Optimisation of a PMAxx™-RT-qPCR Assay and the Preceding Extraction Method to Selectively Detect Infectious Murine Norovirus Particles in Mussels

Human noroviruses are a major cause for gastroenteritis outbreaks. Filter-feeding bivalve molluscs, which accumulate noroviruses in their digestive tissues, are a typical vector for human infection. RT-qPCR, the established method for human norovirus detection in food, does not allow discrimination between infectious and non-infectious viruses and can overestimate potentially infectious viral loads. To develop a more accurate method of infectious norovirus load estimation, we combined intercalating agent propidium monoazide (PMAxx™)-pre-treatment with RT-qPCR assay using in vitro-cultivable murine norovirus. Three primer sets targeting different genome regions and diverse amplicon sizes were used to compare one-step amplification of a short genome fragment to three two-step long-range RT-qPCRs (7 kbp, 3.6 kbp and 2.3 kbp amplicons). Following initial assays performed on untreated infectious, heat-, or ultraviolet-inactivated murine noroviruses in PBS suspension, PMAxx™ RT-qPCRs were implemented to detect murine noroviruses subsequent to their extraction from mussel digestive tissues; virus extraction via anionic polymer-coated magnetic beads was compared with the proteinase K-dependent ISO norm. The long-range RT-qPCR process detecting fragments of more than 2.3 kbp allowed accurate estimation of the infectivity of UV-damaged murine noroviruses. While proteinase K extraction limited later estimation of PMAxx™ pre-treatment effects and was found to be unsuited to the assay, magnetic bead-captured murine noroviruses retained their infectivity. Genome copies of heat-inactivated murine noroviruses differed by 2.3 log₁₀ between RT-qPCR and PMAxx™-RT-qPCR analysis in bivalve molluscs, the PMAxx™ pre-treatment allowing a closer approximation of infectious titres. The combination of bead-based virus extraction and PMAxx™ RT-qPCR thus provides a more accurate model for the estimation of noroviral bivalve mollusc contamination than the conjunction of proteinase K extraction and RT-qPCR and has the potential (once validated utilising infectious human norovirus) to provide an added measure of security to food safety authorities in the hazard assessment of potential bivalve mollusc contamination.

     

Assessment of Viral Contamination of Five Brazilian Artisanal Cheese Produced from Raw Milk: a Randomized Survey

Food and Environmental Virology - Enteric viruses have been described as important contaminants in fresh and ready-to-eat foods such as sandwiches, deli meat and dairy products. This is a...     

The impact of COVID-19 pandemic in medical waste amounts: a case study from a high-populated city of Turkey

Journal of Material Cycles and Waste Management - In this study, the amount of medical waste generated in a high-populated city in Turkey between January 2018 and August 2021 was evaluated, and the...     

The combination of chemical,structural, and functional indicators to evaluate the anthropogenic impacts on agricultural stream ecosystems

Environmental Science and Pollution Research - Freshwater contamination by pesticides in agricultural landscapes is of increasing concern worldwide, with strong pesticide impacts on biodiversity,...     

Distance-based eigenvector maps (DBEM) to analyse metapopulation structure with irregular sampling

Characterizing spatial patterns due to ecological processes is a major issue for analysing and predicting species distributions. Grasping the non-linear nature of population dynamics over networks of discrete suitable sites is here central, as very specific signatures are expected. In the line of promising results from Fourier analysis of metapopulation maps, we found distance-based eigenvector maps (DBEM) to help disentangle the respective signatures of habitat and metapopulation structuring, with the great advantage of being applicable to irregular sampling schemes, a common feature of ecological surveys. A smoothing procedure was required to obtain the distinguishable signatures, and this may be a critical issue for investigating non-contingent and reliable patterns in spatial ecology.     

Studying ecological communities from a neutral standpoint: A review of models’ structure and parameter estimation

Neutral models provide an alternative to niche-based assembly rules of ecological communities by assuming that communities’ properties are shaped by the stochastic interplay between ecological drift, migration and speciation. The recent and ongoing interest about neutral assumptions has produced many developments on the theoretical side, with nevertheless limited echoes in terms of analyses of real-world data. The present review paper aims to help bridge the widening gap between modellers and field ecologists through two objectives. First, to provide a multi-criteria typology of the main neutral models, including those from population genetics that have not yet been transposed to ecology, by considering how the fundamental processes of ecological drift, speciation and migration are modelled and, specifically, how space is taken into account. Second, to review methods recently proposed to estimate models parameters from field data, a point that should be mastered to allow for broader applications.     

Are sea snakes pertinent bio-indicators for coral reefs? a comparison between species and sites

Classical sampling methods often miss important components of coral reef biodiversity, notably organisms that remain sheltered within the coral matrix. Recent studies using sea kraits (sea snakes) as bio-indicators suggest that the guild of predators represented by anguilliform fish (Congridae, Muraenidae, Ophichthidae, henceforth “eels” for simplicity) were far more abundant and diverse than previously suspected. In the current study, eel diversity (similarity and species richness indices) estimated via sea snake sampling (SSS) was compared among six areas of one of the main oceanic biodiversity hotspot of the Pacific Ocean (southwest lagoon of New Caledonia). Based on the eel diversity in the snakes’ diet, the results obtained in six areas, in two snake species, and using different estimates (ANOSIM, Shannon index…) were consistent, suggesting that SSS provided robust information. Analyses also suggested subtle, albeit significant, differences in the eel assemblages among islets. Such spatial differences are discussed in light of local management practices. As SSS is easy to use, cost-effective, and provides the best picture of eel assemblages to date, it can be employed to monitor the eel assemblages in addition to the snakes themselves in many areas of the Indo-Pacific Ocean, thereby providing an index of the top predator biodiversity of many coral reefs.     

An individual-based model to study the reproduction of egg bearing copepods: Application to Eurytemora affinis (Copepoda Calanoida) from the Seine estuary,France

Limited empirical studies have elucidated the daily egg production and associated reproductive processes of egg bearing copepod. Herein, we present an individual-based model which constitutes a realistic representation of the reproduction in egg bearing copepods. The model has been parameterized using an extensive set of experimental data obtained from the literature and from the laboratory and field experiments on the estuarine copepod Eurytemora affinis. The proposed model takes into account the adult female longevity, the clutch size and interclutch duration, which is a function of egg maturation time and latency time required by the female after egg hatching to produce a new clutch. The embryonic development time and hatching success are also taken into account. The effect of temperature on the means and variances of above-mentioned reproductive parameters has been also incorporated. A multi agent system based generic platform “Mobidyc” has been used to generate and calibrate the model. The model demonstrates the reproductive parameters of females of E. affinis which is validated through individual based experiments. Temperature specific simulations provide a dynamical explanation of temperature effect on the cumulative egg production. The daily survival principally affects the number of clutches produced per female during its life span. The results obtained in the present study by combining temperature and survival effects reveal the relatively greater importance of the first factor on the daily egg production of egg-carrying copepods. The present model is generic and hence easily applicable to other animals with comparable reproductive strategy.     

Composition of plastics from waste electrical and electronic equipment (WEEE) by direct sampling

This paper describes a direct analysis study carried out in a recycling unit for waste electrical and electronic equipment (WEEE) in Portugal to characterize the plastic constituents of WEEE. Approximately 3400 items, including cooling appliances, small WEEE, printers, copying equipment, central processing units, cathode ray tube (CRT) monitors and CRT televisions were characterized, with the analysis finding around 6000 kg of plastics with several polymer types. The most common polymers are polystyrene, acrylonitrile-butadiene-styrene, polycarbonate blends, high-impact polystyrene and polypropylene. Additives to darken color are common contaminants in these plastics when used in CRT televisions and small WEEE. These additives can make plastic identification difficult, along with missing polymer identification and flame retardant identification marks. These drawbacks contribute to the inefficiency of manual dismantling of WEEE, which is the typical recycling process in Portugal. The information found here can be used to set a baseline for the plastics recycling industry and provide information for ecodesign in electrical and electronic equipment production.     

The impact of natural and anthropogenic hydrocarbons on the tropospheric budget of carbon monoxide

A method to quantify the relative contributions of surface sources and photochemical production of atmospheric carbon monoxide has been implemented in a three-dimensional chemical-transport model. The impact of biogenic and anthropogenic hydrocarbons has been calculated. The oxidation of isoprene contributes to about 10% of the global tropospheric burden of carbon monoxide, with a maximum contribution over southern America and Africa. Oxidation of methane and terpenes contribute to 28 and 2%, respectively, of the tropospheric burden of CO. The oxidation of the other hydrocarbons, which include ethane, propane, ethylene, propylene and the surrogate hydrocarbon representing other hydrocarbons results in 12% of the CO tropospheric burden, among which 69% results from the oxidation of hydrocarbons of biologic origin. The overall global CO yield from the oxidation of isoprene is estimated to be 23% on a carbon basis. Comparisons between model results and the few available observations of isoprene, terpenes and their oxidation products show that there is no evidence that the current global isoprene emissions proposed in the IGAC/GEIA emissions data base are substantially overestimated, as suggested by previous studies.