Fish health is greatly influenced by the prevailing bacterial communities and the dramatic change in their structure in the aquatic environment. The prevalent environmental bacterial pathogens in aquaculture production in two southern states of Nigeria were characteriz ed and evaluated for antibiotic sensitivity, antimicrobial resistance, and presence of virulence genes.
Liver, kidney, and spleen collected aseptically from apparently health y African catfish (Clarias gariepinus ) farmed in Delta and Ogun states, were subjected to ba cterial culture, isolation, identification , and DNA extraction. Whole-genome sequencing of extracted DNAs was carried out at WorldFish, Malaysia while biochemical and antibiotic susceptibility tests were conducted at Mississippi State University, USA . Ribosomal RNA-containing contigs were identified and their corresponding 16 S rRNA gene was extracted and used to BLAST against the NCBI microbial 16S database. Mass screening of contigs for antimicrobial resistance and virulence genes was performed.
The BlastN revealed Providencia vermicola (99.802%), Providencia rettgeri (99.666%), Proteus mirabilis (99.799%), Proteus vulgaris (99.527% ), Citrobacter freundi (99.667%), Morganella morganii (99.728%), Alcaligenes faecalis (99.316% ), and Pseudomonas qingdaonensis (100%) as the prevalent environmental organisms . Analyses of the isolates showed that only Alcaligenes faecalis had no antimicrobial-resistance (AMR) genes. Providencia spp. (P. vermicola and P. rettgeri) genome had seven AMR genes; tet(57) and tet(59) (tetracycline) , aadA1 and ant(3’’)-Ia (Streptomycin ), sat2_gen (Streptothricin) , catB2 (Chloramphenicol) , and dfrA1 (Trimethoprim) . Proteus (P. mirabilis and P. vulgaris ) possessed fourteen AMR genes: hugA , blaTEM -1, blaTEM-1B (Beta-lactam) , catA1, catA4, and cat (Chloramphenicol) , dfrA7 (Trimethoprim) , sul1, sul2, (Sulfonamide) , aph(3’’)-Ib; aph(6)-Id (Streptomycin) , tet(A), tet(J) (Tetracycline) , and blaTEM -1B (Amoxicillin, Ampicillin, Cephalothin, Piperacillin, Ticarcillin) . Citrobacter freundi possessed four AMR genes; qnrB12 (Quinolone and Ciprofloxacin), blaCMY-159 , blaMOR-2 , and blaCMY-98_1 (Cephalosporin). Morganella morganii had two AMR genes; catA2 (Chloramphenicol) and blaDHA-17 (Cephalosporin, Amoxicillin; Amoxicillin+Clavulanic acid; Ampicillin; Ampicillin+Clavulanic acid; Cefotaxime; Cefoxitin; Ceftazidime; Piperacillin; Piperacillin+Tazobactam; Ticarcillin; Ticarcillin+Clavulanic acid ). Pseudomonas qingdaonensis possessed seven AMR genes; cmlB1 (Chloramphenicol), dfrA1 (Trimethoprim), sat2_gen (Streptothricin ), sul3 (Sulfonamide), aac(3)-IIe (Gentamicin) , aac(3)-IIa (Tobramycin ), aadA1 and ant(3’’)-Ia_1 (Streptomycin).
Out of the eight isolates, only Citrobacter freundi and Pseudomonas qingdaonensis had virulence factors; 11 (csgB, csgD, csgE, csgF, csgG, ompA, entA, entB, entE, fepG , and fepC) and 2 (pilH and algR), respectively.
Opportunistic pathogens are becoming an emerging threat to aquaculture systems because di seases caused by them have been generally overlooked. The p ossession of AMR genes and virulence factors by opportunistic environmental pathogens in farmed fish suggests pollution of aquatic environments with adverse effects on the environment, animals, and public health.
Keywords: Environmental pathogens , Whole-genome sequencing, 16S rRNA, Antimicrobial Resistance Genes , Virulence factors , Clarias gariepinus, Public health