Taxonomic And Functional Proifling Of Gastrointestinal Tract Microbiome Of The Farmed Turbot(Scophtalmus Maximus)

Culture-independent methods and culture-dependent technologies were appliedto unveil the taxonomic composition and functional diversity of the farmed turbot(Scophthalmus maximus) gastrointestinal (GI) microbiome. First, metagenomicscombined with16S rRNA sequence analysis was used to analyze the taxonomicdistribution and a total of33,998contigs and95operational taxonomic units (OTUs)were identified, respectively. Proteobacteria and Firmicutes which existed in both GIcontent and mucus were dominated in the turbot GI microbiome. At the genus level,Vibrio was observed in all parts of turbot GI tract, especially in rectum content.Within Vibrio, the four main species, V. vulnificus, V. cholerae, V. parahaemolyticusand V. harveyi were recognized as potential pathogens which were responsible formany fish diseases. As compared taxonomic distribution between hosts,Gammaproteobacteria and Mollicutes were overabundant in turbot GI microbiomes.Among them, members of Vibrio and Mycoplasma were dominant.16S rRNAsequence analysis also indicated that the bacterial community diversity decreasedalong the turbot GI tract (from stomach to rectum) and the turbot GI tract may harborsome bacteria which originate from associated seawater.Second, to enhance the cultivation efficiency, eighteen agar media were tested forcultivating fish-gut-associated bacteria in farmed adult turbot, including agar mediawith or without1%gastrointestinal (GI) supernatant, and2%and4%GI supernatantagar media. A total of1,711colonies were analyzed, and24operational taxonomicunits (OTUs) were identified. Most colonies and OTUs were obtained from Zobell2216E agar media, while a low diversity was identified from MRS/MRS+agar media.Agar media with GI supernatant (1%,2%, and4%) yielded different profiles of OTUsfrom those of the corresponding original media and provided some substances thatenhanced the cultivation efficiency of bacteria in turbot GI tract.Gammaproteobacteria represented the large majority of the colonies (82%). Firmicutes and Actinobacteria represented15.6%and2.4%colonies, respectively. Atthe genus level,49.4%of all colonies belonged to Vibrio. Other potential pathogensincluding Pseudomonas, Photobacterium, and Enterobacter and potential probioticsincluding Bacillus, Paenibacillus, Pseudomonas and Shewanella were also obtainedfrom agar media. Additionally, most of OTUs identified in this study had highhomology with the species that were first described in turbot GI tract. The impact ofthese species on the turbot physiology and health should be further investigated.Last, functional analyses indicated that the clustering-based subsystem and manymetabolic subsystems were dominated in the turbot GI metagenome, accounting for15.9%of turbot GI metagenome. Compared to other gut metagenomes, quorumsensing and biofilm formation was overabundant in the turbot GI metagenomes.Genes associated with quorum sensing and biofilm formation was found in specieswithin Vibrio, including V. vulnificus, V. cholerae and V. parahaemolyticus. In farmedfish gut metagenomes, the stress response and protein folding subsystems wereover-represented and several genes regarding antibiotic and heavy metal resistancewere also detected. These data suggested that the turbot GI microbiome may beaffected by human factors in aquaculture. Additionally, iron acquisition andmetabolism subsystem was more abundant in the turbot GI metagenome thanfreshwater fish (hybrid striped bass) gut metagenome suggesting that uniquemetabolic potential may be observed in marine animals.