金黄色葡萄球菌耐药基因及新型抗菌药物的研究进展

金黄色葡萄球菌(Staphylococcus aureus,S.aureus),简称金葡菌,是一种临床常见致病菌,可引起皮肤化脓性感染、肺炎、心包炎、败血症及脓毒血症等多种疾病。目前临床上金黄色葡萄球菌的多重耐药情况已十分严峻,特别是耐甲氧西林金黄色葡萄球菌(methicillin-resistant Staphylococcus aureus,MRSA)的出现,已成为临床抗感染治疗的棘手难题,严重危害人类健康。本文主要介绍金黄色葡萄球菌基因水平的耐药性、新型抗菌药物的研发及临床治疗金黄色葡萄球菌感染三方面的进展,为金黄色葡萄球菌的耐药性研究及耐药菌株的临床有效治疗提供科学依据。     

V型羊毛硫肽雷可肽的抑菌机制

[背景]雷可肽(Lexapeptide)为首例V型羊毛硫肽家族化合物,具有较好的抗革兰氏阳性菌活性,对耐甲氧西林金黄色葡萄球菌(Methicillin-Resistant Staphylococcus aureus,MRSA)和表皮葡萄球菌(Methicillin-Resistant Staphylococcus ep...     

紫草素与依布硒啉对金黄色葡萄球菌的协同作用

[目的]探讨紫草素(shikonin,SKN)协同依布硒啉(ebselen,EbSe)抗金黄色葡萄球菌(Staphylococcus aureus,S.aureus)的作用及机制.[方法]分光光度法测定紫草素-依布硒啉抗金黄色葡萄球菌活性;碘化丙啶(propidium iodide,PI)染色检测紫草素-依布硒啉杀菌效...     

2009—2012年金黄色葡萄球菌血液分离株的临床分布和耐药性研究

目的调查和分析金黄色葡萄球菌败血症患者的临床特点和菌株的耐药情况,为临床诊断和合理用药提供参考。方法收集并分析2009年至2012年血培养确诊为金黄色葡萄球菌败血症患者的临床及菌株资料。结果2009年至2012年金黄色葡萄球菌血液分离株共80株,其中甲氧西林耐药金黄色葡萄球菌（methieillin resistant Staphylococcus,MRSA）占50％。2009年至2011年对甲氧西林的耐药率逐年提高,2012年呈下降趋势。其中MRSA血流感染患者多为合并基础疾病的老年人,留置静脉导管及体腔引流管、气管插管、联合使用抗生素、住院时间长为易感因素。结论金黄色葡萄球菌血液分离株中,MRSA检出率高,占50％,临床表现大多较重,合并多种基础疾病。加强抗生素的管理及重视和预防院内感染后能明显降低MRSA的检出率。     

金黄色葡萄球菌病防治研究进展

金黄色葡萄球菌(Staphylococcus aureus, SA)被认为是最常见的食源性致病菌之一,引起人畜的感染性疾病,导致皮肤、软组织和血液感染,引发脓毒症和中毒性休克综合征。随着抗生素的滥用,金黄色葡萄球菌的耐药性逐渐增强,导致耐甲氧西林金黄色葡萄球菌(methicillin resistant Staphylococcus aureus, MRSA)的出现,并且在全球范围内散播,严重危害公共卫生安全。目前亟需有效控制SA感染的新疗法,因此本文对金黄色葡萄球菌防治技术的研究进展进行综述,并对其防治前景进行了分析,以期对金黄色葡萄球菌尤其是MRSA的控制提供理论指导。     

金黄色葡萄球菌预防及治疗研究进展

金黄色葡萄球菌(Staphylococcus aureus)是院内获得性感染的主要病原体之一,金黄色葡萄球菌会引起患者的严重感染,对患者的治疗和预后产生不良影响。现今临床预防和治疗金黄色葡萄球菌感染主要依赖抗生素。近年来,随着金黄色葡萄球菌耐药性的增强,万古霉素(Vancomycin)成为治疗其感染的最后一道抗生素防线。目前迫切需要探寻新的治疗方法来代替抗生素治疗。现归纳针对金黄色葡萄球菌耐药性的最新疗法,旨在为预防和治疗金黄色葡萄球菌感染提供新的思考。     

野生抚育山参内生真菌分离鉴定

采用组织块法对野生抚育山参(Panax ginseng)根中内生真菌进行分离,共分离出16株内生真菌,根据形态鉴定出10株,分别隶属于头孢属(Cephalosporium)、青霉属(Penicillium)、柱孢属(Cylindrocarpon)、轮枝孢属(Verticillium)、单端孢属(Trichothecium)。以金黄色葡萄球菌(Staphylococcus aureus)、大肠杆菌(Escherichia coli)、白色念珠菌(Candida albicans)为指示菌对内生真菌进行抑菌活性筛选,共筛选出2株内生菌,这2株内生菌对金黄色葡萄球菌有较强的抑制作用。     

耐甲氧西林金黄色葡萄球菌及其肠毒素的检测和耐药性分析

目的评价乳胶结合试验检测耐甲氧西林金黄色葡萄球菌(MRSA)及其肠毒素(SE),并进行耐药性分析.方法收集130株金黄色葡萄球菌临床分离株,通过药敏试验将其分为耐甲氧西林金黄色葡萄球菌和甲氧西林敏感金黄色葡萄球菌(MSSA),用反向间接血凝试验(RPHA)检测金黄色葡萄球菌肠毒素.结果67株MR-SA产肠毒素,19株MSSA产肠毒素,MRSA产肠毒素率为100%,MSSA产肠毒素率为30%.结论实验室应重视金黄色葡萄球菌肠毒素的检测.     

类SCC样甲氧西林敏感性金黄色葡萄球菌的基因组学分析

金黄色葡萄球菌(Staphylococcus aureus, S. aureus), 作为一种常见的致病菌常可导致严重感染性疾病. 金黄色葡萄球菌的主要特点是易产生耐药性. 临床上运用多重PCR方法对S. aureus耐药菌株进行分型. SCC作为一种mecA基因的转运载体, 介导金黄色葡萄球菌产生耐药性. 本研究报道了一株新甲氧西林敏感性金黄色葡萄球菌的全基因组序列信息, 即类SCC样MSSA463菌株, 经多重PCR方法, 该菌株被误鉴定为耐甲氧西林金黄色葡萄球菌, 结果与药敏结果相冲突. 为此, 运用焦磷酸测序方法完成了该菌株的全基因组测序, 并与已知金黄色葡萄球菌的基因组序列信息不同, 发现可读框(CZ049; AB037671)存在于attL, attR反向重复序列的临近下游. 这些结果提示, 在金黄色葡萄球菌与其他微生物之间可能存在一种水平基因转移, 并改变了金黄色葡萄球菌对药物的敏感性, 推测attL, attR反向重复序列可能作为外源性基因的插入部位而存在.     

一株金黄色葡萄球菌噬菌体的分离鉴定与生物学特性

[背景]金黄色葡萄球菌作为一种条件致病菌,在临床感染中扮演着重要的角色,需要研究更多更有效的防治手段.[目的]分离金黄色葡萄球菌噬菌体,研究其生物学特性,从而为金黄色葡萄球菌的替代防治提供理论借鉴.[方法]以金黄色葡萄球菌D085为宿主从污水中分离得到一株噬菌体,命名为vB_SauS_SAP3,用PEG8000浓缩、氯...     

Characterization of Virulence Factors and Genetic Background of Staphylococcus aureus Isolated from Peking University People’s Hospital Between 2005 and 2009

The aim of this study was to investigate both the genetic features of MRSA strains and the occurrence of virulence factors produced by Staphylococcus aureus strains isolated from Peking University People’s Hospital in Beijing, China, between 2005 and 2009. A total of 179 S. aureus strains were isolated, 139 of which were MRSA. The MRSA strains were characterized epidemiologically by SCCmec typing, spa typing and agr typing, then were classified into different genetic groups. The prevalence of genes coding for 14 exotoxins and eight adhesion factors among the S. aureus samples was assessed via polymerase chain reaction. Cluster analysis based on virulence factors-encoding gene content was performed to divide the MRSA isolates into valid clusters. Correspondence analysis was done to analyze the correlation between virulence factors clusters and genetic groups. JCSC1716-agrI-t030 (67.6%), SCCmec-IIIA-agrI-t030 (14.4%), SCCmec-IIIA-agrI-t037 (8.6%) and SCCmecII-agrII-t002 (2.2%) were four predominant MRSA clones. PVL was positive only in MSSA strains, there were at least three superantigenic toxins in our HA-MRSA clones, the prevalence of 16 virulence factors genes (sea, seb, sec, sed, seg, sei, sej, pvl, lukE-lukD, eta, bbp, can, ebp, clfA, fib, fnbB) in MRSA and MSSA was found to be significantly different from MSSA (P < 0.05). Results of correspondence analysis among clusters based on virulence factors genes and groups based on genetic typing illustrated not only the correspondence relationship between groups and clusters overall (P < 0.001), but also the genetic diversity of MRSA strains with respect to virulence factors genes.     

Genome sequence of Staphylococcus aureus strain Newman and comparative analysis of staphylococcal genomes: polymorphism and evolution of two major pathogenicity islands

Strains of Staphylococcus aureus, an important human pathogen, display up to 20% variability in their genome sequence, and most sequence information is available for human clinical isolates that have not been subjected to genetic analysis of virulence attributes. S. aureus strain Newman, which was also isolated from a human infection, displays robust virulence properties in animal models of disease and has already been extensively analyzed for its molecular traits of staphylococcal pathogenesis. We report here the complete genome sequence of S. aureus Newman, which carries four integrated prophages, as well as two large pathogenicity islands. In agreement with the view that S. aureus Newman prophages contribute important properties to pathogenesis, fewer virulence factors are found outside of the prophages than for the highly virulent strain MW2. The absence of drug resistance genes reflects the general antibiotic-susceptible phenotype of S. aureus Newman. Phylogenetic analyses reveal clonal relationships between the staphylococcal strains Newman, COL, NCTC8325, and USA300 and a greater evolutionary distance to strains MRSA252, MW2, MSSA476, N315, Mu50, JH1, JH9, and RF122. However, polymorphism analysis of two large pathogenicity islands distributed among these strains shows that the two islands were acquired independently from the evolutionary pathway of the chromosomal backbones of staphylococcal genomes. Prophages and pathogenicity islands play central roles in S. aureus virulence and evolution.     

Comparison of the Virulence of Methicillin-Resistant and Methicillin-Sensitive Staphylococcus aureus

The virulence of methicillin-resistant Staphylococcus aureus (MRSA) was compared with that of methicillin-sensitive S. aureus (MSSA), using 13 MRSA and 7 MSSA strains isolated from clinical specimens. The infectivity and lethality of the two groups were examined as to the inoculum required to infect 50% of guinea pigs (ID₅₀) and to kill 50% of mice (LD₅₀), respectively. The mean ID₅₀ [log₁₀ colony forming units (CFU)] for MRSA strains was 7.1 ± 0.60 standard deviation, which was 1.5 higher than that for MSSA strains (P < 0.001). The mean LD₅₀ (log₁₀ CFU) for MRSA strains was 9.0 ± 0.42, being 1.1 higher than that for MSSA strains (P = 0.001). Pretreatment of mice with cyclophosphamide decreased the mean LD₅₀ for MRSA strains more than that for MSSA strains, resulting in the difference in the mean LD₅₀ being insignificant (P = 0.502). These results indicate that MRSA is less virulent than MSSA in normal hosts, but that they are equally virulent in immunocompromised hosts. The growth of MRSA strains was much slower than that of MSSA strains in the lag phase, although their growth rates were almost the same in the exponential growth phase, suggesting that the difference in virulence between them may be at least partly due to such a difference in growth.     

Molecular Analysis of Human and Canine Staphylococcus aureus Strains Reveals Distinct Extended-Host-Spectrum Genotypes Independent of Their Methicillin Resistance

Staphylococcus aureus causes a wide range of infectious diseases in humans and various animal species. Although presumptive host-specific factors have been reported, certain genetic lineages seem to lack specific host tropism, infecting a broad range of hosts. Such Extended-Host-Spectrum Genotypes (EHSGs) have been described in canine infections, caused by common regional human methicillin-resistant S. aureus (MRSA) lineages. However, information is scarce about the occurrence of methicillin-susceptible S. aureus (MSSA) EHSGs. To gain deeper insight into EHSG MSSA and EHSG MRSA of human and canine origin, a comparative molecular study was carried out, including a convenience sample of 120 current S. aureus (70 MRSA and 50 MSSA) isolates obtained from infected dogs. spa typing revealed 48 different spa types belonging to 16 different multilocus sequence typing clonal complexes (MLST-CCs). Based on these results, we further compared a subset of canine (n = 48) and human (n = 14) strains, including isolates of clonal complexes CC5, CC22, CC8, CC398, CC15, CC45, and CC30 by macrorestriction (pulsed-field gel electrophoresis [PFGE]) and DNA-microarray analysis. None of the methods employed was able to differentiate between clusters of human and canine strains independently of their methicillin resistance. In contrast, DNA-microarray analysis revealed 79% of the 48 canine isolates as carriers of the bacteriophage-encoded human-specific immune evasion cluster (IEC). In conclusion, the high degree of similarity between human and canine S. aureus strains regardless of whether they are MRSA or MSSA envisions the existence of common genetic traits that enable these strains as EHSGs, challenging the concept of resistance-driven spillover of MRSA.     

Exploring extra-cellular proteins in methicillin susceptible and methicillin resistant Staphylococcus aureus by liquid chromatography–tandem mass spectrometry

Staphylococcus aureus (S. aureus) strains cause several diseases in humans from minor skin infections to severe lethal infections. To explore the virulence determinants of this important microorganism, two clinical isolates of methicillin susceptible S. aureus (MSSA) and methicillin resistant S. aureus (MRSA) were subjected to proteomic analysis of their extracellular products using liquid chromatography–tandem mass spectrometry. The numbers of proteins identified in MSSA and MRSA extracellular products were 168 and 261; respectively, from them 117 were shared, while 144 proteins were unique to MRSA. The shared proteins, having a higher protein score with increased number of peptide matches in MRSA over MSSA, reflect the relatively active secretory state of MRSA rather than biased analytical variances. Characteristic determinants for MRSA were identified; mostly found to play a role in the virulence. We conclude that MRSA produces distinct proteins considered as its virulence determinants and we found that the shared extracellular products are more abundant in MRSA than MSSA that supporting the high invasiveness of MRSA over MSSA in pathogenesis.     

苯唑西林敏感、mecA基因阳性的猪源金黄色葡萄球菌的流行、分子分型及耐药性研究

[目的] 为了解我国猪源苯唑西林敏感-mecA阳性金黄色葡萄球菌（Oxacillin-susceptible,mecA-positive Staphylococcus aureus,OS-MRSA）的流行情况、菌株分子特征及耐药性,本研究对我国中西部4个省份（甘肃、陕西、河南和广西）的9个规模化养猪场进行鼻腔拭子样本采集。[方法] 运用PCR扩增nuc和mecA基因及苯唑西林耐药性检测对OS-MRSA菌株进行分离鉴定。然后对分离所得的OS-MRSA菌株进行26种毒素编码基因、16种抗生素耐药性以及spa、MLST和SCCmec分型检测。[结果] 结果表明,采集的884份样本中,67份样本7.6%（67/884）分离到金黄色葡萄球菌,包括50株甲氧西林敏感菌株（Methicillin-sensitive Staphylococcus aureus,MSSA）、8株苯唑西林耐受-mecA阳性金黄色葡萄球菌（Oxacillin-resistant mecA-positive,OR-MRSA）和9株OS-MRSA菌株。26种被检毒素编码基因中有9种毒素编码基因被检出,其中hla基因检出率最高,其次为hld、hlb、hlg、sei、sem、seg、sen和seo。此外,67株分离株中仅有16株携带肠毒素编码基因,其中OR-MRSA和OS-MRSA菌株分别占37.5%（6/16）和50.0%（8/16）,且携带毒素编码基因的菌株克隆型均为ST9-t899。16种所测试抗生素中,菌株对12种抗生素表现为耐药,其中MSSA、OR-MRSA和OS-MRSA分离株分别主要对1-8、10-12和7-11种抗生素耐药。所有分离株共有4种克隆型ST398-t571、ST9-t899、ST398-t034和t11241,其中ST9-t899为MRSA菌株唯一克隆型和ST398-t571为MSSA优势克隆型。除4株分离株未检测到SCCmec分型外,IV_b（76.5%,13/17）是MRSA分离株的唯一分型。[结论] 结果表明,我国猪源MRSA分离株对苯唑西林药物敏感性发生了改变,出现了较多的苯唑西林敏感菌株。此外,MSSA和MRSA分离株优势克隆型分别为ST398-t571和ST9-IVb-t899。研究还发现,克隆型与毒素编码基因有显著相关性,携带毒素编码基因的菌株克隆型均为ST9-t899。通过了解我国猪源MSSA、OR-MRSA和OS-MRSA的流行、分子特征和耐药性,可以为我国猪源金黄色葡萄球菌的防控提供数据支持。     

Synergistic effect between dieckol from <Emphasis Type="Italic">Ecklonia stolonifera</Emphasis> and β-lactams against methicillin-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis>

We have been attempting for some time to discover a compound evidencing antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA). The dieckol isolated from Ecklonia stolonifera has been shown to exhibit antibacterial activity against methicillin-susceptible S. aureus (MSSA) and MRSA. The minimum inhibitory concentrations (MICs) of dieckol were determined in a range of 32 to 64 μg/mL against standard MSSA and MRSA strains. Furthermore, dieckol clearly reversed the high-level ampicillin and penicillin resistance of MRSA. The MICs of ampicillin against two standard strains of MRSA were dramatically reduced from 512 to 0.5 μg/mL in combination with 1/4 MIC of dieckol (16 μg/mL). The fractional inhibitory concentration (FIC) indices of ampicillin and penicillin were measured from 0.066 to 0.266 in combination with 8 or 16 μg/mL of dieckol against all tested MRSA strains, thereby suggesting that dieckol-ampicillin or dieckol-penicillin combinations exert a synergistic effect against MRSA. The results of this study indicate that dieckol, administered in combination with β-lactams, may prove effective in the treatment of MRSA infections. Our finding may also contribute to the development of an alternative phytotherapeutic anti-MRSA agent.     

一株牛源都柏林沙门氏菌的全基因组测序及毒力与耐药性分析

【背景】沙门氏菌(Salmonella spp.)是重要的人畜共患病原菌,其毒力和耐药性的不断增强引起广泛关注。【目的】了解从通辽市一犊牛死亡病例中所分离牛源都柏林沙门氏菌的毒力及耐药性情况。【方法】以病死犊牛肺脏为材料,经细菌分离纯化及16S rRNA基因测序,鉴定病原为沙门氏菌。采用动物试验、药敏试验和PCR方法对分离菌进行毒力、耐药性,以及毒力基因和耐药基因检测,并对其进行全基因组测序分析。【结果】分离菌具有较强毒力,对小鼠半数致死量为2.8×10⁶ CFU/mL。分离菌为多重耐药菌,仅对多粘菌素B和噻孢霉素敏感,对强力霉素和恩诺沙星中度敏感。检测13种沙门氏菌常见毒力基因,检出率为92.3%。对分离菌进行全基因组测序分析,该菌株为都柏林沙门氏菌,基因组大小为4 965 370 bp,GC含量为52.12%,同时携带2个质粒,大小分别为79 524 bp (pTLS-1)和45 301 bp (pTLS-2)。分离菌中共携带996个毒力基因和24个毒力岛;共携带42个耐药基因,其中4个为可水平转移基因,基因组中存在9个可移动遗传元件,包括插入序列和转座子等。【结论】分离牛源都柏林沙门氏菌菌株具有较强毒力且为多重耐药株,携带大量毒力基因及耐药基因。     

Design,synthesis and antistaphylococcal activity of marine pyrrole alkaloid derivatives

Abstract

A novel set of 16 hybrids of bromopyrrole alkaloids with aroyl hydrazone were designed, synthesized and evaluated for antibacterial and antibiofilm activities against methicillin-resistant Staphylococcus aureus (MRSA; ATCC 43866), methicillin-susceptible Staphylococcus aureus (MSSA; ATCC 35556) and Staphylococcus epidermidis (SE, S. epidermidis ATCC 35984). Of the 16 tested hybrids, 14 exhibited equal or superior antibiofilm activity against MSSA and MRSA relative to standard vancomycin. Compound 4m showed highest potency with antibiofilm activity of 0.39?µg/mL and 0.78?µg/mL against MSSA and MRSA, respectively. Thus, this compound could act as a potential lead for further development of new antistaphylococcal drugs.     

A Novel Gene,fudoh, in the SCCmec Region Suppresses the Colony Spreading Ability and Virulence of Staphylococcus aureus

Staphylococcus aureus colonies can spread on soft agar plates. We compared colony spreading of clinically isolated methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA). All MSSA strains showed colony spreading, but most MRSA strains (73%) carrying SCCmec type-II showed little colony spreading. Deletion of the entire SCCmec type-II region from these MRSA strains restored colony spreading. Introduction of a novel gene, fudoh, carried by SCCmec type-II into Newman strain suppressed colony spreading. MRSA strains with high spreading ability (27%) had no fudoh or a point-mutated fudoh that did not suppress colony spreading. The fudoh-transformed Newman strain had decreased exotoxin production and attenuated virulence in mice. Most community-acquired MRSA strains carried SCCmec type-IV, which does not include fudoh, and showed high colony spreading ability. These findings suggest that fudoh in the SCCmec type-II region suppresses colony spreading and exotoxin production, and is involved in S. aureus pathogenesis.