Evaluation of the protection against norovirus afforded by E. coli monitoring of shellfish production areas under EU regulations.

EC Regulation 854/2004 requires the classification of bivalve mollusc harvesting areas according to the faecal pollution status of sites. It has been reported that determination of Escherichia coli in bivalve shellfish is a poor predictor of norovirus (NoV) contamination in individual samples. We explore the correlation of shellfish E. coli data with norovirus presence using data from studies across 88 UK sites (1,184 paired samples). We investigate whether current E. coli legislative standards could be refined to reduce NoV infection risk. A significant relationship between E. coli and NoV was found in the winter months (October to February) using data from sites with at least 10 data pairs (51 sites). We found that the ratio of arithmetic means (log10 E. coli to log10 NoV) at these sites ranged from 0.6 to 1.4. The lower ratios (towards 0.6) might typically indicate situations where the contribution from UV disinfected sewage discharges was more significant. Conversely, higher ratios (towards 1.4) might indicate a prevalence of animal sources of pollution; however, this relationship did not always hold true and so further work is required to fully elucidate the factors of relevance. Reducing the current class B maximum (allowed in 10% of samples) from 46,000 E. coli per 100 g (corresponding NoV value of 75750 ± 103) to 18,000 E. coli per 100 g (corresponding NoV value of 29365 ± 69) reduces maximum levels of NoV by a factor of 2.6 to 1; reducing the upper class B limit to 100% compliance with 4,600 E. coli per 100 g (corresponding NoV value of 7403 ± 39) reduces maximum levels of NoV by a factor of 10.2 to 1. We found using the UK filtered winter dataset that a maximum of 200 NoV corresponded to a maximum of 128 ± 7 E. coli per 100 g. A maximum of 1,000 NoV corresponded to a maximum of 631 ± 14 E. coli per 100 g.

Patterns of Multi-Antibiotic-Resistant Escherichia Coli from Streams with No History of Antimicrobial Inputs.

A growing body of evidence suggests that contaminated environments may harbor a greater proportion of antibiotic-resistant microorganisms than unpolluted reference sites. Here, we report the screening of 427 Escherichia coli strains isolated from 11 locations on nine streams draining the US Department of Energy's Savannah River Site against a panel of five antibiotics. Streams were chosen to capture a wide range of watersheds from minimally disturbed to highly impacted. Overall, higher levels of resistance were found in waterborne E. coli that also generally exhibited low spatial variability. However, 3 of 11 locations also demonstrated elevated resistance levels in sediments. Two of these occurred in highly disturbed tributaries with no obvious sources of antimicrobials. To further investigate these patterns, we screened a subset of isolates obtained from three streams against 23 antibiotics or antibiotic combinations. A large proportion of these isolates (>40 %) demonstrated resistance to 10 or more antimicrobials, suggesting that environmental multi-antibiotic resistance may be prevalent in this bacterial commensal. Only 4 of 87 viable isolates were tested susceptible to all 23 antibiotics and combinations. Among these multi-antibiotic-resistant isolates, several demonstrated resistance to all structural classes of antimicrobial agents tested, including frontline antibiotics such as gatifloxacin and ciprofloxacin.

Detection of the pufferfish toxin tetrodotoxin in European bivalves, England, 2013 to 2014.

We report the first detection of tetrodotoxins (TTX) in European bivalve shellfish. We demonstrate that TTX is present within the temperate waters of the United Kingdom, along the English Channel, and can accumulate in filter-feeding molluscs. The toxin is heat-stable and thus it cannot be eliminated during cooking. While quantified concentrations were low in comparison to published minimum lethal doses for humans, the results demonstrate that the risk to shellfish consumers should not be discarded.

Antimicrobial textiles.

Bacteria have evolved unique mechanisms that allow them survive in the presence of strong selection pressures. Included in these mechanisms is the ability to share genetic determinants among and between species of bacteria thus spreading metal or antibiotic resistance traits quickly. The textile industry in response to demand has developed antimicrobial fabrics by the addition of bactericidal compounds during production. Some of these antimicrobials include metal nanoparticles, quaternary ammonia compounds, and broad spectrum compounds like triclosan. Bacteria have already expressed resistance to each of these bactericides. Here we discuss the evolutionary and ecological consequences of antimicrobial textiles in terms of co-selection. We predict that continued use of such materials could result in increased and widespread resistance to specific antimicrobials, especially metals, with an increased resistance to antibiotics. Such increases have the potential to find their way into other bacterial populations of human pathogens leading to serious and unintended public health consequences.

A seafood risk tool for assessing and mitigating chemical and pathogen hazards in the aquaculture supply chain.

Intricate links between aquatic animals and their environment expose them to chemical and pathogenic hazards, which can disrupt seafood supply. Here we outline a risk schema for assessing potential impacts of chemical and microbial hazards on discrete subsectors of aquaculture-and control measures that may protect supply. As national governments develop strategies to achieve volumetric expansion in seafood production from aquaculture to meet increasing demand, we propose an urgent need for simultaneous focus on controlling those hazards that limit its production, harvesting, processing, trade and safe consumption. Policies aligning national and international water quality control measures for minimizing interaction with, and impact of, hazards on seafood supply will be critical as consumers increasingly rely on the aquaculture sector to supply safe, nutritious and healthy diets.

Vibrio alginolyticus-associated wound infection acquired in British waters, Guernsey, July 2011.

In July 2011, a Vibrio alginolyticus infection was diagnosed in a woman from Guernsey in the Channel Islands, British Isles after sea bathing and application of a natural seaweed dressing to a pre-existing leg wound. Microbiological investigation confirmed Vibrio in the wound and the species of seaweed used for the dressing. The washing of open wounds in seawater and use of unsterilised seaweed dressings should be discouraged, particularly in individuals with underlying risk conditions.

Rapid identification and differentiation of agricultural faecal contamination sources using multiplex PCR.

AIMS: To develop a quick, easy-to-use, robust and sensitive multiplex PCR assay to detect common sources of agricultural faecal contamination using a combination of bacterial and eukaryote-specific PCR targets. METHOD AND RESULTS: A novel multiplex PCR method was developed that utilizes primers specific for a conserved region of the eukaryote cytochrome-B gene as well as a universal 16S rRNA and the E. coli-specific uidA gene. This multiplex PCR assay was capable of identifying faecal amendments from pig, sheep, cow and goat sources in 24/30 (80%) of amended water samples. CONCLUSIONS: The method was capable of accurately identifying common agricultural sources. SIGNIFICANCE AND IMPACT OF THE STUDY: The procedure described here is simple, rapid (<5 h) and can be used as a first step in microbial source tracking studies, particularly where agricultural faecal contamination is suspected.

A pan-European ring trial to validate an International Standard for detection of Vibrio cholerae, Vibrio parahaemolyticus and Vibrio vulnificus in seafoods.

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.

Bioaccumulation and persistence of faecal bacterial and viral indicators in Mytilus edulis and Crassostrea gigas.

This study investigated the response of two shellfish species - mussels (Mytilus edulis) and oysters (Crassostrea gigas) to microbial contamination in order to gain a better understanding of the bioaccumulation and persistence of microorganisms under controlled conditions. M. edulis and C. gigas were placed in sterile laboratory-prepared artificial seawater and initial tests were carried out to ensure both faecal indicator bacteria (FIB) and bacteriophages were below detection limits. FIB and phages were isolated, purified and dosed into experimental tanks containing the shellfish species. The GB124 phages were bioaccumulated to the highest concentration in M. edulis (1.88Log10) and C. gigas (1.27Log10) after 24h. Somatic coliphages were bioaccumulated to the highest concentration in both M. edulis (4.84log10) and C. gigas (1.73log10) after 48h. F-RNA phages were bioaccumulated to the highest concentration in M. edulis (3.51log10) after 6h but were below detection limit in C. gigas throughout the exposure period. Escherichia coli, faecal coliforms and intestinal enterococci were bioaccumulated to the highest concentrations in M. edulis (5.05log10, 5.06log10 and 3.98log10, respectively) after 48h. In C. gigas, E. coli reached a maximum concentration (5.47log10) after 6h, faecal coliforms (5.19log10) after 12h and intestinal enterococci (3.23log10) after 24h.M. edulis bioaccumulated phages to a greater extent than the faecal bacteria, and in both shellfish species, faecal bacteria persisted for longer periods over 48h than the phages. This study highlights significant variation in the levels and rate of accumulation and persistence with respect to both shellfish species and the indicators used to assess risk. The results suggest that phage-based indicators could help elucidate risks to human health associated with pathogenic organisms.