Materials and Methods:
A cross-sectional study was conducted in the Jabalpur region (23.1815°
N, 79.9864° E), Madhya Pradesh between March, 2018- June, 2019 to
investigate the prevalence of CCHF and theileriosis in livestock’s and
its associated risk factors (Fig.1). To cover urban and rural/tribal
areas, five villages were selected purposely from peripheral villages of
study region.. Simple random sampling method was applied to select the
animals from the herd. The sample size was estimated based on the
assumption of 10% sero-positivity rate, the level of confidence as 95
percent and a absolute error of 5 %. A minimum desired sample size was
140, so the desired sample bovine and goat/sheep was 140 each. Overall,
a total of 155 bovine and 238 goats’ blood samples were collected from
the jugular veins following biosafety measures. The pre-designed
structured interview schedule was used to collect information on animal
husbandry activities and risk factors. Sera were separated and subjected
to two commercial ELISA kits (1 for bovine -Catalogue No.CCHFCTIGG-48,
and 1 for goats -Catalogue No. CCHFSGIGG-48) (Zydus Diagnostics,
Ahmedabad, India). The manufacturer’s instructions were followed for
testing samples and based on the ratio >1.5, the positivity
was determined for both the kits. The DNA was isolated from the CCHF IgG
negative bovine blood samples using DNA Sure Mini Kit (Nucleo-pore,
GENETIX, India) as per the manufacturer’s protocol. The isolated DNA
samples were subjected to PCR amplification targeting the cytochrome b
(312 bp segment) gene of the Theileria annulata (Bilgic et al.,
2010). The PCR products were sequenced analyzed and submitted to
GenBank. The necessary ethical permission for the conduct of this study
was obtained from the Institutional Animal Ethics Committee (IAEC) and
Institutional Ethics Committee (NIRTH/IEC/3756/2018).