scholarly journals Dinucleoside polyphosphates act as 5’-RNA caps in Escherichia coli

2019 ◽  
Author(s):  
Oldřich Hudeček ◽  
Roberto Benoni ◽  
Martin Culka ◽  
Martin Hubálek ◽  
Lubomír Rulíšek ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Rna Stability ◽  
E Coli ◽  
Chemical Structures ◽  
Additional Layer ◽  
Similar Chemical ◽  
Decapping Enzyme ◽  
Almost All ◽  
Dinucleoside Polyphosphates ◽  
Rna Caps

Dinucleoside polyphosphates (NpnNs), discovered more than 50 years ago,1 are pleiotropic molecules present in almost all types of cells.2 It has been shown that their intracellular concentration can under stress conditions increase from the µM to mM range 2,3. However, the cellular roles and mechanisms of action of NpnNs are still speculative4,5. They have never been considered as part of the RNA, even though they have similar chemical structures as already known RNA caps, such as the nicotinamide adenine dinucleotide (NAD)6-8 and 7-methylguanylate cap9. Here, we show that both methylated and non-methylated Npn Ns serve as RNA caps in Escherichia coli (E. coli). NpnNs are excellent substrates for T7 and E. coli RNA polymerases (RNAP) and efficiently initiate transcription. Further, we demonstrate that the E. coli decapping enzyme RNA 5’ pyrophosphohydrolase (RppH) is able to remove the NpnNs-cap from the RNA. RppH was, however, not able to cleave the methylated forms of the NpnN-caps, suggesting that the methylation adds an additional layer to the RNA stability regulation. Our work introduces an original perspective on the chemical structure of RNA in prokaryotes and the function of RNA caps. This is the first evidence that small molecules like NpnNs can act in cells via their incorporation into RNA and influence the cellular metabolism.

scholarly journals Phylogenetic Grouping and Virulence Potential of Extended-Spectrum-β-Lactamase-Producing Escherichia coli Strains in Cattle

2012 ◽  
Vol 78 (13) ◽  
pp. 4677-4682 ◽  
Author(s):  
Charlotte Valat ◽  
Frédéric Auvray ◽  
Karine Forest ◽  
Véronique Métayer ◽  
Emilie Gay ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Virulence Determinants ◽  
Phylogenetic Groups ◽  
Content Type ◽  
E Coli ◽  
Extended Spectrum ◽  
Specific Distribution ◽  
Extended Spectrum Beta Lactamases ◽  
Phylogenetic Grouping ◽  
Almost All

ABSTRACTIn line with recent reports of extended-spectrum beta-lactamases (ESBLs) inEscherichia coliisolates of highly virulent serotypes, such as O104:H4, we investigated the distribution of phylogroups (A, B1, B2, D) and virulence factor (VF)-encoding genes in 204 ESBL-producingE. coliisolates from diarrheic cattle. ESBL genes, VFs, and phylogroups were identified by PCR and a commercial DNA array (Alere, France). ESBL genes belonged mostly to the CTX-M-1 (65.7%) and CTX-M-9 (27.0%) groups, whereas those of the CTX-M-2 and TEM groups were much less represented (3.9% and 3.4%, respectively). One ESBL isolate wasstx1andeaepositive and belonged to a major enterohemorrhagicE. coli(EHEC) serotype (O111:H8). Two other isolates wereeaepositive butstxnegative; one of these had serotype O26:H11. ESBL isolates belonged mainly to phylogroup A (55.4%) and, to lesser extents, to phylogroups D (25.5%) and B1 (15.6%), whereas B2 strains were quasi-absent (1/204). The number of VFs was significantly higher in phylogroup B1 than in phylogroups A (P= 0.04) and D (P= 0.02). Almost all of the VFs detected were found in CTX-M-1 isolates, whereas only 64.3% and 33.3% of them were found in CTX-M-9 and CTX-M-2 isolates, respectively. These results indicated that the widespread dissemination of theblaCTX-Mgenes within theE. colipopulation from cattle still spared the subpopulation of EHEC/Shiga-toxigenicE. coli(STEC) isolates. In contrast to other reports on non-ESBL-producing isolates from domestic animals, B1 was not the main phylogroup identified. However, B1 was found to be the most virulent phylogroup, suggesting host-specific distribution of virulence determinants among phylogenetic groups.

scholarly journals Additional Og-Typing PCR Techniques Targeting Escherichia coli-Novel and Shigella-Unique O-Antigen Biosynthesis Gene Clusters

2020 ◽  
Vol 58 (11) ◽  
Author(s):  
Atsushi Iguchi ◽  
Hironobu Nishii ◽  
Kazuko Seto ◽  
Jiro Mitobe ◽  
Kenichi Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Strong Association ◽  
Gene Clusters ◽  
Epidemiological Studies ◽  
Content Type ◽  
E Coli ◽  
O Antigen ◽  
Wide Range ◽  
Biosynthesis Gene ◽  
Almost All

ABSTRACT The O-serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and controls. O-serogroup diversification shows a strong association with the genetic diversity in some O-antigen biosynthesis gene clusters. Through genomic studies, in addition to the types of O-antigen biosynthesis gene clusters (Og-types) from conventional O-serogroup strains, a number of novel Og-types have been found in E. coli isolates. To assist outbreak investigations and surveillance of pathogenic E. coli at inspection institutes, in previous studies, we developed PCR methods that could determine almost all conventional O-serogroups and some novel Og-types. However, there are still many Og-types that may not be determined by simple genetic methods such as PCR. Thus, in the present study, we aimed to develop an additional Og-typing PCR system. Based on the novel Og-types, including OgN32, OgN33, and OgN34, presented in this study, we designed an additional 24 PCR primer pairs targeting 14 novel and 2 diversified E. coli Og-types and 8 Shigella-unique Og-types. Subsequently, we developed 5 new multiplex PCR sets consisting of 33 primers, including the aforementioned 24 primers and 9 primers reported in previous studies. The accuracy and specificity of the PCR system was validated using approximately 260 E. coli and Shigella O-serogroup and Og-type reference strains. The Og-typing PCR system reported here can determine a wide range of Og-types of E. coli and may help epidemiological studies, in addition to the surveillance of pathogenic E. coli.

scholarly journals In vivo characterization of the activities of novel cyclodipeptide oxidases: new tools for increasing chemical diversity of bioproduced 2,5-diketopiperazines in Escherichia coli

2020 ◽  
Vol 19 (1) ◽  
Author(s):  
Fabien Le Chevalier ◽  
Isabelle Correia ◽  
Lucrèce Matheron ◽  
Morgan Babin ◽  
Mireille Moutiez ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biological Activities ◽  
Gene Clusters ◽  
Chemical Diversity ◽  
E Coli ◽  
Chemical Structures ◽  
In Vivo Characterization ◽  
Culture Supernatants

Abstract Background Cyclodipeptide oxidases (CDOs) are enzymes involved in the biosynthesis of 2,5-diketopiperazines, a class of naturally occurring compounds with a large range of pharmaceutical activities. CDOs belong to cyclodipeptide synthase (CDPS)-dependent pathways, in which they play an early role in the chemical diversification of cyclodipeptides by introducing Cα-Cβ dehydrogenations. Although the activities of more than 100 CDPSs have been determined, the activities of only a few CDOs have been characterized. Furthermore, the assessment of the CDO activities on chemically-synthesized cyclodipeptides has shown these enzymes to be relatively promiscuous, making them interesting tools for cyclodipeptide chemical diversification. The purpose of this study is to provide the first completely microbial toolkit for the efficient bioproduction of a variety of dehydrogenated 2,5-diketopiperazines. Results We mined genomes for CDOs encoded in biosynthetic gene clusters of CDPS-dependent pathways and selected several for characterization. We co-expressed each with their associated CDPS in the pathway using Escherichia coli as a chassis and showed that the cyclodipeptides and the dehydrogenated derivatives were produced in the culture supernatants. We determined the biological activities of the six novel CDOs by solving the chemical structures of the biologically produced dehydrogenated cyclodipeptides. Then, we assessed the six novel CDOs plus two previously characterized CDOs in combinatorial engineering experiments in E. coli. We co-expressed each of the eight CDOs with each of 18 CDPSs selected for the diversity of cyclodipeptides they synthesize. We detected more than 50 dehydrogenated cyclodipeptides and determined the best CDPS/CDO combinations to optimize the production of 23. Conclusions Our study establishes the usefulness of CDPS and CDO for the bioproduction of dehydrogenated cyclodipeptides. It constitutes the first step toward the bioproduction of more complex and diverse 2,5-diketopiperazines.

scholarly journals Stress-Induced MazF-Mediated Proteins in Escherichia coli

mBio ◽  
2019 ◽  
Vol 10 (2) ◽  
Author(s):  
Akanksha Nigam ◽  
Tamar Ziv ◽  
Adi Oron-Gottesman ◽  
Hanna Engelberg-Kulka
Keyword(s):  
Escherichia Coli ◽  
Proteomic Analysis ◽  
Normal Growth ◽  
Growth Conditions ◽  
Translation Machinery ◽  
Content Type ◽  
E Coli ◽  
Adverse Conditions ◽  
Almost All ◽  
Induced Proteins

ABSTRACT Escherichia coli mazEF is an extensively studied stress-induced toxin-antitoxin (TA) system. The toxin MazF is an endoribonuclease that cleaves RNAs at ACA sites. By that means, under stress, the induced MazF generates a stress-induced translation machinery (STM) composed of MazF-processed mRNAs and selective ribosomes that specifically translate the processed mRNAs. Here, we performed a proteomic analysis of all the E. coli stress-induced proteins that are mediated through the chromosomally borne mazF gene. We show that the mRNAs of almost all of them are characterized by the presence of an ACA site up to 100 nucleotides upstream of the AUG initiator. Therefore, under stressful conditions, induced MazF processes mRNAs that are translated by STM. Furthermore, the presence of the ACA sites far upstream (up to 100 nucleotides) of the AUG initiator may still permit translation by the canonical translation machinery. Thus, such dual-translation mechanisms enable the bacterium under stress also to prepare proteins for immediate functions while coming back to normal growth conditions. IMPORTANCE The stress response, the strategy that bacteria have developed in order to cope up with all kinds of adverse conditions, is so far understood at the level of transcription. Our previous findings of a uniquely modified stress-induced translation machinery (STM) generated in E. coli under stress by the endoribonucleolytic activity of the toxin MazF opens a new chapter in understanding microbial physiology under stress at the translational level. Here, we performed a proteomic analysis of all the E. coli stress-induced proteins that are mediated by chromosomally borne MazF through STM.

scholarly journals Structure-activity relationships of 3-substituted-5,5- diphenylhydantoins as potential antiproliferative and antimicrobial agents

2011 ◽  
Vol 76 (12) ◽  
pp. 1597-1606 ◽  
Author(s):  
Nemanja Trisovic ◽  
Bojan Bozic ◽  
Ana Obradovic ◽  
Olgica Stefanovic ◽  
Snezana Markovic ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antimicrobial Agents ◽  
Human Colon ◽  
Antibacterial Activities ◽  
E Coli ◽  
Structure Activity Relationships ◽  
Structure Activity ◽  
Almost All

A series of twelve 3-substituted-5,5-diphenylhydantoins was synthesized, including some whose anticonvulsant activities have already been reported in the literature. Their antiproliferative activities against HCT-116 human colon carcinoma cells were evaluated to determine structure-activity relationships. Almost all of the compounds exhibited statistically significant antiproliferative effects at a concentration of 100 ?M, while the derivative bearing a benzyl group was active even at lower concentrations. Moreover, their in vitro antibacterial activities against Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923 and clinical isolates of Escherichia coli, Proteus mirabilis, Pseudomonas aeruginosa, Enterococcus faecalis and Staphylococcus aureus were evaluated. Only the 3-iso-propyl and 3-benzyl derivatives showed weak antibacterial activities against the Gram-positive bacterium E. faecalis and the Gram-negative bacteria E. coli ATCC 25922 and E. coli.

scholarly journals Intron-assisted, viroid-based production of insecticidal circular double-stranded RNA in Escherichia coli

2020 ◽  
Author(s):  
Beltrán Ortolá ◽  
Teresa Cordero ◽  
Xu Hu ◽  
José-Antonio Daròs
Keyword(s):  
Escherichia Coli ◽  
Large Scale ◽  
Rna Stability ◽  
Diabrotica Virgifera Virgifera ◽  
Septate Junction ◽  
Circular Rnas ◽  
Double Stranded Rna ◽  
E Coli ◽  
Group I ◽  
Insecticide Activity

ABSTRACTRNA interference (RNAi) is a natural mechanism for protecting against harmful genetic elements and regulating gene expression, which can be artificially triggered by the delivery of homologous double-stranded RNA (dsRNA). This mechanism can be exploited as a highly specific and environmentally friendly pest control strategy. To this aim, systems for producing large amounts of recombinant dsRNA are necessary. We describe a system to efficiently produce large amounts of circular dsRNA in Escherichia coli and demonstrate the efficient insecticidal activity of these molecules against Western corn rootworm (WCR, Diabrotica virgifera virgifera LeConte), a highly damaging pest of corn crops. In our system, the two strands of the dsRNA are expressed in E. coli embedded within the very stable scaffold of Eggplant latent viroid (ELVd), a small circular non-coding RNA. Stability in E. coli of the corresponding plasmids with long inverted repeats was achieved by using a cDNA coding for a group-I autocatalytic intron from Tetrahymena thermophila as a spacer. RNA circularization and large-scale accumulation in E. coli cells was facilitated by co-expression of eggplant tRNA ligase, the enzyme that ligates ELVd during replication in the host plant. The inserted intron efficiently self-spliced from the RNA product during transcription. Circular RNAs containing a dsRNA moiety homologous to smooth septate junction 1 (DvSSJ1) gene exhibited excellent insecticide activity against WCR larvae. Finally, we show that the viroid scaffold can be separated from the final circular dsRNA product using a second T. thermophila self-splicing intron in a permuted form.

scholarly journals Emergence of Escherichia coli encoding Shiga toxin 2f in human Shiga toxin-producing E. coli (STEC) infections in the Netherlands, January 2008 to December 2011

2014 ◽  
Vol 19 (17) ◽  
Author(s):  
I Friesema ◽  
K van der Zwaluw ◽  
T Schuurman ◽  
M Kooistra-Smid ◽  
E Franz ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
The Netherlands ◽  
Shiga Toxin ◽  
Distinct Group ◽  
Shiga Toxins ◽  
E Coli ◽  
Mild Disease ◽  
Shiga Toxin 2 ◽  
Almost All ◽  
Human Infections

The Shiga toxins of Shiga toxin-producing Escherichia coli (STEC) can be divided into Shiga toxin 1 (Stx1) and Shiga toxin 2 (Stx2) with several sub-variants. Variant Stx2f is one of the latest described, but has been rarely associated with symptomatic human infections. In the enhanced STEC surveillance in the Netherlands, 198 STEC O157 cases and 351 STEC non-O157 cases, including 87 stx2f STEC isolates, were reported between 2008 and 2011. Most stx2f strains belonged to the serogroups O63:H6 (n=47, 54%), O113:H6 (n=12, 14%) and O125:H6 (n=12, 14%). Of the 87 stx2f isolates, 84 (97%) harboured the E. coli attaching and effacing (eae) gene, but not the enterohaemorrhagic E. coli haemolysin (hly) gene. Stx2f STEC infections show milder symptoms and a less severe clinical course than STEC O157 infections. Almost all infections with stx2f (n=83, 95%) occurred between June and December, compared to 170/198 (86%) of STEC O157 and 173/264 (66%) of other STEC non-O157. Stx2f STEC infections in the Netherlands are more common than anticipated, and form a distinct group within STEC with regard to virulence genes and the relatively mild disease.

scholarly journals AcrAB Efflux System: Expression and Contribution to Fluoroquinolone Resistance in Klebsiella spp

2002 ◽  
Vol 46 (12) ◽  
pp. 3984-3986 ◽  
Author(s):  
Annarita Mazzariol ◽  
Jessica Zuliani ◽  
Giuseppe Cornaglia ◽  
Gian Maria Rossolini ◽  
Roberta Fontana
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Oxytoca ◽  
Fluoroquinolone Resistance ◽  
Multidrug Efflux ◽  
Efflux System ◽  
E Coli ◽  
Almost All ◽  
Klebsiella Spp ◽  
Different Levels ◽  
Multidrug Efflux System

ABSTRACT Seven Klebsiella pneumoniae and four Klebsiella oxytoca clinical isolates with different levels of resistance to ciprofloxacin were studied. Mutations in the topoisomerase genes were found in almost all strains, but the contribution of a multidrug efflux system homologous to AcrAB in Escherichia coli was also observed. Overexpression of this efflux system was demonstrated by immunoblotting with antibodies against E. coli AcrA.

Relationship between water quality parameters and the survival of indicator microorganisms – Escherichia coli – in a stormwater wetland

2013 ◽  
Vol 68 (7) ◽  
pp. 1650-1656 ◽  
Author(s):  
Jing Cheng ◽  
Siping Niu ◽  
Youngchul Kim
Keyword(s):  
Escherichia Coli ◽  
Water Quality ◽  
Pearson Correlation ◽  
Dilution Effect ◽  
Quality Parameters ◽  
Water Quality Parameters ◽  
Indicator Microorganisms ◽  
E Coli ◽  
Significant Seasonal Variation ◽  
Almost All

Indicator microorganisms – Escherichia coli – in a stormwater wetland controlling non-point source pollution in the Keum River basin was studied and the relationship between water quality parameters and the survival of E. coli was investigated by Pearson correlation analysis based on 16 sets of data collected on dry days. A significant seasonal variation of the density of E. coli was observed in the influent and effluent. A dominant decay of E. coli was found in almost all the wetland components. In the settling pond, the density was observed to vary negatively with pH and the removal of total suspended solids, and the decay was promoted by solar radiation. In the aeration pond, the survival of E. coli was encouraged negatively by the dilution effect, unfavorable increase of dissolved oxygen (DO) and sedimentation with coarse particles. In the marsh wetland, the reduction of E. coli was positively influenced by UV radiation, obvious increase of pH, DO and sedimentation with algae and particles. In the polishing pond, the introduction of E. coli with avian feces and growth of the bacteria were more prevalent than decay. Although this introduction or growth was almost negligible, further study will be conducted in the future.

Indoxyl-β-D-glucuronide, a novel chromogenic reagent for the specific detection and enumeration of Escherichia coli in environmental samples

1988 ◽  
Vol 34 (5) ◽  
pp. 690-693 ◽  
Author(s):  
Arthur Newton Ley ◽  
Raymond John Bowers ◽  
Saul Wolfe
Keyword(s):  
Escherichia Coli ◽  
Large Scale ◽  
Membrane Filter ◽  
Chloroacetic Acid ◽  
Specific Detection ◽  
E Coli ◽  
Filter Test ◽  
Almost All ◽  
First Time ◽  
Specific Reagent

About 97% of Escherichia coli strains produce β-glucuronidase, but almost all other Enterobacteriaceae lack this enzyme. A D-glucopyranosiduronic acid (glucuronide) possessing a readily detectable β-linked aglycone should, therefore, constitute a specific reagent for the detection of this organism. For this purpose, the title compound has been synthesized for the first time. The synthesis proceeds in eight steps from readily available D-glucuronolactone, anthranilic acid, and chloroacetic acid and can be carried out on a large scale. The compound has the predicted properties: when included in the standard membrane filter test for the analysis of water, indoxyl-β-D-glucuronide allows specific detection of E. coli through the formation of blue colonies that are the result of rapid conversion of the liberated aglycone to indigo. The recovery of E. coli is easily measured and almost quantitative.

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