| Cr(?)pollution seriously threatens environmental safety.Microbial remediation of Cr(?)pollution has become a hotspot in current researches.Researchers have extensively studied the physiological and biochemical mechanisms underlying microbial tolerance and reduction of Cr(?),but the molecular mechanisms are still not fully clear.The development of high-throughput sequencing technology helps to further explore the molecular mechanisms of microbial tolerance to heavy metals.Therefore,this study assessed L.fursiformis 15-4tolerance to heavy metals and its molecular mechanisms.The results showed strong tolerance to various heavy metals,while transcriptome sequencing and q PCR analysis on the Cr(?)-treated L.fursiformis 15-4 strains were used to explore the molecular mechanisms.The main results are as follows:1.The minimal inhibitory concentrations(MICs)of L.fursiformis 15-4 on Pb(?),Cr(?),Cu(?),Co(?),Ni(?),Cd(?),Ag(?),and Hg(?)were 4.60?4.0?2.60?1.36?1.36?0.26?0.28and 0.03 m M at 20?and pH7.0,respectively,which suggests significant tolerance to a variety of heavy metals.The results from the experiments using shaking flask culture showed that L.fursiformis 15-4 could completely reduced 1 m M and 2 m M Cr(?)added in the medium for 36 h and 84 h,respectively,indicating that strain 15-4 was robust to reduce Cr(?)to Cr(?).2.Transmission electron microscopy observation showed that,when cultured in LB liquid medium for 24 h,the cell of L.fursiformis 15-4 remained a rod-shaped and regular and integral structure.Whereas,when incubated in the LB liquid medium containing 1 m M and 2m M Cr(?),the cell displaied a thickened wall,no capsule membrane,and some unknown materials on the surface.3.The strains cultured in LB liquid medium without and with 2 m M Cr(?)for 24 h were performed transcriptome sequencing.The raw sequences were assembled after quality control and were annotated to the reference genome.As result,a total of 4816 genes and 201s RNAs were annotated,of which 3907 genes were expressed in two samples.Using the edger3.24.3 software,1098 differentially expressed genes(DEGs),accounting for 28.1%of the total genes,were identified by comparing the two sample groups,of which 605(55.10%)genes were up-regulated and 493(44.90%)genes were down-regulated.The results of DEGs function annotation showed that the number of genes annotated to GO,COG,KEGG,Nr and Swiss-Prot databases were 883(80.42%),1079(98.27%),630(57.38%),1093(99.55%),900(81.97%),respectively.The GO enrichment analysis showed that the up-regulated DEGs were significantly enriched to cellular process,metabolic process,cellular metabolic process,and organic substance metabolic process,etc.The down-regulated DEGs were mainly enriched to membrane part,intrinsic component of membrane,and integral component of membrane,etc.The KEGG enrichment analysis showed that the up-regulated DEGs were significantly enriched to ABC transporter,ribosome,porphyrin metabolism,aminoacyl t RNA biosynthesis,sulfur metabolism,and oxidative phosphorylation.The down-regulated DEGs were mainly enriched to quorum sensing,histidine metabolism,and atrazine degradation,suggesting that these pathways were significantly affected by Cr(?).4.Further analyses implied the involvement of several genes in Cr(?)reduction of L.fusiformis 15-4,such as genes encoding dihydrolipoate dehydrogenase(lpd),nitroreductase quinone oxidoreductase(qor),FMN reductase(ssu E),isocitrate dehydrogenase encoding gene(icd),LLM flavin-dependent oxidoreductase encoding gene,and ferredoxin/flavinredoxin-NADP~+reductase(fnr).Moreover,L.fusiformis 15-4 employed the metabolic pathways such as ABC transporter,porphyrin metabolism,oxidative phosphorylation,aminoacyl t RNA biosynthesis,ribosome,sulfur metabolism,and TCA cycle to resist Cr(?). |
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