Potential of Marine Strains of Pseudoalteromonas to Improve Resistance of Juvenile Sea Bass to Pathogens and Limit Biofilm Development.

The European sea bass (Dicentrarchus labrax), one of the most produced marine fish species in Europe, is acutely vulnerable to multiple infectious hazards. In this study, we investigated the potential probiotic effect of some marine Pseudoalteromonas bacterial strains against two major pathogens of this species, Vibrio harveyi and the nervous necrosis virus (NNV), and examined their antibiofilm effect. Impregnation phase was done by repeated immersion of juvenile's sea bass during 8 to 12 weeks in seawater containing the probiotic candidates at a concentration of 106 CFU/mL. Four candidates were tested: (1) a combination of two strains producing antimicrobial compounds, hCg-42 and hOe-125; (2) strain 3J6, with known antibiofilm properties; (3) strain RA15, from the same genus, but with no identified probiotic effect; and (4) a control group without probiotics. At the end of the impregnation phase, fish underwent an infection challenge with V. harveyi or with a pathogenic strain of NNV and mortality was monitored. For the V. harveyi challenge, improved survival rates of 10 and 25% were obtained for the RA15 and the mix hCg-42 + hOe-125-impregnated groups, respectively. For the NNV challenge, no significant benefic effect of the probiotics on infection kinetics or cumulative mortality was observed. At the end of the impregnation phase, the maximal thickness of biofilm was significantly lower in the 3J6, double strain, and RA15 groups, compared with the non-impregnated control group. This study highlights the interesting probiotic potential of marine bacteria to limit mortalities induced by bacterial pathogens as well as biofilm development.

Susceptibility of pike Esox lucius to VHSV and IHNV and potential transmission to rainbow trout Oncorhynchus mykiss.

Determining the origin of recurrent outbreaks of fish diseases occurring on fish farms is essential for disease prevention and control measures. In this study, we investigated the potential reservoir role of wild fish species living near salmonid farms which were regularly found to be positive for viral hemorrhagic septicemia virus (VHSV). In addition to VHSV, infectious hematopoietic necrosis virus (IHNV) was also isolated from several pike Esox lucius samples collected from a pond near the salmonid farms of interest. All isolates of VHSV and IHNV analyzed had 100% identical partial glycoprotein gene sequences. VHSV pike strain OO128-25 belonged to the Ia genotype and shared 99.1 to 99.5% nucleotide identity with strains recently isolated from the farms. IHNV pike strain OO121-8, European genotype, appeared to be different from strains from France characterized since the first isolation in 1987. Isolates representative of both viral species were highly virulent in rainbow trout Oncorhynchus mykiss. OO128-25 induced 65% mortality in pike fingerlings, whereas only weak mortality was observed with OO121-8, despite characteristic symptoms in infected fish. High levels of specific antibodies to VHSV and IHNV were detected in adult pike in the absence of clinical signs. Infection of rainbow trout in contact with experimentally VHSV- or IHNV-infected pike fingerlings indicates possible horizontal transmission. These results suggest that pike could act as a reservoir for VHSV and IHNV in the wild, providing additional evidence to explain viral persistence and resurgence in certain areas.

Influence of Cosmetic Type and Distribution Channel on the Presence of Regulated Fragrance Allergens: Study of 2044 Commercial Products.

Contact dermatitis linked to cosmetic products is a very common reason for visits to the dermatologist, and in more than half the cases, it is due to an allergic reaction. Fragrances are most often the culprit. The aim of the study was to describe the common fragrance allergens in different categories of cosmetic products available on the European market. We wanted to assess the influence of cosmetic type and distribution channel on the presence of fragrance allergens. There are the allergens whose concentration exceeds 0.001% in leave-on products and 0.01% in rinse-off products. A total of 2044 commercial hygiene, care, and makeup cosmetic products were analyzed to specifically study regulated fragrance allergens. The influence that the product category and its distribution channel (retail stores and specialized stores such as beauty institutes or hairdressers and pharmacy) have on the prevalence of these allergens was evaluated. The Kruskal-Wallis test has been used for statistical data analysis. There is a wide range of fragrance allergens, the most common being limonene (found in about 30% of products tested), linalool (just over a quarter of the products tested), and benzyl alcohol (approximately 16% of the products tested). The average number of allergens found and their nature varies depending on the type of product in question (maximum number for shampoos and oral care with about 70 allergens and minimum number for nail polish and makeup for eyes with fewer than 10 allergens). In the area of hygiene, deodorants and oral hygiene products are particularly noteworthy, the former for their significantly high number of allergens and the latter for their low number. There is also a significant difference between the number of allergens found in eye makeup and foundations. Our results indicate that the number of regulated fragrance allergens is particularly influenced by the type of products.

Dye residues in aquaculture products: Targeted and metabolomics mass spectrometric approaches to track their abuse.

The aim of the study was to initiate an exhaustive strategy of control by implementing both targeted and non-targeted metabolomics approaches. A LC-MS/MS method including an oxidative step for most of dyes was developed and validated to target the analysis of 14 residues belonging to different families of dyes. The method was suitable for the quantitative confirmation of 13 dyes at low ppb levels. The metabolomics approach objective was to compare fingerprints between farmed fish treated with malachite green and farmed fish treated with victoria pure blue bo. Analytical information on modifications in the metabolome of muscle, liver and plasma was exploited by HRMS following by multivariate statistics and revealed some direct or endogenous metabolites among relevant mass features contributing to the constructed models. These two approaches, either appropriate biomarkers either enlarged targeted dyes are explored concomitantly to help improving the strategy for tracking new illegal practices in aquaculture.

Validation of a KHV antibody enzyme-linked immunosorbent assay (ELISA).

Koi herpesvirus (KHV) causes KHV disease (KHVD). The virus is highly contagious in carp or koi and can induce a high mortality. Latency and, in some cases, a lack of signs presents a challenge for virus detection. Appropriate immunological detection methods for anti-KHV antibodies have not yet been fully validated for KHV. Therefore, it was developed and validated an enzyme-linked immunosorbent assay (ELISA) to detect KHV antibodies. The assay was optimized with respect to plates, buffers, antigens and assay conditions. It demonstrated high diagnostic and analytical sensitivity and specificity and was particularly useful at the pond or farm levels. Considering the scale of the carp and koi industry worldwide, this assay represents an important practical tool for the indirect detection of KHV, also in the absence of clinical signs.

Viral encephalopathy and retinopathy in aquaculture: a review.

Viral encephalopathy and retinopathy (VER), otherwise known as viral nervous necrosis (VNN), is a major devastating threat for aquatic animals. Betanodaviruses have been isolated in at least 70 aquatic animal species in marine and in freshwater environments throughout the world, with the notable exception of South America. In this review, the main features of betanodavirus, including its diversity, its distribution and its transmission modes in fish, are firstly presented. Then, the existing diagnosis and detection methods, as well as the different control procedures of this disease, are reviewed. Finally, the potential of selective breeding, including both conventional and genomic selection, as an opportunity to obtain resistant commercial populations, is examined.

Validation of a serum neutralization test for detection of antibodies specific to cyprinid herpesvirus 3 in infected common and koi carp (Cyprinus carpio).

Cyprinid herpesvirus 3 (CyHV-3) is the aetiological agent of a serious infective, notifiable disease affecting common carp and varieties. In survivors, infection is generally characterized by a subclinical latency phase with restricted viral replication. The CyHV-3 genome is difficult to detect in such carrier fish that represent a potential source of dissemination if viral reactivation occurs. In this study, the analytical and diagnostic performance of an alternative serum neutralization (SN) method based on the detection of CyHV-3-specific antibodies was assessed using 151 serum or plasma samples from healthy and naturally or experimentally CyHV-3-infected carp. French CyHV-3 isolate 07/108b was neutralized efficiently by sera from carp infected with European, American and Taiwanese CyHV-3 isolates, but no neutralization was observed using sera specific to other aquatic herpesviruses. Diagnostic sensitivity, diagnostic specificity and repeatability of 95.9%, 99.0% and 99.3%, respectively, were obtained, as well as a compliance rate of 89.9% in reproducibility testing. Neutralizing antibodies were steadily detected in infected carp subjected to restrictive or permissive temperature variations over more than 25 months post-infection. The results suggest that this non-lethal diagnostic test could be used in the future to improve the epidemiological surveillance and control of CyHV-3 disease.

Comparison of the virucidal efficacy of peracetic acid, potassium monopersulphate and sodium hypochlorite on bacteriophages P001 and MS2.

AIMS: The phagicidal activity of peroxy products against the virulent bacteriophage P001 infecting lactic acid bacteria and bacteriophage MS2 used as a surrogate of enteric viruses (EVs) was evaluated and compared to sodium hypochlorite using the EN 13610 European suspension test and a surface test developed in our laboratories. METHODS AND RESULTS: Infectivity tests were adapted and/or developed to determine the activity of disinfectants against reference P001 phage of Lactoccocus lactis and F-specific RNA phage MS2 of Escherichia coli in conditions simulating practical use. Similar concentrations of sodium hypochlorite were phagicidal against both bacteriophages, either at 0·05-0·125% of active chlorine using the suspension test or at 0·12-0·5% using the surface test. For Potassium monopersulphate (MPS), phagicidal concentrations varied from 0·006 to 0·012% whatever the type of test and phages. However, for peracetic acid products (PAP) used in suspension, concentrations 55 times higher were necessary against MS2 (0·271%) than against P001 (0·005%). With the surface test, 0·089-0·178% concentrations of PAP were effective against MS2, but these concentrations were 16-32 times greater than needed against P001. CONCLUSIONS: Sodium hypochlorite and MPS had similar phagicidal activities against P001 and MS2, but PAP did not. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first comparative study to investigate through suspension and surface tests the difference in resistance to peroxy compounds between a reference bacteriophage (P001) used to evaluate phagicidal concentrations in European standards and a surrogate of EVs (MS2). Results underline the importance of validation tests on pertinent surrogates of viruses or bacteriophages to adjust the concentration of disinfectants for use in the food and water industries.

Comparison of the virucidal efficiency of peracetic acid, potassium monopersulfate and sodium hypochlorite on hepatitis A and enteric cytopathogenic bovine orphan virus.

AIMS: The virucidal activity of peroxy-products was evaluated and compared with sodium hypochlorite using the EN 14675 European suspension test and a surface test developed in our laboratory. The classical approach on infectivity of viruses was complemented with a prospective approach on virus genomes. METHODS AND RESULTS: Both infectivity tests were adapted and/or developed to determine the activity of disinfectants against reference bovine enterovirus type 1 [enteric cytopathogenic bovine orphan virus (ECBO)] and resistant hepatitis A virus (HAV) in conditions simulating practical use. Similar concentrations of active chlorine were virucidal against both viruses, either at 0·062% using the suspension test or at 0·50-1% using the surface test. However, for potassium monopersulfate and peracetic acid products, concentrations of approximately three times (3%) to 72 times (9%) higher were necessary against HAV than ECBO when determined with the suspension test. With the surface test, 4-8% peroxy-products were virucidal against HAV, either 16 times more peroxy-products concentrations than against ECBO. No significant impact on the targeted area of the viral genome measured by real-time RT-PCRs was obtained for ECBO and HAV suspensions treated with disinfectants, even with doses higher than the minimal virucidal concentrations. CONCLUSIONS: Sodium hypochlorite, but not peroxy-products, had similar activity against ECBO and HAV. No relation could be established between infectivity tests and genome destruction. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first comparative study that investigates with novel suspension and surface tests the reduction of infectivity and genome destruction of two resistant viruses by peroxy-compounds. The results and conclusions collected with European standards are discussed.

The impact of temperature on the inactivation of enteric viruses in food and water: a review.

Temperature is considered as the major factor determining virus inactivation in the environment. Food industries, therefore, widely apply temperature as virus inactivating parameter. This review encompasses an overview of viral inactivation and virus genome degradation data from published literature as well as a statistical analysis and the development of empirical formulae to predict virus inactivation. A total of 658 data (time to obtain a first log(10) reduction) were collected from 76 published studies with 563 data on virus infectivity and 95 data on genome degradation. Linear model fitting was applied to analyse the effects of temperature, virus species, detection method (cell culture or molecular methods), matrix (simple or complex) and temperature category (<50 and ≥50°C). As expected, virus inactivation was found to be faster at temperatures ≥50°C than at temperatures <50°C, but there was also a significant temperature-matrix effect. Virus inactivation appeared to occur faster in complex than in simple matrices. In general, bacteriophages PRD1 and PhiX174 appeared to be highly persistent whatever the matrix or the temperature, which makes them useful indicators for virus inactivation studies. The virus genome was shown to be more resistant than infectious virus. Simple empirical formulas were developed that can be used to predict virus inactivation and genome degradation for untested temperatures, time points or even virus strains.

Influence of HCV genotype 1 subtypes on the virus response to PEG interferon alpha-2a plus ribavirin therapy.

New factors that influence the viral response in HCV non-genotype 2/3 patients must be identified in order to optimize anti-HCV treatment. This multicenter prospective study evaluates the influence of HCV variability and pharmacological parameters on the virological response of these patients to pegylated interferon α2a (peg-IFN-α2a: 180 µg/week) and ribavirin (RBV; 800-1,200 mg/day) for 48 weeks. HCV subtypes were identified by sequencing the NS5B region. Serum RBV and peg-IFN-α2a concentrations were measured at weeks 4 and 12. The 115 patients (67 men; median age = 49, range 31-76) included 64 who had never been treated and 27 co-infected with HIV. The mean baseline HCV RNA was 6.30 ± 0.06 log IU/ml and the HCV genotypes were: G1 (n = 93) with 1a (n = 37) and 1b (n = 50), G4 (n = 20) and G5 (n = 2). Most patients (79/108; 73%) had an early virological response. Independent predictors of an early virological response were interferon naive patients (OR= 2.98, 95% CI: 1.15-7.72) and RBV of >2,200 ng/ml at week 12 (OR = 3.41, 95% CI: 1.31-8.90). Forty of 104 patients (38%) had a sustained virological response. The only independent predictors of a sustained virological response were subtype 1b (OR = 6.82, 95% CI: 1.7-26.8), and HCV RNA <15 IU/ml at week 12 (OR = 25, 95% CI: 6.4-97.6). Thus a serum RBV concentration of >2,200 ng/ml was associated with an early virological response and patients infected with HCV subtype 1b had a better chance of a sustained virological response than did those infected with subtype 1a.

A national survey of acute hepatitis E in France.

BACKGROUND: Few data are available on the incidence, risk factors and contamination pathways involved in acute indigenous hepatitis E in developed countries. AIMS: To draw up an overall picture of hepatitis E cases, to confirm whether or not the majority of the cases were indigenous and to attempt to identify the risk factors and contamination pathways involved in hepatitis E. METHODS: This study was performed in the framework of a national network (ANGH) including 96 participating centres. The 19 centres with at least one case of acute HEV reported a total number of 53 cases. RESULTS: A decreasing South-to-North geographic gradient was observed. A nonspecific clinical profile was observed in many cases. Acute hepatitis E was of indigenous origin in 90% of the patients. The most relevant and/or frequent possible risk factors among the 47 indigenous metropolitan cases were water consumption from a personal water supply, uncooked shellfish consumption and the recent acquisition of a pet pig. CONCLUSIONS: This national survey confirmed that acute indigenous hepatitis E is an emerging disease in developed countries such as France, and suggests that various risk factors are responsible for acute indigenous hepatitis E contamination in non-endemic countries.

[Hepatitis caused by clometacin (Dupéran). Retrospective study of 30 cases. A model of autoimmune drug-induced hepatitis?]. / Hépatites à la clométacine (Dupéran). Etude rétrospective de 30 cas. Un modèle d'hépatite auto-immune médicamenteuse?

Thirty cases of clometacin-induced hepatitis were retrospectively collected over a nine-year period in hepatogastroenterological units of non university, public hospitals. There was a strong female predominance (90 percent). Clometacin (Dupéran) was taken because of arthritis in 8 out of 10 cases. Administration was continuous in 85 percent of cases and median duration was 445 days. median dose was 450 mg per day. Jaundice, fatigue, and weight loss were the most frequent symptoms, but edema, ascites and palmar erythema were not uncommon. Thrombopenia (38 percent) was the most frequent hematologic abnormality. Renal failure, always with benign course, was present in 1/4 of cases. Biochemical disorders indicated hepatocellular and cholestatic hepatitis in 3/4 and 1/4 of cases respectively. Hypoprothrombinemia below 50 percent was noted in 1 out of 6 cases, and was associated with death in half cases. Gamma-globulins were increased in 80 percent of cases, with a predominant increase of IgG. Antinuclear or anti-smooth muscle antibodies were present in 60 percent of cases, whereas antimitochondrial and antimicrosomes were absent. Histopathological examination of the liver biopsy specimens obtained in 25 patients showed acute hepatitis in 8 and chronic active hepatitis with fibrosis in 17--including 6 patients with cirrhosis; there were no epidemiological, clinical (except ascites), or biochemical differences between these two groups. Four of the 7 patients tested had HLA B8 antigens; they all had chronic active hepatitis, with autoantibodies in 3 cases. Median duration of hospitalization was 21 days. Hepatitis was directly responsible for death in 3 patients; biochemical sequelae (hypergammaglobulinemia or anicteric cholestasis) were present in 8 patients, 2 of whom most likely had cirrhosis.(ABSTRACT TRUNCATED AT 250 WORDS)