Abstract

MOLECULAR CHARACTERISATION OF CLOSTRIDIUM PERFRINGENS ISOLATED FROM SHEEP USING MICRO-PCR, ELISA AND MALDI-TOF

Sobha Rani.M, BeenaRavi, Suhas.V. Bhandari, Venkatesh. M. D. and Asha Mayanna

ABSTRACT

Enterotoxaemia is a fatal and severe disease of sheep and goats caused by bacteria called Clostridium perfringens – types C and D. They release toxins into the blood stream and over multiply in numbers, which leads to sudden death of the cattle. Till date the diagnosis of this disease is purely based on traditional methods of identification of the bacteria. Using modern technologies in disease diagnosis not only helps in saving time, but also the lives of animals. In the present study both the traditional methods and modern methods like MALDI and ELISA were compared for detection of causative bacteria from the clinical samples. Using the conventional gram staining method and biochemical tests, the isolated bacteria have been identified as Clostridium perfringens. MALDI-TOF analysis also confirmed the presence of Clostridium perfringens. Using ELISA, the sub-types ofthe organism based on the toxin released by Clostridium perfringens type D have been identified. 16S rRNA gene amplification and sequencing yielded a partial sequence of the 16S rRNA gene of the organism. Based on the analysis of this sequence, the organism was identified as Clostridium perfringens strain 13 (GenBank Accession Number: NR 074482.1). Modern methods enabled the identification of isolated bacteria accurately at subtype level in a short time and helped in educing turnaround time required for treatment and other control measures like vaccination of remaining animals in the herd.

Keywords: Enterotoxaemia, Clostridium perfringens, MALDI, ELISA.