scholarly journals Regulation by ToxR-Like Proteins Converges onvttRBExpression To Control Type 3 Secretion System-Dependent Caco2-BBE Cytotoxicity in Vibrio cholerae

2016 ◽  
Vol 198 (11) ◽  
pp. 1675-1682 ◽  
Author(s):  
Kelly A. Miller ◽  
Madeline K. Sofia ◽  
Jacob W. A. Weaver ◽  
Christopher H. Seward ◽  
Michelle Dziejman
Keyword(s):  
Gene Expression ◽  
Cell Death ◽  
Vibrio Cholerae ◽  
Secretion System ◽  
Content Type ◽  
Transcriptional Regulatory ◽  
Type 3 Secretion System ◽  
Type 3

ABSTRACTGenes carried on the type 3 secretion system (T3SS) pathogenicity island ofVibrio choleraenon-O1/non-O139 serogroup strain AM-19226 must be precisely regulated in order for bacteria to cause disease. Previously reported results showed that both T3SS function and the presence of bile are required to cause Caco2-BBE cell cytotoxicity during coculture with strain AM-19226. We therefore investigated additional parameters affectingin vitrocell death, including bacterial load and the role of three transmembrane transcriptional regulatory proteins, VttRA, VttRB, and ToxR. VttRAand VttRBare encoded on the horizontally acquired T3SS genomic island, whereas ToxR is encoded on the ancestral chromosome. While strains carrying deletions in any one of the three transcriptional regulatory genes are unable to cause eukaryotic cell death, the results of complementation studies point to a hierarchy of regulatory control that converges onvttRBexpression. The data suggest both that ToxR and VttRAact upstream of VttRBand that modifying the level of eithervttRAorvttRBexpression can strongly influence T3SS gene expression. We therefore propose a model whereby T3SS activity and, hence,in vitrocytotoxicity are ultimately regulated byvttRBexpression.IMPORTANCEIn contrast to O1 and O139 serogroupV. choleraestrains that cause cholera using two main virulence factors (toxin-coregulated pilus [TCP] and cholera toxin [CT]), O39 serogroup strain AM-19226 uses a type 3 secretion system as its principal virulence mechanism. Although the regulatory network governing TCP and CT expression is well understood, the factors influencing T3SS-associated virulence are not. Using anin vitromammalian cell model to investigate the role of three ToxR-like transmembrane transcriptional activators in causing T3SS-dependent cytotoxicity, we found that expression levels and a hierarchical organization were important for promoting T3SS gene expression. Furthermore, our results suggest that horizontally acquired, ToxR-like proteins act in concert with the ancestral ToxR protein to orchestrate T3SS-mediated pathogenicity.

scholarly journals The cytotoxic type 3 secretion system 1 ofVibriorewires host gene expression to subvert cell death and activate cell survival pathways

2017 ◽  
Vol 10 (479) ◽  
pp. eaal4501 ◽  
Author(s):  
Nicole J. De Nisco ◽  
Mohammed Kanchwala ◽  
Peng Li ◽  
Jessie Fernandez ◽  
Chao Xing ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Death ◽  
Cell Survival ◽  
Secretion System ◽  
Host Gene Expression ◽  
Survival Pathways ◽  
Type 3 Secretion System ◽  
Activate Cell ◽  
System 1 ◽  
Type 3

scholarly journals Genomic Avenue to Avian Colisepticemia

mBio ◽  
2015 ◽  
Vol 6 (1) ◽  
Author(s):  
Sagi Huja ◽  
Yaara Oren ◽  
Eva Trost ◽  
Elzbieta Brzuszkiewicz ◽  
Dvora Biran ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genetic Analysis ◽  
Virulence Factors ◽  
Iron Uptake ◽  
Secretion System ◽  
Economic Losses ◽  
Content Type ◽  
E Coli ◽  
Type 3 Secretion System ◽  
Type 3

ABSTRACTHere we present an extensive genomic and genetic analysis of Escherichia coli strains of serotype O78 that represent the major cause of avian colisepticemia, an invasive infection caused by avian pathogenicEscherichia coli(APEC) strains. It is associated with high mortality and morbidity, resulting in significant economic consequences for the poultry industry. To understand the genetic basis of the virulence of avian septicemic E. coli, we sequenced the entire genome of a clinical isolate of serotype O78—O78:H19 ST88 isolate 789 (O78-9)—and compared it with three publicly available APEC O78 sequences and one complete genome of APEC serotype O1 strain. Although there was a large variability in genome content between the APEC strains, several genes were conserved, which are potentially critical for colisepticemia. Some of these genes are present in multiple copies per genome or code for gene products with overlapping function, signifying their importance. A systematic deletion of each of these virulence-related genes identified three systems that are conserved in all septicemic strains examined and are critical for serum survival, a prerequisite for septicemia. These are the plasmid-encoded protein, the defective ETT2 (E. colitype 3 secretion system 2) type 3 secretion system ETT2sepsis, and iron uptake systems. Strain O78-9 is the only APEC O78 strain that also carried the regulon coding for yersiniabactin, the iron binding system of theYersiniahigh-pathogenicity island. Interestingly, this system is the only one that cannot be complemented by other iron uptake systems under iron limitation and in serum.IMPORTANCEAvian colisepticemia is a severe systemic disease of birds causing high morbidity and mortality and resulting in severe economic losses. The bacteria associated with avian colisepticemia are highly antibiotic resistant, making antibiotic treatment ineffective, and there is no effective vaccine due to the multitude of serotypes involved. To understand the disease and work out strategies to combat it, we performed an extensive genomic and genetic analysis of Escherichia coli strains of serotype O78, the major cause of the disease. We identified several potential virulence factors, conserved in all the colisepticemic strains examined, and determined their contribution to growth in serum, an absolute requirement for septicemia. These findings raise the possibility that specific vaccines or drugs can be developed against these critical virulence factors to help combat this economically important disease.

scholarly journals Role of the acquisition of a type 3 secretion system in the emergence of novel pathogenic strains of Xanthomonas

2018 ◽  
Vol 20 (1) ◽  
pp. 33-50 ◽  
Author(s):  
Valérian Meline ◽  
Wesley Delage ◽  
Chrystelle Brin ◽  
Camille Li-Marchetti ◽  
Daniel Sochard ◽  
...  
Keyword(s):  
Secretion System ◽  
Type 3 Secretion System ◽  
Type 3

scholarly journals OpiA, a Type Six Secretion System Substrate, Localizes to the Cell Pole and Plays a Role in Bacterial Growth and Viability in Francisella tularensis LVS

2020 ◽  
Vol 202 (14) ◽  
Author(s):  
Stuart Cantlay ◽  
Kristen Haggerty ◽  
Joseph Horzempa
Keyword(s):  
Cell Size ◽  
Francisella Tularensis ◽  
Secretion System ◽  
Live Vaccine ◽  
Intracellular Pathogen ◽  
Content Type ◽  
Erythrocyte Invasion ◽  
Host Pathogen

ABSTRACT Francisella tularensis is an intracellular pathogen and the causative agent of tularemia. The F. tularensis type six secretion system (T6SS) is required for a number of host-pathogen interactions, including phagolysosomal escape and invasion of erythrocytes. One known effector of the T6SS, OpiA, has recently been shown to be a phosphatidylinositol-3 kinase. To investigate the role of OpiA in erythrocyte invasion, we constructed an opiA-null mutant in the live vaccine strain, F. tularensis LVS. OpiA was not required for erythrocyte invasion; however, deletion of opiA affected growth of F. tularensis LVS in broth cultures in a medium-dependent manner. We also found that opiA influenced cell size, gentamicin sensitivity, bacterial viability, and the lipid content of F. tularensis. A fluorescently tagged OpiA (OpiA–emerald-green fluorescent protein [EmGFP]) accumulated at the cell poles of F. tularensis, which is consistent with the location of the T6SS. However, OpiA-EmGFP also exhibited a highly dynamic localization, and this fusion protein was detected in erythrocytes and THP-1 cells in vitro, further supporting that OpiA is secreted. Similar to previous reports with F. novicida, our data demonstrated that opiA had a minimal effect on intracellular replication of F. tularensis in host immune cells in vitro. However, THP-1 cells infected with the opiA mutant produced modestly (but significantly) higher levels of the proinflammatory cytokine tumor necrosis factor alpha compared to these host cells infected with wild-type bacteria. We conclude that, in addition to its role in host-pathogen interactions, our results reveal that the function of opiA is central to the biology of F. tularensis bacteria. IMPORTANCE F. tularensis is a pathogenic intracellular pathogen that is of importance for public health and strategic defense. This study characterizes the opiA gene of F. tularensis LVS, an attenuated strain that has been used as a live vaccine but that also shares significant genetic similarity to related Francisella strains that cause human disease. The data presented here provide the first evidence of a T6SS effector protein that affects the physiology of F. tularensis, namely, the growth, cell size, viability, and aminoglycoside resistance of F. tularensis LVS. This study also adds insight into our understanding of OpiA as a determinant of virulence. Finally, the fluorescence fusion constructs presented here will be useful tools for dissecting the role of OpiA in infection.

scholarly journals Salmonella enterica Serovar Typhimurium SPI-1 and SPI-2 Shape the Global Transcriptional Landscape in a Human Intestinal Organoid Model System

mBio ◽  
2021 ◽  
Vol 12 (3) ◽  
Author(s):  
Anna-Lisa E. Lawrence ◽  
Basel H. Abuaita ◽  
Ryan P. Berger ◽  
David R. Hill ◽  
Sha Huang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Intestinal Epithelium ◽  
Salmonella Enterica ◽  
Secretion System ◽  
Host Responses ◽  
Secretion Systems ◽  
Content Type ◽  
Serovar Typhimurium ◽  
Type 3

ABSTRACT The intestinal epithelium is a primary interface for engagement of the host response by foodborne pathogens, like Salmonella enterica Typhimurium. While the interaction of S. Typhimurium with the mammalian host has been well studied in transformed epithelial cell lines or in the complex intestinal environment in vivo, few tractable models recapitulate key features of the intestine. Human intestinal organoids (HIOs) contain a polarized epithelium with functionally differentiated cell subtypes, including enterocytes and goblet cells and a supporting mesenchymal cell layer. HIOs contain luminal space that supports bacterial replication, are more amenable to experimental manipulation than animals and are more reflective of physiological host responses. Here, we use the HIO model to define host transcriptional responses to S. Typhimurium infection, also determining host pathways dependent on Salmonella pathogenicity island-1 (SPI-1)- and -2 (SPI-2)-encoded type 3 secretion systems (T3SS). Consistent with prior findings, we find that S. Typhimurium strongly stimulates proinflammatory gene expression. Infection-induced cytokine gene expression was rapid, transient, and largely independent of SPI-1 T3SS-mediated invasion, likely due to continued luminal stimulation. Notably, S. Typhimurium infection led to significant downregulation of host genes associated with cell cycle and DNA repair, leading to a reduction in cellular proliferation, dependent on SPI-1 and SPI-2 T3SS. The transcriptional profile of cell cycle-associated target genes implicates multiple miRNAs as mediators of S. Typhimurium-dependent cell cycle suppression. These findings from Salmonella-infected HIOs delineate common and distinct contributions of SPI-1 and SPI-2 T3SSs in inducing early host responses during enteric infection and reinforce host cell proliferation as a process targeted by Salmonella. IMPORTANCE Salmonella enterica serovar Typhimurium (S. Typhimurium) causes a significant health burden worldwide, yet host responses to initial stages of intestinal infection remain poorly understood. Due to differences in infection outcome between mice and humans, physiological human host responses driven by major virulence determinants of Salmonella have been more challenging to evaluate. Here, we use the three-dimensional human intestinal organoid model to define early responses to infection with wild-type S. Typhimurium and mutants defective in the SPI-1 or SPI-2 type-3 secretion systems. While both secretion system mutants show defects in mouse models of oral Salmonella infection, the specific contributions of each secretion system are less well understood. We show that S. Typhimurium upregulates proinflammatory pathways independently of either secretion system, while the downregulation of the host cell cycle pathways relies on both SPI-1 and SPI-2. These findings lay the groundwork for future studies investigating how SPI-1- and SPI-2-driven host responses affect infection outcome and show the potential of this model to study host-pathogen interactions with other serovars to understand how initial interactions with the intestinal epithelium may affect pathogenesis.

scholarly journals Complete Genome Sequence of Pseudomonas aeruginosa CMC-115, a Clinical Strain from an Acute Ventilator-Associated Pneumonia Patient

2020 ◽  
Vol 9 (30) ◽  
Author(s):  
Adenike Adenikinju ◽  
Roderick V. Jensen ◽  
Thomas M. Kerkering ◽  
Dorothy C. Garner ◽  
Jayasimha Rao
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Complete Genome ◽  
Secretion System ◽  
Clinical Strain ◽  
Ventilator Associated Pneumonia ◽  
Circular Chromosome ◽  
Content Type ◽  
Pneumonia Patient ◽  
Type 3 Secretion System ◽  
Type 3

ABSTRACT We report the complete genome of clinical strain Pseudomonas aeruginosa CMC-115, which was isolated from an acute ventilator-associated pneumonia patient. Illumina sequencing reads were assembled using Geneious to yield a 6,375,262-bp circular chromosome that exhibited an unusual ferrichrome receptor in the pyoverdine synthesis locus and the absence of type 3 secretion system genes.

scholarly journals Response of Pseudomonas aeruginosa to the Innate Immune System-Derived Oxidants Hypochlorous Acid and Hypothiocyanous Acid

2020 ◽  
Vol 203 (2) ◽  
pp. e00300-20
Author(s):  
Katie V. Farrant ◽  
Livia Spiga ◽  
Jane C. Davies ◽  
Huw D. Williams
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Immune System ◽  
Hypochlorous Acid ◽  
Secretion System ◽  
Content Type ◽  
Type 3 Secretion System ◽  
Lung Infections ◽  
Type 3

ABSTRACTPseudomonas aeruginosa is a significant nosocomial pathogen and is associated with lung infections in cystic fibrosis (CF). Once established, P. aeruginosa infections persist and are rarely eradicated despite host immune cells producing antimicrobial oxidants, including hypochlorous acid (HOCl) and hypothiocyanous acid (HOSCN). There is limited knowledge as to how P. aeruginosa senses, responds to, and protects itself against HOCl and HOSCN and the contribution of such responses to its success as a CF pathogen. To investigate the P. aeruginosa response to these oxidants, we screened 707 transposon mutants, with mutations in regulatory genes, for altered growth following HOCl exposure. We identified regulators of antibiotic resistance, methionine biosynthesis, catabolite repression, and PA14_07340, the homologue of the Escherichia coli HOCl-sensor RclR (30% identical), which are required for protection against HOCl. We have shown that RclR (PA14_07340) protects specifically against HOCl and HOSCN stress and responds to both oxidants by upregulating the expression of a putative peroxiredoxin, rclX (PA14_07355). Transcriptional analysis revealed that while there was specificity in the response to HOCl (231 genes upregulated) and HOSCN (105 genes upregulated), there was considerable overlap, with 74 genes upregulated by both oxidants. These included genes encoding the type 3 secretion system, sulfur and taurine transport, and the MexEF-OprN efflux pump. RclR coordinates part of the response to both oxidants, including upregulation of pyocyanin biosynthesis genes, and, in the presence of HOSCN, downregulation of chaperone genes. These data indicate that the P. aeruginosa response to HOCl and HOSCN is multifaceted, with RclR playing an essential role.IMPORTANCE The bacterial pathogen Pseudomonas aeruginosa causes devastating infections in immunocompromised hosts, including chronic lung infections in cystic fibrosis patients. To combat infection, the host’s immune system produces the antimicrobial oxidants hypochlorous acid (HOCl) and hypothiocyanous acid (HOSCN). Little is known about how P. aeruginosa responds to and survives attack from these oxidants. To address this, we carried out two approaches: a mutant screen and transcriptional study. We identified the P. aeruginosa transcriptional regulator, RclR, which responds specifically to HOCl and HOSCN stress and is essential for protection against both oxidants. We uncovered a link between the P. aeruginosa transcriptional response to these oxidants and physiological processes associated with pathogenicity, including antibiotic resistance and the type 3 secretion system.

scholarly journals Tulane Virus Recognizes the A Type 3 and B Histo-Blood Group Antigens

2014 ◽  
Vol 89 (2) ◽  
pp. 1419-1427 ◽  
Author(s):  
Dongsheng Zhang ◽  
Pengwei Huang ◽  
Lu Zou ◽  
Todd L. Lowary ◽  
Ming Tan ◽  
...  
Keyword(s):  
Cell Culture ◽  
Blood Group ◽  
Culture Method ◽  
Blood Group Antigens ◽  
Content Type ◽  
Tulane Virus ◽  
A Cell ◽  
Type 3

ABSTRACTTulane virus (TV), the prototype of theRecovirusgenus in the calicivirus family, was isolated from the stools of rhesus monkeys and can be cultivatedin vitroin monkey kidney cells. TV is genetically closely related to the genusNorovirusand recognizes the histo-blood group antigens (HBGAs), similarly to human noroviruses (NoVs), making it a valuable surrogate for human NoVs. However, the precise structures of HBGAs recognized by TV remain elusive. In this study, we performed binding and blocking experiments on TV with extended HBGA types and showed that, while TV binds all four types (types 1 to 4) of the B antigens, it recognizes only the A type 3 antigen among four types of A antigens tested. The requirements for HBGAs in TV replication were demonstrated by blocking of TV replication in cell culture using the A type 3/4 and B saliva samples. Similar results were also observed in oligosaccharide-based blocking assays. Importantly, the previously reported, unexplained increase in TV replication by oligosaccharide in cell-based blocking assays has been clarified, which will facilitate the application of TV as a surrogate for human NoVs.IMPORTANCEOur understanding of the role of HBGAs in NoV infection has been significantly advanced in the past decade, but direct evidence for HBGAs as receptors for human NoVs remains lacking due to a lack of a cell culture method. TV recognizes HBGAs and can replicatein vitro, providing a valuable surrogate for human NoVs. However, TV binds to some but not all saliva samples from A-positive individuals, and an unexplained observation of synthetic oligosaccharide blocking of TV binding has been reported. These issues have been resolved in this study.

scholarly journals Glutamate Limitation, BvgAS Activation, and (p)ppGpp Regulate the Expression of the Bordetella pertussis Type 3 Secretion System

2015 ◽  
Vol 198 (2) ◽  
pp. 343-351 ◽  
Author(s):  
Tomoko Hanawa ◽  
Kazunari Kamachi ◽  
Hideo Yonezawa ◽  
Toshiyuki Fukutomi ◽  
Hayato Kawakami ◽  
...  
Keyword(s):  
Bordetella Pertussis ◽  
Protein Profile ◽  
Secretion System ◽  
Transcriptional Level ◽  
Content Type ◽  
Bacterial Physiology ◽  
Number Of Patients ◽  
Type 3 Secretion System ◽  
Type 3

ABSTRACTBordetella pertussisis a bacterium that is considered to be highly adapted to humans, and it has not been isolated from the environment. As this bacterium does not utilize sugars, the abundant supply of glutamate in Stainer Scholte (SS) medium enablesB. pertussisto grow efficiently in liquid culturein vitro, and as such, SS medium is a popular choice for laboratory experiments. However, the concentration of glutamate in thein vivoniche ofB. pertussisis quite low. We investigated the bacterial response to low concentrations of glutamate to elucidate bacterial physiology via the expression of the type 3 secretion system (T3SS), and we discuss its relationship to the Bvg mode in which the two-component regulator of pathogenesis (BvgAS) is activated. Glutamate limitation induced the expression of both the T3SS apparatus and effector genes at the transcriptional level. (p)ppGpp, a modulator of the stringent response, was necessary for maximum expression of the T3SS genes. These observations indicate that the expression of the T3SS is managed by nutrient starvation. In addition, the autoaggregation ability was high in the absence of glutamate and no autoaggregation was observed in glutamate-replete medium. Taken together, glutamate-limited conditions in Bvg+mode elicit the high expression of T3SS genes inB. pertussisand promotes its sessile form.IMPORTANCEBordetella pertussisis a highly contagious pathogen that causes respiratory infectious disease. In spite of the increasing use of vaccination, the number of patients with pertussis is increasing. The proteins producedin vivooften are different from the protein profile under laboratory conditions; therefore, the development of conditions reflecting the host environment is important to understand native bacterial behavior. In the present study, we examined the effect of glutamate limitation, as its concentrationin vivois much lower than that in the culture medium currently used forB. pertussisexperiments. As predicted, the T3SS was induced by glutamate limitation. These results are suggestive of the importance of regulation by nutrient conditions and in the pathogenicity ofB. pertussis.

scholarly journals Regulatory Effects of CsrA in Vibrio cholerae

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Heidi A. Butz ◽  
Alexandra R. Mey ◽  
Ashley L. Ciosek ◽  
Alexander A. Crofts ◽  
Bryan W. Davies ◽  
...  
Keyword(s):  
Gene Expression ◽  
Vibrio Cholerae ◽  
Virulence Gene ◽  
Growth Cycle ◽  
Mammalian Host ◽  
Aqueous Environment ◽  
Content Type ◽  
Human Host

ABSTRACT CsrA is a posttranscriptional global regulator in Vibrio cholerae. Although CsrA is critical for V. cholerae survival within the mammalian host, the regulatory targets of CsrA remain mostly unknown. To identify pathways controlled by CsrA, RNA-seq transcriptome analysis was carried out by comparing the wild type and the csrA mutant grown to early exponential, mid-exponential, and stationary phases of growth. This enabled us to identify the global effects of CsrA-mediated regulation throughout the V. cholerae growth cycle. We found that CsrA regulates 22% of the V. cholerae transcriptome, with significant regulation within the gene ontology (GO) processes that involve amino acid transport and metabolism, central carbon metabolism, lipid metabolism, iron uptake, and flagellum-dependent motility. Through CsrA-RNA coimmunoprecipitation experiments, we found that CsrA binds to multiple mRNAs that encode regulatory proteins. These include transcripts encoding the major sigma factors RpoS and RpoE, which may explain how CsrA regulation affects such a large proportion of the V. cholerae transcriptome. Other direct targets include flrC, encoding a central regulator in flagellar gene expression, and aphA, encoding the virulence gene transcription factor AphA. We found that CsrA binds to the aphA mRNA both in vivo and in vitro, and CsrA significantly increases AphA protein synthesis. The increase in AphA was due to increased translation, not transcription, in the presence of CsrA, consistent with CsrA binding to the aphA transcript and enhancing its translation. CsrA is required for the virulence of V. cholerae and this study illustrates the central role of CsrA in virulence gene regulation. IMPORTANCE Vibrio cholerae, a Gram-negative bacterium, is a natural inhabitant of the aqueous environment. However, once ingested, this bacterium can colonize the human host and cause the disease cholera. In order to successfully transition between its aqueous habitat and the human host, the bacterium must sense changes in its environment and rapidly alter gene expression. Global regulators, including CsrA, play an integral role in altering the expression of a large number of genes to promote adaptation and survival, which is required for intestinal colonization. We used transcriptomics and a directed CsrA-RNA coimmunoprecipitation to characterize the CsrA regulon and found that CsrA alters the expression of more than 800 transcripts in V. cholerae. Processes regulated by CsrA include motility, the rugose phenotype, and virulence pathways. CsrA directly binds to the aphA transcript and positively regulates the production of the virulence regulator AphA. Thus, CsrA regulates multiple processes that have been linked to pathogenesis.

Sign in / Sign up

Export Citation Format

Share Document