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

Avian metapneumovirus (aMPV) causes a highly contagious upper respiratory and reproductive disease in chickens, turkeys, and ducks. Here, complete genome sequences of aMPV-B vaccine strains BR/1890/E1/19 (PL21, Nemovac; Boehringer Ingelheim Animal Health, Brazil) and BR/1891/E2/19 (1062; Hipraviar, France) were sequenced and compared with the pathogenic field strain VCO3/60616.

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

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.

Genome Sequence Variations of Infectious Bronchitis Virus Serotypes From Commercial Chickens in Mexico.

New variants of infectious bronchitis viruses (IBVs; Coronaviridae) continuously emerge despite routine vaccinations. Here, we report genome sequence variations of IBVs identified by random non-targeted next generation sequencing (NGS) of vaccine and field samples collected on FTA cards from commercial flocks in Mexico in 2019-2021. Paired-ended sequencing libraries prepared from rRNA-depleted RNAs were sequenced using Illumina MiSeq. IBV RNA was detected in 60.07% (n = 167) of the analyzed samples, from which 33 complete genome sequences were de novo assembled. The genomes are organized as 5'UTR-[Rep1a-Rep1b-S-3a-3b-E-M-4b-4c-5a-5b-N-6b]-3'UTR, except in eight sequences lacking non-structural protein genes (accessory genes) 4b, 4c, and 6b. Seventeen sequences have auxiliary S2' cleavage site located 153 residues downstream the canonically conserved primary furin-specific S1/S2 cleavage site. The sequences distinctly cluster into lineages GI-1 (Mass-type; n = 8), GI-3 (Holte/Iowa-97; n = 2), GI-9 (Arkansas-like; n = 8), GI-13 (793B; n = 14), and GI-17 (California variant; CAV; n = 1), with regional distribution in Mexico; this is the first report of the presence of 793B- and CAV-like strains in the country. Various point mutations, substitutions, insertions and deletions are present in the S1 hypervariable regions (HVRs I-III) across all 5 lineages, including in residues 38, 43, 56, 63, 66, and 69 that are critical in viral attachment to respiratory tract tissues. Nine intra-/inter-lineage recombination events are present in the S proteins of three Mass-type sequences, two each of Holte/Iowa-97 and Ark-like sequence, and one each of 793B-like and CAV-like sequences. This study demonstrates the feasibility of FTA cards as an attractive, adoptable low-cost sampling option for untargeted discovery of avian viral agents in field-collected clinical samples. Collectively, our data points to co-circulation of multiple distinct IBVs in Mexican commercial flocks, underscoring the need for active surveillance and a review of IBV vaccines currently used in Mexico and the larger Latin America region.

Phylogenetic analysis, molecular changes, and adaptation to chickens of Mexican lineage H5N2 low-pathogenic avian influenza viruses from 1994 to 2019.

The Mexican lineage H5N2 low pathogenic avian influenza viruses (LPAIVs) were first detected in 1994 and mutated to highly pathogenic avian influenza viruses (HPAIVs) in 1994-1995 causing widespread outbreaks in poultry. By using vaccination and other control measures, the HPAIVs were eradicated but the LPAIVs continued circulating in Mexico and spread to several other countries. To get better resolution of the phylogenetics of this virus, the full genome sequences of 44 H5N2 LPAIVs isolated from 1994 to 2011, and 6 detected in 2017 and 2019, were analysed. Phylogenetic incongruence demonstrated genetic reassortment between two separate groups of the Mexican lineage H5N2 viruses between 2005 and 2010. Moreover, the recent H5N2 viruses reassorted with previously unidentified avian influenza viruses. Bayesian phylogeographic results suggested that mechanical transmission involving human activity is the most probable cause of the virus spillover to Central American, Caribbean, and East Asian countries. Increased infectivity and transmission of a 2011 H5N2 LPAIV in chickens compared to a 1994 virus demonstrates improved adaptation to chickens, while low virus shedding, and limited contact transmission was observed in mallards with the same 2011 virus. The sporadic increase in basic amino acids in the HA cleavage site, changes in potential N-glycosylation sites in the HA, and truncations of PB1-F2 should be further examined in relation to the increased infectivity and transmission in poultry. The genetic changes that occur as this lineage of H5N2 LPAIVs continues circulating in poultry is concerning not only because of the effect of these changes on vaccination efficacy, but also because of the potential of the viruses to mutate to the highly pathogenic form. Continued vigilance and surveillance efforts, and the pathogenic and genetic characterization of circulating viruses, are required for the effective control of this virus.