Development of a real-time PCR for the identification of Anaplasma marginale in cattle

Abstract

Anaplasma marginale is a gram-negative, obligate intracellular bacterium that infects cattle, buffalo, deer, and antelope, causing significant economic losses worldwide. In recent years, due to an increase in the area of uncultivated land and uncontrolled use of pastures, there has been a tendency for a sharp increase in the population of blood sucking ticks - carriers of blood-parasitic animal diseases. In this regard, the development of molecular genetics methods for the diagnosis of blood parasitic diseases is relevant for taking effective measures to prevent the spread of tick-borne infections. The lack of sensitivity and specificity of the microscopic and immunological diagnostic methods used, as well as the tendency for a sharp increase in the population of blood-sucking ticks, aggravates the epizootological situation. As part of this research, a real time PCR test system for the identification of Anaplasma marginale was developed. Species-specific primers and a fluorescence-labelled TaqMan probe were developed for the highly conserved gene of the heat shock family protein groEL. The developed realtime PCR test system showed high specificity and sensitivity, allowing the detection of the groEL gene of A. marginale at the level of 8 copies in the reaction. The developed real-time PCR test system can be used for early diagnosis of anaplasmosis in cattle and conducting monitoring studies.