Screening of carbon sources for enrichment and directional isolation ofRhodobacteraceae from the gut of Litopenaeus vannamei

Abstract:

[Objective]Rhodobacteraceae, the dominant group of gut microbiota in Litopenaeus vannamei, usually has higher relative abundance in the gut of healthy shrimps, and some members of this family have been identified as the indicators for shrimp health. Therefore, clarifying the method for the directional enrichment and isolation of Rhodobacteraceae from shrimp gut can provide a basis for the development of probiotics for shrimp farming. [Methods]The 16S rRNA gene high-throughput sequencing was applied in the screening of the suitable carbon sources for the enrichment of Rhodobacteraceae. Then, the bacteria were directionally isolated from the enriched samples via the pure culture method. Finally, the taxonomic status and genetic diversity of the isolates were determined.[Results] The addition of short-chain fatty acids (acetic acid, propionic acid, butyric acid, and valeric acid) and sodium bicarbonate facilitated the enrichment of Rhodobacteraceae, which mainly included Cribrihabitans, Tritonibacter, Rhodovulum, Ruegeria, Sagittula, andThalassobius. A total of 303 bacterial strains belonging to 12 families of 2 phyla were isolated from the samples with high relative abundance of Rhodobacteraceae, and 119 out of the 303 strains were identified as members of Rhodobacteraceae, including 90 strains of Tritonibacter, 25 strains ofPhaeobacter, 1 strain of Sulfitobacter, 1 strain of Ruegeria, 1 strain ofRoseovarius, and 1 strain ofAliiroseovarius. In addition, all the isolates of Rhodobacteraceae had the relative abundance similar to the results of high-throughput sequencing. [Conclusion] We explored the effect of different carbon sources on the enrichment and directional isolation of Rhodobacteraceae by using high-throughput sequencing and screened out five carbon sources for isolating 119 strains of Rhodobacteraceae.

Key words:Litopenaeus vannamei;Rhodobacteraceae;carbon source;16S rRNA gene high-throughput sequencing;directional isolation