Metagenomic sequencing determines complete infectious bronchitis virus (avian Gammacoronavirus) vaccine strain genomes and associated viromes in chicken clinical samples

Abstract

Infectious bronchitis virus (IBV, genus Gammacoronavirus ) causes an economically important and highly contagious disease in chicken. Random primed RNA sequencing was applied to two IBV positive clinical samples and one in ovo-passaged virus. The virome of a cloacal swab pool was dominated by IBV (82% of viral reads) allowing de novo assembly of a GI-13 lineage complete genome with 99.95% nucleotide identity to vaccine strain 793B. In addition, substantial read counts (16% of viral reads) allowed the assembly of a near-complete chicken astrovirus genome, while lower read counts identified the presence of chicken calicivirus and avian leucosis virus. Viral reads in a respiratory/intestinal tissue pool were distributed between IBV (22.53%), Sicinivirus (Picornaviridae, 24%), and avian leucosis virus (37.04%). A complete IBV genome with 99.95% nucleotide identity to vaccine strain H120 (lineage GI-1), as well as a near-complete avian leucosis virus genome and a partial Sicinivirus genome were assembled from the tissue sample data. Lower read counts identified chicken calicivirus, Avibirnavirus (infectious bursal disease virus, assembling to 98.85% of segment A and 69.66% of segment B closely related to D3976/1 from Germany, 2017) and avian orthoreovirus, while three avian orthoavulavirus 1 reads confirmed prior real-time RT-PCR result. IBV sequence variation analysis identified both fixed and minor frequency variations in the tissue sample compared to its in ovo-passaged virus. Metagenomic methods allow the determination of complete coronavirus genomes from clinical chicken samples while providing additional insights in RNA virus sequence diversity and coinfecting viruses potentially contributing to pathogenicity. Supplementary Information The online version contains supplementary material available at 10.1007/s11262-021-01872-7.

Epidemiology of Infectious Bursal Disease Virus in Poland during 2016–2022

“…However, the analysis confirmed the presence of additional eight aa substitutions, but four of them were also found in the Belgian A3B1 reassortant chicken/4439_001/2020 (Accession no. MZ367373 and MZ367374), such as R 133 in VP5, K 359 in VP2, N 787 and V 951 in VP3; and A 667 in VP1 [ 30 ], which showed that there were at least two lineages within this group of reassortants. Unfortunately, only three full genomic sequences of A3B1 strains are available in the GenBank database, making it difficult to determine their diversity at the genetic level.…”

Metagenomic Next-Generation Sequencing of Cerebrospinal Fluid for the Diagnosis of Cerebral Aspergillosis

“…We assessed the probability of A. fumigatus detection given an SDSSRN of 2, the former standard for A. fumigatus detection. We hypothesised that the probability of mapping a single PMseq read to the A. fumigatus genome is a Bernoulli process, as suggested by Ebinger et al, meaning that the read either mapped correctly or incorrectly (Van Borm et al, 2021). Therefore, we expected the detection probability of A. fumigatus to follow a binomial mass function with increasing SDSSRN.…”

An Amplicon-Based Application for the Whole-Genome Sequencing of GI-19 Lineage Infectious Bronchitis Virus Directly from Clinical Samples

“…This method involves random priming and avoids the selection of specific RNA sequences, thus minimizing the biases associated with amplicon-based sequencing and allowing the detection of multiple pathogens [ 22 , 23 ]. However, in clinical samples, the low viral genetic loads and the overwhelming proportion of sequencing reads from the host DNA pose challenges to achieving the complete genome sequences of viruses by metagenomic sequencing [ 29 , 30 , 40 , 41 ]. In this study, the clinical samples, AD04 and AQ10, exhibited relatively low proportions of mapped reads to the reference genome, at 0.85% and 44.63%, respectively, demonstrating the limitation in obtaining the complete genome sequences of IBV directly from clinical samples using SISPA-mNGS.…”

“…Generally, these methods have been applied to IBV isolates amplified by serial passage in embryonated chicken eggs [ 26 , 27 , 28 ]. However, despite the need for the mNGS of viruses in clinical tissue samples, variations in amplification due to the low viral genetic load in tissue samples can pose challenges for metagenomic sequencing [ 29 , 30 ]. To circumvent this difficulty, different enrichment techniques employing specific primers targeting the whole genome of the virus have been developed.…”