The virulence regulator AbsR in avian pathogenic Escherichia coli has pleiotropic effects on bacterial physiology

Avian pathogenic Escherichia coli(APEC)belonging to extraintestinal pathogenic E.coli(ExPEC)can cause severe infections in extraintestinal tissues in birds and humans,such as the lungs and blood.MprA(microcin production regulation,locus A,herein renamed AbsR,a blood survival regulator),a member of the MarR(multiple antibiotic resistance regulator)transcriptional regulator family,governs the expression of capsule biosynthetic genes in human ExPEC and represents a promising druggable target for antimicrobials.However,a deep understanding of the AbsR regulatory mechanism as well as its regulon is lacking.In this study,we present a systems-level analysis of the APEC AbsR regulon using ChIP-Seq(chromatin immunoprecipitation sequencing)and RNA-Seq(RNA sequencing)methods.We found that AbsR directly regulates 99 genes and indirectly regulates 667 genes.Furthermore,we showed that:1)AbsR contributes to antiphagocytotic effects by macrophages and virulence in a mouse model for systemic infection by directly activating the capsular gene cluster;2)AbsR positively impacts biofilm formation via direct regulation of the T2SS(type II secretion system)but plays a marginal role in virulence;and 3)AbsR directly upregulates the acid tolerance signaling system EvgAS to withstand acid stress but is dispensable in ExPEC virulence.Finally,our data indicate that the role of AbsR in virulence gene regulation is relatively conserved in ExPEC strains.Altogether,this study provides a comprehensive analysis of the AbsR regulon and regulatory mechanism,and our data suggest that AbsR likely influences virulence primarily through the control of capsule production.Interestingly,we found that AbsR severely represses the expression of the type I-F CRISPR(clustered regularly interspaced short palindromic repeats)-Cas(CRISPR associated)systems,which could have implications in CRISPR biology and application.

Solar photocatalytic pathogenic disinfection:Fundamentals to state-of-the-art

It is necessary to treat pathogen-infected water before its utilisation.Of conventionally used treatment methods,solar photocatalysis has gained considerable momentum owing to its operational simplicity and capacity to use freely and abundantly available solar energy.This article systematically reviewed the disinfection of water with photocatalysis.It addressed the concerns of microbial infection of water and the fundamentals behind its treatment with photocatalysis.It presented an in-depth description of pathogenic deactivation with powerful reactive oxygen species.Special emphasis was given to process intensification as it is an attractive technique that provides multifunctionality and/or equipment miniaturisation.Solar reactor design regarding mobilised/immobilised photocatalysts and compound parabolic concentrators were elucidated.Finally,key parameters governing photoperformance,corresponding trade-offs,and the need for their optimisation were discussed.Overall,this article is a single point of reference for researchers,environmentalists,and industrialists who address the ever-severing challenge of providing clean water whilst also maintaining energy sustainability.

Isolation and Identification of a Pathogenic E.coli Strain Causing Diarrhea in Foxes

[Objectives] The study aimed to identify the pathogenic E. coli strain that caused diarrhea in foxes and to analyze its drug sensitivity.[Methods] A pathogenic E. coli strain was isolated from dead foxes with diarrhea. By conventional bacterial isolation and culture, morphological observation, pathogenicity test and K-B disc method, the isolated strain was identified as pathogenic E. coli .[Results] The isolated pathogen was highly sensitive to ceftriaxone, cefotaxime, ciprofloxacin and lincomycin, moderately sensitive to enrofloxacin, neomycin, gentamycin, spectinomycin, florfenicol, amikacin and polymyxin, and resistant to ampicillin, amoxicillin and doxycycline.[Conclusions] This study provided reference for the prevention and control of diarrheal diseases in foxes in Qinhuangdao region.

Detection of High Pathogenicity Island(HPI) in Pathogenic Escherichia coli of Mink by PCR

[Objective] The paper was to analyze the carrying status of high pathogenicity island （HPI） in pathogenic Escherichia coli of mink. [Method] Eight strains of E. coli were isolated from dead mink, and conducted pathogenicity test of artificial infection. The carrying status of HPI （irp2, fyuA） was detected by PCR. [Result] Eight strains of E. coli were pathogenic E. coli, and the carrying rate of HPI （irp2, fyuA） was 100%, positively correlated with the pathogenicity. [Conclusion] The results lay a foundation for further exploring the pathogenic mechanism of E. coli..

Pathogenicity of Escherichia coli O_(123) from Rex Rabbit on White Mice

[Objective]The paper was to analyze the pathogenesis of Escherichia coli O123 from rex rabbit. [Method]E. coli O123 isolated from rabbit liver with diarrhea symptom in scale rex rabbit farm was intraperitoneally injected into 18- 22 g Kunming mice,and its pathogenicity was determined by clinical symptoms and pathological examination. [Result]When the inoculation concentration was about 8. 5 × 107 CFU /mL,Kunming mice appeared the clinical symptoms of drooping spirit,diarrhea and gathering,and the mortality reached 50%. Anatomical examination found that intestinal wall was thinning and intestinal mucosa was bleeding. [Conclusion]E. coli from rex rabbit has strong pathogenicity,and establishing animal model with Kunming mice to study its pathogenesis is of great reference significance for diagnosis and prevention of E. coli disease of rex rabbit.

一株致多发性浆膜炎猪源肠外E.coli的分离与鉴定

无菌采集多发性浆膜炎病猪的心血、肝脏,进行细菌分离培养,并对分离菌株进行染色镜检、生化试验、PCR鉴定、小鼠致病性试验和药敏试验。结果显示,分离菌株为肠外致病性大肠杆菌;分离菌株对庆大霉素、恩诺沙星、强力霉素、氨苄西林、磺胺类药物、头孢噻肟等均耐药,对阿米卡星、多粘菌素、美罗培南、亚胺培南敏感。本研究为致多发性浆膜炎的猪大肠杆菌病的诊断与防治提供了参考依据。

Isolation and Characterization of E. Coli O157 : H7 from Infected Newborn Calves in Northeast China

Escherichia coli O157 : H7 is a foodborne pathogen that poses a major threat to public health. Epidemiologic investigations have identified dairy cows, especially calves, are the principal reservoir of E. coli O157 : H7. In this study, based on the results, E. coli O157 : H7 was the main cause of E. coli disease outbreak in late October, 2015, and more than 90% of newborn calves died of serious diarrhea. Through further experiments, the drug sensitivity and resistance of the strain, the expression of the virulence gene and virulence pathogenicity were studied. E. coli O157 : H7 isolates were resistant to 12 antibiotics including penicillin, tetracycline and ampicillin, and were sensitive to eight antibiotics including cefoperazone, ceftazidime and amikacin. Resistance genes included tetB, strB, aadB, aphA, floR, TEM and virulence genes included stx1, eaeA and hlyA. Using specific pathogen free mice, the result showed that the isolate was pathogenic with a median lethal dose of 7.9×107 CFU · mL-1. This study described the pathogenesis and clinical manifestations of E. coli O157 : H7 infection. These results guided the use of antibiotics in prevent and control of bacterial infections in the future.

UV-TiO_2 photocatalytic disinfection and photoreactivation of pathogenic bacterium in municipal wastewater

The disinfected bacteria will be a photoreactivation under the irradiation of the sunlight,and the light intensity plays an important role in the bacteria resurrection.The effect of light intensity on photoreactivation of Escherichia coli(E.coli) and Enterococcus faecalis(E.faecalis) in secondary effluents which were disinfected respectively by pure UV and UV-TiO_2 was investigated.The results show that the disinfection efficiency of UV-TiO_2 is much higher than that of the pure UV disinfection.The photoreactivation rate of E.coli is much higher in pure UV disinfection than in UV-TiO_2 photocatalytic disinfection.Under high light intensity in UV-TiO_2 disinfection,high resurrection rate can be induced.However,a higher resurrection rate can be introduced even under low light intensity in pure UV disinfection alone.Meanwhile,UV-TiO_2 disinfection has a strong inhibition effect on E.faecalis photoreactivation.When the light intensity is lower than 21 μW/cm^2,nearly no resurrection of E.faecalis occurs after 72 h resurrection irradiation,and a little resurrection rate is observed only under a strong photoreactivating light intensity.

A Comparative Investigation on Growth of Three Food Born Pathogenic Bacteria Inoculated with Withania somnifera:an Invitro Experimental Study

Background:Withania somnifera(WS)is proposed as one of the alternatives instead of the antibiotic.This study is aimed to evaluate the inhibitory potency of enzymatic extract of the fruits of the WS.Methods:As an invitro experimental study,the growth rate of Shigella dysenteriae,Salmonella typhimurium,and Escherichia coli inoculated in different concentrations(25%,12.5%,6.25%and 3.125%)of the extract were assessed.A microtitre plate method was conducted.ANOVA was applied to identify statistical differences with p-value<0.05.Results:Different concentrations of extract,in comparison with control,declined the growth rate of all tested bacteria.All concentrations inhibited the growth of S.typhimurium(p<0.05).Compared to the microorganism control,effective concentration of the extract inhibiting the growth of E.coli was 12.5%,and 6.25%,while it was 12.5%,and 6.25%for Sh.dysenteriae(p<0.05).A dose-dependent response of E.coli was observed.The antibacterial activity of the extract tested was found mainly against E.coli and Sh.dysenteriae.The most resistant microorganism compared to E.coli and Sh.dysenteriae was S.typhimurium(p<0.05).25%of the concentration of the extract showed the different inhibitory effect among three tested bacteria(p<0.05).Conclusions:The extract was labeled as an antibacterial agent against the representative of three foodborne bacteria,Invitro.The common effective concentrations of the extract(12.5,and 6.25%)is recommended for further research,as food additive,to remedy digestive ailments related to E.coli,S.typhimurium and Sh.dysenteriae.

The Wheat Pathogenesis Related Protein (TdPR1.2) Ensures Contrasting Behaviors to <i>E. coli</i>Transformant Cells under Stress Conditions

The pathogenesis-related proteins 1 (PR-1) gene family play important roles in the plant metabolism in response to biotic and abiotic stresses. The wheat TdPR1.2 has been previously isolated and characterized. Here we showed by bio-informatic analysis that TdPR1.2 contains six cysteine residues that are conserved between all PR-1 proteins tested. Using ScanProsite tool, we found that TdPR1.2 structure has a CRISP family signature 1 and 2 located at the C-terminal part of the protein. Those two domains are conserved in many identified PR1.2 proteins in plants. Moreover, SignalIP-5.0 analysis revealed that TdPR1.2 contains a putative signal peptide formed by 25 amino acids at the N-terminal extremity. The presence of this signal peptide suggested that the mature proteins will be secreted after the cleavage of the signal sequence. Further, we investigate the role of the TdPR1.2 proteins in the growth of Escherichia coli transformants cells under different abiotic stresses. Our results showed that the full-length form of TdPR1.2 enhanced tolerance of E. coli against salt and osmotic stress but not to KCl. Moreover, TdPR1.2 protein confers bacterial tolerance to heavy metals in solid and liquid mediums. Based on these results, we suggest that the TdPR1.2 protein could play an important role in response to abiotic stress conditions.

产毒性大肠杆菌(ETEC)中毒力岛分布的研究

目的 了解小肠结肠炎耶尔森菌强毒力岛在产毒性大肠杆菌中的分布，以便于进一步深入研究毒力岛在大肠杆菌中的结构的完整性和功能。方法ＰＣＲ扩增和原位杂交及基因序列分析。结果 在检测的９３株产毒性大肠杆菌（ＥＴＥＣ）的毒力岛的检出率为 ３２．２５％（３２／９３），而且这些阳性菌株中的毒力岛大部分连接到 ａｓｎｔＲＮＡ（天门冬氨酸ｔＲＮＡ）位点。结论 产毒性大肠杆菌是致病性较强的病原菌之一，而小肠结肠炎耶尔森菌强毒力岛在产毒性大肠杆菌中阳性率较高的分布．

产NDM ExPEC毒力与其耐药性和分子分型的关系研究

目的对产新德里金属β-内酰胺酶(NDM)肠外致病性大肠埃希菌(ExPEC)菌株进行分子分型,研究产NDM ExPEC毒力与其对碳青霉烯类抗菌药物的耐药性和分子分型之间的关系。方法选取分离自该院的15株产NDM ExPEC菌株(实验组)和28株不产NDM ExPEC菌株(对照组)作为研究对象,对产NDM ExPEC菌株进行系统发育分群和多位点序列分型(MLST)研究,PCR方法检测7种毒力基因在这两组菌株中的分布,χ2检验比较分布差异。结果系统发育分群结果显示,15株产NDM ExPEC菌株中,13株属于A群,2株属于D群;MLST分型结果显示,ST410是检出率最高的ST型;实验组毒力基因fimH检出率最高;对照组毒力基因sat、fyuA和ompT检出率高于实验组,差异有统计学意义(P<0.05)。结论该院绝大多数产NDM ExPEC菌株的毒力较弱,表明细菌耐药性强并不意味着其致病性也强。

肠聚集性大肠杆菌ybtE基因缺失突变株的构建

目的构建肠聚集性大肠杆菌ybtE缺失突变株,为下一步ybtE基因功能的研究奠定基础。方法应用PCR扩增肠聚集性大肠杆菌的ybtE基因,将其克隆至pUC18载体,酶切缺失988bp ybtE片段,并插入998bp的卡那霉素抗性基因,利用定向克隆技术将突变的ybtE片段克隆至自杀质粒pKTN701,形成重组自杀质粒。将携带重组自杀质粒的SM10λpir与EAEC17-2进行接合转移,利用同源重组,根据卡那霉素抗性筛选并鉴定接合子。结果经PCR、酶切和探针杂交鉴定,得到2株肠聚集性大肠杆菌ybtE缺失突变株。结论成功构建肠聚集性大肠杆菌ybtE基因缺失突变株。

Department of Etiology and Department of Pharmacology

In order to understand the role of transmembrane signal transduction of host cells in the early steps of infection,the adherence of E. coli to HEp 2 cells and the change of activity of phospholipase C γ (PLC γ) induced by the adherence were investigated.The adherence of enteropathogenic E.coli (EPEC), strain E.7, induced a significant increase of inositol triphosphat (IP 3) level in HEp 2 cells. The adherence of the bacteria and the increase of IP 3 was kinetically correlated. Whereas the increase of IP 3 level induced by the adherence of the control strain EPEC (H511), a non piliated strain, was much meager than that by E7, a piliated strain. The results highlighted an important role of transmembrane signals like IP 3 in the pathogenesis of EPEC.

Effects of Preparation and Storage of Agar Media on the Sensitivity of Bacterial Forward Scattering Patterns

Recent worldwide foodborne outbreaks emphasize the need for the development of rapid and accurate method for pathogen detection. To address such issues, a new colony based label-free detection method working on the principles of elastic light scattering was introduced. In order to build libraries of scattering images for bacterial pathogens, it is pertinent to determine the effect of preparation and storage of the agar media on the scatter patterns. Scatter patterns of three Escherichia coli serovars (O26, O111 and O157) were studied and used in a model system, after growth on Sorbitol-MacConkey agar plates that were prepared and stored at different conditions in the laboratory. Quantitative image processing software was used to analyze variation in scatter patterns of the same serovar on media prepared under various standard laboratory conditions and to generate a cross-validation matrix for comparison. Based on the results, it was determined that attention should be given during preparation of media so that the agar plates are not air-dried more than 10 - 20 min after solidification at room temperature. The plates could be stored in sealed bags in cold room (4oC - 10oC) for up to a month before use. The findings of this study should provide guidelines in preparation, storage, and handling of media for generation of reproducible scatter patterns of bacterial colonies with the light scattering sensor for pathogen detection.

犊牛源肠外致病性大肠杆菌分离鉴定、致病性及耐药性分析

【目的】确定新疆喀什地区某规模化牛场致犊牛死亡的细菌性病原体及其生物学特性,为该地区规模化牛场细菌病的防治和合理用药提供参考。【方法】对死亡犊牛进行病理解剖及病原分离,挑选肝脏(KS-1)与肛拭子(KS-2)来源的单菌落纯化培养,通过形态观察、生化试验、16S rRNA基因扩增及系统进化分群鉴定分离株,并通过毒力基因检测、致病性试验及药敏试验对分离株的致病性和耐药性进行研究。【结果】死亡犊牛肺脏、肝脏、心血、关节液和肠系膜淋巴结以及健康犊牛肛拭子样品接种后菌落生长形态一致,结合选择性培养基上菌落生长形态、革兰染色镜检形态特点、生化特性及大肠杆菌16S rRNA的验证,鉴定分离菌株为大肠杆菌。系统进化分群试验中,KS-1为A群,KS-2为B1群。毒力基因检测结果显示,KS-1携带crl、papC、hlyE、eaeA、ompA、ompC、iucD和iutA共8种毒力相关基因,KS-2携带crl、hlyE和ompF共3种毒力相关基因。致病性试验结果显示,腹腔注射1×108 CFU分离株感染小鼠,KS-1组小鼠在16 h内全部死亡,KS-2组小鼠无死亡现象出现。药敏试验结果显示,KS-1对哌拉西林/他唑巴坦、头孢唑林、头孢呋辛等17种药物敏感,对氨苄青霉素/舒巴坦和哌拉西林中度敏感,对氨苄西林和复方新诺明耐药;KS-2对哌拉西林/他唑巴坦、头孢替坦、亚胺培南、美罗培南、阿米卡星和呋喃妥英敏感,对妥布霉素中度敏感,对其余14种药物耐药。【结论】本研究分离出2株犊牛源大肠杆菌,其中KS-1毒力较强,且携带丰富的毒力基因,对大部分常用抗生素敏感,对氨苄西林和复方新诺明具有耐药性;KS-2携带毒力基因较少,呈现多重耐药。

黄芩苷对猪源肠外致病性大肠杆菌的体内保护作用

为探究黄芩苷对猪源肠外致病性大肠杆菌(Extraintestinal pathogenic Escherichia coli,ExPEC)感染小鼠后的保护作用,采用小鼠存活率试验和组织载菌量试验评估黄芩苷对小鼠的保护作用,比较感染组和黄芩苷治疗组小鼠临床表现、存活率、组织载菌量及病理变化。结果表明,猪源肠外致病性大肠杆菌感染后小鼠状态萎靡不振、食欲下降,黄芩苷治疗组小鼠状态较感染组状态好;黄芩苷治疗组小鼠存活率相对于感染组有一定提升;黄芩苷治疗组小鼠心脏、肝脏、脾脏和肾脏中组织载菌量及病理变化相对于感染组均极显著降低(P<0.01)。

猪源肠外致病性大肠杆菌的分离与鉴定

目的检测猪源肠外致病性大肠杆菌(ExPEC)的血清型与毒力基因。方法对2013-2014年间秦皇岛地区30份组织病料采用细菌形态观察、培养特性试验、生化试验鉴定出8株肠外致病性大肠杆菌。采用玻片凝集法测定这些大肠杆菌病的血清型,并用PCR扩增法检测9种毒力基因。结果定型菌株5株,分别属于O53、O93和O157 3个血清型,9种毒力基因PCR检测结果表明ler、iutA、irp2、fyuA、astA 5种基因检出率分别为100%、75.0%、50.0%、50.0%、25.0%。结论所检测的猪源肠外致病性大肠杆菌以O53、O93和O157为主要流行血清型,同时携带fyuA、ler和iutA基因的菌株致病性较强。

肠外致病性大肠埃希菌多重耐药性及氨基糖苷类耐药基因分析

目的检测临床分离猪、鸡源肠外致病性大肠埃希菌的耐药性,分析其携带的氨基糖苷类耐药基因,以指导临床用药,探究细菌对氨基糖苷类抗生素的抗性机制。方法应用K-B法测定分离株对19种常用抗生素的耐药情况。用双纸片法检测产ESBLs菌株并进行基因分型。用PCR和测序方法检测氨基糖苷类抗性基因aadA1,aadA2,strA-strB,aadB,aacC2,aac(3)-IV,aph(3′)-Ia,aph(3′)-IIa。结果临床分离株表现多重耐药性,对头孢菌素类药物和阿米卡星的敏感性最高(63%～97%),93%的菌株耐药谱值≥10。共检测到两株产超广谱β-内酰胺酶(ESBLs)菌株,基因型属TEM-1+CTX-M-14。耐药菌株与相应的氨基糖苷类耐药基因符合率高(≥74%)且序列保守。结论猪、鸡源肠外致病性大肠埃希菌呈多重耐药性,其对氨基糖苷类抗生素的抗性机制以产生针对该类抗生素的修饰酶为主。

规模化养鸭场鸭源大肠杆菌分离株的致病性及系统发育分析

为研究鸭源大肠杆菌(E.coli)的致病性,明确鸭病原性E.coli与人和其他动物E.coli的亲缘关系及O血清型与菌种间遗传关系的相关性,本研究从我国西南地区规模化养鸭场患典型E.coli败血症的雏鸭体内分离37个血清型共82个E.coli分离株(其中优势血清型32株,其他血清型50株),以每株0.2 mL(l09 cfu/mL)腿部肌肉接种7日龄健康鸭进行致病性试验;并对其中14个鸭源E.coli代表株进行16S rRNA基因克隆、测序及系统发育分析。结果表明:所有分离株全部为致病性E.coli,其中高致病株、中等致病株和低致病株分别占受试菌株的80.5%(66/82)、17.1%(14/82)和2.4%(2/82),4个优势血清型全部为高致病株和中等致病株,分别占受试菌株的84.4%(27/32)和15.6%(5/32);14个代表株鸭源E.coli的16S rRNA形成2个主要分支,3株新发现的血清型单独形成一个较远的分支,11株常见血清型与人及其他动物源E.coli形成另一个大的分支,其中有6株与人的O157亲缘关系较近,2株与出血性E.coli O157∶H7 sakai聚为一个小分支;从遗传进化的关系分析,这些菌株有可能是人类的潜在病原菌;同时还发现不同血清型的分离株可以聚为一支,而同一血清型的分离株可以处于不同的分支,常规的O血清学分型不能体现菌种间遗传关系的远近。