A anti-phage system depletes nicotinamide adenine dinucleotide to restrict virulent bacteriophages.

Bacteria and their predatory viruses (bacteriophages or phages) are in a perpetual molecular arms race. This has led to the evolution of numerous phage defensive systems in bacteria that are still being discovered, as well as numerous ways of interference or circumvention on the part of phages. Here, we identify a unique molecular battle between the classical biotype of and virulent phages ICP1, ICP2, and ICP3.

A Anti-Phage System Depletes Nicotinamide Adenine Dinucleotide to Restrict Virulent Bacteriophages.

Bacteria and their predatory viruses (bacteriophages or phages) are in a perpetual molecular arms race. This has led to the evolution of numerous phage defensive systems in bacteria that are still being discovered, as well as numerous ways of interference or circumvention on the part of phages. Here, we identify a unique molecular battle between the classical biotype of and virulent phages ICP1, ICP2, and ICP3.

A Tail Fiber Protein and a Receptor-Binding Protein Mediate ICP2 Bacteriophage Interactions with Vibrio cholerae OmpU.

ICP2 is a virulent bacteriophage (phage) that preys on Vibrio cholerae. ICP2 was first isolated from cholera patient stool samples. Some of these stools also contained ICP2-resistant isogenic V.

Author Correction: Antibiotic susceptibility signatures identify potential antimicrobial targets in the Acinetobacter baumannii cell envelope.

A Correction to this paper has been published: https://doi.org/10.1038/s41467-020-20098-z.

Identification of Spacer and Protospacer Sequence Requirements in the Vibrio cholerae Type I-E CRISPR/Cas System.

The prokaryotic adaptive immune system CRISPR/Cas serves as a defense against bacteriophage and invasive nucleic acids. A type I-E CRISPR/Cas system has been detected in classical biotype isolates of , the causative agent of the disease cholera. Experimental characterization of this system revealed a functional immune system that operates using a 5'-TT-3' protospacer-adjacent motif (PAM) for interference.

Antibiotic susceptibility signatures identify potential antimicrobial targets in the Acinetobacter baumannii cell envelope.

A unique, protective cell envelope contributes to the broad drug resistance of the nosocomial pathogen Acinetobacter baumannii. Here we use transposon insertion sequencing to identify A. baumannii mutants displaying altered susceptibility to a panel of diverse antibiotics.

Transposon Mutagenesis Screen of Klebsiella pneumoniae Identifies Multiple Genes Important for Resisting Antimicrobial Activities of Neutrophils in Mice.

is a Gram-negative bacterial pathogen that causes a range of infections, including pneumonias, urinary tract infections, and septicemia, in otherwise healthy and immunocompromised patients. has become an increasing concern due to the rise and spread of antibiotic-resistant and hypervirulent strains. However, its virulence determinants remain understudied.

High-Throughput Analysis of Gene Function in the Bacterial Predator Bdellovibrio bacteriovorus.

is a bacterial predator capable of killing and replicating inside most Gram-negative bacteria, including antibiotic-resistant pathogens. Despite growing interest in this organism as a potential therapeutic, many of its genes remain uncharacterized. Here, we perform a high-throughput genetic screen with using transposon sequencing (Tn-seq) to explore the genetic requirements of predation.

Vibrio cholerae motility exerts drag force to impede attack by the bacterial predator Bdellovibrio bacteriovorus.

The bacterial predator Bdellovibrio bacteriovorus is evolved to attack and kill other bacteria, including the human intestinal pathogen Vibrio cholerae. Although B. bacteriovorus exhibit a broad prey range, little is known about the genetic determinants of prey resistance and sensitivity.

Growth arrest and a persister state enable resistance to osmotic shock and facilitate dissemination of Vibrio cholerae.

Vibrio cholerae is a water-borne bacterial pathogen and causative agent of cholera. Although V. cholerae is a halophile, it can survive in fresh water, and this has a major role in cholera epidemics through consumption of contaminated water and subsequent fecal-oral spread.

Immunity Provided by an Outer Membrane Vesicle Cholera Vaccine Is Due to O-Antigen-Specific Antibodies Inhibiting Bacterial Motility.

An outer membrane vesicle (OMV)-based cholera vaccine is highly efficacious in preventing intestinal colonization in the suckling mouse model. Immunity from OMVs comes from immunoglobulin (Ig), particularly IgG, in the milk of mucosally immunized dams. Anti-OMV IgG renders Vibrio cholerae organisms immotile, thus they pass through the small intestine without colonizing.

A comparative study of ChIP-seq sequencing library preparation methods.

Background: ChIP-seq is the primary technique used to investigate genome-wide protein-DNA interactions. As part of this procedure, immunoprecipitated DNA must undergo "library preparation" to enable subsequent high-throughput sequencing. To facilitate the analysis of biopsy samples and rare cell populations, there has been a recent proliferation of methods allowing sequencing library preparation from low-input DNA amounts.

Transposon-Sequencing Analysis Unveils Novel Genes Involved in the Generation of Persister Cells in Uropathogenic Escherichia coli.

Persister cells are highly tolerant to different antibiotics and are associated with relapsing infections. In order to understand this phenomenon further, we exposed a transposon library to a lethal concentration of ampicillin, and mutants that survived were identified by transposon sequencing (Tn-Seq). We determined that mutations related to carbon metabolism, cell envelope (cell wall generation and membrane proteins), and stress response have a role in persister cell generation.

Global Tn-seq analysis of carbohydrate utilization and vertebrate infectivity of Borrelia burgdorferi.

Borrelia burgdorferi maintains a complex life cycle between tick and vertebrate hosts. Although some genes have been identified as contributing to bacterial adaptation in the different hosts, the list is incomplete. In this manuscript, we report the first use of transposon mutagenesis combined with high-throughput sequencing (Tn-seq) in B.

Carbon catabolite repression by seryl phosphorylated HPr is essential to Streptococcus pneumoniae in carbohydrate-rich environments.

Carbon catabolite repression (CCR) is a regulatory phenomenon implemented by bacteria to hierarchically organize carbohydrate utilization in order to achieve maximal growth. CCR is likely of great importance to Streptococcus pneumoniae because the human host sites inhabited by this pathogen represent complex carbohydrate environments. In this species, inactivation of the prototypical Gram-positive CCR master regulator, ccpA, attenuates virulence in mice but does not relieve CCR of most metabolic enzymes, suggesting CcpA-independent CCR mechanisms predominate.

Genome-Wide Fitness and Genetic Interactions Determined by Tn-seq, a High-Throughput Massively Parallel Sequencing Method for Microorganisms.

The lagging annotation of bacterial genomes and the inherent genetic complexity of many phenotypes is hindering the discovery of new drug targets and the development of new antimicrobial agents and vaccines. This unit presents Tn-seq, a method that has made it possible to quantitatively determine fitness for most genes in a microorganism and to screen for quantitative genetic interactions on a genome-wide scale and in a high-throughput fashion. Tn-seq can thus direct studies on the annotation of genes and untangle complex phenotypes.

Genes contributing to Staphylococcus aureus fitness in abscess- and infection-related ecologies.

Unlabelled: Staphylococcus aureus is a leading cause of both community- and hospital-acquired infections that are increasingly antibiotic resistant. The emergence of S. aureus resistance to even last-line antibiotics heightens the need for the development of new drugs with novel targets.

Genome-Wide Fitness and Genetic Interactions Determined by Tn-seq, a High-Throughput Massively Parallel Sequencing Method for Microorganisms.

The lagging annotation of bacterial genomes and the inherent genetic complexity of many phenotypes is hindering the discovery of new drug targets and the development of new antimicrobial agents and vaccines. This unit presents Tn-seq, a method that has made it possible to quantitatively determine fitness for most genes in a microorganism and to screen for quantitative genetic interactions on a genome-wide scale and in a high-throughput fashion. Tn-seq can thus direct studies on the annotation of genes and untangle complex phenotypes.

Identification of in vivo regulators of the Vibrio cholerae xds gene using a high-throughput genetic selection.

Vibrio cholerae, the causative agent of cholera, remains a threat to public health in areas with inadequate sanitation. As a waterborne pathogen, V. cholerae moves between two dissimilar environments, aquatic reservoirs and the intestinal tract of humans.

Identification of a membrane-bound transcriptional regulator that links chitin and natural competence in Vibrio cholerae.

Vibrio cholerae is naturally competent when grown on chitin. It is known that expression of the major regulator of competence, TfoX, is controlled by chitin; however, the molecular mechanisms underlying this requirement for chitin have remained unclear. In the present study, we identify and characterize a membrane-bound transcriptional regulator that positively regulates the small RNA (sRNA) TfoR, which posttranscriptionally enhances tfoX translation.

Gene fitness landscapes of Vibrio cholerae at important stages of its life cycle.

Vibrio cholerae has evolved to adeptly transition between the human small intestine and aquatic environments, leading to water-borne spread and transmission of the lethal diarrheal disease cholera. Using a host model that mimics the pathology of human cholera, we applied high density transposon mutagenesis combined with massively parallel sequencing (Tn-seq) to determine the fitness contribution of >90% of all non-essential genes of V. cholerae both during host infection and dissemination.

A core microbiome associated with the peritoneal tumors of pseudomyxoma peritonei.

Background: Pseudomyxoma peritonei (PMP) is a malignancy characterized by dissemination of mucus-secreting cells throughout the peritoneum. This disease is associated with significant morbidity and mortality and despite effective treatment options for early-stage disease, patients with PMP often relapse. Thus, there is a need for additional treatment options to reduce relapse rate and increase long-term survival.

Understanding barriers to Borrelia burgdorferi dissemination during infection using massively parallel sequencing.

Borrelia burgdorferi is an invasive spirochete that can cause acute and chronic infections in the skin, heart, joints, and central nervous system of infected mammalian hosts. Little is understood about where the bacteria encounter the strongest barriers to infection and how different components of the host immune system influence the population as the infection progresses. To identify population bottlenecks in a murine host, we utilized Tn-seq to monitor the composition of mixed populations of B.

Characterization of undermethylated sites in Vibrio cholerae.

The activities of DNA methyltransferases are important for a variety of cellular functions in bacteria. In this study, we developed a modified high-throughput technique called methyl homopolymer tail mediated sequencing (methyl HTM-seq) to identify the undermethylated sites in the Vibrio cholerae genome for the two DNA methyltransferases, Dam, an adenine methyltransferase, and VchM, a cytosine methyltransferase, during growth in rich medium in vitro. Many of the undermethylated sites occurred in intergenic regions, and for most of these sites, we identified the transcription factors responsible for undermethylation.

A bacteriophage encodes its own CRISPR/Cas adaptive response to evade host innate immunity.

Bacteriophages (or phages) are the most abundant biological entities on earth, and are estimated to outnumber their bacterial prey by tenfold. The constant threat of phage predation has led to the evolution of a broad range of bacterial immunity mechanisms that in turn result in the evolution of diverse phage immune evasion strategies, leading to a dynamic co-evolutionary arms race. Although bacterial innate immune mechanisms against phage abound, the only documented bacterial adaptive immune system is the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins) system, which provides sequence-specific protection from invading nucleic acids, including phage.