木糖葡萄球菌与金黄色葡萄球菌、表面葡萄球菌的基因组比较(阐述毒力、侵染性,样本是健康人的鼻腔)副本

LETTER TO THE EDITORGenome of Staphylococcus xylosus and Comparison with S.aureusand S.epidermidisStaphylococci are Gram-positive,AT-rich cocci,and often stick together in grape-like clusters.The genus can be classi-fied into two groups based on their ability to produce coagu-lase,an enzyme that causes clotting of blood plasma (Otto,2004).Coagulase-positive Staphylococci include Staphylo-coccus aureus ,a common pathogen of community-acquired and nosocomial infections (Smith et al.,2009).Their inva-siveness is associated with the ability to adhere to host sur-faces (Vuong et al.,2003).Among coagulase-negative Staphylococci (CNS),S.epidermidis is the most frequently found pathogen in humans,and is also a common cause of nosocomial infections (Nostro et al.,2007;Wang et al.,2009).S.epidermidis is believed to account for most of the infections caused by CNS and is highly resistant to many antibiotics including penicillins and cephalosporins (Al-Shuneigat et al.,2005).S.xylosus is also a CNS.It is naturally present in raw meat and milk and is commonly used in starter culture for fermentation (Planchon et al.,2006,2007).This species is normally regarded as non-pathogenic,but a few strains are related to human opportunistic infections (Akhaddar et al.,2010).In addition,some S.xylosus strains have the ability to form biofilm (Planchon et al.,2006).Bacterial virulence genes can be regulated by diffusible signal molecules termed autoinducers (AIs).Because the control of gene expression by AIs is cell-density dependent,this phenomenon has been called quorum sensing (Brelles-Marino and Bedmar,2001;Antunes et al.,2010).In Staphy-lococci,there are two quorum sensing systems,P2and P3.Their regulatory mechanism was described earlier by Liu et al.(2012).Other regulatory genes and virulence factors shared by S.epidermidis and S.aureus have also been reported (Frebourg et al.,2000;Gelosia et al.,2001).However,it is still not clear whether these virulence related genes are also present in non-pathogenic S.xylosus genome.In this study,we isolated S.xylosus NJ from a nasal sample of a healthy person at Jiangsu People’s Hospital in China and determined its genome sequence using whole-genome shotgun sequencing strategy with a Hiseq2000(Illumina,CA,USA)sequencer.The project generated a total of w 2739Mbsequences and w 927folds coverage of the genome.The draft genome data were assembled using the Velvet assembly pro-gram.The assembly generated 45contigs with a size of >200bp,22of which were longer than 500bp with the N 50length of 396,400bp.These 45contigs were deposited in GenBank and annotated using Rapid Annotation using Sub-system Technology (RAST)server.In addition,96tandem repeat sequences were found in these contigs.The draft genome contained a chromosome of 2,940,053bp with a 32.40%G þC content.The general features are listed in Table 1.There are 6predicted rRNA genes and 22tRNA genes,and 83.64%nucleotides are predicted to encode proteins.By a combination of coding potential prediction and homology search,2783coding DNA sequences (CDSs)with an average length of 884bp were identified on the draft genome (Table 1).The 2199CDSs annotated by specific Clusters of Orthologous Groups (COG)function groups can be classified into 21COG categories,and 2456CDSs can be annotated into 1221KEGG orthology by KAAS (Moriya et al.,2007).The organization of the genome of S.xylosus NJ was shown in a circular map in Fig.1A.In addition,phylogenetic dendrogram based on a comparison of 16S rRNA sequences for S.xylosus NJ with members of the Staphylococcus was shown in Fig.S1.There is another S.xylosus draft genome,S.xylosus DMB3-Bh1,in GenBank.Therefore,we compared S.xylosus NJ with S.xylosus DMB3-Bh1using BLAST (version 2.2.26).The result showed that there are 2319CDSs in S.xylosus DMB3-Bh1(89.30%)similar to S.xylosus NJ (Fig.S2).The metabolic network of S.xylosus NJ was constructed using the RAST server with the 411subsystems identified in the genome.There are many carbohydrate subsystem features,including genes involved in organic acids,fermentation,sugar alcohols,di-and oligo-saccharides,central carbohydrates,monosaccharides,and one-carbon metabolisms.Many protein metabolism features are also present,including protein biosynthesis machinery such as the small subunit (SSU)and large subunit (LSU)of the bacterial ribosome.Moreover,we prepared the comparative analysis of this genome with other staphylococcal genomes (S.aureus subsp.aureus N315(S.Available online at ScienceDirectJournal of Genetics and Genomics 41(2014)413e 416JGG1673-8527/$-see front matter Copyright Ó2014,Institute of Genetics and Developmental Biology,Chinese Academy of Sciences,and Genetics Society of China.Published by Elsevier Limited and Science Press.All rights reserved./10.1016/j.jgg.2014.03.007aureus N315)and S.epidermidis ATCC12228).The results showed that there are 404subsystems for S.aureus N315genome and 381subsystems for S.epidermidis ATCC12228genome.A comparative metabolic network analysis showed that D-galacturonate and D-glucuronate utilization subsystem is only present in S.xylosus NJ genome.However,serine-glyoxylate cycle subsystem is absent in S.aureus N315genome.A more detailed comparative analysis of this genome with these two staphylococcal genomes is summarized in Table S1.The genomic organization of S.xylosus NJ was compared with those of S.aureus N315and S.epidermidis ATCC12228(Table 1).There are 1502CDSs (55.20%)in S.xylosus NJ similar to those in S.aureus N315and 1446CDSs (53.14%)similar to those in S.epidermidis ATCC12228.The number of shared genes among S.xylosus NJ,S.aureus N315,and S.epidermidis ATCC12228was shown in Fig.1B.Over 92%of homologous genes were assigned to COG function groups (Fig.S3).In addition,the larger amount of unique proteins is present for S.xylosus NJ with 1145proteins.Meanwhile,unique genes of three strains were assigned to COG function group.The result showed that the numbers of genes related to energy production and conversion,carbohydrate transport and metabolism,and general function in S.xylosus NJ genome are considerably more than other strains (Fig.S4).Compared to S.aureus N315and S.epidermidis ATCC12228genomes,S.xylosus NJ genome possesses some significant differences,such as virulence and defense ability,cell wall and capsule constitution,phages,prophages,trans-posable elements and plasmids,and cell regulation and signaling.In virulence and defense category,adhesins have previously been reported as virulence factors by promoting the accumu-lation phase of biofilm (Corrigan et al.,2007;Kim et al.,2010).In consistent with the non-pathogenic property,no adhesin was detected in S.xylosus NJ.In contrast,six adhesins were present in S.epidermidi s ATCC12228and 22in S.aureus N315(Table S2).Another important gene family,staphylococcal enterotoxins (SEs)encodes powerfulsuperantigens that stimulate non-specific T-cell proliferation (Balaban and Rasooly,2000).However,there was no gene associated with SEs in S.xylosus NJ genome.S.aureus pro-duce a -hemolysin,g -hemolysin,and leukocidin toxins,which function as two component toxins in the disruption and lysis of erythrocytes and leukocytes (Gouaux et al.,1997).Compared with S.aureus N315,there was no gene in the subcategory of toxins and superantigens in S.xylosus NJ.However,S.xylosusTable 1Chromosome features of S.xylosus NJ,S.aureus N315,and S.epidermidis ATCC12228S.xylosus S.aureus S.epidermidisFig.1.The genomic organization of S.xylosus NJ and the comparison with other Staphylococcus .A:Circular representation of the S.xylosus NJ chromosome.GC-skew:sliding window size of 10kb and calculating (G ÀC)/(G þC)in 500bp steps.Circles from outside to inside:1,contigs were arrange in clockwise direction from large to small;2,CDS on forward strand;3,CDS on reverse strand;4,tRNA genes;5,rRNA genes;6,GC-skew (window size of 10kb);and 7,purple indicates C content and yellow indicates G content (step size 500bp).B:Venn diagram for the deduced proteins in S.xylosus NJ,S.aureus N315and S.epidermidis ATCC12228.The total number of all deduced proteins for these three species is 7781.The number of proteins per each chromosome is given.Values were identified by BLASTCLUST (version 2.2.26)using an identity of >60%,an alignment coverage of >70%and an e -value of 1e-6as cut-off.The number of clusters represents the non-redundant protein coding genes for intersection.The overlapping sections indicate shared numbers of proteins.414Letter to the Editor /Journal of Genetics and Genomics 41(2014)413e 416NJ carried fewer antibiotic-resistance genes.For instance, teicoplanin andfluoroquinolones resistance genes were pre-sent in S.xylosus NJ.Meanwhile,multidrug-resistance efflux pumps were also present in S.xylosus NJ,which supports a previous observation made by Piddock(2006)that multidrug-resistance efflux pumps have roles not only for resistance to antibiotics,but also in bacterial pathogenicity.However, methicillin resistance genes were not found in S.xylosus NJ.In cell wall and capsule category,sialic acid is a bioavail-able carbon and nitrogen source that is abundant on mucosal surfaces and in secretions in the commensal environment (Olson et al.,2013).It has been shown by Almagro-Moreno and Boyd(2009)that several pathogens,including S.aureus N315,could utilize sialic acid as a carbon source,which in turn establish a competitive advantage in heavily sialylated environments such as the human gut.Our data indicate that S. xylosus NJ,but not S.epidermidis ATCC12228,contains genes associated with sialic acid metabolism(Table S3).Most Staphylococci produce types5and8microcapsules,which account for75%of human infections(Lowy,1998).Types5 and8cap genes were not found in S.xylosus NJ(Table S3).In category of phages,prophages,transposable elements and plasmids,Tn552is not detected in the genome of S.xylosus NJ. The staphylococcal pathogenicity islands(SaPIs)encoded one or more staphylococcal superantigens.We identified17path-ogenicity islands in S.aureus N315.However,no one was detected in S.xylosus NJ(Table S4).Interestingly,in contrast to S.aureus N315and S.epidermidis ATCC12228,ten copies related to phages and prophages were detected in S.xylosus NJ.In category of cell regulation and signaling,S.xylosus NJ showed significant differences in biofilm formation and staph-ylococcal accessory gene regulator systems compared to S. aureus N315and S.epidermidis ATCC12228.Besides adhesins absence in S.xylosus NJ,entire ica genes involved in biofilm formation were not found.Although a few biofilm-positive strains do not produce detectable polysaccharide intercellular adhesin(PIA),the mechanism was not due to the absence of the ica operon,but was a result of reduced transcription or trans-lation of ica genes(Zhang et al.,2003).In addition,S.epi-dermidis ATCC12228is non-biofilm-forming strain due to lack of ica operon(Zhang et al.,2003).So,we speculated that S. xylosus NJ may not form biofilm.However,staphylococcal accessory regulator A(SarA),biofunctional autolysin Atl,and RNA polymerase sigma factor SigB were present in S.xylosus NJ(Table S5).Therefore,we speculated that the function of these genes may be not for biofilm formation,but for others such as virulence factor production.In staphylococcal acces-sory gene regulator system,there were13genes in S.aureus N315and8in S.epidermidis ATCC12228,but none in S. xylosus NJ(Table S5).This may explain the great difference among these strains in virulence.One of the important factors contributing to staphylococcal toxin is their peptide-based quorum sensing system,encoded by the accessory gene regu-lator(agr)locus(Balaban et al.,2004;Kiran et al.,2008; Antunes et al.,2010).The agr locus(agr ABCD)and related genes were not detected in S.xylosus NJ.The comparative genomic analysis suggests that S.xylosus NJ may be less virulent.In addition,we found that ElaA protein and prophage Clp protease-like protein were only present in S.xylosus NJ.In summary,the comparative genomic analysis of S.xylo-sus NJ with S.aureus N315and S.epidermidis ATCC12228 showed great difference on genome composition within this genus.Particularly,ica operon and agr operon were absent in S.xylosus NJ.Here,we described the genome analysis of S. xylosus NJ to provide a new genome for Staphylococcus genome database.In addition,we analyzed genes in S.xylosus NJ different from those in S.aureus N315and S.epidermidis ATCC12228.Their differences mainly exist in genes associ-ated with virulence and pathogenicity,such as adhesins, superantigens,resistance to antibiotics,capsular poly-saccharide synthesis enzymes,pathogenicity islands,quorum sensing and biofilm formation.This study can help reveal how commensal S.xylosus strains evolved into invasive strains. ACKNOWLEDGMENTSThis work was supported by the grants from the National Natural Science Foundation of China(Nos.81301475, 31170131and31070312)and Jiangsu Qinglan Project,and the Shaoxing Major Scientific and Technological Projects(No. 2011A11013).SUPPLEMENTARY DATASupplementary data related to this article can be found at /10.1016/j.jgg.2014.03.007Xiaojuan Tan a,1,Lin Liu b,c,1,Shili Liu d,Dongting Yang a, Yikun Liu c,Shuang Yang c,Aiqun Jia a,*,Nan Qin b,* a School of Environmental and Biological Engineering,Nanjing University ofScience and Technology,Nanjing210094,Chinab State Key Laboratory for Diagnosis and Treatment of Infectious Disease, The First Affiliated Hospital,Zhejiang University,Hangzhou310003,Chinac NGS Sequencing Department,Beijing Genomics Institute(BGI),Shenzhen518083,China d Institute of Pathogen Microbiology,School of Medicine,Shandong University,Jinan250012,China*Corresponding authors.Tel/fax:þ86057187236421. 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