A mutation of a single core gene, tssM, of Type VI secretion system of Xanthomonas perforans influences virulence, epiphytic survival and transmission during pathogenesis on tomato

2021 ◽  
Author(s):  
Prabha Liyanapathiranage ◽  
Jeffrey B Jones ◽  
Neha Potnis
Keyword(s):  
Disease Severity ◽  
Core Gene ◽  
Secretion System ◽  
Wild Type ◽  
In Planta ◽  
Type Vi Secretion System ◽  
Biotrophic Pathogen ◽  
Infection Period ◽  
Xanthomonas Perforans ◽  
Type Vi Secretion

Xanthomonas perforans is a seed-borne hemi-biotrophic pathogen that successfully establishes infection in the phyllosphere of tomato. While the majority of the studies investigating mechanistic basis of pathogenesis have focused on successful apoplastic growth, factors important during asymptomatic colonization in the early stages of disease development are not well understood. In this study, we show that tssM gene of the type VI secretion system cluster i3* (T6SS-i3*) plays a significant role during initial asymptomatic epiphytic colonization at different stages during the life cycle of the pathogen. Mutation in a core gene, tssM of T6SS-i3*, imparted higher aggressiveness to the pathogen, as indicated by higher overall disease severity, higher in planta growth as well as shorter latent infection period compared to the wild-type upon dip-inoculation of 4-5-week-old tomato plants. Contribution of tssM towards aggressiveness was evident during vertical transmission from seed-to-seedling with wild-type showing reduced disease severity as well as lower in planta populations on seedlings compared to the mutant. Presence of functional TssM offered higher epiphytic fitness as well as higher dissemination potential to the pathogen when tested in an experimental setup mimicking transplant house high-humidity conditions. We showed higher osmotolerance being one mechanism by which TssM offers higher epiphytic fitness. Taken together, these data reveal that functional TssM plays a larger role in offering ecological advantage to the pathogen. TssM prolongs the association of hemi-biotrophic pathogen with the host, minimizing overall disease severity, yet facilitating successful dissemination.

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 VgrG-dependent effectors and chaperones modulate the assembly of the type VI secretion system

2021 ◽  
Vol 17 (12) ◽  
pp. e1010116
Author(s):  
Xiaoye Liang ◽  
Tong-Tong Pei ◽  
Hao Li ◽  
Hao-Yu Zheng ◽  
Han Luo ◽  
...  
Keyword(s):  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Structural Proteins ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Double Deletion ◽  
Type Vi Secretion ◽  
Structural Determinant

The type VI secretion system (T6SS) is a spear-like nanomachine found in gram-negative pathogens for delivery of toxic effectors to neighboring bacterial and host cells. Its assembly requires a tip spike complex consisting of a VgrG-trimer, a PAAR protein, and the interacting effectors. However, how the spike controls T6SS assembly remains elusive. Here we investigated the role of three VgrG-effector pairs in Aeromonas dhakensis strain SSU, a clinical isolate with a constitutively active T6SS. By swapping VgrG tail sequences, we demonstrate that the C-terminal ~30 amino-acid tail dictates effector specificity. Double deletion of vgrG1&2 genes (VgrG3+) abolished T6SS secretion, which can be rescued by ectopically expressing chimeric VgrG3 with a VgrG1/2-tail but not the wild type VgrG3. In addition, deletion of effector-specific chaperones also severely impaired T6SS secretion, despite the presence of intact VgrG and effector proteins, in both SSU and Vibrio cholerae V52. We further show that SSU could deliver a V. cholerae effector VasX when expressing a plasmid-borne chimeric VgrG with VasX-specific VgrG tail and chaperone sequences. Pull-down analyses show that two SSU effectors, TseP and TseC, could interact with their cognate VgrGs, the baseplate protein TssK, and the key assembly chaperone TssA. Effectors TseL and VasX could interact with TssF, TssK and TssA in V. cholerae. Collectively, we demonstrate that chimeric VgrG-effector pairs could bypass the requirement of heterologous VgrG complex and propose that effector-stuffing inside the baseplate complex, facilitated by chaperones and the interaction with structural proteins, serves as a crucial structural determinant for T6SS assembly.

scholarly journals Identification of Escherichia coli ClpAP in regulating susceptibility to type VI secretion system-mediated attack by Agrobacterium tumefaciens

2019 ◽  
Author(s):  
Hsiao-Han Lin (林筱涵) ◽  
Manda Yu (余文廸) ◽  
Manoj Kumar Sriramoju ◽  
Shang-Te Danny Hsu (徐尚德) ◽  
Chi-Te Liu (劉啟德) ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Agrobacterium Tumefaciens ◽  
Secretion System ◽  
Gram Negative Bacteria ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Competition Assay ◽  
E Coli ◽  
Type Vi Secretion ◽  
In Trans

AbstractType VI secretion system (T6SS) is an effector delivery system used by gram-negative bacteria to kill other bacteria or eukaryotic host to gain fitness. In Agrobacterium tumefaciens, T6SS has been shown to kill other bacteria such as Escherichia coli. Interestingly, the A. tumefaciens T6SS killing efficiency differs when using different E. coli strains as recipient cells. Thus, we hypothesize that a successful T6SS killing not only relies on attacker T6SS activity but also depends on recipient factors. A high-throughput interbacterial competition assay was employed to test the hypothesis by screening for mutants with reduced killing outcomes caused by A. tumefaciens strain C58. From the 3909 E. coli Keio mutants screened, 16 candidate mutants were filtered out. One strain, ΔclpP::Kan, showed ten times more resistant to T6SS-mediating killing but restored its susceptibility when complemented with clpP in trans. ClpP is a universal and highly conserved protease that exists in both prokaryotes and eukaryotic organelles. In E. coli, ClpP uses either ClpA or ClpX as an adaptor for substrate specificity. Therefore, the susceptibility of the ΔclpA::Kan and ΔclpX::Kan was also tested. The T6SS attack susceptibility of ΔclpA::Kan is at the same level as ΔclpP::Kan, while ΔclpX::Kan showed no difference as compared to that of wild-type E. coli BW25113. The data also suggest that ClpA-ClpP interaction, rather than its protease activity, is responsible for enhancing susceptibility to T6SS killing. This study highlights the importance of recipient factors in determining the outcome of T6SS killing.

scholarly journals Pathoadaptive Conditional Regulation of the Type VI Secretion System in Vibrio cholerae O1 Strains

2011 ◽  
Vol 80 (2) ◽  
pp. 575-584 ◽  
Author(s):  
Takahiko Ishikawa ◽  
Dharmesh Sabharwal ◽  
Jeanette Bröms ◽  
Debra L. Milton ◽  
Anders Sjöstedt ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Secretion System ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Content Type ◽  
High Osmolarity ◽  
Secretion Pathway ◽  
Type Vi Secretion ◽  
Vibrio Cholerae O1 ◽  
K 12

ABSTRACTThe most recently discovered secretion pathway in Gram-negative bacteria, the type VI secretion system (T6SS), is present in many species and is considered important for the survival of non-O1 non-O139Vibrio choleraein aquatic environments. Until now, it was not known whether there is a functionally active T6SS in wild-typeV. choleraeO1 strains, the cause of cholera disease in humans. Here, we demonstrate the presence of a functionally active T6SS in wild-typeV. choleraeO1 strains, as evidenced by the secretion of the T6SS substrate Hcp, which required several gene products encoded within the putativevasgene cluster. Our analyses showed that the T6SS of wild-typeV. choleraeO1 strain A1552 was functionally activated when the bacteria were grown under high-osmolarity conditions. The T6SS was also active when the bacteria were grown under low temperature (23°C), suggesting that the system may be important for the survival of the bacterium in the environment. A test of the interbacterial virulence ofV. choleraestrain A1552 against anEscherichia coliK-12 strain showed that it was strongly enhanced under high osmolarity and that it depended on thehcpgenes. Interestingly, we found that the newly recognized osmoregulatory protein OscR plays a role in the regulation of T6SS gene expression and secretion of Hcp fromV. choleraeO1 strains.

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 Functional Exploration of the Bacterial Type VI Secretion System in Mutualism: Azorhizobium caulinodans ORS571–Sesbania rostrata as a Research Model

2018 ◽  
Vol 31 (8) ◽  
pp. 856-867 ◽  
Author(s):  
Hsiao-Han Lin ◽  
Hsin-Mei Huang ◽  
Manda Yu ◽  
Erh-Min Lai ◽  
Hsiao-Lin Chien ◽  
...  
Keyword(s):  
Culture Conditions ◽  
Secretion System ◽  
Sesbania Rostrata ◽  
Research Model ◽  
Azorhizobium Caulinodans ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Type Vi Secretion ◽  
Azorhizobium Caulinodans Ors571 ◽  
Bacterial Type

The bacterial type VI secretion system (T6SS) has been considered the armed force of bacteria because it can deliver toxin effectors to prokaryotic or eukaryotic cells for survival and fitness. Although many legume symbiotic rhizobacteria encode T6SS in their genome, the biological function of T6SS in these bacteria is still unclear. To elucidate this issue, we used Azorhizobium caulinodans ORS571 and its symbiotic host Sesbania rostrata as our research model. By using T6SS gene deletion mutants, we found that T6SS provides A. caulinodans with better symbiotic competitiveness when coinfected with a T6SS-lacking strain, as demonstrated by two independent T6SS-deficient mutants. Meanwhile, the symbiotic effectiveness was not affected by T6SS because the nodule phenotype, nodule size, and nodule nitrogen-fixation ability did not differ between the T6SS mutants and the wild type when infected alone. Our data also suggest that under several lab culture conditions tested, A. caulinodans showed no T6SS-dependent interbacterial competition activity. Therefore, instead of being an antihost or antibacterial weapon of the bacterium, the T6SS in A. caulinodans ORS571 seems to participate specifically in symbiosis by increasing its symbiotic competitiveness.

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