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Carbapenems are the last resort antimicrobials for the treatment of extended spectrum β-lactamases (ESBLs) producing Enterobacteriaceae. Emergence of carbapenems resistant group B2 uropathogenic E. coli (UPEC) is a major concern because of their high virulence. Prevalence of these enzymes and multidrug resistance (MDR) among B2 UPEC isolates from Iraqi outpatients with acute urinary tract infection (UTI) was evaluated in this research. Urine cultures were performed and the isolates were identified biochemically. Escherichia coli isolates were tested for phylogroup reference by quadraplex PCR, then B2 isolates were detected for antimicrobial resistance by disc diffusion test and carbapenemase genes by PCR. Escherichia coli was the most prevalent among Gram-negative isolates (66.6%) and B2 was the most detected phylogroup among E. coli isolates (33.9%). Most of B2 isolates showed high resistance rates to tested antimicrobials, especially β-lactams with MDR revealed in 100% of them. Whereas, low resistance rates were noted against carbapenems, aminoglycosides and nitrofurantoin. Carbapenemase genes were detected in 76.3% of B2 isolates. Of which, blaOXA-48 was the most frequent (57.8%), followed by blaPER (47.3%), blaKPC (15.7%), blaVEB and blaVIM (10.5%, for each). Whereas, blaGES and blaIMP genes were not found. Coproduction of these genes occurred among 17 isolates. The combination of blaOXA-48 and blaPER was the most frequent (41.1%). All carbapenemase producing isolates were MDR. These results revealed high prevalence of carbapenemase genes and MDR among B2 UPEC recovered in this study. In the study area. it is strongly advised to use aminoglycosides and nitrofurantoin for empirical treatment of UPEC., Competing Interests: The authors declared that no competing interest exist.

Extended-Spectrum Beta-Lactamase (ESBL) producing Enterobacteriaceae (EPE) is increasing worldwide, though less documented in low-income settings. Here we determined the prevalence of EPE infection and carriage, and patient factors associated with EPE-carriage among pediatric patients in three health care levels in Tanzania. Between January and April 2016, 350 febrile children (median age 21 months) seeking care at a university or a regional referral hospital, or a health centre in Moshi municipality, Tanzania, were included. Socio-demographic characteristics were collected using a questionnaire. Rectal swabs and blood cultures were collected from all children (n = 350) and urinary samples from 259 children at admission. ESBL-phenotype and antimicrobial susceptibility were determined for Klebsiella pneumoniae (K. pneumoniae) and Escherichia coli (E. coli) isolates. Only one EPE case (E. coli) in blood and four in urine (one E. coli and three K. pneumoniae) were found, whereas (n = 90, 26%) of the children were colonized in feces (ESBL-E. coli; n = 76, ESBL-K. pneumoniae, n = 14). High resistance rates were seen in fecal ESBL-E. coli (n = 76) against trimethoprim-sulfamethoxazole (n = 69, 91%), gentamicin (n = 51, 67%), ciprofloxacin (n = 39, 51%) and chloramphenicol (n = 27, 35%) whereas most isolates were sensitive to amikacin (n = 71, 93%). Similar rates were seen for fecal ESBL-K. pneumoniae. Resistance to first line antibiotics were also very high in fecal E. coli not producing ESBL. No sociodemographic factor was associated with EPE-carriage. Children colonized with EPE were younger than 12 months (n = 43, 48%) and often treated with antibiotics (n = 40, 44%) in the previous two months. After adjustment for age children admitted to the intensive care unit had higher odds of EPE fecal carriage compared with those in the general wards (OR = 3.9, 95%CI = 1.4-10.4). Despite comparatively high rates of fecal EPE-carriage and previous antibiotic treatment, clinical EPE cases were rare in the febrile children. The very high resistant rates for the EPE and the non-ESBL producing E. coli to commonly used antibiotics are worrying and demand implementation of antibiotic stewardship programs in all levels of health care in Tanzania., (© 2021. The Author(s).)

Objective: To determine the epidemiology, risk factors, and prognosis of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae bloodstream infections (BSIs) among hematology malignancy (HM) patients in China. Method: From January 2010 to June 2018, we retrospectively collected and analyzed the 449 HM patients with E. coli or K. pneumoniae BSIs from three leading hospitals in Hunan Province, China. Results: Two hundred four (45.4%) patients harbored ESBL-producing bacteremia. The proportion of ESBL-producing bacteremia increased significantly with the growth of the year, with a ratio of 34.47% in 2010-2014 to 54.7% in 2015-2018. Comparing with non-ESBL groups in HM patients, central venous catheter (odds ratio [OR] 1.717, p  = 0.009), previous antibiotic exposure (OR 1.559, p  = 0.035), and E. coli (OR 2.561, p  ≤ 0.001) among ESBL groups were independent risk factors. No significant differences in 30-day mortality were tested in patients with BSI caused by ESBL-producing or non-ESBL-producing E. coli and K. pneumoniae (17.1% vs. 16.7%; p  = 0. 893). The proportion of carbapenem used within 72 hours after the onset of bacteremia in two groups was high, which was routinely used as "last-resort drugs" in Gram-negative bacterial infections. Risk factors associated with 30-day mortality in HM patients with E. coli or K. pneumoniae bacteremia were myelodysplastic syndrome, incomplete remission of the disease, Multinational Association of Supportive Care in Cancer score <21, Pitt bacteremia score ≥4, Charlson comorbidity score >3, catheter insertion, use of vasopressors, and inappropriate antibiotics within 72 hours of BSI onset. Conclusions: The results of this study may provide some references for the whole process management of HM patients with BSIs.

Ribonucleotide reductases (RNRs) catalyze the de novo conversion of nucleotides to deoxynucleotides in all organisms, controlling their relative ratios and abundance. In doing so, they play an important role in fidelity of DNA replication and repair. RNRs' central role in nucleic acid metabolism has resulted in five therapeutics that inhibit human RNRs. In this review, we discuss the structural, dynamic, and mechanistic aspects of RNR activity and regulation, primarily for the human and Escherichia coli class Ia enzymes. The unusual radical-based organic chemistry of nucleotide reduction, the inorganic chemistry of the essential metallo-cofactor biosynthesis/maintenance, the transport of a radical over a long distance, and the dynamics of subunit interactions all present distinct entry points toward RNR inhibition that are relevant for drug discovery. We describe the current mechanistic understanding of small molecules that target different elements of RNR function, including downstream pathways that lead to cell cytotoxicity. We conclude by summarizing novel and emergent RNR targeting motifs for cancer and antibiotic therapeutics.

Carbonic anhydrase II knockout ( Car2 -/- ) mice have depleted numbers of renal intercalated cells, which are increasingly recognized to be innate immune effectors. We compared pyelonephritis susceptibility following reciprocal renal transplantations between Car2 -/- and wild-type mice. We examined the effect of pharmacological CA suppression using acetazolamide in an experimental murine model of urinary tract infection. Car2 -/- versus wild-type mice were compared for differences in renal innate immunity. In our transplant scheme, mice lacking CA-II in the kidney had increased pyelonephritis risk. Mice treated with acetazolamide had lower kidney bacterial burdens at 6 h postinfection, which appeared to be due to tubular flow from diuresis because comparable results were obtained when furosemide was substituted for acetazolamide. Isolated Car2 -/- kidney cells enriched for intercalated cells demonstrated altered intercalated cell innate immune gene expression, notably increased calgizzarin and insulin receptor expression. Intercalated cell number and function along with renal tubular flow are determinants of pyelonephritis risk.

Plasmid-mediated extended-spectrum β-lactamase and AmpC β-lactamase (ESBL/pAmpC) producing bacteria are present at all levels of the broiler production pyramid. Young birds can be found positive for ESBL/pAmpC-producing Escherichia coli shortly after arrival at farm. The aim of this study was to determine the effect of different challenge doses of ESBL/pAmpC-producing E. coli on time-until-colonization and the level of excretion in young broilers. One-day-old broilers (specific-pathogen free (SPF) and conventional Ross 308) were housed in isolators and challenged with 0.5 ml ESBL/pAmpC-producing E. coli strains of varying doses (10 1 -10 5  CFU/ml). Presence and concentration (CFU/gram feces) of ESBL/pAmpC-producing E. coli and total E. coli were determined longitudinally from cloacal swabs, and in cecal content 72 h after challenge. Higher challenge doses resulted in shorter time-until-colonization. However, even the lowest dose (10 1  CFU/ml) resulted in colonization of the broilers which excreted >10 6  CFU/gram feces 72 h after inoculation. Conventional broilers were colonized later than SPF broilers, although within 72 h after challenge all broilers were excreting ESBL/pAmpC-producing E. coli. A probabilistic model was used to estimate the probability of colonization by initial inoculation or transmission. The higher the dose the higher the probability of excreting ESBL/pAmpC-producing E. coli as a result of inoculation. In conclusion, low initial doses of ESBL/pAmpC-producing E. coli can result in rapid colonization of a flock. Interventions should thus be aimed to eliminate ESBL/pAmpC-producing bacteria in the environment of the hatchlings and measures focusing at reducing colonization and transmission of ESBL/pAmpC-producing E. coli should be applied shortly after hatching., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Antibiotic resistance in bacteria is an emerging problem globally. Resistant bacteria are found in human and animal microbiota, as well as in the environment. Wastewater receives bacteria from all these sources and thus can provide a measurement of abundance and diversity of antibiotic-resistant bacteria circulating in communities. In this study, water samples were collected from a wastewater pump station in a Norwegian suburban community over a period of 15 months. A total of 45 daily samples were cultured and analyzed for the presence of Escherichia coli Eighty E. coli -like colonies were collected from each daily sample and then phenotyped and analyzed for antibiotic resistance using the PhenePlate-AREB system. During the sampling period, two unique E. coli phenotypes with resistance to cefotaxime and cefpodoxime indicating carriage of extended-spectrum β-lactamases (ESBL) were observed repeatedly. Whole-genome sequencing of 15 representative isolates from the two phenotypes identified these as two distinct clones belonging to the two globally spread E. coli multilocus sequence types (STs) ST131 and ST648 and carrying bla CTX-M-15 The number of ESBL-positive E. coli strains in the community wastewater pump station was 314 of 3,123 (10%) analyzed E. coli strains. Of the ESBL-positive isolates, 37% belonged to ST648, and 7% belonged to ST131. Repeated findings of CTX-M-15-positive ST648 and ST131 over time indicate that these STs are resident in the analyzed wastewater systems and/or circulate abundantly in the community., (Copyright © 2019 Paulshus et al.)

Background: Evidence suggests that antimicrobial peptides, components of the innate immune response, protect the kidneys and bladder from bacterial challenge. We previously identified ribonuclease 7 (RNase 7) as a human antimicrobial peptide that has bactericidal activity against uropathogenic Escherichia coli (UPEC). Functional studies assessing RNase 7's contributions to urinary tract defense are limited., Methods: To investigate RNase 7's role in preventing urinary tract infection (UTI), we quantified urinary RNase 7 concentrations in 29 girls and adolescents with a UTI history and 29 healthy female human controls. To assess RNase 7's antimicrobial activity in vitro in human urothelial cells, we used siRNA to silence urothelial RNase 7 production and retroviral constructs to stably overexpress RNase 7; we then evaluated UPEC's ability to bind and invade these cells. For RNase 7 in vivo studies, we developed humanized RNase 7 transgenic mice, subjected them to experimental UTI, and enumerated UPEC burden in the urine, bladder, and kidneys., Results: Compared with controls, study participants with a UTI history had 1.5-fold lower urinary RNase 7 concentrations. When RNase 7 was silenced in vitro , the percentage of UPEC binding or invading human urothelial cells increased; when cells overexpressed RNase 7, UPEC attachment and invasion decreased. In the transgenic mice, we detected RNase 7 expression in the kidney's intercalated cells and bladder urothelium. RNase 7 humanized mice exhibited marked protection from UPEC., Conclusions: These findings provide evidence that RNase 7 has a role in kidney and bladder host defense against UPEC and establish a foundation for investigating RNase 7 as a UTI prognostic marker or nonantibiotic-based therapy., (Copyright © 2019 by the American Society of Nephrology.)

Antibiotic stewardship programs have traditionally focused on reducing hospital antibiotic use. However, reducing community antibiotic prescribing could have substantial impacts in both hospital and community settings. We developed a deterministic model of transmission of extended-spectrum beta-lactamase-producing Escherichia coli in both the community and hospitals. We fit the model to existing, national-level antibiotic use and resistance prevalence data from Sweden. Across a range of conditions, a given relative change in antibiotic use in the community had a greater impact on resistance prevalence in both the community and hospitals than an equivalent relative change in hospital use. However, on a per prescription basis, changes in antibiotic use in hospitals had the greatest impact. The magnitude of changes in prevalence were modest, even with large changes in antimicrobial use. These data support the expansion of stewardship programs/interventions beyond the walls of hospitals, but also suggest that such efforts would benefit hospitals themselves., (© The Author(s) 2018. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Carbapenemase-producing Enterobacteriaceae (CPE) are well known to cause many serious infections resulting in increasing mortality rate, treatment cost, and prolonged hospitalization. Among the widely recognized types of carbapenemases, New Delhi β -lactamase (NDM) and Klebsiella pneumoniae carbapenemase (KPC) are the most important enzymes. However, in Vietnam, there are only scattered reports of CPE due to the lack of simple and affordable methods that are suitable to laboratory conditions. This study aims to survey the characteristics of carbapenem-resistant E. coli and K. pneumoniae (CR-E/K) at two hospitals in Southern Vietnam and perform some simple methods to detect the two enzymes. A total of 100 CR-E/K strains were collected from clinical isolates of Gia Dinh People's Hospital and Dong Nai General Hospital, Vietnam, from November 2017 to May 2018. The patient-related information was also included in the analysis. We conducted real-time polymerase chain reaction (PCR), Modified Hodge Test (MHT), and combined disk test (CDT) on all isolates. Carbapenemase-encoding genes were detected in 47 isolates (36 NDM, 10 KPC, and one isolate harboring both genes). The E. coli strain carrying simultaneously these two genes was the first case reported here. Most of isolates were collected from patients in ICU, Infectious Disease Department, and Department of Urologic Surgery. Urine and sputum were two common specimens. The true positive rate (sensitivity, TPR) and specificity (SPC) of the imipenem-EDTA (ethylen diamine tetra acetic acid) for NDM detection and the imipenem-PBA (phenylboronic acid) for KPC detection on E. coli were 93.8%, 97.1% and 66.7%, 95.7%, respectively. Meanwhile, the imipenem-EDTA for NDM detection and the imipenem-PBA for KPC detection among K. pneumonia achieved 90.5%, 100% and 100%, 92.9% TPR and SPC, respectively. However, MHT showed low sensitivity and specificity. Our findings showed that CP-E/K were detected with high prevalence in the two hospitals. We suggest that CDT can be used as a low-priced and accurate method of detection.

The rapid emergence of multiresistant microbial pathogens, dubbed superbugs, is a serious threat to human health. Extended spectrum beta lactamase (ESBL)-producing Escherichia coli is a superbug causing worldwide outbreaks, necessitating timely and accurate tracking of resistant strains. Accordingly, this study was designed to investigate the spread of ESBL-producing Escherichia coli isolates, to analyze the effect of different genotypic and phenotypic factors on in vitro resistance patterns, and to assess the diagnostic value of commonly used ESBL genetic markers. For that purpose, we cultured 250 clinical isolates and screened their susceptibility to beta-lactam antibiotics. Among 12 antibiotics screened, only imipenem seems to have remained resilient. We subsequently analyzed the ESBL phenotype of Escherichia coli isolates and examined potential associations between their resistance phenotypes and patient-related factors. ESBL genotyping of 198 multiresistant isolates indicated that 179 contained at least one bla CTX-M gene. As we statistically dissected the data, we found associations between overall resistance and body site / type of disease. Additionally, we confirmed the diagnostic value of testing both bla CTX-M-1 and bla CTX-M-15 in providing better prediction of overall resistance. Finally, on sequencing the amplification products of detected bla CTX-M genes, we discovered two novel variants, which we named bla CTX-M-14.2 and bla CTX-M-15.2.

Beta (β)-lactamases are the most important agents that confer drug resistance among gram-negative bacteria. Continuous mutations in β-lactamases make them remarkably diverse. We carried out the transcriptome analysis of 10 β-lactamase genes of Extended-Spectrum β-lactamases (ESBL), Metallo β-lactamases (MBL), and AmpC β-lactamases (ABL) in drug-resistant and sensitive diarrheagenic E. coli (DEC) isolates obtained from children up to 5 years of age. Out of the 10 β-lactamase genes, four belonged to ESBL (TEM, SHV, CTX, and OXA); three to MBL (NDM-1, IMP, and VIM); and three to ABL (ACT, DHA and CMY) class of genes. The different categories of DEC were estimated for β-lactamases production using a set of conventional phenotypic tests, followed by detection of their messenger RNA (mRNA) expression. The study revealed a direct correlation between mRNA expression of these genes and the presence of antibiotic resistance; also corroborated by mutation analysis of the AmpC promoter region. All the 10 β-lactamase genes showed a significant increase in their expression levels in resistant isolates, compared to those of the sensitive isolates, indicating their possible role in the disease pathogenesis. Increase in mRNA expression of β-lactamase genes, and thereby virulence, may be due to multifactorial parameters causing phenotypic as well as genotypic changes. Our study highlights the necessity of instantaneous detection of β-lactamase gene expression to curb the overwhelming threat posed by emergence of drug resistance amongst the commensal E. coli strains in children from developing countries for larger public health interest.

Objectives: Drug resistance among gram-negative bacteria is a worldwide challenge. Due to the importance of drug-resistant Klebsiella pneumoniae and Escherichia coli strains in hospital-acquired infections, we aimed to determine the phenotypic and genotypic characteristics of ESBL-, AmpC-, and carbapenemase-producing isolates obtained from hospitalized patients in Tehran and Ilam (Iran)., Materials and Methods: In total, 90 K. pneumoniae isolates and 65 E. coli isolates were collected from various infections. Phenotypic identification of bacterial isolates was performed using standard methods. Phenotypic screening of ESBL, AmpC, and carbapenemase enzymes was carried out. Detection of ESBL, AmpC, and carbapenemase genes was also performed by the PCR method., Results: Phenotypic detection tests showed that 36 (40%) K. pneumoniae and 23 (35.4%) E. coli isolates were ESBL producers. Moreover, 18 (20%) and 6 (9.2%) K. pneumoniae and E. coli isolates were AmpC producers, respectively. Modified Hodge test results indicated that 39 (43.3%) K. pneumoniae and 18 (27.7%) E. coli isolates produced carbapenemase. Molecular tests showed that 40% of K. pneumoniae and 36.9% of E. coli isolates were ESBL positive. AmpC was detected in 24.4 and 13.8% of K. pneumoniae and E. coli isolates. Carbapenemase was detected in 34 (37.8%) K. pneumoniae and 13 (20%) E. coli isolates. -Conclusion: In this study, 3 K. pneumoniae isolates simultaneously carried ESBL, AmpC, and carbapenemase genes. Up-to-date strategies such as combination therapy or utilization of new antimicrobial agents might help to combat such drug-resistant organisms., (© 2019 The Author(s) Published by S. Karger AG, Basel.)

Sepsis is a life-threatening inflammatory syndrome accompanying a bloodstream infection. Frequently secondary to pathogenic bacterial infections, sepsis remains difficult to treat as a singular disease mechanism. We compared the pathogenesis of murine sepsis experimentally elicited by five bacterial pathogens and report similarities among host responses to Gram-negative Salmonella and E. coli. We observed that a host protective mechanism involving de-toxification of lipopolysaccharide by circulating alkaline phosphatase (AP) isozymes was incapacitated during sepsis caused by Salmonella or E. coli through activation of host Toll-like receptor 4, which triggered Neu1 and Neu3 neuraminidase induction. Elevated neuraminidase activity accelerated the molecular aging and clearance of AP isozymes, thereby intensifying disease. Mice deficient in the sialyltransferase ST3Gal6 displayed increased disease severity, while deficiency of the endocytic lectin hepatic Ashwell-Morell receptor was protective. AP augmentation or neuraminidase inhibition diminished inflammation and promoted host survival. This study illuminates distinct routes of sepsis pathogenesis, which may inform therapeutic development., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Enteropathogenic Escherichia coli (EPEC) belongs to a group of enteric human pathogens known as attaching-and-effacing (A/E) pathogens, which utilize a type III secretion system (T3SS) to translocate a battery of effector proteins from their own cytoplasm into host intestinal epithelial cells. Here we identified EspH to be an effector that prompts the recruitment of the tetraspanin CD81 to infection sites. EspH was also shown to be an effector that suppresses the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (Erk) signaling pathway at longer infection times. The inhibitory effect was abrogated upon deletion of the last 38 amino acids located at the C terminus of the protein. The efficacy of EspH-dependent Erk suppression was higher in CD81-deficient cells, suggesting that CD81 may act as a positive regulator of Erk, counteracting Erk suppression by EspH. EspH was found within CD81 microdomains soon after infection but was largely excluded from these domains at a later time. Based on our results, we propose a mechanism whereby CD81 is initially recruited to infection sites in response to EspH translocation. At a later stage, EspH moves out of the CD81 clusters to facilitate effective Erk inhibition. Moreover, EspH selectively inhibits the tumor necrosis factor alpha (TNF-α)-induced Erk signaling pathway. Since Erk and TNF-α have been implicated in innate immunity and cell survival, our studies suggest a novel mechanism by which EPEC suppresses these processes to promote its own colonization and survival in the infected gut., (Copyright © 2018 American Society for Microbiology.)

The aim of this study was to identify risk factors for extended-spectrum β-lactamase (ESBL)-producing Escherichia coli (E coli) bloodstream infection (BSI) among carriers hospitalized between March 2011 and June 2016 at the ICU of the West China Hospital.The cases were patients with at least 1 episode of ESBL-producing E coli BSI within 1 week after a positive rectal swab. Controls were selected randomly 1:2 among ESBL-producing E coli rectal carriers who did not develop BSI.Among 19,429 ICU patients, 9015 (46.4%) had a positive rectal swab for ESBL-producing E coli. Of them, 42 (0.5%) were diagnosed with ESBL-producing E coli BSI. The in-hospital mortality was higher for the BSI patients compared with controls (19.1% vs. 6.0%, P = .031). In the past 72 hours, patients in case group were more likely to use penicillin (odds ratio [OR] = 12.076; 95% confidence interval [CI]: 1.397-104.251, P = .02), cephalosporin (OR = 6.900; 95% CI: 1.493-31.852, P = .01), and carbapenem (OR = 5.422; 95% CI: 1.228-23.907, P = .03) as compared to patients in control group. Also, when compared to patients in control group, patients in case group were likely to stay for a longer time in ICU before positive rectal swab test (OR = 1.041, 95% CI: 1.009-1.075, P = .01) and have higher maximum body temperature before positive rectal swab (OR = 8.014; 95% CI: 2.408-26.620, P = .001).Bacteremia owing to ESBL-producing E coli was associated with high antimicrobial exposure, hospital stay, and maximum body temperature.

Objectives: The incidence of urinary tract infection (UTI) due to extended-spectrum β-lactamase (ESBL)-producing Escherichia coli has increased over recent years. Initial empirical therapy is often ineffective for these resistant isolates resulting in prolonged hospitalization and increased mortality. This study was conducted to determine the risk factors of UTI caused by ESBL E. coli in the emergency department (ED)., Methods: This is a retrospective case-control study at a university hospital in Korea with UTI patients who visited ED between June 2015 and December 2016. We compared case patients with ESBL E. coli UTI (n = 50) to control patients with non-ESBL-producing E. coli UTI (n = 100), which were matched for age and sex. Multivariate logistic regression analysis was used to explore risk factors., Results: Our study showed that hospital-acquired infection (OR = 3.86; 95% CI = 1.26-11.8; p = .017), prior UTI within 1 year (OR = 3.26; 95% CI = 1.32-8.05; p = .010), and underlying cerebrovascular disease (OR = 3.24; 95% CI = 1.45-7.25; p = .004) were independent risk factors for acquisition of ESBL-producing E. coli. Notably, 35 (70%) out of 50 case patients had community-acquired infection, and 68% and 54% of ESBL E. coli were resistance to ciprofloxacin and trimethoprim-sulfamethoxazole, respectively. On the contrary, 98% of ESBL E. coli was susceptible to amikacin., Conclusion: The main risk factors identified in our study should be considered when treating UTI patients in ED. Amikacin may improve the outcome of empirical treatment without increasing carbapenem utilization., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Carbapenems are the preferred β-lactamics for treatment for infections caused by enterobacteria producing extended-spectrum β-lactamases (ESBL); however, clinical studies show effectiveness of piperacillin/tazobactam in certain infections by Escherichia coli ESBL producers., Aim: To determine the clinical and micro-biological cure with piperacillin/tazobactam in patients with infections caused by E. coli ESBL producers, CTXM type., Methods: Retrospective descriptive study with adults hospitalized in a university hospital. We included urinary tract infections (UTI), intra-abdominal infections (IAI), soft tissue infections (STI) and/or bacteremia., Results: We studied 40 patients, where 65% corresponded to UTI, 25% to IAI and 10% were STI. The overall clinical cure was achieved in 89.4%, with the best results in the ITU (100%), followed by STI (80%) and 70% in IAI. The 85% of the strains had minimum inhibitory concentrations (MIC) ≤8 μg/ml and 70% with MIC ≤4 μg/mL, however the rate of failure were high in intra-abdominal infections with high inocula or not controlled; CTX-M-15 was found in the 62.5%., Conclusions: Piperacillin/tazobactam was efficient to obtain clinical and microbiological cure in patients with infections caused by ESBL producers but susceptible E. coli, especially in UTI and STI and to a lesser extent in IAI.

The execution of shock following high dose E. coli lipopolysaccharide (LPS) or bacterial sepsis in mice required pro-apoptotic caspase-8 in addition to pro-pyroptotic caspase-11 and gasdermin D. Hematopoietic cells produced MyD88- and TRIF-dependent inflammatory cytokines sufficient to initiate shock without any contribution from caspase-8 or caspase-11. Both proteases had to be present to support tumor necrosis factor- and interferon-β-dependent tissue injury first observed in the small intestine and later in spleen and thymus. Caspase-11 enhanced the activation of caspase-8 and extrinsic cell death machinery within the lower small intestine. Neither caspase-8 nor caspase-11 was individually sufficient for shock. Both caspases collaborated to amplify inflammatory signals associated with tissue damage. Therefore, combined pyroptotic and apoptotic signaling mediated endotoxemia independently of RIPK1 kinase activity and RIPK3 function. These observations bring to light the relevance of tissue compartmentalization to disease processes in vivo where cytokines act in parallel to execute diverse cell death pathways., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Chronic pelvic pain causes significant patient morbidity and is a challenge to clinicians. Using a murine neurogenic cystitis model that recapitulates key aspects of interstitial cystitis/bladder pain syndrome (IC), we recently showed that pseudorabies virus (PRV) induces severe pelvic allodynia in BALB/c mice relative to C57BL/6 mice. Here, we report that a quantitative trait locus (QTL) analysis of PRV-induced allodynia in F2 CxB progeny identified a polymorphism on chromosome 13, rs6314295 , significantly associated with allodynia (logarithm of odds = 3.11). The nearby gene encoding acyloxyacyl hydrolase ( Aoah) was induced in the sacral spinal cord of PRV-infected mice. AOAH-deficient mice exhibited increased vesicomotor reflex in response to bladder distension, consistent with spontaneous bladder hypersensitivity, and increased pelvic allodynia in neurogenic cystitis and postbacterial chronic pain models. AOAH deficiency resulted in greater bladder pathology and tumor necrosis factor production in PRV neurogenic cystitis, markers of increased bladder mast cell activation. AOAH immunoreactivity was detectable along the bladder-brain axis, including in brain sites previously correlated with human chronic pelvic pain. Finally, AOAH-deficient mice had significantly higher levels of bladder vascular endothelial growth factor, an emerging marker of chronic pelvic pain in humans. These findings indicate that AOAH modulates pelvic pain severity, suggesting that allelic variation in Aoah influences pelvic pain in IC.

G protein-coupled receptor kinase-6 (GRK6) is a serine/threonine kinase that is important in inflammatory processes. In this study, we examined the role of GRK6 in Escherichia coli -induced lung infection and inflammation using GRK6 knockout (KO) and wild-type (WT) mice. Intratracheal instillation of E. coli significantly enhanced bacterial load in the bronchoalveolar lavage (BAL) of KO compared with WT mice. Reduced bacterial clearance in the KO mice was not due to an intrinsic defect in neutrophil phagocytosis or killing but as a result of reduced neutrophil numbers in the KO BAL. Interestingly, neutrophil numbers in the lung were increased in the KO compared with WT mice, suggesting a potential dysfunction in transepithelial migration of neutrophils from the lungs to the bronchoalveolar space. This effect was selective for lung tissue because peritoneal neutrophil numbers were similar between the two genotypes following peritoneal infection. Although neutrophil expression of CXCR2/CXCR3 was similar between WT and KO, IL-17A expression was higher in the KO compared with WT mice. These results suggest that enhanced neutrophil count in the KO lungs but reduced numbers in BAL are likely due to transepithelial migration defect and/or altered chemokines/cytokines. Together, our studies suggest a previously unrecognized and novel role for GRK6 in neutrophil migration specific to pulmonary tissue during bacterial infection., (Copyright © 2017 the American Physiological Society.)

Rab small GTPases control membrane trafficking through effectors that recruit downstream mediators such as motor proteins. Subcellular trafficking typically involves multiple Rabs, with each specific step mediated by a distinct Rab protein. We describe a collaboration between two distinct Rab-protein-orchestrated trafficking circuits in bladder epithelial cells (BECs) that expels intracellular uropathogenic Escherichia coli (UPEC) from their intracellular niche. RAB11a and RAB27b and their trafficking circuitry are simultaneously involved in UPEC expulsion. While RAB11a recruits its effector RAB11FIP3 and cytoskeletal motor Dynein, RAB27b mobilizes the effector MyRIP and motor Myosin VIIa to mediate bacterial expulsion. This collaboration is coordinated by deposition of the exocyst complex on bacteria-containing vesicles, an event triggered by the innate receptor Toll-like receptor 4. Both RAB11a and RAB27b are recruited and activated by the exocyst complex components SEC6/SEC15. Thus, the cell autonomous defense system can mobilize and coalesce multiple subcellular trafficking circuitries to combat infections., (Published by Elsevier Inc.)

As a major diarrheagenic human pathogen, enterohemorrhagic Escherichia coli (EHEC) produce attaching and effacing (A/E) lesions, characterized by the formation of actin pedestals, on mammalian cells. A bacterial T3SS effector NleL from EHEC O157:H7 was recently shown to be a HECT-like E3 ligase in vitro, but its biological functions and host targets remain elusive. Here, we report that NleL is required to effectively promote EHEC-induced A/E lesions and bacterial infection. Furthermore, human c-Jun NH2-terminal kinases (JNKs) were identified as primary substrates of NleL. NleL-induced JNK ubiquitylation, particularly mono-ubiquitylation at the Lys 68 residue of JNK, impairs JNK's interaction with an upstream kinase MKK7, thus disrupting JNK phosphorylation and activation. This subsequently suppresses the transcriptional activity of activator protein-1 (AP-1), which modulates the formation of the EHEC-induced actin pedestals. Moreover, JNK knockdown or inhibition in host cells complements NleL deficiency in EHEC infection. Thus, we demonstrate that the effector protein NleL enhances the ability of EHEC to infect host cells by targeting host JNK, and elucidate an inhibitory role of ubiquitylation in regulating JNK phosphorylation.

Data on community-associated extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli (CA-ESBLEC) infections in Japan are scarce. We compared the clinical and microbiological epidemiology of CA-ESBLEC infections with that of healthcare-associated-ESBLEC infections among 76 patients with ESBLEC infections. We identified a high prevalence (26%) of CA-ESBLEC infections in Japan; only a small proportion (15%) of patients with CA-ESBLEC infections had recent exposure to antibiotics., (Copyright © 2016 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Four trials were conducted to evaluate the impact of Escherichia coli probiotic strain ED1a administration to pigs on the gut carriage or survival in manure of extended-spectrum-β-lactamase-producing E. coli Groups of pigs were orally inoculated with strain E. coli M63 carrying the bla CTX-M-1 gene (n = 84) or used as a control (n = 26). In the first two trials, 24 of 40 E. coli M63-inoculated pigs were given E. coli ED1a orally for 6 days starting 8 days after oral inoculation. In the third trial, 10 E. coli M63-inoculated pigs were given either E. coli ED1a or probiotic E. coli Nissle 1917 for 5 days. In the fourth trial, E. coli ED1a was given to a sow and its 12 piglets, and these 12 piglets plus 12 piglets that had not received E. coli ED1a were then inoculated with E. coli M63. Fecal shedding of cefotaxime-resistant Enterobacteriaceae (CTX-RE) was studied by culture, and bla CTX-M-1 genes were quantified by PCR. The persistence of CTX-RE in manure samples from inoculated pigs or manure samples inoculated in vitro with E. coli M63 with or without probiotics was studied. The results showed that E. coli M63 and ED1a were good gut colonizers. The reduction in the level of fecal excretion of CTX-RE in E. coli ED1a-treated pigs compared to that in nontreated pigs was usually less than 1 log 10 CFU and was mainly observed during the probiotic administration period. The results obtained with E. coli Nissle 1917 did not differ significantly from those obtained with E. coli ED1a. CTX-RE survival did not differ significantly in manure samples with or without probiotic treatment. In conclusion, under our experimental conditions, E. coli ED1a and E. coli Nissle 1917 could not durably prevent CTX-RE colonization of the pig gut., (Copyright © 2016 American Society for Microbiology.)

The aim of this study was to identify the presence of group CTX-M-9 extended spectrum beta-lactamases (ESBL) in clinical Escherichia coli isolates from pediatric patients. A total of 404 non-repeated positive ESBL E. coli isolates were collected from documented clinical infections in pediatric patients over a 2-year period. The identification and susceptibility profiles were determined using an automated system. Isolates that suggested ESBL production based on their resistance profiles to third and fourth generation cephalosporin and monobactam were selected. ESBL production was phenotypically confirmed using a diffusion method with cefotaxime and ceftazidime discs alone and in combination with clavulanic acid. blaESBL gene identification was performed through PCR amplification and sequencing. Pulsed Field Gel Electrophoresis (PFGE) and Multilocus Sequence Typing (MLST) were performed to establish the clonal relationships of the E. coli isolates. CTX-M-9-type ESBLs were detected in 2.5% of the isolates. The subtypes corresponded to blaCTX-M-14 (n = 4) and blaCTX-M-27 (n = 6). Additionally, coexistence with other beta-lactamases was observed. A clonal relationship was established in three isolates; the rest were classified as non-related. We found seven different sequence type (ST) in CTX-M-9- producing E. coli isolates. ST38 was the most frequent. This study is the first report in Mexico to document the presence of group CTX-M-9 ESBLs in E. coli isolates from pediatric patients., Competing Interests: The authors have declared that no competing interests exist.

The objective of this study was to determine whether antibiotic exposure is associated with extended-spectrum-beta-lactamase- or AmpC-producing Escherichia coli or Klebsiella pneumoniae infections in children. We collected extended-spectrum-beta-lactamase- or AmpC-producing E. coli or K. pneumoniae isolates and same-species susceptible controls from normally sterile sites of patients aged ≤21 years, along with associated clinical data, at four free-standing pediatric centers. After controlling for potential confounders, the relative risk of having an extended-spectrum-beta-lactamase-producing isolate rather than a susceptible isolate was 2.2 times higher (95% confidence interval [CI], 1.49 to 3.35) among those with antibiotic exposure in the 30 days prior to infection than in those with no antibiotic exposure. The results were similar when analyses were limited to exposure to third-generation cephalosporins, other broad-spectrum beta-lactams, or trimethoprim-sulfamethoxazole. Conversely, the relative risk of having an AmpC-producing versus a susceptible isolate was not significantly elevated with any antibiotic exposure in the 30 days prior to infection (adjusted relative risk ratio, 1.12; 95% CI, 0.65 to 1.91). However, when examining subgroups of antibiotics, the relative risk of having an AmpC-producing isolate was higher for patients with exposure to third-generation cephalosporins (adjusted relative risk ratio, 4.48; 95% CI, 1.75 to 11.43). Dose-response relationships between antibiotic exposure and extended-spectrum-beta-lactamase-producing or AmpC-producing isolates were not demonstrated. These results reinforce the need to study and implement pediatric antimicrobial stewardship strategies, and they indicate that epidemiological studies of third-generation cephalosporin-resistant E. coli and K. pneumoniae isolates should include resistance mechanisms when possible., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Cathepsin S (CTSS) is a eukaryotic protease mostly expressed in professional antigen presenting cells (APCs). Since CTSS activity regulation plays a role in the pathogenesis of various autoimmune diseases like multiple sclerosis, atherosclerosis, Sjögren's syndrome and psoriasis as well as in cancer progression, there is an ongoing interest in the reliable detection of cathepsin S activity. Various applications have been invented for specific detection of this enzyme. However, most of them have only been shown to be suitable for human samples, do not deliver quantitative results or the experimental procedure requires technical equipment that is not commonly available in a standard laboratory. We have tested a fluorogen substrate, Mca-GRWPPMGLPWE-Lys(Dnp)-DArg-NH2, that has been described to specifically detect CTSS activities in human APCs for its potential use for mouse samples. We have modified the protocol and thereby offer a cheap, easy, reproducible and quick activity assay to detect CTSS activities in mouse APCs. Since most of basic research on CTSS is performed in mice, this method closes a gap and offers a possibility for reliable and quantitative CTSS activity detection that can be performed in almost every laboratory., (Copyright © 2016. Published by Elsevier B.V.)

To investigate the dissemination of ESBL/pAmpC-producing E. coli within the food distribution system of Ho Chi Minh City (HCMC), Vietnam, the prevalence of ESBL/pAmpC-producing E. coli strains in chicken meat, pork, beef, and fish/shrimp samples obtained from slaughterhouses, a wholesale market, and supermarkets was examined. Among the total of 330 collected food samples, ESBL/pAmpC-producing E. coli was detected in 150 samples (45.5%). The highest prevalence of these isolates was in chicken meat (76/82, 92.7%), followed by pork (32/92, 34.8%), beef (18/74, 34.3%), and fish/shrimp (24/82, 29.3%). A total of 342 strains of ESBL/pAmpC-producing E. coli were isolated from 150 positive food samples. The most prevalent genes responsible for ESBL or pAmpC activity belonged to the CTX-M-9 (110/342, 31.2%), CTX-M-1 (102/342, 29.8%), and CIT (118/342, 34.5%) groups. To our knowledge, this is the first report of the high occurrence of pAmpC (37.1%) in animal-based food in Vietnam. Among the 342 total ESBL/pAmpC-producing E. coli isolates, 276 (80.7%) were resistant to at least 6 antibiotic agents. Notably, high percentages of resistance to ciprofloxacin and fosfomycin were found in isolates from chicken (80.5% and 50.8%, resp.). These findings demonstrate that animal-based food products in HCMC represent a major reservoir of ESBL/pAmpC-producing E. coli.

Extended-spectrum β-lactamases (ESBLs) are rapidly evolving plasmid transferrable enzymes that confer unique patterns of antibiotic resistance on various bacterial species. Such organisms pose special challenges to laboratory identification, as well as antibiotic selection, administration, and follow-up. Although such infections are increasingly common in the obstetric population, issues surrounding ESBLs are not widely recognized by practicing obstetricians, and controversies exist regarding diagnosis and management. This article provides the practitioner with a summary of clinically pertinent information that will assist in the proper care of pregnant patients with ESBL infection., (Copyright © 2015 Elsevier Inc. All rights reserved.)

A CMY-2-producing capsular type K2 Klebsiella pneumoniae strain (TVGHKP93) with multidrug resistance was isolated from a recurrent liver abscess in a patient who also carried a CMY-2-producing Escherichia coli strain (TVGHEC01) in the stool. TVGHKP93 retained its high virulence compared with that of the isogenic strain (TVGHKP60) with wild-type resistance from the first liver abscess. Our conjugation experiment showed the successful transfer of the blaCMY-2-carrying plasmid from TVGHEC01 into TVGHKP60. The transconjugant showed both high virulence and the multidrug-resistant phenotype, as did TVGHKP93., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

The presence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli was investigated in input (manure from livestock husbandry) and output samples of six German biogas plants in 2012 (one sampling per biogas plant) and two German biogas plants investigated in an annual cycle four times in 2013/2014. ESBL-producing Escherichia coli were cultured by direct plating on CHROMagar ESBL from input samples in the range of 100 to 104 colony forming units (CFU) per g dry weight but not from output sample. This initially indicated a complete elimination of ESBL-producing E. coli by the biogas plant process. Detected non target bacteria were assigned to the genera Acinetobacter, Pseudomonas, Bordetella, Achromobacter, Castellaniella, and Ochrobactrum. A selective pre-enrichment procedure increased the detection efficiency of ESBL-producing E. coli in input samples and enabled the detection in five of eight analyzed output samples. In total 119 ESBL-producing E. coli were isolated from input and 46 from output samples. Most of the E. coli isolates carried CTX-M-type and/or TEM-type beta lactamases (94%), few SHV-type beta lactamase (6%). Sixty-four blaCTX-M genes were characterized more detailed and assigned mainly to CTX-M-groups 1 (85%) and 9 (13%), and one to group 2. Phylogenetic grouping of 80 E. coli isolates showed that most were assigned to group A (71%) and B1 (27%), only one to group D (2%). Genomic fingerprinting and multilocus sequence typing (MLST) showed a high clonal diversity with 41 BOX-types and 19 ST-types. The two most common ST-types were ST410 and ST1210. Antimicrobial susceptibility testing of 46 selected ESBL-producing E. coli revealed that several isolates were additionally resistant to other veterinary relevant antibiotics and some grew on CHROMagar STEC but shiga-like toxine (SLT) genes were not detected. Resistance to carbapenems was not detected. In summary the study showed for the first time the presence of ESBL-producing E. coli in output samples of German biogas plants.

Introduction: Multidrug resistance (MDR) and emergence of extended-spectrum β-lactamases (ESBLs) that mediate resistance to β-lactam drugs among Escherichia coli and other uropathogens have been reported worldwide. However, there is little information on the detection of ESBLs genes in E. coli from patients with urinary tract infections (UTIs) in the Arab countries using polymerase chain reaction (PCR), and in Libya such information is lacking., Methods: All patients attending Zawiya Teaching Hospital in Zawiya city between November 2012 and June 2013 suspected of having UTIs and from whom midstream urine samples were taken as part of the clinical workup were included in this prospective study. Samples were examined for uropathogens by standard bacteriological procedures. VITEK-2 automated microbiology system was used to identify the isolated uropathogens and determine the susceptibility of E. coli and Klebsiella spp. isolates to antimicrobials. In addition, phenotypically ESBLs-positive E. coli isolates were tested for ESBLs genes by PCR., Results: The present study enrolled 1,790 patients with UTIs. Uropathogens were found in 371 (20.7%) urine specimens examined. Mixed pathogens were detected in two specimens with 373 total pathogens isolated. E. coli and Klebsiella spp. were the predominant uropathogens at 55.8% (208/373) and 18.5% (69/373), respectively. Other pathogens were detected in 25.7% (96/373) of urine samples. Of the E. coli and Klebsiella spp. tested, 69.2 and 100% were resistant to ampicillin, 6.7 and 33.3% to ceftriaxone, and 23.1 and 17.4% to ciprofloxacin, respectively. MDR (resistance to ≥3 antimicrobial groups) was found in 69 (33.2%) of E. coli and in 29 (42%) of Klebsiella spp. isolates. ESBLs were detected phenotypically in 14 (6.7%) of E. coli and in 15 (21.7%) of Klebsiella spp. isolates. Thirteen out of the 14 phenotypically ESBL-positive E. coli were positive for ESBL genes by PCR. bla TEM gene was detected in seven isolates, bla OXA gene in 10 isolates and bla CTX-M gene in six isolates. bla SHV gene was not detected in the present study., Conclusion: The isolation of MDR ESBL-producing uropathogens undoubtedly will limit the choices clinicians have to treat their patients with UTIs. Therefore, there is an urgent need for surveillance studies on antimicrobial resistance and prevalence of ESBLs among uropathogens to guide the clinical treatment of UTIs in Libya in the future.

Extended-spectrum β-lactamases (ESBLs) pose a threat to public health because of their ability to confer resistance to extended-spectrum cephalosporins such as cefotaxime. The CTX-M β-lactamases are the most widespread ESBL enzymes among antibiotic resistant bacteria. Many of the active site residues are conserved between the CTX-M family and non-ESBL β-lactamases such as TEM-1, but the residues Ser237 and Arg276 are specific to the CTX-M family, suggesting that they may help to define the increased specificity for cefotaxime hydrolysis. To test this hypothesis, site-directed mutagenesis of these positions was performed in the CTX-M-14 β-lactamase. Substitutions of Ser237 and Arg276 with their TEM-1 counterparts, Ala237 and Asn276, had a modest effect on cefotaxime hydrolysis, as did removal of the Arg276 side chain in an R276A mutant. The S237A:R276N and S237A:R276A double mutants, however, exhibited 29- and 14-fold losses in catalytic efficiency for cefotaxime hydrolysis, respectively, while the catalytic efficiency for benzylpenicillin hydrolysis was unchanged. Therefore, together, the Ser237 and Arg276 residues are important contributors to the cefotaximase substrate profile of the enzyme. High-resolution crystal structures of the CTX-M-14 S70G, S70G:S237A, and S70G:S237A:R276A variants alone and in complex with cefotaxime show that residues Ser237 and Arg276 in the wild-type enzyme promote the expansion of the active site to accommodate cefotaxime and favor a conformation of cefotaxime that allows optimal contacts between the enzyme and substrate. The conservation of these residues, linked to their effects on structure and catalysis, imply that their coevolution is an important specificity determinant in the CTX-M family.

The hallmark of enteropathogenic Escherichia coli (EPEC) infection is the formation of actin-rich pedestal-like structures, which are generated following phosphorylation of the bacterial effector Tir by cellular Src and Abl family tyrosine kinases. This leads to recruitment of the Nck-WIP-N-WASP complex that triggers Arp2/3-dependent actin polymerization in the host cell. The same phosphorylation-mediated signalling network is also assembled downstream of the Vaccinia virus protein A36 and the phagocytic Fc-gamma receptor FcγRIIa. Here we report that the EPEC type-III secretion system effector EspJ inhibits autophosphorylation of Src and phosphorylation of the Src substrates Tir and FcγRIIa. Consistent with this, EspJ inhibits actin polymerization downstream of EPEC, Vaccinia virus and opsonized red blood cells. We identify EspJ as a unique adenosine diphosphate (ADP) ribosyltransferase that directly inhibits Src kinase by simultaneous amidation and ADP ribosylation of the conserved kinase-domain residue, Src E310, resulting in glutamine-ADP ribose.

Background: Resistance to cephalosporins in Enterobacteriaceae is mainly due to the production of extended-spectrum beta-lactamase (ESBL). Little is known about ESBL-producing bacteria in Bangladesh. Therefore, the study presents results of phenotypic and molecular characterization of ESBL-producing Escherichia coli from hospitals in Bangladesh., Methods: A total of 339 E. coli isolated from patients with urinary tract and wound infections attending three different medical hospitals in urban and rural areas of Bangladesh between 2003-2007 were screened for ESBL-production by the double disk diffusion test. Isolates with ESBL-phenotype were further characterized by antibiotic susceptibility testing, PCR and sequencing of different β-lactamase and virulence genes, serotyping, and XbaI-macrorestriction followed by pulsed-field gel electrophoresis (PFGE)., Results: We identified 40 E. coli with ESBL phenotype. These isolates were resistant to ceftriaxone, ceftazidime, cefotaxime, aztreonam, cefepime, and nalidixic acid but remained susceptible to imipenem. All but one isolate were additionally resistant to ciprofloxacin, and 3 isolates were resistant to cefoxitin. ESBL genes of blaCTX-M-1-group were detected in all isolates; blaTEM-type and blaOXA-1-type genes were detected in 33 (82.5%) and 19 (47.5%) isolates, respectively. Virulence genes that are present in diarrhoeagenic E. coli were not found. Class-1 integron was present in 20 (50%) isolates. All the ESBL-producing E. coli isolates harbored plasmids ranging between 1.1 and 120 MDa. PFGE-typing revealed 26 different pulsotypes, but identical pulsotype showed 6 isolates of serotype O25:H4., Conclusion: The prevalence of multidrug-resistant ESBL-producing E. coli isolates appears to be high and the majority of the isolates were positive for blaCTX-M. Although there was genetic heterogeneity among isolates, presence of a cluster of isolates belonging to serotype O25:H4 indicates dissemination of the pandemic uropathogenic E. coli clone in Bangladesh.

Inflammasome-mediated host defenses have been extensively studied in innate immune cells. Whether inflammasomes function for innate defense in intestinal epithelial cells, which represent the first line of defense against enteric pathogens, remains unknown. We observed enhanced Salmonella enterica serovar Typhimurium colonization in the intestinal epithelium of caspase-11-deficient mice, but not at systemic sites. In polarized epithelial monolayers, siRNA-mediated depletion of caspase-4, a human ortholog of caspase-11, also led to increased bacterial colonization. Decreased rates of pyroptotic cell death, a host defense mechanism that extrudes S. Typhimurium-infected cells from the polarized epithelium, accounted for increased pathogen burdens. The caspase-4 inflammasome also governs activation of the proinflammatory cytokine, interleukin (IL)-18, in response to intracellular (S. Typhimurium) and extracellular (enteropathogenic Escherichia coli) enteric pathogens, via intracellular LPS sensing. Therefore, an epithelial cell-intrinsic noncanonical inflammasome plays a critical role in antimicrobial defense at the intestinal mucosal surface., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Two reports in this issue of Cell Host & Microbe (Sellin et al., 2014; Knodler et al., 2014) establish the cell-intrinsic inflammasome-induced extrusion of infected enterocytes as a general defense mechanism against acute bacterial infections., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objective: To investigate CTX-M genotypes among extended-spectrum β-lactamase-producing Escherichia coli (ESBL-EC) isolated from patients with community-onset and hospital-onset infections in China, their clonality and the distribution of CTX-M variants in different specimens of community-onset and hospital-onset infections., Methods: ESBL-EC isolates were collected from general hospitals from 2011 to 2012 in China. Broth microdilution method antimicrobial susceptibility testing of 16 antibiotics was performed. Clinical data from community-onset and hospital-onset infections due to ESBL-EC were analyzed. ESBL-encoding genes were amplified by PCR and sequenced, and multilocus sequence typing (MLST) was performed for a random selection of predominant CTX-M type strains identified., Results: A total of 1,168 ESBL-EC isolates were obtained from various clinical specimens, 41.7% of which were responsible for causing community-onset infections. The presence of urinary calculi was higher in community-onset infections, whereas malignancy, cardiovascular and cerebrovascular diseases, dementia, chronic renal disease, diabetes mellitus and surgical treatment were found to have higher proportions in hospital-onset infections. There was no significant difference in trauma between community-onset and hospital-onset infections. 96.2% of the isolates were detected to harbor blaCTX-M genes. blaCTX-M-1 group and blaCTX-M-9 group were detected at 40.7% and 48.7% respectively, and both positive group accounted for 10.6%. blaCTX-M-55 (24.8%) and blaCTX-M-15 (18.2%) were the major genotypes in blaCTX-M-1 group while blaCTX-M-14 (46.8%) was predominant in blaCTX-M-9 group. A comparison of blaCTX-M distribution in different specimens between ESBL-EC causing community-onset and hospital-onset infection showed no significant difference. A total of 229 isolates were tested for MLST. ST131 (14%) was the predominant type. ST648, ST405 and ST1193 were also detected., Conclusions: Community-onset ESBL-EC has emerged as a common pathogen in China. CTX-M-14 is the most commonly encountered, CTX-M-55 and CTX-M-15 have spread rapidly. ST131 is the predominant clonal group, and the great diversity of CTX-M-producing isolates of E. coli has emerged in China.

Components of fatty acid biosynthetic pathway have been identified as attractive targets for the development of new antibacterial agents. Compounds of series A (4a-4 g) and series B (5a-5 g) were synthesized by the formation of an amine bond between aromatic acid and 4-phenylthiazol-2-amine or 4-(4-bromophenyl)thiazol-2-amine. These thiazole derivatives have evaluated as potent FabH inhibitors. Nineteen compounds (4b-4h, 4 k, 4 l, 5a-5h, 5k, 5l) are reported for the first time. Most of the synthesized compounds exhibited antibacterial activity in the MTT assay. The MIC value of these compounds ranged from 1.56 μg/mL to 100 μg/mL. Moreover, the tested compounds also showed FabH inhibition ability with IC50 value ranging from 5.8 μM to 48.1 μM. The IC50 values are near the MIC values. Compound 5f has exhibited the best antibacterial and Escherichia coli FabH inhibitory activity. Docking simulation and the QSAR study was conducted for learning about binding mode and the relationship between structure and activity., (Copyright © 2013 Elsevier Masson SAS. All rights reserved.)

The spread of carbapenemase-producing Enterobacteriaceae is a global problem; however, no exact data on the epidemiology of carbapenemase in the Baltic countries and St. Petersburg area is available. We aimed to evaluate the epidemiology of carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in the Baltic States and St. Petersburg, Russia, and to compare the different methods for carbapenemase detection. From January to May 2012, all K. pneumoniae (n = 1983) and E. coli (n = 7774) clinical isolates from 20 institutions in Estonia, Latvia, Lithuania, and St. Petersburg, Russia were screened for carbapenem susceptibility. The IMP, VIM, GIM, NDM, KPC, and OXA-48 genes were detected using real-time PCR and the ability to hydrolyze ertapenem was determined using MALDI-TOF MS. Seventy-seven strains were found to be carbapenem nonsusceptible. From these, 15 K. pneumoniae strains hydrolyzed ertapenem and carried the bla NDM gene. All of these strains carried integron 1 and most carried integron 3 as well as genes of the CTX-M-1 group. No carbapenemase-producing E. coli or K. pneumoniae strains were found in Estonia, Latvia, or Lithuania; however, NDM-positive K. pneumoniae was present in the hospital in St. Petersburg, Russia. A MALDI-TOF MS-based assay is a suitable and cost-effective method for the initial confirmation of carbapenemase production.

Objectives: The aim of this study was to evaluate if xanthine oxidase and myeloperoxidase levels quantitation method may alternate routine culture method, which takes more time in the diagnosis of urinary tract infections., Material and Methods: Five hundred and forty-nine outpatients who had admitted to Clinic Microbiology Laboratory were included in the study. The microorganisms were identified by using VITEK System. The urine specimens that were negative from the quantitative urine culture were used as controls. The activities of MPO and XO in spot urine were measured by spectrophotometric method., Results: Through the urine cultures, 167 bacteria were isolated from 163 urine specimens; 386 cultures yielded no bacterial growth. E. coli was the most frequent pathogen. In infection with E. coli both XO and MPO levels were increased the most. The sensitivity, specificity, positive predictive value, and negative predictive value for XO were 100%, 100%, 100%, and 100%, respectively. These values for MPO were 87%, 100%, 100%, and 94%, respectively., Conclusion: These data obtained suggest that urine XO and MPO levels may be new markers in the early detection of UTI.

The prevalence of extended-spectrum-β-lactamase-producing Enterobacteriaceae (ESBLPE) was studied in stool samples from 125 8- to 16-month-old healthy children. Twenty-four percent of them and 10.7% of the 318 fecal samples studied yielded extended-spectrum-β-lactamase-producing Escherichia coli, with the types being SHV-12, CTX-M-1, CTX-M-14, and TEM-52, the most common types of β-lactamases. This high prevalence of ESBLPE in healthy people, which is to our knowledge the highest currently reported in Europe, may represent a risk for increased infections by these organisms in the future.

Background & Aims: Inducible chitinase 3-like-1 is expressed by intestinal epithelial cells (IECs) and adheres to bacteria under conditions of inflammation. We performed a structure-function analysis of the chitin-binding domains encoded by the chiA gene, which mediates the pathogenic effects of adherent invasive Escherichia coli (AIEC)., Methods: We created AIEC (strain LF82) with deletion of chiA (LF82-ΔchiA) or that expressed chiA with specific mutations. We investigated the effects of infecting different IEC lines with these bacteria compared with nonpathogenic E coli; chitinase activities were measured using the colloidal chitin-azure method. Colitis was induced in C57/Bl6 mice by administration of dextran sodium sulfate, and mice were given 10(8) bacteria for 15 consecutive days by gavage. Stool/tissue samples were collected and analyzed., Results: LF82-ΔchiA had significantly less adhesion to IEC lines than LF82. Complementation of LF82-ΔchiA with the LF82 chiA gene, but not chiA from nonpathogenic (K12) E coli, increased adhesion. We identified 5 specific polymorphisms in the chitin-binding domain of LF82 chiA (at amino acids 362, 370, 378, 388, and 548) that differ from chiA of K12 and were required for LF82 to interact directly with IECs. This interaction was mediated by an N-glycosylated asparagine in chitinase 3-like-1 (amino acid 68) on IECs. Mice infected with LF82, or LF82-ΔchiA complemented with LF82 chiA, developed more severe colitis after administration of dextran sodium sulfate than mice infected with LF82-ΔchiA or LF82 that expressed mutant forms of chiA., Conclusions: AIEC adheres to an N-glycosylated chitinase 3-like-1 on IECs via the chitin-binding domain of chiA. This mechanism promotes the pathogenic effects of AIEC in mice with colitis., (Copyright © 2013 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Reactive oxygen species (ROS) from nicotinamide adenine dinucleotide phosphate (NADPH) oxidases and their related dual oxidases are known to have significant roles in innate immunity and cell proliferation. In this study, the 5,545 bp cDNA of the silkworm Bombyx mori dual oxidase (BmDuox) gene containing a full-length open reading frame was cloned. It was shown to include an N-terminal signal peptide consisting of 28 amino acid residues, a 240 bp 5'-terminal untranslated region (5'-UTR), an 802 bp 3'-terminal region (3'-UTR), which contains nine ATTTA motifs, and a 4,503 bp open reading frame encoding a polypeptide of 1,500 amino acid residues. Structural analysis indicated that BmDuox contains a typical peroxidase domain at the N-terminus followed by a calcium-binding domain, a ferric-reducing domain, six transmembrane regions and binding domains for flavin adenine dinucleotide (FAD) and nicotinamide adenine dinucleotide (NAD). Transcriptional analysis revealed that BmDuox mRNA was expressed more highly in the head, testis and trachea compared to the midgut, hemocyte, Malpighian tube, ovary, fat bodies and silk glands. BmDuox mRNA was expressed during all the developmental stages of the silkworm. Subcellular localization revealed that BmDoux was present mainly in the periphery of the cells. Some cytoplasmic staining was detected, with rare signals in the nucleus. Expression of BmDuox was induced significantly in the larval midgut upon challenge by Escherichia coli and Bombyx mori nucleopolyhedrovirus (BmNPV). BmDuox-deleted larvae showed a marked increase in microbial proliferation in the midgut after ingestion of fluorescence-labeled bacteria compared to the control. We conclude that reducing BmDuox expression greatly increased the bacterial load, suggesting BmDuox has an important role in inhibiting microbial proliferation and the maintenance of homeostasis in the silkworm midgut.