Uso de diferentes métodos de conservación de garrapatas Rhipicephalus sanguineus para mejorar la extracción de ADN de bacterias rickettsiales

Resumen

 
 
 

The Rhipicephalus sanguineus tick is a hemoparasite of importance in public health because it is the vector of rickettsiosis, ehrlichiosis, and anaplasmosis in humans. The identification of these agents by molecular diagnosis depends of an efficient obtaining of nucleic acids. The objective of the study was to evaluate different tick conservation and maceration methods for their subsequent use in obtaining DNA. Twenty-six replicates of ten ticks per replica we used for each preservation and maceration method. For the first method, we use liquid nitrogen, in the second, freezing at -80 °C was used, and in the third, it was agreed to dissect and macerate at room temperature. Subsequently, we used a commercial DNA extraction kit and the quantity, quality and purity of the DNA was measured by spectrophotometry and agarose gel. The conservation and maceration of the ticks in liquid nitrogen achieved the highest concentration (400 ng / µl), quality and purity of DNA and amplified 100% of the replicas. With the method of freezing at -80 ° C, up to 210 ng / µl and an amplification of 92.3% were obtained, the procedure carried out at room temperature being in third place, with which a concentration of 100 ng / µl was obtained and an 84.6% amplification. Only the genera Ehrlichia and Anaplasma were positive by PCR. Therefore, it is concluded that with the method of conservation and maceration in liquid nitrogen, DNA will be obtained in quantity and quality that will favor the detection of zoonotic diseases transmitted by ticks.