# | Rank | Similarity | Title + Abs. | Year | PMID |
|---|---|---|---|---|---|
| 0 | 1 | 2 | 3 | 4 | 5 |
| 8239 | 0 | 0.9962 | Surviving bacterial sibling rivalry: inducible and reversible phenotypic switching in Paenibacillus dendritiformis. Natural habitats vary in available nutrients and room for bacteria to grow, but successful colonization can lead to overcrowding and stress. Here we show that competing sibling colonies of Paenibacillus dendritiformis bacteria survive overcrowding by switching between two distinct vegetative phenotypes, motile rods and immotile cocci. Growing colonies of the rod-shaped bacteria produce a toxic protein, Slf, which kills cells of encroaching sibling colonies. However, sublethal concentrations of Slf induce some of the rods to switch to Slf-resistant cocci, which have distinct metabolic and resistance profiles, including resistance to cell wall antibiotics. Unlike dormant spores of P. dendritiformis, the cocci replicate. If cocci encounter conditions that favor rods, they secrete a signaling molecule that induces a switch to rods. Thus, in contrast to persister cells, P. dendritiformis bacteria adapt to changing environmental conditions by inducible and reversible phenotypic switching. IMPORTANCE: In favorable environments, species may face space and nutrient limits due to overcrowding. Bacteria provide an excellent model for analyzing principles underlying overcrowding and regulation of density in nature, since their population dynamics can be easily and accurately assessed under controlled conditions. We describe a newly discovered mechanism for survival of a bacterial population during overcrowding. When competing with sibling colonies, Paenibacillus dendritiformis produces a lethal protein (Slf) that kills cells at the interface of encroaching colonies. Slf also induces a small proportion of the cells to switch from motile, rod-shaped cells to nonmotile, Slf-resistant, vegetative cocci. When crowding is reduced and nutrients are no longer limiting, the bacteria produce a signal that induces cocci to switch back to motile rods, allowing the population to spread. Genes encoding components of this phenotypic switching pathway are widespread among bacterial species, suggesting that this survival mechanism is not unique to P. dendritiformis. | 2011 | 21628502 |
| 241 | 1 | 0.9961 | A color-based competition assay for studying bacterial stress responses in Micrococcus luteus. Competition assays measure differences between populations of bacteria after stress adaptation, populations of different bacteria and mutations in antibiotic resistance genes. We have developed a competition-based assay to evaluate if genes upregulated under starvation are important for bacterial survival. Stress responses are critical for survival in non-pathogenic and pathogenic bacteria alike including Mycobacterium tuberculosis, Enterococcus fecaelis, Escherichia coli and Staphylococcus aureus. Unfortunately, most stress-survival proteins are poorly understood because suitable model bacteria and techniques are limited. To address this problem, we have engineered Micrococcus luteus NCTC 2665 (M. luteus) for competition assays by inactivating the sarcinaxanthin biosynthesis gene crtE (ΔcrtE), changing M. luteus colonies from yellow to white. This change allows easy identification in mixed cultures. The crtE knockout is relatively neutral for growth in complex and minimal acetate media and shows a measured fitness of one in competition with yellow wild-type bacteria. The ΔcrtE M. luteus competition assay identified a competition defect in a M. luteus strain when a specific universal stress protein was inactivated, suggesting a negative survival phenotype for this protein. We anticipate this competition assay can identify defects in other gene knockouts and mutational studies in M. luteus and will enhance our understanding of bacterial survival mechanisms. | 2019 | 30865770 |
| 9073 | 2 | 0.9958 | EpitoCore: Mining Conserved Epitope Vaccine Candidates in the Core Proteome of Multiple Bacteria Strains. In reverse vaccinology approaches, complete proteomes of bacteria are submitted to multiple computational prediction steps in order to filter proteins that are possible vaccine candidates. Most available tools perform such analysis only in a single strain, or a very limited number of strains. But the vast amount of genomic data had shown that most bacteria contain pangenomes, i.e., their genomic information contains core, conserved genes, and random accessory genes specific to each strain. Therefore, in reverse vaccinology methods it is of the utmost importance to define core proteins and core epitopes. EpitoCore is a decision-tree pipeline developed to fulfill that need. It provides surfaceome prediction of proteins from related strains, defines core proteins within those, calculate their immunogenicity, predicts epitopes for a given set of MHC alleles defined by the user, and then reports if epitopes are located extracellularly and if they are conserved among the core homologs. Pipeline performance is illustrated by mining peptide vaccine candidates in Mycobacterium avium hominissuis strains. From a total proteome of ~4,800 proteins per strain, EpitoCore predicted 103 highly immunogenic core homologs located at cell surface, many of those related to virulence and drug resistance. Conserved epitopes identified among these homologs allows the users to define sets of peptides with potential to immunize the largest coverage of tested HLA alleles using peptide-based vaccines. Therefore, EpitoCore is able to provide automated identification of conserved epitopes in bacterial pangenomic datasets. | 2020 | 32431712 |
| 9410 | 3 | 0.9958 | Comparative genomics and an insect model rapidly identify novel virulence genes of Burkholderia mallei. Burkholderia pseudomallei and its host-adapted deletion clone Burkholderia mallei cause the potentially fatal human diseases melioidosis and glanders, respectively. The antibiotic resistance profile and ability to infect via aerosol of these organisms and the absence of protective vaccines have led to their classification as major biothreats and select agents. Although documented infections by these bacteria date back over 100 years, relatively little is known about their virulence and pathogenicity mechanisms. We used in silico genomic subtraction to generate their virulome, a set of 650 putative virulence-related genes shared by B. pseudomallei and B. mallei but not present in five closely related nonpathogenic Burkholderia species. Although most of these genes are clustered in putative operons, the number of targets for mutant construction and verification of reduced virulence in animal models is formidable. Therefore, Galleria mellonella (wax moth) larvae were evaluated as a surrogate host; we found that B. pseudomallei and B. mallei, but not other phylogenetically related bacteria, were highly pathogenic for this insect. More importantly, four previously characterized B. mallei mutants with reduced virulence in hamsters or mice had similarly reduced virulence in G. mellonella larvae. Site-specific inactivation of selected genes in the computationally derived virulome identified three new potential virulence genes, each of which was required for rapid and efficient killing of larvae. Thus, this approach may provide a means to quickly identify high-probability virulence genes in B. pseudomallei, B. mallei, and other pathogens. | 2008 | 18223084 |
| 9231 | 4 | 0.9957 | CRISPR: new horizons in phage resistance and strain identification. Bacteria have been widely used as starter cultures in the food industry, notably for the fermentation of milk into dairy products such as cheese and yogurt. Lactic acid bacteria used in food manufacturing, such as lactobacilli, lactococci, streptococci, Leuconostoc, pediococci, and bifidobacteria, are selectively formulated based on functional characteristics that provide idiosyncratic flavor and texture attributes, as well as their ability to withstand processing and manufacturing conditions. Unfortunately, given frequent viral exposure in industrial environments, starter culture selection and development rely on defense systems that provide resistance against bacteriophage predation, including restriction-modification, abortive infection, and recently discovered CRISPRs (clustered regularly interspaced short palindromic repeats). CRISPRs, together with CRISPR-associated genes (cas), form the CRISPR/Cas immune system, which provides adaptive immunity against phages and invasive genetic elements. The immunization process is based on the incorporation of short DNA sequences from virulent phages into the CRISPR locus. Subsequently, CRISPR transcripts are processed into small interfering RNAs that guide a multifunctional protein complex to recognize and cleave matching foreign DNA. Hypervariable CRISPR loci provide insights into the phage and host population dynamics, and new avenues for enhanced phage resistance and genetic typing and tagging of industrial strains. | 2012 | 22224556 |
| 8396 | 5 | 0.9957 | Screening and Functional Analyses of Novel Cecropins from Insect Transcriptome. Antibiotic resistance is a significant and growing threat to global public health. However, antimicrobial peptides (AMPs) have shown promise as they exhibit a broad spectrum of antibacterial activities with low potential for resistance development. Insects, which inhabit a wide range of environments and are incredibly diverse, remain largely unexplored as a source of novel AMPs. To address this, we conducted a screening of the representative transcriptomes from the 1000 Insect Transcriptome Evolution (1KITE) dataset, focusing on the homologous reference genes of Cecropins, the first identified AMPs in insects known for its high efficiency. Our analysis identified 108 Cecropin genes from 105 insect transcriptomes, covering all major hexapod lineages. We validated the gene sequences and synthesized mature peptides for three identified Cecropin genes. Through minimal inhibition concentration and agar diffusion assays, we confirmed that these peptides exhibited antimicrobial activities against Gram-negative bacteria. Similar to the known Cecropin, the three Cecropins adopted an alpha-helical conformation in membrane-like environments, efficiently disrupting bacterial membranes through permeabilization. Importantly, none of the three Cecropins demonstrated cytotoxicity in erythrocyte hemolysis tests, suggesting their safety in real-world applications. Overall, this newly developed methodology provides a high-throughput bioinformatic pipeline for the discovery of AMP, taking advantage of the expanding genomic resources available for diverse organisms. | 2023 | 37887806 |
| 3765 | 6 | 0.9956 | An allelic exchange system for compliant genetic manipulation of the select agents Burkholderia pseudomallei and Burkholderia mallei. Burkholderia pseudomallei and B. mallei are Gram-negative bacterial pathogens that cause melioidosis in humans and glanders in horses, respectively. Both bacteria are classified as category B select agents in the United States. Due to strict select-agent regulations, the number of antibiotic selection markers approved for use in these bacteria is greatly limited. Approved markers for B. pseudomallei include genes encoding resistance to kanamycin (Km), gentamicin (Gm), and zeocin (Zeo); however, wild type B. pseudomallei is intrinsically resistant to these antibiotics. Selection markers for B. mallei are limited to Km and Zeo resistance genes. Additionally, there are few well developed counter-selection markers for use in Burkholderia. The use of SacB as a counter-selection method has been of limited success due to the presence of endogenous sacBC genes in the genomes of B. pseudomallei and B. mallei. These impediments have greatly hampered the genetic manipulation of B. pseudomallei and B. mallei and currently few reliable tools for the genetic manipulation of Burkholderia exist. To expand the repertoire of genetic tools for use in Burkholderia, we developed the suicide plasmid pMo130, which allows for the compliant genetic manipulation of the select agents B. pseudomallei and B. mallei using allelic exchange. pMo130 harbors an aphA gene which allows for Km selection, the reporter gene xylE, which allows for reliable visual detection of Burkholderia transformants, and carries a modified sacB gene that allows for the resolution of co-integrants. We employed this system to generate multiple unmarked and in-frame mutants in B. pseudomallei, and one mutant in B. mallei. This vector significantly expands the number of available tools that are select-agent compliant for the genetic manipulation of B. pseudomallei and B. mallei. | 2009 | 19010402 |
| 5079 | 7 | 0.9956 | Development of a Rapid, Culture-Free, Universal Microbial Identification System Using Internal Transcribed Spacer Targeting Primers. The indiscriminate administration of broad-spectrum antibiotics is a primary contributor to the increasing prevalence of antibiotic resistance. Unfortunately, culture, the gold standard for bacterial identification is a time intensive process. Due to this extended diagnostic period, broad-spectrum antibiotics are generally prescribed to prevent poor outcomes. To overcome the deficits of culture-based methods, we have developed a rapid universal bacterial identification system. The platform uses a unique universal polymerase chain reaction primer set that targets the internal transcribed spacer regions between conserved bacterial genes, creating a distinguishable amplicon signature for every bacterial species. Bioinformatic simulation demonstrates that nearly every bacteria in a set of 45 commonly isolated pathogenic species can be uniquely identified using this approach. We experimentally confirmed these predictions on a representative set of pathogenic bacterial species. We further showed that the system can determine the corresponding concentration of each pathogen. Finally, we validated performance in clinical urinary tract infection samples. | 2025 | 39503259 |
| 4353 | 8 | 0.9956 | Bioinformatics-driven discovery of skin microbiota bacteriocins as potential antibiotics and probiotics. The human skin microbiota, comprising a diverse range of microorganisms, including bacteria, viruses, and fungi, plays an important role in maintaining skin health and protecting against pathogenic invasions. Among these microorganisms, certain bacteria produce bacteriocins, which are ribosomal peptides with potent antimicrobial properties. This study presents a novel computational approach to identify and predict bacteriocins from microbial genomes comprising sebaceous region of the skin, aiming to explore their therapeutic potential. Through genome analysis using advanced bioinformatics tools, we identified potential genes, operons, open reading frames (ORFs), and promoter regions linked to bacteriocin production. The BAGEL4 platform was employed to detect structural bacteriocin genes, while modelling bacterial growth and bacteriocin expression under various environmental conditions was conducted using MATLAB's SimBiology application. The results revealed the optimal conditions for bacteriocin production and highlighted promising candidates for further experimental validation. These findings underscore the significance of skin microbiota as a source of novel bacteriocins, offering potential alternatives to traditional antibiotics amidst rising antimicrobial resistance. | 2025 | 40702306 |
| 9088 | 9 | 0.9956 | Cocrystallizing and Codelivering Complementary Drugs to Multidrugresistant Tuberculosis Bacteria in Perfecting Multidrug Therapy. Bacteria cells exhibit multidrug resistance in one of two ways: by raising the genetic expression of multidrug efflux pumps or by accumulating several drug-resistant components in many genes. Multidrug-resistive tuberculosis bacteria are treated by multidrug therapy, where a few certain antibacterial drugs are administered together to kill a bacterium jointly. A major drawback of conventional multidrug therapy is that the administration never ensures the reaching of different drug molecules to a particular bacterium cell at the same time, which promotes growing drug resistivity step-wise. As a result, it enhances the treatment time. With additional tabletability and plasticity, the formation of a cocrystal of multidrug can ensure administrating the multidrug chemically together to a target bacterium cell. With properly maintaining the basic philosophy of multidrug therapy here, the synergistic effects of drug molecules can ensure killing the bacteria, even before getting the option to raise the drug resistance against them. This can minimize the treatment span, expenditure and drug resistance. A potential threat of epidemic from tuberculosis has appeared after the Covid-19 outbreak. An unwanted loop of finding molecules with the potential to kill tuberculosis, getting their corresponding drug approvals, and abandoning the drug after facing drug resistance can be suppressed here. This perspective aims to develop the universal drug regimen by postulating the principles of drug molecule selection, cocrystallization, and subsequent harmonisation within a short period to address multidrug-resistant bacteria. | 2023 | 37150990 |
| 9079 | 10 | 0.9956 | Review, Evaluation, and Directions for Gene-Targeted Assembly for Ecological Analyses of Metagenomes. Shotgun metagenomics has greatly advanced our understanding of microbial communities over the last decade. Metagenomic analyses often include assembly and genome binning, computationally daunting tasks especially for big data from complex environments such as soil and sediments. In many studies, however, only a subset of genes and pathways involved in specific functions are of interest; thus, it is not necessary to attempt global assembly. In addition, methods that target genes can be computationally more efficient and produce more accurate assembly by leveraging rich databases, especially for those genes that are of broad interest such as those involved in biogeochemical cycles, biodegradation, and antibiotic resistance or used as phylogenetic markers. Here, we review six gene-targeted assemblers with unique algorithms for extracting and/or assembling targeted genes: Xander, MegaGTA, SAT-Assembler, HMM-GRASPx, GenSeed-HMM, and MEGAN. We tested these tools using two datasets with known genomes, a synthetic community of artificial reads derived from the genomes of 17 bacteria, shotgun sequence data from a mock community with 48 bacteria and 16 archaea genomes, and a large soil shotgun metagenomic dataset. We compared assemblies of a universal single copy gene (rplB) and two N cycle genes (nifH and nirK). We measured their computational efficiency, sensitivity, specificity, and chimera rate and found Xander and MegaGTA, which both use a probabilistic graph structure to model the genes, have the best overall performance with all three datasets, although MEGAN, a reference matching assembler, had better sensitivity with synthetic and mock community members chosen from its reference collection. Also, Xander and MegaGTA are the only tools that include post-assembly scripts tuned for common molecular ecology and diversity analyses. Additionally, we provide a mathematical model for estimating the probability of assembling targeted genes in a metagenome for estimating required sequencing depth. | 2019 | 31749830 |
| 8378 | 11 | 0.9956 | Genome mining reveals unlocked bioactive potential of marine Gram-negative bacteria. BACKGROUND: Antibiotic resistance in bacteria spreads quickly, overtaking the pace at which new compounds are discovered and this emphasizes the immediate need to discover new compounds for control of infectious diseases. Terrestrial bacteria have for decades been investigated as a source of bioactive compounds leading to successful applications in pharmaceutical and biotech industries. Marine bacteria have so far not been exploited to the same extent; however, they are believed to harbor a multitude of novel bioactive chemistry. To explore this potential, genomes of 21 marine Alpha- and Gammaproteobacteria collected during the Galathea 3 expedition were sequenced and mined for natural product encoding gene clusters. RESULTS: Independently of genome size, bacteria of all tested genera carried a large number of clusters encoding different potential bioactivities, especially within the Vibrionaceae and Pseudoalteromonadaceae families. A very high potential was identified in pigmented pseudoalteromonads with up to 20 clusters in a single strain, mostly NRPSs and NRPS-PKS hybrids. Furthermore, regulatory elements in bioactivity-related pathways including chitin metabolism, quorum sensing and iron scavenging systems were investigated both in silico and in vitro. Genes with siderophore function were identified in 50% of the strains, however, all but one harboured the ferric-uptake-regulator gene. Genes encoding the syntethase of acylated homoserine lactones were found in Roseobacter-clade bacteria, but not in the Vibrionaceae strains and only in one Pseudoalteromonas strains. The understanding and manipulation of these elements can help in the discovery and production of new compounds never identified under regular laboratory cultivation conditions. High chitinolytic potential was demonstrated and verified for Vibrio and Pseudoalteromonas species that commonly live in close association with eukaryotic organisms in the environment. Chitin regulation by the ChiS histidine-kinase seems to be a general trait of the Vibrionaceae family, however it is absent in the Pseudomonadaceae. Hence, the degree to which chitin influences secondary metabolism in marine bacteria is not known. CONCLUSIONS: Utilizing the rapidly developing sequencing technologies and software tools in combination with phenotypic in vitro assays, we demonstrated the high bioactive potential of marine bacteria in an efficient, straightforward manner - an approach that will facilitate natural product discovery in the future. | 2015 | 25879706 |
| 9078 | 12 | 0.9956 | MetaCherchant: analyzing genomic context of antibiotic resistance genes in gut microbiota. MOTIVATION: Antibiotic resistance is an important global public health problem. Human gut microbiota is an accumulator of resistance genes potentially providing them to pathogens. It is important to develop tools for identifying the mechanisms of how resistance is transmitted between gut microbial species and pathogens. RESULTS: We developed MetaCherchant-an algorithm for extracting the genomic environment of antibiotic resistance genes from metagenomic data in the form of a graph. The algorithm was validated on a number of simulated and published datasets, as well as applied to new 'shotgun' metagenomes of gut microbiota from patients with Helicobacter pylori who underwent antibiotic therapy. Genomic context was reconstructed for several major resistance genes. Taxonomic annotation of the context suggests that within a single metagenome, the resistance genes can be contained in genomes of multiple species. MetaCherchant allows reconstruction of mobile elements with resistance genes within the genomes of bacteria using metagenomic data. Application of MetaCherchant in differential mode produced specific graph structures suggesting the evidence of possible resistance gene transmission within a mobile element that occurred as a result of the antibiotic therapy. MetaCherchant is a promising tool giving researchers an opportunity to get an insight into dynamics of resistance transmission in vivo basing on metagenomic data. AVAILABILITY AND IMPLEMENTATION: Source code and binaries are freely available for download at https://github.com/ctlab/metacherchant. The code is written in Java and is platform-independent. COTANCT: ulyantsev@rain.ifmo.ru. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online. | 2018 | 29092015 |
| 8402 | 13 | 0.9955 | Exploring phage-host interactions in Burkholderia cepacia complex bacterium to reveal host factors and phage resistance genes using CRISPRi functional genomics and transcriptomics. Complex interactions of bacteriophages with their bacterial hosts determine phage host range and infectivity. While phage defense systems and host factors have been identified in model bacteria, they remain challenging to predict in non-model bacteria. In this paper, we integrate functional genomics and transcriptomics to investigate phage-host interactions, revealing active phage resistance and host factor genes in Burkholderia cenocepacia K56-2. Burkholderia cepacia complex species are commonly found in soil and are opportunistic pathogens in immunocompromised patients. We studied infection of B. cenocepacia K56-2 with Bcep176, a temperate phage isolated from Burkholderia multivorans. A genome-wide dCas9 knockdown library targeting B. cenocepacia K56-2 was constructed, and a pooled infection experiment identified 63 novel genes or operons coding for candidate host factors or phage resistance genes. The activities of a subset of candidate host factor and resistance genes were validated via single-gene knockdowns. Transcriptomics of B. cenocepacia K56-2 during Bcep176 infection revealed that expression of genes coding for host factor and resistance candidates identified in this screen was significantly altered during infection by 4 h post-infection. Identifying which bacterial genes are involved in phage infection is important to understand the ecological niches of B. cenocepacia and its phages, and for designing phage therapies.IMPORTANCEBurkholderia cepacia complex bacteria are opportunistic pathogens inherently resistant to antibiotics, and phage therapy is a promising alternative treatment for chronically infected patients. Burkholderia bacteria are also ubiquitous in soil microbiomes. To develop improved phage therapies for pathogenic Burkholderia bacteria, or engineer phages for applications, such as microbiome editing, it's essential to know the bacterial host factors required by the phage to kill bacteria, as well as how the bacteria prevent phage infection. This work identified 65 genes involved in phage-host interactions in Burkholderia cenocepacia K56-2 and tracked their expression during infection. These findings establish a knowledge base to select and engineer phages infecting or transducing Burkholderia bacteria. | 2025 | 41036840 |
| 8399 | 14 | 0.9955 | SYN-View: A Phylogeny-Based Synteny Exploration Tool for the Identification of Gene Clusters Linked to Antibiotic Resistance. The development of new antibacterial drugs has become one of the most important tasks of the century in order to overcome the posing threat of drug resistance in pathogenic bacteria. Many antibiotics originate from natural products produced by various microorganisms. Over the last decades, bioinformatical approaches have facilitated the discovery and characterization of these small compounds using genome mining methodologies. A key part of this process is the identification of the most promising biosynthetic gene clusters (BGCs), which encode novel natural products. In 2017, the Antibiotic Resistant Target Seeker (ARTS) was developed in order to enable an automated target-directed genome mining approach. ARTS identifies possible resistant target genes within antibiotic gene clusters, in order to detect promising BGCs encoding antibiotics with novel modes of action. Although ARTS can predict promising targets based on multiple criteria, it provides little information about the cluster structures of possible resistant genes. Here, we present SYN-view. Based on a phylogenetic approach, SYN-view allows for easy comparison of gene clusters of interest and distinguishing genes with regular housekeeping functions from genes functioning as antibiotic resistant targets. Our aim is to implement our proposed method into the ARTS web-server, further improving the target-directed genome mining strategy of the ARTS pipeline. | 2020 | 33396183 |
| 6306 | 15 | 0.9955 | Retrieval of Enterobacteriaceae drug targets using singular value decomposition. MOTIVATION: The identification of potential drug target proteins in bacteria is important in pharmaceutical research for the development of new antibiotics to combat bacterial agents that cause diseases. RESULTS: A new model that combines the singular value decomposition (SVD) technique with biological filters composed of a set of protein properties associated with bacterial drug targets and similarity to protein-coding essential genes of Escherichia coli (strain K12) has been created to predict potential antibiotic drug targets in the Enterobacteriaceae family. This model identified 99 potential drug target proteins in the studied family, which exhibit eight different functions and are protein-coding essential genes or similar to protein-coding essential genes of E.coli (strain K12), indicating that the disruption of the activities of these proteins is critical for cells. Proteins from bacteria with described drug resistance were found among the retrieved candidates. These candidates have no similarity to the human proteome, therefore exhibiting the advantage of causing no adverse effects or at least no known adverse effects on humans. CONTACT: rita_silverio@hotmail.com. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online. | 2015 | 25480374 |
| 6308 | 16 | 0.9955 | A shotgun antisense approach to the identification of novel essential genes in Pseudomonas aeruginosa. BACKGROUND: Antibiotics in current use target a surprisingly small number of cellular functions: cell wall, DNA, RNA, and protein biosynthesis. Targeting of novel essential pathways is expected to play an important role in the discovery of new antibacterial agents against bacterial pathogens, such as Pseudomonas aeruginosa, that are difficult to control because of their ability to develop resistance, often multiple, to all current classes of clinical antibiotics. RESULTS: We aimed to identify novel essential genes in P. aeruginosa by shotgun antisense screening. This technique was developed in Staphylococcus aureus and, following a period of limited success in Gram-negative bacteria, has recently been used effectively in Escherichia coli. To also target low expressed essential genes, we included some variant steps that were expected to overcome the non-stringent regulation of the promoter carried by the expression vector used for the shotgun antisense libraries. Our antisense screenings identified 33 growth-impairing single-locus genomic inserts that allowed us to generate a list of 28 "essential-for-growth" genes: five were "classical" essential genes involved in DNA replication, transcription, translation, and cell division; seven were already reported as essential in other bacteria; and 16 were "novel" essential genes with no homologs reported to have an essential role in other bacterial species. Interestingly, the essential genes in our panel were suggested to take part in a broader range of cellular functions than those currently targeted by extant antibiotics, namely protein secretion, biosynthesis of cofactors, prosthetic groups and carriers, energy metabolism, central intermediary metabolism, transport of small molecules, translation, post-translational modification, non-ribosomal peptide synthesis, lipopolysaccharide synthesis/modification, and transcription regulation. This study also identified 43 growth-impairing inserts carrying multiple loci targeting 105 genes, of which 25 have homologs reported as essential in other bacteria. Finally, four multigenic growth-impairing inserts belonged to operons that have never been reported to play an essential role. CONCLUSIONS: For the first time in P. aeruginosa, we applied regulated antisense RNA expression and showed the feasibility of this technology for the identification of novel essential genes. | 2014 | 24499134 |
| 9405 | 17 | 0.9955 | Functional Metagenomic Screening for Antimicrobial Resistance in the Oral Microbiome. A large proportion of bacteria, from a multitude of environments, are not yet able to be grown in the laboratory, and therefore microbiological and molecular biological investigations of these bacteria are challenging. A way to circumvent this challenge is to analyze the metagenome, the entire collection of DNA molecules that can be isolated from a particular environment or sample. This collection of DNA molecules can be sequenced and assembled to determine what is present and infer functional potential, or used as a PCR template to detect known target DNA and potentially unknown regions of DNA nearby those targets; however assigning functions to new or conserved hypothetical, functionally cryptic, genes is difficult. Functional metagenomics allows researchers to determine which genes are responsible for selectable phenotypes, such as resistance to antimicrobials and metabolic capabilities, without the prerequisite needs to grow the bacteria containing those genes or to already know which genes are of interest. It is estimated that a third of the resident species of the human oral cavity is not yet cultivable and, together with the ease of sample acquisition, makes this metagenome particularly suited to functional metagenomic studies. Here we describe the methodology related to the collection of saliva samples, extraction of metagenomic DNA, construction of metagenomic libraries, as well as the description of functional assays that have previously led to the identification of new genes conferring antimicrobial resistance. | 2021 | 34410638 |
| 9448 | 18 | 0.9955 | Fresh Ideas Bloom in Gut Healthcare to Cross-Fertilize Lake Management. Harmful bacteria may be the most significant threat to human gut and lake ecosystem health, and they are often managed using similar tools, like poisoning with antibiotics or algicides. Out-of-the-box thinking in human microbiome engineering is leading to novel methods, like engineering bacteria to kill pathogens, "persuade" them not to produce toxins, or "mop up" their toxins. The bacterial agent can be given a competitive edge via an exclusive nutrient, and they can be engineered to commit suicide once their work is done. Viruses can kill pathogens with specific DNA sequences or knock out their antibiotic resistance genes using CRISPR technology. Some of these ideas may work for lakes. We critically review novel methods for managing harmful bacteria in the gut from the perspective of managing toxic cyanobacteria in lakes, and discuss practical aspects such as modifying bacteria using genetic engineering or directed evolution, mass culturing and controlling the agents. A key knowledge gap is in the ecology of strains, like toxigenic vs nontoxigenic Microcystis, including allelopathic and Black Queen interactions. Some of the "gut methods" may have future potential for lakes, but there presently is no substitute for established management approaches, including reducing N and P nutrient inputs, and mitigating climate change. | 2019 | 31647664 |
| 8387 | 19 | 0.9955 | Construction and Analysis of Two Genome-Scale Deletion Libraries for Bacillus subtilis. A systems-level understanding of Gram-positive bacteria is important from both an environmental and health perspective and is most easily obtained when high-quality, validated genomic resources are available. To this end, we constructed two ordered, barcoded, erythromycin-resistance- and kanamycin-resistance-marked single-gene deletion libraries of the Gram-positive model organism, Bacillus subtilis. The libraries comprise 3,968 and 3,970 genes, respectively, and overlap in all but four genes. Using these libraries, we update the set of essential genes known for this organism, provide a comprehensive compendium of B. subtilis auxotrophic genes, and identify genes required for utilizing specific carbon and nitrogen sources, as well as those required for growth at low temperature. We report the identification of enzymes catalyzing several missing steps in amino acid biosynthesis. Finally, we describe a suite of high-throughput phenotyping methodologies and apply them to provide a genome-wide analysis of competence and sporulation. Altogether, we provide versatile resources for studying gene function and pathway and network architecture in Gram-positive bacteria. | 2017 | 28189581 |