The Role of Viral Population Diversity in Adaptation of Bovine Coronavirus to New Host Environments

Abstract

The high mutation rate of RNA viruses enables a diverse genetic population of viral genotypes to exist within a single infected host. In-host genetic diversity could better position the virus population to respond and adapt to a diverse array of selective pressures such as host-switching events. Multiple new coronaviruses, including SARS, have been identified in human samples just within the last ten years, demonstrating the potential of coronaviruses as emergent human pathogens. Deep sequencing was used to characterize genomic changes in coronavirus quasispecies during simulated host-switching. Three bovine nasal samples infected with bovine coronavirus were used to infect human and bovine macrophage and lung cell lines. The virus reproduced relatively well in macrophages, but the lung cell lines were not infected efficiently enough to allow passage of non lab-adapted samples. Approximately 12 kb of the genome was amplified before and after passage and sequenced at average coverages of nearly 950×(454 sequencing) and 38,000×(Illumina). The consensus sequence of many of the passaged samples had a 12 nucleotide insert in the consensus sequence of the spike gene, and multiple point mutations were associated with the presence of the insert. Deep sequencing revealed that the insert was present but very rare in the unpassaged samples and could quickly shift to dominate the population when placed in a different environment. The insert coded for three arginine residues, occurred in a region associated with fusion entry into host cells, and may allow infection of new cell types via heparin sulfate binding. Analysis of the deep sequencing data indicated that two distinct genotypes circulated at different frequency levels in each sample, and support the hypothesis that the mutations present in passaged strains were “selected” from a pre-existing pool rather than through de novo mutation and subsequent population fixation.

Ultra-Deep Sequencing of Intra-host Rabies Virus Populations during Cross-species Transmission

“…Sequencing Center. Sequencing was performed as described previously [17,18]. For all samples sequenced by Illumina (pairedend read technology), overlapping read pairs (ORPs), generated by combining short fragment libraries with long sequencing reads, was used to reduce sequencing errors and improve rare variant detection accuracy.…”

Genomic and single nucleotide polymorphism analysis of infectious bronchitis coronavirus

“…Another member of this genus, the recently-discovered Middle East Respiratory Syndrome (MERS) coronavirus (MERS-CoV) has claimed 88 human lives from 212 cases since April 2012, and dromedary camels are the suspected reservoir (Briese et al, 2014). Genus Gammacoronavirinae includes strains infecting birds and whales (Woo et al, 2012;McBride et al, 2014;Borucki et al, 2013) and deltacoronaviruses have been described in birds, swine and cats (Woo et al, 2012). The diversity of hosts and genomic features amongst CoVs have been attributed to their unique mechanism of viral recombination, a high frequency of recombination, and an inherently high mutation rate (Lai and Cavanagh, 1997).…”

Molecular stability of a vaccine strain of Canine coronavirus after serial passages in A72 cells

“…Though the region of the S gene targeted in this study is responsible for membrane fusion and syncytial formation and has a higher mutation pressure than the S1 region, responsible for binding to cellular receptors (LAI; CAVANAGH, 1997), nucleotide polymorphism may also be found in S2 region (GALLAGHER; BUCHMEIER, 2001) allowing also virus entry into a variety of types of cells in trypsin-independent way (BELOUZARD et al, 2009;BORUCKI et al, 2013).…”