TaqMan-Based Real-Time PCR for Diagnosis of Multiple Tick-Transmitted Diseases among the Bovine Population in Thrissur and Wayanad Districts, Kerala State, India | Chapter 3 | Microbiology and Biotechnology Research: An Overview Vol. 4

TaqMan PCR (Polymerase Chain Reaction) is a type of real-time PCR, and it uses a nucleic-acid probe complementary to an internal segment of the target DNA. The present study aimed to use TaqMan-based qualitative Real Time PCR (qPCR) as an aid in accurate and confirmatory diagnosis up to species level for multiple tick-transmitted pathogens affecting the bovine population. Conventional diagnostic methods based on microscopy will not be sufficient to identify multiple tick-transmitted pathogens, pathogens having similar infective stages and their identification up to species level. A total of 60 peripheral blood samples in EDTA were collected randomly from five different private cattle farms in Wayanad and Thrissur districts, Kerala state. Peripheral blood in EDTA was stored at-80°C until usage and processed for preparation of template DNA as per the protocols described in the High Pure PCR template preparation kit. Primer efficiency testing was conducted by plotting a standard curve. Custom-synthesised positive clone in pUGM plasmid (with initial concentration 95ng/µl for A. phagocytophilum, and 10ng/µl for others) was used for primer efficiency testing and standardisation of cycling conditions. The average Ct value observed for 38 positive samples for the Theileria genus was 12.79 cycles, ranging from 4.1 to 36.19 cycles. The average Ct value observed for 31 positive samples for M. haemocanis was 22.42 cycles, ranging from 2.66 to 37.6 cycles. The average Ct value observed for 22 positive samples for P. multocida was 20.20 cycles, ranging from 2.74 to 35.72 cycles. Lower Ct/Cq values corresponded to a greater amount of initial template. In most of the tick-transmitted diseases among the bovine population presented for treatment, P. multocida is causing severe problems as a secondary pathogen; the reason may be attributed to suppression of adaptive immune response by the primary tick-transmitted pathogens. This data should give an insight while planning mass vaccination strategies for bovine population in the state of Kerala; by Department of Animal Husbandry because a cock tail of tick transmitted pathogens in a single animal will interfere with the future sero conversion and duration of immunity.

 

Author(s) Details

 Sunitha Karunakaran
Department of Animal Husbandry, Clinical Laboratory, Animal Disease Control Project Paravattani, Thrissur, East P.O-680005, Kerala, India.

Hareesh Sasidharan
Department of Animal Husbandry, Clinical Laboratory, Animal Disease Control Project Paravattani, Thrissur, East P.O-680005, Kerala, India.

 

Gopika Thattat Gopinathan
Department of Animal Husbandry, Clinical Laboratory, Animal Disease Control Project Paravattani, Thrissur, East P.O-680005, Kerala, India.

 

Please see the book here:- https://doi.org/10.9734/bpi/mbrao/v4/5690