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https://www.readbyqxmd.com/read/29286406/high-throughput-parallel-sequencing-to-measure-fitness-of-leptospira-interrogans-transposon-insertion-mutants-during-golden-syrian-hamster-infection
#1
Kristel Lourdault, James Matsunaga, Karen V Evangelista, David A Haake
In this manuscript, we describe a transposon sequencing (Tn-Seq) technique to identify and quantify Leptospira interrogans mutants altered in fitness during infection of Golden Syrian hamsters. Tn-Seq combines random transposon mutagenesis with the power of high-throughput sequencing technology. Animals are challenged with a pool of transposon mutants (input pool), followed by harvesting of blood and tissues a few days later to identify and quantify the number of mutants in each organ (output pools). The output pools are compared to the input pool to evaluate the in vivo fitness of each mutant...
December 18, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29230204/a-comprehensive-overview-of-online-resources-to-identify-and-predict-bacterial-essential-genes
#2
REVIEW
Chong Peng, Yan Lin, Hao Luo, Feng Gao
Genes critical for the survival or reproduction of an organism in certain circumstances are classified as essential genes. Essential genes play a significant role in deciphering the survival mechanism of life. They may be greatly applied to pharmaceutics and synthetic biology. The continuous progress of experimental method for essential gene identification has accelerated the accumulation of gene essentiality data which facilitates the study of essential genes in silico. In this article, we present some available online resources related to gene essentiality, including bioinformatic software tools for transposon sequencing (Tn-seq) analysis, essential gene databases and online services to predict bacterial essential genes...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/29213059/a-comprehensive-assessment-of-the-genetic-determinants-in-salmonella-typhimurium-for-resistance-to-hydrogen-peroxide-using-proteogenomics
#3
Sardar Karash, Rohana Liyanage, Abdullah Qassab, Jackson O Lay, Young Min Kwon
Salmonella is an intracellular pathogen infecting a wide range of hosts and can survive in macrophages. An essential mechanism used by macrophages to eradicate Salmonella is production of reactive oxygen species. Here, we used proteogenomics to determine the candidate genes and proteins that have a role in resistance of S. Typhimurium to H2O2. For Tn-seq, a saturated Tn5 insertion library was grown in vitro under either 2.5 (H2O2L) or 3.5 mM H2O2 (H2O2H). We identified two sets of overlapping genes required for resistance of S...
December 6, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29208119/comparative-analysis-of-the-burkholderia-cenocepacia-k56-2-essential-genome-reveals-cell-envelope-functions-that-are-uniquely-required-for-survival-in-species-of-the-genus-burkholderia
#4
April S Gislason, Keith Turner, Mike Domaratzki, Silvia T Cardona
Burkholderia cenocepacia K56-2 belongs to the Burkholderia cepacia complex, a group of Gram-negative opportunistic pathogens that have large and dynamic genomes. In this work, we identified the essential genome of B. cenocepacia K56-2 using high-density transposon mutagenesis and insertion site sequencing (Tn-seq circle). We constructed a library of one million transposon mutants and identified the transposon insertions at an average of one insertion per 27 bp. The probability of gene essentiality was determined by comparing of the insertion density per gene with the variance of neutral datasets generated by Monte Carlo simulations...
November 2017: Microbial Genomics
https://www.readbyqxmd.com/read/29186514/permissive-zones-for-the-centromere-binding-protein-parb-on-the-caulobacter-crescentus-chromosome
#5
Ngat T Tran, Clare E Stevenson, Nicolle F Som, Anyarat Thanapipatsiri, Adam S B Jalal, Tung B K Le
Proper chromosome segregation is essential in all living organisms. In Caulobacter crescentus, the ParA-ParB-parS system is required for proper chromosome segregation and cell viability. The bacterial centromere-like parS DNA locus is the first to be segregated following chromosome replication. parS is bound by ParB protein, which in turn interacts with ParA to partition the ParB-parS nucleoprotein complex to each daughter cell. Here, we investigated the genome-wide distribution of ParB on the Caulobacter chromosome using a combination of in vivo chromatin immunoprecipitation (ChIP-seq) and in vitro DNA affinity purification with deep sequencing (IDAP-seq)...
November 23, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29184024/identification-of-fitness-determinants-during-energy-limited-growth-arrest-in-pseudomonas-aeruginosa
#6
David W Basta, Megan Bergkessel, Dianne K Newman
Microbial growth arrest can be triggered by diverse factors, one of which is energy limitation due to scarcity of electron donors or acceptors. Genes that govern fitness during energy-limited growth arrest and the extent to which they overlap between different types of energy limitation are poorly defined. In this study, we exploited the fact that Pseudomonas aeruginosa can remain viable over several weeks when limited for organic carbon (pyruvate) as an electron donor or oxygen as an electron acceptor. ATP values were reduced under both types of limitation, yet more severely in the absence of oxygen...
November 28, 2017: MBio
https://www.readbyqxmd.com/read/29184015/molecular-basis-of-bacterial-longevity
#7
Kieran B Pechter, Liang Yin, Yasuhiro Oda, Larry Gallagher, Jianming Yang, Colin Manoil, Caroline S Harwood
It is well known that many bacteria can survive in a growth-arrested state for long periods of time, on the order of months or even years, without forming dormant structures like spores or cysts. How is such longevity possible? What is the molecular basis of such longevity? Here we used the Gram-negative phototrophic alphaproteobacterium Rhodopseudomonas palustris to identify molecular determinants of bacterial longevity. R. palustris maintained viability for over a month after growth arrest due to nutrient depletion when it was provided with light as a source of energy...
November 28, 2017: MBio
https://www.readbyqxmd.com/read/29162049/rna-seq-and-tn-seq-reveal-fitness-determinants-of-vancomycin-resistant-enterococcus-faecium-during-growth-in-human-serum
#8
Xinglin Zhang, Vincent de Maat, Ana M Guzmán Prieto, Tomasz K Prajsnar, Jumamurat R Bayjanov, Mark de Been, Malbert R C Rogers, Marc J M Bonten, Stéphane Mesnage, Rob J L Willems, Willem van Schaik
BACKGROUND: The Gram-positive bacterium Enterococcus faecium is a commensal of the human gastrointestinal tract and a frequent cause of bloodstream infections in hospitalized patients. The mechanisms by which E. faecium can survive and grow in blood during an infection have not yet been characterized. Here, we identify genes that contribute to growth of E. faecium in human serum through transcriptome profiling (RNA-seq) and a high-throughput transposon mutant library sequencing approach (Tn-seq)...
November 21, 2017: BMC Genomics
https://www.readbyqxmd.com/read/29131301/genome-wide-identification-of-tolerance-mechanisms-towards-p-coumaric-acid-in-pseudomonas-putida
#9
Patricia Calero, Sheila I Jensen, Klara Bojanovič, Rebecca Lennen, Anna Koza, Alex T Nielsen
The soil bacterium Pseudomonas putida KT2440 has gained increasing biotechnological interest due to its ability to tolerate different types of stress. Here, the tolerance of P. putida KT2440 towards eleven toxic chemical compounds was investigated. P. putida was found to be significantly more tolerant towards three of the eleven compounds when compared to Escherichia coli. Increased tolerance was for example found towards p-coumaric acid, an interesting precursor for polymerization with a significant industrial relevance...
November 13, 2017: Biotechnology and Bioengineering
https://www.readbyqxmd.com/read/29109185/genome-wide-screens-reveal-new-gene-products-that-influence-genetic-competence-in-streptococcus-mutans
#10
Robert C Shields, Greg O'Brien, Natalie Maricic, Alexandria Kesterson, Megan Grace, Stephen J Hagen, Robert A Burne
A network of genes and at least two peptide signaling molecules tightly control when Streptococcus mutans becomes competent to take up DNA from its environment. Widespread changes in the expression of genes occur when S. mutans is presented with competence signal peptides in vitro, including increased production of the alternative sigma factor, ComX, which activates late competence genes. Still, the way that gene products that are regulated by competence peptides influence DNA uptake and cellular physiology are not well understood...
November 6, 2017: Journal of Bacteriology
https://www.readbyqxmd.com/read/29109175/the-streptococcus-agalactiae-stringent-response-enhances-virulence-and-persistence-in-human-blood
#11
Thomas A Hooven, Andrew J Catomeris, Maryam Bonakdar, Luke J Tallon, Ivette Santana-Cruz, Sandra Ott, Sean C Daugherty, Hervé Tettelin, Adam J Ratner
Streptococcus agalactiae (group B Streptococcus, GBS) causes serious infection in neonates. We previously reported a Tn-seq system for performing genome-wide assessment of gene fitness in GBS. In order to identify molecular mechanisms required for GBS to transition from a mucosal commensal lifestyle to bloodstream invasion, we performed Tn-seq on GBS coincubated with human whole blood. Our analysis identified 16 genes conditionally essential for GBS survival in blood, of which 75% were members of the capsular polysaccharide (cps) operon...
November 6, 2017: Infection and Immunity
https://www.readbyqxmd.com/read/29099305/peripheral-blood-vessels-are-a-niche-for-blood-borne-meningococci
#12
Elena Capel, Jean-Philippe Barnier, Aldert L Zomer, Christine Bole-Feysot, Thomas Nussbaumer, Anne Jamet, Hervé Lécuyer, Daniel Euphrasie, Zoé Virion, Eric Frapy, Philippe Pélissier, Olivier Join-Lambert, Thomas Rattei, Sandrine Bourdoulous, Xavier Nassif, Mathieu Coureuil
Neisseria meningitidis is the causative agent of cerebrospinal meningitis and that of a rapidly progressing fatal septic shock known as purpura fulminans. Meningococcemia is characterized by bacterial adhesion to human endothelial cells of the microvessels. Host specificity has hampered studies on the role of blood vessels colonization in N. meningitidis associated pathogenesis. In this work, using a humanized model of SCID mice allowing the study of bacterial adhesion to human cells in an in vivo context we demonstrate that meningococcal colonization of human blood vessels is a prerequisite to the establishment of sepsis and lethality...
November 3, 2017: Virulence
https://www.readbyqxmd.com/read/29032547/genome-wide-mutagenesis-in-borrelia-burgdorferi
#13
Tao Lin, Lihui Gao
Signature-tagged mutagenesis (STM) is a functional genomics approach to identify bacterial virulence determinants and virulence factors by simultaneously screening multiple mutants in a single host animal, and has been utilized extensively for the study of bacterial pathogenesis, host-pathogen interactions, and spirochete and tick biology. The signature-tagged transposon mutagenesis has been developed to investigate virulence determinants and pathogenesis of Borrelia burgdorferi. Mutants in genes important in virulence are identified by negative selection in which the mutants fail to colonize or disseminate in the animal host and tick vector...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28994778/creation-of-a-dense-transposon-insertion-library-using-bacterial-conjugation-in-enterobacterial-strains-such-as-escherichia-coli-or-shigella-flexneri
#14
Nikki E Freed
Transposon mutagenesis is a method that allows gene disruption via the random genomic insertion of a piece of DNA called a transposon. The protocol below outlines a method for high efficiency transfer between bacterial strains of a plasmid harboring a transposon containing a kanamycin resistance marker. The plasmid-borne transposase is encoded by a variant tnp gene that inserts the transposon into the genome of the recipient strain with very low insertional bias. This method thus allows the creation of large mutant libraries in which transposons have been inserted into unique genomic positions in a recipient strain of either Escherichia coli or Shigella flexneri bacteria...
September 23, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28943871/global-screening-of-salmonella-enterica-serovar-typhimurium-genes-for-desiccation-survival
#15
Rabindra K Mandal, Young M Kwon
Salmonella spp., one of the most common foodborne bacterial pathogens, has the ability to survive under desiccation conditions in foods and food processing facilities for years. This raises the concerns of Salmonella infection in humans associated with low water activity foods. Salmonella responds to desiccation stress via complex pathways involving immediate physiological actions as well as coordinated genetic responses. However, the exact mechanisms of Salmonella to resist desiccation stress remain to be fully elucidated...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/28900609/genes-contributing-to-porphyromonas-gingivalis-fitness-in-abscess-and-epithelial-cell-colonization-environments
#16
Daniel P Miller, Justin A Hutcherson, Yan Wang, Zuzanna M Nowakowska, Jan Potempa, Deborah R Yoder-Himes, David A Scott, Marvin Whiteley, Richard J Lamont
Porphyromonas gingivalis is an important cause of serious periodontal diseases, and is emerging as a pathogen in several systemic conditions including some forms of cancer. Initial colonization by P. gingivalis involves interaction with gingival epithelial cells, and the organism can also access host tissues and spread haematogenously. To better understand the mechanisms underlying these properties, we utilized a highly saturated transposon insertion library of P. gingivalis, and assessed the fitness of mutants during epithelial cell colonization and survival in a murine abscess model by high-throughput sequencing (Tn-Seq)...
2017: Frontiers in Cellular and Infection Microbiology
https://www.readbyqxmd.com/read/28855313/complexity-of-complement-resistance-factors-expressed-by-acinetobacter-baumannii-needed-for-survival-in-human-serum
#17
Amaro F Sanchez-Larrayoz, Noha M Elhosseiny, Marc G Chevrette, Yang Fu, Peter Giunta, Raúl G Spallanzani, Keerthikka Ravi, Gerald B Pier, Stephen Lory, Tomás Maira-Litrán
Acinetobacter baumannii is a bacterial pathogen with increasing impact in healthcare settings, due in part to this organism's resistance to many antimicrobial agents, with pneumonia and bacteremia as the most common manifestations of disease. A significant proportion of clinically relevant A. baumannii strains are resistant to killing by normal human serum (NHS), an observation supported in this study by showing that 12 out of 15 genetically diverse strains of A. baumannii are resistant to NHS killing. To expand our understanding of the genetic basis of A...
October 15, 2017: Journal of Immunology: Official Journal of the American Association of Immunologists
https://www.readbyqxmd.com/read/28832676/genome-wide-discovery-of-novel-m1t1-group-a-streptococcal-determinants-important-for-fitness-and-virulence-during-soft-tissue-infection
#18
Yoann Le Breton, Ashton T Belew, Jeffrey A Freiberg, Ganesh S Sundar, Emrul Islam, Joshua Lieberman, Mark E Shirtliff, Hervé Tettelin, Najib M El-Sayed, Kevin S McIver
The Group A Streptococcus remains a significant human pathogen causing a wide array of disease ranging from self-limiting to life-threatening invasive infections. Epithelium (skin or throat) colonization with progression to the subepithelial tissues is the common step in all GAS infections. Here, we used transposon-sequencing (Tn-seq) to define the GAS 5448 genetic requirements for in vivo fitness in subepithelial tissue. A near-saturation transposon library of the M1T1 GAS 5448 strain was injected subcutaneously into mice, producing suppurative inflammation at 24 h that progressed to prominent abscesses with tissue necrosis at 48 h...
August 2017: PLoS Pathogens
https://www.readbyqxmd.com/read/28806924/essential-genome-of-campylobacter-jejuni
#19
Rabindra K Mandal, Tieshan Jiang, Young Min Kwon
BACKGROUND: Campylobacter species are a leading cause of bacterial foodborne illness worldwide. Despite the global efforts to curb them, Campylobacter infections have increased continuously in both developed and developing countries. The development of effective strategies to control the infection by this pathogen is warranted. The essential genes of bacteria are the most prominent targets for this purpose. In this study, we used transposon sequencing (Tn-seq) of a genome-saturating library of Tn5 insertion mutants to define the essential genome of C...
August 14, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28765224/genome-wide-sensitivity-analysis-of-the-microsymbiont-sinorhizobium-meliloti-to-symbiotically-important-defensin-like-host-peptides
#20
Markus F F Arnold, Mohammed Shabab, Jon Penterman, Kevin L Boehme, Joel S Griffitts, Graham C Walker
The model legume species Medicago truncatula expresses more than 700 nodule-specific cysteine-rich (NCR) signaling peptides that mediate the differentiation of Sinorhizobium meliloti bacteria into nitrogen-fixing bacteroids. NCR peptides are essential for a successful symbiosis in legume plants of the inverted-repeat-lacking clade (IRLC) and show similarity to mammalian defensins. In addition to signaling functions, many NCR peptides exhibit antimicrobial activity in vitro and in vivo Bacterial resistance to these antimicrobial activities is likely to be important for symbiosis...
August 1, 2017: MBio
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