Diversity and functional enzymes of halophilic bacteria in the Da Qaidam Salt Lake

Abstract:

Objective To explore the diversity of halophilic bacteria in the magnesium sulfate-subtype Da Qaidam Salt Lake, compare the effects of different culture conditions on the diversity of halophilic bacteria, and screen the extracellular functional enzymes of halophilic bacteria.Methods Illumina MiSeq was used to analyze the diversity of bacteria in the Da Qaidam Salt Lake. Thirteen media, 2 salinities, 8 enrichment culture periods, and 6 dilution gradients were selected to isolate halophilic bacteria, and the taxonomic status of the strains was determined by 16S rRNA gene sequencing and BLAST sequence alignment. According to the sequencing results, 45 representative strains of different species belonging to 18 genera were selected and 7 media were used to screen the strains with activities of functional enzymes including protease, cellulase, amylase, and esterase.Results A total of 244 bacterial OTUs were obtained by culture-free high-throughput sequencing in the Da Qaidam Salt Lake. The strains with clear taxonomic status were annotated to 153 genera, 133 families, 92 orders, 53 classes of 19 phyla, with Pseudomonadota and Actinomycetota being the dominant phyla. A total of 593 strains of halophilic bacteria were isolated from the mixed water and mud samples of the Da Qaidam Salt Lake, belonging to 22 genera, 12 families, 8 orders, 5 classes of 4 phyla, of which 11 strains may belong to potential new species. Pseudomonadota and Bacillota were the dominant phyla, and Halomonas, Virgibacillus, and Bacillus were the dominant genera. The number of halophilic bacteria isolated under the culture with 10% NaCl was significantly higher than that with 18% NaCl, indicating that moderately halophilic bacteria were dominant among culturable halophilic bacteria. The media with better isolation performance were 2216E, 1/2 R2A, 1/10 2216E, and 1/10 TSA, all of which are oligotrophic media. The optimal enrichment culture periods ranged from 7 to 30 days. The undiluted samples obtained the best isolation results, followed by dilution gradients of 10^-1 and 10^-2. Among the 45 representative strains, 40.0%, 31.1%, 40.0%, and 82.22% of the strains had activities of protease, cellulase, amylase, and esterase, respectively.Conclusion Optimization of the isolation and culture conditions can significantly improve the diversity of halophilic bacteria that can be cultured in salt lakes. The high diversity and high halophilic enzyme activities of the culturable halophilic bacteria from the Da Qaidam Salt Lake provide a basis for further application of these halophilic bacteria.