Development and Evaluation of High-Density SNP Arrays for the Eastern Oyster Crassostrea virginica.

The eastern oyster Crassostrea virginica is a major aquaculture species for the USA. The sustainable development of eastern oyster aquaculture depends upon the continued improvement of cultured stocks through advanced breeding technologies. The Eastern Oyster Breeding Consortium (EOBC) was formed to advance the genetics and breeding of the eastern oyster. To facilitate efficient genotyping needed for genomic studies and selection, the consortium developed two single-nucleotide polymorphism (SNP) arrays for the eastern oyster: one screening array with 566K SNPs and one breeders' array with 66K SNPs. The 566K screening array was developed based on whole-genome resequencing data from 292 oysters from Atlantic and Gulf of Mexico populations; it contains 566,262 SNPs including 47K from protein-coding genes with a marker conversion rate of 48.34%. The 66K array was developed using best-performing SNPs from the screening array, which contained 65,893 oyster SNPs including 22,984 genic markers with a calling rate of 99.34%, a concordance rate of 99.81%, and a much-improved marker conversion rate of 92.04%. Null alleles attributable to large indels were found in 13.1% of the SNPs, suggesting that copy number variation is pervasive. Both arrays provided easy identification and separation of selected stocks from wild progenitor populations. The arrays contain 31 mitochondrial SNPs that allowed unambiguous identification of Gulf mitochondrial genotypes in some Atlantic populations. The arrays also contain 756 probes from 13 oyster and human pathogens for possible detection. Our results show that marker conversion rate is low in high polymorphism species and that the two-step process of array development can greatly improve array performance. The two arrays will advance genomic research and accelerate genetic improvement of the eastern oyster by delineating genetic architecture of production traits and enabling genomic selection. The arrays also may be used to monitor pedigree and inbreeding, identify selected stocks and their introgression into wild populations, and assess the success of oyster restoration.

Are Patients Discharged on the HEART Pathway Following Up?

INTRODUCTION: The HEART (history, electrocardiogram [ECG], age, risk factors, troponin) pathway is a useful tool in the emergency department to identify patients that are safe for outpatient evaluation of chest pain. A dedicated HEART Clinic to follow-up versus primary care remains a topic that requires further delineation. We sought to identify how many patients discharged on the HEART pathway specifically followed up with the established HEART Clinic. MATERIALS AND METHODS: This is a secondary analysis of a previously published dataset. In an initial validation study of the HEART Pathway, 625 consecutive subjects were identified via chart review, 449 of which were included. We identified subjects for inclusion in this study if they were found to have a HEART score of 3 or less. Subjects were excluded if they were admitted or if their follow-up was beyond 6 weeks. RESULTS: Of the 449 subjects, 185 met criteria for study inclusion. 125 (67.6%) had follow-up with an average time of 7.94 days (95% CI: 6.54-9.34). Of those, half had additional testing such as ECG, cardiac computed tomography angiography, and treadmill stress testing. The most common clinics for follow-up were the Family Medicine, Internal Medicine, and HEART Clinic representing 35.8, 29, and 18% of the follow-ups, respectively. No subject died, had a myocardial infarction, or required reperfusion. CONCLUSIONS: Of the subjects discharged on the HEART Pathway, 67.6% followed up. Of those subjects that followed up, 18% did so at the HEART Clinic.

Partial validation of a TaqMan real-time quantitative PCR assay for the detection of Panulirus argus virus 1.

The Caribbean spiny lobster Panulirus argus supports important fisheries throughout the greater Caribbean and is also the only known host for the pathogenic virus Panulirus argus virus 1 (PaV1). While discovered nearly 2 decades ago, gaps still exist in our knowledge of PaV1, such as the dose required to establish infection and its viability outside of the host. To help answer such questions and to enhance diagnostic capabilities, we developed a TaqMan real-time quantitative polymerase chain reaction (qPCR) assay for PaV1. Of the advantages offered by qPCR, one of the most important benefits is its ability to accurately quantify viral DNA copies in a clinical sample. The qPCR assay was found to be efficient (mean ± SD: 99.19 ± 4.67%) and sensitive, detecting as few as 10 copies of PaV1 plasmid DNA. Its diagnostic sensitivity and specificity determined using a set of 165 lobster samples (138 from Florida, USA, and 27 from across the Caribbean) were 100 and 84%, respectively. The qPCR assay should thus prove useful as a research tool and for detecting and quantifying PaV1 infection severity in Caribbean spiny lobsters.

Mitotic instability in triploid and tetraploid one-year-old eastern oyster, Crassostrea virginica, assessed by cytogenetic and flow cytometry techniques.

For commercial oyster aquaculture, triploidy has significant advantages. To produce triploids, the principal technology uses diploid × tetraploid crosses. The development of tetraploid brood stock for this purpose has been successful, but as more is understood about tetraploids, it seems clear that chromosome instability is a principal feature in oysters. This paper is a continuation of work to investigate chromosome instability in polyploid Crassostrea virginica. We established families between tetraploids-apparently stable (non-mosaic) and unstable (mosaic)-and normal reference diploids, creating triploid groups, as well as tetraploids between mosaic and non-mosaic tetraploids. Chromosome loss was about the same for triploid juveniles produced from either mosaic or non-mosaic tetraploids or from either male or female tetraploids. However, there was a statistically significant difference in chromosome loss in tetraploid juveniles produced from mosaic versus non-mosaic parents, with mosaics producing more unstable progeny. These results confirm that chromosome instability, as manifested in mosaic tetraploids, is of little concern for producing triploids, but it is clearly problematic for tetraploid breeding. Concordance between the results from cytogenetics and flow cytometry was also tested for the first time in oysters, by assessing the ploidy of individuals using both techniques. Results between the two were non-concordant.