大肠杆菌变种的两种分子生物学检测方法分析

目的:对大肠杆菌的一种重要的变种--肠出血性大肠杆菌O157-H7的几种检测方法进行比较研究.方法:以自动免疫磁珠收集系统(AIMS)、自动酶标免疫测试系统(VIDAS)与传统常规的分离方法进行对比分析.结果:运用自动免疫磁珠收集系统(AIMS)方法对80份可能含有肠出血性大肠杆菌O157-H7的实样进行检测,检出份数为6份,检出率为7.5％,而且在一周之内可以全部对上述检出实样进行鉴定.AIMS法能够检出浓度在10cfu/mL模拟实样之中的肠出血性大肠杆菌O157-H7,然后将此法与CHROMagar 0157琼脂板相结合,其效果则更为明显.而自动酶标免疫测试系统(VIDAS)与传统与的分离方法则检测的效果不佳,检出率为0.自动免疫磁珠收集系统(AIMS)检测方法与自动酶标免疫测试系统(VIDAS)、传统与的分离方法在检出率方面存在显著的统计学差异,P＜0.01.结论:运用自动免疫磁珠收集系统(AIMS)结合CHROMagar 0157琼脂板对出血性大肠杆菌O157-H7进行检测,检出率较高、灵敏度较高且快速便捷,可以用于O157-H7外环境检测与食品污染源的实际调查之中,应该对其加以广泛地推广并应用.     

Transduction of resistance to cyanide

Путем пассажей лизогенного штамма Escherichia coli K 12 на средах, содержавших повышающиеся кондентрадии дианистого калия, удалось вырастить варианты, резистентные к 0,4 мол. Кондент KCN. Полученные резистентые штаммы сохраняют свою резистентность и при пассажах на средах без дианистого калия. С приобретением устойчивости к дианистому калиию у этих штаммов изменились и другие свойства в сравнении с исходным штаммом. Некоторые изменения оказались обратимыми, другие, — как биохимические изменения, изменения чувствительности к ?агам и в продукции ?ага, — сохранялись стабильно и после 50 пассажей на средах без цианистого калия. С помощью ?ага, выделенного из штамма К 12, устойчивого к 0,4 мол. концентрации цианистого калия, удалась трансдукция не только устойчивости к цианистому калию, но одновременно и чувствительности к ?агам группы coli Т и утраты способности сбраживать дульцит. Однако связь между этими особенностями не была стопроцентной, так как перенесенная устойчивость к дианистому калию оказалась несколько более низкой, чем исходная (0,3 мол. Вместо 0,4 мол.), а бактерии после трансдукдии продуцировали не только ?аг α, как бактериидоноры, но и преимущественно первоначальный ?аг.     

Localization of proteases in cells ofEscherichia coli andBacillus megaterium

1. (1)
   Изучалася локализация протеаз в клетках Bacillus megaterium и Escherichia coli. B. megaterium—это типичный протеолитический микроорганизм, тогда как E. coli вообще не выделяет протеазы в среду.     
   
   

The influence of rifampicin resistant mutations on the biosynthesis of exopolysaccharides by strain Escherichia coli K-12 lon

Applied Biochemistry and Microbiology - The influence of RNA polymerase (rif) mutations on the yield of capsular exopolysaccharide— colanic acid (CA) of Escherichia coli K-12 lon strain was...     

加压CO2对大肠杆菌细胞膜的损伤作用

[目的]细菌细胞膜的损伤可以表现在细菌细胞内物质泄漏和细菌细胞吸收染料.与巴氏杀菌(63℃C、30 min)比较,研究加压CO2对大肠杆菌细胞膜的损伤作用,目的是分析出大肠杆菌死亡与细胞膜损伤的关系.[方法]检测大肠杆菌细胞膜通透性的改变情况,大肠杆菌内蛋白质和核酸的泄漏程度,并通过透射电镜观察大肠杆菌形态的改变情况.[结果]在研究范围内,加压CO2处理使大肠杆菌细胞膜通透性发生改变;加压CO2处理时虽然发生了胞内蛋白质泄漏,但发生泄漏的时间明显滞后于99％以上菌体死亡时间,因此并不是大肠杆菌死亡的原因,只是大肠杆菌死亡后的继发现象;大肠杆菌死亡与加压CO2处理导致的胞内核酸泄漏有关;大肠杆菌死亡与加压CO2处理导致的菌体形态改变有关.[结论]加压CO2对大肠杆菌细胞膜的损伤作用与菌体死亡有直接关系.     

Characterization of transmissible genetic elements from sucrose-fermenting Salmonella strains.

Two of seven sucrose-fermenting Salmonella strains obtained from clinical sources were found capable of conjugal transfer of the sucrose fermentation (Scr+) property to the Escherichia coli K-12 strain WR3026. The genetic elements conferring this Scr+ property, designated scr-53 and scr-94, were then conjugally transmissible from Escherichia coli WR3026 Scr+ exconjugants to other strains of Escherichia coli at frequences of 5 times 10- minus 6 to 5 times 10- minus 3 for the scr-53 element and 10- minus 6 to 10- minus 5 for the scr-94 element. In Escherichia coli hosts, both of these elements were compatible with F-lac and with each of six previously characterized transmissible lac elements. No antibiotic resistance characteristics or colicin production were discovered to be associated with either scr-53 or scr-94. Neither scr element generated a male host response to the female-specific phage phiII, but the scr-53 element rendered its Escherichia coli host sensitive to the male-specific phage R-17. Escherichia coli hosts containing scr-53 were susceptible to lysis by P1vir, and transduction of the scr-53 element was accomplished with this phage. The scr-53 element was isolated from Escherichia coli WR3026, Scr+ transductants, and Escherichia coli WR2036 Scr+ exconjugants as a covalently closed circular deoxyribonucleic acid molecule with a molecular weight (determined by electron microscopy) of approximately 52 times 10-6. Receipt of the scr-94 element rendered Escherichia coli hosts of this element unsusceptible to lysis by P1vir, although adsorption of the phage by an Escherichia coli WR3026 exconjugant containing scr-94 occurred as efficiently as it did on WR3026 itself. Repeated examination of Escherichia coli strains harboring scr-94, as well as of the Salmonella strain which initially contained it, did not reveal the presence of circular deoxyribonucleic acid. The synthesis of the sucrose cleaving enzyme was inducible in Escherichia coli exconjugants containing either scr-53 or scr-94.     

猪源大肠杆菌耐药质粒图谱及耐药性分析

目的:探讨猪大肠杆菌的耐药质粒图谱、耐药性及耐药基因之间的关系。方法:从湖南省株洲、益阳的四个猪场分离出9株大肠杆菌,进行质粒电泳图谱分析、用PCR法检测耐喹诺酮类耐药基因Gyr A、Par C和耐四环素类耐药基因Tet A、Tet B,并采用Kirby-bauer法对这9株大肠杆菌进行药敏(18种抗生素)试验。结果:其中9株大肠杆菌含有三条或者三条以上的质粒条带,且其质粒谱型均不相同;9株大肠杆菌均检测出4种耐药基因Gyr A、Par C、Tet A和Tet B;9株大肠杆菌对所选用的抗生素存在不同程度的耐药性,其中7株大肠杆菌对10种或10种以上的抗生素耐药,最高对13种抗生素耐药,氨苄西林、青霉素、阿莫西林、红霉素的耐药率达100%,对四环素、多西环素的耐药率达到88.9%,而多粘菌素B、阿奇霉素、大观霉素耐药率较低。结论:耐药性与质粒条带数、耐药基因之间并无明显的相关性;猪大肠杆菌呈多重耐药之势,在治疗大肠杆菌病时最好根据药敏实验结果选用合适的抗生素。     

Pterostilbene production by microorganisms expressing resveratrol O-methyltransferase

Pterostilbene (3,5-dimethoxy-4′-hydroxyl-trans-stilbene)—a derivative of resveratrol—is a natural dietary compound and the primary antioxidant component in berries. Pterostilbene has significant advantages over resveratrol in bioavailability, half-life in the body, cellular uptake, oral absorption and metabolic stability. Here, we expressed the resveratrol O-methyltransferase (ROMT) gene (VvROMT) from grape (Vitis vinifera) in Escherichia coli and Saccharomyces cerevisiae and confirmed its specific ability to catalyze the production of pterostilbene from resveratrol. By co-expressing an additional two genes from the resveratrol biosynthetic pathway—4-coumarate CoA-ligase (4CL) and stilbene synthase (STS)—a large amount of pterostilbene was produced, with a trace amount of pinostilbene detected. To understand the molecular basis of the catalytic activity, four key amino acid residues were identified in a 3D-model of VvROMT and mutagenized and assayed for augmented catalytic activity. Our results demonstrate the potential utility of the engineered microorganisms for pterostilbene production and provide protein engineering targets that will hopefully lead to increased activity of the ROMT enzyme.     

大肠埃希菌耐药性及其基因同源性分析

目的 研究临床分离的大肠埃希菌对常用抗生索的耐药性及其基因分型,了解其耐药性趋势与传播流行情况,为临床合理治疗大肠埃希菌引起的感染提供参考依据。方法 采用常规鉴定技术鉴定细菌;采用K—B纸片扩散法测定77株大肠埃希菌对19种药物的耐药性;K—B法鉴定产超广谱β-内酰胺酶（ESBLs）;通过脉冲场凝胶电泳（PFGE）法对其进行基因分型以确定菌株之间的亲缘关系;FINGERPRINT Ⅱ软件进行细菌基因指纹图谱分析。结果 大肠埃希菌对青霉素类、喹诺酮类药物和氨曲南的耐药性明显增高,亚胺培南和美罗培南是大肠埃希菌感染患者的首选药物;经ESBLs确证试验,ESBLs阳性率为28．60％（22／77）;产ESBLs大肠埃希菌经PFGE指纹图谱分析,除第62株和第70株相似性系数为78．27％外,其余相似度均低于70．0％;ESBLs大肠埃希菌阴性株中除少数几对菌株相似性系数较高外,其余呈散在分布,且电泳带存有6条以上的不同条带,为流行病学无关的不同克隆。结论 大肠埃希菌对常用抗生索耐药性明显增高,且呈多重耐药趋势;该研究尚不能证明存在大肠埃希菌爆发性流行感染,提示可能存在院内感染大肠埃希菌的优势克隆;PFGE基因分型方法是耐药性与流行状况分析的有效手段。     

Comparative biochemical characterization of two GroEL homologs from the cyanobacterium Synechococcus elongatus PCC 7942

Unlike Escherichia coli, cyanobacteria generally contain two GroEL homologs. The chaperone function of cyanobacterial GroELs was examined in vitro for the first time with GroEL1 and GroEL2 of Synechococcus elongatus PCC 7942. Both GroELs prevented aggregation of heat-denatured proteins. The ATPase activity of GroEL1 was approximately one-sixth that of Escherichia coli GroEL, while that of GroEL2 was insignificant. The activities of both GroELs were enhanced by GroES, while that of Escherichia coli GroEL was suppressed. The ATPase activity of GroEL1 was greatly enhanced in the presence of GroEL2, but the folding activities of GroEL1 and GroEL2 were much lower than that of Escherichia coli GroEL, regardless of the co-presence of the counterpart or GroES. Both native and recombinant GroEL1 forms a tetradecamer like Escherichia coli GroEL, while GroEL2 forms a heptamer or dimer, but the GroEL1 and GroEL2 oligomers were extremely unstable. In sum, we concluded that the cyanobacterial GroELs are mutually distinct and different from Escherichia coli GroEL.     

Expression of Erwinia chrysanthemi ENA49 uvr gene in Escherichia coli K12 cells

The uvrA gene of Erwinia chrysanthemi ENA49 similar to uvrA gene of Escherichia coli K12 has been cloned in vivo in Escherichia coli AB1886 uvrA6 cells using the plasmid pULB113 (RP4mini Mu). The presence of pULB113 carrying uvrA gene of Erwinia in Escherichia coli K12 uvrA- cells resulted in suppression of this mutation while uvrB and uvrC are not suppressed by this locus. The genetic control of excision repair of UV-damage in Erwinia chrysanthemi ENA49 is concluded to be similar to the one in Escherichia coli K12.     

少量制备大肠杆菌感受态细胞条件探索

目的:为了获得重复性好、转化率高的少量制备感受态细胞的方法,利用不同生长时期的大肠杆菌感受态细胞,进行转化比较。方法:根据普通实验室的实验条件,常规方法提取质粒,氯化钙法转化不同生长时期的大肠杆菌感受态细胞,比较转化率。结果:大肠杆菌感受态细胞的转化率与OD值显著相关,在OD600nm为0.39和0.55时转化率最高,在OD600nm为0.28~0.55之间均可得到理想的转化效果。结论:少量制备感受态细胞方法操作中无需添加任何保护试剂和细胞复苏培养,操作简便、重复性好,实验成本低廉。     

大肠埃希菌在肿瘤患者肠外的分布及耐药谱分析

目的：了解大肠埃希菌在肿瘤患者肠外的分布和感染情况及耐药性。方法：参照全国临床检验操作规程,采用K-B法对云南省肿瘤医院58例肿瘤患者继发大肠埃希菌感染进行分析及对9种抗生素的耐药谱测定。结果：各类肿瘤患者中,以宫颈癌及继发大肠埃希菌感染多见,其中,宫颈癌为28.30%。肺癌为26.87%。从标本来源来看,以尿液标本最多,为63.79%,其次为痰液12.07%及分泌物12.07%。结论：大肠埃希菌在肿瘤患者肠外分布广泛,所致感染较严重,经耐药谱测定发现大肠埃希菌多重耐药类型多,提示对肿瘤患者治疗应重视局部微生态平衡及控制感染。     

重庆地区儿童感染患者11 039株大肠埃希菌的临床分布及耐药性

目的 分析2010—2015年重庆地区儿童感染的11039株大肠埃希菌的临床分布特征及耐药性,为合理应用抗菌药物和预防控制医院感染提供依据。方法 分析大肠埃希菌对19种抗菌药物的耐药性,采用BD Phoenix 100 MIC法结合K-B纸片扩散法进行药敏试验,按美国临床实验室标准化委员会（CLSI）标准判断结果。结果 大肠埃希菌总检出率为13.53%;大肠埃希菌在呼吸病房和新生儿病房的检出率高于其他病房;以痰标本多见,占62.22%（6869/11039）;大肠埃希菌对氨苄青霉素耐药率最高（90.33%）,对美罗培南、亚胺培南的耐药率低,分别为0.81%和0.78%。ESBLs平均检出率为50.98%（5628/11039）。结论 大肠埃希菌的耐药性呈上升趋势,呼吸病房和新生儿病房是预防控制的重点科室。     

The product of the avian erythroblastosis virus erbB locus is a glycoprotein

Avian erythroblastosis virus (AEV) induces both erythroblastosis and fibrosarcomas in susceptible birds. A locus, v-erbB, within the viral genome has been implicated in AEV-mediated oncogenesis. We report here the detection and partial characterization of the protein product of the v-erbB oncogene in AEV-transformed cells. We obtained the antisera necessary for our analysis by expressing a portion of the molecularly cloned v-erbB locus in Escherichia coli and immunizing rabbits with the resulting bacterial erbB polypeptide. Antisera directed against the bacterial polypeptide reacted with v-erbB proteins obtained from virus-infected avian cells. By three criteria—tunicamycin inhibition, lectin binding and metabolic labeling with radioactive sugar precursors—the product of the v-erbB gene appears to be a glycoprotein.     

产ESBL克雷伯菌与大肠埃希菌质粒分布的初步研究

比较产超广谱 β 内酰胺酶克雷伯菌与大肠埃希菌质粒的表型与分布。用BlaTEM引物经PCR技术将 5 1株克雷伯菌与 2 9株大肠埃希菌编码产超广谱 β 内酰胺酶的质粒扩增 ,琼脂糖凝胶电泳分离目的片段后得到完整的质粒电泳图谱。克雷伯菌与大肠埃希菌存在有相同大小的产超广谱 β 内酰胺酶的质粒。 2种细菌有产ESBL相同质粒 ,PCR技术可用于产超广谱 β 内酰胺酶细菌质粒的分析。     

Metabolic regulation of the histidine operon in Escherichia coli and Salmonella typhimurium

Expression of the histidine operon in Escherichia coli cells in contrast to the one in Salmonella typhimurium is changed proportionally to cells growth rate on the different carbon sources. The specific activity of histidinol-dehydrogenase is repressed by addition of 19 amino acids both in Escherichia coli and Salmonella typhimurium independent of the growth medium used. Using of Escherichia coli and Salmonella typhimurium strains containing the heterologous histidine operons made possible to demonstrate the dependence of the histidine operon metabolic regulation to be determined by the operon itself but not by the specificity of the recipient cells. ppGpp was shown to be a positive regulator of the histidine operon expression in Escherichia coli.     

Carbohydrates as recognition determinants in phagocytosis and in lectin-mediated killing of target cells

Carbohydrate-lectin interactions serve as the basis of recognition by phagocytic cells of particles and of various target cells. Such interactions occur in the following systems: between sugars on the surface of the phagocytic cells and lectins on the surface of other cells—the best studied example is the binding of mannose-specific Escherichia coli and related organisms via their surface lectins to oligo-mannose residues on macrophages; between lectins on the surface of phagocytic cells and sugars on particles or other cells—phagocytosis of zymosan and of sialidase-treated erythrocytes, mediated respectively by mannose-specific and galactose-specific lectins on macrophages, belongs to this category; by extracellular lectins that form bridges between sugars on both types of cell—as shown by enhancement of phagocytosis of staphylococci by wheat germ agglutinin, and by lectin-dependent killing of target cells by macrophages. These interactions may play an important role in the activities of phagocytic cells in vivo. They may provide an initial host defense mechanism immediately after microbial infection, operate in tissues where phagocytic activity is poor, and participate in tumor rejection.     

产超广谱β-内酰胺酶大肠埃希菌对喹诺酮类抗菌药物的耐药性检测

目的 了解临床分离产超广谱β-内酰胺酶（ESBLs）大肠埃希菌对喹诺酮类等抗菌药物的耐药性。方法 NCCLS表型确证试验（纸片增强法）检测出临床分离大肠埃希菌中产ESBLs菌株,琼脂稀释法测定产ESBLs菌株对喹诺酮类等抗菌药物的耐药性。结果 临床分离大肠埃希菌中产ESBLs菌株的检出率为40．2％（92／229）,产ESBLs菌株以尿标本多见,对6种喹诺酮类抗菌药物的耐药率均在89％以上,哌拉西林的耐药率为100％;对头孢噻肟、头孢他啶和哌拉西林-三唑巴坦的耐药率分别为77．2％、1．1％和21．7％,对亚胺培南极其敏感,耐药率为0％。结论 产ESBLs大肠埃希菌发生率较高,对喹诺酮类抗菌药物耐药显著,临床应加强检测和监测。     

血培养阳性报警时间联合降钙素原、中性粒细胞/淋巴细胞比值对大肠埃希菌血流感染患者死亡风险的预测价值

摘要 目的：探讨血培养阳性报警时间（TTP）联合降钙素原（PCT）、中性粒细胞/淋巴细胞比值（NLR）对大肠埃希菌血流感染（BSI）患者死亡风险的预测价值。方法：选取2020年1月～2022年6月我院收治的223例大肠埃希菌BSI患者,根据入院后28 d内是否死亡分为死亡组和存活组。收集患者临床资料和血培养TTP,检测PCT、NLR。采用多因素Logistic回归分析大肠埃希菌BSI患者死亡的影响因素。采用受试者工作特征（ROC）曲线分析TTP、PCT、NLR对大肠埃希菌BSI患者死亡风险的预测价值。结果：223例大肠埃希菌BSI患者入院后28 d内死亡率为30.04％（67/223）。死亡组TTP短于存活组,PCT、NLR高于存活组（P均＜0.001）。多因素Logistic回归分析显示,年龄增加、入住重症监护病房（ICU）、气管插管/切开、PCT升高、NLR升高为大肠埃希菌BSI患者死亡的独立危险因素,TTP延长为独立保护因素（P＜0.05）。ROC曲线分析显示,TTP、PCT、NLR联合预测大肠埃希菌BSI患者死亡的曲线下面积大于各指标单独预测。结论：TTP缩短和PCT、NLR升高与大肠埃希菌BSI患者死亡风险增加相关,TTP、PCT、NLR联合预测大肠埃希菌BSI患者死亡风险的价值较高。