Validación de la técnica de PCR en tiempo real para el diagnóstico de Coxiella Burnetii como agente causal de fiebre Q

Abstract

Q fever, caused by Coxiella burnetii, is a zoonotic disease that affects both humans and animals. It is primarily transmitted through the inhalation of contaminated aerosols. Although most cases are asymptomatic or mild, it can lead to serious complications such as pneumonia or endocarditis. Serological diagnosis presents limitations, as specific antibodies appear only weeks after infection, making detection in the acute phase difficult. Real-time PCR (qPCR) offers a more effective alternative, allowing for early and accurate identification of C. burnetii. This study validated the use of qPCR as a diagnostic method in Ecuador, a country with limited information about the bacterium. Primers and a probe were designed from sequences of the IS1111 gene using bioinformatics tools such as Primer-BLAST to select specific sequences. Relevant genes, such as isocitrate dehydrogenase (icd), com1, and others, were chosen for primer design. The sequences were processed using the UNGEN software, allowing for the selection of the most precise sequences. The tests were conducted at the National Institute of Public Health and Research (INSPI), using samples of mouse liver and bovine blood. The optimization of qPCR showed a linear relationship between the concentration of the analyte and the threshold cycle, confirming the sensitivity and reliability of the method. Although the evaluated samples did not show positive amplification, the positive control of C. burnetii amplified correctly, demonstrating that the selected primers are suitable. The qPCR is a promising tool for the early detection of C. burnetii in Ecuador. The optimization of the primers and the validation of the method strengthen its potential to improve surveillance and diagnosis of Q fever, contributing to better control of the disease in the country.