Detection and Genome Sequence Analysis of Avian Metapneumovirus Subtype A Viruses Circulating in Commercial Chicken Flocks in Mexico

Abstract

Avian metapneumoviruses (aMPV subtypes A-D) are respiratory and reproductive pathogens of poultry. Since aMPV-A was initially reported in Mexico in 2014, there have been no additional reports of its detection in the country. Using nontargeted next-generation sequencing (NGS) of FTA card-spotted respiratory samples from commercial chickens in Mexico, seven full genome sequences of aMPV-A (lengths of 13,288–13,381 nucleotides) were de novo assembled. Additionally, complete coding sequences of genes N (n = 2), P and M (n = 7 each), F and L (n = 1 each), M2 (n = 6), SH (n = 5) and G (n = 2) were reference-based assembled from another seven samples. The Mexican isolates phylogenetically group with, but in a distinct clade separate from, other aMPV-A strains. The genome and G-gene nt sequences of the Mexican aMPVs are closest to strain UK/8544/06 (97.22–97.47% and 95.07–95.83%, respectively). Various amino acid variations distinguish the Mexican isolates from each other, and other aMPV-A strains, most of which are in the G (n = 38), F (n = 12), and L (n = 19) proteins. Using our sequence data and publicly available aMPV-A data, we revised a previously published rRT-PCR test, which resulted in different cycling and amplification conditions for aMPV-A to make it more compatible with other commonly used rRT-PCR diagnostic cycling conditions. This is the first comprehensive sequence analysis of aMPVs in Mexico and demonstrates the value of nontargeted NGS to identify pathogens where targeted virus surveillance is likely not routinely performed.

Unique Variants of Avian Coronaviruses from Indigenous Chickens in Kenya

“…Raw NGS data were preprocessed and analyzed using a nontargeted taxonomic classification and assembly pipeline developed by BASE₂BIO LLC (Oshkosh, WI, USA), as recently described [ 35 ]. The pipeline utilized megahit v 1.2.9 for de novo assembly using default parameters [ 36 ].…”

Complete Genome Sequences of Avian Metapneumovirus Subtype B Vaccine Strains from Brazil

“…Genomic data from nontargeted next-generation sequencing (NGS) of clinical samples can simultaneously identify and pathotype viruses, predict their likelihood of causing severe disease, and provide information about the appropriate vaccines for disease control ( 1 ). During a project on improving NGS for diagnostics of avian pathogens in South American commercial chickens, all samples were preserved by spotting onto Whatman FTA cards (Millipore-Sigma) and shipped at room temperature to USDA-ARS Southeast Poultry Research Laboratory (Athens, GA), and avian metapneumovirus (aMPV; family Pneumoviridae ) was commonly identified ( 2 ). The aMPVs are important respiratory pathogens consisting of subtypes A to D and two unclassified subtypes ( 3 – 8 ).…”

“…Variant sites ( 12 ) with a minor frequency of ≥10% were replaced with IUPAC codes. The assembled consensus sequences were annotated using Geneious Prime v2023.1.1 as recently described ( 2 , 13 ), aligned with published sequences using MAFFT v7.490 ( 14 ), trimmed using trimAl v1.3 ( 15 ), and subjected to phylogenetic analysis using MEGA (maximum likelihood [ML] method; 1,000 bootstrap replicates) ( 16 ). Unless stated otherwise, default parameters were used for all tools.…”

Molecular Characterization of Complete Genome Sequence of an Avian Coronavirus Identified in a Backyard Chicken from Tanzania

“…Quality of the assembly was assessed using BWA-MEM [29] and coverage/artifacts inspected using IGV [30]. The assembled consensus sequences were annotated using Geneious Prime ® v2023.1.2 (www.geneious.com (accessed on 18 January 2023)), as recently described [31,32]. Sequences from this study and other AvCoVs (retrieved from GenBank using BLASTn algorithm) were aligned using MAFFT v7.490 [33], trimmed using trimAl v1.2 [34] and used for Maximum Likelihood phylogenetic analysis in MEGA 11 (1000 bootstrap replicates) [35].…”