Immunogenic Proteins Characterization in the Larvae of Oestrus ovis by Western Blot Analysis

Abstract

The present study was undertaken to characterize and identify immunogenic proteins of Oestrus ovis in Telangana. Oestrus ovis is the most predominant nasal bot fly causing nasal myiasis in India. The collected larvae from Jiyaguda slaughter house, Hyderabad are seperated into L2 and L3 larvae were washed with normal saline and PBS pH 7.4 and preserved at -70°C for antigen preparation and some were preserved in 70 % ethanol for identification. The larvae were speciated by light microscopic examination based on morphological features such as ventral spines, anterior hooks and “D” shaped posterior spiracles. In this study, an antigen was prepared from second and third instar larvae of Oestrus ovis using a soluble buffer solution. The resulting Crude Somatic Larval (CSL) antigen had a protein concentration of 27.4 mg per ml. The CSL antigen was resolved in 8% SDS-PAGE, Immunoblotting was performed by electro transferring the separated polypeptides onto a nitrocellulose (NC) membrane. Treatment with specific reagents and antibodies resulted in the identification of 19 immunoreactive polypeptides, with molecular weights matching those observed in SDS-PAGE. Four polypeptides with molecular weights of 58.56, 20.74, 19.09, and 28.91 kDa exhibited the strongest reaction towards the antibodies raised against CSL antigen. These four polypeptides are considered promising candidates for future immunizations and testing their protective efficacy through in vivo assays. These findings have implications for advancing strategies to combat myiasis, a parasitic disease caused by Oestrus ovis larvae.