抗生素对溶藻弧菌SXT/R391元件转移频率的影响北大核心CSCD

【目的】研究萘啶酸、诺氟沙星、卡那霉素3种抗生素对溶藻弧菌(Vibrio alginolyticus)SXT/R391元件ICEVal A056-1转移频率的影响。【方法】利用PCR检测溶藻弧菌A056中ICEVal A056-1的自我剪切、转移潜力。通过溶藻弧菌A056与大肠杆菌菌株VB111的接合实验,研究溶藻弧菌分别在含不同浓度萘啶酸、诺氟沙星、卡那霉素的LB培养基中培养15 min或30 min后,ICEVal A056-1转移频率的变化规律。【结果】溶藻弧菌A056细胞中有环状形式的ICEVal A056-1分子存在,具有水平转移潜力;溶藻弧菌A056在含40μg/m L萘啶酸的LB中培养30 min后,ICEVal A056-1转移频率是对照组的19.59倍;在含50μg/m L诺氟沙星的LB中培养15 min后,ICEVal A056-1转移频率是对照组的31.25倍;在含不同浓度卡那霉素的LB中培养30 min后,ICEVal A056-1转移频率与对照组没有显著差别。【结论】部分抗生素的使用可以明显促进溶藻弧菌ICEVal A056-1向大肠杆菌的转移,因此海洋环境中抗生素的滥用及随意排放很可能加剧ICEs(integrating conjugative elements)从溶藻弧菌到其他细菌的传播。     

A novel ICE in the genome of Shewanella putrefaciens W3-18-1: comparison with the SXT/R391 ICE-like elements

A novel R391-like ICE (integrating conjugative element) has been detected in the 4.2 MB genome of Shewanella putrefaciens W3-18-1 located on three different contigs. Assembly of the ICE encoding contigs based on similarity with R391 revealed a mosaic element of plasmid, phage and transposon-like sequences typical of SXT/R391 ICE-like elements. The element, which is 110 057 bp in length, was highly similar to R391 sequences, with most related ORFs showing >96% amino acid sequence identity. The element, designated ICESpuPO1, contained a number of inserts determining resistance to copper and other heavy metals and a broad-spectrum RND efflux pump similar to antibiotic efflux systems. The element was integrated into the Shewanella prfC gene in a manner similar to related ICE-like elements. The chromosomal element junctions contained a 17-bp SXT/R391-like attL and attR site and an unannotated ORF between attL and the ICE integrase encoding a putative recombinational directional factor necessary for excision, with 100% amino acid identity to the R391 ORF4 product.     

Genomic analysis of a novel integrative conjugative element in Vibrio cholerae

Integrative conjugative elements (ICEs) are a class of self-transmissible mobile elements that mediate horizontal gene transfer in bacteria, and play an important role in bacterial evolution. Since 1992, ICEs of the SXT/R391 family have been found to be widely distributed among Vibrio cholerae strains isolated in Asian countries. Here we describe ICEVchB33, an ICE found in the genomes of two V. cholerae O1 Eltor strains, one isolated in India, 1994, and the other from Mozambique, 2004. ICEVchB33 revealed a new genetic organization, different from other ICEs of the SXT/R391 family, demonstrating the genomic plasticity of these elements.     

副溶血弧菌TF2中整合接合元件ICEVpaTF2的生物信息学特征和剪切、环化活性

[目的] 从副溶血弧菌TF2基因组框架序列中拼接获得整合性接合元件（ICE）ICEVpaTF2的全序列,分析其基因组学特征;研究ICEVpaTF2是否具有从基因组中剪切、环化活性以及剪切、环化时attB和attP位点如何形成。[方法] 对副溶血弧菌TF2基因组框架序列进行RAST注释,发现其基因组中可能存在一个完整的ICE元件,命名为ICEVpaTF2,经过人工拼接和PCR扩增测序验证,得到完整的ICEVpaTF2序列,并对ICEVpaTF2再次进行RAST注释和ICE元件部分特征分析。通过PCR检测,探索ICEVpaTF2是否能够从基因组剪切并环化。通过attL、attR、attB、attP位点序列比较,探索attB和attP重组特征。[结果] ICEVpaTF2全长83588 bp,包含SXT/R391家族ICE元件的52个保守核心基因,它们与ICE切除、整合、自我转移和调节机制相关。ICEVpaTF2也包含5个外源基因插入的热点区（HS）、2个可变区（VR）以及3个非典型插入位点。HS和VR包含了大量可变基因,它们负责编码限制性修饰系统、DNA修复系统或毒素-抗毒素系统等,赋予宿主广泛适应性功能,ICEVpaTF2也包含独特未知功能基因。通过对int、xis二个核心保守基因种系发生分析,发现ICEVpaTF2的int、xis分别属于以R391和SXT为代表的亚群。ICEVpaTF2在attL和attR处发生剪切并完成环化,新形成杂合重组的attB和attP位点。[结论] ICEVpaTF2属于SXT/R391家族,是一个具备自我剪切和环化能力的完整ICE元件,剪切后新形成的attB和attP位点由attL、attR杂合重组形成。     

Molecular characterization of <Emphasis Type="Italic">Vibrio cholerae</Emphasis> isolates from cholera outbreaks in north India

Vibrio cholerae isolates recovered from cholera outbreaks in Bhind district of Madhya Pradesh and Delhi, Northern India were characterized. The O1 serogroup isolates from Bhind outbreak were of Inaba serotype whereas both Ogawa and Inaba serotypes were recovered from Delhi. PCR analysis revealed that only O1 serogroup V. cholerae isolates carried the virulence-associated genes like ctxA, tcpA, ace, and zot. Molecular typing by repetitive sequence based ERIC, VCR1, and VC1 PCR’s revealed similar DNA profile for both Inaba and Ogawa serotypes. A discrete VC1-PCR band identified among the El Tor strains had greater similarity (>97%) to the V. cholerae genome sequence and therefore has the potential to be used as a marker for the identification of the V. cholerae strains. Non-O1 strains recovered from Bhind region differed among themselves as well as from that of the O1 isolates. All the O1 serogroup isolates possessed SXT element and were uniformly resistant to the antibiotics nalidixic acid, polymyxin-B, furazolidone, cloxacilin, trimethoprim-sulfamethaxazole, and vibriostatic agent 0129. Inaba strains from both Delhi and Bhind differed from Ogawa strains by their resistance to streptomycin despite sharing similar DNA patterns in all the three rep-PCRs. Though Delhi and Bhind are separate geographical regions in Northern India, Inaba strains from both these places appear to be closely related owing to their similarity in antibiogram and genetic profile.     

A variant type of Vibrio cholerae SXT element in a multidrug-resistant strain of Vibrio fluvialis

Vibrio fluvialis strain H-08942 was isolated from an infant aged 6 months who was suffering from cholera-like diarrhea in India. This strain showed the typical multidrug-resistance phenotype of an SXT element. It was resistant to sulfamethoxazole (Su), trimethoprim (Tm), chloramphenicol (Cm) and streptomycin (Sm), in addition to other antibiotics such as ampicillin (Am), furazolidone (Fz), nalidixic acid (Na), and gentamicin (Gm). The SXT element is a Vibrio cholerae-derived integrative and conjugative element (ICE) that has also been referred to as a conjugative transposon. Our goal was to find a relationship between these resistant phenotypes and the presence of the SXT element in this unique strain. By using PCR, we detected the antibiotic resistance genes, the integrase gene and the attP attachment site of SXT element. Cloning and DNA sequencing results showed that both the SXT integrase gene and its attP site of V. fluvialis were similar but not identical to those of V. cholerae. The SXT integrase gene of V. fluvialis has a 99% identity to that of V. cholerae, and the attP site of SXT of V. fluvialis is variant and shorter (641 bp) than that of V. cholerae (785 bp). It was possible for the SXT of V. fluvialis to be transferred by conjugation to a laboratory strain of Escherichia coli. Here, we report the detection of a variant SXT element in species other than V. cholerae, with molecular characterization and analysis of its integrase gene and its attP site.