistence of the pig-associated markers in river waters. Significance and Impact of the Study: The persistence profiles of the Pig-1-Bac, Pig-2-Bac and Lact. amylovorus markers in addition to their high specificity and sensitivity support their use as relevant markers to identify pig faecal contamination in river waters.

The present study focused on the isolation of culturable bacteria from mussels and sea water to identify Vibrionaceae potentially pathogenic for humans. Three sites located on the French Atlantic coast were monitored monthly (twice each month during summer) for 1 year. Environmental parameters were surveyed (water temperature, salinity, turbidity, chlorophyll a) and bacteria were detected by culture and identified by API 20E(R) systems (BioMerieux) and PCR. A total of seven species were detected (Grimontia hollisae, Photobacterium damselae, Vibrio alginolyticus, V. cholerae, V. fluvialis, V. vulnificus and V. parahaemolyticus) and species diversity was higher at the end of summer. Surprisingly, V. cholerae non-O1/non-O139 was detected in spring. No site effect was detected. Using Sorensen similarity indices and statistical analyses, we showed that chlorophyll a had a significant influence on the bacterial community detected in mussels and assemblages were more similar to one another when chlorophyll a values were above 20 mu g l(-1). No significant effect of any parameter was found on the community detected in water samples. Such surveys are essential for the understanding of sanitary crises and detection of emerging pathogens.

A real-time reverse transcription-PCR method was developed to determine whether the recovery of culturability of viable but nonculturable (VBNC) Vibrio parahaemolyticus induced the expression of virulence genes coding for the thermostable direct hemolysin and for type III secretion system 2 (TTSS2). The culturability of clinical strain Vp5 of V. parahaemolyticus in artificial seawater at 4°C was monitored, and the VBNC state was obtained. One day after entry into the VBNC state, temperature upshifts to 20 and 37°C allowed the recovery of culturability. Standard curves for the relative quantification of expression of the housekeeping genes rpoS , pvsA , fur , and pvuA ; the tdh2 gene; and the TTSS2 genes (VPA1354, VPA1346, and VPA1342) were established. The levels of expression of the pvsA and pvuA genes were stable and were used to normalize the levels of expression of the other genes. No transcriptional induction of the selected virulence genes under the temperature conditions used to recover the culturability of the VBNC bacteria was observed. The study results demonstrate that the recovery of culturability of VBNC cells of pathogenic V. parahaemolyticus is restricted to regrowth, without correlation with the induction of virulence gene expression. Disease induction would depend mainly on host-pathogen interactions that allow the expression of the virulence genes. This is the first time that the use of mRNA to detect viable cells was evaluated by computing the half-lives of multiple mRNA species under conditions inducing the VBNC state.

The objective of this study was to explore the recovery of culturability of viable but nonculturable ( VBNC) Vibrio parahaemolyticus after temperature upshift and to determine whether regrowth or resuscitation occurred. A clinical strain of V. parahaemolyticus Vp5 was rendered VBNC by exposure to artificial seawater ( ASW) at 4 degrees C. Aliquots of the ASW suspension of cells ( 0.1, 1 and 10 ml) were subjected to increased temperatures of 20 degrees C and 37 degrees C. Culturability of the cells in the aliquots was monitored for colony formation on a rich medium and changes in morphology were measured by scanning ( SEM) and transmission ( TEM) electron microscopy. Samples of VBNC cells were fixed and examined by SEM, revealing a heterogeneous population comprising small cells and larger, flattened cells. Forty-eight hours after temperature upshift to 20 degrees C or 37 degrees C, both elongation and division by binary fission of the cells were observed, employing SEM and TEM, but only in the 10-ml aliquots. The results suggest that a portion of VBNC cells is able to undergo cell division. It is concluded that a portion of VBNC cells of V. parahaemolyticus subjected to cold temperatures remain viable. After temperature upshift, regrowth of those cells, rather than resuscitation of all bacteria of the initial inoculum, appears to be responsible for recovery of culturability of VBNC cells of V. parahaemolyticus. Nutrient in filtrates of VBNC cells is hypothesized to allow growth of the temperature-responsive cells, with cell division occurring via binary fission, but also including an atypical, asymmetric cell division.

Vibrio parahaemolyticus , Vibrio vulnificus , and Vibrio cholerae of the non-O1/non-O139 serotype are present in coastal lagoons of southern France. In these Mediterranean regions, the rivers have long low-flow periods followed by short-duration or flash floods during and after heavy intense rainstorms, particularly at the end of the summer and in autumn. These floods bring large volumes of freshwater into the lagoons, reducing their salinity. Water temperatures recorded during sampling (15 to 24°C) were favorable for the presence and multiplication of vibrios. In autumn 2011, before heavy rainfalls and flash floods, salinities ranged from 31.4 to 36.1‰ and concentrations of V. parahaemolyticus , V. vulnificus , and V. cholerae varied from 0 to 1.5 × 10 3 most probable number (MPN)/liter, 0.7 to 2.1 × 10 3 MPN/liter, and 0 to 93 MPN/liter, respectively. Following heavy rainstorms that generated severe flash flooding and heavy discharge of freshwater, salinity decreased, reaching 2.2 to 16.4‰ within 15 days, depending on the site, with a concomitant increase in Vibrio concentration to ca. 10 4 MPN/liter. The highest concentrations were reached with salinities between 10 and 20‰ for V. parahaemolyticus , 10 and 15‰ for V. vulnificus , and 5 and 12‰ for V. cholerae . Thus, an abrupt decrease in salinity caused by heavy rainfall and major flooding favored growth of human-pathogenic Vibrio spp. and their proliferation in the Languedocian lagoons. Based on these results, it is recommended that temperature and salinity monitoring be done to predict the presence of these Vibrio spp. in shellfish-harvesting areas of the lagoons.

Aims: This work investigates the maintenance of viability and potential virulence of Vibrio parahaemolyticus in a viable but nonculturable population (VBNC) state by reverse transcription-polymerase chain reaction (RT-PCR). Methods and Results: Housekeeping genes, 16S-23S rDNA and rpoS, as well as virulence genes, tdh1 and tdh2, were selected and detected by PCR in a pathogenic strain of V. parahaemolyticus (Vp4). Their expression was then studied by RT-PCR in V. parahaemolyticus Vp4 cultivated in rich medium at 37degreesC. The 16S-23S rDNA and rpoS, tdh1, tdh2 genes were transcripted at the mid-logarithmic, stationary and late stationary phases, corresponding to various physiological states. The expression of these genes was also studied by RT-PCR in a VBNC population of V. parahaemolyticus Vp4 in artificial seawater (ASW). The effect of temperature (washing of bacterial culture and microcosms) on the attaining VBNC bacteria was first considered. Washing of V. parahaemolyticus Vp4, collected at the mid-logarithmic phase, at 10 or 4degreesC before inoculation in ASW at 4degreesC allowed bacteria entered the VBNC state between 22 and 31 days. The 16S-23S rDNA and rpoS gene were expressed in the VBNC bacteria whereas no expression of the tdh1 and tdh2 genes was observed in the same populations. Conclusion: The two selected housekeeping genes, 16S-23S rDNA and rpoS, proved to be good viability markers for V. parahaemolyticus Vp4 in culturable and VBNC states. These first data indicated that the pathogenic strain Vp4 would not maintain the expression of the virulence genes, tdh1 and tdh2, in VBNC state. Significance and Impact of the Study: Use of RT-PCR for investigating the maintenance or not of viability and potential virulence in VBNC V. parahaemolyticus will facilitate further study to evaluate the potential risk presented by this pathogen in the environment.

The objective of this study was to develop a molecular detection method that better estimates the potential risk associated with the presence of Vibrio vulnificus. For that purpose, we applied seminested reverse transcription-PCR (RT-PCR) to viable but nonculturable (VBNC) populations of V. vulnificus and targeted the cytotoxin-hemolysin virulence gene vvhA. Three strains, two environmental, IF Vv10 and IF Vv18, and one clinical, C7184, were used in this study. Artificial seawater, inoculated with mid-log-phase cells, was maintained at 4°C. VBNC cells resulted after 3, 6, and 14 days for C7184, IF Vv18, and IF Vv10, respectively. Our data indicate that seminested RT-PCR is sensitive for the detection of vvhA mRNA in artificial seawater when exclusively nonculturable bacteria are present. This is the first report of the expression of a toxin gene in VBNC V. vulnificus. Moreover, vvhA transcripts were shown to persist in nonculturable populations over a 4.5-month period, with a progressive decline of the signal over time. This result indicates that special attention should be given to the presence of potentially pathogenic VBNC cells in environmental samples when assessing public health risk. Vibrio vulnificus is a gram-negative estuarine bacterium that has been shown to be responsible for human gastroenteritis, wound and/or soft tissue infections, and fatal primary septicemia, with mortality exceeding 50%. Infections are acquired after consumption of raw or undercooked contaminated seafood or after wound exposure to seawater, and they preferentially affect persons having underlying hepatic diseases or immunocompromised individuals (21). It has been shown that V. vulnificus enters into a viable but

Aims To describe the diversity of the culturable mesophilic and potentially pathogenic vibrios isolated at 22 and 37 degrees C on TCBS medium, in September 2009 from seawater and surface sediments. Methods and Results q-PCR assays previously selected for the identification of bacterial strains isolated at 37 degrees C were used in combination with the partial sequencing of two housekeeping genes, pyrH and toxR, to identify 315 strains isolated at 22 degrees C. The great majority of the 37 degrees C strains was identified by q-PCR assays, (five of the six species) with the predominance of Vibrio alginolyticus (85 center dot 9%) and V.harveyi (10 center dot 7%). The human pathogens V.parahaemolyticus and V.cholerae were rarely detected (two strains each). The 22 degrees C strains were successfully identified by the phylogeny analysis of pyrH and toxR genes, revealing 20 Vibrio species, with the predominance of the clam pathogen V.celticus (36 center dot 8%). The Splendidus and the Harveyi groups represented the main Vibrio group at 22 degrees C (80%) and 37 degrees C (99 center dot 5%), respectively. Conclusions The combination of q-PCR assays and the sequencing of pyrH and toxR genes highlighted two different Vibrio communities at 22 and 37 degrees C both dominated by pathogenic species for marine organisms. Significance and Impact of the Study The sequencing of the pyrH gene revealed to be a valuable tool to identify environmental Vibrio spp. strains isolated at 22 degrees C, as 92 center dot 3% of them were identified in this study.

International audience; Vibrio parahaemolyticus, Vibrio vulnificus and Vibrio cholerae are human pathogens. Little is known about these Vibrio spp. in the coastal lagoons of France. The purpose of this study was to investigate their incidence in water, shellfish and sediment of three French Mediterranean coastal lagoons using the most probable number-polymerase chain reaction (MPN-PCR). In summer, the total number of V. parahaemolyticus in water, sediment, mussels and clams collected from the three lagoons varied from 1 to >1.1 × 103 MPN/l, 0.09 to 1.1 × 103 MPN/ml, 9 to 210 MPN/g and 1.5 to 2.1 MPN/g, respectively. In winter, all samples except mussels contained V. parahaemolyticus, but at very low concentrations. Pathogenic (tdh- or trh2-positive) V. parahaemolyticus were present in water, sediment and shellfish samples collected from these lagoons. The number of V. vulnificus in water, sediment and shellfish samples ranged from 1 to 1.1 × 103 MPN/l, 0.07 to 110 MPN/ml and 0.04 to 15 MPN/g, respectively, during summer. V. vulnificus was not detected during winter. V. cholerae was rarely detected in water and sediment during summer. In summary, results of this study highlight the finding that the three human pathogenic Vibrio spp. are present in the lagoons and constitute a potential public health hazard

Aims: To identify Vibrio vulnificus, Vibrio cholerae and Vibrio alginolyticus using standardized DNA extraction method and real-time PCR assays, among a large number of bacterial strains isolated from marine environment. Methods and Results: Methods for DNA extraction and real-time PCR were standardized to identify a large number of Vibrio spp. strains isolated through regular collection campaigns of environmental samples. Three real-time PCR assays were developed from a multiplex PCR, targeting V. vulnificus, V. cholerae and V. alginolyticus on the dnaJ gene. After testing their specificity, these systems were applied for the identification of 961 strains isolated at 22°C (446 strains) and 37°C (515 strains) in September 2009. The predominance of V. alginolyticus (82·6%) among the Vibrio spp. strains isolated at 37°C was shown. At 22°C, only 1·6% of the strains were identified by PCR and they were V. alginolyticus. Conclusions: Reproducible and specific real-time PCR assays combined to a DNA extraction method on microplates were used to constitute a large environmental Vibrio strains collection and to identify and detect potential human pathogenic Vibrio isolated at 37°C. For environmental strains isolated at 22°C, because of the higher species diversity, other approaches, like sequencing, should be chosen for identification. Significance and Impact of the Study: The protocol developed in this study provides an appropriate and rapid screening tool to identify a large number of bacterial strains routinely isolated from the environment in long-term studies.

This chapter focuses on the enteric viruses, hepatitis A virus and norovirus, responsible for the main outbreaks linked to shellfish consumption. One major drawback for viral contamination estimation in shellfish is the lack of indicator or standardised method, preventing systematic control. After a review of the main sources of pollution, we identified the conditions responsible for shellfish contamination. This will help to set up potential strategies to reduce contamination in harvesting areas such as the limitation of faecal input or the improvement of tools for management. As knowledge is progressing risk management, strategies will help to protect the consumer and will also led to improve regulation. Rapid alert systems need to be set up to prevent coastal area contamination and promising examples of coastal management have proved their efficacy to reduce faecal load and thus, improve water quality. Shellfish have long been recognised as being beneficial to human health and this benefit should also be taken into consideration in managing coastal areas and preserving the water quality.

A number of bacterial illnesses may arise from the consumption of seafood that has either been contaminated at source or which becomes contaminated during the processing and retail chain. Such illnesses may arise from infection with the bacteria themselves or by the ingestion of toxins formed in the foodstuff prior to consumption. This division is actually too simplistic: for example, the toxins of Clostridium botulinum and Staphylococcus aureus are preformed in food during bacterial growth, the toxin of Clostridium perfringens is usually only formed when the bacteria sporulate in the intestinal tract while the toxin of Vibrio cholerae O1 (and O139) is produced when the bacteria multiply in the intestinal tract.

Introduction : "The occurrence of pathogenic microorganisms in seawater or in shellfish could exist anytime sewage from human or animal origin would be discharged to the coast" (Metcalf, 1982). According to the diseases occurring in the human population or in animals, pathogens might be present in recreational waters or in shellfish. Thus, the presence of human enteric viruses (norovirus, astrovirus, rotavirus, hepatitisAvirus (HAV)) and pathogenic bacteria (Salmonella, Listeria monocytogenes, Shiga-toxin-producing Escherichia coli (STEC), Vibrio cholerae, Vibrio parahaemolyticus, etc.) has been reported in coastal areas for a long time (Colwell, 1978; Metcalf, 1978; Melnick et al., 1979; Grimes, 1991; Bosch et al., 2001;Kong et al., 2002). These microorganisms have been implicated in gastrointestinal and respiratory illnesses and other infections (skin, eyes, etc.), (Griffin et al., 2003). Using risk-assessment models for viruses, maximum risks were estimated to be 1.3 infections per 100 swimmers (Colwell et al., 1996). To evaluate the risk due to the presence of these pathogens in the environment, certain criteria have to be determined. Among them, the infectious dosewould be of a greatest importance (Table 14.1). Even if the infectious dose vary with the strains, the age of the patient, or other parameters, some pathogens are highly dangerous for men even at low concentrations (HAV, E. coli O157:H7, V. cholerae), whereas others have to be ingested in high concentrations to be harmful (V. parahaemolyticus) or are highly infectious but not very dangerous (norovirus). Thus, for some pathogens, a low contamination in seafood, for example, is not acceptable, based on risk-assessment models (Colwell et al., 1996). Among bacteria, the Vibrio family plays an important role in infections, waterborne or seafood diseases, especially in countries surrounded by warm marine waters. Toxigenic V. cholerae O1 and O139 are the causative agents of cholera in developing countries, whereas, non-O1 and non-O139 strains may also be pathogenic but are rarely associated with epidemics (Anonymous, 2002). V. parahaemolyticus has been recognized as a major cause of foodborne gastroenteritis in Japan and Eastern countries and has been linked to seafood consumption. In the United States, more than 700 cases of illness due to V. parahaemolyticus and associated to raw oyster consumption were reported between 1997 and 1998. However, only a few outbreaks have been reported in Australia (1990-1992) and France (2001). The difference between the countries could be attributed to the ecology of this bacteria that requires elevated temperature to grow (>18?C). Vibrio vulnificus associated with primary septicemia, has also been detected in a variety of raw seafood products in Korea and Japan (Anonymous, 2002). Concerning enteric bacteria discharged into marine waters, bacterial control of seafood lowered the risk of Salmonella outbreaks. The occurrence of other bacteria such as Campylobacter jejuni and L. monocytogenes has rarely been reported in shellfish (Feldhusen, 2000; Montfort et al., 1998; Dupray et al., 1999; Federighi et al., 1998). However, L. monocytogenes have been implicated with smoked fish consumption (Rocourt et al., 2000). Human enteric viruses are involved in more than 30% of the outbreaks linked to oyster consumption. Despite the variety of viruses that can communicate shellfish only, HAV and norovirus have been epidemiologically linked to disease (Lees, 2000; Le Guyader et al., 2003). The objective of this review is to gather information on the input and behavior of the main enteric microorganisms-Vibrio family excepted-via the water route and the factors able to affect the seawater and seafood quality.

Aims: This study was carried out to investigate the occurrence of potentially pathogenic species of Vibrio in French marine and estuarine environments. Methods and Results: Samples of coastal waters and mussels collected between July and September 1999 were analysed by culture, using selective media including thiosulphate-citrate-bile salts-sucrose and modified cellobiose-polymixin B-colistin agar. Presumptive Vibrio colonies were isolated and identified using selected biochemical tests. Specific primers based on flanking sequences of the cytolysin, vvhA gene, pR72H DNA fragment and 16S–23S rRNA intergenic spacer region (ISR) were used in a polymerase chain reaction (PCR) to confirm the identification of Vibrio vulnificus, V. parahaemolyticus and V. cholerae, respectively. In this study, V. alginolyticus (99 of 189) was the predominant species, followed by V. parahaemolyticus (41 of 189), V. vulnificus (20 of 189) and non-O1/non-O139 V. cholerae (three of 189). All 20 V. vulnificus isolates showed PCR amplification of the vvhA gene, 16 of which had been isolated from estuarine water. The PCR amplification of the pR72H DNA fragment in 41 V. parahaemolyticus isolates generated two unique amplicons of 387 and 320 bp. The latter, present in 24·4% of these isolates, had not previously been found in V. parahaemolyticus strains examined to date. Amplification of the trh gene in two of the isolates suggested these to be virulent strains. Three strains identified as V. cholerae by amplification of the 16S–23S rRNA ISR were confirmed to be non-cholera (non-O1/non-O139) strains. Conclusions: The results of this study demonstrated the presence of pathogenic Vibrio species in French coastal waters. Furthermore, the PCR approach proved useful for the rapid and reliable confirmation of species identification. Significance and Impact of the Study: These findings indicate the potential sanitary risk associated with the presence of pathogenic Vibrio spp. in cultivated mussels and in the aquatic environment. The PCR can be used to detect pathogenic vibrios directly in environmental samples.

Prod 2017-108 EA BIOmE CT ? INRA; National audience; Quand l'écologie pose son diagnostic. L'actualité nous le rappelle sans cesse : les maladies comme le paludisme, le Sida, ébola ou zika sont difficiles à combattre, et les maladies chroniques de type cancer, diabète ou maladies cardio-vasculaires toujours plus meurtrières. Pour la première fois de son histoire, l'humanité voit même son espérance de vie diminuer. Pour comprendre les raisons de cette crise sanitaire, l'écologie scientifique pose un autre regard sur la santé qui prend en considération les transformations de l'environnement et l'évolution de nos modes de vie. Cette nouvelle approche évolutive et écologique des problèmes de santé tente d'élucider les mécanismes en jeu dans la transmission et l'émergence de nouveaux pathogènes, mais aussi dans la résistance toujours croissante aux antibiotiques. Elle permet de mesurer l'effet des facteurs environnementaux sur la santé : concentration urbaine et industrielle et nouveaux comportements induits en matière d'alimentation et d'activités physiques. Cet ouvrage rédigé par des spécialistes issus des laboratoires du CNRS ou associés fait le point sur les recherches en écologie de la santé. Il permet à chacun de comprendre pourquoi il est essentiel de remonter aux sources environnementales et comportementales de nos maux, et de ne pas se limiter aux symptômes.