基于高通量测序的花马盐湖原核微生物及其耐盐基因

Microbial community and salt-tolerant genes of hypersaline sediment from Huama lake revealed by metagenomic sequencing

【目的】揭示陕北花马盐湖沉积物原核微生物群落组成,并分析其潜在的耐盐功能基因。【方法】构建盐湖沉积物宏基因组16S r RNA文库和fosmid文库,利用Illumina HiSeq高通量测序及生物信息技术分析细菌古菌群落组成和耐盐菌株(5-5)外源宏基因组的潜在耐盐基因。【结果】获得18978条有效的16Sr RNA序列,共5221个OTUs,包括23个门,155个属,其中广古菌门(Euryarchaeota)和变形菌门(Proteobacteria)为优势菌门,盐杆状菌属(Halorhabdus)、盐红菌属(Halorubrum)及假单胞菌属(Pseudomonas)等16个属为优势属,以及嗜盐单胞菌属(Halomonas)、冷弯菌属(Psychroflexus)及不动细菌属(Acinetobacter)等139个属为非优势属。从4126个fosmid文库菌株中筛选出37株耐盐菌株,其中菌株5-5、2E4和2F4对不同浓度的NaCl、CuSO_4、ZnSO_4及CdSO_4具有耐受性,从5-5的外源宏基因组序列中获得61个Unigene,其中12个Unigene的同源基因编码的蛋白质如无机焦磷酸酶、转座酶、亚碲酸钾抗性蛋白及钙调蛋白等广泛参与其他生物的耐盐逆境。【结论】盐湖沉积物中蕴藏着丰富多样的细菌古菌类群以及潜在耐盐功能基因资源。

[Objective] The aim of this study was to investigate prokaryotic microbial community structures and salt-tolerant genes in hypersaline sediments from Huama salt lake, Northern Shaanxi, China. [Methods] We constructed 16S rRNA library and fosmid library using metagenome of the sediment, and detected prokaryotic microbial distribution and salt-tolerant genes by high-throughput pyrosequencing and bioinformatic analysis. [Results] In total 18978 high-quality reads for 16S rRNA were assigned to 23 phyla, 155 genera and 5221 OTUs. Euryarchaeota and Proteobacteria were the dominant phyla. Sixteen dominant genera such as Halorhabdus, Halorubrum and Pseudomonas and 139 infrequent ones like Halomonas, Psychroflexus and Acinetobacter were explored. A total of 37 salt-tolerant strains were detected from the metagenomic DNA fosmid library containing 4126 strains. The salt-tolerant strains like 5-5, 2E4 and 2F4 exhibited strong tolerance against stresses caused by NaCl, CuSO_4, ZnSO_4 and CdSO_4. High-throughput pyrosequencing of the metagenomic fragment in the strain 5-5 explored 61 unigenes, 23 homogenous genes encoding salt-, or other stress-tolerant proteins in other biological cells, such as inorganic pyrophosphatase, transposase, tellurite resistance protein and calmodulin. [Conclusion] Hypersaline sediments in salt lakes harbor diverse bacterial and arhaeal community and rich salt-tolerant genes.