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Members of the genus Metallosphaera are widely found in sulfur-rich and metal-laden environments, but their physiological and ecological roles remain poorly understood. Here, we sequenced Metallosphaera tengchongensis Ric-A, a strain isolated from the Tengchong hot spring in Yunnan Province, China, and performed a comparative genome analysis with other Metallosphaera genomes. The genome of M. tengchongensis had an average nucleotide identity (ANI) of approximately 70% to that of Metallosphaera cuprina. Genes sqr, tth, sir, tqo, hdr, tst, soe, and sdo associated with sulfur oxidation, and gene clusters fox and cbs involved in iron oxidation existed in all Metallosphaera genomes. However, the adenosine-5-phosphosulfate (APS) pathway was only detected in Metallosphaera sedula and Metallosphaera yellowstonensis, and several subunits of fox cluster were lost in M. cuprina. The complete 3hydroxypropionate/4-hydroxybutyrate cycle and dicarboxylate/4-hydroxybutyrate cycle involved in carbon fixation were found in all Metallosphaera genomes. A large number of gene family gain events occurred in M. yellowstonensis and M. sedula, whereas gene family loss events occurred frequently in M. cuprina. Pervasive strong purifying selection was found acting on the gene families of Metallosphaera, of which transcriptionrelated genes underwent the strongest purifying selection. In contrast, genes related to prophages, transposons, and defense mechanisms were under weaker purifying pressure. Taken together, this study expands knowledge of the genomic traits of Metallosphaera species and sheds light on their evolution.

Background: Antimonite [Sb(III)]-oxidizing bacterium has great potential in the environmental bioremediation of Sb-polluted sites. Bacillus sp. S3 that was previously isolated from antimony-contaminated soil displayed high Sb(III) resistance and Sb(III) oxidation efficiency. However, the genomic information and evolutionary feature of Bacillus sp. S3 are very scarce. Results: Here, we identified a 5,579,638 bp chromosome with 40.30% GC content and a 241,339 bp plasmid with 36.74% GC content in the complete genome of Bacillus sp. S3. Genomic annotation showed that Bacillus sp. S3 contained a key aioB gene potentially encoding As(III)/Sb(III) oxidase, which was not shared with other Bacillus strains. Further, a series of genes associated with Sb(III) and other heavy metal(loid)s were also ascertained in Bacillus sp. S3, reflecting its adaptive advantage for growth in the harsh eco-environment. Based on the analysis of phylogenetic relationship and the average nucleotide identities (ANI), we found that Bacillus sp. S3 was a novel species within the Bacillus genus. The majority of mobile genetic elements (MGEs) mainly distributed on chromosomes within the Bacillus genus. Pan-genome analysis showed that the 45 genomes contained 554 core genes and many unique genes were dissected in analyzed genomes. Whole genomic alignment showed that Bacillus genus underwent frequently large-scale evolutionary events. In addition, the origin and evolution analysis of Sb(III)-resistance genes revealed that evolutionary relationships and horizontal gene transfer (HGT) events among the Bacillus genus. The assessment of functionality of heavy metal(loid)s resistance genes emphasized its indispensable roles in the harsh eco-environment of Bacillus genus. The real-time Quantitative PCR (RT-qPCR) results of Sb(III)-related genes indicated that the Sb(III) resistance was constantly increased under the Sb(III) stress. Conclusions: The results in this study shed light on the molecular mechanisms of Bacillus sp. S3 coping with Sb(III), extended our understanding on the evolutionary relationship between Bacillus sp. S3 and other closely related species, and further enriched the Sb(III) resistance genetic data sources.

Background: Antimonite [Sb(III)]-oxidizing bacterium have great potential in the environmental bioremediation of Sb-polluted sites. Bacillus sp. S3 isolated from antimony-contaminated soil showed high Sb(III) resistance and Sb(III) oxidation efficiency. However, very little genomic information and evolutionary relationships that bacterial oxidation of Sb(III) is available. Results: Here, we identified a 5579638 bp chromosome with 40.30% GC content and a 241339 bp plasmid with 36.74% GC content in the complete genome of Bacillus sp. S3. Genomic annotation showed that Bacillus sp. S3 contains key aioB gene potentially encoding As(III)/Sb(III) oxidase, is not shared with other Bacillus strains. Further, a series of genes associated with Sb(III) and other heavy metal(loid) were also ascertained, reflecting adaptive advantage for growth in the harsh eco-environment. It is noteworthy that Bacillus sp. S3 is a novel species within the Bacillus genus as indicated by phylogenetic relationship and the average nucleotide identities (ANI) analysis. The presence of genomic plasticity demonstrated a high number of mobile genetic elements (MGEs) that were mainly distributed on chromosomes within the Bacillus genus. The core genome contained 554 core genes and many unique genes were dissected in analyzed genomes, indicating a conserved core but adaptive pan repertoire. Whole genomic alignment indicates that frequently genomic reshuffling and rearrangements, genetic gain and loss, and other recombination events occurred during the evolutionary history of Bacillus genus. In addition, the origin and evolution analysis of Sb(III)-resistance genes revealed that evolutionary relationships and events of horizontal gene transfer (HGT) among the Bacillus genus. The assessment of functionality of heavy metal(loid) resistance genes emphasized its indispensable roles in the harsh eco-environment of Bacillus genus. The real-time Quantitative PCR(RT-qPCR) results of Sb(III)-related genes indicated that the Sb(III) resistance was constantly increased under the Sb(III) stress. Conclusions: The insights provided in this study shed light on the molecular details of Bacillus sp. S3 coping with Sb(III), which extended our understanding on the evolutionary relationship between Bacillus sp. S3 and other closely related species and will enrich the Sb(III) resistance genetic data sources.