| Sphingomonads(including Sphingomonas,Sphingobium,etc.)can often be isolated from the environments contaminated by organic pollutants and generally have the capability to degrade a variety of xenobiotics,suggesting that they adapt well to contaminated environments and play important roles in environmental remediation.Different from typical Gram-negative bacteria,they contain sphingoglycolipids(SGLs)instead of lipopolysaccharides(LPSs)in their outer membranes.Since the carbohydrate parts of SGLs are much shorter than those of LPSs,sphingomonads are speculated to have higher cell surface hydrophobicity(CSH),which promote the degradation of hydrophobic xenobiotics.However,the CSH of sphingomonads has not been studied systematically and deeply.Its roles in environmental adaptation,influence factors,molecular mechanisms and evolution process are unclear.Sphingobium hydrophobicum C1,isolated from electronic waste(e-waste,containing various heavy metals and toxic organic compounds)contaminated river sediment by our group,is the most hydrophobic sphingomonad ever known.It can effectively degrade deca-brominated diphenyl ether(BDE-209)with zero-valent iron.This study investigated the advantageous characteristics of strain C1 involved in habitat adaptation,especially the roles of high CSH;determined the material bases and related genes of high CSH;analyzed the characteristics of the C1 genome and the mechanisms driving adaptive evolution.The main results were as follows:(1)Compared with hydrophilic mutant C2,S.xenophagum NBRC 107872(a hydrophilic and closely related strain)and E.coli ATCC 25922(a hydrophilic and well-characterized strain which widely used as a control for various laboratory experiments),hydrophobic strain C1 exhibited stronger adhesion to kaolinite and montmorillonite(the common components of soil and sediment),and had higher biofilm-forming ability,indicating that high CSH contributed to the colonization of strain C1 in sediment.Strain C1 could tolerate high concentrations of polybrominated diphenyl ethers and heavy metals,and could feed on several kinds of e-waste-related xenobiotics,such as phthalate esters,biphenyl and diphenyl ether.Its high CSH was related to the great heavy metal resistance to some extent,and promoted the adsorption of hydrophobic xenobiotic BDE-209 to improve the bioavailability.These characteristics provide strain C1 survival advantage in ewaste contaminated river sediment.(2)When strain C1 was exposed to environmental stress,such as lack of available nutrients,high temperature,acid or alkaline p H and heavy metals,its CSH would increase significantly,and more selfaggregation and more adsorption on the wall appeared,which might help to resist external environmental stress.When strain C1 was continuously passaged in nutrient-rich LB liquid medium,hydrophilic mutants would increase gradually in the culture.These results indicated that the high CSH of strain C1 was related to some adverse situations,and was a response to environmental stress.(3)By comparing the differences between strain C1 and hydrophilic mutant C2 in lipophilic dyes staining,infrared spectroscopy,Raman spectroscopy and cell surface components including membrane proteins,polar lipids,fatty acids and extracellular polymeric substances,and evaluating the effect of protease digestion on the CSH,the outer membrane proteins were found to be an important material basis of the high CSH.(4)The C1 and C2 genomes both consisted of two chromosomes and five large plasmids.Their main difference was copy number of the three smallest endogenous plasmids(P3,P4,P5),and the copy number in strain C2 was 2?3 times more than that in strain C1.The deletion of the replication initiation protein gene on P3 in C2(C2??rep)caused the decease of P3 copy number and the significant increase of CSH(69.2±3.9% vs 17.6±2.1%).Moreover,mutant C1??P3 which lost P3 was accidentally isolated in the culture of strain C1,and it harbored slightly higher CSH than strain C1(72.8±2.5% vs 62.8±4.0%).These results verified that the CSH was closely associated with P3.The transcriptome analysis of strains C1,C2 and C2??rep found that the variation of P3 could affect the expression of the genes on P3 coding the enzymes involved in fatty acid metabolism and glycosyl synthesis,an Ton B dependent receptor protein,an Tetr family regulatory protein(the expression regulation of some outer membrane protein genes),and many hypothetical proteins,which would further affect the global metabolic network and the expression of outer membrane protein genes.These ultimately would lead to the change of CSH.In addition,the high CSH of C2??rep and the long cell pellet pattern after centrifugation might also be related to the loss of the genes on P5 involved in glycosyl synthesis,transfer and polysaccharide secretion which might lead to defects in the synthesis of carbohydraterelated compounds and the formation of hydrophobic glycolipids.(5)Although the genes coding the outer membrane protein Oap,quorum sensing system and type IV secretion system were significantly differentially expressed between strains C1 and C2(The transcriptome and proteome results from other members of our group),the knockout of related genes did not change the CSH,indicating that they were not the decisive factor for the CSH.(6)The C1 genome contained many mobile elements.The comparison with 19 other Sphingobium genomes found the obvious sequence differences and many unique genes in the C1 genome.Moreover,the frequent genome rearrangements were observed in isolates from laboratory cultures of strain C1.These reflected high genome plasticity of strain C1.The abundant genes involved in environmental adaptation,such as xenobiotic degradation and heavy metal resistance,were located on the accessory chromosome Chr2,plasmids and genomic islands.Strain C1 might provide beneficial genes to surrounding microorganisms to promote community evolution by multiple sets of type IV secretion systems.According to the isolation time of closely related strains and the starting time of e-waste pollution,it was inferred that the genome of the C1 ancestor already contained most sequences of Chr1 and Chr2 before e-waste pollution,which included protocatechuic acid degradation gene clusters,multidrug efflux pump genes,etc.Positive selection,gene replication and horizontal gene transfer(HGT),especially the latter,had driven the adaptive evolution of strain C1.Many xenobiotic degradation genes,heavy metal resistance genes and CSH-related genes were derived from HGT.In summary,this study revealed the roles of high CSH of strain C1 in habitat adaptation,the material bases and related genes of high CSH,and the adaptive evolution process and driving forces of strain C1,which provide new insights into the relationship between microbial CSH and environmental adaptation,and the microbial adaptive evolution in e-waste contaminated environment,and provide theoretical guidance for the CSH regulation of sphingomonads and other microorganisms,as well as the bioremediation of contaminated environments. |
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