Antibacterial mechanism of kojic acid and tea polyphenols against Escherichia coli O157:H7 through transcriptomic analysis

Escherichia coli O157:H7 is one of the major foodborne pathogenic bacterial that cause infectious diseases in humans.The previous found that a combination of kojic acid and tea polyphenols exhibited better activity against E.coli O157:H7 than using either alone.This study aimed to explore responses underlying the antibacterial mechanisms of kojic acid and tea polyphenols from the gene level.The functional enrichment analysis by comparing kojic acid and tea polyphenols individually or synergistically against E.coli O157:H7 found that acid resistance systems in kojic acid were activated,and the cell membrane and genomic DNA were destructed in the cells,resulting in“oxygen starvation”.The oxidative stress response triggered by tea polyphenols inhibited both sulfur uptake and the synthesis of ATP,which affected the bacteria's life metabolic process.Interestingly,we found that kojic acid combined with tea polyphenols hindered the uptake of iron that played an essential role in the synthesis of DNA,respiration,tricarboxylic acid cycle.The results suggested that the iron uptake pathways may represent a novel approach for kojic acid and tea polyphenols synergistically against E.coli O157:H7 and provided a theoretical basis for bacterial pathogen control in the food industry.

Preclinical and clinical evidence of the association of colibactinproducing Escherichia coli with anxiety and depression in colon cancer

BACKGROUND The association between the intestinal microbiota and psychiatric disorders is becoming increasingly apparent.The gut microbiota contributes to colorectal carcinogenesis(CRC),as demonstrated with colibactin-producing Escherichia coli(CoPEC).AIM To evaluate the association between CoPEC prevalence and anxiety-and depressive-like behaviors with both preclinical and clinical approaches.METHODS Patients followed after a CRC surgery and for whom the prevalence of CoPEC has been investigated underwent a psychiatric interview.Results were compared according to the CoPEC colonization.In parallel C57BL6/J wild type mice and mice with a CRC susceptibility were chronically infected with a CoPEC strain.Their behavior was assessed using the Elevated Plus Maze test,the Forced Swimming Test and the Behavior recognition system PhenoTyper®.RESULTS In a limited cohort,all patients with CoPEC colonization presented with psychiatric disorders several years before cancer diagnosis,whereas only one patient(17%)without CoPEC did.This result was confirmed in C57BL6/J wildtype mice and in a CRC susceptibility mouse model(adenomatous polyposis colimultiple intestinal neoplasia/+).Mice exhibited a significant increase in anxiety-and depressive-like behaviors after chronic infection with a CoPEC strain.CONCLUSION This finding provides the first evidence that CoPEC infection can induce microbiota-gut-brain axis disturbances in addition to its procarcinogenic properties.

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.

Prevalence of Drug Resistant Uropathogenic Escherichia coli from Immunocompromised Diabetic Patients Attending Selected Health Facilities in Benue State

Escherichia coli is the commonest bacterial uropathogen of UTIs, the commonest infections in immunocompromised diabetic patients. Better understanding of their main resistance mechanisms to commonly used antibacterial agents will help to reduce the burden of this infection. The prevalence of drug resistant uropathogenic Escherichia coli isolates from immunocompromised diabetic patients attending selected health facilities in Benue State was investigated. Two hundred and ninety-six midstream urine samples were collected for both study and control diabetic patients. Bacterial isolation was done using semi-quantitative method. Drug resistant Escherichia coli were identified as multidrug resistant (MDR), extensive drug resistant (XDR) and pan-drug resistant organisms (PDR). Statistical significance was considered at p E. coli isolates from the study and control subjects with overall prevalence of 20.9% and 8.4% respectively. The isolates were highly resistant to penicillin (ampicillin), monobactam (aztreonam), older quinolone (nalidixic acid) whereas the majority of them showed high susceptibility to aminoglycoside (streptomycin), cephalosporin (cefotaxime) and carbapenem (imipenem). None showed complete susceptibility to all the tested antibiotics. Twenty-five E. coli were identified in this MDR, eight, XDR while 5 were PDR. High numbers of drug resistant E. coli isolates were identified in the study group of which 25 were MDR, 8 XDR while 5 were PDR isolates. High prevalence of UTI and drug resistant isolates occur in diabetic patients with hyperglycemic condition.

Impact of Genetic Diversity of Uropathogenic Escherichia coli and Klebsiella pneumoniae Strains on the Dissemination of Extended Spectrum Beta-Lactam Resistance Genes in Côte d’Ivoire

The increase and spread of bacterial resistance to extended-spectrum beta-lactam antibiotics are reported in many infections and are a real public health problem worldwide. Drug pressure is a factor that favors the emergence of a population of better adapted bacteria. However, there is no literature highlighting the genetic diversity and evolutionary structure of E. coli and K. pneumoniae in an environment with high selection pressure in Côte d’Ivoire. The objective of this study was to evaluate the genetic diversity of E. coli and K. pneumoniae strains circulating at the HKB Hospital in Abobo and at the Daloa Regional Hospital and its impact on the dissemination of extended spectrum beta-lactam resistance genes. A total of 39 strains isolated from the urinary tract of infected patients, including 30 strains of E. coli and 9 strains of K. pneumoniae were studied. A total of 39 strains isolated from the urinary tract of infected patients, including 30 strains of E. coli and 9 strains of K. pneumoniae were studied. From genomic DNA extracts, ESBL resistance genes were amplified by PCR and sequenced, in addition to genetic typing by ERIC-PCR. The data obtained were submitted to genetic and bioinformatics analyses. The results have shown a genetic diversity important in E. coli and K. pneumoniae with diversity indexs (SID) ranging from 0.5 to 0.77. The genetic structure of the bacterial species studied has shown a clonal distribution of strains with clones expressing TEM-9 and CTX-M-15 variants. Also, this clonal structure was correlated with the spread of resistance genes in E. coli and K. pneumoniae. The spread of resistant clones is a factor that might limit the fight against antibiotic resistance.

Phytochemicals of Aloe barbadensis miller as Potential Inhibitors of Uropathogenic Escherichia coli for Urinary Tract Infection Therapy: An in Silico Approach

Urinary tract infections (UTIs) are common infections caused by normal skin or rectum bacteria that get into the urethra and infect the urinary tract. Although the infection can affect various parts of the tract, bladder infections are the most prevalent kind. Uropathogenic Escherichia Coli (UPEC) is the most common pathogen associated with UTI development. Therefore, inhibiting the UPEC protein target (PDB ID: 8BVD) appears to be a promising therapeutic strategy. Therefore, in this study, molecular docking and dynamics were conducted to examine the antibacterial activity of Aloe barbadensis miller against UPEC bacteria. The Aloe barbadensis miller natural compounds licochalcone A, palmidin B and palmidin C were downloaded from PubChem with amoxicillin, which was used as a control drug and studied against the target molecule. The potential parameters examined were the docking scores, absorption, distribution, metabolism, excretion, toxicity (ADMET), bioavailability, root mean square deviation (RMSD), root mean square fluctuation (RMSF), hydrogen bonding, radius of gyration, and potential energy of the system. Docking scores showed that all ligands demonstrated an admirable candidature as an inhibitor to 8BVD molecule, and the score hierarchy is licochalcone A (-6.4 kcal/mol), palmidin C (-6.1 kcal/mol), palmidin B (-6.0 kcal/mol), and amoxicillin (-5.9 kcal/mol). All ligands appeared to have good drug-like properties and oral bioavailability. Molecular dynamic studies showed that all ligands exhibited an excellent nominee as inhibitors in their vicinity at 20 ns. However, there is a relatively high fluctuation of palmidin B compared with other compounds, which seems to be more stable. This work suggests that the selected phytoconstituents could be used as inhibitors of the 8BVD protein in the fight against UTIs. However, further investigation on the clinical and experimental validation of UTI treatment’s specific mechanisms and effects is still welcomed.

Multidrug-Resistant of Escherichia coli and Salmonella spp. Strains in Chicken Feces Intended for Consumption in Open Spaces of Ouagadougou, Burkina Faso

Resistant bacteria can be transmitted to humans through feces or contaminated meat from local chickens. Bacterial strains were isolated from the intestinal contents of 400 local chicken samples from various sales sites. These strains were then characterized using bacteriological and biochemical methods to identify resistant strains. In a study conducted in Ouagadougou, we systematically collected chicken fecal samples from 20 locations across the city, followed by isolation and identification of Salmonella spp. using specific enrichment and culture methods, as well as Escherichia coli. Bacterial strains were characterized using antibiotic resistance profiles were determined through agar diffusion tests, revealing sensitivity or resistance to a range of antibiotics based on established scientific criteria. The results showed that out of the 400 samples collected, 81.25% and 63.5% were contaminated by Escherichia coli and Salmonella spp., respectively. Among these, 86.15% of identified Escherichia coli and 50.78% of Salmonella spp. displayed resistance to at least one tested antibiotic. Among 280 Escherichia coli isolates identified resistant to at least one antibiotic, 31.07% were resistant to cefotaxime (CTX), 20.35% to ceftazidime (CAZ), 21.07% to ceftriaxone (CTR), 75% to amoxicillin clavulanic acid (AMC), 23.57% aztreoname (ATM) and 27.14% were resistant to imipenem (IMP). In the case of the 129 Salmonella spp. isolates resistant to at least one tested antibiotic, 34.88% were resistant to CTX;41.08% to CAZ;35.65% to CTR, 92% to AMC, 39.53% to ATM and finally 47.28% were resistant to IMP. Our study revealed high prevalence of resistance in bacterial strains isolated from local chickens sold outdoors in Ouagadougou. These findings raise significant public health concerns, due to the possible transmission of these resistant strains to humans through the consumption of contaminated meat, thus complicating the treatment of bacterial 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.

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.

Oleanolic acid improved intestinal immune function by activating and potentiating bile acids receptor signaling in E. coli-challenged piglets

Background Infection with pathogenic bacteria during nonantibiotic breeding is one of the main causes of animal intestinal diseases.Oleanolic acid(OA)is a pentacyclic triterpene that is ubiquitous in plants.Our previous work demonstrated the protective effect of OA on intestinal health,but the underlying molecular mechanisms remain unclear.This study investigated whether dietary supplementation with OA can prevent diarrhea and intestinal immune dysregulation caused by enterotoxigenic Escherichia coli(ETEC)in piglets.The key molecular role of bile acid receptor signaling in this process has also been explored.Results Our results demonstrated that OA supplementation alleviated the disturbance of bile acid metabolism in ETEC-infected piglets(P<0.05).OA supplementation stabilized the composition of the bile acid pool in piglets by regulating the enterohepatic circulation of bile acids and significantly increased the contents of UDCA and CDCA in the ileum and cecum(P<0.05).This may also explain why OA can maintain the stability of the intestinal microbiota structure in ETEC-challenged piglets.In addition,as a natural ligand of bile acid receptors,OA can reduce the severity of intestinal inflammation and enhance the strength of intestinal epithelial cell antimicrobial programs through the bile acid receptors TGR5 and FXR(P<0.05).Specifically,OA inhibited NF-κB-mediated intestinal inflammation by directly activating TGR5 and its downstream c AMP-PKA-CREB signaling pathway(P<0.05).Furthermore,OA enhanced CDCA-mediated MEK-ERK signaling in intestinal epithelial cells by upregulating the expression of FXR(P<0.05),thereby upregulating the expression of endogenous defense molecules in intestinal epithelial cells.Conclusions In conclusion,our findings suggest that OA-mediated regulation of bile acid metabolism plays an important role in the innate immune response,which provides a new diet-based intervention for intestinal diseases caused by pathogenic bacterial infections in piglets.

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.

E.coli HPI通过NLRP3/caspase-1信号通路诱导细胞焦亡而促进肠道炎症

目的:探究大肠埃希菌(Escherichia coli,E.coli)强毒力岛(high-pathogenicity island,HPI)对细胞焦亡及肠道炎症的影响。方法:用含HPI的E.coli株(HPI+)、HPI缺失的E.coli株(△HPI)和脂多糖(lipopolysaccharide,LPS)分别处理昆明小鼠和IPEC-J2细胞(猪小肠上皮细胞)。测定小鼠肠道乳酸脱氢酶(lactate dehydrogenase,LDH)活性、超氧化物歧化酶(superoxide dismutase,SOD)活性、IgA表达和分泌性IgA(secretory IgA,SIgA)含量;HE和TUNEL染色观察肠道损伤;RT-qPCR、免疫组织化学染色和Western blot检测小鼠肠组织和IPEC-J2细胞中核苷酸结合寡聚化结构域样受体蛋白3(nucleotide-binding oligomerization domain-like receptor protein 3,NLRP3)/caspase-1信号通路关键调控点的表达;ELISA检测小鼠血清和IPEC-J2细胞培养上清液中白细胞介素1β(interleukin-1β,IL-1β)和IL-18含量;共聚焦显微镜观察NLRP3与caspase-1的共定位。siRNA沉默IPEC-J2细胞的NLRP3,验证NLRP3在E.coli HPI感染中的关键调控功能。结果:相对于△HPI感染,E.coli HPI显著降低小鼠肠道SOD活性,增加IgA+B细胞,并促进LDH的释放和SIgA的分泌;ELISA、HE染色和TUNEL染色结果显示,E.coli HPI促进了小鼠肠道上皮细胞DNA断裂、组织损伤和炎症;Western blot结果显示,相对于△HPI,E.coli HPI感染使得小鼠肠道消皮素D的N端片段(gasdermin D N-terminal fragment,GSDMD-N)蛋白水平显著升高;RT-qPCR和免疫组织化学染色结果显示,E.coli HPI显著促进了小鼠肠道和IPEC-J2细胞中NLRP3、含caspase募集结构域的凋亡相关斑点样蛋白(apoptosis-associated speck-like protein containing a caspase recruitment domain,ASC)、caspase-1、GSDMD、IL-1β和IL-18的mRNA和蛋白表达,IL-1β和IL-18分泌增加;共聚焦显微镜观察发现,相对于△HPI感染,E.coli HPI显著促进了NLRP3炎症小体的组装,使得NLRP3与caspase-1发生共定位;此外,在NLRP3沉默的IPEC-J2细胞中观察到E.coli HPI诱导的细胞炎症、细胞损伤及NLRP3/caspase-1信号通路的激活被缓解。结论:HPI的存在增强了E.coli的毒力水平,促进肠道炎症;NLRP3/caspase-1信号通路调控的细胞焦亡参与了E.coli HPI诱导的肠道损伤。

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.

亚胺培南对携带bla_(NDM-1)耐药基因的Escherichia coli耐药性及内膜secY、secE和secG转录水平的影响

目的探讨亚胺培南(IPM)对bla_(NDM-1)阳性Escherichia coli耐药性及其内膜secY、secE和secG转录水平的影响。方法以重组质粒pET28a(+)-bla_(NDM-1)转化菌株E.coli DH5α-bla_(NDM-1)和E.coli BL21(DE3)-bla_(NDM-1)为研究对象,在梯度浓度增加或撤消IPM暴露下传代培养菌株,检测17种抗生素对IPM暴露菌株的MIC值;SDS-PAGE凝胶电泳检测NDM-1表达;蛋白活性实验检测NDM-1活性;qRT-PCR检测bla_(NDM-1)及内膜secY、secE和secG转录水平。结果12μg/mL和0_(20)μg/mL IPM暴露的E.coli DH5α-bla_(NDM-1)菌株CFZ、CXM、FOX、CRO、CAZ、FEP等头孢类药物的MIC值(≥8μg/mL/≥8μg/mL、≥32μg/mL/≥32μg/mL、≥32μg/mL/≥32μg/mL、≥64μg/mL/=32μg/mL、≥32μg/mL/≥32μg/mL、≥32μg/mL/=8μg/mL)与0μg/mL IPM暴露MIC值(≤2μg/mL、=16μg/mL、≤8μg/mL、≤1μg/mL、≤4μg/mL、≤2μg/mL)相比均显著增高,而IPM和MEM的MIC值与0μg/mL IPM暴露(≤1μg/mL和≤1μg/mL)相比也显著增高(=8μg/mL/=8μg/mL和=4μg/mL/=4μg/mL)。12μg/mL和0_(20)μg/mL IPM暴露的E.coli BL21(DE3)-bla_(NDM-1)菌株FOX、CRO、FEP等头孢类药物的MIC值(≥32μg/mL/≥32μg/mL、≥64μg/mL/≥64μg/mL、=16μg/mL/=16μg/mL)与0μg/mL IPM暴露(≤8μg/mL、≤1μg/mL、≤2μg/mL)相比也均显著增高,而IPM和MEM的MIC值与0μg/mL IPM暴露(≤1μg/mL和≤1μg/mL)相比也均显著增高(≥16μg/mL/≥16μg/mL和≥16μg/mL/≥16μg/mL)。SDS-PAGE显示,随12μg/mL IPM暴露时间的延长,菌株NDM-1水解IPM活性增加。qRT-PCR显示,12μg/mL IPM暴露的E.coli DH5α-bla_(NDM-1)和E.coli BL21(DE3)-bla_(NDM-1)的bla_(NDM-1)、secY、secE和secG转录水平分别上调2.31/2.5、3.05/1.96、2.83/1.24和2.71/1.45倍。结论IPM可使bla_(NDM-1)阳性E.coli由碳青霉烯类敏感变为耐药且保持稳定,细菌内膜SecYEG跨膜通道蛋白参与菌株耐药性调控,这为认识抗生素压力下肠杆菌科细菌耐药性变化及指导临床合理用药提供理论支撑。

E.coli O78和LAB对牦牛肠道上皮细胞MAPK/Zonulin通路的影响

为探究牦牛致病性大肠杆菌(E.coli O78)和乳酸杆菌(LAB)对牦牛肠道上皮屏障MAPK/Zonulin通路的影响作用,采用牦牛上皮细胞作为实验模型,Transwell构建单层上皮细胞屏障,设立阴性对照组(Control组)、阳性对照组(Model组,加入1×10^(5)CFU E.coli O78)、低剂量组乳酸杆菌组(LLAB组,加入1×10^(5)CFU E.coli O78和1×10^(5)CFU LAB)、高剂量乳酸杆菌组(HLAB组,加入加入1×10^(5)CFU E.coli O78和1×10^(7)CFU LAB),采用RT-qPCR和Western-blot技术测定各组P38、ERK、JNK、Zonulin的基因和蛋白表达水平。结果:与Control组相比,Model组的P38、ERK、JNK、Zonulin的基因表达水平极显著上升(P<0.01),而其蛋白表达水平呈现显著上升的趋势(P<0.05);与Model组相比,LLAB组P38的基因表达水平极显著下降(P<0.01),ERK、JNK、Zonulin的基因表达水平表现显著下降的趋势(P<0.05),P38、ERK、Zonulin的蛋白表达水平则呈现极显著下降的趋势(P<0.01),JNK的蛋白表达水平表现为显著下降(P<0.05);与Model组相比,HALB组P38、ERK、JNK的基因的表达水平表现为显著下降(P<0.05),Zonulin的基因表达水平表现为极显著下降(P<0.01),P38、ERK、JNK、Zonulin的蛋白表达水平呈现极显著下降的趋势(P<0.01)。结论:E.coli O78可通过上调MAPK通路关键基因P38、ERK、JNK的基因表达水平,同时上调Zonulin蛋白使肠道屏障受损;LAB可以减轻E.coli O78引起的肠道屏障受损,这一作用是通过下调MAPK通路关键基因P38、ERK、JNK的基因表达水平,同时下调Zonulin蛋白表达水平来实现的。

Phytogenic feed additives alleviate pathogenic Escherichia coli-induced intestinal damage through improving barrier integrity and inhibiting inflammation in weaned pigs

Background:This study was conducted to investigate the effects of each phytogenic feed additive(PFA;PFA1,bitter citrus extract;PFA2,a microencapsulated blend of thymol and carvacrol;PFA3,a mixture of bitter citrus extract,thymol,and carvacrol;PFA4,a premixture of grape seed,grape marc extract,green tea,and hops;PFA5,fenugreek seed powder)on the growth performance,nutrient digestibility,intestinal morphology,and immune response in weaned pigs infected with Escherichia coli(E.coli).Results:A total of 634-week-old weaned pigs were placed in individual metabolic cages and assigned to seven treatment groups.The seven treatments were as follows:1)NC;basal diet without E.coli challenge,2)PC;basal diet with E.coli challenge,3)T1;PC+0.04%PFA1,4)T2;PC+0.01%PFA2,5)T3;PC+0.10%PFA3,6)T4;PC+0.04%PFA4,7)T5;PC+0.10%PFA5.The experiments lasted in 21 d,including 7 d before and 14 d after the first E.coli challenge.In the E.coli challenge treatments,all pigs were orally inoculated by dividing a total of 10 mL of E.coli F18 for 3 consecutive days.The PFA-added groups significantly increased(P<0.05)average daily gain and feed efficiency and decreased(P<0.05)the fecal score at d 0 to 14 post-inoculation(PI).Tumor necrosis factorαwas significantly lower(P<0.05)in the PFA-added groups except for T1 in d 14 PI compared to the PC treatment.The T3 had a higher(P<0.05)immunoglobulin G and immunoglobulin A concentration compared to the PC treatment at d 7 PI.Also,T3 showed significantly higher(P<0.05)villus height:crypt depth and claudin 1 expression in ileal mucosa,and significantly downregulated(P<0.05)the expression of calprotectin compared to the PC treatment.Conclusions:Supplementation of PFA in weaned pigs challenged with E.coli alleviated the negative effects of E.coli and improved growth performance.Among them,the mixed additive of bitter citrus extract,thymol,and carvacrol showed the most effective results,improving immune response,intestinal morphology,and expression of tight junctions.

Characterization of a bla_(CTX-M-3),bla_(KPC-2)and bla_(TEM-1B)co-producing IncN plasmid in Escherichia coli of chicken origin

An extensively drug-resistant(XDR)Escherichia coli strain 258E was isolated from an anal swab sample of a chicken farm of Anhui province in China.Genomic analyses indicated that the strain 258E harbors an incompatibility group N(IncN)plasmid pEC258-3,which co-produces bla_(CTX-M-3),bla_(KPC-2),bla_(TEM-1B),qnrS1,aac(6')-Ib-cr,dfrA14,arr-3,and aac(6')-Ib3.Multiple genome arrangement analyses indicated that pEC258-3 is highly homologous with pCRKP-1-KPC discovered in Klebsiella pneumoniae from a patient.Furthermore,conjugation experiments proved that plasmid pEC258-3 can be transferred horizontally and may pose a significant potential threat in animals,community and hospital settings.