致病菌基因组岛的研究进展

细菌基因组岛是细菌基因组上的特定区域,和水平基因转移相关,具有一定的结构特点,常携带致病、耐药及与适应性等功能相关的基因。通过基因组岛在细菌间的移动,可以造成相关基因在细菌间的传播,在细菌生存和致病等过程中具有重要作用。目前已经可通过生物信息和分子生物学实验等方法对基因组岛进行预测和验证。通过对致病菌基因组岛的研究,可以阐释细菌致病性和耐药等重要功能的获得,对疾病进行溯源,在传染病预防控制中具有重要意义。     

细菌中基因组岛的转移与调控机制研究进展

基因水平转移可导致细菌不同种属间个体DNA的交换,从而使细菌对环境的适应性增强,是细菌进化的重要途径之一。基因组岛是基因水平转移的重要载体,可移动的基因组岛能够整合到宿主的染色体上,并在特定的条件下切除,进而通过转化、接合或转导等方式转移到新的宿主中。基因组岛具有多种生物学功能,如抗生素抗性、致病性、异源物质降解、重金属抗性等。基因组岛的转移造成可变基因在不同种属细菌间的广泛传播,例如毒力和耐药基因的传播导致了多重耐药细菌的产生,威胁人类健康。基因组岛由整合酶介导转移,同时在转移的过程受到多种不同转录因子的调控。本文对细菌中基因组岛的结构特点、转移和调控机制以及预测等方面进行了综述,并最终阐明基因组岛的转移及其调控机制是遏制基因组岛传播的重要策略。     

细菌中整合性接合元件的研究进展

整合性接合元件是近年来在细菌中发现的一种可移动的基因元件,它位于染色体上,可通过接合转移的方式介导细菌间基因的水平转移。这种基因的水平转移有助于细菌适应特定的环境条件,但许多整合性接合元件包含耐药基因,这些遗传元件的水平转移极大地加速了耐药基因在同种及不同种属之间的传播,造成细菌的耐药以至多重耐药问题日益严重,耐药机制日趋复杂;同时整合性接合元件与基因岛有着密切的联系,因此对其特征及转移机制进行研究很有必要。     

利用CRISPR-Cas系统防控细菌耐药性的研究进展

细菌多重耐药是医药健康、农林牧渔、生态环境等多领域共同面临的全球性挑战.抗生素耐药基因跨物种跨区域传播是导致细菌多重耐药形成的重要原因.然而,目前尚无有效方案解决日益严峻的细菌多重耐药问题.由规律成簇间隔短回文重复序列和与之相关的蛋白组成的CRISPR-Cas系统,可靶向切割进入细菌的外源核酸,具有防控耐药基因转移导致...     

病原菌毒力岛研究进展

毒力岛作为基因组岛的一种亚类,是细菌染色体上具有特定结构和功能特征的可移动基因大片段,经基因水平转移（转导、接合或转化）获得,可使细菌基因组进化在短期内发生“量的飞跃”,直接或间接增强细菌的生态适应性,与病原菌的致病性密切相关。毒力岛存在于多种动植物病原细菌中,对于细菌的毒力变异、遗传进化甚至新病原亚种形成有重要意义。简要综述了病原菌毒力岛的研究进展,介绍了毒力岛的结构、功能特征及其在病原菌进化中作用。     

产超广谱β-内酰胺酶细菌中I类整合子的研究进展

产超广谱β-内酰胺酶(Extended-spectrum beta-lactamase, ESBLs)细菌的多重耐药性是临床用药的一大难题, 近年研究发现其耐药性的产生与整合子密切相关, 其中临床最常见、研究最深入的是I类整合子。整合子是一种可移动基因元件, 在整合酶的作用下捕捉外源基因盒并使之表达, 是具有基因整合和切除功能的天然克隆和表达系统。研究表明I类整合子可连续捕捉和整合多种耐药基因, 以质粒或转座子为载体在细菌之间传播耐药性, 使ESBLs细菌多重耐药趋势十分严峻。本文就I类整合子的结构特征、I类整合子对耐药基因盒的整合作用及其与ESBLs细菌耐药性的关系等方面进行综述。     

质粒介导的喹诺酮耐药基因qnr的分类、耐药机制及其在国内的流行状况北大核心CSCD

喹诺酮类抗菌药物从早期主要用于治疗尿道感染发展到后来治疗肠道感染和呼吸道感染,目前已在临床、畜牧业和水产业中广泛使用,细菌对其耐药性也逐渐呈蔓延趋势,耐药机制日趋复杂。喹诺酮类耐药机制主要分为染色体介导的耐药和质粒介导的耐药,后者对细菌耐药性的广泛传播起着重要作用。1998年首次报道了质粒介导的喹诺酮类耐药机制,即质粒上qnr基因介导的细菌对氟喹诺酮耐药机制,qnr基因可在不同细菌中迅速水平传播,引发的感染不易控制,使得院内感染大范围的流行。此外,qnr基因通常与β-内酰胺类耐药基因相关或存在于复杂整合子中与其它多重耐药基因共同整合,缩小了临床医生治疗相关细菌感染时选药或联合用药的空间,给我们带来了严峻的挑战。本文就qnr基因的发现历史、耐药机理及在国内的流行状况做了详细概述。     

细菌中介导多重耐药的Tn7转座子研究进展

转座子是介导细菌耐药性传播的重要可移动遗传元件。Tn7转座子与细菌耐药密切相关,其携带转座模块和Ⅱ类整合子系统。Tn7编码转座相关蛋白TnsABCDE进行“剪切-粘贴”机制转座,转座核心TnsABC也可与三链DNA或Cas-RNA复合物结合实现转座。近年来新发现了多种介导多重耐药的Tn7转座子,其在介导细菌抗生素、消毒剂和重金属抗性基因的获得、传播扩散等方面发挥了重要作用。本文综述了细菌中Tn7转座子的遗传结构、转座机制、流行以及新发现的介导多重耐药的Tn7转座子,以期为细菌中Tn7转座子的深入研究提供参考。     

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。     

细菌自然转化的机制及影响因素

水平基因转移对耐药基因传播、编码毒素基因质粒的扩散和毒力岛的转移等过程具有重要的生物学意义。自然转化是指具有感受态的细菌从外界摄取并整合裸露DNA,是水平基因转移的方式之一。细菌发生自然转化极大地促进了耐药基因在不同细菌间的播散,导致细菌对抗生素耐药,给临床治疗带来极大的困难。许多细菌具备自然转化能力,但不同细菌自然转化过程存在着差异。细菌自然感受及转化的诱发及效率亦受到多种因素的影响。文中着重于阐述不同细菌的自然转化机制及其影响因素。     

Proteomic analysis of Proteus mirabilis outer membrane proteins reveals differential expression in vivo vs. in vitro conditions

Proteus mirabilis is an opportunistic pathogen that frequently causes complicated urinary tract infections. Among a wide spectrum of potential virulence factors, outer membrane proteins (OMPs) are critical for bacterial interactions and survival in different environments. In this work, we used a proteomic approach to assess P. mirabilis in vivo OMPs expression compared to in vitro, including iron replete and iron-restricted conditions. Three putative iron receptors, IreA, PMI0842, and PMI2596, were detected both in bacterium grown in vivo and in vitro under iron-restricted conditions. A prophage gene product, PMI1721, was detected only on in vivo growing bacterium, suggesting a potential role yet to be disclosed on the surface of P. mirabilis. Plasminogen, a host protein, was co-purified with OMPs of in vivo grown bacteria, which is in accordance with previous observations and suggests that plasminogen bound to P. mirabilis surface may be associated to virulence as seen in other bacterial pathogens. Western blots using sera of experimentally challenged mice showed that iron-regulated proteins are expressed and highly immunogenic during infection. This work confirms observations made by others for P. mirabilis and reveals details not yet described, suggesting new aspects of the bacterium pathogenesis that remain unknown.     

A Comparative Categorization of Protein Function Encoded in Bacterial or Archeal Genomic Islands

Genomes of prokaryotes harbor genomic islands (GIs), which are frequently acquired via horizontal gene transfer (HGT). Here I present an analysis of GIs with respect to gene-encoded functions. GIs were identified by statistical analysis of codon usage and clustering. Genes classified as putatively alien (pA) or putatively native (pN) were categorized according to the COG database. Among pA and pN genes, the distribution of COG functions and classes were studied for different groupings of prokaryotes. Groups were formed according to taxonomical relation or habitats. In all groups, genes related to class L (replication, recombination, and repair) were statistically significantly overrepresented in GIs. GIs of bacteria and archaea showed a distinct pattern of preferences. In archeal GIs, genes belonging to COG class M (cell wall/membrane/envelope biogenesis) or Q (secondary metabolites biosynthesis, transport, and catabolism) were more frequent. In bacterial GIs, genes of classes U (intracellular trafficking, secretion, and vesicular transport), N (cell motility), and V (defense mechanisms) were predominant. Underrepresentation was strongest for genes belonging to class J (translation, ribosomal structure, and biogenesis). Among single COG functions overrepresented in GIs were transferases and transporters. In both superkingdoms, HGT enhances genomic content by meeting demands that are independent of the studied habitats. These findings are in agreement with the complexity theory, which predicts the preferential import of operational genes. However, only specific subsets of operational genes were enriched in GIs. Modification of the cell envelope, cell motility, secretion, and protection of cellular DNA are major issues in HGT. [Reviewing Editor: Dr. Siv Andersson]     

革兰氏阴性菌AcrAB-TolC多药外排泵结构数据对抑制剂研发的启示

革兰氏阴性菌的多重耐药性已成为全球广泛聚焦的问题。近年研究发现,耐药结节细胞分化(resistance-nodulation-cell division,RND)家族外排泵的过表达,与革兰氏阴性菌的多重耐药性密切相关。在RND家族中,广泛存在于革兰氏阴性菌中的AcrAB-TolC外排泵被认为是导致多重耐药性的主要原因之一。为了开发有效的抑制剂,需要对AcrAB-TolC外排泵的结构有一个清晰的认识。以往对该外排泵结构的研究主要局限于体外采用X射线晶体学技术或冷冻电镜单颗粒分析技术来解析其单个组分或全泵的结构。细胞冷冻电子断层扫描技术为揭示AcrAB-TolC外排泵在天然细胞膜环境中的组装和运行机制提供了新的见解,本文综述了AcrAB-TolC不同层级的结构数据在研发外排泵抑制剂方面的贡献。     

A Novel Method to Predict Genomic Islands Based on Mean Shift Clustering Algorithm

Genomic Islands (GIs) are regions of bacterial genomes that are acquired from other organisms by the phenomenon of horizontal transfer. These regions are often responsible for many important acquired adaptations of the bacteria, with great impact on their evolution and behavior. Nevertheless, these adaptations are usually associated with pathogenicity, antibiotic resistance, degradation and metabolism. Identification of such regions is of medical and industrial interest. For this reason, different approaches for genomic islands prediction have been proposed. However, none of them are capable of predicting precisely the complete repertory of GIs in a genome. The difficulties arise due to the changes in performance of different algorithms in the face of the variety of nucleotide distribution in different species. In this paper, we present a novel method to predict GIs that is built upon mean shift clustering algorithm. It does not require any information regarding the number of clusters, and the bandwidth parameter is automatically calculated based on a heuristic approach. The method was implemented in a new user-friendly tool named MSGIP—Mean Shift Genomic Island Predictor. Genomes of bacteria with GIs discussed in other papers were used to evaluate the proposed method. The application of this tool revealed the same GIs predicted by other methods and also different novel unpredicted islands. A detailed investigation of the different features related to typical GI elements inserted in these new regions confirmed its effectiveness. Stand-alone and user-friendly versions for this new methodology are available at http://msgip.integrativebioinformatics.me.     

Dissemination and loss of a biofilm‐related genomic island in marine Pseudoalteromonas mediated by integrative and conjugative elements

Acquisition of genomic islands (GIs) plays a central role in the diversification and adaptation of bacteria. Some GIs can be mobilized in trans by integrative and conjugative elements (ICEs) or conjugative plasmids if the GIs carry specific transfer‐related sequences. However, the transfer mechanism of GIs lacking such elements remains largely unexplored. Here, we investigated the transmissibility of a GI found in a coral‐associated marine bacterium. This GI does not carry genes with transfer functions, but it carries four genes required for robust biofilm formation. Notably, this GI is inserted in the integration site for SXT/R391 ICEs. We demonstrated that acquisition of an SXT/R391 ICE results in either a tandem GI/ICE arrangement or the complete displacement of the GI. The GI displacement by the ICE greatly reduces biofilm formation. In contrast, the tandem integration of the ICE with the GI in cis allows the GI to hijack the transfer machinery of the ICE to excise, transfer and re‐integrate into a new host. Collectively, our findings reveal that the integration of an ICE into a GI integration site enables rapid genome dynamics and a new mechanism by which SXT/R391 ICEs can augment genome plasticity.     

一株奇异变形杆菌对铜绿假单胞菌多细胞行为的抑制作用

目的：抗生素耐药性成为了全球性的健康问题。研究发现病原菌的多细胞行为在抗生素的耐药性中起着至关重要的作用（尤其是生物膜）,因而通过抑制多细胞行为而控制耐药性成为当务之急。本文以奇异变形杆菌（Proteus mirabilis）为研究对象,考察它的发酵滤液对一种机会致病菌——铜绿假单胞菌（Pseudomonas aeruginosa）多细胞行为的作用,以期得到一株多细胞行为抑制菌：在不影响Paerugiliosa生长的前提下,抑制生物膜形成、EPS产生以及定向丛集运动,解除保护,减缓扩散,为降低Paemgi—nosa耐药性,增强抗生素作用效果提供可能。方法：采用结晶紫生物膜测定法、蒽酮一硫酸法、平板检测法,探究Pmirabilis发酵滤液对Paemginosa生物膜、胞外多聚物、定向丛集运动和生长的影响。结果：Pmirabilis发酵滤液能显著抑制Paeruginosa生物膜量,在体积百分比浓度为1％时,抑制率可达60．9％。该菌的发酵滤液还能阻碍Paeruginosa的定向丛集运动,减弱它的吸附和扩散运动;同时,也减少了Pacrugillosa胞外多聚物的产量,在滤液体积百分比浓度为1％时,抑制率达到45．9％。更重要的是,固体平板实验证明该发酵滤液对P．aemginosa的生长没有影响。结论：Pmirabilis在不影响病原菌生长的前提下,对病原菌的多细胞行为有一定的控制作用。其发酵滤液中存在着抑制微生物膜、定向丛集运动等的成分,在治疗细菌感染性疾病和降低抗生素耐药性方面有潜在应用价值。     

克雷伯氏菌属(Klebsiella)细菌比较基因组学分析揭示多复制子抗性质粒介导广泛的抗性基因传播

[目的] 研究克雷伯氏菌与多复制子抗性质粒间的关系,分析细菌携带多复制子质粒对抗生素环境的响应机制。[方法] 以2018-2020年分离的56株不同来源克雷伯氏菌（Klebsiella sp.）分离株为研究对象,利用微量肉汤稀释法评估其多重耐药表型,对分离菌株进行全基因组测序（WGS）,通过细菌全基因组关联分析（BGWAS）技术和比较基因组学方法深入解析多复制子抗性质粒形成的机制。[结果] 耐药表型分析发现野生动物来源的菌株具有更广的耐药谱系,总体Klebsiella sp.对氨苄西林表现出很高的耐药率（80.36%）,尤其是马来穿山甲来源菌株对头孢类抗生素高度耐受,同时对氯霉素、左氧氟沙星和复方新诺明等药物耐受,基因组分析发现这些菌株携带了抗性质粒和更多的抗生素抗性基因。进一步对69个质粒序列分析,发现有28个质粒为多复制子质粒,主要携带bla_CTX-M-15、bla_CTX-M-14、bla_CTX-M-55、bla_OXA-1和bla_TEM-1等β-内酰胺酶基因。细菌携带质粒类型分析认为Klebsiella pneumoniae可能是多复制子质粒的重要宿主,质粒骨架与结构分析发现多复制子质粒多由2个或2个以上单个质粒融合而成,携带此类质粒的菌株不仅获得了更广的耐药表型,而且在全球传播扩散分布逐年增加,因此产生对抗生素环境更强的适应性。[结论] 多重耐药性细菌呈现的表型与携带的多复制子质粒有关,相比较下多复制子质粒比非多复制子质粒有更强的抗性基因携带能力,或许是细菌在强大的抗生素压力下产生的重要响应机制。本研究对于未来探索细菌抗性基因的传播扩散机制具有重要意义。     

人工哺育期腹泻婴猴奇异变形杆菌的分离与鉴定

目的对一株人工哺育期引发恒河猴婴猴腹泻的奇异变形杆菌进行了鉴定,为实验猕猴疾病检测、鉴别诊断提供参考依据。方法通过培养特性、菌落形态、染色、生化试验和血清学诊断鉴别等检查,对分离菌株进行初步鉴定,同时,对分离菌株进行致病性试验及药敏试验。结果通过表型生物学特性鉴定,并结合血清学诊断鉴别方法,确证该分离菌株为奇异变形杆菌,应用药敏试验筛选出了高度敏感的抗菌药,控制了该病的继续发生,致病性试验证明,该分离菌株对小白鼠有高致病性。结论分离到的奇异变形杆菌是导致本次婴猴腹泻死亡的病原菌,该菌为条件致病菌,对实验猕猴和研究人员均有潜在的危害,尽管该菌不是国家标准要求排除的病原菌,但该菌引发的传染病将对动物实验造成严重影响,故应引起高度重视。     

GI-POP: A combinational annotation and genomic island prediction pipeline for ongoing microbial genome projects

Sequencing of microbial genomes is important because of microbial-carrying antibiotic and pathogenetic activities. However, even with the help of new assembling software, finishing a whole genome is a time-consuming task. In most bacteria, pathogenetic or antibiotic genes are carried in genomic islands. Therefore, a quick genomic island (GI) prediction method is useful for ongoing sequencing genomes. In this work, we built a Web server called GI-POP (http://gipop.life.nthu.edu.tw) which integrates a sequence assembling tool, a functional annotation pipeline, and a high-performance GI predicting module, in a support vector machine (SVM)-based method called genomic island genomic profile scanning (GI-GPS). The draft genomes of the ongoing genome projects in contigs or scaffolds can be submitted to our Web server, and it provides the functional annotation and highly probable GI-predicting results. GI-POP is a comprehensive annotation Web server designed for ongoing genome project analysis. Researchers can perform annotation and obtain pre-analytic information include possible GIs, coding/non-coding sequences and functional analysis from their draft genomes. This pre-analytic system can provide useful information for finishing a genome sequencing project.