Molecular detection of Ehrlichia canis in dogs and organs of their respective ticks

Author(s)

Bruno César Miranda Oliveira

• Document type: Master's Dissertation
• Press: Araçatuba. 2016-05-30.
• Institution: Universidade Estadual Paulista (Unesp). Faculdade de Medicina Veterinária. Araçatuba
• Defense date: 2016-05-02
• Advisor: Katia Denise Saraiva Bresciani

Abstract

The canine erlichiosis presents a high level of occurrence in the routine of the medical clinic of small animals. This bacteria multiplies in the epithelial cells of the intestine, in the hemocytes and in the salivary glands of the tick Riphicephalus sanguineus, where there is a transestadial transmission and probably does not occur a transovarial transmission. The objective of our study was to molecularly detect Ehrlichia canis in samples of lymph nodes and bone marrow of dogs and in the intestines, ovaries and salivary glands of their respective ticks R. sanguineus. Therefore, 720 female arthropods (ten of each animal) were removed from the 72 evaluated dogs. Next, aspirative punches were made of lymph nodes e bone marrow. After the dissection of the ticks, their organs (intestines, ovaries and salivary glands), like the punches samples of lymph nodes and bone marrow of the dogs were tested by Polymerase Chain Reaction (nested-PCR) for amplification of the gene 16S rRNA of E. canis and also tested by the Quantitative Real Time PCR (qPCR) to E. canis, based on a fragment of the gene dsb. The detection of the refer pathogen by the nested-PCR was 80,5% in lymph nodes and 44,4% in bone marrows, showing a significant difference (p<0,05). ). In relation to the nested-PCRs of the tick’s organs, we observed positivity of 22% on the intestines, 11% on the ovaries and 7% on the salivary glands. In the qPCR the detection was of 70,8% and 44,4% on the lymph nodes e bone marrows, respectively (p<0,05) and of 31,9% on the intestines, 10,0% on the ovaries and 15, 2% on the salivary glands of the studied ticks. The absolute quantification on the lymph nodes and bone marrow of the dogs was of 1473,79 Copies of a fragment of the gene dsb of E. canis/µL, in bone marrows of 2080,09 copies/µL and in the intestines, ovaries and salivary glands the quantification were of 1211,53 copies/µL, 2602,01 copies/µL and of 49,23 copies/µL respectively. We concluded that there was a major molecular detection on lymph nodes on the nested-PCR as much as the qPCR and that the bacteria was present and in quantifiable levels for the first time in ovaries of R. sanguineus sensu lato. (AU)

• FAPESP's process: 14/26461-7 - Molecular and serological detection of Ehrlichia canis in dogs and their respective ticks in two endemic regions
• Grantee: Bruno César Miranda Oliveira
• Support Opportunities: Scholarships in Brazil - Master