Table 1: Primers, their sequences and their application in this study.

Abstract

Motile Aeromonas septicaemia (MAS), caused by Aeromonas hydrophila, is one of the most significant bacterial disease responsible for mortality in Indian catfish, Clarias magur, a potential aquaculture species in the Indian subcontinent. In fish, innate immunity elicited by pathogen recognition receptors (PRRs) plays an important role in providing protection against bacterial infection. Information on PRRs including Toll-like receptors (tlrs) and their response to bacterial pathogens remains unexplored in magur. Toll-like receptor 2 (tlr2), a phylogenetically conserved germ-line encoded PRR recognizes specific microbial structure and trigger MyD88-dependent signaling pathway to induce release of various cytokines responsible for innate immune response. In the present study, tlr2 gene of magur was characterized and downstream signaling was studied following challenge with A. hydrophila. The full-length cDNA of magur tlr2 (mtlr2) comprised of 3,066 bp with a single open reading frame of 2,373 bp encoding 790 amino acids having a theoretical pI value of 6.11 and molecular weight of 90 kDa. Structurally, it comprised of signal peptide (1–42aa), one leucine-rich repeat region (LRR) at N-terminal (LRR1-NT: 50–73 aa) and C-terminal (LRR-CT: 588–608 aa), twenty LRRs in between, one trans-membrane (Tm) domain (609–631aa) followed by cytoplasmic TIR domain (670–783aa). Phylogenetically, mtlr2 is closely related to pangasius and channel catfish. Highest basal expression of mtlr2, myd88 and il-1β in spleen, nf-kb in anterior kidney was observed. Lowest basal expression of mtlr2 in skin and myd88, nf-kb and il-1β in muscle was detected. Significant up-regulation of mtlr2 and downstream expression occurred at 3, 8, 24 h post infection to A. hydrophila in important immune organs such as liver, spleen, intestine and kidney. These findings highlight the vital role of tlr2 in eliciting innate immune defence against A. hydrophila infection.