Sterilization Effects on Nitric Oxide-Releasing Glucose Sensors.

Nitric oxide (NO) release from S-nitrosothiol-modified mesoporous silica nanoparticles imbedded in the diffusion limiting layer of a glucose sensor has been demonstrated as an effective strategy for mitigating the foreign body response common to sensor implantation, resulting in improved analytical performance. With respect to potential clinical translation of this approach, the effects of sterilization on NO-releasing biosensors require careful evaluation, as NO donor chemistry is sensitive to temperature and environment. Herein, we evaluated the influence of multiple sterilization methods on 1) sterilization success; 2) NO payload; and 3) sensor performance to establish the commercialization potential of NO-releasing glucose sensors. Sensors were treated with ethylene oxide gas, the most common sterilization method for intricate medical devices, which led to undesirable (i.e., premature) release of NO. To reduce NO loss, alternative sterilization methods that were studied included exposure to ultraviolet (UV) light and immersion in 70% ethanol (EtOH). Sterilization cycle times required to reach a 10-6 sterility assurance level were determined for both UV light and 70% EtOH against Gram-negative and -positive bacteria. The longest sterilization cycle times (258 s and 628 s for 70% EtOH and UV light, respectively) resulted in a negligible impact on benchtop sensor performance. However, sterilization with 70% ethanol resulted in a reduced NO-release duration. Ultraviolet light exposure for ~10 min proved successful at eliminating bacteria without compromising NO payloads or durations and presents as the most promising method for sterilization of these sensors. In addition, storage of NO-releasing sensor membranes at -20 and -80°C resulted in preservation of NO release for 6 and 12 months, respectively.

A consortia of clinical E. coli strains with distinct in vitro adherent/invasive properties establish their own co-colonization niche and shape the intestinal microbiota in inflammation-susceptible mice.

BACKGROUND: Inflammatory bowel disease (IBD) patients experience recurrent episodes of intestinal inflammation and often follow an unpredictable disease course. Mucosal colonization with adherent-invasive Escherichia coli (AIEC) are believed to perpetuate intestinal inflammation. However, it remains unclear if the 24-year-old AIEC in vitro definition fully predicts mucosal colonization in vivo. To fill this gap, we have developed a novel molecular barcoding approach to distinguish strain variants in the gut and have integrated this approach to explore mucosal colonization of distinct patient-derived E. coli isolates in gnotobiotic mouse models of colitis. RESULTS: Germ-free inflammation-susceptible interleukin-10-deficient (Il10-/-) and inflammation-resistant WT mice were colonized with a consortium of AIEC and non-AIEC strains, then given a murine fecal transplant to provide niche competition. E. coli strains isolated from human intestinal tissue were each marked with a unique molecular barcode that permits identification and quantification by barcode-targeted sequencing. 16S rRNA sequencing was used to evaluate the microbiome response to E. coli colonization. Our data reveal that specific AIEC and non-AIEC strains reproducibly colonize the intestinal mucosa of WT and Il10-/- mice. These E. coli expand in Il10-/- mice during inflammation and induce compositional dysbiosis to the microbiome in an inflammation-dependent manner. In turn, specific microbes co-evolve in inflamed mice, potentially diversifying E. coli colonization patterns. We observed no selectivity in E. coli colonization patterns in the fecal contents, indicating minimal selective pressure in this niche from host-microbe and interbacterial interactions. Because select AIEC and non-AIEC strains colonize the mucosa, this suggests the in vitro AIEC definition may not fully predict in vivo colonization potential. Further comparison of seven E. coli genomes pinpointed unique genomic features contained only in highly colonizing strains (two AIEC and two non-AIEC). Those colonization-associated features may convey metabolic advantages (e.g., iron acquisition and carbohydrate consumption) to promote efficient mucosal colonization. CONCLUSIONS: Our findings establish the in vivo mucosal colonizer, not necessarily AIEC, as a principal dysbiosis driver through crosstalk with host and associated microbes. Furthermore, we highlight the utility of high-throughput screens to decode the in vivo colonization dynamics of patient-derived bacteria in murine models. Video Abstract.

Nitric Oxide-Induced Morphological Changes to Bacteria.

Antimicrobial resistance poses a serious threat to global health, necessitating research for alternative approaches to treating infections. Nitric oxide (NO) is an endogenously produced molecule involved in multiple physiological processes, including the response to pathogens. Herein, we employed microscopy- and fluorescence-based techniques to investigate the effects of NO delivered from exogenous NO donors on the bacterial cell envelopes of pathogens, including resistant strains. Our goal was to assess the role of NO donor architecture (small molecules, oligosaccharides, dendrimers) on bacterial wall degradation to representative Gram-negative bacteria (Klebsiella pneumoniae, Pseudomonas aeruginosa) and Gram-positive bacteria (Staphylococcus aureus, Enterococcus faecium) upon treatment. Depending on the NO donor, bactericidal NO doses spanned 1.5-5.5 mM (total NO released). Transmission electron microscopy of bacteria following NO exposure indicated extensive membrane damage to Gram-negative bacteria with warping of the cellular shape and disruption of the cell wall. Among the small-molecule NO donors, those providing a more extended release (t1/2 = 120 min) resulted in greater damage to Gram-negative bacteria. In contrast, rapid NO release (t1/2 = 24 min) altered neither the morphology nor the roughness of these bacteria. For Gram-positive bacteria, NO treatments did not result in any drastic change to cellular shape or membrane integrity, despite permeation of the cell wall as measured by depolarization assays. The use of positively charged quaternary ammonium (QA)-modified NO-releasing dendrimer proved to be the only NO donor system capable of penetrating the thick peptidoglycan layer of Gram-positive bacteria.

Biofilm Dispersal, Reduced Viscoelasticity, and Antibiotic Sensitization via Nitric Oxide-Releasing Biopolymers.

Compared to planktonic bacteria, biofilms are notoriously difficult to eradicate due to their inherent protection against the immune response and antimicrobial agents. Inducing biofilm dispersal to improve susceptibility to antibiotics is an attractive therapeutic avenue for eradicating biofilms. Nitric oxide (NO), an endogenous antibacterial agent, has previously been shown to induce biofilm dispersal, but with limited understanding of the effects of NO-release properties. Herein, the antibiofilm effects of five promising NO-releasing biopolymer candidates were studied by assessing dispersal, changes in biofilm viscoelasticity, and increased sensitization to tobramycin after treatment with NO. A threshold level of NO was needed to achieve biofilm dispersal, with longer-releasing systems requiring lower concentrations. The most positively charged NO-release systems (from the presence of primary amines) led to the greatest reduction in viscoelasticity of Pseudomonas aeruginosa biofilms. Co-treatment of tobramycin with the NO-releasing biopolymer greatly decreased the dose of tobramycin required to eradicate tobramycin-susceptible and -resistant biofilms in both cellular and tissue models.

A consortia of clinical E. coli strains with distinct in-vitro adherent/invasive properties establish their own co-colonization niche and shape the intestinal microbiota in inflammation-susceptible mice.

Background: Inflammatory bowel disease (IBD) patients experience recurrent episodes of intestinal inflammation and often follow an unpredictable disease course. Mucosal colonization with adherent-invasive Escherichia coli (AIEC) are believed to perpetuate intestinal inflammation. However, it remains unclear if the 24-year-old AIEC in-vitro definition fully predicts mucosal colonization in-vivo. To fill this gap, we have developed a novel molecular barcoding approach to distinguish strain variants in the gut and have integrated this approach to explore mucosal colonization of distinct patient-derived E. coli isolates in gnotobiotic mouse models of colitis. Results: Germ-free inflammation-susceptible interleukin-10-deficient (Il10-/-) and inflammation-resistant WT mice were colonized with a consortia of AIEC and non-AIEC strains, then given a murine fecal transplant to provide niche competition. E. coli strains isolated from human intestinal tissue were each marked with a unique molecular barcode that permits identification and quantification by barcode-targeted sequencing. 16S rRNA sequencing was used to evaluate the microbiome response to E. coli colonization. Our data reveal that specific AIEC and non-AIEC strains reproducibly colonize the intestinal mucosa of WT and Il10-/- mice. These E. coli expand in Il10-/- mice during inflammation and induce compositional dysbiosis to the microbiome in an inflammation-dependent manner. In turn, specific microbes co-evolve in inflamed mice, potentially diversifying E. coli colonization patterns. We observed no selectivity in E. coli colonization patterns in the fecal contents, indicating minimal selective pressure in this niche from host-microbe and interbacterial interactions. Because select AIEC and non-AIEC strains colonize the mucosa, this suggests the in vitro AIEC definition may not fully predict in vivo colonization potential. Further comparison of seven E. coli genomes pinpointed unique genomic features contained only in highly colonizing strains (two AIEC and two non-AIEC). Those colonization-associated features may convey metabolic advantages (e.g., iron acquisition and carbohydrate consumption) to promote efficient mucosal colonization. Conclusions: Our findings establish the in-vivo mucosal colonizer, not necessarily AIEC, as a principal dysbiosis driver through crosstalk with host and associated microbes. Furthermore, we highlight the utility of high-throughput screens to decode the in-vivo colonization dynamics of patient-derived bacteria in murine models.

Role of Nitric Oxide-Releasing Glycosaminoglycans in Wound Healing.

Two glycosaminoglycan (GAG) biopolymers, hyaluronic acid (HA) and chondroitin sulfate (CS), were chemically modified via carbodiimide chemistry to facilitate the loading and release of nitric oxide (NO) to develop a multi-action wound healing agent. The resulting NO-releasing GAGs released 0.2-0.9 µmol NO mg-1 GAG into simulated wound fluid with NO-release half-lives ranging from 20 to 110 min. GAGs containing alkylamines with terminal primary amines and displaying intermediate NO-release kinetics exhibited potent, broad spectrum bactericidal action against three strains each of Pseudomonas aeruginosa and Staphylococcus aureus ranging in antibiotic resistance profile. NO loading of the GAGs was also found to decrease murine TLR4 activation, suggesting that the therapeutic exhibits anti-inflammatory mechanisms. In vitro adhesion and proliferation assays utilizing human dermal fibroblasts and human epidermal keratinocytes displayed differences as a function of the GAG backbone, alkylamine identity, and NO-release properties. In combination with antibacterial properties, the adhesion and proliferation profiles of the GAG derivatives enabled the selection of the most promising wound healing candidates for subsequent in vivo studies. A P. aeruginosa-infected murine wound model revealed the benefits of CS over HA as a pro-wound healing NO donor scaffold, with benefits of accelerated wound closure and decreased bacterial burden attributable to both active NO release and the biopolymer backbone.

A nadA Mutation Confers Nicotinic Acid Auxotrophy in Pro-carcinogenic Intestinal Escherichia coli NC101.

Inflammatory bowel diseases (IBDs) and inflammation-associated colorectal cancer (CRC) are linked to blooms of adherent-invasive Escherichia coli (AIEC) in the intestinal microbiota. AIEC are functionally defined by their ability to adhere/invade epithelial cells and survive/replicate within macrophages. Changes in micronutrient availability can alter AIEC physiology and interactions with host cells. Thus, culturing AIEC for mechanistic investigations often involves precise nutrient formulation. We observed that the pro-inflammatory and pro-carcinogenic AIEC strain NC101 failed to grow in minimal media (MM). We hypothesized that NC101 was unable to synthesize a vital micronutrient normally found in the host gut. Through nutrient supplementation studies, we identified that NC101 is a nicotinic acid (NA) auxotroph. NA auxotrophy was not observed in the other non-toxigenic E. coli or AIEC strains we tested. Sequencing revealed NC101 has a missense mutation in nadA, a gene encoding quinolinate synthase A that is important for de novo nicotinamide adenine dinucleotide (NAD) biosynthesis. Correcting the identified nadA point mutation restored NC101 prototrophy without impacting AIEC function, including motility and AIEC-defining survival in macrophages. Our findings, along with the generation of a prototrophic NC101 strain, will greatly enhance the ability to perform in vitro functional studies that are needed for mechanistic investigations on the role of intestinal E. coli in digestive disease.

Yersiniabactin-Producing Adherent/Invasive Escherichia coli Promotes Inflammation-Associated Fibrosis in Gnotobiotic Il10-/- Mice.

Fibrosis is a significant complication of intestinal disorders associated with microbial dysbiosis and pathobiont expansion, notably Crohn's disease (CD). Mechanisms that favor fibrosis are not well understood, and therapeutic strategies are limited. Here we demonstrate that colitis-susceptible Il10-deficient mice develop inflammation-associated fibrosis when monoassociated with adherent/invasive Escherichia coli (AIEC) that harbors the yersiniabactin (Ybt) pathogenicity island. Inactivation of Ybt siderophore production in AIEC nearly abrogated fibrosis development in inflamed mice. In contrast, inactivation of Ybt import through its cognate receptor FyuA enhanced fibrosis severity. This corresponded with increased colonic expression of profibrogenic genes prior to the development of histological disease, therefore suggesting causality. fyuA-deficient AIEC also exhibited greater localization within subepithelial tissues and fibrotic lesions that was dependent on Ybt biosynthesis and corresponded with increased fibroblast activation in vitro Together, these findings suggest that Ybt establishes a profibrotic environment in the host in the absence of binding to its cognate receptor and indicate a direct link between intestinal AIEC and the induction of inflammation-associated fibrosis.

A Klebsiella pneumoniae Regulatory Mutant Has Reduced Capsule Expression but Retains Hypermucoviscosity.

The polysaccharide capsule is an essential virulence factor for Klebsiella pneumoniae in both community-acquired hypervirulent strains as well as health care-associated classical strains that are posing significant challenges due to multidrug resistance. Capsule production is known to be transcriptionally regulated by a number of proteins, but very little is known about how these proteins collectively control capsule production. RmpA and RcsB are two known regulators of capsule gene expression, and RmpA is required for the hypermucoviscous (HMV) phenotype in hypervirulent K. pneumoniae strains. In this report, we confirmed that these regulators performed their anticipated functions in the ATCC 43816 derivative, KPPR1S: rcsB and rmpA mutants are HMV negative and have reduced capsule gene expression. We also identified a novel transcriptional regulator, RmpC, encoded by a gene near rmpA The ΔrmpC strain has reduced capsule gene expression but retains the HMV phenotype. We further showed that a regulatory cascade exists in which KvrA and KvrB, the recently characterized MarR-like regulators, and RcsB contribute to capsule regulation through regulation of the rmpA promoter and through additional mechanisms. In a murine pneumonia model, the regulator mutants have a range of colonization defects, suggesting that they regulate virulence factors in addition to capsule. Further testing of the rmpC and rmpA mutants revealed that they have distinct and overlapping functions and provide evidence that HMV is not dependent on overproduction of capsule. This distinction will facilitate a better understanding of HMV and how it contributes to enhanced virulence of hypervirulent strains.IMPORTANCEKlebsiella pneumoniae continues to be a substantial public health threat due to its ability to cause health care-associated and community-acquired infections combined with its ability to acquire antibiotic resistance. Novel therapeutics are needed to combat this pathogen, and a greater understanding of its virulence factors is required for the development of new drugs. A key virulence factor for K. pneumoniae is the capsule, and community-acquired hypervirulent strains produce a capsule that causes hypermucoidy. We report here a novel capsule regulator, RmpC, and provide evidence that capsule production and the hypermucoviscosity phenotype are distinct processes. Infection studies showing that this and other capsule regulator mutants have a range of phenotypes indicate that additional virulence factors are in their regulons. These results shed new light on the mechanisms controlling capsule production and introduce targets that may prove useful for the development of novel therapeutics for the treatment of this increasingly problematic pathogen.

A Serendipitous Mutation Reveals the Severe Virulence Defect of a Klebsiella pneumoniae fepB Mutant.

Klebsiella pneumoniae is considered a significant public health threat because of the emergence of multidrug-resistant strains and the challenge associated with treating life-threatening infections. Capsule, siderophores, and adhesins have been implicated as virulence determinants of K. pneumoniae, yet we lack a clear understanding of how this pathogen causes disease. In a previous screen for virulence genes, we identified a potential new virulence locus and constructed a mutant (smr) with this locus deleted. In this study, we characterize the smr mutant and show that this mutation renders K. pneumoniae avirulent in a pneumonia model of infection. The smr mutant was expected to have a deletion of three genes, but subsequent genome sequencing indicated that a much larger deletion had occurred. Further analysis of the deleted region indicated that the virulence defect of the smr mutant could be attributed to the loss of FepB, a periplasmic protein required for import of the siderophore enterobactin. Interestingly, a ΔfepB mutant was more attenuated than a mutant unable to synthesize enterobactin, suggesting that additional processes are affected. As FepB is highly conserved among the members of the family Enterobacteriaceae, therapeutic targeting of FepB may be useful for the treatment of Klebsiella and other bacterial infections. IMPORTANCE In addition to having a reputation as the causative agent of several types of hospital-acquired infections, Klebsiella pneumoniae has gained widespread attention as a pathogen with a propensity for acquiring antibiotic resistance. It is capable of causing a range of infections, including urinary tract infections, pneumonia, and sepsis. Because of the rapid emergence of carbapenem resistance among Klebsiella strains, there is a dire need for a better understanding of virulence mechanisms and identification of new drug targets. Here, we identify the periplasmic transporter FepB as one such potential target.

The "PepSAVI-MS" Pipeline for Natural Product Bioactive Peptide Discovery.

The recent increase in extensively drug-resistant bacterial pathogens and the associated increase of morbidity and mortality demonstrate the immediate need for new antibiotic backbones with novel mechanisms of action. Here, we report the development of the PepSAVI-MS pipeline for bioactive peptide discovery. This highly versatile platform employs mass spectrometry and statistics to identify bioactive peptide targets from complex biological samples. We validate the use of this platform through the successful identification of known bioactive peptides from a botanical species, Viola odorata. Using this pipeline, we have widened the known antimicrobial spectrum for V. odorata cyclotides, including antibacterial activity of cycloviolacin O2 against A. baumannii. We further demonstrate the broad applicability of the platform through the identification of novel anticancer activities for cycloviolacins by their cytotoxicity against ovarian, breast, and prostate cancer cell lines.

In vivo transcriptional profiling of Yersinia pestis reveals a novel bacterial mediator of pulmonary inflammation.

UNLABELLED: Inhalation of Yersinia pestis results in primary pneumonic plague, a highly lethal and rapidly progressing necrotizing pneumonia. The disease begins with a period of extensive bacterial replication in the absence of disease symptoms, followed by the sudden onset of inflammatory responses that ultimately prove fatal. Very little is known about the bacterial and host factors that contribute to the rapid biphasic progression of pneumonic plague. In this work, we analyzed the in vivo transcription kinetics of 288 bacterial open reading frames previously shown by microarray analysis to be dynamically regulated in the lung. Using this approach combined with bacterial genetics, we were able to identify five Y. pestis genes that contribute to the development of pneumonic plague. Deletion of one of these genes, ybtX, did not alter bacterial survival but attenuated host inflammatory responses during late-stage disease. Deletion of ybtX in another lethal respiratory pathogen, Klebsiella pneumoniae, also resulted in diminished host inflammation during infection. Thus, our in vivo transcriptional screen has identified an important inflammatory mediator that is common to two Gram-negative bacterial pathogens that cause severe pneumonia. IMPORTANCE: Yersinia pestis is responsible for at least three major pandemics, most notably the Black Death of the Middle Ages. Due to its pandemic potential, ease of dissemination by aerosolization, and a history of its weaponization, Y. pestis is categorized by the Centers for Disease Control and Prevention as a tier 1 select agent most likely to be used as a biological weapon. To date, there is no licensed vaccine against Y. pestis. Importantly, an early "silent" phase followed by the rapid onset of nondescript influenza-like symptoms makes timely treatment of pneumonic plague difficult. A more detailed understanding of the bacterial and host factors that contribute to pathogenesis is essential to understanding the progression of pneumonic plague and developing or enhancing treatment options.

Complete Genome Sequence of Klebsiella pneumoniae Strain ATCC 43816 KPPR1, a Rifampin-Resistant Mutant Commonly Used in Animal, Genetic, and Molecular Biology Studies.

Klebsiella pneumoniae is an urgent public health threat due to the spread of carbapenem-resistant strains causing serious, and frequently fatal, infections. To facilitate genetic, molecular, and immunological studies of this pathogen, we report the complete chromosomal sequence of a genetically tractable, prototypical strain used in animal models.

Klebsiella: a long way to go towards understanding this enigmatic jet-setter.

Klebsiella pneumoniae is the causative agent of a variety of diseases, including pneumonia, urinary tract infections, septicemia, and the recently recognized pyogenic liver abscesses (PLA). Renewed efforts to identify and understand the bacterial determinants required to cause disease have come about because of the worldwide increase in the isolation of strains resistant to a broad spectrum of antibiotics. The recent increased isolation of carbapenem-resistant strains further reduces the available treatment options. The rapid geographic spread of the resistant isolates and the spread to other pathogens are of particular concern. For many years, the best characterized virulence determinants were capsule, lipopolysaccharide, siderophores, and types 1 and 3 fimbriae. Recent efforts to expand this list include in vivo screens and whole-genome sequencing. However, we still know little about how this bacterium is able to cause disease. Some recent clonal analyses of K. pneumoniae strains indicate that there are distinct clonal groups, some of which may be associated with specific disease syndromes. However, what makes one clonal group more virulent and what changes the disease pattern are not yet clear and remain important questions for the future.

Whole-Genome Draft Sequences of Three Multidrug-Resistant Klebsiella pneumoniae Strains Available from the American Type Culture Collection.

Infection with multidrug-resistant Klebsiella pneumoniae is a significant problem worldwide, requiring a better understanding of how various strains are able to defeat current antibiotic therapies and cause disease. Here, we report the draft genome sequences of three K. pneumoniae strains harboring the SHV-18, KPC-2, or NDM-1 ß-lactamases.

Type III effector VopC mediates invasion for Vibrio species.

Vibrio spp. are associated with infections caused by contaminated food and water. A type III secretion system (T3SS2) is a shared feature of all clinical isolates of V. parahaemolyticus and some V. cholerae strains. Despite its being responsible for enterotoxicity, no molecular mechanism has been determined for the T3SS2-dependent pathogenicity. Here, we show that although Vibrio spp. are typically thought of as extracellular pathogens, the T3SS2 of Vibrio mediates host cell invasion, vacuole formation, and replication of intracellular bacteria. The catalytically active effector VopC is critical for Vibrio T3SS2-mediated invasion. There are other marine bacteria encoding VopC homologs associated with a T3SS; therefore, we predict that these bacteria are also likely to use T3SS-mediated invasion as part of their pathogenesis mechanisms. These findings suggest a new molecular paradigm for Vibrio pathogenicity and modify our view of the roles of T3SS effectors that are translocated during infection.

Vibrio parahaemolyticus cell biology and pathogenicity determinants.

Vibrio parahaemolyticus is a significant cause of gastroenteritis worldwide. Characterization of this pathogen has revealed a unique repertoire of virulence factors that allow for colonization of the human host and disease. The following describes the known pathogenicity determinants while establishing the need for continued research.

Shotgun proteomics of Xanthobacter autotrophicus Py2 reveals proteins specific to growth on propylene.

Coenzyme M (CoM, 2-mercaptoethanesulfonate), once thought to be exclusively produced by methanogens, is now known to be the central cofactor in the metabolism of short-chain alkenes by a variety of aerobic bacteria. There is little evidence to suggest how, and under what conditions, CoM is biosynthesized by these organisms. A shotgun proteomics approach was used to investigate CoM-dependent propylene metabolism in the Gram-negative bacterium Xanthobacter autotrophicus Py2. Cells were grown on either glucose or propylene, and the soluble proteomes were analyzed. An average of 395 proteins was identified from glucose-grown replicates, with an average of 419 identified from propylene-grown replicates. A number of linear megaplasmid (pXAUT01)-encoded proteins were found to be specifically produced by growth on propylene. These included all known to be crucial to propylene metabolism, in addition to an aldehyde dehydrogenase, a DNA-binding protein, and five putative CoM biosynthetic enzymes. This work has provided fresh insight into bacterial alkene metabolism and has generated new targets for future studies in X. autotrophicus Py2 and related CoM-dependent alkene-oxidizing bacteria.

A Vibrio effector protein is an inositol phosphatase and disrupts host cell membrane integrity.

The marine bacterium Vibrio parahaemolyticus causes gastroenteritis in humans and encodes the type III effector protein VPA0450, which contributes to host cell death caused by autophagy, cell rounding, and cell lysis. We found that VPA0450 is an inositol polyphosphate 5-phosphatase that hydrolyzed the D5 phosphate from the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate. VPA0450 disrupted cytoskeletal binding sites on the inner surface of membranes of human cells and caused plasma membrane blebbing, which compromised membrane integrity and probably contributed to cell death by facilitating lysis. Thus, bacterial pathogens can disrupt adaptor protein-binding sites required for proper membrane and cytoskeleton dynamics by altering the homeostasis of membrane-bound inositol-signaling molecules.

Tipping the balance by manipulating post-translational modifications.

Bacteria use a variety of mechanisms during infection to ensure their survival including the delivery of virulence factors via a type III secretion system into the infected cell. The factors exhibit diverse activities that in many cases mimic eukaryotic mechanisms used by the host to defend against infection. Herein we describe a class of effectors that use post-translational modifications, some reversible and others irreversible, to manipulate host signaling systems to subvert the host response.