COVID-19: Viral whole genome sequencing protocols for routine application at veterinary diagnostic laboratories

Project Summary DNA or RNA sequencing is an important tool for viral strain identification and characterization. During the last ten years advances in next generation sequencing (NGS) technologies have enhanced the capabilities of viral sequencing. However, NGS is expensive and requires advanced equipment and bioinformatics expertise. For those reasons, its adoption as a routine diagnostic service at veterinary diagnostic laboratories (VDLs) has been slow. An additional complication of the implementation of NGS in VDLs is the variety of platforms available, and the lack of practical information available to a laboratory to select the right platform. Some platforms can be less expensive upfront but require a larger expense in supplies per sample. Other platforms may be an inexpensive option, but are not well suited to run multiple samples at a time. On the other hand, some platforms may require more labor than others. Unfortunately, there is probably not a single platform or a single protocol that suits each VDL. There are hundreds of studies being published every week on NGS. However, they are mainly research papers, with results that cannot be readily applied to routine diagnostics. Therefore, there is a need for studies that provide practical recommendations for the use of NGS technologies in routine veterinary diagnostics. The University of Minnesota VDL is well positioned to provide this information. Our current NGS section has access to multiple sequencing platforms, as well as expertise in molecular diagnostics and bioinformatics. The objective of this study is to develop practical recommendations for the application of viral whole genome sequencing at VDLs. These recommendations will be applicable to VDLs in the VetLIRN network and will enhance the sequencing capabilities of the network as a whole. Three different viruses will be sequenced in this study: Severe Acute Respiratory Syndrome Cornavirus-2 (SARS-CoV-2), Porcine Epidemic Diarrhea Virus (PEDV) and Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). Three different platforms will be assessed: Oxford Nanopore's MinIon, Illumina's iSeq and Illumina's MiSeq. Three different batch sizes will be evaluated: 1 sample, 4 samples and 8 samples. These are sample sizes that are realistic for a VDL that is planning on performing one or two runs per week. Three main types of outcome will be evaluated for each different protocol: a) Accuracy; b) Cost: equipment, maintenance, labor (molecular diagnostics technician), labor (bioinformatician); and c) Turnaround time: sample processing time, library preparation time, sequencing time, analysis time. The final report will include tabulated expected outcomes (cost and turnaround time) for each combination of sequencing platform and sample size.