Comparison of the Effect of Corn-fermented Protein and Traditional Ingredients on the Fecal Microbiota of Dogs.

Corn-fermented protein (CFP), a co-product from the ethanol industry, is produced using post-fermentation technology to split the protein and yeast from fiber prior to drying. The objective of this study was to determine the effect of CFP compared to traditional ingredients on the fecal microbiota of dogs. The four experimental diets included a control with no yeast and diets containing either 3.5% brewer's dried yeast, 2.5% brewer's dried yeast plus 17.5% distiller's dried grains with solubles, or 17.5% CFP. The experimental diets were fed to adult dogs (n = 12) in a 4 × 4 replicated Latin square design. Fresh fecal samples (n = 48) were analyzed by 16S metagenomic sequencing. Raw sequences were processed through mothur. Community diversity was evaluated in R. Relative abundance data were analyzed within the 50 most abundant operational taxonomic units using a mixed model of SAS. Alpha and beta diversity were similar for all treatments. Predominant phyla among all samples were Firmicutes (73%), Bacteroidetes (15%), Fusobacteria (8%), and Actinobacteria (4%). There were no quantifiable (p > 0.05) shifts in the predominant phyla among the treatments. However, nine genera resulted in differences in relative abundance among the treatments. These data indicate that compared to traditional ingredients, CFP did not alter the overall diversity of the fecal microbiota of healthy adult dogs over 14 days.

Effect of the host genotype at a Porcine Reproductive and Respiratory Syndrome (PRRS) resistance marker on evolution of the modified-live PRRS vaccine virus in pigs.

Porcine reproductive and respiratory syndrome; Porcine reproductive and respiratory syndrome virus; PRRS; Whole genome sequencing; Quasispecies; WUR allele.

Assessment of porcine Rotavirus-associated virome variations in pigs with enteric disease.

Enteric disease is the predominant cause of morbidity and mortality in young mammals including pigs. Viral species involved in porcine enteric disease complex (PEDC) include rotaviruses, coronaviruses, picornaviruses, astroviruses and pestiviruses among others. The virome of three groups of swine samples submitted to the Kansas State University Veterinary Diagnostic Laboratory for routine testing were assessed, namely, a Rotavirus A positive (RVA) group, a Rotavirus co-infection (RV) group and a Rotavirus Negative (RV Neg) group. All groups were designated by qRT-PCR test results for Porcine Rotavirus A, B, C and H such that samples positive for RVA only went in the RVA group, samples positive for > 1 rotavirus went in the RV group and samples negative for all were grouped in the RVNeg group. All of the animals had clinical enteric disease resulting in scours and swollen joints/lameness, enlarged heart and/or a cough. All samples were metagenomic sequenced and analyzed for viral species composition that identified 14 viral species and eight bacterial viruses/phages. Sapovirus and Escherichia coli phages were found at a high prevalence in RVA and RV samples but were found at low or no prevalence in the RVNeg samples. Picobirnavirus was identified at a high proportion and prevalence in RVNeg and RV samples but at a low prevalence in the RVA group. Non-rotaviral diversity was highest in RVA samples followed by RV then RV Neg samples. A sequence analysis of the possible host of Picobirnaviruses revealed fungi as the most likely host. Various sequences were extracted from the sample reads and a phylogenetic update was provided showing a high prevalence of G9 and P[23] RVA genotypes. These data are important for pathogen surveillance and control measures.

Using Next Generation Sequencing to Identify Mutations Associated with Repair of a CAS9-induced Double Strand Break Near the CD4 Promoter.

Double strand breaks (DSBs) in DNA are the most cytotoxic type of DNA damage. Because a myriad of insults can result in these lesions (e.g., replication stress, ionizing radiation, unrepaired UV damage), DSBs occur in most cells each day. In addition to cell death, unrepaired DSBs reduce genome integrity and the resulting mutations can drive tumorigenesis. These risks and the prevalence of DSBs motivate investigations into the mechanisms by which cells repair these lesions. Next generation sequencing can be paired with the induction of DSBs by ionizing radiation to provide a powerful tool to precisely define the mutations associated with DSB repair defects. However, this approach requires computationally challenging and cost prohibitive whole genome sequencing to detect the repair of the randomly occurring DSBs associated with ionizing radiation. Rare cutting endonucleases, such as I-Sce1, provide the ability to generate a single DSB, but their recognition sites must be inserted into the genome of interest. As a result, the site of repair is inherently artificial. Recent advances allow guide RNA (sgRNA) to direct a Cas9 endonuclease to any genome locus of interest. This could be applied to the study of DSB repair making next generation sequencing more cost effective by allowing it to be focused on the DNA flanking the Cas9-induced DSB. The goal of the manuscript is to demonstrate the feasibility of this approach by presenting a protocol that can define mutations that stem from the repair of a DSB upstream of the CD4 gene. The protocol can be adapted to determine changes in the mutagenic potential of DSB associated with exogenous factors, such as repair inhibitors, viral protein expression, mutations, and environmental exposures with relatively limited computation requirements. Once an organism's genome has been sequenced, this method can be theoretically employed at any genomic locus and in any cell culture model of that organism that can be transfected. Similar adaptations of the approach could allow comparisons of repair fidelity between different loci in the same genetic background.

Near-Complete Genome of SARS-CoV-2 Delta (AY.3) Variant Identified in a Dog in Kansas, USA.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) descriptions of infection and transmission have been increasing in companion animals in the past year. Although canine susceptibility is generally considered low, their role in the COVID-19 disease cycle remains unknown. In this study, we detected and sequenced a delta variant (AY.3) from a 12-year-old Collie living with owners that previously tested positive for SARS-CoV-2. It is unclear if the dogs' symptoms were related to SARS-CoV-2 infection or underlying conditions. The whole genome sequence obtained from the dog sample had several unique consensus level changes not previously identified in a SARS-CoV-2 genome that may play a role in the rapid adaptation from humans to dogs. Within the spike coding region, 5/7 of the subconsensus variants identified in the dog sequence were also identified in the closest in-house human reference case. Taken together, the whole genome sequence, and phylogenetic and subconsensus variant analyses indicate the virus infecting the animal originated from a local outbreak cluster. The results of these analyses emphasize the importance of rapid detection and characterization of SARS-CoV-2 variants of concern in companion animals.

Near-Complete Genome Sequences of Vesicular Stomatitis Virus Isolates from the 2020 Outbreak in Kansas.

Here, we report the near-complete genome sequences of vesicular stomatitis virus (VSV) serotype Indiana isolates from the 2020 U.S. outbreak. The sequences were obtained from swabs collected from Kansas horses in July and August. The four genome sequences help improve our understanding of VSV outbreak dynamics in the United States.