一株猪源化脓隐秘杆菌的鉴定、生物学特性研究与基因组分析

[背景] 化脓隐秘杆菌作为一种条件致病菌,常与蓝耳病病毒、猪圆环病病毒、巴氏杆菌等混合感染猪,引起各种非特异性化脓性感染,部分临床症状与副猪嗜血杆菌相似。本试验从江西某猪场一起因呼吸道症状急性死亡病猪的肺脏分离到一株具有β溶血特性、疑似化脓隐秘杆菌的细菌病原。[目的] 鉴定该病原的菌属种类、生物学特性及基因组特征,为该菌疾病的基础研究与临床防控奠定理论基础。[方法] 采用透射电镜、扫描电镜结合PCR鉴定等方法对菌属种类进行鉴定;通过动物实验、药敏试验,分析菌株的生物学特性;借助全基因组测序、生物信息学等技术发掘该菌的基因组特征。[结果] 形态观察、16S rRNA基因鉴定及系统发育分析证实,该分离菌株是化脓隐秘杆菌,命名为JX18;药敏试验表明,该菌株对克林霉素和林可霉素不敏感,对其余所选药物均敏感;动物实验结果显示,对小鼠腹腔攻毒后,小鼠腹部出现明显脓肿病灶,最高剂量组的小鼠全部死亡;全基因组测序结果表明,该菌株携带plo、nanH、nanP、fimA、fimC、fimE等重要毒力基因,并且与其他化脓隐秘杆菌菌株毒力基因的相似性较高;根据同源基因数据库（Cluster of Orthologous Groups,COG）预测,菌株JX18中参与碳水化合物代谢的基因占比最高;通过毒力基因数据库（virulence factor database,VFDB）、UniProtKB/Swiss-Prot等数据库预测,JX18菌株基因组中携带有多个组氨酸激酶、反应调节因子等与细菌双组分调控系统相关基因,以及多种与粘附、侵入及分泌系统相关的毒力基因。[结论] 在江西省分离鉴定出一株猪源强致病性化脓隐秘杆菌,丰富了猪源化脓隐秘杆菌病原信息,为进一步开展猪源化脓隐秘杆菌相关研究与防控奠定了理论依据。     

致病性大肠杆菌和蜡样芽胞杆菌的分离鉴定

【背景】猪只消化道疾病是养猪业上一大重要疾病,给养猪业带来一定的经济损失。大肠杆菌是引起猪腹泻的一种常见病原菌,可以引起不同日龄的猪腹泻,但主要以幼龄猪为主。【目的】旨在分离鉴定引起四川省眉山市一规模化养猪场病猪大规模腹泻的病原菌。【方法】采用常规细菌分离方法结合16S rRNA基因序列的分析方法从发病猪肝脏、胃以及污染的饲料分离鉴定细菌,并对分离株进行小鼠致病性试验、16S rRNA基因遗传进化树分析、毒力基因的检测、药物敏感试验。【结果】从腹泻猪肝脏中分离到一株致病性大肠杆菌,胃中分离到一株蜡样芽胞杆菌,并且追溯到传染源是该猪场饲料。通过检测这两株菌相应的毒力基因发现大肠杆菌不属于肠外致病性型,蜡样芽胞杆菌检测到了nheA、nheB、nheC、bceT、entFM 5种毒力基因;药敏试验表明常规的氨基糖苷类和头孢类抗生素对大肠杆菌抑菌效果较好,红霉素、氟苯尼考、头孢氨苄、头孢哌酮对蜡样芽胞杆菌抑菌效果较好,而蜡样芽胞杆菌对青霉素、阿莫西林等常规药物不敏感。【结论】饲料存在大肠杆菌和蜡样芽胞杆菌的混合污染。     

炭疽杆菌毒力基因及其表达调控的研究进展

炭疽杆菌是炭疽病的病原体,致病性主要与细菌合成毒素和英膜形成能力有关。炭疽杆菌的毒力基因表达调控涉及细菌质粒和染色体成分,细菌生长环境对毒力基因表达也有影响。     

一株食源性蜡样芽孢杆菌的分离及其对小鼠肠黏膜免疫相关因子和肠道菌群的影响

蜡样芽孢杆菌(Bacillus cereus)是一种常见的食源性致病菌,误食被其污染的食品会引起呕吐或者腹泻,严重者还会死亡。本研究利用划线培养法从腐败米饭中分离培养出一株蜡样芽孢杆菌。通过药敏试验分析及毒力相关基因PCR扩增对其致病性及耐药性进行分析,并将细菌培养物经过腹腔注射进行小鼠感染试验,检测其对小鼠肠黏膜免疫相关因子和肠道菌群的影响,从而分析该病原菌的致病途径及防治手段。结果显示,本研究所分离的食源性蜡样芽孢杆菌,对诺氟沙星、呋喃妥因、四环素、米诺环素、环丙沙星、大观霉素、克林霉素、红霉素、克拉霉素、氯霉素、左氟沙星和万古霉素敏感,对复方新诺明、苯唑西林和青霉素G具有耐受性。该菌株携带hblA、hblC、hblD、nheA、nheB、nheC和entFM 7种毒力相关基因,可产生致腹泻型毒素。小鼠感染后,出现腹泻,肠黏膜免疫球蛋白和炎性因子表达水平显著上调;肠道菌群检测结果表明,经该蜡样芽孢杆菌感染后,小鼠肠道菌群组成发生变化,标志着机体健康的拟杆菌门(Bacteroidetes)中的uncultured_bacterium_f_Muribaculaceae丰度显著下降,而标志着菌群失调的变形菌门(Proteobacteria)条件致病菌uncultured_bacterium_f_Enterobacteriaceae丰度显著升高,且与IgM和IgG呈显著正相关(P<0.05)。这些研究结果表明,携带致腹泻型毒力相关基因的致病性蜡样芽孢杆菌感染机体后,可以通过影响肠道菌群激活免疫系统。     

紫色色杆菌感染临床特症及耐药谱分析

目的 研究紫色色杆菌感染的临床特征与病原体的生物学特性以及耐药分析,以提高临床对紫色色杆菌感染的诊断和治疗.方法 将患者的血液、浓液标本进行培养,分离并把分离获得的菌株在VITEK-32微生物鉴定仪中进行种的鉴定及药物敏感性试验,用小白鼠进行该病原体的毒力试验.结果 12种抗菌药物中舒普深、亚胺培南、特治星（派拉西林/他唑巴坦）、马斯平（头孢吡肟）及喹喏酮类抗菌药物对紫色色杆菌均有较强的抗菌作用,而氨苄西林、头孢唑啉、庆大霉素、妥布霉素、氨曲南、头孢曲松和头孢他啶等抗菌药物抗菌效果不佳.毒力试验结果表明该菌有较强毒力.结论 紫色色杆菌是一种革兰阴性杆菌,毒力强.可引起感染部位脓肿,并可侵入体内各脏器及血液引发脏器的脓肿,脓毒血症是该菌感染的临床特症,重者可引起各脏器功能衰退,中毒性休克甚至死亡.紫色色杆菌感染病情急,进展快,死亡率高,因此早发现,及早有效性治疗极为重要.     

猪链球菌毒力因子和鉴定方法的研究进展

猪链球菌(Streptococcus suis)是一种重要的人畜共患病病原体,能够引起猪疫病,人类感染该菌可导致脑膜炎、败血症甚至死亡。鉴于其对养猪业的巨大经济影响和对公共卫生事业的威胁,关于猪链球菌的研究日益深入,即对其毒力相关蛋白和鉴定方法的研究进展做一综述。     

致病性弧菌主要毒力相关因子的研究进展

弧菌(Vibrio)是一种革兰氏阴性短杆菌,其广泛分布于自然界,以水中最多.弧菌是引起人类、水生动物细菌性疾病的重要病原之一.弧菌的致病性与各个毒力基因的协同作用密切相关.弧菌的毒力因子主要可分为粘附素、内毒素、外毒素等.本文主要对致病性较强的毒力因子粘附素和外毒素进行阐述,以期为弧菌致病机理的研究及弧菌病的防治提供参考和借鉴.     

2016年辽宁省婴幼儿配方食品及 谷类辅助食品中蜡样芽胞杆菌毒力基因的鉴定

摘要：目的 了解婴幼儿配方食品和谷类辅助食品中蜡样芽胞杆菌的毒力基因携带特点,对辽宁省婴幼儿配方食品和谷类辅助食品中蜡样芽胞杆菌的污染状况进行调查。方法 依据GB4789.14?2014《食品安全国家标准 食品微生物学检验 蜡样芽胞杆菌检验》及采用PCR扩增技术和血平板检测的方法对2016年采自辽宁省15个监测点,收集的176份乳源性食品中检出的22株蜡样芽胞杆菌进行10种毒力基因检测。结果 婴幼儿配方食品和谷类辅助食品蜡样芽胞杆菌检出率为12.5%（22/176）,非溶血性的肠毒素Nhe基因、溶血素BL基因、肠毒素T基因和细胞毒素K基因是辽宁省乳源性蜡样芽胞杆菌的主要毒力基因,至少携带2种毒力基因的菌株达到检出菌总数的100.0%。结论 研究结果证实辽宁省婴幼儿配方食品及谷类辅助食品存在蜡样芽胞杆菌污染情况,严格监控婴幼儿配方食品及谷类辅助食品的蜡样芽胞杆菌污染,对于生产出优质婴幼儿配方食品及谷类辅助食品具有重要意义,以期提高婴幼儿食品的质量安全。     

非洲猪瘟病毒的非必需基因：真的可有可无吗？

非洲猪瘟（African swine fever,ASF）是由非洲猪瘟病毒（African swine fever virus,ASFV）引起的一种出血性、致死性的猪烈性传染病。ASF在全球广泛传播,给养猪业造成重大的经济损失。ASFV基因组庞大,可编码150多种蛋白,一些非必需基因编码的蛋白与调控病毒毒力、复制和免疫逃逸等相关。通过删除ASFV毒力相关的非必需基因所构建的减毒株是当前比较有前景的疫苗,然而其安全性有待提高。系统地鉴定ASFV非必需基因及其功能,不仅有助于ASF基因缺失疫苗的研发,也有益于ASFV致病机制研究。本文对目前已鉴定的ASFV非必需基因及其功能研究进行了总结分析,着重讨论了影响ASFV毒力、调控病毒复制、参与免疫逃逸的非必需基因及其编码蛋白的功能,旨在加深对ASFV病原学的认识,为新的ASFV非必需基因的鉴定和功能研究提供参考。     

鼠疫菌毒力调控研究

鼠疫菌通过一系列转录调控子(如CRP、PhoP、RovA和Fur)控制着一些关键毒力因子(如Pla、强毒力岛、III型分泌系统等)的基因表达。鼠疫菌可感应宿主体内信号刺激,紧密调控毒力因子的表达。在这个紧密调控过程中,调控子、毒力相关基因构成了一个动态网络。鼠疫菌在从假结核菌祖先演化的进程中,基因表达调控网络的重塑在鼠疫菌毒力进化过程中发挥着不可取代的作用。     

Clostridioides difficile from Brazilian hospitals: characterization of virulence genes by whole genome sequencing

Clostridioides difficile (CD) is the most frequent cause of healthcare related diarrhea and its severity has increased in the last decade by the spread of hypervirulent strains. Most important CD virulence factor is toxin production; however, not only toxins are responsible for Clostridioides virulence. We sequenced 38 strains and analyzed the presence and integrity of 24 virulence (including toxin) genes. We identified 28 toxigenic strains, six also presented the cdt genes. Only six strains didn't present all others genes searched. All absent genes were adhesion related. Understand others CD virulence factors can lead to a best understanding on this matter.     

Immune defence,extra-pair paternity,and sexual selection in birds

Secondary sexual characters have been suggested to reliably reflect the ability of individuals to resist debilitating parasites, and females may gain direct or indirect fitness benefits from preferring the most extravagantly ornamented males. Extra-pair paternity provides an estimate of an important component of sexual selection in birds. Species with a high frequency of extra-pair paternity have a variance in realized reproductive success that is greater than the variance in apparent reproductive success, and extra-pair copulations and hence extra-pair paternity by females are often directly associated with the expression of male secondary sexual characters. If sexually dichromatic species have experienced a long period of antagonistic coevolution with their parasites, such species should have evolved larger immune defence organs than sexually monochromatic species. Bird species with sexual dichromatism had larger spleens for their body size than monochromatic species in a comparative analysis. Furthermore, species with a high frequency of extra-pair paternity were sexually dichromatic and had large spleens for their body size. These results are consistent with the hypothesis that females of dichromatic bird species seek extra-pair copulations to obtain indirect fitness benefits in terms of superior resistance of their offspring to virulent parasites.     

Iron depletion and virulence in Staphylococcus aureus

Abstract Staphylococcus aureus is able to grow in the presence of extremely low iron concentrations (0.04 μM). In iron-limiting conditions, this species develops alternative metabolic strategies such as highly efficient iron-uptake mechanisms which are only partially shared with S. epidermidis . Here we summarize the mechanisms induced by iron starvation in S. aureus in order to elucidate the virulence characteristics of this bacterium.     

湖北地区暴发病池塘中嗜水气单胞菌的遗传多样性和毒力特征研究

为了调查引起鱼类运动型气单胞菌败血症(俗称暴发病)的嗜水气单胞菌的遗传多样性和毒力特征, 阐明其流行规律, 研究于2006-2009年度从湖北省内3个不同地区的6个发病鱼塘中分离了30株嗜水气单胞菌, 其中20株为临床株(分离自血液、肝脏、肾脏或腹水), 6株为肠道株, 4株为池水株。基于所有菌株gyrB基因序列, 构建了系统发育树; 通过ERIC (Enterobacterial repetitive intergenic consensus, 肠道细菌基因间重复序列)指纹图谱进行菌株的遗传分型; 用PCR方法检测了7个毒力基因在菌株中的分布模式。这7个基因包括气溶素(aerA)、溶血素(hlyA)、热不稳定性细胞兴奋性肠毒素(alt)、热稳定性细胞兴奋性肠毒素(ast)、弹性蛋白酶(ahpB)、脂酶(lip)和鞭毛基因(fla)。此外, 以斑马鱼为感染对象, 通过腹腔注射测定了15株代表菌株的毒力。结果表明: 不同来源的20株临床株、1株肠道株和3株池水株具有相同的遗传特性, 体现为在系统树上聚为一枝, 序列相似性为100%, 具有相同的ERIC指纹图谱, 毒力基因分布模式为: aerA+hlyA+alt+ast+ahpB+lip+fla+, 且均为强毒株(LD50 9.74104cfu/尾)。与临床株相比, 其余5株肠道株和1株池水株或具有不同的ERIC指纹图谱或具有不同的毒力基因分布模式, 显示出了遗传多样性, 且毒力均弱于临床株(LD501.01106cfu/尾)。这说明在一定时间、一定区域内, 作为暴发病病原的嗜水气单胞菌为同一克隆系在流行, 不存在明显的变异或遗传多样性。此结果有助于阐明嗜水气单胞菌引起的暴发病的流行规律, 制定相应的防御措施。多种毒力基因在致病性菌株中的联合流行为发病机理的解析奠定了基础。此外, 鉴于毒力基因谱与致病性之间的相关性, 表明毒力基因可作为标记基因, 用于致病性菌株的检测。       

Definitions of pathogenicity and virulence in invertebrate pathology

Accurate definition and usage of terminology are critical to effective communication in science. In a recently published article, the clarity and consistency of the terms pathogenicity and virulence as used in invertebrate pathology were called into question, and a revision of these terms was proposed. Our objective was to examine definitions of pathogenicity and virulence and their use in invertebrate pathology, and respond to this article. Although usage of the terms pathogenicity and virulence varies, we found considerable consistency in the published definitions of these terms in the invertebrate pathology literature throughout the history of the discipline, as well as among related disciplines such as medicine and microbiology. We did not find the established definitions to be lacking in clarity or utility. Therefore, we recommend that the definition and use of these terms adhere to precedence. Specifically, pathogenicity is the quality or state of being pathogenic, the potential ability to produce disease, whereas virulence is the disease producing power of an organism, the degree of pathogenicity within a group or species. Pathogenicity is a qualitative term, an “all-or-none” concept, whereas virulence is a term that quantifies pathogenicity.     

A single amino acid substitution beyond the C2H2-zinc finger in Ros derepresses virulence and T-DNA genes in Agrobacterium tumefaciens

Ros is a chromosomally-encoded repressor containing a novel C2H2 zinc finger in Agrobacterium tumefaciens. Ros regulates the expression of six virulence genes and an oncogene on the Ti plasmid. Constitutive expression of these genes occurs in the spontaneous mutant 4011R derived from the octopine strain Ach-5, resulting in T-DNA processing in the absence of induction, and in the biosynthesis of cytokinin. Interestingly, the mutation in 4011R is an Arg to Cys conversion at amino acid residue 125 near the C-terminus well outside the zinc finger of Ros. Yet, Ros bearing this mutation is unable to bind to the Ros-box and is unable to complement other ros mutants.