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Автор: Grafiati

Опубліковано: 6 вересня 2023

Оновлено: 7 вересня 2023

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Aguirre-Ramírez, Marisela, Jesús Ramírez-Santos, Laurence Van Melderen, and M. Carmen Gómez-Eichelmann. "Expression of the F plasmid ccd toxin–antitoxin system in Escherichia coli cells under nutritional stress." Canadian Journal of Microbiology 52, no. 1 (January 1, 2006): 24–30. http://dx.doi.org/10.1139/w05-107.

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The ccd system of the F plasmid encodes CcdB, a protein toxic to DNA-gyrase, and CcdA, its antitoxin. The function attributed to this system is to contribute to plasmid stability by killing bacteria that lose the plasmid during cell division. However, the function of ccd in resting bacteria is not clear. Results presented show that ccd transcription increases as bacteria enter stationary phase and that the amount of the Ccd proteins is higher in bacteria under nutritional stress than in growing bacteria. Moreover, an increase in the frequency of Lac+ "adaptive" mutations was observed in stationary-phase bacteria that over-express the Ccd proteins.Key words: ccd system, nutritional stress, adaptive mutation.

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Burger, Virginia M., Alexandra Vandervelde, Jelle Hendrix, Albert Konijnenberg, Frank Sobott, Remy Loris, and Collin M. Stultz. "Hidden States within Disordered Regions of the CcdA Antitoxin Protein." Journal of the American Chemical Society 139, no. 7 (February 8, 2017): 2693–701. http://dx.doi.org/10.1021/jacs.6b11450.

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Madl, Tobias, Laurence Van Melderen, Natacha Mine, Michal Respondek, Monika Oberer, Walter Keller, Leila Khatai, and Klaus Zangger. "Structural Basis for Nucleic Acid and Toxin Recognition of the Bacterial Antitoxin CcdA." Journal of Molecular Biology 364, no. 2 (November 2006): 170–85. http://dx.doi.org/10.1016/j.jmb.2006.08.082.

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Lepka, Daniela, Tobias Kerrinnes, Evelyn Skiebe, Birgitt Hahn, Angelika Fruth, and Gottfried Wilharm. "Adding toYersinia enterocoliticaGene Pool Diversity: Two Cryptic Plasmids from a Biotype 1A Isolate." Journal of Biomedicine and Biotechnology 2009 (2009): 1–10. http://dx.doi.org/10.1155/2009/398434.

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We report the nucleotide sequence of two novel cryptic plasmids (4357 and 14 662 base pairs) carried by aYersinia enterocoliticabiotype 1A strain isolated from pork. As distinguished from most biotype 1A strains, this isolate, designated 07-04449, exhibited adherence to eukaryotic cells. The smaller plasmid pYe4449-1 carries five attributable open reading frames (ORFs) encoding the first CcdA/CcdB-like antitoxin/toxin system described for aYersiniaplasmid, a RepA-like replication initiation protein, and mobilizing factors MobA and MobC. The deduced amino acid sequences showed highest similarity to proteins described inSalmonella(CcdA/B),Klebsiella(RepA), andPlesiomonas(MobA/C) indicating genomic fluidity among members of theEnterobacteriaceae. One additional ORF with unknown function, termed ORF5, was identified with an ancestry distinct from the rest of the plasmid. While the C+G content of ORF5 is 38.3%, the rest of pYe4449-1 shows a C+G content of 55.7%. The C+G content of the larger plasmid pYe4449-2 (54.9%) was similar to that of pYe4449-1 (53.7%) and differed from that of theY. enterocoliticagenome (47.3%). Of the 14 ORFs identified on pYe4449-2, only six ORFs showed significant similarity to database entries. For three of these ORFs likely functions could be ascribed: a TnpR-like resolvase and a phage replication protein, localized each on a low C+G island, and DNA primase TraC. Two ORFs of pYe4449-2, ORF3 and ORF7, seem to encode secretable proteins. Epitope-tagging of ORF3 revealed protein expression at4°Cbut not at or above27°Csuggesting adaptation to a habitat outside swine. The hypothetical protein encoded by ORF7 is the member of a novel repeat protein family sharing theDxxGN(x)nDxxGNmotif. Our findings illustrate the exceptional gene pool diversity within the speciesY. enterocoliticadriven by horizontal gene transfer events.

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Kwak, Yee Gyung, George A. Jacoby, and David C. Hooper. "Effect of Qnr on Plasmid Gyrase Toxins CcdB and ParE." Antimicrobial Agents and Chemotherapy 59, no. 8 (June 8, 2015): 5078–79. http://dx.doi.org/10.1128/aac.00524-15.

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ABSTRACTPlasmid toxins CcdB and ParE are part of addiction systems promoting plasmid maintenance. Both target host DNA gyrase, as do quinolones and plasmid-determined Qnr proteins that protect gyrase from quinolone inhibition. We clonedqnrB4,qnrS1,ccdB,parE, and the antitoxin-encoding genesccdAandparDon compatible plasmids and tested them in combination. CcdB and ParE had no specific effect on quinolone susceptibility or Qnr protection, and Qnr did not act as a CcdB or ParE antitoxin.

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Cotrim, Camila Aparecida, Saulo Santesso Garrido, Eliane Trovatti, and Reinaldo Marchetto. "Síntese, caracterização e estudos de interação de um análogo da antitoxina CcdA empregando fluorescência no estado estacionário." Química Nova 33, no. 4 (2010): 841–45. http://dx.doi.org/10.1590/s0100-40422010000400014.

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Wu, Alma Y., Muhammad Kamruzzaman, and Jonathan R. Iredell. "Specialised functions of two common plasmid mediated toxin-antitoxin systems, ccdAB and pemIK, in Enterobacteriaceae." PLOS ONE 15, no. 6 (June 30, 2020): e0230652. http://dx.doi.org/10.1371/journal.pone.0230652.

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Isaeva, A. S., E. E. Kulikov, N. V. Ravin, B. D. Dorokhov, K. K. Tarasyan, and A. V. Letarov. "Application of the new ccdAB-type natural toxin-antitoxin module for stabilization of inheritance of expressive plasmid vectors based on the bacteriophage N15 replicon in Escherichia coli cells." Microbiology 79, no. 5 (October 2010): 638–45. http://dx.doi.org/10.1134/s0026261710050085.

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Hudson, Lauren K., Lisha Constantine-Renna, Linda Thomas, Christina Moore, Xiaorong Qian, Katie Garman, John R. Dunn, and Thomas G. Denes. "Genomic characterization and phylogenetic analysis of Salmonella enterica serovar Javiana." PeerJ 8 (November 20, 2020): e10256. http://dx.doi.org/10.7717/peerj.10256.

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Salmonella enterica serovar Javiana is the fourth most reported serovar of laboratory-confirmed human Salmonella infections in the U.S. and in Tennessee (TN). Although Salmonella ser. Javiana is a common cause of human infection, the majority of cases are sporadic in nature rather than outbreak-associated. To better understand Salmonella ser. Javiana microbial population structure in TN, we completed a phylogenetic analysis of 111 Salmonella ser. Javiana clinical isolates from TN collected from Jan. 2017 to Oct. 2018. We identified mobile genetic elements and genes known to confer antibiotic resistance present in the isolates, and performed a pan-genome-wide association study (pan-GWAS) to compare gene content between clades identified in this study. The population structure of TN Salmonella ser. Javiana clinical isolates consisted of three genetic clades: TN clade I (n = 54), TN clade II (n = 4), and TN clade III (n = 48). Using a 5, 10, and 25 hqSNP distance threshold for cluster identification, nine, 12, and 10 potential epidemiologically-relevant clusters were identified, respectively. The majority of genes that were found to be over-represented in specific clades were located in mobile genetic element (MGE) regions, including genes encoding integrases and phage structures (91.5%). Additionally, a large portion of the over-represented genes from TN clade II (44.9%) were located on an 87.5 kb plasmid containing genes encoding a toxin/antitoxin system (ccdAB). Additionally, we completed phylogenetic analyses of global Salmonella ser. Javiana datasets to gain a broader insight into the population structure of this serovar. We found that the global phylogeny consisted of three major clades (one of which all of the TN isolates belonged to) and two cgMLST eBurstGroups (ceBGs) and that the branch length between the two Salmonella ser. Javiana ceBGs (1,423 allelic differences) was comparable to those from other serovars that have been reported as polyphyletic (929–2,850 allelic differences). This study demonstrates the population structure of TN and global Salmonella ser. Javiana isolates, a clinically important Salmonella serovar and can provide guidance for phylogenetic cluster analyses for public health surveillance and response.

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Chandra, Soumyanetra, Kritika Gupta, Shruti Khare, Pehu Kohli, Aparna Asok, Sonali Vishwa Mohan, Harsha Gowda, and Raghavan Varadarajan. "The high mutational sensitivity of ccdA antitoxin is linked to codon optimality." Molecular Biology and Evolution, September 7, 2022. http://dx.doi.org/10.1093/molbev/msac187.

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Abstract Deep mutational scanning studies suggest that synonymous mutations are typically silent and that most exposed, non active-site residues are tolerant to mutations. Here we show that the ccdA antitoxin component of the E.coli ccdAB toxin-antitoxin system is unusually sensitive to mutations when studied in the operonic context. A large fraction (∼80%) of single-codon mutations, including many synonymous mutations in the ccdA gene shows inactive phenotype, but they retain native-like binding affinity towards cognate toxin, CcdB. Therefore, the observed phenotypic effects are largely not due to alterations in protein structure/stability, consistent with a large region of CcdA being intrinsically disordered. E. coli codon preference and strength of ribosome-binding associated with translation of downstream ccdB gene are found to be major contributors of the observed mutant phenotypes. In select cases, proteomics studies reveal altered ratios of CcdA:CcdB protein levels in vivo, suggesting that the ccdA mutations likely alter relative translation efficiencies of the two genes in the operon. We extend these results by studying single-site synonymous mutations that lead to loss of function phenotypes in the relBE operon upon introduction of rarer codons. Thus, in their operonic context, genes are likely to be more sensitive to both synonymous and non-synonymous point mutations than inferred previously.

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McCuaig, Frazer, Jody Winter, Jonathan Thomas, Gareth McVicker, and Lesley Hoyles. "Genotypic and phenotypic characterisation of the Klebsiella oxytoca complex." Access Microbiology 2, no. 7A (July 1, 2020). http://dx.doi.org/10.1099/acmi.ac2020.po0163.

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Klebsiella spp. are associated with 3 to 7% of nosocomial infections and can be responsible for a range of conditions including pneumonia, bloodstream infections, meningitis, and necrotizing enterocolitis in infants. The role of Klebsiella pneumoniae in causing disease is well-characterised but, to date, the closely related species Klebsiella oxytoca has not received the same attention, despite often encoding extended-spectrum beta-lactamases and carbapenemases in clinical settings. K. oxytoca is the causative agent of Clostridiodes difficile-negative antibiotic-associated haemorrhagic colitis, a rare condition seen in some individuals receiving antibiotics. Whole-genome sequence analyses have shown K. oxytoca to be a complex comprising at least six species (K. oxytoca, K. michiganensis, K. grimontii, K. huaxiensis, ‘K. pasteurii’, ‘K. spallanzanii’). Our study aims to better characterise the K. oxytoca complex using a polyphasic approach. Preliminary investigations into the genomes of three K. michiganensis clinical isolates revealed the presence of a plasmid-borne ccdABlocus. ccdAB is a toxin-antitoxin (TA) system known to maintain plasmids in other pathogenic enterobacteria. We aim to functionally validate this TA system by cloning and conducting toxicity assays on the CcdB toxin, and cloning and assessing the ability of CcdA to function as an antidote. We also aim to sequence and generate Illumina/Oxford Nanopore hybrid genome assemblies of a larger collection of K. oxytoca complex clinical isolates and investigate their plasmids and TA systems in the same manner.

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Zavrtanik, Uroš, San Hadži, and Jurij Lah. "Unraveling the Thermodynamics of Ultra-tight Binding of Intrinsically Disordered Proteins." Frontiers in Molecular Biosciences 8 (August 31, 2021). http://dx.doi.org/10.3389/fmolb.2021.726824.

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Protein interactions mediated by the intrinsically disordered proteins (IDPs) are generally associated with lower affinities compared to those between globular proteins. Here, we characterize the association between the intrinsically disordered HigA2 antitoxin and its globular target HigB2 toxin from Vibrio cholerae using competition ITC experiments. We demonstrate that this interaction reaches one of the highest affinities reported for IDP-target systems (KD = 3 pM) and can be entirely attributed to a short, 20-residue-long interaction motif that folds into α-helix upon binding. We perform an experimentally based decomposition of the IDP-target association parameters into folding and binding contributions, which allows a direct comparison of the binding contribution with those from globular ultra-high affinity binders. We find that the HigA2-HigB2 interface is energy optimized to a similar extent as the interfaces of globular ultra-high affinity complexes, such as barnase-barstar. Evaluation of other ultra-high affinity IDP-target systems shows that a strategy based on entropy optimization can also achieve comparably high, picomolar affinities. Taken together, these examples show how IDP-target interactions achieve picomolar affinities either through enthalpy optimization (HigA2-HigB2), resembling the ultra-high affinity binding of globular proteins, or via bound-state fuzziness and entropy optimization (CcdA-CcdB, histone H1-prothymosin α).

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Gupta, Kritika, Arti Tripathi, Alishan Sahu, and Raghavan Varadarajan. "Contribution of the Chromosomal ccdAB Operon to Bacterial Drug Tolerance." Journal of Bacteriology 199, no. 19 (July 3, 2017). http://dx.doi.org/10.1128/jb.00397-17.

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ABSTRACT One of the first identified and best-studied toxin-antitoxin (TA) systems in Escherichia coli is the F-plasmid-based CcdAB system. This system is involved in plasmid maintenance through postsegregational killing. More recently, ccdAB homologs have been found on the chromosome, including in pathogenic strains of E. coli and other bacteria. However, the functional role of chromosomal ccdAB genes, if any, has remained unclear. We show that both the native ccd operon of the E. coli O157 strain (ccd O157) and the ccd operon from the F plasmid (ccd F), when inserted on the E. coli chromosome, lead to protection from cell death under multiple antibiotic stress conditions through formation of persisters, with the O157 operon showing higher protection. While the plasmid-encoded CcdB toxin is a potent gyrase inhibitor and leads to bacterial cell death even under fully repressed conditions, the chromosomally encoded toxin leads to growth inhibition, except at high expression levels, where some cell death is seen. This was further confirmed by transiently activating the chromosomal ccd operon through overexpression of an active-site inactive mutant of F-plasmid-encoded CcdB. Both the ccd F and ccd O157 operons may share common mechanisms for activation under stress conditions, eventually leading to multidrug-tolerant persister cells. This study clearly demonstrates an important role for chromosomal ccd systems in bacterial persistence. IMPORTANCE A large number of free-living and pathogenic bacteria are known to harbor multiple toxin-antitoxin systems, on plasmids as well as on chromosomes. The F-plasmid CcdAB system has been extensively studied and is known to be involved in plasmid maintenance. However, little is known about the function of its chromosomal counterpart, found in several pathogenic E. coli strains. We show that the native chromosomal ccd operon of the E. coli O157 strain is involved in drug tolerance and confers protection from cell death under multiple antibiotic stress conditions. This has implications for generation of potential therapeutics that target these TA systems and has clinical significance because the presence of persisters in an antibiotic-treated population can lead to resuscitation of chronic infection and may contribute to failure of antibiotic treatment.

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Al Marjania, Mohammed F., Ebrahim Kouhsari, Fatima S. Ali, and Sawsan H. Authman. "Evaluation of type II toxin-antitoxin systems, antibiotic resistance profiles, and biofilm quorum sensing genes in Acinetobacter baumannii isolates in Iraq." Infectious Disorders - Drug Targets 20 (May 25, 2020). http://dx.doi.org/10.2174/1871526520666200525170318.

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Background: Bacterial Toxin-Antitoxin (TAs) systems are extensive two-component elements in the bacterial genome, which involved in many key biological functions including growth arrest, survival, biofilm formation, plasmid maintenance, defense against phages, persistence and virulence. Aim: This study aimed to assess the molecular determinants involved in TAs, biofilm quorum sensing, and antibiotic resistance profiles in Acinetobacter baumannii isolated from Baghdad`s hospitals in Iraq. Methods: A total of 127 A. baumannii isolates were collected from 2160 different clinical samples. The antimicrobial susceptibility test was performed using disk diffusion test. All isolates were characterized for molecular determinants involved in TAs and biofilm formation using the well-known PCR-based sequencing assay. Results: A high multi-drug resistant (MDR) (96.06%; 122/127 ) and imipenem resistance (84.25%; 107/127 ) rates were observed from A.baumannii isolates. Results showed the presence of rhlIR gene in three isolates (2.36%), and lasIR gene appeared in two isolates (1.57%) isolates, whilst, mazEF, ccdAB, and relBE genes have not detected among isolates. Conclusion: A high MDR and imipenem resistance rates within a low prevalence of rhlIR, and lasIR genes could be found in clinical A. baumannii isolates from some of Iraqi hospitals.

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Fan, Lin-Ping, Yang Yu, Shanshan Huang, Wenjian Liao, Qi-Sen Huang, Fang-Ling Du, Tian-xin Xiang, et al. "Genetic characterization and passage instability of a novel hybrid virulence plasmid in a ST23 hypervirulent Klebsiella pneumoniae." Frontiers in Cellular and Infection Microbiology 12 (July 28, 2022). http://dx.doi.org/10.3389/fcimb.2022.870779.

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Hypervirulent variants of Klebsiella pnuemoniae (hvKP), which causes life-threatening infections, is a global priority pathogen and frequently harbours virulence plasmids. The virulence plasmids have emerged as the predominant vehicles carrying the major pathogenic determinants of hypermucoviscosity and hypervirulence phenotypes. In the present study, we characterized a novel virulence plasmid in AP8555, an ST23 hvKP strain, which induced a metastatic infection and fatal septic shock in a critically ill patient. The serum killing assay, the quantitative biofilm formation assay, the G.mellonella infection model, and the mouse lethality assay demonstrated that AP8555 was almost as virulent as the hvKP strain NUTH-K2044. The plasmid pAP855 could be conjugated to Klebsiella quasipneumoniae ATCC700603 and E. coli J53 at a frequency of 7.2× 10−5 and 8.7× 10−7, respectively. Whole-genome sequencing and bioinformatics analysis confirmed that the plasmid was novel, clustered to the incompatibility type of IncHI1B/IncFIB/IncFII and presented high similarity to the pK2044 plasmid. In contrast, a 130-kb large-fragment insertion was observed on the plasmid, which introduced a genetic hybrid zone with multiple conjugation-related genes of type IV secretion systems (T4SS) and CcdAB toxin-antitoxin systems (TAS) to the plasmid. In the transconjugants, the presence of pAP855 had a negative impact on bacterial fitness, but enhancing the virulence-associated phenotypes. In vitro evolution experiments showed that pAP855 in the transconjugants could not be stably inherited after 10 days of passage. Our study not only reports a novel hybrid plasmid but also highlights the putative pathway of conjugative virulence plasmid formation and evolution by means of genetic rearrangement through sequence insertion. These findings indicate that structural versatility could contribute to the dissemination of cointegrate virulence plasmid, although the plasmid incurred a fitness cost. Therefore, continuous monitoring the acquisition of conjugative virulence plasmids may have critical value for plasmid research and increase awareness of hvKP.

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