Study On Polyphasic Taxonomic Identification And Antimony Resistance Mechanism Of Novel Species Flavobacterium Potami

Excessive mining,metallurgy,combustion of fossil fuels,and weathering of antimony worldwide have caused serious antimony pollution.Antimony pollution in China is particularly serious because China’s antimony reserves and production are the largest in the world‘There is no substitute for ecology,and it is hard to use it without realizing that it is lost’.While controlling and eliminating the sources of pollution,discovering and researching resources,methods and technologies with high efficiency,green,inexpensive and lasting environmental pollution remediation is especially significant for our country,where protecting the environment and saving resources are the basic national policies.Microbial remediation of environmental pollution has the advantages of environmental friendliness,economic savings,and long-lasting circulation.It has excellent potential for application in situ remediation of antimony(Ⅲ)pollution,which has become a research hotspot in environmental remediation.A variety of microorganisms have been found in the environment capable of oxidizing the highly toxic valence state of antimony(Ⅲ)to the less toxic valence state of antimony(V),but the germplasm resources of such microorganisms are poorly reported,and the related studies on microbial antimony(Ⅲ)resistance and transformation mechanisms are seriously inadequate.Therefore,a potentially novel species of Flavobacterium sp.17A~T with antimony(Ⅲ)resistance was isolated from the sediment at the bottom of Fuyang River polluted by various heavy metals and antibiotics,and a polyphasic taxonomy identification was conducted to determine its taxonomic status,and its potential antimony(Ⅲ)resistance genes and related resistance mechanisms were studied by whole-genome sequencing and antimony(Ⅲ)-induced transcriptome sequencing.The main findings are as follows:(1)Polyphasic taxonomy of antimony(Ⅲ)-resistant Flavobacterium potami:Among the 80 bacterial strains isolated in the sample,a strain 16S r RNA gene had the highest similarity with the strain‘Flavobacterium panacis’DCY106~T(97.5%)of the genus Flavobacterium in the type strain database,which was lower than the standard threshold for determining novel species similarity(98.7%).Subsequently,a polyphasic taxonomy study including morphological taxonomy,molecular taxonomy,physiological and biochemical taxonomy,chemical taxonomy and genomic analysis was conducted.Compared with the reference strains,strain 17A~T had both typical characteristics and significant differences of the genus Flavobacterium,and was identified as a new species of Flavobacterium.Finally,a new species name was proposed internationally and named as Flavobacterium potami 17A~T.(2)Genomic study of antimony(Ⅲ)-resistant F.potami:F.potami 17A~T showed multi-heavy metal resistance and was resistant to antibiotics.The genome of F.potami17A~T was sequenced and annotated,and its mobile elements,drug resistance genes,hydrocarbon-active enzymes,secondary metabolites and heavy metal(antimony)resistance genes were predicted.Eleven genes were predicted to be potentially involved in antimony/arsenic translocation or transformation,including three metalloregulatory protein Ars R/Smt B family transcription factors(ars R),three Ars R family transcriptional regulators,two arsenate reductases(ars C),one ACR3 family arsenite efflux protein(ars B),and two iron-sulfur cluster assembly-related proteins(isc S and isc A).(3)Antimony(Ⅲ)resistance transcriptomic study of Flavobacterium potami:The antimony(Ⅲ)inducible transcriptome study of Flavobacterium potami 17AT was carried out based on genome sequencing.Five of the antimony(Ⅲ)resistance genes predicted from the genome were present in the significantly differentially expressed genes sequenced from the antimony(Ⅲ)-inducible transcriptome,encoding Ars R/Smt B family transcription factors(ars R),arsenate reductases(ars C),ACR3 family arsenite efflux proteins(ars B)and iron-sulfur cluster assembly-related proteins(isc S and isc A).Among the higher significantly differentially expressed individual genes,there are also several genes that may be the focus of research on new mechanisms of antimony(Ⅲ)resistance,including proteins of unknown function.In addition,GO and KEGG enrichment analyses revealed that genes related to antioxidant activity and intramolecular oxidoreductase activity in the strain may also have a role in the antimony(Ⅲ)resistance,the exact function of which needs to be further verified.In this study,the taxonomic status of the novel antimony(Ⅲ)-resistant Flavobacterium potami 17A~T was determined by polyphasic taxonomy techniques,which enriched the species diversity of the genus Flavobacterium and antimony(Ⅲ)-resistant strains.The study of antimony(Ⅲ)resistance mechanism of type strain 17A~T is representative,and also provides germplasm resources for antimony(Ⅲ)pollution remediation;The functional genes and possible molecular mechanisms related to antimony(Ⅲ)resistance of Flavobacterium potami 17A~T under antimony-induced conditions were analyzed by whole genome and transcriptome studies,which laid the theoretical foundation and data support for the subsequent enrichment of antimony(Ⅲ)resistance/oxidation regulation mechanism of microorganisms.