Use of F-Specific RNA Bacteriophage to Estimate Infectious Norovirus Levels in Oysters.

Contamination of bivalve shellfish, particularly oysters, with norovirus is recognised as a significant food safety risk. Methods for quantification of norovirus in oysters using the quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) are well established, and various studies using RT-qPCR have detected norovirus in a considerable proportion of oyster samples, both in the UK and elsewhere. However, RT-qPCR detects viral genome, and by its nature is unable to discriminate between positive results caused by infectious viruses and those caused by non-infectious remnants including damaged virus particles and naked RNA. As a result, a number of alternative or complementary approaches to RT-qPCR testing have been proposed, including the use of infectious viral indicator organisms, most frequently F-specific RNA bacteriophage (F-RNA phage). In this study, we investigated the relationships between F-RNA phage and norovirus in digestive tissues from two sets of oyster samples, one randomly collected at retail (630 samples), and one linked to suspected norovirus illness outbreaks (nine samples). A positive association and correlation between PCR-detectable levels of genogroup II F-RNA bacteriophage (associated with human faecal contamination) and norovirus was found in both sets of samples, with more samples positive for genogroup II phage, at generally higher levels than norovirus. Levels of both viruses were higher in outbreak-related than retail samples. Infectious F-RNA phage was detected in 47.8% of all retail samples, and for a subset of 224 samples where characterisation of phage was carried out, infectious GII phage was detected in 30.4%. Infectious GII phage was detected in all outbreak-related samples. Determination of infectivity ratios by comparing levels of PCR-detectable (copies/g) and infectious GII phage (pfu/g) revealed that in the majority of cases less than 10% of virus detected by RT-qPCR was infectious. Application of these ratios to estimate infectious norovirus levels indicated that while 77.8% of outbreak-related samples contained > 5 estimated infectious norovirus/g, only 13.7% of retail samples did. Use of a combination of levels of PCR-detectable norovirus and infectious F-RNA phage showed that while only 7.0% of retail samples contained both > 100 copies/g norovirus and > 10 pfu/g F-RNA phage, these combined levels were present in 77.8% of outbreak-related samples, and 75.9% of retail samples with > 5 estimated infectious norovirus/g. We therefore suggest that combining RT-qPCR testing with a test for infectious F-RNA phage has the potential to better estimate health risks, and to better predict the presence of infectious norovirus than RT-qPCR testing alone.

The use of capture-recapture methods to provide better estimates of the burden of norovirus outbreaks from seafood in England, 2004-2011.

Norovirus (NoV) is the greatest cause of infectious intestinal disease in the UK. The burden associated with foodborne outbreaks is underestimated in part because data are dispersed across different organisations. Each looks at outbreaks through a different lens. To estimate the burden of NoV from seafood including shellfish we used a capture-recapture technique using datasets from three different organisations currently involved in collecting information on outbreaks. The number of outbreaks of NoV related to seafood including shellfish in England was estimated for the period of 2004-2011. The combined estimates were more than three times as high (N = 360 using Chao's sample coverage approach) as the individual count from organisation three (N = 115), which captured more outbreaks than the other two organisations. The estimates were calculated for both independence and dependence between the datasets. There was evidence of under-reporting of NoV outbreaks and inconsistency of reporting between organisations, which means that, currently, more than one data source needs to be used to estimate as accurately as possible the total number of NoV outbreaks and associated cases. Furthermore, either the integration of reporting mechanisms or simplifying the process of reporting outbreaks to organisations is essential for understanding and, hence, controlling disease burden.

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.

A One-Year Survey of Norovirus in UK Oysters Collected at the Point of Sale.

Contamination of bivalve shellfish, particularly oysters, with norovirus is recognised as a food safety risk and a potential contributor to the overall burden of gastroenteritis in the community. The United Kingdom (UK) has comprehensive national baseline data on the prevalence, levels, and seasonality of norovirus in oysters in production areas resulting from a previous two-year study (2009-2011). However, previously, data on final product as sold to the consumer have been lacking. As part of a wider project to establish the overall burden of foodborne norovirus in the UK, this study aimed to address this data gap. A one-year survey of oysters collected from the point-of-sale to the consumer was carried out from March 2015 to March 2016. A total of 630 samples, originating in five different European Union Member States, were collected from 21 regions across the UK using a randomised sampling plan, and tested for norovirus using a method compliant with ISO 15216-1, in addition to Escherichia coli as the statutory indicator of hygiene status. As in the previous production area study, norovirus RNA was detected in a high proportion of samples (68.7%), with a strong winter seasonality noted. Some statistically significant differences in prevalences and levels in oysters from different countries were noted, with samples originating in the Netherlands showing lower prevalences and levels than those from either the UK or Ireland. Overall, levels detected in positive samples were considerably lower than seen previously. Investigation of potential contributing factors to this pattern of results was carried out. Application of normalisation factors to the data from the two studies based on both the numbers of norovirus illness reports received by national surveillance systems, and the national average environmental temperatures during the two study periods resulted in a much closer agreement between the two data sets, with the notably different numbers of illness reports making the major contribution to the differences observed in norovirus levels in oysters. The large majority of samples (76.5%) contained no detectable E. coli; however, in a small number of samples (2.4%) levels above the statutory end product standard (230 MPN/100 g) were detected. This study both revealed the high prevalence of norovirus RNA in oysters directly available to the UK consumer, despite the high level of compliance with the existing E. coli-based health standards, while also highlighting the difficulty in comparing the results of surveys carried out in different time periods, due to variability in risk factors.

A single blind randomised controlled trial of the impact on patient-reported pain of arm elevation versus exsanguination prior to tourniquet inflation.

AIMS: The aim of this study was to compare the pain caused by the application of a tourniquet after exsanguination of the upper limb with that occurring after simple elevation. PATIENTS AND METHODS: We used 26 healthy volunteers (52 arms), each of whom acted as their own matched control. The primary outcome measure was the total pain experienced by each volunteer while the tourniquet was inflated for 20 minutes. This was calculated as the area under the pain curve for each individual subject. Secondary outcomes were pain at each time point; the total pain experienced during the recovery phase; the ability to tolerate the tourniquet and the time for full recovery after deflation of the tourniquet. RESULTS: There was a significant difference in the area under the pain curves in favour of exsanguination (mean difference 8.4; 95% confidence interval (CI) 3.0 to 13.7, p = 0.004). There was no difference between the dominant and non-dominant arms (mean difference -0.2; 95% CI -23.2 to 22.8, p = 0.99). The area under both recovery curves were similar (mean difference 0.7; 95% CI -6.0 to 4.6, p = 0.78). There was no statistical difference in recovery time, the actual mean difference being 30 seconds longer in the elevation group (p = 0.06). CONCLUSION: Many orthopaedic and plastic surgery procedures are done under local anaesthetic or regional block where a bloodless field and a motionless patient are essential. Optimising patient comfort during surgery with the tourniquet inflated is thus a priority. This study is useful in that it compares two common methods of preparation of the upper limb prior to tourniquet inflation and which have not previously been compared in this context. Following on the results of this study, we can confidently conclude that exsanguinating the upper limb before inflating a tourniquet is more comfortable than simply elevating the arm for patients undergoing a procedure under local or regional block, both during the procedure and in the recovery phase. TAKE HOME MESSAGE: Exsanguination rather than elevation is recommended in order to minimise patient discomfort and optimise the surgical field.

Mitogenomics of 'Old World Acraea' butterflies reveals a highly divergent 'Bematistes'.

Afrotropical Acraeini butterflies provide a fascinating potential model system to contrast with the Neotropical Heliconiini, yet their phylogeny remains largely unexplored by molecular methods and their generic level nomenclature is still contentious. To test the potential of mitogenomes in a simultaneous analysis of the radiation, we sequenced the full mitochondrial genomes of 19 African species. Analyses show the potential of mitogenomic phylogeny reconstruction in this group. Inferred relationships are largely congruent with a previous multilocus study. We confirm a monophyletic Telchinia to include the Asiatic Pareba with a complicated paraphylum, traditional (sub)genus Acraea, toward the base. The results suggest that several proposed subgenera and some species groups within Telchinia are not monophyletic, while two other (sub)genera could possibly be combined. Telchinia was recovered without strong support as sister to the potentially interesting system of distasteful model butterflies known as Bematistes, a name that is suppressed in some treatments. Surprisingly, we find that this taxon has remarkably divergent mitogenomes and unexpected synapomorphic tRNA rearrangements. These gene order changes, combined with evidence for deviating dN/dS ratios and evidence for episodal diversifying selection, suggest that the ancestral Bematistes mitogenome has had a turbulent past. Our study adds genetic support for treating this clade as a distinct genus, while the alternative option, adopted by some authors, of Acraea being equivalent to Acraeini merely promotes redundancy. We pave the way for more detailed mitogenomic and multi-locus molecular analyses which can determine how many genera are needed (possibly at least six) to divide Acraeini into monophyletic groups that also facilitate communication about their biology.

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.

Generation of point-mutant FAK knockin mice.

Focal adhesion kinase is a non-receptor protein tyrosine kinase with signaling functions downstream of integrins and growth factor receptors. In addition to its role in adhesion, migration, and proliferation it also has non-kinase scaffolding functions in the nucleus. Focal adhesion kinase (FAK) activation involves the following: (1) ligand bound growth factors or clustered integrins activate FAK kinase domain; (2) FAK autophosphorylates tyrosine (Y) 397; (3) Src binds pY397 and phosphorylates FAK at various other sites including Y861; (4) downstream signaling of activated FAK elicits changes in cellular behavior. Although many studies have demonstrated roles for the kinase domain, Y397 and Y861 sites, in vitro much less is known about their functions in vivo. Here, we report the generation of a series of FAK-mutant knockin mice where mutant FAK, either kinase dead, non-phosphorylatable mutants Y397F and Y861F, or mutant Y397E-containing a phosphomimetic site that results in a constitutive active Y397, can be expressed in a Cre inducible fashion driven by the ROSA26 promoter. In future studies, intercrossing these mice with FAKflox/flox mice and inducible cre-expressing mice will enable the in vivo study of mutant FAK function in the absence of endogenous FAK in a spatially and temporally regulated fashion within the whole organism.

Is tibial tuberosity-trochlear groove distance an appropriate measure for the identification of knees with patellar instability?

PURPOSE: Tibial tuberosity-trochlear groove distance (TT-TG) has been regarded as a useful tool for establishing therapeutic choices for patellar instability. Recently, it has been shown that TT-TG negatively correlated with the quadriceps angle, suggesting that if used individually, neither provide a valid measure of instability. This study aimed to compare TT-TG distance between both knees in patients with unilateral instability to assess whether this measurement is a decisive element in the management decisions for patellar instability. METHODS: Sixty-two patients (18 male and 44 female), reporting to a specialist patella clinic for recurrent unilateral patellar instability, were included in the study. Patients underwent bilateral long leg computed tomography scan to determine TT-TG distance in both knees. Tibial TT-TG in symptomatic and asymptomatic knees in the same individual was compared statistically. RESULTS: Mean TT-TG distance in the symptomatic knee was 16.9 (±4.9) mm, compared to 15.6 (±5.6) mm in the asymptomatic knee. Tibial TT-TG was not significantly different between stable and unstable knees (n.s.). CONCLUSIONS: The lack of difference in TT-TG distance between stable and unstable knees suggests that TT-TG distance alone may not be a decisive element in establishing therapeutic choices for patellar instability. It should, therefore, be interpreted with caution during clinical evaluations. LEVEL OF EVIDENCE: II.

DNA barcoding reveals a largely unknown fauna of Gracillariidae leaf-mining moths in the Neotropics.

Higher taxa often show increasing species richness towards tropical low latitudes, a pattern known as the latitudinal biodiversity gradient (LBG). A rare reverse LBG (with greater richness towards temperate high latitudes) is exhibited by Gracillariidae leaf-mining moths, in which most described species occur in northern temperate areas. We carried out the first assessment of gracillariid species diversity in two Neotropical regions to test whether the relatively low tropical species diversity of this family is genuine or caused by insufficient sampling and a strong taxonomic impediment. Field surveys in six French Guianan and one Ecuadorian site produced 516 gracillariid specimens that were DNA barcoded to facilitate identification and to match larvae inside leaf mines with adults. Species delineation from sequence data was approximated using Automatic Barcode Gap Discovery and Refined Single Linkage Analysis through the Barcode Index Number system, and the proportion of described/undescribed species was estimated after comparison with types of 83% of described species. Locally, alpha-diversity far exceeds that of any known temperate fauna, with as many as 108 candidate species (59.3% as singletons) collected at one site, and with an estimated species richness lower bound of 240 species. Strikingly, at least 85% of the species collected as adults were found to be undescribed. Our sampling represents the most thorough survey of gracillariid species diversity in the Neotropics to date and the results from both our molecular and morphological analyses indicate that the current reverse LBG seen in this group is an artefact of insufficient sampling and a strong description deficit in the Neotropics.

The effect of surgical factors on early patient-reported outcome measures (PROMS) following total knee replacement.

Patient-reported outcome measures (PROMs) are increasingly being used to assess functional outcome and patient satisfaction. They provide a framework for comparisons between surgical units, and individual surgeons for benchmarking and financial remuneration. Better performance may bring the reward of more customers as patients and commissioners seek out high performers for their elective procedures. Using National Joint Registry (NJR) data linked to PROMs we identified 22,691 primary total knee replacements (TKRs) undertaken for osteoarthritis in England and Wales between August 2008 and February 2011, and identified the surgical factors that influenced the improvements in the Oxford knee score (OKS) and EuroQol-5D (EQ-5D) assessment using multiple regression analysis. After correction for patient factors the only surgical factors that influenced PROMs were implant brand and hospital type (both p < 0.001). However, the effects of surgical factors upon the PROMs were modest compared with patient factors. For both the OKS and the EQ-5D the most important factors influencing the improvement in PROMs were the corresponding pre-operative score and the patient's general health status. Despite having only a small effect on PROMs, this study has shown that both implant brand and hospital type do influence reported subjective functional scores following TKR. In the current climate of financial austerity, proposed performance-based remuneration and wider patient choice, it would seem unwise to ignore these effects and the influence of a range of additional patient factors.

The development of LENTICULES™ as reference materials for noroviruses.

AIMS: To investigate the potential for LENTICULES™ to act as reference materials (RMs) for noroviruses (NoV) [genogroups I (GI) and II (GII)] by determining their homogeneity and stability characteristics. METHODS AND RESULTS: NoV used in this study originated from human faecal material, screened for the absence of other faecally transmitted pathogens. The norovirus strains present in the faecal material were characterized by sequencing, and samples containing GI and GII strains representative of genotypes commonly circulating in the community were selected. RMs were produced utilizing modified lenticulating technology. A batch comprising 500 LENTICULES™ containing both norovirus genogroups was produced according to ISO Guide 34. The batch was tested and quantified using an ISO 17025 accredited quantitative real-time RT-PCR assay. Sufficient homogeneity was established using procedures described by Fearn and Thompson (2010), while stability at less than -15°C and ambient temperature (17-22°C) was assessed over 52 weeks and 7 days, respectively. CONCLUSIONS: Lenticulation was shown to be an effective means of preservation of detectable NoV. LENTICULES™ were sufficiently homogeneous and stable throughout medium-term frozen and short-term storage at room temperature to serve as RMs. Virus LENTICULES™ have the advantages of being easy to manipulate, provide assigned values and do not require the manipulation of high titre clinical material. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study show that norovirus LENTICULES™ can be used as stable RMs for quantitative real-time RT-PCR assays. They can be utilized as in-run positive extraction controls and potentially for method calibration and to enable more easy comparison of data generated by the variety of differing norovirus determination methods that have emerged in recent years. LENTICULES™ have the potential to provide essential elements of laboratory quality assurance systems for laboratories implementing these new methods for virus testing in foodstuffs and for those running routine analyses.

Lower rates of dislocation with increased femoral head size after primary total hip replacement: a five-year analysis of NHS patients in England.

Increased femoral head size may reduce dislocation rates following total hip replacement. The National Joint Registry for England and Wales has highlighted a statistically significant increase in the use of femoral heads ≥ 36 mm in diameter from 5% in 2005 to 26% in 2009, together with an increase in the use of the posterior approach. The aim of this study was to determine whether rates of dislocation have fallen over the same period. National data for England for 247 546 procedures were analysed in order to determine trends in the rate of dislocation at three, six, 12 and 18 months after operation during this time. The 18-month revision rates were also examined. Between 2005 and 2009 there were significant decreases in cumulative dislocations at three months (1.12% to 0.86%), six months (1.25% to 0.96%) and 12 months (1.42% to 1.11%) (all p < 0.001), and at 18 months (1.56% to 1.31%) for the period 2005 to 2008 (p < 0.001). The 18-month revision rates did not significantly change during the study period (1.26% to 1.39%, odds ratio 1.10 (95% confidence interval 0.98 to 1.24), p = 0.118). There was no evidence of changes in the coding of dislocations during this time. These data have revealed a significant reduction in dislocations associated with the use of large femoral head sizes, with no change in the 18-month revision rate.

Human norovirus RNA persists in seawater under simulated winter conditions but does not bioaccumulate efficiently in Pacific Oysters (Crassostrea gigas).

Norovirus (NoV) is the principal agent of bivalve molluscan shellfish-associated gastroenteric illness worldwide. Currently, noncultivable human NoVs can be detected in bivalve molluscan shellfish by using molecular methods such as real-time reverse transcription PCR assays (qRT-PCR). In addition to infectious viruses, this methodology may also detect noninfectious NoV, including fragments of the NoV genome. This study addresses, in part, the implications of qRT-PCR results for the detection of NoV in shellfish in the absence of an infectivity assay. To evaluate environmental persistence, the stability of a short fragment of the NoV genome, spanning the qRT-PCR target in the open reading frame 1/2 junction, was assessed in seawater under artificial environmental conditions simulating winter in the United Kingdom (1 mW/cm² UV irradiation, 8°C) during a 4-week period. Detectable RNA levels decreased exponentially (T90 of approximately 141 h); however, sequences were still detectable for up to 2 weeks. The ability of Pacific oysters (Crassostrea gigas) to bioaccumulate NoV particles (from human feces) and RNA fragments was also compared using qRT-PCR. Oysters exposed to NoV particles subsequently were positive for NoV by qRT-PCR at levels several orders of magnitude in excess of the theoretical limit of detection, whereas oysters exposed to similar quantities of NoV RNA were either negative or positive at significantly lower levels. Therefore, although noninfectious fragments of NoV RNA may persist in the environment under winter conditions, this type of material will not be efficiently bioaccumulated by Pacific oysters and should not significantly contribute to positive qRT-PCR results.

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.

On the skin sensitisation potential of rosin and oxidised rosin.

In the EU rosin is classified as a skin sensitiser, apparently on the basis of its oxidation to sensitising agents. Rosin (gum, tall oil or wood) is not a skin sensitiser when examined in the guinea pig maximisation test (GPMT). Oxidised rosins are sensitisers in the GPMT. Oxidised gum rosin was further tested in the mouse local lymph node assay (LLNA) and the Buehler test, but is not a sensitiser in either of these tests. Further, the outcome of the LLNA can be used to assess the potency of oxidised rosin as an inducing agent in humans, and oxidised rosin is, at most, a weak sensitiser in this test. Thus, oxidised rosin is not a potent inducing agent for skin sensitisation unless the dermal barrier is bypassed and/or there is deliberate use of Freund's Complete Adjuvant to induce greater susceptibility. The material used for human patch testing ('colophony') is in oxidised form. A re-examination of epidemiological studies suggests that patients in dermatological clinics show higher response rates than do the general population or those occupationally exposed to presumably oxidised rosin. Thus, the differences seen in susceptibility in the regulatory tests may be reflected in the human population. These results are discussed in terms of possible testing and classification strategies for dealing with existing chemicals, with particular reference to the new European Union legislation.

Aligning conservation priorities across taxa in Madagascar with high-resolution planning tools.

Globally, priority areas for biodiversity are relatively well known, yet few detailed plans exist to direct conservation action within them, despite urgent need. Madagascar, like other globally recognized biodiversity hot spots, has complex spatial patterns of endemism that differ among taxonomic groups, creating challenges for the selection of within-country priorities. We show, in an analysis of wide taxonomic and geographic breadth and high spatial resolution, that multitaxonomic rather than single-taxon approaches are critical for identifying areas likely to promote the persistence of most species. Our conservation prioritization, facilitated by newly available techniques, identifies optimal expansion sites for the Madagascar government's current goal of tripling the land area under protection. Our findings further suggest that high-resolution multitaxonomic approaches to prioritization may be necessary to ensure protection for biodiversity in other global hot spots.

A sensitive and reliable reverse transcriptase PCR-enzyme-linked immunosorbent assay for the detection of human pathogenic viruses in bivalve molluscs.

A colorimetric method, reverse transcriptase PCR with an enzyme-linked immunosorbent assay (RT-PCR-ELISA) was evaluated for ease of use, reliability, and sensitivity when detecting known human pathogenic virus present in shellfish, using a traditional polyethylene precipitation or immunocapture virus concentration method. The newly developed ELISA method could successfully detect enteroviruses and noroviruses in artificially and naturally contaminated shellfish. Overall, ELISA was shown to be a robust and sensitive method, which had a detection limit of 10 to 100 50% tissue culture infective dose enterovirus per gram of Crassostrea gigas (Pacific oyster) digestive gland and whole Mytilus edulis (common blue mussel). The technique was easily established in a new laboratory and required no specialized equipment. The method had a high sample throughput capable of screening 96 samples per run, making the technique extremely time efficient. RT-PCR-ELISA is a safe, quick, reliable technique, which has the potential for use as a standard virus detection method.

Disinfection of feline calicivirus (a surrogate for Norovirus) in wastewaters.

AIMS: To compare the inactivation of feline calicivirus (FCV) (a surrogate for Norovirus, NV) with the reduction of a bacterial water quality indicator (Escherichia coli), a human enteric virus (poliovirus) and a viral indicator (MS2, FRNA bacteriophage), following the disinfection of wastewaters. METHODS AND RESULTS: Bench-scale disinfection experiments used wastewater (sterilized by gamma-irradiation) seeded with laboratory-cultured organisms. Seeded primary effluent was treated with different doses of applied free chlorine (8, 16 and 30 mg l(-1)). FCV and E. coli were easily inactivated by >4 log10, within 5 min with a dose of 30 mg l(-1) of applied chlorine. Poliovirus was more resistant and a reduction of 2.85 log10 was seen after 30 min, MS2 was the most resistant organism (1 log10 inactivation). In further experiments seeded secondary effluent was treated with different doses of u.v. irradiation. To achieve a 4-log10 reduction of E. coli, FCV, poliovirus and MS2 doses of 5.32, 19.04, 27.51 and 62.50 mW s cm(-2), respectively, were required. CONCLUSIONS: Feline calicivirus and E. coli seeded in primary wastewater were very susceptible to chlorination compared with poliovirus and MS2. In contrast, FCV seeded in secondary wastewater was more resistant to u.v. irradiation than E. coli but more sensitive than poliovirus and MS2. SIGNIFICANCE AND IMPACT OF THE STUDY: FRNA phage was more resistant to inactivation than all the viruses tested. This suggests FRNA phage would be a useful and conservative indicator of virus inactivation following disinfection of wastewaters with chlorination or u.v. irradiation.

Probing Norwalk-like virus presence in shellfish, using artificial neural networks.

A database was examined using artificial neural network (ANN) models to investigate the efficacy of predicting PCR-identified Norwalk-like virus presence and absence in shellfish. The relative importance of variables in the model and the predictive power obtained by application of ANN modelling methods were compared with previously developed logistic regression models. In addition, two country-specific datasets were analysed separately with ANN models to determine if the relative importance of the input variables was similar for geographically diverse regions. The results of this analysis found that ANN models predicted Norwalk-like virus presence and absence in shellfish with equivalent, and better, precision than logistic regression models. For overall classification performance, ANN modelling had a rate of 93%, vs 75% for the logistic regression. ANN models were able to illuminate the site-specific relationships between indicators and pathogens.