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.

Distribution of Virulence-Associated Genes of Avian Pathogenic Escherichia coli Isolates in China

216 avian pathogenic Escherichia coli （APEC） isolates were obtained from poultry with colibacillosis in different areas of China. Among them, 195 were serotyped as 078, 088, and 093. Thirteen virulence-associated genes, including fimC, iucD, iss, tsh, fyuA, irp2, eaeA, hlyE, colV, papC, stx2f, vat, and astA, were submitted to PCR amplification. The fimC gene was the most prevalent with a detection rate of 93.6%, followed by iucD （70.8%）, iss （58.8%）, and tsh （51.4%） in APEC isolates. The detection rate of high pathogenicity islands （HPI）-associatedfyuA and irp2 genes were both 44.9%, with no LEE （the locus of enterocyte effacement） island-associated gene eaeA detected. In terms of distribution patterns of the 13 virulence-associated genes, 5 isolates harborbed 10 genes, 19 isolates contained onlyfimC gene, and only 4 isolates had no virulence-associated gene detected. Different correlations of the virulence-associated genes with O serotypes were also investigated and 50% 078 isolates had a gene distribution patterns of fimC^＋iucD^＋irp2^＋fyuA^＋iss^＋colV^＋tsh^＋.

重组体pET-28a-alkB/E.coli降解页岩油泥石油烃的功能研究

为了降解页岩油泥中的石油烃,构建基因工程菌,表达石油烃降解关键酶以提高油泥的降解效果。以Pseudomonas qingdaonensis基因组为模板扩增alkB基因,连接至载体pET-28a,转化至E.coli BL21(DE3)中,SDS-PAGE和Western-blot鉴定表达的外源蛋白。pET-28a-alkB/E.coli处理原油和页岩油泥,采用气相色谱法评估降解效果。发现alkB基因在大肠杆菌中能表达外源蛋白,且14 d原油及页岩油泥的降解率分别为47.5%和47.1%,表明重组体pET-28a-alkB/E.coli具备降解页岩油泥石油烃的功能。

Exploring the modulatory role of bovine lactoferrin on the microbiome and the immune response in healthy and Shiga toxin‑producing E.coli challenged weaned piglets

Background Post-weaned piglets suffer from F18+Escherichia coli(E.coli)infections resulting in post-weaning diar-rhoea or oedema disease.Frequently used management strategies,including colistin and zinc oxide,have contrib-uted to the emergence and spread of antimicrobial resistance.Novel antimicrobials capable of directly interacting with pathogens and modulating the host immune responses are being investigated.Lactoferrin has shown promising results against porcine enterotoxigenic E.coli strains,both in vitro and in vivo.Results We investigated the influence of bovine lactoferrin(bLF)on the microbiome of healthy and infected weaned piglets.Additionally,we assessed whether bLF influenced the immune responses upon Shiga toxin-producing E.coli(STEC)infection.Therefore,2 in vivo trials were conducted:a microbiome trial and a challenge infection trial,using an F18+STEC strain.BLF did not affect theα-andβ-diversity.However,bLF groups showed a higher relative abundance(RA)for the Actinobacteria phylum and the Bifidobacterium genus in the ileal mucosa.When analysing the immune response upon infection,the STEC group exhibited a significant increase in F18-specific IgG serum levels,whereas this response was absent in the bLF group.Conclusion Taken together,the oral administration of bLF did not have a notable impact on theα-andβ-diversity of the gut microbiome in weaned piglets.Nevertheless,it did increase the RA of the Actinobacteria phylum and Bifi-dobacterium genus,which have previously been shown to play an important role in maintaining gut homeostasis.Furthermore,bLF administration during STEC infection resulted in the absence of F18-specific serum IgG responses.

Differentiation of Avian Pathogenic <i>Escherichia coli</i>Strains from Broiler Chickens by Multiplex Polymerase Chain Reaction (PCR) and Random Amplified Polymorphic (RAPD) DNA

We examined 50 Escherichia coli (E. coli) strains isolated from broiler chickens between January 2013 to March 2014 in order to evaluate the epidemiological prevalence of avian pathogenic E. coli (APEC) in Jordan by multiplex PCR and random amplification of polymorphic DNA (RAPD) tests. The multiplex polymerase chain reaction (PCR) which was used as tentative criteria of APEC targets 8 virulence associated genes;enteroaggregative toxin (astA), Type 1 fimbria adhesion (fimH), iron-repressible protein (irp2), P fimbriae (papC), aerobactin (iucD), temperature-sensitive hemagglutinin (tsh), vacuolating autotransporter toxin (vat), and colicin V plasmid operon (cva/cvi) genes. The number of detected genes could be used as a reliable index of their virulence. E. coli strains already typed as an APEC always harbor 5 to 8 genes, but non-APEC strains harbor less than 4 genes. Assuming the criteria of an APEC is possession of 5 or more virulence associated genes;we found that all 50 E. coli strains were classified as APEC strains. The RAPD analysis showed that the E. coli strains could be grouped into 35 of RAPD types by using these two different RAPD primer sets, RAPD analysis primer 4 5'AAGAGCCCGT5', and RAPD analysis primer 6 5'CCCGTCAGCA3'. The current study confirmed the endemic nature of APEC in broiler flocks in Jordan. It is essential that the biosecurity on poultry farms should be improved to prevent the introduction and dissemination of APEC and other agents. Furthermore, farmers need to be educated about the signs, lesions, and the importance of this agent.

Drug Susceptibility Test of Volatile Oil of Artemisiaargyi to Avian Pathogenic Escherichia coli

The volatile oil of Artemisia argyi was extracted by ultrasonic assisted extraction, and the extraction rate of volatile oil was 0.68%. Thevolatile oil of A. argyi was emulsified with 1% Tween-80, and drug susceptibility test was conducted with avian Escherichia coli. The results showedthat the volatile oil of A. argyi had antibacterial effect against avian E. coli, and the minimal inhibitory concentration was 50 mg/mL. Taking sixcommon antibiotics as the control, drug susceptibility test was conducted with volatile oil of A. argyi. The results showed that 10 strains of E. coliwere sensitive to the volatile oil of A. argyi, three of which had different degrees of resistance and one had the tendency of resistance.

Impact of an oligosaccharide-based polymer on the metabolic profiles and microbial ecology of weanling pigs experimentally infected with a pathogenic E.coli

Background Our previous study has reported that supplementation of oligosaccharide-based polymer enhances gut health and disease resistance of pigs infected with enterotoxigenic E.coli(ETEC)F18 in a manner similar to carbadox.The objective of this study was to investigate the impacts of oligosaccharide-based polymer or antibiotic on the host metabolic profiles and colon microbiota of weaned pigs experimentally infected with ETEC F18.Results Multivariate analysis highlighted the differences in the metabolic profiles of serum and colon digesta which were predominantly found between pigs supplemented with oligosaccharide-based polymer and antibiotic.The relative abundance of metabolic markers of immune responses and nutrient metabolisms,such as amino acids and carbohydrates,were significantly differentiated between the oligosaccharide-based polymer and antibiotic groups(q<0.2 and fold change>2.0).In addition,pigs in antibiotic had a reduced(P<0.05)relative abundance of Lachnospiraceae and Lactobacillaceae,whereas had greater(P<0.05)Clostridiaceae and Streptococcaceae in the colon digesta on d 11 post-inoculation(PI)compared with d 5 PI.Conclusions The impact of oligosaccharide-based polymer on the metabolic and microbial profiles of pigs is not fully understood,and further exploration is needed.However,current research suggest that various mechanisms are involved in the enhanced disease resistance and performance in ETEC-challenged pigs by supplementing this polymer.

Characteristics of β-Lactamase Synthesis in E. coli and K. pneumanie Strains in Nosocomial Infections

Background: Recently micro-organisms that synthesize extended-spectrum β-lactamase (ESBLs) were increased. The peculiarities of ESBL synthesis of Escherichia coli and Klebsiella pneumoniae strains that cause nosocomial urinary tract infections, surgical site infections and pneumonia in surgical clinic were studied. ESBL synthesis were observed 38.9% of E. coli strains obtained from urine, 92.3% of strains obtained from surgical site infections, and 50% of strains obtained from sputum. ESBL synthesis were observed 37.5% of K. pneumoniae strains obtained from urine, 85.7% of strains obtained from surgical site infections, and 60% of strains obtained from sputum. Different levels of ESBL synthesize of E. coli and K. pneumoniae strains isolated from different pattern is discussed. Conclusion. ESBL synthesis is common in E. coli and K. pneumoniae strains, which cause nosocomial infections. The frequency of occurrence of ESBL s synthesis among of these strains depends on clinical forms of nosocomial infections.

The Transport and Persistence of Escherichia coli in Leachate from Poultry Litter Amended Soils

Fecal coliform bacteria such as Escherichia coli (E. coli) are one of the main sources of groundwater pollution. An assessment of the transport and Persistence of E. coli in poultry litter amended Decatur silty Clay soil and Hartsells Sandy soil was conducted using soil columns and simulated groundwater leaching. Enumeration of initial E. coli was determined to range from 2.851 × 103 to 3.044 × 103 CFU per gram of soil. These results have been used in a batch study to determine the persistence rate of E. coli in Decatur silty Clay soil and Hartsells Sandy soil. Results prove that E. coli survival growth rate increases for clay soil later than and at a higher rate than sandy soil. The column study has determined that E. coli was transported at a rate of 3.7 × 106 CFU for Decatur silty loam and 6.3 × 106 CFU for Hartsells sandy per gram of soil. Further, linear regression analysis predictions show higher porosity and soil moisture content affect transport, and Hartsells sandy soil has higher transport of E. coli due to its higher porosity and lower volumetric water content.

Growth Inhibition of Escherichia coli and Staphylococcus epidermidis from Various Types of Honey

Honey has long been considered a wound treatment used to keep cuts and other epidermal injuries clean. This study tested that claim by comparing manuka honey used in medicine today, local unprocessed honey taken straight from a hive, and pasteurized honey found at a store, on strains of E. coli and S. epidermidis. The study evaluated the effects these honeys had on bacterial growth to determine which had the greatest inhibition of bacterial growth. To determine this, plates streaked with strains of E. coli or S. epidermidis bacteria and agar wells filled with one of the honeys were incubated and subsequently the diameter of the zone of inhibition was measured. After 20 trials using each honey and bacteria type, manuka and unprocessed were shown to have a statistically significant advantage over the pasteurized honey at inhibiting the growth of E. coli and S. epidermidis, though it was variable whether manuka had an advantage over the unprocessed honey.

Evaluating the Potential of Nitrofurantoin and Fosfomycin for E.coli UTIs: A Susceptibility Study

This study was designed to find the susceptibility of Nitrofurantoin and Fosfomycin among urinary isolates of Escherichia.coli.Four hundred(400)urine samples were collected for susceptibility of nitrofurantoin and fosfomycin among urinary isolates of E.coli.All indoor and outdoor patients'urinary samples yielded growth of E.coli.Mid-stream urine specimens were inoculated on blood agar and CLED agar and incubated at 35±2°C.Growth was observed,and Escherichia coli was identified by Gram staining,Catalase,Motility test and API 20E(Bio murex)as per standard procedure.Antimicrobial susceptibility testing of isolates for nitrofurantoin and fosfomycin was carried out by the modified Kirby-Bauer disc diffusion method according to CLSI guidelines ATCC 25922.E.coli was used as a quality control strain.A total of 400 samples were tested susceptibility of nitrofurantoin and fosfomycin among urinary isolates of E.coli during this period.A total of 400 samples yielded the growth of E.coli,out of which 178(44.5%)were male and 222(55.5%)were female samples.Among males,18(10%)were tolerant to nitrofurantoin,and 2(1.1%)were tolerant to fosfomycin.Among females,9(4.09%)were susceptible to nitrofurantoin while 6(2.72%)were susceptible to fosfomycin.Among age groups below 45 years old,6(4.76%)were tolerant to nitrofurantoin,and 2(1.58%)were sensitive to fosfomycin.Between 46-66 years old,4(2.81%)were sensitive to nitrofurantoin,and 3(2.11%)were sensitive to fosfomycin.Between 67-90 years old,17(12.87%)were sensitive to nitrofurantoin,and 4(3.03%)were tolerant to fosfomycin.Fosfomycin and nitrofurantoin showed good susceptibility in urinary isolates of E.coli and can be used empirically in our setup.

Fosfomycin Resistance in Avian Pathogenic Escherichia coli Isolates

Fosfomycin, a broad-spectrum antibiotic against both Gram-positive and Gram-negative bacteria, is very important in the clinic but many fosfomycin-resistant bacteria have been isolated from patients. In this study, the resistance mechanism of three fosfomycin-resistant avian pathogenic Escherichia coli （APEC） strains （JE1, IF7 and CD11） isolated from septicemic chickens were analyzed. The results showed that their fosfomycin-resistance mechanisms were different. An alteration in the glpT transport system was the main reason of the fosfomycin-resistance mechanisms of strain IF7. Compared with the control stain BL21, the capacity of fosfomycin-uptake was low in all these three stains （JE 1 〉IFT〉CD 11）. Sequence results of murA showed that there were more than 10 sites of nucleotide mutation, but only one amino acid mutation T116A showed in CD11. Real-time detection test showed that the expression level of the murA gene of the three stains was significantly increased （four times increase in strain CD11 and two times increase in strains JE1 and IFT）. The transformation and recombinant test showed that the recombinant bacteria with the murA of JE1 and CD11 showed high minimal inhibitory concentration （MIC） against fosfomycin. From the results of this research, it showed that most of the fosfomycin- resistance mechanisms once showed in patient bacteria have appeared in the APEC strains and the fosfomycin-resistance mechanism of the three APEC isolates was different.

Screening of Chinese Herbal Medicines against Avian Infectious Laryngotracheitis Virus and Escherichia coli

[ Objective] This study aimed to screen Chinese herbal medicines against avian infectious laryngotracheitis virus and Escherichia coli. [ Method ] Conventional Chinese medicine plate dilution method, plate punching method and test tube method were applied to screen Chinese herbal medicines against avian infectious laryngotracheitis virus based on chicken embryo inoculation experiment. [ Result ] Forsythia suspensa, Radix lsatidis, Isatis iadigotica, Lonicera japonica, Codonopsis pilosula, Astragalus membranaceus, Rhizoma Atractylodis Macrocephalae, C, tyeyrrhiza uralensis and Pericarpium granati had relatively strong anti-ILTV effect; among the Chinese herbal medicines against avian E. coli, Sanguisorba offwinalis, Fructus Mume, Rheum palmatum, Anemarrhena asphodeloides, Radix Scutellariae and Fagopyrum cymosum had relatively strong effect against avian E. coli Os, while other Chinese herbal medicines had relatively weak or no inhibitory effect on avian E. coli 0s and 05. [ Conclusion] This study laid the foundation for further development of Chinese herbal medicine compound preparations to treat avian infectious laryngotracheitis, avian colibacillosis and other viral diseases and bacterial diseases.

Isolation and Identification and Drug Resistance Analysis of Avian Escherichia coli and In Vitro Competition with Lactobacillus

[Objective]The paper was to understand the resistance of commonly used clinical antibiotics to avian Escherichia coli and the inhibition of lactobacillus against pathogenic E.coli.[Method]The strain was isolated and identified by SS medium culture,microscope examination,PCR amplification and biochemical test,and the sensitivity of avian E.coli to eight antibiotics was determined by disk diffusion method.Twelve strains of avian E.coli and one strain of lactobacillus were divided into five group using in vitro liquid culture method,namely simultaneous mixed culture group of lactobacillus and E.coli,first colonization group of E.coli,first colonization group of lactobacillus,E.coli control group and lactobacillus control group,for competitive inhibition test between E.coli and lactobacillus.[Result]Totally 34 strains with similar characteristics to E.coli strains were preliminarily screened;the target band amplified by PCR was 293 bp,which was consistent with the expected size.The isolated strain could decompose maltose,sucrose,lactose,glucose,arabinose,mannitol and sorbitol;the nitrate reduction test was positive,while citrate and V-P test was negative,consistent with the biochemical characteristics of E.coli.The resistance rate of the isolates to cefalexin and tylosin was over 65%,and that to enrofloxacin and doxycycline was up to 50%;the isolates were more sensitive to apramycin,tilmicosin,neomycin and colistin,and the total resistance rate was below 15%.The inhibitory effects of mixed culture of lactobacillus and E.coli against E.coli was better than first colonization of lactobacillus at 24 h;the inhibitory effects of first colonization of lactobacillus against E.coli was better than that of mixed culture of lactobacillus and E.coli at 36 h;the inhibitory effects of mixed culture of lactobacillus and E.coli against E.coli was the best at 48 h.[Conclusion]The isolated strains of E.coli were generally resistant to eight commonly used antibiotics.Lactobacillus could inhibit the growth of avian E.coli in vitro,and first colonization of lactobacillus received the best effort.

Detection of new antibiotic resistance gene profile in Escherichia coliassociated with avian leukosis virus infection from broiler chickens

The Escherichia coli(E.coli)is prevailing worldwide,but the epidemiology of E.coli infections feature regional distribution characteristics to some extent.E.coli,as a zoonotic pathogen,can be transferred from animals to humans through food chain or via contact with wounds,causing a public health risk.We reported the swelling of proventriculus and tracheal bleeding following the death in two broiler chickens(Gallus gallus domesticus)from Beijing,China.To investigate whether a virus was involved in the infection,Madin Darby Bovine Kidney(MDCK)cells were co-cultured with supernatants of proventriculus,trachea and spleen homogenates.The avian leucosis virus was detected in the samples of proventriculus and trachea,but the avian influenza virus,the Newcastle disease virus and the avian infectious laryngotracheitis virus were not detected.E.coli isolates were resistant to almost all the antimicrobial as tested except for the combinations of amoxicillin/clavulanic acid and sulfamethoxazole/trimethoprim.PCR tests demonstrated the presence of antibiotic resistance genes in these E.coli isolates and further research revealed a novel gene profile with the presence of CTX-M-1,gyrA,gyrB,oqxA,oqxB,parC and Sul2 antibiotic resistance genes in a strain isolated from a proventriculus sample.These results demonstrated that the presence of antibiotic resistant E.coli would not necessarily cause outbreak of large-scale disease.However,when the bacteria carrying new antibiotic resistance genes enter the environment,it may result in the development of more virulent strains which will potentially impact human and animal health.

重组E.coli工程菌高密度培养生产人源型胶原蛋白

Several technical parameters were studied during the fermentation of recombinant E.coli for the production of collagen-like biopolymer.The effects of dissolved oxygen as well as glucose concentration on fermentation were observed.The OD 600 value could reach 98 when dissolved oxygen was controlled at 50% and glucose around 1%.The production of human-like collagen with a yield of 29.4% was obtained.

养殖场空气中E.coli磺胺类抗生素的抗性

本文从养殖场空气中分离出360株E.coli(大肠杆菌,Escherichia coli),应用肉汤微量稀释法和PCR方法,分离磺胺甲唑敏感菌株,检测抗生素抗性和抗性基因.在分离的E.coli中,对磺胺甲唑敏感菌株为95株(26.4%),有48株含有青霉素、氯霉素、四环素、环丙沙星、庆大霉素和利福平的抗性,而47株未含有抗性.其中,7株菌株含有1种抗生素抗性、11株菌株含有2种抗生素抗性、17株菌株含有3种抗生素抗性、6株菌株含有4种抗生素抗性、4株菌株含有5种抗生素抗性、3株菌株含有6种抗生素抗性.对抗生素的耐受能力依次为:氯霉素、青霉素、四环素、环丙沙星、庆大霉素、利福平.磺胺甲唑敏感菌株共检出163个抗性基因,sul1、int1、sul2、Int2、sul3检出数量依次为49、44、29、20和19;含一种、二种、三种、四种、五种抗性基因菌株分别为45、22、10、7、2;但有6株未检测出抗性基因.结果表明养殖场建场时间、抗生素使用、养殖规模等与抗生素抗性菌的抗性呈正相关;养殖场空气中分离的E.coli抗生素抗性较高,且具有多重抗性;抗生素抗性的表现型与其基因型之间出现不完全吻合现象.

秦皇岛地区狐源致病性E.coli对四环素类药物耐药性和耐药基因的检测

为了确定秦皇岛地区狐源大肠杆菌(E.coli)对四环素类药物的耐药性和耐药基因分布,采用常规的鉴定方法,从不同养狐场送检的腹泻的狐狸体内分离鉴定出20株E.coli。致病性试验表明,该菌为致病性E.coli。药敏试验结果表明:分离菌株对四环素和强力霉素药率分别达到95%和90%。通过PCR方法检测分离菌株四环素类药物的耐药基因,结果显示,tet A和tet B基因的检出率分别为100%和95%。本研究为秦皇岛地区防治狐源致病性大肠杆菌病提供实验基础。

中药三黄汤、小檗碱对E.coli生长抑制作用与庆大霉素的比较

庆大霉素对E .coli的抑菌作用强 ,试管稀释法、琼脂稀释法和平板抑菌圈法抑菌试验得到一致的MIC(3～5 μg/mL) ;三黄汤、小檗碱抑菌作用弱而稳定 ,试管稀释法测得的MIC分别为 5 0 0mg/mL和 2 .5mg/mL ,而用平板抑菌圈法得不到明显的抑菌圈 .在亚MIC药物液体分批培养时 ,庆大霉素抑菌曲线起初 2h迅速下降 ,而后回升到初始浓度 ,而三黄汤抑菌曲线平缓稳定 ,说明中药和抗生素的抑菌机制明显不同 .庆大霉素与小檗碱混合使用时 ,与庆大霉素单独使用相近 ,而与三黄混合使用时 ,则主要表现三黄汤的抑菌作用 .因此中西药结合时需要慎用 .在接近MIC药物培养基中连续传代 2 0次后 ,庆大霉素MIC提高 8倍多 ,而三黄汤和小檗碱MIC无显著变化 ,无抗药性产生 ,也不产生对庆大霉素的交叉抗药性 .研究结果还表明 ,在 1/ 4MIC三黄汤中连续传代 2 0次 ,能消除E .coli已形成的庆大霉素抗性 ,这为解决抗生素抗药性提供了线索 .图 4参 2

蜂胶佐剂对E．coli多价灭活苗免疫小鼠效果的实验研究

目的:铝胶苗和蜂胶佐剂苗对E.coli免疫原性的比较。方法:选用菌株O2、O78及O157菌株血清型,研制的多价大肠杆菌氢氧化铝胶和蜂胶佐剂灭活苗,对同血清型菌株的攻击具有良好的保护作用。结果:通过用铝胶苗和蜂胶佐剂苗试验结果表明,对于大肠杆菌病而言,用蜂胶佐剂苗的效果优于铝胶苗。结论:蜂胶佐剂苗的效果优于铝胶苗。