ORCID Profile
0000-0003-2031-9679
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In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Microbial Genetics | Microbiology | Biochemistry and Cell Biology | Genomics | Bacteriology | Synthetic biology | Receptors and Membrane Biology | Genetics | Synthetic Biology | Systems Biology | Microbial Ecology | Microbial genetics | Bioinformatics | Genomics
Expanding Knowledge in the Biological Sciences | Environmentally Sustainable Plant Production not elsewhere classified | Wheat | Manufacturing not elsewhere classified | Rehabilitation of Degraded Urban and Industrial Environments | Control of Plant Pests, Diseases and Exotic Species in Farmland, Arable Cropland and Permanent Cropland Environments | Vegetables | Pome Fruit, Pip Fruit | Infectious Diseases |
Publisher: Microbiology Society
Date: 17-11-2021
Abstract: Competitive behaviours of plant growth promoting rhizobacteria (PGPR) are integral to their ability to colonize and persist on plant roots and outcompete phytopathogenic fungi, oomycetes and bacteria. PGPR engage in a range of antagonistic behaviours that have been studied in detail, such as the production and secretion of compounds inhibitory to other microbes. In contrast, their defensive activities that enable them to tolerate exposure to inhibitory compounds produced by their neighbours are less well understood. In this study, the genes involved in the Pseudomonas protegens Pf-5 response to metabolites from eight erse rhizosphere competitor organisms, Fusarium oxysporum , Rhizoctonia solani , Gaeumannomyces graminis var. tritici , Pythium spinosum , Bacillus subtilis QST713, Pseudomonas sp. Q2-87, Streptomyces griseus and Streptomyces bikiniensis subspecies bikiniensi , were examined. Proximity induced excreted metabolite responses were confirmed for Pf-5 with all partner organisms through HPLC before culturing a dense Pf-5 transposon mutant library adjacent to each of these microbes. This was followed by transposon-directed insertion site sequencing (TraDIS), which identified genes that influence Pf-5 fitness during these competitive interactions. A set of 148 genes was identified that were associated with increased fitness during competition, including cell surface modification, electron transport, nucleotide metabolism, as well as regulatory genes. In addition, 51 genes were identified for which loss of function resulted in fitness gains during competition. These included genes involved in flagella biosynthesis and cell ision. Considerable overlap was observed in the set of genes observed to provide a fitness benefit during competition with all eight test organisms, indicating commonalities in the competitive response to phylogenetically erse micro-organisms and providing new insight into competitive processes likely to take place in the rhizosphere.
Publisher: American Society for Microbiology
Date: 30-06-2020
DOI: 10.1128/MSYSTEMS.00106-20
Abstract: The threat of antimicrobial resistance to human health has prompted interest in complex, natural products with antimicrobial activity. Honey has been an effective topical wound treatment throughout history, predominantly due to its broad-spectrum antimicrobial activity. Unlike traditional antibiotics, honey-resistant bacteria have not been reported however, honey remains underutilized in the clinic in part due to a lack of understanding of its mechanism of action. Here, we demonstrate that honey affects multiple processes in bacteria, and this is not explained by its major antibacterial components. Honey also uniquely affects bacterial membranes, and this can be exploited for combination therapy with antibiotics that are otherwise ineffective on their own. We argue that honey should be included as part of the current array of wound treatments due to its effective antibacterial activity that does not promote resistance in bacteria.
Publisher: American Society for Microbiology
Date: 07-2013
DOI: 10.1128/IAI.00065-13
Abstract: Acinetobacter baumannii has become a major problem in the clinical setting with the prevalence of infections caused by multidrug-resistant strains on the increase. Nevertheless, only a limited number of molecular mechanisms involved in the success of A. baumannii as a human pathogen have been described. In this study, we examined the virulence features of a hypermotile derivative of A. baumannii strain ATCC 17978, which was found to display enhanced adherence to human pneumocytes and elevated levels of lethality toward Caenorhabditis elegans nematodes. Analysis of cellular lipids revealed modifications to the fatty acid composition, providing a possible explanation for the observed changes in hydrophobicity and subsequent alteration in adherence and motility. Comparison of the genome sequences of the hypermotile variant and parental strain revealed that an insertion sequence had disrupted an hns -like gene in the variant. This gene encodes a homologue of the histone-like nucleoid structuring (H-NS) protein, a known global transcriptional repressor. Transcriptome analysis identified the global effects of this mutation on gene expression, with major changes seen in the autotransporter Ata, a type VI secretion system, and a type I pilus cluster. Interestingly, isolation and analysis of a second independent hypermotile ATCC 17978 variant revealed a mutation to a residue within the DNA binding region of H-NS. Taken together, these mutants indicate that the phenotypic and transcriptomic differences seen are due to loss of regulatory control effected by H-NS.
Publisher: Wiley
Date: 06-2018
Abstract: Pseudomonas is a large and erse genus of Gammaproteobacteria. To provide a framework for discovery of evolutionary and taxonomic relationships of these bacteria, we compared the genomes of type strains of 163 species and 3 additional subspecies of Pseudomonas, including 118 genomes sequenced herein. A maximum likelihood phylogeny of the 166 type strains based on protein sequences of 100 single-copy orthologous genes revealed thirteen groups of Pseudomonas, composed of two to sixty three species each. Pairwise average nucleotide identities and alignment fractions were calculated for the data set of the 166 type strains and 1224 genomes of Pseudomonas available in public databases. Results revealed that 394 of the 1224 genomes were distinct from any type strain, suggesting that the type strains represent only a fraction of the genomic ersity of the genus. The core genome of Pseudomonas was determined to contain 794 genes conferring primarily housekeeping functions. The results of this study provide a phylogenetic framework for future studies aiming to resolve the classification and phylogenetic relationships, identify new gene functions and phenotypes, and explore the ecological and metabolic potential of the Pseudomonas spp.
Publisher: Cold Spring Harbor Laboratory
Date: 09-01-2021
DOI: 10.1101/2021.01.08.425997
Abstract: Plants live in association with microorganisms that positively influence plant development, vigor, and fitness in response to pathogens and abiotic stressors. The bulk of the plant microbiome is concentrated belowground at the plant root-soil interface. Plant roots secrete carbon-rich rhizodeposits containing primary and secondary low molecular-weight metabolites, lysates, and mucilages. These exudates provide nutrients for soil microorganisms and modulate their affinity to host plants, but molecular details of this process are largely unresolved. We addressed this gap by focusing on the molecular dialogue between eight well-characterized beneficial strains of the Pseudomonas fluorescens group and Brachypodium distachyon , a model for economically important food, feed, forage, and biomass crops of the grass family. We collected and analyzed root exudates of B. distachyon and demonstrated the presence of multiple carbohydrates, amino acids, organic acids and phenolic compounds. The subsequent screening of bacteria by Biolog Phenotype MicroArrays revealed that many of these metabolites provide carbon and energy for the Pseudomonas strains. RNA-seq profiling of bacterial cultures amended with root exudates revealed changes in the expression of genes encoding numerous catabolic and anabolic enzymes, transporters, transcriptional regulators, stress response, and conserved hypothetical proteins. Almost half of the differentially expressed genes mapped to the variable part of the strains’ pangenome, reflecting the importance of the variable gene content in the adaptation of P. fluorescens to the rhizosphere lifestyle. Our results collectively reveal the ersity of cellular pathways and physiological responses underlying the establishment of mutualistic interactions between these beneficial rhizobacteria and their plant hosts.
Publisher: American Society for Microbiology
Date: 21-12-2022
DOI: 10.1128/SPECTRUM.03593-22
Abstract: Gram-negative bacteria are problematic for antibiotic development due to the low permeability of their cell envelopes. To rationally design new antibiotics capable of breaching this barrier, more information is required about the specific components of the cell envelope that prevent the passage of compounds with different physiochemical properties. Ampicillin and benzylpenicillin are β-lactam antibiotics with identical chemical structures except for a clever synthetic addition of a primary amine group in icillin, which promotes its accumulation in Gram-negatives. Previous work showed that icillin is better able to pass through the outer membrane porin OmpF in Escherichia coli compared to benzylpenicillin. It is not known, however, how the primary amine may affect interaction with other cell envelope components. This study applied TraDIS to identify genes that affect E. coli fitness in the presence of equivalent subinhibitory concentrations of icillin and benzylpenicillin, with a focus on the cell envelope. Insertions that compromised the outer membrane, particularly the lipopolysaccharide layer, were found to decrease fitness under benzylpenicillin exposure, but had less effect on fitness under icillin treatment. These results align with expectations if benzylpenicillin is poorly able to pass through porins. Disruption of genes encoding the AcrAB-TolC efflux system were detrimental to survival under both antibiotics, but particularly icillin. Indeed, insertions in these genes and regulators of
Publisher: Elsevier
Date: 2019
Publisher: S. Karger AG
Date: 2011
DOI: 10.1159/000325367
Abstract: i Background: /i i Acinetobacter /i i baumannii /i is a major cause of nosocomial infections worldwide due to its fitness within clinical settings and recalcitrance to conventional therapies. The drug: H sup + /sup antiporter 2 (DHA2) family export systems encoded by i A. baumannii /i were investigated for their roles in promoting the success ofthis organism as a human pathogen. i Methods: /i Bioinformatic tools were used to identify the DHA2 family transporters encoded by i Acinetobacter /i spp. and establish their phylogenetic relationships. The drug resistance phenotypes conferred by the transporters were tested using both heterologously expressed proteins in i Escherichia coli /i and i Acinetobacter /i deletion mutants. The transcriptional responses of DHA2 family transporter genes to their substrates were established by qRT-PCR. i Results: /i Six highly conserved DHA2 family proteins were identified in i A. baumannii /i . Drug resistance phenotypes were established for two DHA2 family transporters. The expression of a third DHA2 family protein is highly responsive to the availability of iron. The gene encoding this protein is located within a putative siderophore biosynthesis locus, suggesting a physiological role in iron uptake, possibly via the export of a siderophore. i Conclusions: /i These results highlight functions for DHA2 family proteins in both drug resistance and the maintenance of stable cellular physiology, emphasizing their importance in i A. baumannii /i infections.
Publisher: Wiley
Date: 09-12-2014
Publisher: Wiley
Date: 29-03-2010
DOI: 10.1111/J.1462-2920.2009.02134.X
Abstract: The GacS/GacA signal transduction system is a central regulator in Pseudomonas spp., including the biological control strain P. fluorescens Pf-5, in which GacS/GacA controls the production of secondary metabolites and exoenzymes that suppress plant pathogens. A whole genome oligonucleotide microarray was developed for Pf-5 and used to assess the global transcriptomic consequences of a gacA mutation in P. fluorescens Pf-5. In cultures at the transition from exponential to stationary growth phase, GacA significantly influenced transcript levels of 635 genes, representing more than 10% of the 6147 annotated genes in the Pf-5 genome. Transcripts of genes involved in the production of hydrogen cyanide, the antibiotic pyoluteorin and the extracellular protease AprA were at a low level in the gacA mutant, whereas those functioning in siderophore production and other aspects of iron homeostasis were significantly higher in the gacA mutant than in wild-type Pf-5. Notable effects of gacA inactivation were also observed in the transcription of genes encoding components of a type VI secretion system and cytochrome c oxidase subunits. Two novel gene clusters expressed under the control of gacA were identified from transcriptome analysis, and we propose global-regulator-based genome mining as an approach to decipher the secondary metabolome of Pseudomonas spp.
Publisher: Public Library of Science (PLoS)
Date: 13-05-2009
Publisher: American Society for Microbiology
Date: 29-06-2021
Abstract: Antimicrobial resistance is an emerging global health crisis. Consequently, we have a critical need to prolong our current arsenal of antibiotics, in addition to the development of novel treatment options.
Publisher: Springer Science and Business Media LLC
Date: 09-10-2023
Publisher: American Society for Microbiology
Date: 15-12-2007
DOI: 10.1128/JB.01492-07
Abstract: An acidic residue in transmembrane segment (TMS) 10 is important for recognition of bivalent cationic substrates by the QacA multidrug transporter. Remarkably, an acidic residue in TMS 12 compensated for the absence of such a residue in TMS 10, suggesting that TMS 12 is a component of the bivalent cation-binding region.
Publisher: Springer Science and Business Media LLC
Date: 23-02-2011
Abstract: Iron acquisition systems are important virulence factors in pathogenic bacteria. To identify these systems in Acinetobacter baumannii , the transcriptomic response of the completely sequenced strain ATCC 17978 under iron limiting conditions was investigated using a genomic microarray that contained probes for all annotated open reading frames. Under low iron conditions, transcription levels were more than 2-fold up-regulated for 463 genes, including 95 genes that were up-regulated more than 4-fold. Of particular significance, three siderophore biosynthesis gene clusters, including one novel cluster, were highly up-regulated. Binding sites for the ferric uptake regulator were identified in the promoter regions of many up-regulated genes, suggesting a prominent role for this regulator in the Acinetobacter iron acquisition response. Down-regulation under iron limitation was less dramatic as the transcription of only 202 genes varied more than 2-fold. Various genes involved in motility featured prominently amongst the genes down-regulated when iron was less readily available. Motility assays confirmed that these transcriptional changes are manifested at the phenotypic level. The siderophore biosynthesis gene clusters were further investigated by means of comparative genomic analysis of 10 sequenced Acinetobacter isolates. These analyses revealed important roles for mobile genetic elements in shaping the siderophore meditated iron acquisition mechanisms between different Acinetobacter strains. A. baumannii grown under iron limited conditions resulted in major transcriptional changes of not only many iron acquisition related genes, but also genes involved in other processes such as motility. Overall, this study showed that A. baumannii is well adaptable to growth in an environment which has limiting iron availability.
Publisher: Oxford University Press (OUP)
Date: 10-2006
DOI: 10.1111/J.1574-6968.2006.00411.X
Abstract: The QacA multidrug transporter is encoded on Staphylococcus aureus multidrug resistance plasmids and confers broad-range antimicrobial resistance to more than 30 monovalent and bivalent lipophilic, cationic compounds from at least 12 different chemical classes. QacA contains 10 proline residues predicted to be within transmembrane regions, several of which are conserved in related export proteins. Proline residues are classically known as helix-breakers and are highly represented within the transmembrane helices of membrane transport proteins, where they can mediate the formation of structures essential for protein stability and transport function. The importance of these 10 intramembranous proline residues for QacA-mediated transport function was determined by examining the functional effect of substituting these residues with glycine, alanine or serine. Several proline-substituted QacA mutants failed to confer high-level resistance to selected QacA substrates. However, no single proline mutation, including those at conserved positions, significantly disrupted QacA protein expression or QacA-mediated resistance to all representative substrates, suggesting that these residues are not essential for the formation of structures requisite to the QacA substrate transport mechanism.
Publisher: Public Library of Science (PLoS)
Date: 18-06-2012
Publisher: Springer Science and Business Media LLC
Date: 25-11-2014
Publisher: Public Library of Science (PLoS)
Date: 03-03-2011
Publisher: Frontiers Media SA
Date: 24-08-2021
Publisher: American Society for Microbiology
Date: 26-02-2019
Abstract: A shift in the Western diet since the industrial revolution has resulted in a dramatic increase in the consumption of omega-6 fatty acids, with a concurrent decrease in the consumption of omega-3 fatty acids. This decrease in omega-3 fatty acid consumption has been associated with significant disease burden, including increased susceptibility to infectious diseases. Here we provide evidence that DHA, an omega-3 fatty acid, has superior antimicrobial effects upon the highly drug-resistant pathogen Acinetobacter baumannii , thereby providing insights into one of the potential health benefits of omega-3 fatty acids. The identification and characterization of two novel bacterial membrane protective mechanisms against host fatty acids provide important insights into A. baumannii adaptation during disease. Furthermore, we describe a novel role for the major multidrug efflux system AdeIJK in A. baumannii membrane maintenance and lipid transport. This core function, beyond drug efflux, increases the appeal of AdeIJK as a therapeutic target.
Publisher: Elsevier BV
Date: 2023
Publisher: American Society for Microbiology
Date: 02-11-2016
Abstract: Multidrug efflux pumps provide clinically significant levels of drug resistance in a number of Gram-negative hospital-acquired pathogens. These pathogens frequently carry dozens of genes encoding putative multidrug efflux pumps. However, it can be difficult to determine how many of these pumps actually mediate antimicrobial efflux, and it can be even more challenging to identify the regulatory proteins that control expression of these pumps. In this study, we developed an innovative high-throughput screening method, combining transposon insertion sequencing and cell sorting methods (TraDISort), to identify the genes encoding major multidrug efflux pumps, regulators, and other factors that may affect the permeation of antimicrobials, using the nosocomial pathogen Acinetobacter baumannii . A dense library of more than 100,000 unique transposon insertion mutants was treated with ethidium bromide, a common substrate of multidrug efflux pumps that is differentially fluorescent inside and outside the bacterial cytoplasm. Populations of cells displaying aberrant accumulations of ethidium were physically enriched using fluorescence-activated cell sorting, and the genomic locations of transposon insertions within these strains were determined using transposon-directed insertion sequencing. The relative abundance of mutants in the input pool compared to the selected mutant pools indicated that the AdeABC, AdeIJK, and AmvA efflux pumps are the major ethidium efflux systems in A. baumannii . Furthermore, the method identified a new transcriptional regulator that controls expression of amvA . In addition to the identification of efflux pumps and their regulators, TraDISort identified genes that are likely to control cell ision, cell morphology, or aggregation in A. baumannii . IMPORTANCE Transposon-directed insertion sequencing (TraDIS) and related technologies have emerged as powerful methods to identify genes required for bacterial survival or competitive fitness under various selective conditions. We applied fluorescence-activated cell sorting (FACS) to physically enrich for phenotypes of interest within a mutant population prior to TraDIS. To our knowledge, this is the first time that a physical selection method has been applied in parallel with TraDIS rather than a fitness-induced selection. The results demonstrate the feasibility of this combined approach to generate significant results and highlight the major multidrug efflux pumps encoded in an important pathogen. This FACS-based approach, TraDISort, could have a range of future applications, including the characterization of efflux pump inhibitors, the identification of regulatory factors controlling gene or protein expression using fluorescent reporters, and the identification of genes involved in cell replication, morphology, and aggregation.
Publisher: Springer Science and Business Media LLC
Date: 09-02-2023
DOI: 10.1038/S41467-023-36252-2
Abstract: Acinetobacter baumannii and Klebsiella pneumoniae are opportunistic pathogens frequently co-isolated from polymicrobial infections. The infections where these pathogens co-exist can be more severe and recalcitrant to therapy than infections caused by either species alone, however there is a lack of knowledge on their potential synergistic interactions. In this study we characterise the genomes of A. baumannii and K. pneumoniae strains co-isolated from a single human lung infection. We examine various aspects of their interactions through transcriptomic, phenomic and phenotypic assays that form a basis for understanding their effects on antimicrobial resistance and virulence during co-infection. Using co-culturing and analyses of secreted metabolites, we discover the ability of K. pneumoniae to cross-feed A. baumannii by-products of sugar fermentation. Minimum inhibitory concentration testing of mono- and co-cultures reveals the ability for A. baumannii to cross-protect K. pneumoniae against the cephalosporin, cefotaxime. Our study demonstrates distinct syntrophic interactions occur between A. baumannii and K. pneumoniae , helping to elucidate the basis for their co-existence in polymicrobial infections.
Publisher: Cold Spring Harbor Laboratory
Date: 16-07-2020
DOI: 10.1101/2020.07.16.205906
Abstract: Gene essentiality studies have been performed on numerous bacterial pathogens, but essential gene sets have been determined for only a few plant-associated bacteria. Pseudomonas protegens Pf-5 is a plant-commensal, biocontrol bacteria that can control disease-causing pathogens on a wide range of crops. Work on Pf-5 has mostly focused on secondary metabolism and biocontrol genes, but genome-wide approaches such as high-throughput transposon mutagenesis have not yet been used in this species. Here we generated a dense P. protegens Pf-5 transposon mutant library and used transposon-directed insertion site sequencing (TraDIS) to identify 446 genes essential for growth on rich media. Genes required for fundamental cellular machinery were enriched in the essential gene set, while genes related to nutrient biosynthesis, stress responses and transport were under-represented. Comparison of the essential gene sets of Pf-5 and P. aeruginosa PA14, an opportunistic human pathogen, provides insight into the biological processes important for their different lifestyles. Key differences include cytochrome c biogenesis, formation of periplasmic disulfide bonds, lipid biosynthesis, ribonuclease activity, lipopolysaccharides and cell surface structures. Comparison of the Pf-5 in silico predicted and in vitro determined essential gene sets highlighted the essential cellular functions that are over- and underestimated by each method. Expanding essentiality studies into bacteria with a range of lifestyles can improve our understanding of the biological processes important for survival and growth in different environmental niches. Essential genes are those crucial for survival or normal growth rates in an organism. Essential gene sets have been identified in numerous bacterial pathogens, but only a few plant-associated bacteria. Employing genome-wide approaches, such as transposon insertion sequencing, allows for the concurrent analysis of all genes of a bacterial species and rapid determination of essential gene sets. We have used transposon insertion sequencing to systematically analyze thousands of Pseudomonas protegens Pf-5 genes and gain insights into gene functions and interactions that are not readily available using traditional methods. Comparing Pf-5 essential genes with those of P. aeruginosa PA14, an opportunistic human pathogen, provides insight into differences in gene essentiality which may be linked to their different lifestyles.
Publisher: Proceedings of the National Academy of Sciences
Date: 15-08-2019
Abstract: Multidrug efflux pumps are highly promiscuous determinants of antimicrobial resistance in bacterial pathogens. Since efflux pumps evolved long before the widespread use of antimicrobials, drug transport is likely to be a side reaction in many pumps, fortuitously beneficial to bacteria in hospitals. The AceI efflux protein from Acinetobacter baumannii is the prototype for the proteobacterial antimicrobial compound efflux (PACE) family. AceI was only known to transport the synthetic biocide chlorhexidine, which was incongruous with its ancient origin. Here we demonstrate that short-chain diamines are the physiological substrates of AceI and other PACE members, and that transport is energized by an electrochemical gradient of protons. These observations are important, because diamines play vital roles in bacterial physiology and virulence and have significant commercial uses.
Publisher: Springer Science and Business Media LLC
Date: 22-09-2021
DOI: 10.1038/S42003-021-02629-6
Abstract: Antimicrobial resistance genes, including multidrug efflux pumps, evolved long before the ubiquitous use of antimicrobials in medicine and infection control. Multidrug efflux pumps often transport metabolites, signals and host-derived molecules in addition to antibiotics or biocides. Understanding their ancestral physiological roles could inform the development of strategies to subvert their activity. In this study, we investigated the response of Acinetobacter baumannii to polyamines, a widespread, abundant class of amino acid-derived metabolites, which led us to identify long-chain polyamines as natural substrates of the disinfectant efflux pump AmvA. Loss of amvA dramatically reduced tolerance to long-chain polyamines, and these molecules induce expression of amvA through binding to its cognate regulator AmvR. A second clinically-important efflux pump, AdeABC, also contributed to polyamine tolerance. Our results suggest that the disinfectant resistance capability that allows A. baumannii to survive in hospitals may have evolutionary origins in the transport of polyamine metabolites.
Publisher: Springer Science and Business Media LLC
Date: 06-2003
DOI: 10.1007/S00251-003-0565-X
Abstract: We have used the polymerase chain reaction (PCR) in an attempt to clone and sequence the exons and hitherto unavailable contiguous flanks of all members of the small V(H) 9 germline gene family from inbred mouse strains and sublines that have had a common ancestry within the last century, and to analyze the molecular evolution of these sequences. Fifteen genuine germline genes were isolated (designated V(H) 9.1 through V(H) 9.15) from strains and sublines of DBA, BALB, 129 and C57BL inbred mice. Of the 15 genuine isolates, nine are novel: seven sequences from DBA strains and sublines ( V(H) 9.3 to V(H) 9.9) and two sequences from C57BL strains ( V(H) 9.13 and V(H) 9.14). We have identified sequencing errors and PCR recombinant artefacts in previously published sequences. We detected no sequence ergence of in idual genes shared by the strains and sublines studied. However, we isolated two genes from DBA strains and sublines, V(H) 9.1 and V(H) 9.3, that differ only by five nucleotides encoding three amino acid changes that are concentrated within a 33 nucleotide (11 codon) region. Of these 11 codons, eight encode a putative antigen binding site. There were no differences in the remaining 733 nucleotides sequenced (including both 5' and 3' flanking regions). Potential explanations for the generation of V(H) 9.1 and V(H) 9.3 are discussed.
Publisher: S. Karger AG
Date: 2023
DOI: 10.1159/000529038
Abstract: sc l /sc -cysteine biosynthesis from inorganic sulfur represents a major mechanism by which reduced sulfur is incorporated into organic compounds. Cysteine biosynthesis and regulation is well characterized in i Escherichia coli /i . However, the regulation of sulfur metabolism in i Acinetobacter baumannii /i is only partly understood, with the LysR-type regulator, GigC known to control some aspects of sulfur reduction. In this study, we have used transcriptomics and bioinformatic analyses to characterize a novel LysR-type transcriptional regulator encoded by ABUW_1016 ( i cbl /i ), in a highly multidrug resistant and virulent isolate of i A. baumannii. /i We have shown that Cbl is involved in controlling expression of the genes required for uptake and reduction of various sulfur sources in i A. baumannii /i . Collectively, we have identified the global regulon of Cbl and proposed a model of cysteine biosynthesis and its regulation by Cbl and GigC in i A. baumannii /i .
Publisher: American Society for Microbiology
Date: 08-03-2021
DOI: 10.1128/JB.00432-20
Abstract: Essential genes are those crucial for survival or normal growth rates in an organism. Essential gene sets have been identified in numerous bacterial pathogens but only a few plant-associated bacteria.
Publisher: American Society for Microbiology
Date: 27-02-2015
Abstract: Multidrug efflux systems are a major cause of resistance to antimicrobials in bacteria, including those pathogenic to humans, animals, and plants. These proteins are ubiquitous in these pathogens, and five families of bacterial multidrug efflux systems have been identified to date. By using transcriptomic and biochemical analyses, we recently identified the novel AceI ( Acinetobacter chlorhexidine efflux) protein from Acinetobacter baumannii that conferred resistance to the biocide chlorhexidine, via an active efflux mechanism. Proteins homologous to AceI are encoded in the genomes of many other bacterial species and are particularly prominent within proteobacterial lineages. In this study, we expressed 23 homologs of AceI and examined their resistance and/or transport profiles. MIC analyses demonstrated that, like AceI, many of the homologs conferred resistance to chlorhexidine. Many of the AceI homologs conferred resistance to additional biocides, including benzalkonium, dequalinium, proflavine, and acriflavine. We conducted fluorimetric transport assays using the AceI homolog from Vibrio parahaemolyticus and confirmed that resistance to both proflavine and acriflavine was mediated by an active efflux mechanism. These results show that this group of AceI homologs represent a new family of bacterial multidrug efflux pumps, which we have designated the proteobacterial antimicrobial compound efflux (PACE) family of transport proteins. IMPORTANCE Bacterial multidrug efflux pumps are an important class of resistance determinants that can be found in every bacterial genome sequenced to date. These transport proteins have important protective functions for the bacterial cell but are a significant problem in the clinical setting, since a single efflux system can mediate resistance to many structurally and mechanistically erse antibiotics and biocides. In this study, we demonstrate that proteins related to the Acinetobacter baumannii AceI transporter are a new class of multidrug efflux systems which are very common in Proteobacteria : the proteobacterial antimicrobial compound efflux (PACE) family. This is the first new family of multidrug efflux pumps to be described in 15 years.
Publisher: Informa UK Limited
Date: 31-03-2017
Publisher: Springer Science and Business Media LLC
Date: 17-10-0003
Publisher: American Society for Microbiology
Date: 05-2013
DOI: 10.1128/AEM.03101-12
Abstract: Tannins are a erse group of plant-produced, polyphenolic compounds with metal-chelating and antimicrobial properties that are prevalent in many soils. Using transcriptomics, we determined that tannic acid, a form of hydrolysable tannin, broadly affects the expression of genes involved in iron and zinc homeostases, sulfur metabolism, biofilm formation, motility, and secondary metabolite biosynthesis in the soil- and rhizosphere-inhabiting bacterium Pseudomonas protegens Pf-5.
Publisher: Oxford University Press (OUP)
Date: 28-11-2016
DOI: 10.1093/NAR/GKW1068
Publisher: Frontiers Media SA
Date: 22-04-2015
Publisher: Proceedings of the National Academy of Sciences
Date: 25-11-2013
Abstract: Drug resistance is an increasing problem in clinical settings with some bacterial pathogens now resistant to virtually all available drugs. Chlorhexidine is a commonly used antiseptic and disinfectant in hospital environments, and there is increasing resistance to chlorhexidine seen in some pathogenic bacteria, such as Acinetobacter baumannii . This paper examines the global gene expression of A. baumannii in response to chlorhexidine exposure and identifies a gene that we demonstrate to mediate chlorhexidine resistance. Biochemical investigation reveals that this gene encodes a previously uncharacterized type of drug efflux pump that actively transports chlorhexidine out of the cell.
Publisher: Elsevier BV
Date: 05-2015
DOI: 10.1016/J.RESMIC.2014.10.003
Abstract: Clostridium perfringens can be isolated from a range of environments, including soil, marine and fresh water sediments, and the gastrointestinal tracts of animals and humans. Some C. perfringens strains have attractive industrial applications, e.g., in the degradation of waste products or the production of useful chemicals. However, C. perfringens has been most studied as the causative agent of a range of enteric and soft tissue infections of varying severities in humans and animals. Host preference and disease type in C. perfringens are intimately linked to the production of key extracellular toxins and on this basis toxigenic C. perfringens strains have been classified into five toxinotypes (A-E). To date, twelve genome sequences have been generated for a erse collection of C. perfringens isolates, including strains associated with human and animal infections, a human commensal strain, and a strain with potential industrial utility. Most of the sequenced strains are classified as toxinotype A. However, genome sequences of representative strains from each of the other four toxinotypes have also been determined. Analysis of this collection of sequences has highlighted a lack of features differentiating toxinotype A strains from the other isolates, indicating that the primary defining characteristic of toxinotype A strains is their lack of key plasmid-encoded extracellular toxin genes associated with toxinotype B to E strains. The representative B-E strains sequenced to date each harbour many unique genes. Additional genome sequences are needed to determine if these genes are characteristic of their respective toxinotypes.
Publisher: Microbiology Society
Date: 30-05-2023
DOI: 10.1099/MIC.0.001341
Abstract: Genes encoding a novel multidrug efflux pump, AadT, from the Drug:H + antiporter 2 family, were discovered in Acinetobacter multidrug resistance plasmids. Here, we profiled the antimicrobial resistance potential, and examined the distribution of these genes. aadT homologs were found in many Acinetobacter and other Gram-negative species and were typically adjacent to novel variants of adeAB(C ), which encodes a major tripartite efflux pump in Acinetobacter . The AadT pump decreased bacterial susceptibility to at least eight erse antimicrobials, including antibiotics (erythromycin and tetracycline), biocides (chlorhexidine), and dyes (ethidium bromide and DAPI) and was able to mediate ethidium transport. These results show that AadT is a multidrug efflux pump in the Acinetobacter resistance arsenal and may cooperate with variants of AdeAB(C).
Publisher: American Society for Microbiology
Date: 20-10-2022
DOI: 10.1128/MRA.00317-22
Abstract: Here, we report the 7.7-Mbp genome sequence of Rhodococcus sp. strain 9, which was isolated from Australian groundwater contaminated with phenols and trichloroethylene. This strain has previously been shown to efficiently degrade p -nitrophenol and high-molecular-weight polycyclic aromatic hydrocarbons (PAHs).
Publisher: Cold Spring Harbor Laboratory
Date: 12-01-2023
DOI: 10.1101/2023.01.12.523705
Abstract: Swimming motility is a key bacterial trait, important to success in many niches, including assisting in colonization of host surfaces. Biocontrol bacteria, such as Pseudomonas protegens Pf-5 are increasingly being used as an agricultural tool to control crop diseases, where motility is a factor in successful colonization of the plant rhizosphere. Swimming motility has been studied in a range of bacteria and typically involves a suite of flagella and chemotaxis genes, however the specific gene set employed for both regulation and biogenesis can differ substantially between organisms. Here we used transposon directed insertion site sequencing (TraDIS), a genome-wide approach, to identify 249 genes involved in P. protegens Pf-5 swimming motility. As expected, flagella and chemotaxis genes comprised a large proportion of these genes. However we also identified a suite of additional genes important for swimming, including genes related to peptidoglycan turnover, O-antigen biosynthesis, cell ision, signal transduction, c-di-GMP turnover and phosphate transport, along with 27 conserved hypothetical proteins. Experimental gene knockout mutants and TraDIS data together suggest that defects in the Pst phosphate transporter lead to enhanced swimming motility. Overall, this study expands our knowledge of pseudomonad motility and highlights the utility of a TraDIS-based approach for systematically analyzing the functions of thousands of genes. This work sets a foundation for understanding how swimming motility may be related to the inconsistency in biocontrol bacteria effectiveness and reliability in the field. Biocontrol bacteria, such as Pseudomonas protegens Pf-5 are increasingly being used as an agricultural tool to control crop diseases, and motility is a key factor in their successful colonization of plant surfaces. Here we use a high-throughput approach to identify the suite of genes important for swimming motility in P. protegens Pf-5. These included flagella and chemotaxis genes, as well as a variety of cell surface, cell ision and signalling genes. We also show that defects in the Pst phosphate transporter lead to enhanced swimming motility, a hitherto unreported link between phosphate transport and swimming motility. Understanding the genetic basis of swimming motility enhances our knowledge of key processes in biocontrol bacteria that are needed to ensure their competitive success. This will contribute to developing strategies to increase the utility of biocontrol bacteria in agricultural settings to prevent crop losses.
Publisher: American Society for Microbiology
Date: 28-10-2021
DOI: 10.1128/AEM.01718-21
Abstract: Cadmium toxicity is a widespread problem, yet the interaction of this heavy metal with biological systems is poorly understood. Some microbes have evolved traits to proactively counteract cadmium toxicity, including Acinetobacter baumannii , which is notorious for persisting in harsh environments.
Publisher: Microbiology Society
Date: 18-01-2023
Abstract: Membrane transporters are a large group of proteins that span cell membranes and contribute to critical cell processes, including delivery of essential nutrients, ejection of waste products, and assisting the cell in sensing environmental conditions. Obtaining an accurate and specific annotation of the transporter proteins encoded by a micro-organism can provide details of its likely nutritional preferences and environmental niche(s), and identify novel transporters that could be utilized in small molecule production in industrial biotechnology. The Transporter Automated Annotation Pipeline (TransAAP) ( ransportDB2/TransAAP_login.html ) is a fully automated web service for the prediction and annotation of membrane transport proteins in an organism from its genome sequence, by using comparisons with both curated databases such as the TCDB (Transporter Classification Database) and TDB, as well as selected Pfams and TIGRFAMs of transporter families and other methodologies. TransAAP was used to annotate transporter genes in the prokaryotic genomes in the National Center for Biotechnology Information (NCBI) RefSeq these are presented in the transporter database TransportDB ( www.membranetransport.org ) website, which has a suite of data visualization and analysis tools. Creation and maintenance of a bioinformatic database specific for transporters in all genomic datasets is essential for microbiology research groups and the general research/biotechnology community to obtain a detailed picture of membrane transporter systems in various environments, as well as comprehensive information on specific membrane transport proteins.
Publisher: American Society for Microbiology
Date: 02-05-2013
Abstract: The neurotoxins produced by Clostridium botulinum strains are among the world's most potent toxins and are the causative agents of paralytic botulism. Here, we present the draft genome sequence of the group III C. botulinum strain Eklund-C, including a pseudolysogen-like bacteriophage that harbors the type C neurotoxin operon.
Publisher: American Chemical Society (ACS)
Date: 12-2006
DOI: 10.1021/BI0614380
Abstract: The staphylococcal TetA(K) tetracycline exporter is classified within the major facilitator superfamily of transport proteins and contains 14 alpha-helical transmembrane segments (TMS). Using cysteine-scanning mutagenesis, 27 amino acid residues across and flanking putative TMS 10 of the TetA(K) transporter were in idually replaced with cysteine. The level of solvent accessibility to each of the targeted amino acid positions was determined as a measure of fluorescein maleimide reactivity and demonstrated that TMS 10 of TetA(K) has a cytoplasmic boundary at G313 and is likely to extend from at least V298 on the periplasmic side. TMS 10 was found to be hiphilic containing at least partially solvent accessible amino acid residues along the length of one helical face, suggesting that this helix may line a solvent-exposed channel. Functional analyses of these cysteine mutants demonstrated a significant role for a number of amino acid residues, including a predominance of glycine residues which were further analyzed by alanine substitution. These residues are postulated to allow interhelical interactions between TMS 10 and distal parts of TetA(K) that are likely to be required for the tetracycline transport mechanism in TetA(K) and may be a general feature required by bacterial tetracycline transporters for activity.
Publisher: Frontiers Media SA
Date: 14-04-2021
DOI: 10.3389/FMICB.2021.651282
Abstract: Plants live in association with microorganisms that positively influence plant development, vigor, and fitness in response to pathogens and abiotic stressors. The bulk of the plant microbiome is concentrated belowground at the plant root-soil interface. Plant roots secrete carbon-rich rhizodeposits containing primary and secondary low molecular weight metabolites, lysates, and mucilages. These exudates provide nutrients for soil microorganisms and modulate their affinity to host plants, but molecular details of this process are largely unresolved. We addressed this gap by focusing on the molecular dialog between eight well-characterized beneficial strains of the Pseudomonas fluorescens group and Brachypodium distachyon , a model for economically important food, feed, forage, and biomass crops of the grass family. We collected and analyzed root exudates of B. distachyon and demonstrated the presence of multiple carbohydrates, amino acids, organic acids, and phenolic compounds. The subsequent screening of bacteria by Biolog Phenotype MicroArrays revealed that many of these metabolites provide carbon and energy for the Pseudomonas strains. RNA-seq profiling of bacterial cultures amended with root exudates revealed changes in the expression of genes encoding numerous catabolic and anabolic enzymes, transporters, transcriptional regulators, stress response, and conserved hypothetical proteins. Almost half of the differentially expressed genes mapped to the variable part of the strains’ pangenome, reflecting the importance of the variable gene content in the adaptation of P. fluorescens to the rhizosphere lifestyle. Our results collectively reveal the ersity of cellular pathways and physiological responses underlying the establishment of mutualistic interactions between these beneficial rhizobacteria and their plant hosts.
Publisher: Public Library of Science (PLoS)
Date: 21-07-2016
Publisher: Cold Spring Harbor Laboratory
Date: 13-02-2020
DOI: 10.1101/2020.02.12.946830
Abstract: Manuka honey has broad-spectrum antimicrobial activity and unlike traditional antibiotics, resistance to its killing effects has not been reported. However, its mechanism of action remains unclear. Here we investigated the mechanism of action of manuka honey and its key antibacterial components using a transcriptomic approach in a model organism, Pseudomonas aeruginosa. We show that no single component of honey can account for its total antimicrobial action, and that honey affects the expression of genes in the SOS response, oxidative damage and quorum sensing. Manuka honey uniquely affects genes involved in the explosive cell lysis process and in maintaining the electron transport chain, causing protons to leak across membranes and collapsing the proton motive force and induces membrane depolarisation and permeabilisation in P. aeruginosa . These data indicate that the activity of manuka honey comes from multiple mechanisms of action that do not engender bacterial resistance. The threat of antimicrobial resistance to human health has prompted interest in complex, natural products with antimicrobial activity. Honey has been an effective topical wound treatment throughout history, predominantly due to its broad-spectrum antimicrobial activity. Unlike traditional antibiotics, honey-resistant bacteria have not been reported, however, honey remains underutilised in the clinic in part due to a lack of understanding of its mechanism of action. Here we demonstrate that honey affects multiple processes in bacteria, and this is not explained by its major antibacterial components. Honey also uniquely affects bacterial membranes and this can be exploited for combination therapy with antibiotics that are otherwise ineffective on their own. We argue that honey should be included as part of the current array of wound treatments due to its effective antibacterial activity that does not promote resistance in bacteria.
Publisher: Public Library of Science (PLoS)
Date: 19-03-2013
Publisher: Wiley
Date: 26-06-2023
Abstract: Antimicrobial resistance (AMR) is predicted to cause a worldwide annual toll of 10 million deaths by 2050. This looming public health threat has been linked to antibiotic overuse and pollution, which places selective pressures on AMR maintenance and transfer in and between microbial populations. We examined the distribution, ersity and potential mobility of AMR genes in cyanobacteria. While cyanobacteria are not pathogenic, we hypothesised that they could be a major environmental reservoir for AMR genes. Genes encoding AMR to seven antimicrobial drug classes were found in 10% of cyanobacterial genomes. AMR genes were found in 13% of freshwater, 19% of terrestrial, 34% of symbiotic, 2% of thermal spring, and 3% of marine genomes. AMR genes were found in five cyanobacterial orders with 23% of Nostocales and 8% of Oscillatoriales strains containing AMR genes. The most frequently observed alleles were ansamycin resistance genes, which were present in 7% of strains. AMR genes responsible for resistance to broad‐spectrum β‐lactams, chlor henicols, tetracyclines, macrolides, and aminoglycosides were associated with mobile genetic elements or plasmid replicons or both. These results suggest that cyanobacteria are an extensive reservoir, and potential vector, for AMR genes in erse terrestrial and aquatic habitats.
Publisher: Cold Spring Harbor Laboratory
Date: 03-10-2020
DOI: 10.1101/2020.10.02.324624
Abstract: Multidrug efflux pumps are important drivers of antibiotic resistance in Acinetobacter baumannii and other pathogens, however their ‘natural’ roles beyond transport of clinical antimicrobials are poorly described. Polyamines are an ancient class of molecules with broad roles in all three kingdoms of life, and are the likely natural substrate of at least one efflux pump family. We have defined the transcriptome of A. baumannii following treatment with high levels of the polyamines putrescine, cadaverine, spermidine and spermine. These molecules influenced expression of multiple gene classes in A. baumannii including those associated with virulence, and the four polyamines induced distinct but overlapping transcriptional responses. Polyamine shock also induced expression of the MFS-family efflux pump gene amvA and its repressor gene amvR . Loss of amvA dramatically reduced tolerance to the long-chain triaamine spermidine, but caused only modest changes in resistance to known AmvA substrates such as acriflavine. We confirmed reduced accumulation of spermidine in amvA -deficient A. baumannii , and showed that its expression is induced by long-chain polyamines through its cognate regulator AmvR. Our findings suggest that the conserved A. baumannii efflux pump AmvA has evolved to export spermidine from the cell, but that its substrate recognition promiscuity also allows activity against clinically-important biocides and antibiotics. AMR genes, including multidrug efflux pumps, evolved long before the ubiquitous use of antimicrobials in medicine and infection control. Multidrug efflux pumps often transport metabolites, signals and host-derived molecules in addition to antibiotics or biocides. Understanding the ancestral physiological roles of multidrug efflux pumps could help to inform the development of strategies to subvert their activity. In this study, we investigated the response of Acinetobacter baumannii to polyamines, a widespread, abundant class of amino acid-derived metabolites, which led us to identify long-chain polyamines as natural substrates of the disinfectant efflux pump AmvA. A second clinically-important efflux pump, AdeABC, also contributed to polyamine tolerance. Our results suggest that the disinfectant resistance capability that allows A. baumannii to survive in hospitals may have evolutionary origins in the transport of polyamine metabolites.
Publisher: American Society for Microbiology
Date: 22-12-2021
DOI: 10.1128/SPECTRUM.01455-21
Abstract: The global distribution of multidrug resistance in A. baumannii has necessitated seeking not only alternative therapeutic approaches but also the means to limit the development of resistance in clinical settings. Highly abundant host bioactive compounds, such as polyunsaturated fatty acids, are readily acquired by A. baumannii during infection and have been illustrated to impact the bacterium’s membrane composition and antibiotic resistance.
Publisher: American Society for Microbiology
Date: 26-06-2014
Abstract: Clostridium strains from six phylogenetic groups, C. botulinum groups I to IV, C. baratii , and C. butyricum , display the capacity to produce botulinum neurotoxin. Here, we present the genome sequence of a C. butyricum isolate, the neurotoxigenic strain 5521, which encodes the type E botulinum neurotoxin.
Publisher: Frontiers Media SA
Date: 27-10-2020
Publisher: Frontiers Media SA
Date: 23-09-2015
Publisher: Public Library of Science (PLoS)
Date: 05-07-2012
Publisher: Oxford University Press (OUP)
Date: 31-01-2016
DOI: 10.1093/JAC/DKV460
Abstract: Drug efflux pumps are one of the key machineries in bacterial drug resistance. Although quite a few of these transport systems have been functionally characterized in various organisms, due to large-scale genome sequencing efforts and improved prediction pipelines there are increasing numbers of putative drug efflux genes annotated. For phenotype identification of the proteins encoded by these genes, we developed a novel high-throughput phenotype screening strategy and demonstrated its utility in identifying phenotypes for putative efflux systems encoded by the human pathogen Acinetobacter baumannii. Seventeen putative drug efflux systems from A. baumannii were heterologously expressed in Escherichia coli. For rapid and economical phenotype screening we employed a combination of multiplexed Biolog Phenotype Microarrays and quantitative PCR. Using this method we screened these putative drug efflux pumps against 240 antimicrobial conditions, equating to 4080 simultaneous phenotypic tests. Of the 17 putative drug efflux systems, phenotypes were confirmed for two pumps and novel drug resistance phenotypes were identified for three new A. baumannii drug transporters, which exemplified the power of this method as a high-throughput screening technique. One of the phenotypically characterized putative drug efflux systems was classified within the ATP-binding cassette superfamily of transport proteins and represents the first drug resistance protein characterized from this superfamily in A. baumannii. The remaining two proteins were members of the major facilitator superfamily of efflux pumps. This method has broad potential for high-throughput phenotype characterization of putative drug efflux systems in a range of organisms.
Publisher: Frontiers Media SA
Date: 2013
Publisher: Public Library of Science (PLoS)
Date: 04-05-2017
Publisher: Public Library of Science (PLoS)
Date: 08-2014
Publisher: Oxford University Press (OUP)
Date: 09-08-2011
DOI: 10.1111/J.1574-6968.2011.02362.X
Abstract: Acinetobacter baumannii continues to be a major health problem especially in hospital settings. Herein, features that may play a role in persistence and disease potential were investigated in a collection of clinical A. baumannii strains from Australia. Twitching motility was found to be a common trait in A. baumannii international clone I strains and in abundant biofilm formers, whereas swarming motility was only observed in isolates not classified within the international clone lineages. Bioinformatic analysis of the type IV fimbriae revealed a correlation between PilA sequence homology and motility. A high level of variability in adherence to both abiotic surfaces and epithelial cells was found. We report for the first time the motility characteristics of a large number of A. baumannii isolates and present a direct comparison of A. baumannii binding to nasopharyngeal and lung epithelial cells.
Publisher: Elsevier BV
Date: 04-2009
DOI: 10.1016/J.PEP.2008.11.009
Abstract: The plasmid-encoded QacA multidrug transport protein confers high-level resistance to a range of commonly used antimicrobials and is carried by widespread clinical strains of the human pathogen Staphylococcus aureus making it a potential target for future drug therapies. In order to obtain a sufficient yield of QacA protein for structural and biophysical studies, an optimized strategy for QacA overexpression was developed. QacA expression, directed from several vector systems in Escherichia coli, was tested under various growth and induction conditions and a synthetic qacA gene, codon-optimized for expression in E. coli was developed. Despite the extreme hydrophobicity and potential toxicity of the QacA secondary transport protein, a strategy based on the pBAD expression system, yielding up to four milligrams of approximately 95% pure QacA protein per litre of liquid culture, was devised. Purified QacA protein was examined using circular dichroism spectroscopy and displayed a secondary structure akin to that predicted from in silico analyses. Additionally, detergent solubilized QacA protein was shown to bind its fluorescent substrate rhodamine 6G with micro-molar affinity using a fluorescence polarization-based binding assay, similar to other multidrug transport proteins. To check the applicability of the expression urification system described for QacA to other staphylococcal secondary transporters, the gene encoding the TetA(K) tetracycline efflux protein, which was previously recalcitrant to overexpression, was incorporated into the pBAD-based system and shown to be readily produced at easily detectable levels. Therefore, this expression system could be of general use for the production of secondary transport proteins in E. coli.
Publisher: Elsevier BV
Date: 12-2021
DOI: 10.1016/J.MIB.2021.08.005
Abstract: Since the late 1990's the genome sequences for thousands of species of bacteria have been released into public databases. The release of each new genome sequence typically revealed the presence of tens to hundreds of uncharacterised genes encoding putative membrane proteins and more recently, microbial metagenomics has revealed countless more of these uncharacterised genes. Given the importance of small molecule efflux in bacteria, it is likely that a significant proportion of these genes encode for novel efflux proteins, but the elucidation of these functions is challenging. We used transcriptomics to predict that the function of a gene encoding a hypothetical membrane protein is in efflux-mediated antimicrobial resistance. We subsequently confirmed this function and the likely native substrates of the pump by using detailed biochemical and biophysical analyses. Functional studies of homologs of the protein from other bacterial species determined that the protein is a prototype for a family of multidrug efflux pumps - the Proteobacterial Antimicrobial Compound Efflux (PACE) family. The general functional genomics approach used here, and its expansion to functional metagenomics, will very likely reveal the identities of more efflux pumps and other transport proteins of scientific, clinical and commercial interest in the future.
Publisher: American Chemical Society (ACS)
Date: 24-03-2021
Publisher: American Society for Microbiology
Date: 09-2013
DOI: 10.1128/AEM.01354-13
Abstract: Conjugative plasmids are known to facilitate the acquisition and dispersal of genes contributing to the fitness of Pseudomonas spp. Here, we report the characterization of pA506, the 57-kb conjugative plasmid of Pseudomonas fluorescens A506, a plant epiphyte used in the United States for the biological control of fire blight disease of pear and apple. Twenty-nine of the 67 open reading frames (ORFs) of pA506 have putative functions in conjugation, including a type IV secretion system related to that of MOB P6 family plasmids and a gene cluster for type IV pili. We demonstrate that pA506 is self-transmissible via conjugation between A506 and strains of Pseudomonas spp. or the Enterobacteriaceae . The origin of vegetative replication ( oriV ) of pA506 is typical of those in pPT23A family plasmids, which are present in many pathovars of Pseudomonas syringae , but pA506 lacks repA , a defining locus for pPT23A plasmids, and has a novel partitioning region. We selected a plasmid-cured derivative of A506 and compared it to the wild type to identify plasmid-encoded phenotypes. pA506 conferred UV resistance, presumably due to the plasmid-borne rulAB genes, but did not influence epiphytic fitness of A506 on pear or apple blossoms in the field. pA506 does not appear to confer resistance to antibiotics or other toxic elements. Based on the conjugative nature of pA506 and the large number of its genes that are shared with plasmids from erse groups of environmental bacteria, the plasmid is likely to serve as a vehicle for genetic exchange between A506 and its coinhabitants on plant surfaces.
Publisher: S. Karger AG
Date: 2007
DOI: 10.1159/000099640
Abstract: Drug resistance mediated by integral membrane transporters is an important mode of cellular resistance to cytotoxic agents across all classes of living organisms. Gram-positive bacteria, such as staphylococcal species, are not encapsulated by a selective outer membrane permeability barrier. Therefore, these organisms often employ integral membrane drug transport systems to maintain cellular concentrations of antimicrobials at subtoxic levels. Staphylococcal species, including the opportunistic human pathogen i Staphylococcus aureus /i , encode a multitude of drug exporters, encompassing transporters from each of the five currently recognized families of bacterial drug resistance transporters. A number of these transporters are chromosomally encoded and allow the host cell to realize clinically significant levels of drug resistance after minor mutations to regulatory regions. Others are plasmid-encoded and can be easily passed between staphylococcal strains and species, or acquired from other Gram-positive genera. In combination, staphylococcal drug transporters potentiate resistance to a vast array of antimicrobial compounds, including macrolide, quinolone, tetracycline and streptogramin antibiotics, as well as a broad range of biocides, such as quaternary ammonium compounds, biguanidines and diamidines. An understanding of the genetic and molecular properties of drug transporters will lead to effective treatments of staphylococcal infections. Here we provide a detailed review of the active drug transporters of the staphylococci.
Publisher: Oxford University Press (OUP)
Date: 21-02-2018
DOI: 10.1093/JAC/DKY034
Publisher: Public Library of Science (PLoS)
Date: 04-11-2013
Publisher: American Society for Microbiology
Date: 07-2011
DOI: 10.1128/JB.05089-11
Abstract: Vibrio rotiferianus is a marine pathogen capable of causing disease in various aquatic organisms. We announce the genome sequence of V. rotiferianus DAT722, which has a large chromosomal integron containing 116 gene cassettes and is a model organism for studying the role of this system in vibrio evolution.
Publisher: Springer New York
Date: 25-11-2017
DOI: 10.1007/978-1-4939-7454-2_12
Abstract: The core genomes of most bacterial species include a large number of genes encoding putative efflux pumps. The functional roles of most of these pumps are unknown, however, they are often under tight regulatory control and expressed in response to their substrates. Therefore, one way to identify pumps that function in antimicrobial resistance is to examine the transcriptional responses of efflux pump genes to antimicrobial shock. By conducting complete transcriptomic experiments following antimicrobial shock treatments, it may be possible to identify novel drug efflux pumps encoded in bacterial genomes. In this chapter we describe a complete workflow for conducting transcriptomic analyses by RNA sequencing, to determine transcriptional changes in bacteria responding to antimicrobials.
Publisher: Research Square Platform LLC
Date: 08-02-2022
DOI: 10.21203/RS.3.RS-1184776/V1
Abstract: Acinetobacter baumannii and Klebsiella pneumoniae are opportunistic pathogens frequently co-isolated from polymicrobial infections. The infections where these pathogens co-exist can be more severe and recalcitrant to therapy than infections caused by either species alone, however there is a lack of knowledge on their potential synergistic interactions. In this study we characterised the genomes of A. baumannii and K. pneumoniae strains co-isolated from a single human lung infection. We examined various aspects of their interactions through transcriptomic, phenomic and phenotypic assays that form a basis for understanding their effects on antimicrobial resistance and virulence during co-infection. Using co-culturing and analyses of secreted metabolites, we discovered the ability of K. pneumoniae to cross-feed A. baumannii by-products of sugar fermentation. Minimum inhibitory concentration testing of mono- and co-cultures revealed the ability for A. baumannii to cross-protect K. pneumoniae against the cephalosporin, cefotaxime. Our study demonstrates distinct syntrophic interactions occur between A. baumannii and K. pneumoniae , helping to elucidate the basis for their co-existence in polymicrobial infections.
Publisher: American Society for Microbiology
Date: 18-05-2021
DOI: 10.1128/AAC.01400-20
Abstract: Fluoroquinolones are one of the most prescribed broad-spectrum antibiotics. However, their effectiveness is being compromised by high rates of resistance in clinically important organisms, including Acinetobacter baumannii .
Publisher: S. Karger AG
Date: 2011
DOI: 10.1159/000329836
Abstract: Heterologous expression of membrane proteins in i Escherichia coli /i often requires optimization to overcome problems with toxicity of the recombinant protein to the host cell. A number of Gateway-based destination vectors were constructed to investigate expression of membrane proteins using a high-throughput approach. These vectors were tested using putative drug transporter proteins from the multidrug and toxic compound extrusion (MATE) family and the resistance-nodulation-cell ision superfamily encoded by the human pathogen i Acinetobacter baumannii /i . Active transport of antibiotics and antiseptics mediated by efflux proteins contributes to the high level of multidrug resistance observed in i A. baumannii /i . Substrates for 4 of the 5 putative efflux proteins investigated were identified using the expression vectors designed in this study. Additionally, a Gateway-based suicide vector was designed for construction of specific i A. baumannii /i insertion disruption mutants. This knockout cloning strategy was tested and shown to be successful in inactivating AbeM4, a putative MATE family protein. Therefore, we have shown that the Gateway-based vectors constructed in this study are versatile tools that can be used for manipulation and characterization of membrane proteins.
Publisher: Humana Press
Date: 2014
DOI: 10.1007/978-1-62703-712-9_10
Abstract: Biolog Phenotype MicroArrays for microorganisms provide a high-throughput method for the global analysis of microbial growth phenotypes. Using a colorimetric reaction that is indicative of respiration, these microplate assays measure the response of an in idual strain or microbial community to a large and erse range of nutrients and chemicals. Phenotype MicroArrays have been used to study gene function and to improve genome annotation in single microorganisms and for physiological profiling of bacterial communities. The microplate system can be used to obtain a comprehensive overview of metabolic capability, or it can be tailored, through the use of subsets of plates, to address specific research needs.
Publisher: Elsevier BV
Date: 09-2018
Publisher: American Chemical Society (ACS)
Date: 28-10-2019
Publisher: Cold Spring Harbor Laboratory
Date: 21-10-2020
DOI: 10.1101/2020.10.20.348086
Abstract: Acinetobacter species are ubiquitous Gram-negative bacteria that can be found in water, soil and as commensals of the human skin. The successful inhabitation of Acinetobacter species in erse environments is primarily attributable to the expression of an arsenal of stress resistance determinants, which includes an extensive repertoire of metal ion efflux systems. Although metal ion homeostasis in the hospital pathogen Acinetobacter baumannii is known to contribute to pathogenesis, insights into its metal ion transporters for environmental persistence are lacking. Here, we studied the impact of cadmium stress on A. baumannii . Our functional genomics and independent mutant analyses revealed a primary role for CzcE, a member of the cation diffusion facilitator (CDF) superfamily, in resisting cadmium stress. Further, we show that the CzcCBA heavy metal efflux system also contributes to cadmium efflux. Analysis of the A. baumannii metallome under cadmium stress showed zinc depletion and copper enrichment, which are likely to influence cellular fitness. Overall, this work expands our understanding of the role of membrane transporters in A. baumannii metal ion homeostasis. Cadmium toxicity is a widespread problem, yet the interaction of this heavy metal with biological systems is poorly understood. Some microbes have evolved traits to proactively counteract cadmium toxicity, which includes Acinetobacter baumannii . Here we show that A. baumannii utilises a dedicated cadmium efflux protein in concert with a system that is primarily attuned to zinc efflux, to efficiently overcome cadmium stress. The molecular characterization of A. baumannii under cadmium stress revealed how active cadmium efflux plays a key role in preventing the dysregulation of bacterial metal ion homeostasis, which appeared to be the primary means by which cadmium exerts toxicity upon the bacterium.
Publisher: Springer Science and Business Media LLC
Date: 15-09-2015
Publisher: American Society for Microbiology
Date: 2011
DOI: 10.1128/JB.00895-10
Abstract: Pseudomonas fluorescens Q8r1-96 represents a group of rhizosphere strains responsible for the suppressiveness of agricultural soils to take-all disease of wheat. It produces the antibiotic 2,4-diacetylphloroglucinol and aggressively colonizes the roots of cereal crops. In this study, we analyzed the genome of Q8r1-96 and identified a type III protein secretion system (T3SS) gene cluster that has overall organization similar to that of the T3SS gene cluster of the plant pathogen Pseudomonas syringae . We also screened a collection of 30 closely related P. fluorescens strains and detected the T3SS genes in all but one of them. The Q8r1-96 genome contained ropAA and ropM type III effector genes, which are orthologs of the P. syringae effector genes hopAA1 - 1 and hopM1 , as well as a novel type III effector gene designated ropB . These type III effector genes encoded proteins that were secreted in culture and injected into plant cells by both P. syringae and Q8r1-96 T3SSs. The Q8r1-96 T3SS was expressed in the rhizosphere, but mutants lacking a functional T3SS were not altered in their rhizosphere competence. The Q8r1-96 type III effectors RopAA, RopB, and RopM were capable of suppressing the hypersensitive response and production of reactive oxygen species, two plant immune responses.
Publisher: Public Library of Science (PLoS)
Date: 11-05-2018
Publisher: MDPI AG
Date: 29-01-2019
DOI: 10.3390/IJMS20030575
Abstract: Acinetobacter baumannii has emerged as one of the leading causative agents of nosocomial infections. Due to its high level of intrinsic and adapted antibiotic resistance, treatment failure rates are high, which allows this opportunistic pathogen to thrive during infection in immune-compromised patients. A. baumannii can cause infections within a broad range of host niches, with pneumonia and bacteraemia being associated with the greatest levels of morbidity and mortality. Although its resistance to antibiotics is widely studied, our understanding of the mechanisms required for dealing with environmental stresses related to virulence and hospital persistence, such as copper toxicity, is limited. Here, we performed an in silico analysis of the A. baumannii copper resistome, examining its regulation under copper stress. Using comparative analyses of bacterial P-type ATPases, we propose that A. baumannii encodes a member of a novel subgroup of P1B-1 ATPases. Analyses of three putative inner membrane copper efflux systems identified the P1B-1 ATPase CopA as the primary mediator of cytoplasmic copper resistance in A. baumannii. Using a murine model of A. baumannii pneumonia, we reveal that CopA contributes to the virulence of A. baumannii. Collectively, this study advances our understanding of how A. baumannii deals with environmental copper toxicity, and it provides novel insights into how A. baumannii combats adversities encountered as part of the host immune defence.
Publisher: Springer Science and Business Media LLC
Date: 16-09-2008
Abstract: The staphylococcal QacA multidrug efflux protein confers resistance to an exceptional number of structurally unrelated antimicrobial compounds. Aromatic amino acid residues have been shown to be highly important for the transport function of several multidrug transporters and are intimately involved in multidrug binding. This study investigated the structural and functional importance of the seven tyrosine residues in QacA by examining the phenotypic effect of incorporating conservative (aromatic) and non-conservative (non-aromatic) substitutions for these residues. Determination of the resistance profiles and analysis of drug transport assays revealed that non-conservative substitutions for most tyrosine residues influenced the QacA drug recognition spectrum. However, an aromatic residue at three tyrosine positions, 63, 410 and 429, was of importance for QacA-mediated transport and resistance to the majority of substrates tested. A tyrosine or phenylalanine residue at amino acid positions corresponding to 63 of QacA in related drug efflux proteins is found to be highly conserved. Therefore, an aromatic side chain at this position is likely to partake in a function common to these drug transporters, such as proton translocation or essential intramolecular contacts, whereas aromatic residues at the non-conserved 410 and 429 positions are expected to mediate a QacA-specific function, possibly forming or stabilising part of the QacA drug binding region.
Publisher: Public Library of Science (PLoS)
Date: 07-02-2013
Publisher: Wiley
Date: 19-08-2012
DOI: 10.1111/J.1462-2920.2012.02849.X
Abstract: Zinc is an important nutrient but can be lacking in some soil environments, influencing the physiology of soil-dwelling bacteria. Hence, we studied the global effect of zinc limitation on the transcriptome of the rhizosphere biocontrol strain Pseudomonas protegens Pf-5 (formerly Pseudomonas fluorescens). We observed that the expression of the putative zinc uptake regulator (Zur) gene was upregulated, and we mapped putative Zur binding sites in the Pf-5 genome using bioinformatic approaches. In line with the need to regulate intracellular zinc concentrations, an array of potential zinc transporter genes was found to be zinc-regulated. To adapt to low-zinc conditions, a gene cluster encoding non-zinc-requiring paralogues of zinc-dependent proteins was also significantly upregulated. Similarly, transcription of genes encoding non-zinc-requiring paralogues of ribosomal proteins L31 and L36 was increased by zinc limitation. A strong transcriptional downregulation of the putative copper chaperone gene (copZ) was also observed, suggesting interplay between zinc and copper homeostasis. Importantly, zinc also affected biocontrol attributes in Pf-5, most notably reducing the expression of the gene cluster responsible for biosynthesis of the antibiotic 2,4-diacetylphloroglucinol (DAPG) under zinc limitation. This study clearly defines changes to the molecular physiology of Pf-5 that enable it to survive under zinc limitation.
Publisher: American Society for Microbiology
Date: 04-2008
DOI: 10.1128/JB.01864-07
Abstract: Tryptophan residues can possess a multitude of functions within a multidrug transport protein, e.g., mediating interactions with substrates or distal parts of the protein, or fulfilling a structural requirement, such as guiding the depth of membrane insertion. In this study, the nine tryptophan residues of the staphylococcal QacA multidrug efflux protein were in idually mutated to alanine and phenylalanine, and the functional consequences of these changes were determined. Phenylalanine substitutions for each tryptophan residue were functionally tolerated. However, alanine modifications revealed an important functional role for three tryptophan residues, W58, W149, and W173, each of which is well conserved among QacA-related transport proteins in the major facilitator superfamily. The most functionally compromising mutation, an alanine substitution for W58, likely to be located at the extracellular interface of transmembrane segment 2, abolished all detectable QacA-mediated resistance and transport function. Second-site suppressor analyses identified several mutations that rescued the function of the W58A QacA mutant. Remarkably, all of these suppressor mutations were shown to be located in cytoplasmic loops between transmembrane helices 2 and 3 or 12 and 13, demonstrating novel functional associations between amino acid positions on opposite sides of the membrane and in distal N- and C-terminal regions of the QacA protein.
No related organisations have been discovered for Karl Hassan.
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