ABSTRACT Respiratory tract infections in poultry have become serious problems with negative consequences for poultry production. Among these diseases, mycoplasmosis and avian pneumoviruses stand out, which, despite being relevant in poultry health, have not been systematically investigated in Brazil, especially regarding the interaction between these agents or the occurrence of co-infection in broiler chickens. The present study investigated the occurrence of infection by Mycoplasma gallisepticum (MG) and subtypes A and B of avian metapneumovirus (aMPV) in broiler chickens from commercial poultry flocks kept on farms with high biosecurity standards and located in the Southeast and Midwest regions of Brazil. Polymerase chain reaction (PCR) and reverse transcription nested-PCR techniques were applied to detect and/or identify, respectively, MG and aMPV in samples of nasal and tracheal swabs collected between 2017 and 2018, from 87 batches of broiler chickens from 15 commercial production farms that showed respiratory clinical signs. Two out of 87 batches sampled (2.3%) were positive for MG, while none of them were positive for aMPV. The low or no incidence of these pathogens can be explained by the adoption of increasingly effective health control measures for these agents on farms with high biosecurity standards. In addition, there are clues that other bacterial and viral infectious agents may be involved in the etiology of respiratory problems of these broiler chickens, which showed clinical signs of respiratory diseases upon sample collection. Brazil coinfection co (MG (aMPV PCR (PCR nestedPCR nested andor identify respectively 201 2018 8 1 2.3% 23 2 3 (2.3% addition collection 20 2.3 (2.3 2. (2. (2 (

Anticorpos monoclonais constituem a base de vários testes usados na detecção e na identificação de antígenos. Nesse contexto, tais imuno-reagentes têm sido extensivamente empregados na identificação de estirpes virais envolvidas na etiologia de surtos de bronquite infecciosa a campo, permitindo o aperfeiçoamento das técnicas de detecção e caracterização antigênica do vírus da bronquite infecciosa das galinhas (VBI). No presente estudo, uma biblioteca de fragmentos de anticorpos de galinha originalmente preparada por phage display contra a estirpe vacinal (H120) do VBI foi usada para a seleção de fragmentos de anticorpos recombinantes com reatividade cruzada para as estirpes heterólogas IBVPR01 e IBVPR05, isoladas de surtos a campo no Brasil e a estirpe SE-17, isolada nos Estados Unidos. Após três ciclos de panning, foi identificado, pelo ELISA, um conjunto de 15 anticorpos scFv expressos em fagos e com reatividade cruzada para essas mesmas estirpes do VBI. A análise por Western-blotting revelou que dois desses clones apresentavam fagos expressando fragmentos de anticorpos monoclonais com reatividade cruzada para a nucleoproteína N das três estirpes do VBI e também para a forma recombinante dessa nucleoproteína derivada da estirpe M41. Concluindo, os fragmentos de anticorpos monoclonais recombinantes scFv-N produzidos em fagos interagem com um epítopo mais conservado da proteína N do VBI e apresentam um grande potencial para utilização na detecção e no diagnóstico direto desse vírus.

Monoclonal antibodies are the basis of various techniques used for antigen detection or characterization, and their use is specially recommended for the identification of viral strains involved in the etiology of infectious bronchitis outbreaks. These antibodies are homogeneous, highly specific and fully characterizable, allowing the improvement of immunological techniques detection and antigenic characterization of avian infectious bronchitis virus strains (IBV). A phage display library was used, which was prepared previously against the IBV vaccine strain (H120) for the selection of new scFv antibody fragments specific for heterologous IBV strains isolated from outbreaks in Brazil (IBVPR01, IBVPR05) and USA (SE-17). After three cycles of panning, a set of 15 scFv antibodies were expressed in phages and cross-reacted in ELISA with these three viral strains. Western-blotting analysis showed that two of the clones were expressing scFv specific for the nucleoprotein of these IBV strains, as well as to the recombinant form of this protein derived from M41. In conclusion, the recombinant fragments of monoclonal antibodies expressed in phage have a great potential for future use in immunodiagnostic techniques and to study the evolution of infectious bronchitis virus.