Abstract: This paper reports the investigation of necrotic enteritis (NE) in six broiler chickens of age two to three weeks old, died in Jowai poultry farm, Meghalaya, India. Initially, scanning electron microscopy (SEM) was performed to observe the morphological changes within the intestine. Intestinal contents and liver samples from dead chicken were investigated for isolation of bacteria and their virulence determinant. The SEM analysis of infected intestine revealed massive necrosis and complete destruction of the intestinal villi within the intestinal mucosa. Bacterial isolation confirmed the causative agent as C. perfringens in NE. All the isolates harboured single and double plasmid deoxyribonucleic acid with identical 45.2kb common plasmid. In polymerase chain reaction (PCR) assay all 10 clinical isolates harboured alpha toxin gene (cpa) of C. perfringens, however, four isolates also carried additional beta2 toxin gene (cpb2). None of the isolates were positive for beta, epsilon, iota and enterotoxin genes. PCR analysis revealed that all isolates derived from NE belonged to C. perfringens type A. The partial cpa gene sequence analysis showed 97.6 to 100% homology among the C. perfringens isolates. The study confirmed that C. perfringens type A is the most predominant one associated with necrotic enteritis in broiler chickens in Meghalaya, India and the alpha toxin (CPA) might play a significant role in the pathogenesis of the disease in broiler chicken.