Congresso Brasileiro de Microbiologia 2023

Congresso Brasileiro de Microbiologia 2023

Resumo: 811-1

811-1

Detection of the IS6110 insertion sequence in the blood of cattle non-responder to the tuberculin skin test.

Autores:

Micaela Encinas (IABIMO - Instituto de Agrobiotecnología y Biología Molecular) ; Sergio Garbaccio (IPVET - Instituto de Patobiología Veterinaria ) ; Romina Sammarruco (IPVET - Instituto de Patobiología Veterinaria ) ; Victoria Ruiz Menna (IPVET - Instituto de Patobiología Veterinaria ) ; Carlos Garro (IPVET - Instituto de Patobiología Veterinaria ) ; Fernando Delgado (IPVET - Instituto de Patobiología Veterinaria ) ; Ximena Ferrara Muñiz (IABIMO - Instituto de Agrobiotecnología y Biología Molecular) ; Martín Zumárraga (IABIMO - Instituto de Agrobiotecnología y Biología Molecular) ; Maria Emilia Eirin (IABIMO - Instituto de Agrobiotecnología y Biología Molecular)

Resumo:

In Argentina, bovine tuberculosis (BTB), caused by Mycobacterium bovis (M. bovis), is an endemic disease that affects livestock species and humans. The tuberculin skin test (TST) is used for the diagnosis and control of BTB. There are infected animals that do not respond to TST, which constitute a challenge for diagnosis and a limitation in sanitation programs since they represent sources of infection in herds. The objective of this work was to determine the presence of the insertion sequence IS6110 in the blood of bovines with confirmed infection but with a non-responder profile for the TST, in local herds. Blood sample was obtained from the jugular vein of 70 dairy cattle of the Holando Argentino breed, negative for the TST. DNA was extracted with the PuriPrep-S kit (InbioHighway) and quantified by spectrophotometry (A260nm). The quality (A260nm/A280nm) and integrity of the genetic material were evaluated by electrophoresis in 0.8 % agarose gel and by PCR amplification of a fraction (450bp) of the mitochondrial 16S ribosomal RNA gene (16SARNrmt). The presence of IS6110 was studied by PCR-touchdown (245bp) followed by 2 % agarose gel electrophoresis. Amplification products were visualized by ethidium bromide staining and exposure to an ultraviolet light source. Out of the total, the presence of disseminated lesions was recorded in the 64 % of the cases, and in a lower proportion (18 %) lesions were confined to one organ. In turn, in 15 % of the samples the presence of M. bovis genetic material was detected by PCR IS6110 in blood. In those samples that were positive by PCR in blood, 81.8 % presented macroscopic lesions compatible with tuberculosis with confirmation of infection by histopathology/Ziehl Neelsen staining or by tissue PCR. In conclusion, the application of the IS6110 PCR-touchdown technique in blood suggest that it could be used as a complementary diagnostic tool in the identification of infected animals that do not react to TST.

Palavras-chave:

bovine tuberculosis, PCR, diagnosis, anergy, blood