Frequency of enterotoxins, toxic shock syndrome toxin-1, and biofilm formation genes in Staphylococcus aureus isolates from cows with mastitis in the Northeast of Brazil.

Staphylococcus aureus is among the microorganisms more frequently associated with subclinical bovine mastitis. S. aureus may produce several virulence factors. This study aimed at determining the frequency of virulence factors such as enterotoxins, toxic shock syndrome toxin 1, and ica adhesion genes. In addition, we assessed antimicrobial drug resistance in S. aureus isolated from clinical and subclinical cases of mastitis. A total of 88 cows with clinical or subclinical mastitis were sampled, resulting in 38 S. aureus isolates, from which 25 (65.78%) carried toxin genes, including seb, sec, sed, tst, and icaD adhesion gene. These S. aureus isolates belong to 21 ribotypes and three S. aureus strains belonged to the same ribotype producing ica adhesion gene. Approximately 90% of S. aureus strains obtained in our study demonstrated multiple resistance to different antimicrobial agents. The most efficacious antimicrobial agents against the isolates were gentamicin, amoxicillin, and norfloxacin. Gentamicin was the most efficacious agent inhibiting 78.95% of the S. aureus isolates. The least efficacious were penicillin, streptomycin, and ampicillin. Our results can help in understanding the relationship between virulence factors and subclinical mastitis caused by S. aureus. Further research about diversity of S. aureus isolates and genes responsible for the pathogenicity of subclinical mastitis is essential.

Molecular characterization of Corynebacterium pseudotuberculosis isolated from goats using ERIC-PCR.

Corynebacterium pseudotuberculosis, the infectious agent of caseous lymphadenitis (CLA), is responsible for substantial economic losses in goat and sheep production. Molecular characterization of C. pseudotuberculosis isolates by enterobacterial repetitive intergenic consensus (ERIC)-PCR has shown promising results in genotyping strains isolated from sheep with CLA. We evaluated the genetic diversity of C. pseudotuberculosis isolates collected from the Sertão region of the Pernambuco (PE) State, Brazil, and investigated the potential of ERIC-PCR as a tool for the molecular typing of strains of C. pseudotuberculosis isolated from goats. Thirty-two C. pseudotuberculosis strains isolated from goats in the municipalities of Floresta and Ibimirim, PE, C. pseudotuberculosis type strain ATCC 19410, the 1002 vaccine strain, and a field isolate of Rhodococcus equi were fingerprinted using the primers ERIC-1R and ERIC-2 and the primer pair ERIC- 1R+ERIC-2. Using 100% similarity as the cutoff, 8, 10, and 7 genotypes were obtained with ERIC-1-PCR, ERIC-2-PCR, and ERIC-1+2-PCR, respectively. The Hunter-Gaston discriminatory index calculated for the ERIC-1-PCR was 0.75. The index for the ERIC-2-PCR was 0.88, and the index for the ERIC-1+2-PCR was 0.79. Among goat isolates of C. pseudotuberculosis, three, two and four genotypes (found by ERIC-1-PCR, ERIC-2-PCR, and ERIC-1+2-PCR, respectively) had been previously described among sheep isolates from Minas Gerais State, Brazil. These results showed that ERIC-PCR has good discriminatory power and typeability, making it a useful tool for discrimination among C. pseudotuberculosis isolates from goats.

Padronizacão da técnica de imunoistoquímica para o diagnóstico etiológico de rotina da diarréia bovina a vírus / Bovine viral diarrhea diagnostic: immunohistochemistry standardization for routine

The immunohistochemistry standardization for bovine viral diarrhea virus (BVDV) diagnostic was described. The formalin-fixed tissue samples from a heifer with mucosal disease were used as positive control. The validation of the first phase results was performed using samples from an aborted fetus and a calf infected with reference strains of BVDV. The best results were seen using monoclonal antibodies and a commercial kit consisting of labelled streptavidin biotin (LSAB) reagents and the diaminobenzidine (DAB) substrate-chromogen reagent. The immunohistochemistry demonstrated to be an useful method for routine diagnosis for the controll and detection of BVDV infection.

Padronizaçäo da técnica de imunoperoxidase para detecçäo do vírus da diarréia bovina a vírus em cultura de células / Standardization of immunoperoxidase test to detection bovine viral diarrhea virus in cell culture

Este estudo teve como objetivo a padronizaçäo do ensaio de imunoperoxidase em monocamada de células (IPM) para o diagnóstico etiológico da diarréia bovina a vírus (DBV). O teste foi padronizado em monocamada de cultivo primário de pulmäo fetal bovino (PFB) inoculada com as amostras clássicas, citopatogênica (CP) e näo citopatogênica (NCP), do vírus da DBV e testado em amostras biológicas suspeitas processadas no teste clássico de isolamento viral (IV). O método de IPM identificou o vírus da DBV, apresentando melhores resultados com a utilizaçäo do calor como agente fixador, a soroalbumina bovina a 4 por cento em PBS como bloqueador e a revelaçäo com o cromógeno 3-amino-9-etil-carbazol (AEC). Como anticorpos primários, tanto o anticorpo policlonal como o monoclonal forneceram bons resultados

Padronização da técnica de imunoperoxidase para detecção do vírus da diarréia bovina a vírus em cultura de células: Standardization of immunoperoxidase test to detection bovine viral diarrhea virus in cell culture

The aim of this study was to standardize the immunoperoxidase in cell monolayer assay (IPMA) for the etiological diagnosis of bovine viral diarrhea (BVD). The method was standardized in monolayer of primary bovine fetal lung culture inoculated with cytophatic and non-cytophatic classical strains of BVD virus and tested using samples that were considered suspected in the classical technique of viral isolation. The IPMA successfully identified BVD virus and presented better results when heat was used for fixation, BSA 4% solution in PBS was used for blocking and AEC chromogen was used for revelation. Both monoclonal and polycloral antibodies gave good results when used as primary antibodies.

Este estudo teve como objetivo a padronização do ensaio de imunoperoxidase em monocamada de células (IPM) para o diagnóstico etiológico da diarréia bovina a vírus (DBV). O teste foi padronizado em monocamada de cultivo primário de pulmão fetal bovino (PFB) inoculada com as amostras clássicas, citopatogênica (CP) e não citopatogênica (NCP), do vírus da DBV e testado em amostras biológicas suspeitas processadas no teste clássico de isolamento viral (IV). O método de IPM identificou o vírus da DBV, apresentando melhores resultados com a utilização do calor como agente fixador, a soroalbumina bovina a 4% em PBS como bloqueador e a revelação com o cromógeno 3-amino-9-etil-carbazol (AEC). Como anticorpos primários, tanto o anticorpo policlonal como o monoclonal forneceram bons resultados.