A Genome-Wide Association Study for Host Resistance to Ostreid Herpesvirus in Pacific Oysters (Crassostrea gigas).

Ostreid herpesvirus (OsHV) can cause mass mortality events in Pacific oyster aquaculture. While various factors impact on the severity of outbreaks, it is clear that genetic resistance of the host is an important determinant of mortality levels. This raises the possibility of selective breeding strategies to improve the genetic resistance of farmed oyster stocks, thereby contributing to disease control. Traditional selective breeding can be augmented by use of genetic markers, either via marker-assisted or genomic selection. The aim of the current study was to investigate the genetic architecture of resistance to OsHV in Pacific oyster, to identify genomic regions containing putative resistance genes, and to inform the use of genomics to enhance efforts to breed for resistance. To achieve this, a population of ∼1,000 juvenile oysters were experimentally challenged with a virulent form of OsHV, with samples taken from mortalities and survivors for genotyping and qPCR measurement of viral load. The samples were genotyped using a recently-developed SNP array, and the genotype data were used to reconstruct the pedigree. Using these pedigree and genotype data, the first high density linkage map was constructed for Pacific oyster, containing 20,353 SNPs mapped to the ten pairs of chromosomes. Genetic parameters for resistance to OsHV were estimated, indicating a significant but low heritability for the binary trait of survival and also for viral load measures (h2 0.12 - 0.25). A genome-wide association study highlighted a region of linkage group 6 containing a significant QTL affecting host resistance. These results are an important step toward identification of genes underlying resistance to OsHV in oyster, and a step toward applying genomic data to enhance selective breeding for disease resistance in oyster aquaculture.

A highly specific Escherichia coli qPCR and its comparison with existing methods for environmental waters.

The presence of Escherichia coli in environmental waters is considered as evidence of faecal contamination and is therefore commonly used as an indicator in both water quality and food safety analysis. The long period of time between sample collection and obtaining results from existing culture based methods means that contamination events may already impact public health by the time they are detected. The adoption of molecular based methods for E. coli could significantly reduce the time to detection. A new quantitative real-time PCR (qPCR) assay was developed to detect the ybbW gene sequence, which was found to be 100% exclusive and inclusive (specific and sensitive) for E. coli and directly compared for its ability to quantify E. coli in environmental waters against colony counts, quantitative real-time NASBA (qNASBA) targeting clpB and qPCR targeting uidA. Of the 87 E. coli strains tested, 100% were found to be ybbW positive, 94.2% were culture positive, 100% were clpB positive and 98.9% were uidA positive. The qPCR assays had a linear range of quantification over several orders of magnitude, and had high amplification efficiencies when using single isolates as a template. This compared favourably with qNASBA which showed poor linearity and amplification efficiency. When the assays were applied to environmental water samples, qNASBA was unable to reliably quantify E. coli while both qPCR assays were capable of predicting E. coli concentrations in environmental waters. This study highlights the inability of qNASBA targeting mRNA to quantify E. coli in environmental waters, and presents the first E. coli qPCR assay with 100% target exclusivity. The application of a highly exclusive and inclusive qPCR assay has the potential to allow water quality managers to reliably and rapidly detect and quantify E. coli and therefore take appropriate measures to reduce the risk to public health posed by faecal contamination.