Showing papers on "Escherichia coli published in 2018"

Optically-controlled bacterial metabolite for cancer therapy.

Abstract: Bacteria preferentially accumulating in tumor microenvironments can be utilized as natural vehicles for tumor targeting. However, neither current chemical nor genetic approaches alone can fully satisfy the requirements on both stability and high efficiency. Here, we propose a strategy of "charging" bacteria with a nano-photocatalyst to strengthen their metabolic activities. Carbon nitride (C3N4) is combined with Escherichia coli (E. coli) carrying nitric oxide (NO) generation enzymes for photo-controlled bacterial metabolite therapy (PMT). Under light irradiation, photoelectrons produced by C3N4 can be transferred to E. coli to promote the enzymatic reduction of endogenous NO3- to cytotoxic NO with a 37-fold increase. In a mouse model, C3N4 loaded bacteria are perfectly accumulated throughout the tumor and the PMT treatment results in around 80% inhibition of tumor growth. Thus, synthetic materials-remodeled microorganism may be used to regulate focal microenvironments and increase therapeutic efficiency.

Characterization of poxtA, a novel phenicol-oxazolidinone-tetracycline resistance gene from an MRSA of clinical origin.

Abstract: Objectives To characterize a novel phenicol-oxazolidinone-tetracycline resistance gene, named poxtA, identified in a previously described MRSA strain that was highly resistant to linezolid and also carried the cfr gene. Methods The poxtA gene was identified by bioinformatic analysis of the whole genome sequence of Staphylococcus aureus AOUC-0915. The poxtA gene was cloned in a shuttle plasmid vector and expressed in Escherichia coli, S. aureus and Enterococcus faecalis to investigate the protein function. Comparative sequence analyses at the protein and genetic levels were carried out using standard procedures. Results The poxtA gene encodes a protein that is 32% identical to OptrA and exhibits structural features typical of the F lineage of the ATP-binding cassette (ABC) protein superfamily that cause antibiotic resistance by ribosomal protection. Expression of poxtA in E. coli, S. aureus and E. faecalis was able to decrease susceptibility to phenicols, oxazolidinones and tetracyclines. A database search identified the presence of poxtA in E. faecalis, Enterococcus faecium and Pediococcus acidilactici strains, mostly of animal origin, and revealed the presence of poxtA homologues in the genomes of some Clostridiales. Analysis of the genetic context revealed that poxtA was located in a composite transposon-like structure containing two IS1216 elements. Conclusions A novel resistance gene, named poxtA, encoding a protein of the antibiotic resistance (ARE) ABC-F lineage, was identified in the genome of an MRSA of clinical origin. PoxtA can confer decreased susceptibility to phenicols, oxazolidinones and tetracyclines and is associated with a putative mobile element that could contribute to its horizontal dissemination.

Colibactin: More Than a New Bacterial Toxin.

Abstract: Cyclomodulins are bacterial toxins that interfere with the eukaryotic cell cycle. A new cyclomodulin called colibactin, which is synthetized by the pks genomic island, was discovered in 2006. Despite many efforts, colibactin has not yet been purified, and its structure remains elusive. Interestingly, the pks island is found in members of the family Enterobacteriaceae (mainly Escherichia coli and Klebsiella pneumoniae) isolated from different origins, including from intestinal microbiota, septicaemia, newborn meningitis, and urinary tract infections. Colibactin-producing bacteria induce chromosomal instability and DNA damage in eukaryotic cells, which leads to senescence of epithelial cells and apoptosis of immune cells. The pks island is mainly observed in B2 phylogroup E. coli strains, which include extra-intestinal pathogenic E. coli strains, and pks E. coli are over-represented in biopsies isolated from colorectal cancer. In addition, pks E. coli bacteria increase the number of tumours in diverse colorectal cancer mouse models. Thus, colibactin could have a major impact on human health. In the present review, we will focus on the biological effects of colibactin, the distribution of the pks island, and summarize what is currently known about its synthesis and its structure.

The Colibactin Genotoxin Generates DNA Interstrand Cross-Links in Infected Cells

Abstract: Colibactins are hybrid polyketide-nonribosomal peptides produced by Escherichia coli, Klebsiella pneumoniae, and other Enterobacteriaceae harboring the pks genomic island. These genotoxic metabolites are produced by pks-encoded peptide-polyketide synthases as inactive prodrugs called precolibactins, which are then converted to colibactins by deacylation for DNA-damaging effects. Colibactins are bona fide virulence factors and are suspected of promoting colorectal carcinogenesis when produced by intestinal E. coli. Natural active colibactins have not been isolated, and how they induce DNA damage in the eukaryotic host cell is poorly characterized. Here, we show that DNA strands are cross-linked covalently when exposed to enterobacteria producing colibactins. DNA cross-linking is abrogated in a clbP mutant unable to deacetylate precolibactins or by adding the colibactin self-resistance protein ClbS, confirming the involvement of the mature forms of colibactins. A similar DNA-damaging mechanism is observed in cellulo, where interstrand cross-links are detected in the genomic DNA of cultured human cells exposed to colibactin-producing bacteria. The intoxicated cells exhibit replication stress, activation of ataxia-telangiectasia and Rad3-related kinase (ATR), and recruitment of the DNA cross-link repair Fanconi anemia protein D2 (FANCD2) protein. In contrast, inhibition of ATR or knockdown of FANCD2 reduces the survival of cells exposed to colibactin-producing bacteria. These findings demonstrate that DNA interstrand cross-linking is the critical mechanism of colibactin-induced DNA damage in infected cells. IMPORTANCE Colorectal cancer is the third-most-common cause of cancer death. In addition to known risk factors such as high-fat diets and alcohol consumption, genotoxic intestinal Escherichia coli bacteria producing colibactin are proposed to play a role in colon cancer development. Here, by using transient infections with genotoxic E. coli, we showed that colibactins directly generate DNA cross-links in cellulo. Such lesions are converted into double-strand breaks during the repair response. DNA cross-links, akin to those induced by metabolites of alcohol and high-fat diets and by widely used anticancer drugs, are both severely mutagenic and profoundly cytotoxic lesions. This finding of a direct induction of DNA cross-links by a bacterium should facilitate delineating the role of E. coli in colon cancer and engineering new anticancer agents.

Global landscape of cell envelope protein complexes in Escherichia coli

Abstract: Bacterial cell envelope protein (CEP) complexes mediate a range of processes, including membrane assembly, antibiotic resistance and metabolic coordination. However, only limited characterization of relevant macromolecules has been reported to date. Here we present a proteomic survey of 1,347 CEPs encompassing 90% inner- and outer-membrane and periplasmic proteins of Escherichia coli. After extraction with non-denaturing detergents, we affinity-purified 785 endogenously tagged CEPs and identified stably associated polypeptides by precision mass spectrometry. The resulting high-quality physical interaction network, comprising 77% of targeted CEPs, revealed many previously uncharacterized heteromeric complexes. We found that the secretion of autotransporters requires translocation and the assembly module TamB to nucleate proper folding from periplasm to cell surface through a cooperative mechanism involving the β-barrel assembly machinery. We also establish that an ABC transporter of unknown function, YadH, together with the Mla system preserves outer membrane lipid asymmetry. This E. coli CEP 'interactome' provides insights into the functional landscape governing CE systems essential to bacterial growth, metabolism and drug resistance.

Surface Modified with a Host Defense Peptide-Mimicking β-Peptide Polymer Kills Bacteria on Contact with High Efficacy

Abstract: Methicillin-resistant Staphylococcus aureus (MRSA) has been one of the major nosocomial pathogens to cause frequent and serious infections that are associated with various biomedical surfaces. This study demonstrated that surface modified with host defense peptide-mimicking β-peptide polymer, has surprisingly high bactericidal activities against Escherichia coli (E. coli) and MRSA. As surface-tethered β-peptide polymers cannot move freely to adopt the collaborative interactions with bacterial membrane and are too short to penetrate the cell envelop, we proposed a mode of action by diffusing away the cell membrane-stabilizing divalent ions, Ca2+ and Mg2+. This hypothesis was supported by our study that Ca2+ and Mg2+ supplementation in the assay medium causes up to 80% loss of bacterial killing efficacy and that the addition of divalent ion chelating ethylenediaminetetraacetic acid into the above assay medium leads to significant recovery of the bacterial killing efficacy. In addition to its potent bacteria...

Repurposing type III polyketide synthase as a malonyl-CoA biosensor for metabolic engineering in bacteria

Abstract: Malonyl-CoA is an important central metabolite for the production of diverse valuable chemicals including natural products, but its intracellular availability is often limited due to the competition with essential cellular metabolism Several malonyl-CoA biosensors have been developed for high-throughput screening of targets increasing the malonyl-CoA pool However, they are limited for use only in Escherichia coli and Saccharomyces cerevisiae and require multiple signal transduction steps Here we report development of a colorimetric malonyl-CoA biosensor applicable in three industrially important bacteria: E coli, Pseudomonas putida, and Corynebacterium glutamicum RppA, a type III polyketide synthase producing red-colored flaviolin, was repurposed as a malonyl-CoA biosensor in E coli Strains with enhanced malonyl-CoA accumulation were identifiable by the colorimetric screening of cells showing increased red color Other type III polyketide synthases could also be repurposed as malonyl-CoA biosensors For target screening, a 1,858 synthetic small regulatory RNA library was constructed and applied to find 14 knockdown gene targets that generally enhanced malonyl-CoA level in E coli These knockdown targets were applied to produce two polyketide (6-methylsalicylic acid and aloesone) and two phenylpropanoid (resveratrol and naringenin) compounds Knocking down these genes alone or in combination, and also in multiple different E coli strains for two polyketide cases, allowed rapid development of engineered strains capable of enhanced production of 6-methylsalicylic acid, aloesone, resveratrol, and naringenin to 4403, 309, 518, and 1038 mg/L, respectively The malonyl-CoA biosensor developed here is a simple tool generally applicable to metabolic engineering of microorganisms to achieve enhanced production of malonyl-CoA–derived chemicals

The Role of Outer Membrane Proteins and Lipopolysaccharides for the Sensitivity of Escherichia coli to Antimicrobial Peptides

Abstract: Bacterial resistance to classical antibiotics is emerging worldwide. The number of infections caused by multidrug resistant bacteria is increasing and becoming a serious threat for human health globally. In particular, Gram-negative pathogens including multidrug resistant Escherichia coli are of serious concern being resistant to the currently available antibiotics. All Gram-negative bacteria are enclosed by an outer membrane which acts as an additional protection barrier preventing the entry of toxic compounds including antibiotics and antimicrobial peptides (AMPs). In this study we report that the outer membrane component lipopolysaccharide (LPS) plays a crucial role for the antimicrobial susceptibility of E. coli BW25113 against the cationic AMPs Cap18, Cap11, Cap11-1-18m2, melittin, indolicidin, cecropin P1, cecropin B, and the polypeptide antibiotic colistin, whereas the outer membrane protease OmpT and the lipoprotein Lpp only play a minor role for the susceptibility against cationic AMPs. Increased susceptibility toward cationic AMPs was found for LPS deficient mutants of E. coli BW25113 harboring deletions in any of the genes required for the inner part of core-oligosaccharide of the LPS, waaC, waaE, waaF, waaG, and gmhA. In addition, our study demonstrates that the antimicrobial activity of Cap18, Cap11, Cap11-1-18m2, cecropin B, and cecropin P1 is not only dependent on the inner part of the core oligosaccharide, but also on the outer part and its sugar composition. Finally, we demonstrated that the antimicrobial activity of selected Cap18 derivatives harboring amino acid substitutions in the hydrophobic interface, are non-active against wild-type E. coli ATCC29522. By deleting waaC, waaE, waaF, or waaG the antimicrobial activity of the non-active derivatives can be partially or fully restored, suggesting a very close interplay between the LPS core oligosaccharide and the specific Cap18 derivative. Summarizing, this study implicates that the nature of the outer membrane component LPS has a big impact on the antimicrobial activity of cationic AMPs against E. coli. In particular, the inner as well as the outer part of the core oligosaccharide are important elements determining the antimicrobial susceptibility of E. coli against cationic AMPs.

Antibiotic resistance pattern and virulence genes content in avian pathogenic Escherichia coli (APEC) from broiler chickens in Chitwan, Nepal.

Abstract: Avian pathogenic Escherichia coli (APEC) are causative agent of extraintestinal infections, collectively known as colibacillosis, which results significant losses in poultry industries. The extraintestinal survival of E. coli is facilitated by numerous virulence factors which are coded by virulence genes. This study was conducted to find out the pattern of antibiotic resistance and virulence genes content in the APEC strains isolated from broiler chickens at National Avian Disease Investigation Laboratory and Veterinary Teaching Hospital, Rampur, Chitwan, Nepal. A total of 50 E. coli strains were isolated from 50 colibacillosis suspected broiler chickens. Out of 50 isolates of E. coli, 47 (94%) showed resistant to three or more antimicrobials. The highest levels (22%) of multidrug-resistant E. coli were observed for five different types of antimicrobials. Antibiogram profiles of 50 E. coli strains showed the maximum resistance to ampicillin (98%), followed by co-trimoxazole (90%), and doxycycline (62%). The highest intermediate resistance was shown by colistin (50%) and the highest sensitivity was against amikacin (84%), followed by nitrofurantoin (55%). Based on the genetic criteria, 45 (90%) E. coli isolates were considered as pathogenic (APEC) which contained more than five virulence genes. Out of total APEC genes detected, we found the combination of iss, iucD, hlyF, ompT, iroN, and iutA genes were mostly associated with the APEC and additionally, to some lesser extent irp2, papC, Cva/cvi, and tsh genes showed the critical role for virulent traits of APEC strains. In this study, high prevalent of antimicrobial resistant pattern was found with avian pathogenic E. coli strains isolated from broiler chickens. To our knowledge, this is the first molecular analysis which confirmed the prevalence of APEC strains in poultry sector in Nepal. These finding suggest the need of surveillance and intervention system to control misuse of antibiotics and APEC outbreak in the poultry farm.

Metabolic engineering of Escherichia coli for de novo biosynthesis of vitamin B 12

Abstract: The only known source of vitamin B12 (adenosylcobalamin) is from bacteria and archaea. Here, using genetic and metabolic engineering, we generate an Escherichia coli strain that produces vitamin B12 via an engineered de novo aerobic biosynthetic pathway. In vitro and/or in vivo analysis of genes involved in adenosylcobinamide phosphate biosynthesis from Rhodobacter capsulatus suggest that the biosynthetic steps from co(II)byrinic acid a,c-diamide to adocobalamin are the same in both the aerobic and anaerobic pathways. Finally, we increase the vitamin B12 yield of a recombinant E. coli strain by more than ∼250-fold to 307.00 µg g−1 DCW via metabolic engineering and optimization of fermentation conditions. Beyond our demonstration of E. coli as a microbial biosynthetic platform for vitamin B12 production, our study offers an encouraging example of how the several dozen proteins of a complex biosynthetic pathway can be transferred between organisms to facilitate industrial production. Vitamin B12 is an essential nutrient with limited natural sources. Here the authors transfer 28 pathway synthesis genes from several bacteria including R. capsulatus to E. coli and, using metabolic engineering and optimised fermentation conditions, achieve high yields.

Antibiotic-resistant Escherichia coli from retail poultry meat with different antibiotic use claims.

Abstract: We sought to determine if the prevalence of antibiotic-resistant Escherichia coli differed across retail poultry products and among major production categories, including organic, “raised without antibiotics”, and conventional. We collected all available brands of retail chicken and turkey—including conventional, “raised without antibiotic”, and organic products—every two weeks from January to December 2012. In total, E. coli was recovered from 91% of 546 turkey products tested and 88% of 1367 chicken products tested. The proportion of samples contaminated with E. coli was similar across all three production categories. Resistance prevalence varied by meat type and was highest among E. coli isolates from turkey for the majority of antibiotics tested. In general, production category had little effect on resistance prevalence among E. coli isolates from chicken, although resistance to gentamicin and multidrug resistance did vary. In contrast, resistance prevalence was significantly higher for 6 of the antibiotics tested—and multidrug resistance—among isolates from conventional turkey products when compared to those labelled organic or “raised without antibiotics”. E. coli isolates from chicken varied strongly in resistance prevalence among different brands within each production category. The high prevalence of resistance among E. coli isolates from conventionally-raised turkey meat suggests greater antimicrobial use in conventional turkey production as compared to “raised without antibiotics” and organic systems. However, among E. coli from chicken meat, resistance prevalence was more strongly linked to brand than to production category, which could be caused by brand-level differences during production and/or processing, including variations in antimicrobial use.

"It's a gut feeling" - Escherichia coli biofilm formation in the gastrointestinal tract environment.

Abstract: Escherichia coli can commonly be found, either as a commensal, probiotic or a pathogen, in the human gastrointestinal (GI) tract. Biofilm formation and its regulation is surprisingly variable, although distinct regulatory pattern of red, dry and rough (rdar) biofilm formation arise in certain pathovars and even clones. In the GI tract, environmental conditions, signals from the host and from commensal bacteria contribute to shape E. coli biofilm formation within the multi-faceted multicellular communities in a complex and integrated fashion. Although some major regulatory networks, adhesion factors and extracellular matrix components constituting E. coli biofilms have been recognized, these processes have mainly been characterized in vitro and in the context of interaction of E. coli strains with intestinal epithelial cells. However, direct observation of E. coli cells in situ, and the vast number of genes encoding surface appendages on the core or accessory genome of E. coli suggests the complexity of the biofilm process to be far from being fully understood. In this review, we summarize biofilm formation mechanisms of commensal, probiotic and pathogenic E. coli in the context of the gastrointestinal tract.

Esterase-Catalyzed Siderophore Hydrolysis Activates an Enterobactin-Ciprofloxacin Conjugate and Confers Targeted Antibacterial Activity.

Abstract: Enteric Gram-negative bacteria, including Escherichia coli, biosynthesize and deploy the triscatecholate siderophore enterobactin (Ent) in the vertebrate host to acquire iron, an essential nutrient. We report that Ent–Cipro, a synthetic siderophore–antibiotic conjugate based on the native Ent platform that harbors an alkyl linker at one of the catechols with a ciprofloxacin cargo attached, affords targeted antibacterial activity against E. coli strains that express the pathogen-associated iroA gene cluster. Attachment of the siderophore to ciprofloxacin, a DNA gyrase inhibitor and broad-spectrum antibiotic that is used to treat infections caused by E. coli, generates an inactive prodrug and guides the antibiotic into the cytoplasm of bacteria that express the Ent uptake machinery (FepABCDG). Intracellular hydrolysis of the siderophore restores the activity of the antibiotic. Remarkably, Fes, the cytoplasmic Ent hydrolase expressed by all E. coli, does not contribute to Ent–Cipro activation. Instead, this ...

Absence of Curli in Soil-Persistent Escherichia coli Is Mediated by a C-di-GMP Signaling Defect and Suggests Evidence of Biofilm-Independent Niche Specialization.

Abstract: Escherichia coli is commonly viewed as a gastrointestinal commensal or pathogen although an increasing body of evidence suggests that it can persist in non-host environments as well. Curli are a major component of biofilm in many enteric bacteria including E. coli and are important for adherence to different biotic and abiotic surfaces. In this study we investigated curli production in a unique collection of soil-persistent E. coli isolates and examined the role of curli formation in environmental persistence. Although most soil-persistent E. coli were curli-positive, 10% of isolates were curli-negative (17 out of 170). Curli-producing E. coli (COB583, COB585, and BW25113) displayed significantly more attachment to quartz sand than the curli-negative strains. Long-term soil survival experiments indicated that curli production was not required for long-term survival in live soil (over 110 days), as a curli-negative mutant BW25113ΔcsgB had similar survival compared to wild type BW25113. Mutations in two genes associated with c-di-GMP metabolism, dgcE and pdeR, correlated with loss of curli in eight soil-persistent strains, although this did not significantly impair their survival in soil compared to curli-positive strains. Overall, the data indicate that curli-deficient and biofilm-defective strains, that also have a defect in attachment to quartz sand, are able to reside in soil for long periods of time thus pointing to the possibility that niches may exist in the soil that can support long-term survival independently of biofilm formation.

Rapid Growth of Uropathogenic Escherichia coli during Human Urinary Tract Infection

Abstract: Uropathogenic Escherichia coli (UPEC) strains cause most uncomplicated urinary tract infections (UTIs). These strains are a subgroup of extraintestinal pathogenic E. coli (ExPEC) strains that infect extraintestinal sites, including urinary tract, meninges, bloodstream, lungs, and surgical sites. Here, we hypothesize that UPEC isolates adapt to and grow more rapidly within the urinary tract than other E. coli isolates and survive in that niche. To date, there has not been a reliable method available to measure their growth rate in vivo Here we used two methods: segregation of nonreplicating plasmid pGTR902, and peak-to-trough ratio (PTR), a sequencing-based method that enumerates bacterial chromosomal replication forks present during cell division. In the murine model of UTI, UPEC strain growth was robust in vivo, matching or exceeding in vitro growth rates and only slowing after reaching high CFU counts at 24 and 30 h postinoculation (hpi). In contrast, asymptomatic bacteriuria (ABU) strains tended to maintain high growth rates in vivo at 6, 24, and 30 hpi, and population densities did not increase, suggesting that host responses or elimination limited population growth. Fecal strains displayed moderate growth rates at 6 hpi but did not survive to later times. By PTR, E. coli in urine of human patients with UTIs displayed extraordinarily rapid growth during active infection, with a mean doubling time of 22.4 min. Thus, in addition to traditional virulence determinants, including adhesins, toxins, iron acquisition, and motility, very high growth rates in vivo and resistance to the innate immune response appear to be critical phenotypes of UPEC strains.IMPORTANCE Uropathogenic Escherichia coli (UPEC) strains cause most urinary tract infections in otherwise healthy women. While we understand numerous virulence factors are utilized by E. coli to colonize and persist within the urinary tract, these properties are inconsequential unless bacteria can divide rapidly and survive the host immune response. To determine the contribution of growth rate to successful colonization and persistence, we employed two methods: one involving the segregation of a nonreplicating plasmid in bacteria as they divide and the peak-to-trough ratio, a sequencing-based method that enumerates chromosomal replication forks present during cell division. We found that UPEC strains divide extraordinarily rapidly during human UTIs. These techniques will be broadly applicable to measure in vivo growth rates of other bacterial pathogens during host colonization.

Enhancing Production of Pinene in Escherichia coli by Using a Combination of Tolerance, Evolution, and Modular Co-culture Engineering

Abstract: alpha-Pinene is a natural and active monoterpene, which is widely used as a flavoring agent and in fragrances, pharmaceuticals and biofuels. Although it has been successfully produced by genetically engineered microorganisms, the production level of pinene is much lower than that of hemiterpene (isoprene) and sesquiterpenes (farnesene) to date. We first improved pinene tolerance to 2.0% and pinene production by adaptive laboratory evolution after atmospheric and room temperature plasma (ARTP) mutagenesis and overexpression of the efflux pump to obtain the pinene tolerant strain Escherichia coli YZFP, which is resistant to fosmidomycin. Through error-prone PCR and DNA shuffling, we isolated an Abies grandis geranyl pyrophosphate synthase variant that outperformed the wild-type enzyme. To balance the expression of multiple genes, a tunable intergenic region (TIGR) was inserted between A. grandis GPPSD90G/L175P and Pinus taeda Pt1Q457L. In an effort to improve the production, an E. coli-E. coli modular co-culture system was engineered to modularize the heterologous mevalonate (MEV) pathway and the TIGR-mediated gene cluster of A. grandis GPPSD90G/L175P and P. taeda Pt1Q457L. Specifically, the MEV pathway and the TIGR-mediated gene cluster were integrated into the chromosome of the pinene tolerance strain E. coli YZFP and then evolved to a higher gene copy number by chemically induced chromosomal evolution, respectively. The best E. coli-E. coli co-culture system of fermentation was found to improve pinene production by 1.9-fold compared to the mono-culture approach. The E. coli-E. coli modular co-culture system of whole-cell biocatalysis further improved pinene production to 166.5 mg/L.

Production of itaconic acid from acetate by engineering acid-tolerant Escherichia coli W.

Abstract: Utilization of abundant and cheap carbon sources can effectively reduce the production cost and enhance the economic feasibility. Acetate is a promising carbon source to achieve cost-effective microbial processes. In this study, we engineered an Escherichia coli strain to produce itaconic acid from acetate. As acetate is known to inhibit cell growth, we initially screened for a strain with a high tolerance to 10 g/L of acetate in the medium, and the W strain was selected as the host. Subsequently, the WC strain was obtained by overexpression of cad (encoding cis-aconitate decarboxylase) using a synthetic promoter and 5' UTR. However, the WC strain produced only 0.13 g/L itaconic acid because of low acetate uptake. To improve the production, the acetate assimilating pathway and glyoxylate shunt pathway were amplified by overexpression of pathway genes as well as its deregulation. The resulting strain, WCIAG4 produced 3.57 g/L itaconic acid (16.1% of theoretical maximum yield) after 88 hr of fermentation with rapid acetate assimilation. These efforts support that acetate can be a potential feedstock for biochemical production with engineered E. coli.

Mechanisms of ultraviolet disinfection and chlorination of Escherichia coli: Culturability, membrane permeability, metabolism, and genetic damage

Abstract: Traditional culture methods may underestimate the tolerance of microorganisms to disinfectants because of the existence of viable but nonculturable or sublethally injured cells after disinfection. The selection of a strict method is crucial for the evaluation of disinfection performance. The actions of 2 typical disinfectants - ultraviolet (UV) and chlorine - on the fecal indicator Escherichia coli were investigated by the detection of culturability, membrane permeability, metabolic activity, deoxyribonucleic acid (DNA), and messenger ribonucleic acid (mRNA). During UV disinfection, the irreversible damages in the cell membrane and cellular adenosine triphosphate (ATP) were negligible at low UV doses ( 5mg/L) treatments. The decay of mRNA was more rapid than that of DNA. The degradation level of mRNA depended on the choice of target genes. After exposure to 50mJ/cm2 UV dose or 5mg/L chlorine for 30min, the DNA damage repair function (RecA mRNA) was inhibited. The mRNA involved in the DNA damage repair function can be a potential indicator of bacterial viability.

Outer membrane vesicles from β-lactam-resistant Escherichia coli enable the survival of β-lactam-susceptible E . coli in the presence of β-lactam antibiotics

Abstract: Outer membrane vesicles (OMVs) containing various bacterial compounds are released from mainly gram-negative bacteria. Secreted OMVs play important roles in the ability of a bacterium to defend itself, and thus contribute to the survival of bacteria in a community. In this study, we collected OMVs from β-lactam antibiotic-resistant Escherichia coli established by conjugation assay and the parental β-lactam antibiotic-susceptible strain, and performed comparative proteomic analysis to examine whether these OMVs carried β-lactam-resistant compounds. We also investigated whether both types of OMVs could protect susceptible cells from β-lactam-induced death and/or directly degrade β-lactam antibiotics. Several proteins that can be involved in degrading β-lactam antibiotics were more abundant in OMVs from β-lactam-resistant E. coli, and thus OMVs from β-lactam resistant E. coli could directly and dose-dependently degrade β-lactam antibiotics and fully rescue β-lactam-susceptible E. coli and other bacterial species from β-lactam antibiotic-induced growth inhibition. Taken together, present study demonstrate that OMVs from β-lactam-resistant E. coli play important roles in survival of antibiotic susceptible bacteria against β-lactam antibiotics. This finding may pave the way for new efforts to combat the current global spread of antibiotic resistances, which is considered to be a significant public health threat.

Mechanism of Antibacterial Activity of Choline-Based Ionic Liquids (CAGE)

Abstract: The continued emergence of antibiotic-resistant organisms has severely depleted our arsenal of effective antimicrobials. Ionic liquids (ILs) show great promise as antibacterial agents but understanding the mechanism of attack on bacterial cells is key to ensuring that design of IL-based biocides impart maximum efficacy with minimal toxicity, while also avoiding the potential for the target organisms to become resistant. Here we report the antibacterial attributes of a set of choline and geranate (CAGE)-based ILs and identify the mechanism by which they interact with the Gram-negative cell wall of Escherichia coli. CAGE is envisaged as an antimicrobial agent to treat topical infections in skin. Our earlier work has shown that CAGE is highly effective across a breadth of bacterial, fungal, and viral species and is benign to human cells. This combination makes CAGE an ideal antimicrobial for human use. Four CAGE variants with varying ratios of choline and geranic acid were synthesized and tested for their an...

Identification and pathogenomic analysis of an Escherichia coli strain producing a novel Shiga toxin 2 subtype.

Abstract: Shiga toxin (Stx) is the key virulent factor in Shiga toxin-producing Escherichia coli (STEC). To date, three Stx1 subtypes and seven Stx2 subtypes have been described in E. coli, which differed in receptor preference and toxin potency. Here, we identified a novel Stx2 subtype designated Stx2h in E. coli strains isolated from wild marmots in the Qinghai-Tibetan plateau, China. Stx2h shares 91.9% nucleic acid sequence identity and 92.9% amino acid identity to the nearest Stx2 subtype. The expression of Stx2h in type strain STEC299 was inducible by mitomycin C, and culture supernatant from STEC299 was cytotoxic to Vero cells. The Stx2h converting prophage was unique in terms of insertion site and genetic composition. Whole genome-based phylo- and patho-genomic analysis revealed STEC299 was closer to other pathotypes of E. coli than STEC, and possesses virulence factors from other pathotypes. Our finding enlarges the pool of Stx2 subtypes and highlights the extraordinary genomic plasticity of E. coli strains. As the emergence of new Shiga toxin genotypes and new Stx-producing pathotypes pose a great threat to the public health, Stx2h should be further included in E. coli molecular typing, and in epidemiological surveillance of E. coli infections.

Current Trends in Antimicrobial Resistance of Escherichia coli.

Abstract: Escherichia coli is the most common Gram-negative bacterial pathogen, presenting both a clinical and an epidemiological challenge. In the last decade, several successful multidrug-resistant high-risk strains, such as strain E. coli ST131 have evolved, mainly due to the growing selective pressure of antimicrobial use. These strains present enhanced fitness and pathogenicity, effective transmission and colonization abilities, global distribution due to efficient dissemination, and resistance to various antimicrobial resistances. Here, we describe the emerging trends and epidemiology of resistant E. coli, including carbapenemase-producing E. coli, E. coli ST131 and colistin resistant E. coli.

Prevalence, identification of virulence factors, O-serogroups and antibiotic resistance properties of Shiga-toxin producing Escherichia coli strains isolated from raw milk and traditional dairy products

Abstract: Shiga-toxigenic Escherichia coli strains are one of the most important foodborne bacteria with an emergence of antibiotic resistance. Foodborne STEC strains are mainly associated with presence of certain virulence factors and O-seogroups. The present investigation was done to study the distribution of virulence factors, O-serogroups and antibiotic resistance properties of Shiga-toxigenic Escherichia coli isolated from milk and dairy products. Six-hundred samples were randomly collected and immediately transferred to laboratory. All samples were cultured and E. coli strains were isolated. STEC strains were identified based on the presence of putative virulence factors and subtypes. STEC isolates were subjected to multiplex PCR and disk diffusion methods. One-hundred and eighty-one out of 600 samples (30.16%) harbored E. coli. Prevalence of STEC strains was 10.66%. O157 (43.75%) and O26 (37.50%) were the most frequently identified serogroups. Aac(3)-IV (100%), CITM (96.87%) and tetA (76.56%) were the most commonly detected antibiotic resistance genes. STEC strains had the highest prevalence of resistance against ampicillin (100%), gentamicin (100%) and tetracycline (96.87%). Kashk and dough were negative for presence of E. coli strains. High prevalence of resistant-O157 strains and simultaneous presence of multiple virulence factors pose an important public health problem regarding the consumption of raw milk and dairy products.