Phylodynamic analysis and molecular diversity of the avian infectious bronchitis virus of chickens in Brazil.

Avian infectious bronchitis virus (IBV) is the etiological agent of a highly contagious disease, which results in severe economic losses to the poultry industry. The spike protein (S1 subunit) is responsible for the molecular diversity of the virus and many sero/genotypes are described around the world. Recently a new standardized classification of the IBV molecular diversity was conducted, based on phylogenetic analysis of the S1 gene sequences sampled worldwide. Brazil is one of the biggest poultry producers in the world and the present study aimed to review the molecular diversity and reconstruct the evolutionary history of IBV in the country. All IBV S1 gene sequences, with local and year of collection information available on GenBank, were retrieved. Phylogenetic analyses were carried out based on a maximum likelihood method for the classification of genotypes occurring in Brazil, according to the new classification. Bayesian phylogenetic analyses were performed with the Brazilian clade and related international sequences to determine the evolutionary history of IBV in Brazil. A total of 143 Brazilian sequences were classified as GI-11 and 46 as GI-1 (Mass). Within the GI-11 clade, we have identified a potential recombinant strain circulating in Brazil. Phylodynamic analysis demonstrated that IBV GI-11 lineage was introduced in Brazil in the 1950s (1951, 1917-1975 95% HPD) and population dynamics was mostly constant throughout the time. Despite the national vaccination protocols, our results show the widespread dissemination and maintenance of the IBV GI-11 lineage in Brazil and highlight the importance of continuous surveillance to evaluate the impact of currently used vaccine strains on the observed viral diversity of the country.

A Real-Time Reverse-Transcription Polymerase Chain Reaction for Differentiation of Massachusetts Vaccine and Brazilian Field Genotypes of Avian Infectious Bronchitis Virus.

The avian infectious bronchitis virus is classified into serotypes or genotypes (or both) in different poultry-producing countries of the world. In Brazil, Massachusetts type (Mass), used as a live vaccine, and local field Brazilian variants (genotypes; BR) predominate in the commercial poultry flocks. This study describes the development and validation of two real-time reverse-transcription polymerase chain reactions (RT-qPCR) for the specific detection of Mass and BR genotypes in allantoic fluids and clinical samples. Genotype-specific primers, combined with a generic probe targeted to the S1 gene, originated Mass RT-qPCR and BR RT-qPCR-specific assays. Analytical sensitivity and linearity of these assays were determined in comparison with an IBV generic real-time RT-PCR based on the 5' untranslated region (5'UTR RT-qPCR). Mass RT-qPCR detected five Mass field isolates, three vaccine samples, and one coinfected sample (BR and Mass) while BR RT-qPCR detected 16 BR field isolates. Both assays were linear (R(2) > 0.98), reproducible, and as sensitive as the classical 5'UTR RT-qPCR used to detect IBV. In the analysis of 141 IBV clinical samples, 8 were positive for Mass RT-qPCR, 76 for BR RT-qPCR, and 2 for both assays. In the remaining 55 samples, 25 were positive only for 5'UTR RT-qPCR and 30 were negative for the three assays. In conclusion, both assays were able to detect Mass and BR genotypes, allowing rapid and easy IBV molecular typing from allantoic fluids and clinical samples.