First isolation and whole genome characterization of Porcine
Deltacoronavirus from pigs in Peru
Abstract
Porcine Deltacoronavirus is a newly emergent enteric pathogen affecting
swine farms worldwide. It has been detected in several countries in
Europe, Asia and North America; yet, it has not been reported in South
America. In November 2019, an enteric disease outbreak in a pig farm
located in San Martin, Peru; was reported along with submission of three
intestinal samples from pigs who succumbed to the disease. Samples were
processed for molecular detection by qRT-PCR, viral isolation and
further sequencing analysis. A taqman-based RT-PCR was performed to
differentiate among the most relevant swine enteric coronaviruses
described to date. All samples were positive to Porcine Deltacoronavirus
with a cycle threshold (Ct) value between 9-14, revealing a high viral
load, while testing negative to Porcine Epidemic diarrhea and
Transmissible Gastroenteritis viruses. Following detection, viral
isolation was performed using PK-15 and Vero cell lines. After 5 days of
inoculation, no cytopathic effect was observed. A second blind passage
allowed the observation of cytopathic effect on PK-15 cells, while it
remained absent in Vero cells. A fluorescence test using an anti-N
monoclonal antibody confirmed viral replication. One sample was
processed for whole genome sequencing (NGS). In short, raw reads were
imported into CLC genomics and assembled de novo. Out of 479k
reads generated from the sample, 436k assembled into a 25501 bp contig
which was 99.5% identical to a reference Porcine Deltacoronavirus
strain from US within the North American phylogroup. Yet, there are
relevant differences at the nucleotide and amino acid levels compared to
previously described Porcine Deltacoronavirus strains. Altogether, our
findings represent the first report of Porcine Deltacoronavirus in South
America, its genomic characterization, which provides information of its
evolutionary origin. Thus, this study offers new insights into the
molecular epidemiology of Porcine Deltacoronavirus infections in the
swine industry.