Isolation of Orthopoxvirus vaccinia during an outbreak of vesicular disease in dairy cattle from Viçosa, state of Minas Gerais, southeast Brazil.

In August 2023, an outbreak of vesicular disease clinically suggestive of poxvirus infections, was reported in dairy cows in the municipality of Viçosa, state of Minas Gerais, southeast Brazil. A total of 4 cows, one calf, and one milker were clinically affected in a rural region with history of bovine vaccinia cases. The main objective of the present study was to confirm Orthopoxvirus vaccinia (VACV) infection as the main cause of this outbreak. Swab from the lesions, mostly located in teats, were collected from all 5 non-human animals. Swab samples were submitted to virus isolation in Vero cell cultures and observed daily for cytopathic effect (CPE). Three (60%) out of five samples presented CPE at 48 h post-infection. Cell suspensions and supernatants from all cultures exhibiting CPE were respectively subjected to transmission electron microscopy (TEM), polymerase chain reaction (PCR), and Sanger sequencing for further characterization. Two isolates presented brick-shaped viral particles suggestive of orthopoxvirus infection by TEM. All isolates tested positive for VACV by PCR and were confirmed by nucleotide sequencing. The alignment of the nucleotide sequences of the gene encoding the hemagglutinin protein revealed that all samples were identical among each other. To evaluate the exposure to VACV of other dairy cows in the farm, blood samples of nine cows were collected a month later and their sera tested for neutralizing antibodies for one of the VACV isolates. Two thirds of the cows were seropositive by plaque reduction neutralization test, with PRNT50 titers varying from 20 to ≥ 80, confirming their exposure. Serological and molecular evidences presented here confirmed VACV as the cause of the vesicular outbreak reported in Viçosa. These findings associated to other studies show that VACV remains causing disease in dairy cows and people in the state of Minas Gerais, the largest milk producer in Brazil. Bovine vaccinia has generated economic losses, affecting both cows and their herders. Educational, zootechnical management measures, and viral surveillance remain essential strategies for controlling and preventing new cases in cattle and humans among small-scale dairy producers in Brazil.