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Aeromonas hydrophila infection induces Toll-like receptor 2 (tlr2) and associated downstream signaling in Indian catfish, Clarias magur (Hamilton, 1822).

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.

SUBMITTER: Muduli C

PROVIDER: S-EPMC8641487 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Antimicrobial peptides (AMPs) constitute a broad range of bioactive compounds in diverse organisms, including fish. They are effector molecules for the innate immune response, against pathogens, tissue damage and infections. Still, AMPs from African Catfish, Clarias gariepinus, skin mucus are largely unexplored despite their possible therapeutic role in combating antimicrobial resistance. In this study, African Catfish Antimicrobial peptides (ACAPs) were identified from the skin mucus of African Catfish, C. gariepinus. Native peptides were extracted from fish mucus scrapings in 10% acetic acid (v/v) and ultra-filtered using 5 kDa molecular weight cut-off membrane. The extract was purified using C18 Solid-Phase Extraction. The antibacterial activity was determined using the Agar Well Diffusion method and broth-dilution method utilizing Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922). Thereafter, Sephadex G-25 gel filtration was further utilized in bio-guided isolation of the most active fractions prior to peptide identification using Orbitrap Fusion Lumos Tribrid Mass Spectrometry. The skin mucus extracted from African Catfish from all the three major lakes of Uganda exhibited antimicrobial activity on E. coli and S. aureus. Lake Albert's C. gariepinus demonstrated the best activity with the lowest MIC of 2.84 and 0.71 μg/ml on S. aureus and E. coli, respectively. Sephadex G-25 peak I mass spectrometry analysis (Data are available via ProteomeXchange with identifier PXD029193) alongside in silico analysis revealed seven short peptides (11-16 amino acid residues) of high antimicrobial scores (0.561-0.905 units). In addition, these peptides had a low molecular weight (1005.57-1622.05 Da) and had percentage hydrophobicity above 54%. Up to four of these AMPs demonstrated α-helix structure conformation, rendering them amphipathic. The findings of this study indicate that novel AMPs can be sourced from the skin mucus of C. gariepinus. Such AMPs are potential alternatives to the traditional antibiotics and can be of great application to food and pharmaceutical industries; however, further studies are still needed to establish their drug-likeness and safety profiles.

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Aeromonas hydrophila infection induces Toll-like receptor 2 (tlr2) and associated downstream signaling in Indian catfish, Clarias magur (Hamilton, 1822).

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