Numerous, ongoing outbreaks in Brazilian swine herds have been characterized by vesicular lesions in sows and acute losses of neonatal piglets. The complete genome of Seneca Valley virus (SVV) was identified in vesicular fluid and sera of sows, providing evidence of association between SVV and vesicular disease and viraemia in affected animals.

Porcine epidemic diarrhea virus (PEDV) has caused severe economic losses both recently in the United States (US) and historically throughout Europe and Asia. Traditionally, analysis of the spike gene has been used to determine phylogenetic relationships between PEDV strains. We determined the complete genomes of 93 PEDV field samples from US swine and analyzed the data in conjunction with complete genome sequences available from GenBank (n=126) to determine the most variable genomic areas. Our results indicate high levels of variation within the ORF1 and spike regions while the C-terminal domains of structural genes were highly conserved. Analysis of the Receptor Binding Domains in the spike gene revealed a limited number of amino acid substitutions in US strains compared to Asian strains. Phylogenetic analysis of the complete genome sequence data revealed high rates of recombination, resulting in differing evolutionary patterns in phylogenies inferred for the spike region versus whole genomes. These finding suggest that significant genetic events outside of the spike region have contributed to the evolution of PEDV.

Indexing individual template molecules with a unique identifier (UID) before PCR and deep sequencing is promising for detecting low frequency mutations, as true mutations could be distinguished from PCR errors or sequencing errors based on consensus among reads sharing same index. In an effort to develop a robust assay to detect from urine low-abundant bladder cancer cells carrying well-documented mutations, we have tested the idea first on a set of mock templates, with wild type and known mutants mixed at defined ratios. We have measured the combined error rate for PCR and Illumina sequencing at each nucleotide position of three exons, and demonstrated the power of a UID in distinguishing and correcting errors. In addition, we have demonstrated that PCR sampling bias, rather than PCR errors, challenges the UID-deep sequencing method in faithfully detecting low frequency mutation.

Porcine deltacoronavirus (PDCoV) was identified in multiple states across the United States (US) in 2014. In this study, we investigate the presence of PDCoV in diagnostic samples, which were further categorized by case identification (ID), and the association between occurrence, age, specimen and location between March and September 2014. Approximately, 7% of the case IDs submitted from the US were positive for PDCoV. Specimens were categorized into eight groups, and the univariate analysis indicated that oral fluids had 1.89 times higher odds of detecting PDCoV compared to feces. While the 43-56 day age group had the highest percentage of PDCoV positives (8.4%), the univariate analysis indicated no significant differences between age groups. However, multivariable analysis for age adjusted by specimen indicated the >147 day age group had 59% lower odds than suckling pigs of being positive for PDCoV. The percentage of PDCoV in diagnostic samples decreased to <1% in September 2014. In addition, 19 complete PDCoV genomes were sequenced, and Bayesian analysis was conducted to estimate the emergence of the US clade. The evolutionary rate of the PDCoV genome is estimated to be 3.8×10(-4) substitutions/site/year (2.3×10(-4)-5.4×10(-4), 95% HPD). Our results indicate that oral fluids continue to be a valuable specimen to monitor swineherd health, and PDCoV has been circulating in the US prior to 2014.

Clinical laboratories have adopted next generation sequencing (NGS) as a gold standard for the diagnosis of hereditary disorders because of its analytic accuracy, high throughput, and potential for cost-effectiveness. We describe the implementation of a single broad-based NGS sequencing assay to meet the genetic testing needs at the University of Minnesota. A single hybrid capture library preparation was used for each test ordered, data was informatically blinded to clinically-ordered genes, and identified variants were reviewed and classified by genetic counselors and molecular pathologists. We performed 2509 sequencing tests from August 2012 till December 2017. The diagnostic yield has remained steady at 25%, but the number of variants of uncertain significance (VUS) included in a patient report decreased over time with 50% of the patient reports including at least one VUS in 2012 and only 22% of the patient reports reporting a VUS in 2017 (p = .002). Among the various clinical specialties, the diagnostic yield was highest in dermatology (60% diagnostic yield) and ophthalmology (42% diagnostic yield) while the diagnostic yield was lowest in gastrointestinal diseases and pulmonary diseases (10% detection yield in both specialties). Deletion/duplication analysis was also implemented in a subset of panels ordered, with 9% of samples having a diagnostic finding using the deletion/duplication analysis. We have demonstrated the feasibility of this broad-based NGS platform to meet the needs of our academic institution by aggregating a sufficient sample volume from many individually rare tests and providing a flexible ordering for custom, patient-specific panels.