Molecular nature of methicillin-resistant Staphylococcus aureus derived from explosive nosocomial outbreaks of the 1980s in Japan

Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) with Panton-Valentine leukocidin (PVL) genes is increasing worldwide. Nosocomial outbreak-derived (hospital-acquired) MRSA (HA-MRSA) in Japan in the 1980s was also largely PVL(+). PVL(+) HA-MRSA and CA-MRSA shared the same multi-locus sequence type (ST30) and methicillin resistance cassette (SCCmecIV), but were divergent in oxacillin resistance, spa typing, PFGE analysis or clfA gene analysis. PVL(+) HA-MRSA, which probably originated in PVL(+)S. aureus ST30, was highly adhesive (carrying cna and bbp genes), highly-toxic (carrying luk(PV) and sea genes) and highly drug-resistant. PVL(+) HA-MRSA was once replaced by other PVL(-) HA-MRSA (e.g., ST5), and is re-emerging as CA-MRSA.     

马源马链球菌兽疫亚种3株新疆分离株基因型的鉴定及MLST分析

【背景】马链球菌兽疫亚种(Streptococcus equi subsp. zooepidemicus,SeZ)是引起马腺疫的主要病原,还可引起猪链球菌病,加强该菌的地方株分子流行病学监测对有效防控相关疫病十分必要。【目的】对新疆地区2个马场SeZ分离株进行鉴定和药敏特性分析,并分析3株新疆分离株的分子流行与菌株的遗传进化特征。【方法】对分离纯化的3株病原菌(ZHZ113、ZHZ211和ZHZ523)进行染色观察、生化及药敏特性检测,对16S rRNA和SeM基因进行遗传进化分析,以链球菌7个管家基因arcC、nrdE、proS、spi、tdk、tpi和yqiL为目的基因对3株分离菌进行多位点序列分型(Multilocus Sequence Typing,MLST)研究。【结果】3株SeZ的药敏结果显示这3株分离菌对不同抗生素的耐药程度不同,但均对头孢西丁、庆大霉素、链霉素、红霉素、左氧氟沙星、环丙沙星、土霉素等11种药物敏感。16SrRNA基因序列分析显示这3株分离菌均属于Ⅱ群(兽疫链球菌)。3株菌的MLST分型结果分别为ST39、ST419、ST421型,其中ST419和ST42...     

不同毒力差异基因型的新生隐球菌MLST分型及交配型鉴定研究

目的：了解及比较两组毒力差异明显的新生隐球菌格鲁比变种的多位点序列分型（MLST）的特点并进行交配型鉴定。方法采用多位点序列分型（MLST）的方法,设计7个看家基因（CAP59,GPD1,LAC1,PLB1,SOD1,URA5和IGS1）的引物,扩增并分析来源分别为环境和临床的各10株新生隐球菌格鲁比变种的基因型,并鉴定实验菌株交配型,与多位点微卫星分型（MLMT）结果对比,比较不同基因分型方法在分类中的稳定性和可靠性。结果在微卫星分型中为MLMT-36的10株环境分离株,MLST分型为ST-15,而在微卫星分型中为MLMT-13型的10株临床分离株,MLST分型为ST-32,所有菌株交配型均为MAT-α。结论MLST分型结果与MLMT分型结果高度一致,提示以上两种分子分型技术在真菌分类鉴定研究中可显示对于其分离背景及进化来源的高分辨率及稳定性。     

安徽部分地区动物源弯曲菌的分离鉴定及多位点序列分型

【背景】弯曲菌是一种重要的食源性人兽共患病原菌,革兰氏阴性、微需氧、弯曲螺旋状。【目的】为了解安徽地区弯曲菌流行状况和分子遗传特征,对安徽6个不同地区动物源的弯曲菌进行分离鉴定,并研究分离株分子分型。【方法】通过形态学及培养特性观察、生化试验、PCR方法对菌株进行鉴定。以弯曲菌7个管家基因asp A、gln A、glt A、gly A、pgm、tkt和unc A为目的基因对分离株进行多位点序列分型,并制成遗传进化树。【结果】共分离到42株弯曲菌菌株,源自6个地区的分离株具有较为一致的形态特性和相似的生化特性。多位点序列分型结果显示,本研究中共获得32种ST型,共发现9种新的ST型(8190、8222、8223、8831、8833、8841、8832、8834和8843)和6个新的等位基因(gln A606、gln A607、glt A518、gly A680、pgm863和unc A541)。进化树结果显示,空肠弯曲菌与结肠弯曲菌遗传关系相差甚远,聚集归为两个大群,分别有5个分支和3个分支。【结论】安徽6个地区不同来源的空肠弯曲菌与结肠弯曲菌均有丰富的基因型,且没有明显优势的基因型。从遗传变异的角度来看,空肠弯曲菌复杂多样,结肠弯曲菌相对保守。     

A multilocus sequence typing (MLST) approach to diminish the problems that are associated with DNA barcoding: A reply to Stahlhut et al. (2012)

In a recent Perspective, Stahlhut et al. (2012) argued that potential Wolbachia-induced effects on inheritance patterns of mitochondrial DNA do not significantly affect DNA barcoding efforts. Since this hypothesis can be readily tested, we suggest to do so by including multiple, nuclear markers in DNA barcoding studies.     

Chlamydia buteonis,a new Chlamydia species isolated from a red-shouldered hawk

Chlamydiaceae are obligate intracellular bacterial pathogens for humans and animals. A recent study highlighted that a Chlamydiaceae intermediary between C. psittaci and C. abortus can infect hawks. Here, an isolate was obtained upon passage of cloacal and conjunctival sac material collected from a female hatch-year red-shouldered hawk (Buteo lineatus) in cultured cells. The diseased bird, one of 12 birds housed in a rehabilitation center, developed conjunctivitis and later died. Swabs from both sites tested positive for Chlamydia using the QuickVue Chlamydia test. The isolate, named RSHA, tested negative in qPCR assays specific for C. psittaci and C. abortus, respectively. Analysis of the 16S rRNA, 23S rRNA and whole genome sequences as well as MLST, ANIb and TETRA values reveal that C. psittaci and C. abortus are the closest relatives of RSHA. However, the overall results strongly suggest a phylogenetic intermediate position between these two species. Therefore, we propose the introduction of a new species designated Chlamydia buteonis with RSHA^T as the type strain.     

山东省某地区奶牛源大肠埃希菌的血清型、耐药特性及分子特性

【目的】旨在对从山东省某地区4个健康奶牛养殖场分离到的大肠埃希菌进行优势血清型、耐药特性、Ⅰ类整合子基因盒携带情况以及系统进化群分析。【方法】采集194份来自山东省某地区4个规模化奶牛场奶牛新鲜粪便样品,进行大肠埃希菌分离和鉴定,利用常用大肠埃希菌诊断血清进行血清型鉴定;利用10%的绵羊血平板检测溶血性;利用K-B法检测对14种常规抗菌药物的敏感性;利用聚合酶链式反应(PCR)检测革兰阴性菌常见的6大类24种耐药基因、Ⅰ类整合子基因盒结构并对目的条带测序分析;利用细菌多位点序列分型(Multilocus sequence typing,MLST)技术分析大肠埃希菌的ST型并使用eBURST v3软件分析菌株之间的克隆关系。【结果】从194份新鲜粪便样品中分离到171株大肠埃希菌,其中主要为致病性(19.9%)和侵袭性大肠埃希菌(17.0%),优势血清型分别为O128:K67(12/171)和O143:K7(12/171)。另外,具有溶血性的大肠埃希菌阳性率为9.4%(16/171);药敏试验结果显示多重耐药菌株的比率为22.2%,其中对氨苄西林耐药率最高为33.9%,四环素次之,为24.0%;PCR检测耐药基因和整合子结果显示,59.1%的菌株携带β-内酰胺类耐药基因blaTEM,59.1%的菌株携带氨基糖苷类耐药基因ant(2′),未检测到四环素耐药基因tetA和tetB;Ⅰ类整合子的阳性率为4.1%(7/171),dfrA12-aadA2-sul1为优势基因盒结构(4/171);MLST将大肠埃希菌分为8种ST型,其中,ST155(10/171)和ST58(45/171)形成一个克隆复合物且没有发现新的ST型。【结论】本研究证实,从该地区规模化健康奶牛场新鲜粪便中分离到的大肠埃希菌优势血清型为O128:K67和O143:K7;少部分大肠埃希菌具有溶血性;仅对氨苄西林、四环素等具有较高的耐药率;优势基因盒结构为dfrA12-aadA2-sul1;MLST分型显示不同奶牛场分离出亲缘关系较近的菌株,其分布具有多态性,血清型与ST型之间无相关性。本研究表明源自表观健康的奶牛的大肠埃希菌存在多重耐药现象,具有食品公共卫生安全隐患,该研究对于提升规模化奶牛场奶制品的安全生产与质量评估具有一定的理论指导意义。     

Multilocus sequence typing of Campylobacter jejuni, and the correlation between clonal complex and pulsed-field gel electrophoresis macrorestriction profile

The characterization of Campylobacter jejuni has been significantly improved by the use of multilocus sequence typing (MLST), which allows the relationship between isolates to be determined. The sequence types (STs) of 261 isolates of C. jejuni from New Zealand were determined. Isolates were obtained from a range of sources including chicken meat, cattle, pigs, duck, sheep, water and human infections. Thirty-two new alleles and 44 new STs were identified. Comparison of the MLST data and pulsed-field gel electrophoresis macrorestriction profiles showed that the macrorestriction profiles were good predictors of the clonal complex (CC) but not ST. All the major CCs identified elsewhere in the world were found in New Zealand as well as the association of certain CCs with particular animal niches. The majority of new STs identified were from river water isolates.     

Presence of pathogenic cryptococci on trees situated in two recreational areas in South Africa

Knowledge of the environmental prevalence of members of the Cryptococcus neoformans/Cryptococcus gattii species complex is important, since cryptococcal infection is acquired from the environment. We determined whether trees located in two South African recreational areas harboured pathogenic cryptococci and compared the isolates to clinical isolates obtained from Western Cape hospitals with molecular typing techniques. The majority of isolates originating from trees in a public park in Cape Town (PPCT) were C. gattii sensu stricto, followed by C. neoformans sensu stricto genotype AFLP1/VNI. The PPCT trees might be a source of infection, since all genotype AFLP1/VNI isolates from these trees and one clinical isolate belonged to the same sequence type (ST), i.e. ST23. Recombination and basidiospore production might be occurring in PPCT trees that contained C. gattii s.s. isolates belonging to both mating types. The presence of C. gattii s.s. in PPCT trees might therefore pose a risk to human health.     

Molecular epidemiology of clinically high-risk Pseudomonas aeruginosa strains: Practical overview

In recent years, numerous outbreaks of multidrug-resistant Pseudomonas aeruginosa have been reported across the world. Once an outbreak occurs, besides routinely testing isolates for susceptibility to antimicrobials, it is required to check their virulence genotypes and clonality profiles. Replacing pulsed-field gel electrophoresis DNA fingerprinting are faster, easier-to-use, and less expensive polymerase chain reaction (PCR)-based methods for characterizing hospital isolates. P. aeruginosa possesses a mosaic genome structure and a highly conserved core genome displaying low sequence diversity and a highly variable accessory genome that communicates with other Pseudomonas species via horizontal gene transfer. Multiple-locus variable-number tandem-repeat analysis and multilocus sequence typing methods allow for phylogenetic analysis of isolates by PCR amplification of target genes with the support of Internet-based services. The target genes located in the core genome regions usually contain low-frequency mutations, allowing the resulting phylogenetic trees to infer evolutionary processes. The multiplex PCR-based open reading frame typing (POT) method, integron PCR, and exoenzyme genotyping can determine a genotype by PCR amplifying a specific insertion gene in the accessory genome region using a single or a multiple primer set. Thus, analyzing P. aeruginosa isolates for their clonality, virulence factors, and resistance characteristics is achievable by combining the clonality evaluation of the core genome based on multiple-locus targeting methods with other methods that can identify specific virulence and antimicrobial genes. Software packages such as eBURST, R, and Dendroscope, which are powerful tools for phylogenetic analyses, enable researchers and clinicians to visualize clonality associations in clinical isolates.