scholarly journals Transcriptome and Comparative Genomics Analyses Reveal New Functional Insights on Key Determinants of Pathogenesis and Interbacterial Competition inPectobacteriumandDickeyaspp.

2018 ◽  
Vol 85 (2) ◽  
Author(s):  
Daniel Bellieny-Rabelo ◽  
Collins K. Tanui ◽  
Nikki Miguel ◽  
Stanford Kwenda ◽  
Divine Y. Shyntum ◽  
...  
Keyword(s):  
Comparative Genomics ◽  
Plant Pathogens ◽  
Ectopic Expression ◽  
Soft Rot ◽  
Secretion System ◽  
Specific Gene ◽  
Type I ◽  
Genomic Information ◽  
Type Vi Secretion System ◽  
Content Type

ABSTRACTSoft-rotEnterobacteriaceae(SRE), typified byPectobacteriumandDickeyagenera, are phytopathogenic bacteria inflicting soft-rot disease in crops worldwide. By combining genomic information from 100 SRE with whole-transcriptome data sets, we identified novel genomic and transcriptional associations among key pathogenicity themes in this group. Comparative genomics revealed solid linkage between the type I secretion system (T1SS) and the carotovoricin bacteriophage (Ctv) conserved in 96.7% ofPectobacteriumgenomes. Moreover, their coactivation during infection indicates a novel functional association involving T1SS and Ctv. Another bacteriophage-borne genomic region, mostly confined to less than 10% ofPectobacteriumstrains, was found, presumably comprising a novel lineage-specific prophage in the genus. We also detected the transcriptional coregulation of a previously predicted toxin/immunity pair (WHH and SMI1_KNR4 families), along with the type VI secretion system (T6SS), which includeshcpand/orvgrGgenes, suggesting a role in disease development as T6SS-dependent effectors. Further, we showed that another predicted T6SS-dependent endonuclease (AHH family) exhibited toxicity in ectopic expression assays, indicating antibacterial activity. Additionally, we report the striking conservation of the group 4 capsule (GFC) cluster in 100 SRE strains which consistently features adjacently conserved serotype-specific gene arrays comprising a previously unknown organization in GFC clusters. Also, extensive sequence variations found ingfcAorthologs suggest a serotype-specific role in the GfcABCD machinery.IMPORTANCEDespite the considerable loss inflicted on important crops yearly byPectobacteriumandDickeyadiseases, investigations on key virulence and interbacterial competition assets relying on extensive comparative genomics are still surprisingly lacking for these genera. Such approaches become more powerful over time, underpinned by the growing amount of genomic information in public databases. In particular, our findings point to new functional associations among well-known genomic themes enabling alternative means of neutralizing SRE diseases through disruption of pivotal virulence programs. By elucidating novel transcriptional and genomic associations, this study adds valuable information on virulence candidates that could be decisive in molecular applications in the near future. The utilization of 100 genomes ofPectobacteriumandDickeyastrains in this study is unprecedented for comparative analyses in these taxa, and it provides novel insights on the biology of economically important plant pathogens.

scholarly journals Transcriptome and Comparative Genomics analyses reveal new functional insights on key determinants of pathogenesis and interbacterial competition inPectobacteriumandDickeyaspp

2018 ◽  
Author(s):  
Daniel Bellieny-Rabelo ◽  
Collins K. Tanui ◽  
Nikki Miguel ◽  
Stanford Kwenda ◽  
Divine Y. Shyntum ◽  
...  
Keyword(s):  
Comparative Genomics ◽  
Ectopic Expression ◽  
Soft Rot ◽  
Genomic Region ◽  
Specific Gene ◽  
Type Vi Secretion System ◽  
Group 4 ◽  
Dependent Endonuclease ◽  
Sequence Variations ◽  
Rot Disease

SummarySoft-rotEnterobacteriaceae(SRE) typified byPectobacteriumandDickeyagenera are phytopathogenic bacteria inflicting soft-rot disease in crops worldwide. By combining genomic information from 100 SRE with whole-transcriptome datasets, we identified novel genomic and transcriptional associations amongst key pathogenicity themes in this group. Comparative genomics revealed solid linkage between thetypeIsecretionsystem (T1SS) and the carotovoricin bacteriophage (Ctv) conserved in 96.7% ofPectobacteriumgenomes. Moreover, their co-activation during infection might indicate a novel functional association involving T1SS/Ctv. Another bacteriophage-borne genomic region mostly confined in less than 10% ofPectobacteriumorganisms was found, presumably comprising a novel lineage-specific prophage in the genus. We also detected the transcriptional co-regulation of a previously predicted toxin/immunity pair (WHH and SMI1_KNR4 families) along with type VI secretion system (T6SS) andhcp/vgrGgenes suggesting a role in disease development as T6SS-dependent effectors. Further, we showed that another predicted T6SS-dependent endonuclease (AHH-family) exhibited toxicity in ectopic expression assays indicating antibacterial activity. Additionally, we report the striking conservation of group-4-capsule (GFC) cluster in 100 SRE strains which consistently features adjacently conserved serotype-specific gene-arrays comprising a previously unknown organization in GFC clusters. Also, extensive sequence variations found ingfcAorthologs suggest a serotype-specific role in the GfcABCD machinery.

scholarly journals The impact of type VI secretion system, bacteriocins and antibiotics on competition amongst Soft-RotEnterobacteriaceae: Regulation of carbapenem biosynthesis by iron and the transcriptional regulator Fur

2018 ◽  
Author(s):  
Divine Yutefar Shyntum ◽  
Ntombikayise Nkomo ◽  
Alessandro Rino Gricia ◽  
Ntwanano Luann Shigange ◽  
Daniel Bellieny-Rabelo ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Plant Pathogens ◽  
Soft Rot ◽  
Secretion System ◽  
Economic Losses ◽  
Computational Techniques ◽  
In Planta ◽  
Type Vi Secretion System ◽  
Type Vi Secretion

AbstractPlant microbial communities’ complexity provide a rich model for investigation on biochemical and regulatory strategies involved in interbacterial competition. Within these niches, the soft rotEnterobacteriaceae(SRE) comprise an emerging group of plant-pathogens inflicting soft rot/black-leg diseases and causing economic losses worldwide in a variety of crops. In this report, a range of molecular and computational techniques are utilized to survey the contribution of antimicrobial factors such as bacteriocins, carbapenem antibiotic and type VI secretion system (T6SS) in interbacterial competition among plant-pathogens/endophytes using an aggressive SRE as a case study (Pectobacterium carotovorumsubsp.brasiliensestrain PBR1692 –Pcb1692). A preliminary screening using next-generation sequencing of 16S rRNA comparatively analysing healthy and diseased potato tubers, followed byin vitrocompetition assays, corroborated the aggressiveness ofPcb1692 against several relevant taxa sharing this niche ranging from Proteobacteria toFirmicutes. The results showed growth inhibition of several Proteobacteria by Pcb1692 depends either on carbapenem or pyocin production. Whereas for targetedFirmicutes, only pyocin seems to play a role in growth inhibition byPcb1692. Further analyses elucidated that although T6SS confers no relevant advantage duringin vitrocompetition, a significant attenuation in competition by the mutant strain lacking a functional T6SS was observedin planta. Furthermore, production of carbapenem byPcb1692 was observably dependent on the presence of environmental iron and oxygen. Additionally, upon deletion offur, slyA andexpI regulators, carbapenem production ceased, implying a complex regulatory mechanism involving these three genes. Potential Fur binding sites found upstream ofslyA,carR andexpR inPectobacteriumgenomes harboring carbapenem-associated genes further suggests a conserved regulatory pattern in the genus, in which carbapenem might be modulated in response to iron through the control exerted by Fur over secondary regulators. Furthermore, we unveiled the striking role played by S-pyocin in growth inhibition within the SRE group.Authors SummaryFor many phytopathogenic bacteria, more is known about interactions within the host and virulence factors used for host colonisation while relatively less is known about microbe-microbe interactions and factors that shape niche colonisation. The soft rotEnterobacteriaceae(SRE) comprise an emerging group of phytopathogens causing soft rot/black-leg diseases in a variety of crops leading to huge economic losses worldwide. In this report, a range of molecular and computational techniques are utilized to survey the contribution of antimicrobial factors such as bacteriocins, carbapenem antibiotic and type VI secretion system (T6SS) in interbacterial competition among plant-pathogens/endophytes using an aggressive SRE as a case study (Pcb1692). Our results show thatPcb1692 inhibits growth of other SRE and several potato endophytes using either the type VI secretion, carbapenem or bacteriocins. Carbapenem plays a role in both inter and intrabacterial competitionin vitro, while thePcb1692T6SS plays a role in interbacterial competitionin planta(in potato tubers). We also demonstrate that carbapenem regulation requires the presence of environmental iron and oxygen in a complex network consisting ofPcb1692 Fur, SlyA, and ExpI. The presence of these gene homologs in several SREs suggests that they too can deploy similar antimicrobials to target other bacteria.

scholarly journals Draft Genome Sequences of Type VI Secretion System-Encoding Vibrio fischeri Strains FQ-A001 and ES401

2019 ◽  
Vol 8 (20) ◽  
Author(s):  
Katherine M. Bultman ◽  
Andrew G. Cecere ◽  
Tim Miyashiro ◽  
Alecia N. Septer ◽  
Mark J. Mandel
Keyword(s):  
Comparative Genomics ◽  
Direct Contact ◽  
Vibrio Fischeri ◽  
Draft Genome ◽  
Secretion System ◽  
Euprymna Scolopes ◽  
Type Vi Secretion System ◽  
Genome Sequences ◽  
Content Type ◽  
Type Vi Secretion

The type VI secretion system (T6SS) facilitates lethal competition between bacteria through direct contact. Comparative genomics has facilitated the study of these systems in Vibrio fischeri, which colonizes the squid host Euprymna scolopes.

Pangenomic Analysis of Dickeya dianthicola Strains Related to the Outbreak of Blackleg and Soft Rot of Potato in USA

Plant Disease ◽  
2021 ◽  
Author(s):  
Tongling Ge ◽  
He Jiang ◽  
Ek Han Tan ◽  
Steven B Johnson ◽  
Robert Larkin ◽  
...  
Keyword(s):  
Plant Cell Wall ◽  
Gene Clusters ◽  
Soft Rot ◽  
Secretion System ◽  
Phylogenomic Analysis ◽  
Type I ◽  
Type Vi Secretion System ◽  
Secretion Systems ◽  
Type Iv ◽  
The Usa

Dickeya dianthicola has caused an outbreak of blackleg and soft rot of potato in the eastern half of the USA since 2015. To investigate genetic diversity of the pathogen, a comparative analysis was conducted on genomes of D. dianthicola strains. Whole genomes of 16 strains from the USA outbreak were assembled and compared to 16 previously sequenced genomes of D. dianthicola isolated from potato or carnation. Among the 32 strains, eight distinct clades were distinguished based on phylogenomic analysis. The outbreak strains were grouped into three clades, with the majority of the strains in clade I. Clade I strains were unique and homogeneous, suggesting a recent incursion of this strain into potato production from alternative hosts or environmental sources. Pangenome of the 32 strains contained 6693 genes, 3377 of which were core genes. By screening primary protein subunits associated with virulence from all USA strains, we found many virulence-related gene clusters, such as plant cell wall degrading enzyme genes, flagellar and chemotaxis related genes, two-component regulatory genes, and type I/II/III secretion system genes were highly conserved but type IV and type VI secretion system genes varied. The virulent clade I strains encoded two clusters of type IV secretion systems, while clade II and III strains encoded only one cluster. Clade I and II strains encoded one more VgrG/PAAR spike protein than clade III. Thus, we predicted that the presence of additional virulence-related genes may have enabled the unique clade I strain to become predominant source in the USA outbreak.

scholarly journals Virulence genes and previously unexplored gene clusters in four commensal Neisseria spp. isolated from the human throat expand the neisserial gene repertoire

2020 ◽  
Vol 6 (9) ◽  
Author(s):  
Alan Calder ◽  
Chukwuma Jude Menkiti ◽  
Aylin Çağdaş ◽  
Jefferson Lisboa Santos ◽  
Ricarda Streich ◽  
...  
Keyword(s):  
Type Species ◽  
Gene Clusters ◽  
Secretion System ◽  
Specific Gene ◽  
Gene Repertoire ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Link Type ◽  
Type Iv ◽  
System A

Commensal non-pathogenic Neisseria spp. live within the human host alongside the pathogenic Neisseria meningitidis and Neisseria gonorrhoeae and due to natural competence, horizontal gene transfer within the genus is possible and has been observed. Four distinct Neisseria spp. isolates taken from the throats of two human volunteers have been assessed here using a combination of microbiological and bioinformatics techniques. Three of the isolates have been identified as Neisseria subflava biovar perflava and one as Neisseria cinerea . Specific gene clusters have been identified within these commensal isolate genome sequences that are believed to encode a Type VI Secretion System, a newly identified CRISPR system, a Type IV Secretion System unlike that in other Neisseria spp., a hemin transporter, and a haem acquisition and utilization system. This investigation is the first to investigate these systems in either the non-pathogenic or pathogenic Neisseria spp. In addition, the N. subflava biovar perflava possess previously unreported capsule loci and sequences have been identified in all four isolates that are similar to genes seen within the pathogens that are associated with virulence. These data from the four commensal isolates provide further evidence for a Neisseria spp. gene pool and highlight the presence of systems within the commensals with functions still to be explored.

scholarly journals The Type VI Secretion System Modulates Flagellar Gene Expression and Secretion in Citrobacter freundii and Contributes to Adhesion and Cytotoxicity to Host Cells

2015 ◽  
Vol 83 (7) ◽  
pp. 2596-2604 ◽  
Author(s):  
Liyun Liu ◽  
Shuai Hao ◽  
Ruiting Lan ◽  
Guangxia Wang ◽  
Di Xiao ◽  
...  
Keyword(s):  
Secretion System ◽  
Host Cells ◽  
Target Cells ◽  
Deletion Mutants ◽  
Citrobacter Freundii ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Type Vi Secretion ◽  
Mutant Strains

The type VI secretion system (T6SS) as a virulence factor-releasing system contributes to virulence development of various pathogens and is often activated upon contact with target cells.Citrobacter freundiistrain CF74 has a complete T6SS genomic island (GI) that containsclpV,hcp-2, andvgrT6SS genes. We constructedclpV,hcp-2,vgr, and T6SS GI deletion mutants in CF74 and analyzed their effects on the transcriptome overall and, specifically, on the flagellar system at the levels of transcription and translation. Deletion of the T6SS GI affected the transcription of 84 genes, with 15 and 69 genes exhibiting higher and lower levels of transcription, respectively. Members of the cell motility class of downregulated genes of the CF74ΔT6SS mutant were mainly flagellar genes, including effector proteins, chaperones, and regulators. Moreover, the production and secretion of FliC were also decreased inclpV,hcp-2,vgr, or T6SS GI deletion mutants in CF74 and were restored upon complementation. In swimming motility assays, the mutant strains were found to be less motile than the wild type, and motility was restored by complementation. The mutant strains were defective in adhesion to HEp-2 cells and were restored partially upon complementation. Further, the CF74ΔT6SS, CF74ΔclpV, and CF74Δhcp-2mutants induced lower cytotoxicity to HEp-2 cells than the wild type. These results suggested that the T6SS GI in CF74 regulates the flagellar system, enhances motility, is involved in adherence to host cells, and induces cytotoxicity to host cells. Thus, the T6SS plays a wide-ranging role inC. freundii.

scholarly journals Fur-Dam Regulatory Interplay at an Internal Promoter of the Enteroaggregative Escherichia coli Type VI Secretion sci1 Gene Cluster

2020 ◽  
Vol 202 (10) ◽  
Author(s):  
Yannick R. Brunet ◽  
Christophe S. Bernard ◽  
Eric Cascales
Keyword(s):  
Escherichia Coli ◽  
Gene Cluster ◽  
Gene Clusters ◽  
Secretion System ◽  
Target Cells ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Internal Promoter ◽  
E Coli ◽  
Type Vi Secretion

ABSTRACT The type VI secretion system (T6SS) is a weapon for delivering effectors into target cells that is widespread in Gram-negative bacteria. The T6SS is a highly versatile machine, as it can target both eukaryotic and prokaryotic cells, and it has been proposed that T6SSs are adapted to the specific needs of each bacterium. The expression of T6SS gene clusters and the activation of the secretion apparatus are therefore tightly controlled. In enteroaggregative Escherichia coli (EAEC), the sci1 T6SS gene cluster is subject to a complex regulation involving both the ferric uptake regulator (Fur) and DNA adenine methylase (Dam)-dependent DNA methylation. In this study, an additional, internal, promoter was identified within the sci1 gene cluster using +1 transcriptional mapping. Further analyses demonstrated that this internal promoter is controlled by a mechanism strictly identical to that of the main promoter. The Fur binding box overlaps the −10 transcriptional element and a Dam methylation site, GATC-32. Hence, the expression of the distal sci1 genes is repressed and the GATC-32 site is protected from methylation in iron-rich conditions. The Fur-dependent protection of GATC-32 was confirmed by an in vitro methylation assay. In addition, the methylation of GATC-32 negatively impacted Fur binding. The expression of the sci1 internal promoter is therefore controlled by iron availability through Fur regulation, whereas Dam-dependent methylation maintains a stable ON expression in iron-limited conditions. IMPORTANCE Bacteria use weapons to deliver effectors into target cells. One of these weapons, the type VI secretion system (T6SS), assembles a contractile tail acting as a spring to propel a toxin-loaded needle. Its expression and activation therefore need to be tightly regulated. Here, we identified an internal promoter within the sci1 T6SS gene cluster in enteroaggregative E. coli. We show that this internal promoter is controlled by Fur and Dam-dependent methylation. We further demonstrate that Fur and Dam compete at the −10 transcriptional element to finely tune the expression of T6SS genes. We propose that this elegant regulatory mechanism allows the optimum production of the T6SS in conditions where enteroaggregative E. coli encounters competing species.

scholarly journals Comparative Genomics and Phylogenetic Analyses Suggest Several Novel Species within the Genus Clavibacter, Including Nonpathogenic Tomato-Associated Strains

2020 ◽  
Vol 86 (6) ◽  
Author(s):  
Ebrahim Osdaghi ◽  
Touraj Rahimi ◽  
S. Mohsen Taghavi ◽  
Maryam Ansari ◽  
Sadegh Zarei ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Comparative Genomics ◽  
Plant Pathogens ◽  
Phylogenetic Analyses ◽  
Taxonomic Status ◽  
Sensu Stricto ◽  
Bacterial Canker ◽  
Agricultural Crops ◽  
Content Type ◽  
Canker Disease

ABSTRACT Members of the genus Clavibacter are economically important bacterial plant pathogens infecting a set of diverse agricultural crops (e.g., alfalfa, corn, potato, tomato, and wheat). Tomato-associated Clavibacter sp. strains account for a great portion of the genetic diversity of the genus, and C. michiganensis sensu stricto (formerly C. michiganensis subsp. michiganensis), causing bacterial canker disease, is considered one of the most destructive seed-borne agents for the crop worldwide. However, current taxonomic descriptions of the genus do not reflect the existing diversity of the strains, resulting in unsatisfactory results in quarantine surveys for the pathogens. In this study, we used all the available genome sequences of Clavibacter sp. strains, including the type strains of newly described subspecies, to provide precise insight into the diversity of tomato-associated members of the genus and further clarify the taxonomic status of the strains using genotypic and phenotypic features. The results of phylogenetic analyses revealed the existence of nine hypothetical new species among the investigated strains. None of the three new subspecies (i.e., C. michiganensis subsp. californiensis, C. michiganensis subsp. chilensis, and C. michiganensis subsp. phaseoli) is included within the tomato-pathogenic C. michiganensis sensu stricto lineage. Although comparative genomics revealed the lack of chp and tomA pathogenicity determinant gene clusters in the nonpathogenic strains, a number of pathogenicity-related genes were noted to be present in all the strains regardless of their pathogenicity characteristics. Altogether, our results indicate a need for a formal taxonomic reconsideration of tomato-associated Clavibacter sp. strains to facilitate differentiation of the lineages in quarantine inspections. IMPORTANCE Clavibacter spp. are economically important bacterial plant pathogens infecting a set of diverse agricultural crops, such as alfalfa, corn, pepper, potato, tomato, and wheat. A number of plant-pathogenic members of the genus (e.g., C. michiganensis sensu stricto and C. sepedonicus, infecting tomato and potato plants, respectively) are included in the A2 (high-risk) list of quarantine pathogens by the European and Mediterranean Plant Protection Organization (EPPO). Although tomato-associated members of Clavibacter spp. account for a significant portion of the genetic diversity in the genus, only the strains belonging to C. michiganensis sensu stricto (formerly C. michiganensis subsp. michiganensis) cause bacterial canker disease of tomato and are subjected to the quarantine inspections. Hence, discrimination between the pathogenic and nonpathogenic Clavibacter sp. strains associated with tomato seeds and transplants plays a pivotal role in the accurate detection and cost-efficient management of the disease. On the other hand, detailed information on the genetic contents of different lineages of the genus would lead to the development of genome-informed specific detection techniques. In this study, we have provided an overview of the phylogenetic and genomic differences between the pathogenic and nonpathogenic tomato-associated Clavibacter sp. strains. We also noted that the taxonomic status of newly introduced subspecies of C. michiganensis (i.e., C. michiganensis subsp. californiensis, C. michiganensis subsp. chilensis, and C. michiganensis subsp. phaseoli) should be reconsidered.

scholarly journals Type VI Secretion System Dynamics Reveals a Novel Secretion Mechanism in Pseudomonas aeruginosa

2018 ◽  
Vol 200 (11) ◽  
Author(s):  
Jacqueline Corbitt ◽  
Jun Seok Yeo ◽  
C. Ian Davis ◽  
Michele LeRoux ◽  
Paul A. Wiggins
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Vibrio Cholerae ◽  
Conformational Change ◽  
Secretion System ◽  
Force Generation ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Evolutionary Diversification ◽  
Type Vi Secretion ◽  
Generation Mechanisms

ABSTRACT The type VI secretion system (T6SS) inhibits the growth of neighboring bacterial cells through a contact-mediated mechanism. Here, we describe a detailed characterization of the protein localization dynamics in the Pseudomonas aeruginosa T6SS. It has been proposed that the type VI secretion process is driven by a conformational-change-induced contraction of the T6SS sheath. However, although the contraction of an optically resolvable TssBC sheath and the subsequent localization of ClpV are observed in Vibrio cholerae , coordinated assembly and disassembly of TssB and ClpV are observed without TssB contraction in P. aeruginosa . These dynamics are inconsistent with the proposed contraction sheath model. Motivated by the phenomenon of dynamic instability, we propose a new model in which ATP hydrolysis, rather than conformational change, generates the force for secretion. IMPORTANCE The type VI secretion system (T6SS) is widely conserved among Gram-negative bacteria and is a central determinant of bacterial fitness in polymicrobial communities. The secretion system targets bacteria and secretes effectors that inhibit the growth of neighboring cells, using a contact-mediated-delivery system. Despite significant homology to the previously characterized Vibrio cholerae T6SS, our analysis reveals that effector secretion is driven by a distinct force generation mechanism in Pseudomonas aeruginosa . The presence of two distinct force generation mechanisms in T6SS represents an example of the evolutionary diversification of force generation mechanisms.

scholarly journals Vibrio cholerae Requires the Type VI Secretion System Virulence Factor VasX To Kill Dictyostelium discoideum

2011 ◽  
Vol 79 (7) ◽  
pp. 2941-2949 ◽  
Author(s):  
Sarah T. Miyata ◽  
Maya Kitaoka ◽  
Teresa M. Brooks ◽  
Steven B. McAuley ◽  
Stefan Pukatzki
Keyword(s):  
Vibrio Cholerae ◽  
Dictyostelium Discoideum ◽  
Virulence Factor ◽  
Membrane Lipids ◽  
Gene Clusters ◽  
Secretion System ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Enzymatic Function ◽  
Type Vi Secretion

ABSTRACTThe type VI secretion system (T6SS) is recognized as an important virulence mechanism in several Gram-negative pathogens. InVibrio cholerae, the causative agent of the diarrheal disease cholera, a minimum of three gene clusters—one main cluster and two auxiliary clusters—are required to form a functional T6SS apparatus capable of conferring virulence toward eukaryotic and prokaryotic hosts. Despite an increasing understanding of the components that make up the T6SS apparatus, little is known about the regulation of these genes and the gene products delivered by this nanomachine. VasH is an important regulator of theV. choleraeT6SS. Here, we present evidence that VasH regulates the production of a newly identified protein, VasX, which in turn requires a functional T6SS for secretion. Deletion ofvasXdoes not affect export or enzymatic function of the structural T6SS proteins Hcp and VgrG-1, suggesting that VasX is dispensable for the assembly of the physical translocon complex. VasX localizes to the bacterial membrane and interacts with membrane lipids. We present VasX as a novel virulence factor of the T6SS, as aV. choleraemutant lackingvasXexhibits a phenotype of attenuated virulence towardDictyostelium discoideum.

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