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1
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Liu X, Cao Y, Hu X, Lv A. Rahnella aquatilis VgrG-mediated PANoptosis in macrophages of Carassius auratus by dual RNA-seq analysis. FISH & SHELLFISH IMMUNOLOGY 2025; 158:110155. [PMID: 39864564 DOI: 10.1016/j.fsi.2025.110155] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/25/2024] [Revised: 01/18/2025] [Accepted: 01/21/2025] [Indexed: 01/28/2025]
Abstract
Rahnella aquatilis is an emerging opportunistic pathogen that usually causes septicemia in fish and poses a potential threat to human health. VgrG gene is an important virulence factor of type VI secretion system in R. aquatilis, but its regulatory mechanism underlying PANoptosis is still unknown. Here, VgrG deletion mutant strain of R. aquatilis (ΔVgrG-RA) and recombinant plasmid pET32a-VgrG were respectively constructed, and immunohistochemistry for VgrG as well as PANoptosis features were evaluated. Moreover, the interaction transcriptome of ΔVgrG-mediated pathogen and host was determined by dual RNA-seq using an in vitro model of the primary macrophage cells from crucian carp Carassius auratus, and a total of 889 and 3765 differentially expressed genes (DEGs) were identified in pathogen-host interaction genes, respectively. Notably, GO and KEGG enrichment analysis were significantly involved in the PANoptosis pathways in ΔVgrG mutant-infected macrophages. The regulatory relationship of potential PANoptosis-related genes (PRGs) were analysed comprehensively, and their binding interaction of several hub proteins (eg., YcgR, Bcl2a, Calr3a, IL-1β) were determined by molecular docking analysis. To our best knowledge, this is first report of R. aquatilis VgrG-mediated interactions between pathogen and host macrophage cells, which will provide a new reference for understanding of molecular mechanism underlying PANoptosis in fish.
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Affiliation(s)
- Xiaoran Liu
- Tianjin Key Lab of Aqua-Ecology and Aquaculture, College of Fisheries, Tianjin Agricultural University, Tianjin, 300392, China
| | - Yanyan Cao
- Tianjin Key Lab of Aqua-Ecology and Aquaculture, College of Fisheries, Tianjin Agricultural University, Tianjin, 300392, China
| | - Xiucai Hu
- Tianjin Key Lab of Aqua-Ecology and Aquaculture, College of Fisheries, Tianjin Agricultural University, Tianjin, 300392, China.
| | - Aijun Lv
- Tianjin Key Lab of Aqua-Ecology and Aquaculture, College of Fisheries, Tianjin Agricultural University, Tianjin, 300392, China.
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2
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Hasan A, Ahmmed MT, Prapti BBR, Rahman A, Islam T, Chouhan CS, Rahman AKMA, Siddique MP. First report of MDR virulent Pseudomonas aeruginosa in apparently healthy Japanese quail (Coturnix japonica) in Bangladesh. PLoS One 2025; 20:e0316667. [PMID: 39854517 PMCID: PMC11761672 DOI: 10.1371/journal.pone.0316667] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2024] [Accepted: 12/14/2024] [Indexed: 01/26/2025] Open
Abstract
Pseudomonas aeruginosa (P. aeruginosa) is a major pathogen associated conditions like septicaemia, respiratory disorders, and diarrhoea in poultry, particularly in Japanese quail (Coturnix japonica). The infection causes huge economical losses due to its high transmissibility, mortality and zoonotic potential. This study aimed to isolate, identify, detect virulence genes, and profile multidrug resistance (MDR) of P. aeruginosa from Japanese quail. Oral and rectal swabs were collected from 110 apparently healthy quail birds across various districts in Bangladesh. Bacterial isolation and identification were performed using cultural, morphological, biochemical, and polymerase chain reaction (PCR) methods. Antibiotic susceptibility was assessed using the disc diffusion method, and virulence genes were detected through PCR. Multivariable logistic regression was used to identify risk factors for P. aeruginosa infection. Both conventional and PCR methods revealed that 25 (22.73%) of the quail birds were positive for P. aeruginosa. The results showed that quail birds in Narsingdi were five times more likely to harbor Pseudomonas species (OR: 5.02; 95% CI: 1.34-18.84) compared to those in Mymensingh Sadar. Additionally, quail birds younger than eight weeks had nearly six times higher odds (OR: 5.93; 95% CI: 1.96-17.91) of carrying Pseudomonas compared to older birds. Female quail birds had almost four times higher odds (OR: 3.77; 95% CI: 1.30-10.93) of harboring Pseudomonas species than males. All 25 P. aeruginosa isolates exhibited multi drug-resistance (MDR) patterns. Virulence gene analysis revealed the consistent presence of exoA and rhlR in all isolates, while exoS, exoY, rhlI, and rhlAB showed variable distribution. The high prevalence of MDR and virulent P. aeruginosa in apparently healthy quail birds particularly in Mymensingh and Dhaka divisions, highlights the urgent need for a comprehensive 'One Health' approach to proactively address and mitigate the potential risk this organism poses to both quail and human populations.
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Affiliation(s)
- Alamgir Hasan
- Department of Microbiology and Hygiene, Mymensingh, Bangladesh
| | | | | | - Aminur Rahman
- Department of Microbiology and Hygiene, Mymensingh, Bangladesh
| | - Tasnim Islam
- Department of Microbiology and Hygiene, Mymensingh, Bangladesh
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Habich A, Chaves Vargas V, Robinson LA, Allsopp LP, Unterweger D. Distribution of the four type VI secretion systems in Pseudomonas aeruginosa and classification of their core and accessory effectors. Nat Commun 2025; 16:888. [PMID: 39837841 PMCID: PMC11751169 DOI: 10.1038/s41467-024-54649-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2023] [Accepted: 11/14/2024] [Indexed: 01/23/2025] Open
Abstract
Bacterial type VI secretion systems (T6SSs) are puncturing molecular machines that transport effector proteins to kill microbes, manipulate eukaryotic cells, or facilitate nutrient uptake. How and why T6SS machines and effectors differ within a species is not fully understood. Here, we applied molecular population genetics to the T6SSs in a global population of the opportunistic pathogen Pseudomonas aeruginosa. We reveal varying occurrence of up to four distinct T6SS machines. Moreover, we define conserved core T6SS effectors, likely critical for the biology of P. aeruginosa, and accessory effectors that can exhibit mutual exclusivity between strains. By ancestral reconstruction, we observed dynamic changes in the gain and loss of effector genes in the species' evolutionary history. Our work highlights the potential importance of T6SS intraspecific diversity in bacterial ecology and evolution.
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Affiliation(s)
- Antonia Habich
- Institute for Experimental Medicine, Kiel University, Kiel, Germany
- Max Planck Institute for Evolutionary Biology, Plön, Germany
| | - Verónica Chaves Vargas
- Institute for Experimental Medicine, Kiel University, Kiel, Germany
- Max Planck Institute for Evolutionary Biology, Plön, Germany
| | - Luca A Robinson
- National Heart and Lung Institute, Imperial College London, London, UK
| | - Luke P Allsopp
- National Heart and Lung Institute, Imperial College London, London, UK
| | - Daniel Unterweger
- Institute for Experimental Medicine, Kiel University, Kiel, Germany.
- Max Planck Institute for Evolutionary Biology, Plön, Germany.
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4
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Han J, Aljahdali N, Zhao S, Tang H, Harbottle H, Hoffmann M, Frye JG, Foley SL. Infection biology of Salmonella enterica. EcoSal Plus 2024; 12:eesp00012023. [PMID: 38415623 PMCID: PMC11636313 DOI: 10.1128/ecosalplus.esp-0001-2023] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2023] [Accepted: 07/31/2023] [Indexed: 02/29/2024]
Abstract
Salmonella enterica is the leading cause of bacterial foodborne illness in the USA, with an estimated 95% of salmonellosis cases due to the consumption of contaminated food products. Salmonella can cause several different disease syndromes, with the most common being gastroenteritis, followed by bacteremia and typhoid fever. Among the over 2,600 currently identified serotypes/serovars, some are mostly host-restricted and host-adapted, while the majority of serotypes can infect a broader range of host species and are associated with causing both livestock and human disease. Salmonella serotypes and strains within serovars can vary considerably in the severity of disease that may result from infection, with some serovars that are more highly associated with invasive disease in humans, while others predominantly cause mild gastroenteritis. These observed clinical differences may be caused by the genetic make-up and diversity of the serovars. Salmonella virulence systems are very complex containing several virulence-associated genes with different functions that contribute to its pathogenicity. The different clinical syndromes are associated with unique groups of virulence genes, and strains often differ in the array of virulence traits they display. On the chromosome, virulence genes are often clustered in regions known as Salmonella pathogenicity islands (SPIs), which are scattered throughout different Salmonella genomes and encode factors essential for adhesion, invasion, survival, and replication within the host. Plasmids can also carry various genes that contribute to Salmonella pathogenicity. For example, strains from several serovars associated with significant human disease, including Choleraesuis, Dublin, Enteritidis, Newport, and Typhimurium, can carry virulence plasmids with genes contributing to attachment, immune system evasion, and other roles. The goal of this comprehensive review is to provide key information on the Salmonella virulence, including the contributions of genes encoded in SPIs and plasmids during Salmonella pathogenesis.
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Affiliation(s)
- Jing Han
- National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, Arkansas, USA
| | - Nesreen Aljahdali
- National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, Arkansas, USA
- Biological Science Department, College of Science, King Abdul-Aziz University, Jeddah, Saudi Arabia
| | - Shaohua Zhao
- Center for Veterinary Medicine, U.S. Food and Drug Administration, Rockville, Maryland, USA
| | - Hailin Tang
- National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, Arkansas, USA
| | - Heather Harbottle
- Center for Veterinary Medicine, U.S. Food and Drug Administration, Rockville, Maryland, USA
| | - Maria Hoffmann
- Center for Food Safety and Applied Nutrition, U.S. Food and Drug Administration, College Park, Maryland, USA
| | - Jonathan G. Frye
- Agricutlutral Research Service, U.S. Department of Agriculture, Athens, Georgia, USA
| | - Steven L. Foley
- National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, Arkansas, USA
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Danov A, Pollin I, Moon E, Ho M, Wilson BA, Papathanos PA, Kaplan T, Levy A. Identification of novel toxins associated with the extracellular contractile injection system using machine learning. Mol Syst Biol 2024; 20:859-879. [PMID: 39069594 PMCID: PMC11297309 DOI: 10.1038/s44320-024-00053-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2024] [Revised: 06/06/2024] [Accepted: 06/27/2024] [Indexed: 07/30/2024] Open
Abstract
Secretion systems play a crucial role in microbe-microbe or host-microbe interactions. Among these systems, the extracellular contractile injection system (eCIS) is a unique bacterial and archaeal extracellular secretion system that injects protein toxins into target organisms. However, the specific proteins that eCISs inject into target cells and their functions remain largely unknown. Here, we developed a machine learning classifier to identify eCIS-associated toxins (EATs). The classifier combines genetic and biochemical features to identify EATs. We also developed a score for the eCIS N-terminal signal peptide to predict EAT loading. Using the classifier we classified 2,194 genes from 950 genomes as putative EATs. We validated four new EATs, EAT14-17, showing toxicity in bacterial and eukaryotic cells, and identified residues of their respective active sites that are critical for toxicity. Finally, we show that EAT14 inhibits mitogenic signaling in human cells. Our study provides insights into the diversity and functions of EATs and demonstrates machine learning capability of identifying novel toxins. The toxins can be employed in various applications dependently or independently of eCIS.
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Affiliation(s)
- Aleks Danov
- Department of Plant Pathology and Microbiology, Institute of Environmental Sciences, The Robert H. Smith Faculty of Agriculture, Food & Environment, The Hebrew University of Jerusalem, Rehovot, 7610001, Israel
| | - Inbal Pollin
- Department of Plant Pathology and Microbiology, Institute of Environmental Sciences, The Robert H. Smith Faculty of Agriculture, Food & Environment, The Hebrew University of Jerusalem, Rehovot, 7610001, Israel
| | - Eric Moon
- Department of Microbiology, University of Illinois Urbana-Champaign, 601 South Goodwin Ave, Urbana, 61801, IL, USA
| | - Mengfei Ho
- Department of Microbiology, University of Illinois Urbana-Champaign, 601 South Goodwin Ave, Urbana, 61801, IL, USA
| | - Brenda A Wilson
- Department of Microbiology, University of Illinois Urbana-Champaign, 601 South Goodwin Ave, Urbana, 61801, IL, USA
| | - Philippos A Papathanos
- Department of Entomology, Institute of Environmental Sciences, The Robert H. Smith Faculty of Agriculture, Food & Environment, The Hebrew University of Jerusalem, Rehovot, 7610001, Israel
| | - Tommy Kaplan
- School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel
- Department of Developmental Biology and Cancer Research, Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Asaf Levy
- Department of Plant Pathology and Microbiology, Institute of Environmental Sciences, The Robert H. Smith Faculty of Agriculture, Food & Environment, The Hebrew University of Jerusalem, Rehovot, 7610001, Israel.
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Abdelhamed H, Mannan SB, Riman MM, Tekedar HC, Lawrence ML. Comparative analysis of three plasmids from Plesiomonas shigelloides strain MS-17-188 and their role in antimicrobial resistance. JAC Antimicrob Resist 2024; 6:dlae109. [PMID: 39035015 PMCID: PMC11258559 DOI: 10.1093/jacamr/dlae109] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2024] [Accepted: 06/25/2024] [Indexed: 07/23/2024] Open
Abstract
Background Plesiomonas shigelloides strain MS-17-188 was isolated from a deceased catfish from East Mississippi and showed resistance to florfenicol, tetracyclines and a sulphonamide. WGS of strain MS-17-188 revealed three plasmids (pPSMS-171881, pPSMS-171882 and pPSMS-171883). Objectives To accurately determine the impact of three plasmids found in P. shigelloides strain MS-17-188 on the dissemination of antibiotic resistance genes and to provide insights into the molecular structure of these plasmids. Methods The genetic features of these plasmids in terms of genes associated with antimicrobial resistance (AMR), virulence, transfer, maintenance and replication were identified using bioinformatic tools. Additionally, we investigated the in vitro mobilization and stability of plasmid-mediated resistance. The Comprehensive Antibiotic Resistance Database and Virulence Factors Database were used to detect the AMR genes and virulence genes of P. shigelloides plasmids. Moreover, plasmid mobility was evaluated by a filter-mating assay using strain MS-17-188 as a donor and azide-resistant Escherichia coli J53 as a recipient strain. A stability experiment was conducted to explore the persistence of plasmid-mediated antibiotic resistance in strain MS-17-188 in the absence and presence of selection. Results pPSMS-171881 harboured multidrug efflux complex (adeF) and two genes responsible for arsenic resistance (arsB and arsC). pPSMS-171882 had a region of 7085 bp encoding type IV secretion system proteins. pPSMS-171883 carried the tetracycline resistance genes tet(A) and tet(R), and a phenicol resistance gene (floR), which were flanked by two transposable elements and mobilization proteins, suggesting that there is a conjugative mechanism by which this plasmid can be mobilized. Results from the stability experiment indicated that pPSMS-171883 is lost over time in the absence of selective pressure. Moreover, pPSMS-171883 is more stable in P. shigelloides at growth temperatures of 30°C and 37°C compared with 40°C and 43°C. After intraperitoneal injection in catfish, P. shigelloides strain MS-17-188 resulted in no mortalities. Conclusions This is the first study to report plasmid-mediated AMR in Plesiomonas isolated from cultured fish, which needs continued monitoring. This study will provide an understanding of the genetic mechanisms of AMR and virulence of P. shigelloides.
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Affiliation(s)
- Hossam Abdelhamed
- Department of Comparative Biomedical Sciences, College of Veterinary Medicine, Mississippi State University, Mississippi State, MS, USA
| | - Shahnewaj Bin Mannan
- Department of Comparative Biomedical Sciences, College of Veterinary Medicine, Mississippi State University, Mississippi State, MS, USA
| | - Munshi Mustafiz Riman
- Department of Comparative Biomedical Sciences, College of Veterinary Medicine, Mississippi State University, Mississippi State, MS, USA
| | - Hasan C Tekedar
- Department of Comparative Biomedical Sciences, College of Veterinary Medicine, Mississippi State University, Mississippi State, MS, USA
| | - Mark L Lawrence
- Department of Comparative Biomedical Sciences, College of Veterinary Medicine, Mississippi State University, Mississippi State, MS, USA
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Che S, Sun C, Yang L, Zhou M, Xia L, Yan J, Jiang M, Wang J, Wang H, Zhao W, Toth I, Hu B, Guo T, Fan J. T6SS and T4SS Redundantly Secrete Effectors to Govern the Virulence and Bacterial Competition in Pectobacterium PccS1. PHYTOPATHOLOGY 2024; 114:1926-1939. [PMID: 38749069 DOI: 10.1094/phyto-11-23-0455-r] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/06/2024]
Abstract
Previous studies revealed that the type VI secretion system (T6SS) has an essential role in bacterial competition and virulence in many gram-negative bacteria. However, the role of T6SS in virulence in Pectobacterium atrosepticum remains controversial. We examined a closely related strain, PccS1, and discovered that its T6SS comprises a single-copy cluster of 17 core genes with a higher identity to homologs from P. atrosepticum. Through extensive phenotypic and functional analyses of over 220 derivatives of PccS1, we found that three of the five VgrGs could be classified into group I VgrGs. These VgrGs interacted with corresponding DUF4123 domain proteins, which were secreted outside of the membrane and were dependent on either the T6SS or type IV secretion system (T4SS). This interaction directly governed virulence and competition. Meanwhile, supernatant proteomic analyses with strains defective in the T6SS and/or T4SS confirmed that effectors, such as FhaB, were secreted redundantly to control the virulence and suppress host callose deposition in the course of infection. Notably, this redundant secretion mechanism between the T6SS and T4SS is believed to be the first of its kind in bacteria.
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Affiliation(s)
- Shu Che
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
- Cell and Molecular Science, James Hutton Institute, Invergowrie, Dundee DD2 5DA, United Kingdom
| | - Chen Sun
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Liuke Yang
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Min Zhou
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Lingyan Xia
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Jingyuan Yan
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Mengyi Jiang
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Jiaju Wang
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Huan Wang
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
- Cell and Molecular Science, James Hutton Institute, Invergowrie, Dundee DD2 5DA, United Kingdom
- Institute of Agricultural Science of Suzhou, Taihu Lake District, Suzhou 215155, China
| | - Wenjun Zhao
- CAIQ Center for Biosafety, Sanya 572024, China
| | - Ian Toth
- Cell and Molecular Science, James Hutton Institute, Invergowrie, Dundee DD2 5DA, United Kingdom
| | - Baishi Hu
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
| | - Tao Guo
- Southern Breeding Administrate Office of Hainan Province, Sanya 572000, China
| | - Jiaqin Fan
- Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, China
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Yu X, Yan Y, Zeng J, Liu Y, Sun X, Wang Z, Li L. T6SS nuclease effectors in Pseudomonas syringae act as potent antimicrobials in interbacterial competition. J Bacteriol 2024; 206:e0027323. [PMID: 38717111 PMCID: PMC11332151 DOI: 10.1128/jb.00273-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2023] [Accepted: 04/09/2024] [Indexed: 06/21/2024] Open
Abstract
Type VI secretion system (T6SS) is a potent weapon employed by various Pseudomonas species to compete with neighboring microorganisms for limited nutrients and ecological niches. However, the involvement of T6SS effectors in interbacterial competition within the phytopathogen Pseudomonas syringae remains unknown. In this study, we examined two T6SS clusters in a wild-type P. syringae MB03 and verified the involvement of one cluster, namely, T6SS-1, in interbacterial competition. Additionally, our results showed that two T6SS DNase effectors, specifically Tde1 and Tde4, effectively outcompeted antagonistic bacteria, with Tde4 playing a prominent role. Furthermore, we found several cognate immunity proteins, including Tde1ia, Tde1ib, and Tde4i, which are located in the downstream loci of their corresponding effector protein genes and worked synergistically to protect MB03 cells from self-intoxication. Moreover, expression of either Tde1 or C-terminus of Tde4 in Escherichia coli cells induced DNA degradation and changes in cell morphology. Thus, our results provide new insights into the role of the T6SS effectors of P. syringae in the interbacterial competition in the natural environment. IMPORTANCE The phytopathogen Pseudomonas syringae employs an active type VI secretion system (T6SS) to outcompete other microorganisms in the natural environment, particularly during the epiphytic growth in the phyllosphere. By examining two T6SS clusters in P. syringae MB03, T6SS-1 is found to be effective in killing Escherichia coli cells. We highlight the excellent antibacterial effect of two T6SS DNase effectors, namely, Tde1 and Tde4. Both of them function as nuclease effectors, leading to DNA degradation and cell filamentation in prey cells, ultimately resulting in cell death. Our findings deepen our understanding of the T6SS effector repertoires used in P. syringae and will facilitate the development of effective antibacterial strategies.
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Affiliation(s)
- Xun Yu
- National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China
- Cooperative Innovation Center of Industrial Fermentation (Ministry of Education and Hubei Province), Hubei University of Technology, Wuhan, China
| | - Yubo Yan
- National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China
| | - Jie Zeng
- National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China
| | - Yongxuan Liu
- National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China
| | - Xiaowen Sun
- National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China
| | - Zhiyong Wang
- College of Biological and Food Engineering, Hubei Minzu University, Enshi, China
| | - Lin Li
- National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China
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9
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Kim JH, Dong J, Le BH, Lonergan ZR, Gu W, Girke T, Zhang W, Newman DK, Martins-Green M. Pseudomonas aeruginosa Activates Quorum Sensing, Antioxidant Enzymes and Type VI Secretion in Response to Oxidative Stress to Initiate Biofilm Formation and Wound Chronicity. Antioxidants (Basel) 2024; 13:655. [PMID: 38929094 PMCID: PMC11200925 DOI: 10.3390/antiox13060655] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2024] [Revised: 04/29/2024] [Accepted: 05/13/2024] [Indexed: 06/28/2024] Open
Abstract
Pseudomonas aeruginosa (PA) is an opportunistic pathogen frequently isolated from cutaneous chronic wounds. How PA, in the presence of oxidative stress (OS), colonizes chronic wounds and forms a biofilm is still unknown. The purpose of this study is to investigate the changes in gene expression seen when PA is challenged with the high levels of OS present in chronic wounds. We used a biofilm-forming PA strain isolated from the chronic wounds of our murine model (RPA) and performed a qPCR to obtain gene expression patterns as RPA developed a biofilm in vitro in the presence of high levels of OS, and then compared the findings in vivo, in our mouse model of chronic wounds. We found that the planktonic bacteria under OS conditions overexpressed quorum sensing genes that are important for the bacteria to communicate with each other, antioxidant stress genes important to reduce OS in the microenvironment for survival, biofilm formation genes and virulence genes. Additionally, we performed RNAseq in vivo and identified the activation of novel genes/pathways of the Type VI Secretion System (T6SS) involved in RPA pathogenicity. In conclusion, RPA appears to survive the high OS microenvironment in chronic wounds and colonizes these wounds by turning on virulence, biofilm-forming and survival genes. These findings reveal pathways that may be promising targets for new therapies aimed at disrupting PA-containing biofilms immediately after debridement to facilitate the treatment of chronic human wounds.
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Affiliation(s)
- Jane H. Kim
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, CA 92521, USA
| | - Julianna Dong
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, CA 92521, USA
| | - Brandon H. Le
- Institute for Integrative Genome Biology, University of California, Riverside, CA 92521, USA
- Department of Botany and Plant Sciences, University of California, Riverside, CA 92521, USA
| | - Zachery R. Lonergan
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA
| | - Weifeng Gu
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, CA 92521, USA
| | - Thomas Girke
- Institute for Integrative Genome Biology, University of California, Riverside, CA 92521, USA
- Department of Botany and Plant Sciences, University of California, Riverside, CA 92521, USA
| | - Wei Zhang
- Institute for Integrative Genome Biology, University of California, Riverside, CA 92521, USA
- Department of Botany and Plant Sciences, University of California, Riverside, CA 92521, USA
| | - Dianne K. Newman
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA
- Geological and Planetary Sciences, California Institute of Technology, Pasadena, CA 91125, USA
| | - Manuela Martins-Green
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, CA 92521, USA
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10
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Chen S, Xie ZX, Yan KQ, Chen JW, Li DX, Wu PF, Peng L, Lin L, Dong CM, Zhao Z, Fan GY, Liu SQ, Herndl GJ, Wang DZ. Functional vertical connectivity of microbial communities in the ocean. SCIENCE ADVANCES 2024; 10:eadj8184. [PMID: 38781332 PMCID: PMC11114224 DOI: 10.1126/sciadv.adj8184] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/25/2023] [Accepted: 04/15/2024] [Indexed: 05/25/2024]
Abstract
Sinking particles are a critical conduit for the transport of surface microbes to the ocean's interior. Vertical connectivity of phylogenetic composition has been shown; however, the functional vertical connectivity of microbial communities has not yet been explored in detail. We investigated protein and taxa profiles of both free-living and particle-attached microbial communities from the surface to 3000 m depth using a combined metaproteomic and 16S rRNA amplicon sequencing approach. A clear compositional and functional vertical connectivity of microbial communities was observed throughout the water column with Oceanospirillales, Alteromonadales, and Rhodobacterales as key taxa. The surface-derived particle-associated microbes increased the expression of proteins involved in basic metabolism, organic matter processing, and environmental stress response in deep waters. This study highlights the functional vertical connectivity between surface and deep-sea microbial communities via sinking particles and reveals that a considerable proportion of the deep-sea microbes might originate from surface waters and have a major impact on the biogeochemical cycles in the deep sea.
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Affiliation(s)
- Shi Chen
- State Key Laboratory of Marine Environmental Science/College of the Environment and Ecology, Xiamen University, Xiamen 361005, China
- Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Zhuhai 519082, China
| | - Zhang-Xian Xie
- School of Resource and Environmental Sciences, Quanzhou Normal University, Quanzhou 362000, China
| | - Ke-Qiang Yan
- BGI-Shenzhen, Shenzhen 518083, China
- College of Life Sciences, University of Chinese Academy of Sciences, Beijing 100049, China
| | - Jian-Wei Chen
- Qingdao Key Laboratory of Marine Genomics, BGI-Qingdao, BGI-Shenzhen, Qingdao 266555, China
- Qingdao-Europe Advanced Institute for Life Sciences, BGI-Shenzhen, Qingdao 266555, China
| | - Dong-Xu Li
- State Key Laboratory of Marine Environmental Science/College of the Environment and Ecology, Xiamen University, Xiamen 361005, China
| | - Peng-Fei Wu
- State Key Laboratory of Marine Environmental Science/College of the Environment and Ecology, Xiamen University, Xiamen 361005, China
| | - Ling Peng
- Qingdao Key Laboratory of Marine Genomics, BGI-Qingdao, BGI-Shenzhen, Qingdao 266555, China
| | - Lin Lin
- State Key Laboratory of Marine Environmental Science/College of the Environment and Ecology, Xiamen University, Xiamen 361005, China
- Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Zhuhai 519082, China
| | - Chun-Ming Dong
- Key Laboratory of Marine Genetic Resources, Third Institute of Oceanography, Ministry of Natural Resources, No. 184, Daxue Road, Siming District, Xiamen 361005, Fujian, China
| | - Zihao Zhao
- Department of Functional and Evolutionary Ecology, Bio-Oceanography and Marine Biology Unit, University of Vienna, Djerassiplatz 1, 1030 Vienna, Austria
| | - Guang-Yi Fan
- BGI-Shenzhen, Shenzhen 518083, China
- Qingdao Key Laboratory of Marine Genomics, BGI-Qingdao, BGI-Shenzhen, Qingdao 266555, China
- Qingdao-Europe Advanced Institute for Life Sciences, BGI-Shenzhen, Qingdao 266555, China
| | - Si-Qi Liu
- BGI-Shenzhen, Shenzhen 518083, China
- College of Life Sciences, University of Chinese Academy of Sciences, Beijing 100049, China
| | - Gerhard J. Herndl
- Department of Functional and Evolutionary Ecology, Bio-Oceanography and Marine Biology Unit, University of Vienna, Djerassiplatz 1, 1030 Vienna, Austria
- NIOZ, Department of Marine Microbiology and Biogeochemistry, Royal Netherlands Institute for Sea Research, Utrecht University, 1790 AB Den Burg, Texel, Netherlands
| | - Da-Zhi Wang
- State Key Laboratory of Marine Environmental Science/College of the Environment and Ecology, Xiamen University, Xiamen 361005, China
- Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Zhuhai 519082, China
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11
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Fecht S, Paracuellos P, Subramoni S, Tan CAZ, Ilangovan A, Costa TRD, Filloux A. Functionality of chimeric TssA proteins in the type VI secretion system reveals sheath docking specificity within their N-terminal domains. Nat Commun 2024; 15:4283. [PMID: 38769318 PMCID: PMC11106082 DOI: 10.1038/s41467-024-48487-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2023] [Accepted: 05/01/2024] [Indexed: 05/22/2024] Open
Abstract
The genome of Pseudomonas aeruginosa encodes three type VI secretion systems, each comprising a dozen distinct proteins, which deliver toxins upon T6SS sheath contraction. The least conserved T6SS component, TssA, has variations in size which influence domain organisation and structure. Here we show that the TssA Nt1 domain interacts directly with the sheath in a specific manner, while the C-terminus is essential for oligomerisation. We built chimeric TssA proteins by swapping C-termini and showed that these can be functional even when made of domains from different TssA sub-groups. Functional specificity requires the Nt1 domain, while the origin of the C-terminal domain is more permissive for T6SS function. We identify two regions in short TssA proteins, loop and hairpin, that contribute to sheath binding. We propose a docking mechanism of TssA proteins with the sheath, and a model for how sheath assembly is coordinated by TssA proteins from this position.
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Affiliation(s)
- Selina Fecht
- CBRB Centre for Bacterial Resistance Biology, Department of Life Sciences, Imperial College London, London, SW7 2AZ, UK
| | - Patricia Paracuellos
- CBRB Centre for Bacterial Resistance Biology, Department of Life Sciences, Imperial College London, London, SW7 2AZ, UK
| | - Sujatha Subramoni
- Singapore Centre for Environmental Life Sciences Engineering, Nanyang Technological University, Singapore, 637551, Singapore
| | - Casandra Ai Zhu Tan
- Singapore Centre for Environmental Life Sciences Engineering, Nanyang Technological University, Singapore, 637551, Singapore
| | - Aravindan Ilangovan
- School of Biological and Behavioural Sciences, Queen Mary University of London, London, E1 4NS, UK
| | - Tiago R D Costa
- CBRB Centre for Bacterial Resistance Biology, Department of Life Sciences, Imperial College London, London, SW7 2AZ, UK
| | - Alain Filloux
- CBRB Centre for Bacterial Resistance Biology, Department of Life Sciences, Imperial College London, London, SW7 2AZ, UK.
- Singapore Centre for Environmental Life Sciences Engineering, Nanyang Technological University, Singapore, 637551, Singapore.
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12
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Wierz JC, Dirksen P, Kirsch R, Krüsemer R, Weiss B, Pauchet Y, Engl T, Kaltenpoth M. Intracellular symbiont Symbiodolus is vertically transmitted and widespread across insect orders. THE ISME JOURNAL 2024; 18:wrae099. [PMID: 38874172 PMCID: PMC11322605 DOI: 10.1093/ismejo/wrae099] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/06/2024] [Revised: 04/05/2024] [Accepted: 06/05/2024] [Indexed: 06/15/2024]
Abstract
Insects engage in manifold interactions with bacteria that can shift along the parasitism-mutualism continuum. However, only a small number of bacterial taxa managed to successfully colonize a wide diversity of insects, by evolving mechanisms for host-cell entry, immune evasion, germline tropism, reproductive manipulation, and/or by providing benefits to the host that stabilize the symbiotic association. Here, we report on the discovery of an Enterobacterales endosymbiont (Symbiodolus, type species Symbiodolus clandestinus) that is widespread across at least six insect orders and occurs at high prevalence within host populations. Fluorescence in situ hybridization in several Coleopteran and one Dipteran species revealed Symbiodolus' intracellular presence in all host life stages and across tissues, with a high abundance in female ovaries, indicating transovarial vertical transmission. Symbiont genome sequencing across 16 host taxa revealed a high degree of functional conservation in the eroding and transposon-rich genomes. All sequenced Symbiodolus genomes encode for multiple secretion systems, alongside effectors and toxin-antitoxin systems, which likely facilitate host-cell entry and interactions with the host. However, Symbiodolus-infected insects show no obvious signs of disease, and biosynthetic pathways for several amino acids and cofactors encoded by the bacterial genomes suggest that the symbionts may also be able to provide benefits to the hosts. A lack of host-symbiont cospeciation provides evidence for occasional horizontal transmission, so Symbiodolus' success is likely based on a mixed transmission mode. Our findings uncover a hitherto undescribed and widespread insect endosymbiont that may present valuable opportunities to unravel the molecular underpinnings of symbiosis establishment and maintenance.
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Affiliation(s)
- Jürgen C Wierz
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
| | - Philipp Dirksen
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
- Department of Evolutionary Ecology, Institute of Organismic and Molecular Evolution, Johannes Gutenberg University, 55128 Mainz, Germany
| | - Roy Kirsch
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
| | - Ronja Krüsemer
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
| | - Benjamin Weiss
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
| | - Yannick Pauchet
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
| | - Tobias Engl
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
| | - Martin Kaltenpoth
- Department of Insect Symbiosis, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany
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13
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Durán D, Vazquez-Arias D, Blanco-Romero E, Garrido-Sanz D, Redondo-Nieto M, Rivilla R, Martín M. An Orphan VrgG Auxiliary Module Related to the Type VI Secretion Systems from Pseudomonas ogarae F113 Mediates Bacterial Killing. Genes (Basel) 2023; 14:1979. [PMID: 38002922 PMCID: PMC10671463 DOI: 10.3390/genes14111979] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2023] [Revised: 10/17/2023] [Accepted: 10/20/2023] [Indexed: 11/26/2023] Open
Abstract
The model rhizobacterium Pseudomonas ogarae F113, a relevant plant growth-promoting bacterium, encodes three different Type VI secretion systems (T6SS) in its genome. In silico analysis of its genome revealed the presence of a genetic auxiliary module containing a gene encoding an orphan VgrG protein (VgrG5a) that is not genetically linked to any T6SS structural cluster, but is associated with genes encoding putative T6SS-related proteins: a possible adaptor Tap protein, followed by a putative effector, Tfe8, and its putative cognate immunity protein, Tfi8. The bioinformatic analysis of the VgrG5a auxiliary module has revealed that this cluster is only present in several subgroups of the P. fluorescens complex of species. An analysis of the mutants affecting the vgrG5a and tfe8 genes has shown that the module is involved in bacterial killing. To test whether Tfe8/Tfi8 constitute an effector-immunity pair, the genes encoding Tfe8 and Tfi8 were cloned and expressed in E. coli, showing that the ectopic expression of tfe8 affected growth. The growth defect was suppressed by tfi8 ectopic expression. These results indicate that Tfe8 is a bacterial killing effector, while Tfi8 is its cognate immunity protein. The Tfe8 protein sequence presents homology to the proteins of the MATE family involved in drug extrusion. The Tfe8 effector is a membrane protein with 10 to 12 transmembrane domains that could destabilize the membranes of target cells by the formation of pores, revealing the importance of these effectors for bacterial interaction. Tfe8 represents a novel type of a T6SS effector present in pseudomonads.
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Affiliation(s)
- David Durán
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049 Madrid, Spain; (D.D.); (D.V.-A.); (E.B.-R.); (D.G.-S.); (M.R.-N.); (R.R.)
| | - David Vazquez-Arias
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049 Madrid, Spain; (D.D.); (D.V.-A.); (E.B.-R.); (D.G.-S.); (M.R.-N.); (R.R.)
| | - Esther Blanco-Romero
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049 Madrid, Spain; (D.D.); (D.V.-A.); (E.B.-R.); (D.G.-S.); (M.R.-N.); (R.R.)
| | - Daniel Garrido-Sanz
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049 Madrid, Spain; (D.D.); (D.V.-A.); (E.B.-R.); (D.G.-S.); (M.R.-N.); (R.R.)
- Department of Fundamental Microbiology, University of Lausanne, 1015 Lausanne, Switzerland
| | - Miguel Redondo-Nieto
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049 Madrid, Spain; (D.D.); (D.V.-A.); (E.B.-R.); (D.G.-S.); (M.R.-N.); (R.R.)
| | - Rafael Rivilla
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049 Madrid, Spain; (D.D.); (D.V.-A.); (E.B.-R.); (D.G.-S.); (M.R.-N.); (R.R.)
| | - Marta Martín
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049 Madrid, Spain; (D.D.); (D.V.-A.); (E.B.-R.); (D.G.-S.); (M.R.-N.); (R.R.)
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14
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Rudzite M, Subramoni S, Endres RG, Filloux A. Effectiveness of Pseudomonas aeruginosa type VI secretion system relies on toxin potency and type IV pili-dependent interaction. PLoS Pathog 2023; 19:e1011428. [PMID: 37253075 PMCID: PMC10281587 DOI: 10.1371/journal.ppat.1011428] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2022] [Revised: 06/20/2023] [Accepted: 05/17/2023] [Indexed: 06/01/2023] Open
Abstract
The type VI secretion system (T6SS) is an antibacterial weapon that is used by numerous Gram-negative bacteria to gain competitive advantage by injecting toxins into adjacent prey cells. Predicting the outcome of a T6SS-dependent competition is not only reliant on presence-absence of the system but instead involves a multiplicity of factors. Pseudomonas aeruginosa possesses 3 distinct T6SSs and a set of more than 20 toxic effectors with diverse functions including disruption of cell wall integrity, degradation of nucleic acids or metabolic impairment. We generated a comprehensive collection of mutants with various degrees of T6SS activity and/or sensitivity to each individual T6SS toxin. By imaging whole mixed bacterial macrocolonies, we then investigated how these P. aeruginosa strains gain a competitive edge in multiple attacker/prey combinations. We observed that the potency of single T6SS toxin varies significantly from one another as measured by monitoring the community structure, with some toxins acting better in synergy or requiring a higher payload. Remarkably the degree of intermixing between preys and attackers is also key to the competition outcome and is driven by the frequency of contact as well as the ability of the prey to move away from the attacker using type IV pili-dependent twitching motility. Finally, we implemented a computational model to better understand how changes in T6SS firing behaviours or cell-cell contacts lead to population level competitive advantages, thus providing conceptual insight applicable to all types of contact-based competition.
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Affiliation(s)
- Marta Rudzite
- MRC Centre for Molecular Bacteriology and Infection, Department of Life Sciences, Imperial College London, London, United Kingdom
| | - Sujatha Subramoni
- Singapore Centre for Environmental Life Sciences Engineering, Nanyang Technological University, Singapore
| | - Robert G. Endres
- Centre for Integrative Systems Biology and Bioinformatics, Department of Life Sciences, Imperial College London, London, United Kingdom
| | - Alain Filloux
- MRC Centre for Molecular Bacteriology and Infection, Department of Life Sciences, Imperial College London, London, United Kingdom
- Singapore Centre for Environmental Life Sciences Engineering, Nanyang Technological University, Singapore
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15
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Kim H, Kim ES, Cho JH, Song M, Cho JH, Kim S, Keum GB, Kwak J, Doo H, Pandey S, Park SH, Lee JH, Jung H, Hur TY, Kim JK, Oh KK, Kim HB, Lee JH. Exploring the Microbial Community and Functional Characteristics of the Livestock Feces Using the Whole Metagenome Shotgun Sequencing. J Microbiol Biotechnol 2023; 33:51-60. [PMID: 36517072 PMCID: PMC9896000 DOI: 10.4014/jmb.2209.09013] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2022] [Revised: 11/17/2022] [Accepted: 12/06/2022] [Indexed: 12/23/2022]
Abstract
The foodborne illness is the important public health concerns, and the livestock feces are known to be one of the major reservoirs of foodborne pathogens. Also, it was reported that 45.5% of foodborne illness outbreaks have been associated with the animal products contaminated with the livestock feces. In addition, it has been known that the persistence of a pathogens depends on many potential virulent factors including the various virulent genes. Therefore, the first step to understanding the public health risk of livestock feces is to identify and describe microbial communities and potential virulent genes that contribute to bacterial pathogenicity. We used the whole metagenome shotgun sequencing to evaluate the prevalence of foodborne pathogens and to characterize the virulence associated genes in pig and chicken feces. Our data showed that the relative abundance of potential foodborne pathogens, such as Bacillus cereus was higher in chickens than pigs at the species level while the relative abundance of foodborne pathogens including Campylobacter coli was only detected in pigs. Also, the microbial functional characteristics of livestock feces revealed that the gene families related to "Biofilm formation and quorum sensing" were highly enriched in pigs than chicken. Moreover, the variety of gene families associated with "Resistance to antibiotics and toxic compounds" were detected in both animals. These results will help us to prepare the scientific action plans to improve awareness and understanding of the public health risks of livestock feces.
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Affiliation(s)
- Hyeri Kim
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Eun Sol Kim
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Jin Ho Cho
- Department of Animal Science, Chungbuk National University, Cheongju, Republic of Korea
| | - Minho Song
- Division of Animal and Dairy Science, Chungnam National University, Daejeon, Republic of Korea
| | - Jae Hyoung Cho
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Sheena Kim
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Gi Beom Keum
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Jinok Kwak
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Hyunok Doo
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Sriniwas Pandey
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea
| | - Seung-Hwan Park
- Korean Collection for Type Cultures, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Jeongeup, Republic of Korea
| | - Ju Huck Lee
- Korean Collection for Type Cultures, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Jeongeup, Republic of Korea
| | - Hyunjung Jung
- Animal Nutrition & Physiology Division, National Institute of Animal Science, RDA, Wanju, Republic of Korea
| | - Tai Young Hur
- Animal Diseases & Health Division, National Institute of Animal Science, RDA, Wanju, Republic of Korea
| | - Jae-Kyung Kim
- Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup, Republic of Korea
| | - Kwang Kyo Oh
- Microbial Safety Division, National Institute of Agricultural Sciences, Rural Development Administration, Wanju, 55365, Republic of Korea
| | - Hyeun Bum Kim
- Department of Animal Resources Science, Dankook University, Cheonan, Republic of Korea,Corresponding authors H.B. Kim Phone: +82-41-550-3653 E-mail:
| | - Ju-Hoon Lee
- Department of Food Animal Biotechnology, Department of Agricultural Biotechnology, Center for Food and Bioconvergence, Seoul National University, Seoul 08826, Republic of Korea,J.H. Lee Phone: +82-2-880-4854 E-mail:
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16
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An Important Role of the Type VI Secretion System of Pseudomonas aeruginosa Regulated by Dnr in Response to Anaerobic Environments. Microbiol Spectr 2022; 10:e0153322. [PMID: 36301114 PMCID: PMC9769707 DOI: 10.1128/spectrum.01533-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
Abstract
The type VI secretion system (T6SS) is capable of secreting a variety of metal-binding proteins involved in metal ion uptake, and it mediates an active metal ion transport system that contributes to competition between bacteria. Pseudomonas aeruginosa H2-T6SS can increase molybdenum ion acquisition and enhance bacterial survival advantage by promoting the secretion of the molybdate-binding protein ModA, in which the expression of H2-T6SS core genes hcp2, hsiA2, and clpV2 is activated by anaerobic conditions and are all regulated by the global regulator Anr. Here, we report the regulation of T6SS by Dnr, a dedicated dissimilatory nitrate respiration regulator in P. aeruginosa. Of the three distinct T6SS loci carried by P. aeruginosa, only the anaerobic expression of H2-T6SS was activated by Dnr; H1-T6SS or H3-T6SS did not respond to anaerobically induced activation. We also demonstrated that Dnr promotes the anaerobic secretion of ModA, which acts as a potential substrate for H2-T6SS, providing an advantage not only for the anaerobic growth of bacteria but also for functional competition. Overall, this study elucidates the important role played by Dnr in mediating the anaerobic expression of T6SS in P. aeruginosa, indicating that the functional advantage of H2-T6SS in response to anaerobic induction may be a conditional environmental adaptation. It also extends our understanding of the function of Dnr as a specific regulator of dissimilatory nitrate respiration. IMPORTANCE The type VI secretion system (T6SS) plays an important role in bacterial competition by mediating the transport of active metal ions. Pseudomonas aeruginosa carries three distinct T6SS loci (H1-, H2-, and H3-T6SS). The H2-T6SS promotes the secretion of the molybdate-binding protein ModA for the acquisition of molybdenum ions to adapt to anaerobic survival. Here, we report that the specialized dissimilatory nitrate respiration regulator Dnr in P. aeruginosa controls the anaerobic expression of H2-T6SS and that this regulation is essential for ModA protein secretion, anaerobic growth, and bacterial competition. This study elucidates the regulatory mechanism of Dnr on H2-T6SS in P. aeruginosa, revealing an important role played by H2-T6SS in adapting to an anaerobic environment.
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17
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Virulence Induction in Pseudomonas aeruginosa under Inorganic Phosphate Limitation: a Proteomics Perspective. Microbiol Spectr 2022; 10:e0259022. [PMID: 36354317 PMCID: PMC9769906 DOI: 10.1128/spectrum.02590-22] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
Inorganic phosphate (Pi) is a central nutrient and signal molecule for bacteria. Pi limitation was shown to increase the virulence of several phylogenetically diverse pathogenic bacteria with different lifestyles. Hypophosphatemia enhances the risk of death in patients due to general bacteremia and was observed after surgical injury in humans. Phosphate therapy, or the reduction of bacterial virulence by the administration of Pi or phosphate-containing compounds, is a promising anti-infective therapy approach that will not cause cytotoxicity or the emergence of antibiotic-resistant strains. The proof of concept of phosphate therapy has been obtained using primarily Pseudomonas aeruginosa (PA). However, a detailed understanding of Pi-induced changes at protein levels is missing. Using pyocyanin production as proxy, we show that the Pi-mediated induction of virulence is a highly cooperative process that occurs between 0.2 to 0.6 mM Pi. We present a proteomics study of PA grown in minimal medium supplemented with either 0.2 mM or 1 mM Pi and rich medium. About half of the predicted PA proteins could be quantified. Among the 1,471 dysregulated proteins comparing growth in 0.2 mM to 1 mM Pi, 1,100 were depleted under Pi-deficient conditions. Most of these proteins are involved in general and energy metabolism, different biosynthetic and catabolic routes, or transport. Pi depletion caused accumulation of proteins that belong to all major families of virulence factors, including pyocyanin synthesis, secretion systems, quorum sensing, chemosensory signaling, and the secretion of proteases, phospholipases, and phosphatases, which correlated with an increase in exoenzyme production and antibacterial activity. IMPORTANCE Antibiotics are our main weapons to fight pathogenic bacteria, but the increase in antibiotic-resistant strains and their consequences represents a major global health challenge, revealing the necessity to develop alternative antimicrobial strategies that do not involve the bacterial killing or growth inhibition. P. aeruginosa has been placed second on the global priority list to guide research on the development of new antibiotics. One of the most promising alternative strategies is the phosphate therapy for which the proof of concept has been obtained for P. aeruginosa. This article reports the detailed changes at the protein levels comparing P. aeruginosa grown under Pi-abundant and Pi-depleted conditions. These data describe in detail the molecular mechanisms underlying phosphate therapy. Apart from Pi, several other phosphate-containing compounds have been used for phosphate therapy and this study will serve as a reference for comparative studies aimed at evaluating the effect of alternative compounds.
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18
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González-Magaña A, Altuna J, Queralt-Martín M, Largo E, Velázquez C, Montánchez I, Bernal P, Alcaraz A, Albesa-Jové D. The P. aeruginosa effector Tse5 forms membrane pores disrupting the membrane potential of intoxicated bacteria. Commun Biol 2022; 5:1189. [PMID: 36335275 PMCID: PMC9637101 DOI: 10.1038/s42003-022-04140-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2021] [Accepted: 10/20/2022] [Indexed: 11/08/2022] Open
Abstract
The type VI secretion system (T6SS) of Pseudomonas aeruginosa injects effector proteins into neighbouring competitors and host cells, providing a fitness advantage that allows this opportunistic nosocomial pathogen to persist and prevail during the onset of infections. However, despite the high clinical relevance of P. aeruginosa, the identity and mode of action of most P. aeruginosa T6SS-dependent effectors remain to be discovered. Here, we report the molecular mechanism of Tse5-CT, the toxic auto-proteolytic product of the P. aeruginosa T6SS exported effector Tse5. Our results demonstrate that Tse5-CT is a pore-forming toxin that can transport ions across the membrane, causing membrane depolarisation and bacterial death. The membrane potential regulates a wide range of essential cellular functions; therefore, membrane depolarisation is an efficient strategy to compete with other microorganisms in polymicrobial environments.
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Affiliation(s)
- Amaia González-Magaña
- Fundación Biofísica Bizkaia/Biofisika Bizkaia Fundazioa (FBB) and Departamento de Bioquímica y Biología Molecular, Instituto Biofisika (CSIC, UPV/EHU), University of the Basque Country, 48940, Leioa, Spain
| | - Jon Altuna
- Fundación Biofísica Bizkaia/Biofisika Bizkaia Fundazioa (FBB) and Departamento de Bioquímica y Biología Molecular, Instituto Biofisika (CSIC, UPV/EHU), University of the Basque Country, 48940, Leioa, Spain
| | - María Queralt-Martín
- Laboratory of Molecular Biophysics, Department of Physics, University Jaume I, 12071, Castellón, Spain
| | - Eneko Largo
- Fundación Biofísica Bizkaia/Biofisika Bizkaia Fundazioa (FBB) and Departamento de Bioquímica y Biología Molecular, Instituto Biofisika (CSIC, UPV/EHU), University of the Basque Country, 48940, Leioa, Spain
- Departamento de Inmunología, Microbiología y Parasitología, University of the Basque Country, 48940, Leioa, Spain
| | - Carmen Velázquez
- Fundación Biofísica Bizkaia/Biofisika Bizkaia Fundazioa (FBB) and Departamento de Bioquímica y Biología Molecular, Instituto Biofisika (CSIC, UPV/EHU), University of the Basque Country, 48940, Leioa, Spain
| | - Itxaso Montánchez
- Departamento de Inmunología, Microbiología y Parasitología, University of the Basque Country, 48940, Leioa, Spain
| | - Patricia Bernal
- Departamento de Microbiología, Facultad de Biología, Universidad de Sevilla, 41012, Sevilla, Spain
| | - Antonio Alcaraz
- Laboratory of Molecular Biophysics, Department of Physics, University Jaume I, 12071, Castellón, Spain
| | - David Albesa-Jové
- Fundación Biofísica Bizkaia/Biofisika Bizkaia Fundazioa (FBB) and Departamento de Bioquímica y Biología Molecular, Instituto Biofisika (CSIC, UPV/EHU), University of the Basque Country, 48940, Leioa, Spain.
- Ikerbasque, Basque Foundation for Science, 48013, Bilbao, Spain.
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19
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de Oliveira HL, Dias GM, Neves BC. Genome sequence of Pseudomonas aeruginosa PA1-Petro—A role model of environmental adaptation and a potential biotechnological tool. Heliyon 2022; 8:e11566. [DOI: 10.1016/j.heliyon.2022.e11566] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2022] [Revised: 08/12/2022] [Accepted: 11/07/2022] [Indexed: 11/16/2022] Open
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20
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Huang Y, Han Y, Li Z, Li X, Li Z, Liu P, Liu X, Cheng Q, Fan F, Kan B, Liang W. TssI2-TsiI2 of Vibrio fluvialis VflT6SS2 delivers pesticin domain-containing periplasmic toxin and cognate immunity that modulates bacterial competitiveness. Gut Microbes 2022; 14:2136460. [PMID: 36288406 PMCID: PMC9620997 DOI: 10.1080/19490976.2022.2136460] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/04/2023] Open
Abstract
Vibrio fluvialis is a halophilic Gram-negative bacterium regarded as an emerging unusual enteric pathogen of increasing public health concern. Our previous work has identified two type VI secretion systems (T6SSs) in V. fluvialis, VflT6SS1, and VflT6SS2, and the latter is functional in mediating interbacterial competitiveness. However, its antibacterial effectors remain to be clarified. In this work, we focused on a new potential effector/immunity pair TssI2/TsiI2. Bioinformatics analysis revealed that the C-terminal domain of TssI2 belongs to a widespread family of pesticin, and its antibacterial toxicity and corresponding protection by TsiI2 were proved via bacterial killing assays, and their action sites were localized to the periplasm of bacterial cells. The interaction of TssI2 and TsiI2 was demonstrated by the bacterial adenylate cyclase two-hybrid, protein pull-down and isothermal titration calorimetry assays. Site-directed mutagenesis demonstrated that, in addition to Glu-844, Thr-863, and Asp-869, which correspond to three reported residues in pesticin of Yersinia pestis, additional residues including Phe-837, Gly-845, Tyr-851, Gly-867, Gln-963, Trp-975, and Arg-1000 were also proved to be crucial to the bactericidal activity of TssI2. Muramidase/lysozyme-related peptidoglycan (PG) hydrolase activities of TssI2 and its variants were validated with permeabilized Escherichia coli cells and purified PG substrate. Based on sequence homologies at C-terminals in various V. fluvialis isolates, TssI2 was subdivided into five clusters (12-22% identity among them), and the antibacterial activities of representative effectors from other four Clusters were also confirmed through periplasmic over-expression in E. coli host. Two selected cognate immunities were proved to confer protection against the toxicities of their effectors. Additionally, TsiI2, which belongs to Cluster I, exhibited cross-protection to effector from Cluster V. Together, current findings expand our knowledge of the diversity and consistency of evolved VgrG effectors in V. fluvialis and on how VflT6SS2 mediates a competitive advantage to gain a better survival.
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Affiliation(s)
- Yuanming Huang
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Yu Han
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Zhenpeng Li
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Xiaorui Li
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Zhe Li
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Ping Liu
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Xiaoshu Liu
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Qian Cheng
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Fenxia Fan
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Biao Kan
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China,CONTACT Biao Kan
| | - Weili Liang
- State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China,Weili Liang State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
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21
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Carobbi A, Di Nepi S, Fridman CM, Dar Y, Ben‐Yaakov R, Barash I, Salomon D, Sessa G. An antibacterial T6SS in Pantoea agglomerans pv. betae delivers a lysozyme-like effector to antagonize competitors. Environ Microbiol 2022; 24:4787-4802. [PMID: 35706135 PMCID: PMC9796082 DOI: 10.1111/1462-2920.16100] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2021] [Accepted: 06/10/2022] [Indexed: 12/30/2022]
Abstract
The type VI secretion system (T6SS) is deployed by numerous Gram-negative bacteria to deliver toxic effectors into neighbouring cells. The genome of Pantoea agglomerans pv. betae (Pab) phytopathogenic bacteria contains a gene cluster (T6SS1) predicted to encode a complete T6SS. Using secretion and competition assays, we found that T6SS1 in Pab is a functional antibacterial system that allows this pathogen to outcompete rival plant-associated bacteria found in its natural environment. Computational analysis of the T6SS1 gene cluster revealed that antibacterial effector and immunity proteins are encoded within three genomic islands that also harbour arrays of orphan immunity genes or toxin and immunity cassettes. Functional analyses indicated that VgrG, a specialized antibacterial effector, contains a C-terminal catalytically active glucosaminidase domain that is used to degrade prey peptidoglycan. Moreover, we confirmed that a bicistronic unit at the end of the T6SS1 cluster encodes a novel antibacterial T6SS effector and immunity pair. Together, these results demonstrate that Pab T6SS1 is an antibacterial system delivering a lysozyme-like effector to eliminate competitors, and indicate that this bacterium contains additional novel T6SS effectors.
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Affiliation(s)
- Andrea Carobbi
- School of Plant Sciences and Food Security, The George S. Wise Faculty of Life SciencesTel‐Aviv UniversityTel‐Aviv
| | - Simone Di Nepi
- School of Plant Sciences and Food Security, The George S. Wise Faculty of Life SciencesTel‐Aviv UniversityTel‐Aviv
| | - Chaya M. Fridman
- Department of Clinical Microbiology and Immunology, Sackler Faculty of MedicineTel Aviv UniversityTel Aviv
| | - Yasmin Dar
- Department of Clinical Microbiology and Immunology, Sackler Faculty of MedicineTel Aviv UniversityTel Aviv
| | - Rotem Ben‐Yaakov
- Department of Clinical Microbiology and Immunology, Sackler Faculty of MedicineTel Aviv UniversityTel Aviv
| | - Isaac Barash
- School of Plant Sciences and Food Security, The George S. Wise Faculty of Life SciencesTel‐Aviv UniversityTel‐Aviv
| | - Dor Salomon
- Department of Clinical Microbiology and Immunology, Sackler Faculty of MedicineTel Aviv UniversityTel Aviv
| | - Guido Sessa
- School of Plant Sciences and Food Security, The George S. Wise Faculty of Life SciencesTel‐Aviv UniversityTel‐Aviv
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22
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Genetic Identification of Pseudomonas aeruginosa Virulence Genes Associated with Keratitis in Egyptian Population. JOURNAL OF PURE AND APPLIED MICROBIOLOGY 2022. [DOI: 10.22207/jpam.16.3.12] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Infectious keratitis continues to be a prominent cause of vision impairment worldwide through a variety of causes. Pseudomonas aeruginosa is a Gram-negative bacterium that frequently causes vision-threatening microbial keratitis. P. aeruginosa contains a diverse array of virulence factors, including exoA, exoS, nan1, and lasB, some of which may contribute to its pathogenicity. Because the clinical characteristics of bacterial keratitis vary, making a quick differential diagnosis can be difficult, resulting in a delay in diagnosis and worse outcome. In this study, we performed multiplex polymerase chain reaction to detect the presence of nan1, toxA, exoS, and lasB, and determine their association with distinct clinical presentations of P. aeruginosa-related keratitis. We also performed antibiotic susceptibility testing of the isolates. A total of 49 P. aeruginosa strains were obtained from individuals with keratitis between May 2021 and December 2021 from the Research Institute of Ophthalmology, Giza, Egypt. Results showed that lasB was most expressed gene (81.8%), followed by tox (63.6%) and exoS (31.8%), whereas nan1 was the least commonly expressed gene 1316 (22.7%). The antibiotic susceptibility profile showed that TOB was the least sensitive antibiotic (26.5%), followed by CIP (34.7%), whereas CT was the most sensitive antibiotic (89.8%), followed by GAT (83.7%) and PB (81.6%). Several virulence genes were identified in P. aeruginosa isolates, suggesting that these genes are associated with varying degrees of intrinsic virulence and pathogenicity. Substantial associations between specific virulence genes and the source of infection imply that infection control measures can aid in regulating the distribution of virulence genes among P. aeruginosa strains.
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23
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Filloux A. Bacterial protein secretion systems: Game of types. MICROBIOLOGY (READING, ENGLAND) 2022; 168. [PMID: 35536734 DOI: 10.1099/mic.0.001193] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Protein trafficking across the bacterial envelope is a process that contributes to the organisation and integrity of the cell. It is the foundation for establishing contact and exchange between the environment and the cytosol. It helps cells to communicate with one another, whether they establish symbiotic or competitive behaviours. It is instrumental for pathogenesis and for bacteria to subvert the host immune response. Understanding the formation of envelope conduits and the manifold strategies employed for moving macromolecules across these channels is a fascinating playground. The diversity of the nanomachines involved in this process logically resulted in an attempt to classify them, which is where the protein secretion system types emerged. As our knowledge grew, so did the number of types, and their rightful nomenclature started to be questioned. While this may seem a semantic or philosophical issue, it also reflects scientific rigour when it comes to assimilating findings into textbooks and science history. Here I give an overview on bacterial protein secretion systems, their history, their nomenclature and why it can be misleading for newcomers in the field. Note that I do not try to suggest a new nomenclature. Instead, I explore the reasons why naming could have escaped our control and I try to reiterate basic concepts that underlie protein trafficking cross membranes.
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Affiliation(s)
- Alain Filloux
- MRC Centre for Molecular Bacteriology and Infection, Department of Life Sciences, Imperial College London, London, SW7 2AZ, UK
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24
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Ray S, Pandey NK, Kushwaha GS, Das S, Ganguly AK, Vashi N, Kumar D, Suar M, Bhavesh NS. Structural investigation on SPI-6 associated Salmonella Typhimurium VirG-like stress protein that promotes pathogen survival in macrophages. Protein Sci 2022; 31:835-849. [PMID: 34997791 DOI: 10.1002/pro.4272] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2021] [Revised: 12/29/2021] [Accepted: 12/30/2021] [Indexed: 11/10/2022]
Abstract
Enteric microbial pathogenesis, remarkably a complex process, is achieved by virulence factors encoded by genes located within regions of the bacterial genome termed pathogenicity islands. Salmonella pathogenicity islands (SPI) encodes proteins, that are essential virulence determinants for pathogen colonization and virulence. In addition to the well-characterized SPI-1 and SPI-2 proteins, which are required for bacterial invasion and intracellular replication, respectively, SPI-6 (formerly known as Salmonella enterica centisome 7 island; SCI) encoding proteins are also known to play pivotal role in Salmonella pathogenesis. However, the underlying molecular mechanism of these proteins remained elusive. To gain molecular insights into SPI-6 associated proteins, in this study, a SPI-6 Salmonella Typhimurium VirG-like protein (STV) is characterized using interdisciplinary experimental approaches including X-ray crystallography, NMR spectroscopy, infection assays, and mice model. The high-resolution crystal structure, determined by the single-wavelength anomalous dispersion (SAD) method, reveals that STV belongs to the LTxxQ motif family. Solution-state NMR spectroscopy studies reveal that STV form a dimer involving interconnected helices. Interestingly, functional studies shows that STV influence pathogen persistence inside macrophages in vitro at later stages of infection. Altogether, our findings suggest that STV, a member of the LTxxQ stress protein family, modulates bacterial survival mechanism in macrophages through SPI-1 and SPI-2 genes, respectively. This article is protected by copyright. All rights reserved.
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Affiliation(s)
- Shilpa Ray
- School of Biotechnology, Kalinga Institute of Industrial Technology (KIIT), Deemed to be university, Bhubaneswar, India
| | - Nishant Kumar Pandey
- School of Biotechnology, Kalinga Institute of Industrial Technology (KIIT), Deemed to be university, Bhubaneswar, India.,Transcription Regulation group, International Centre of Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi, India
| | - Gajraj Singh Kushwaha
- Transcription Regulation group, International Centre of Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi, India.,KIIT-Technology Business Incubator, Kalinga Institute of Industrial Technology (KIIT), Deemed to be university, Bhubaneswar, India
| | - Susmita Das
- School of Biotechnology, Kalinga Institute of Industrial Technology (KIIT), Deemed to be university, Bhubaneswar, India
| | - Akshay Kumar Ganguly
- Transcription Regulation group, International Centre of Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi, India
| | - Nimi Vashi
- Cellular Immunology group, International Centre of Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi, India
| | - Dhiraj Kumar
- Cellular Immunology group, International Centre of Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi, India
| | - Mrutyunjay Suar
- School of Biotechnology, Kalinga Institute of Industrial Technology (KIIT), Deemed to be university, Bhubaneswar, India.,KIIT-Technology Business Incubator, Kalinga Institute of Industrial Technology (KIIT), Deemed to be university, Bhubaneswar, India
| | - Neel Sarovar Bhavesh
- Transcription Regulation group, International Centre of Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi, India
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25
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Caseinolytic Proteins (Clp) in the Genus Klebsiella: Special Focus on ClpK. Molecules 2021; 27:molecules27010200. [PMID: 35011428 PMCID: PMC8746953 DOI: 10.3390/molecules27010200] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2021] [Revised: 11/15/2021] [Accepted: 12/15/2021] [Indexed: 11/16/2022] Open
Abstract
Caseinolytic proteins (Clp), which are present in both prokaryotes and eukaryotes, play a major role in cell protein quality control and survival of bacteria in harsh environmental conditions. Recently, a member of this protein family, ClpK was identified in a pathogenic strain of Klebsiella pneumoniae which was responsible for nosocomial infections. ClpK is linked to the thermal stress survival of this pathogen. The genome wide analysis of Clp proteins in Klebsiella spp. indicates that ClpK is present in only 34% of the investigated strains. This suggests that the uptake of the clpk gene is selective and may only be taken up by a pathogen that needs to survive harsh environmental conditions. In silico analyses and molecular dynamic simulations show that ClpK is mainly α-helical and is highly dynamic. ClpK was successfully expressed and purified to homogeneity using affinity and anion exchange chromatography. Biophysical characterization of ClpK showed that it is predominantly alpha-helical, and this is in agreement with in silico analysis of the protein structure. Furthermore, the purified protein is biologically active and hydrolyses ATP in a concentration- dependent manner.
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26
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Pan X, Tang M, You J, Osire T, Sun C, Fu W, Yi G, Yang T, Yang ST, Rao Z. PsrA is a novel regulator contributes to antibiotic synthesis, bacterial virulence, cell motility and extracellular polysaccharides production in Serratia marcescens. Nucleic Acids Res 2021; 50:127-148. [PMID: 34893884 PMCID: PMC8754645 DOI: 10.1093/nar/gkab1186] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2020] [Revised: 11/13/2021] [Accepted: 12/03/2021] [Indexed: 12/23/2022] Open
Abstract
Serratia marcescens is a Gram-negative bacterium of the Enterobacteriaceae family that can produce numbers of biologically active secondary metabolites. However, our understanding of the regulatory mechanisms behind secondary metabolites biosynthesis in S. marcescens remains limited. In this study, we identified an uncharacterized LysR family transcriptional regulator, encoding gene BVG90_12635, here we named psrA, that positively controlled prodigiosin synthesis in S. marcescens. This phenotype corresponded to PsrA positive control of transcriptional of the prodigiosin-associated pig operon by directly binding to a regulatory binding site (RBS) and an activating binding site (ABS) in the promoter region of the pig operon. We demonstrated that L-proline is an effector for the PsrA, which enhances the binding affinity of PsrA to its target promoters. Using transcriptomics and further experiments, we show that PsrA indirectly regulates pleiotropic phenotypes, including serrawettin W1 biosynthesis, extracellular polysaccharide production, biofilm formation, swarming motility and T6SS-mediated antibacterial activity in S. marcescens. Collectively, this study proposes that PsrA is a novel regulator that contributes to antibiotic synthesis, bacterial virulence, cell motility and extracellular polysaccharides production in S. marcescens and provides important clues for future studies exploring the function of the PsrA and PsrA-like proteins which are widely present in many other bacteria.
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Affiliation(s)
- Xuewei Pan
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China
| | - Mi Tang
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China
| | - Jiajia You
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China
| | - Tolbert Osire
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China
| | - Changhao Sun
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China
| | - Weilai Fu
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China.,Fujian Dabeinong Aquatic Sci. & Tech. Co., Ltd., Zhangzhou 363500, China
| | - Ganfeng Yi
- Fujian Dabeinong Aquatic Sci. & Tech. Co., Ltd., Zhangzhou 363500, China
| | - Taowei Yang
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China
| | - Shang-Tian Yang
- Department of Chemical and Biomolecular Engineering, The Ohio State University, Columbus, OH 43210, USA
| | - Zhiming Rao
- Key Laboratory of Industrial Biotechnology of the Ministry of Education, Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, Wuxi 214122, China
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27
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Iyer S, Das C. The unity of opposites: Strategic interplay between bacterial effectors to regulate cellular homeostasis. J Biol Chem 2021; 297:101340. [PMID: 34695417 PMCID: PMC8605245 DOI: 10.1016/j.jbc.2021.101340] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2020] [Revised: 10/15/2021] [Accepted: 10/20/2021] [Indexed: 11/23/2022] Open
Abstract
Legionella pneumophila is a facultative intracellular pathogen that uses the Dot/Icm Type IV secretion system (T4SS) to translocate many effectors into its host and establish a safe, replicative lifestyle. The bacteria, once phagocytosed, reside in a vacuolar structure known as the Legionella-containing vacuole (LCV) within the host cells and rapidly subvert organelle trafficking events, block inflammatory responses, hijack the host ubiquitination system, and abolish apoptotic signaling. This arsenal of translocated effectors can manipulate the host factors in a multitude of different ways. These proteins also contribute to bacterial virulence by positively or negatively regulating the activity of one another. Such effector-effector interactions, direct and indirect, provide the delicate balance required to maintain cellular homeostasis while establishing itself within the host. This review summarizes the recent progress in our knowledge of the structure-function relationship and biochemical mechanisms of select effector pairs from Legionella that work in opposition to one another, while highlighting the diversity of biochemical means adopted by this intracellular pathogen to establish a replicative niche within host cells.
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Affiliation(s)
- Shalini Iyer
- Department of Chemistry, Purdue University, West Lafayette, Indiana, USA.
| | - Chittaranjan Das
- Department of Chemistry, Purdue University, West Lafayette, Indiana, USA.
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28
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Nolan LM, Cain AK, Clamens T, Furniss RCD, Manoli E, Sainz-Polo MA, Dougan G, Albesa-Jové D, Parkhill J, Mavridou DA, Filloux A. Identification of Tse8 as a Type VI secretion system toxin from Pseudomonas aeruginosa that targets the bacterial transamidosome to inhibit protein synthesis in prey cells. Nat Microbiol 2021; 6:1199-1210. [PMID: 34413503 PMCID: PMC7611593 DOI: 10.1038/s41564-021-00950-8] [Citation(s) in RCA: 28] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2018] [Accepted: 07/15/2021] [Indexed: 02/07/2023]
Abstract
The Type VI secretion system (T6SS) is a bacterial nanomachine that delivers toxic effectors to kill competitors or subvert some of their key functions. Here, we use transposon directed insertion-site sequencing to identify T6SS toxins associated with the H1-T6SS, one of the three T6SS machines found in Pseudomonas aeruginosa. This approach identified several putative toxin-immunity pairs, including Tse8-Tsi8. Full characterization of this protein pair demonstrated that Tse8 is delivered by the VgrG1a spike complex into prey cells where it targets the transamidosome, a multiprotein complex involved in protein synthesis in bacteria that lack either one, or both, of the asparagine and glutamine transfer RNA synthases. Biochemical characterization of the interactions between Tse8 and the transamidosome components GatA, GatB and GatC suggests that the presence of Tse8 alters the fine-tuned stoichiometry of the transamidosome complex, and in vivo assays demonstrate that Tse8 limits the ability of prey cells to synthesize proteins. These data expand the range of cellular components targeted by the T6SS by identifying a T6SS toxin affecting protein synthesis and validate the use of a transposon directed insertion site sequencing-based global genomics approach to expand the repertoire of T6SS toxins in T6SS-encoding bacteria.
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Affiliation(s)
- Laura M. Nolan
- MRC Centre for Molecular Bacteriology and Infection (CMBI), Department of Life Sciences, Imperial College London, London, SW7 2AZ, United Kingdom
| | - Amy K. Cain
- Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, CB10 1SA, United Kingdom
| | - Thomas Clamens
- MRC Centre for Molecular Bacteriology and Infection (CMBI), Department of Life Sciences, Imperial College London, London, SW7 2AZ, United Kingdom
| | - R. Christopher D. Furniss
- MRC Centre for Molecular Bacteriology and Infection (CMBI), Department of Life Sciences, Imperial College London, London, SW7 2AZ, United Kingdom
| | - Eleni Manoli
- MRC Centre for Molecular Bacteriology and Infection (CMBI), Department of Life Sciences, Imperial College London, London, SW7 2AZ, United Kingdom
| | - Maria A. Sainz-Polo
- Structural Biology Unit, CIC bioGUNE, Bizkaia Technology Park, 48160 Derio, Spain
| | - Gordon Dougan
- Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, CB10 1SA, United Kingdom
| | - David Albesa-Jové
- Structural Biology Unit, CIC bioGUNE, Bizkaia Technology Park, 48160 Derio, Spain,IKERBASQUE, Basque Foundation for Science, Bilbao, Spain
| | - Julian Parkhill
- Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, CB10 1SA, United Kingdom
| | - Despoina A.I. Mavridou
- MRC Centre for Molecular Bacteriology and Infection (CMBI), Department of Life Sciences, Imperial College London, London, SW7 2AZ, United Kingdom,Department of Molecular Biosciences, University of Texas at Austin, Austin, 78712, Texas, USA,Correspondence to Alain Filloux: ; Despoina Mavridou:
| | - Alain Filloux
- MRC Centre for Molecular Bacteriology and Infection (CMBI), Department of Life Sciences, Imperial College London, London, SW7 2AZ, United Kingdom,Correspondence to Alain Filloux: ; Despoina Mavridou:
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29
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CHOPRA MEENU, BANDYOPADHYAY SAMIRAN, BHATTACHARYA DEBARAJ, BANERJEE JAYDEEP, SINGH RAVIKANT, SWARNKAR MOHIT, SINGH ANILKUMAR, DE SACHINANDAN. Genome based phylogeny and virulence factor analysis of mastitis causing Escherichia coli isolated from Indian cattle. THE INDIAN JOURNAL OF ANIMAL SCIENCES 2021; 90:1577-1583. [DOI: https:/doi.org/10.56093/ijans.v90i12.113165] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/30/2023]
Abstract
Mastitis is a highly infectious disease prevalent in dairy cattle and it is majorly caused by Escherichia coli (E. coli). The objective of present study is to investigate the occurrence of virulence genes, antimicrobial susceptibility and comparative analysis of E. coli (IVRI KOL CP4 and CM IVRI KOL-1) isolates from mastitis infected animal. Whole-genome sequencing (WGS) was performed using a PacBio RS II system and de novo assembled using Hierarchical Genome Assembly Process (HGAP3). Bacterial Pan Genome Analysis Pipeline (BPGA) was used for pangenome analysis. A set of 50 E. coli isolates were used for comparative analysis (48 collected from the database and 2 reference sequences). Core genes were further concatenated for phylogenetic analyses. In silico analysis was performed for antibiotic resistance and virulence gene identification. Both of the E. coli isolates carried many resistance genes including, b-lactamase, quinolones, rifampicin, macrolide, aminoglycoside and phenicols resistance. We detected 39 virulence genes in IVRI KOL CP4 and 52 in CM IVRI KOL-1 which include toxins, adhesions, invasins, secretion machineries or iron acquisition system. High prevalence of mastitis strains belongs to phylogroups A, although few isolates were also assigned to phylogenetic groups B1 and B2. In conclusion, the present study reported the presence of genes involved in Adherence, Iron acquisition, secretion system and toxins which shown to be crucial in MPEC pathogenicity. This is the first whole genome analysis of MPEC strains to be carried out in Indian isolate to highlights the spread of resistance and virulence genes in food animals.
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30
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Eddie BJ, Malanoski AP, Onderko EL, Phillips DA, Glaven SM. Marinobacter atlanticus electrode biofilms differentially regulate gene expression depending on electrode potential and lifestyle. Biofilm 2021; 3:100051. [PMID: 34195607 PMCID: PMC8233155 DOI: 10.1016/j.bioflm.2021.100051] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/01/2021] [Revised: 05/13/2021] [Accepted: 06/07/2021] [Indexed: 11/18/2022] Open
Abstract
Marinobacter spp. are opportunitrophs with a broad metabolic range including interactions with metals and electrodes. Marinobacter atlanticus strain CP1 was previously isolated from a cathode biofilm microbial community enriched from a sediment microbial fuel cell. Like other Marinobacter spp., M. atlanticus generates small amounts of electrical current when grown as a biofilm on an electrode, which is enhanced by the addition of redox mediators. However, the molecular mechanism resulting in extracellular electron transfer is unknown. Here, RNA-sequencing was used to determine changes in gene expression in electrode-attached and planktonic cells of M. atlanticus when grown at electrode potentials that enable current production (310 and 510 mV vs. SHE) compared to a potential that enables electron uptake (160 mV). Cells grown at current-producing potentials had increased expression of genes for molybdate transport, regardless of planktonic or attached lifestyle. Electrode-attached cells at current-producing potentials showed increased expression of the major export protein for the type VI secretion system. Growth at 160 mV resulted in an increase in expression of genes related to stress response and DNA repair including both RecBCD and the LexA/RecA regulatory network, as well as genes for copper homeostasis. Changes in expression of proteins with PEP C-terminal extracellular export motifs suggests that M. atlanticus is remodeling the biofilm matrix in response to electrode potential. These results improve our understanding of the physiological adaptations required for M. atlanticus growth on electrodes, and suggest a role for metal acquisition, either as a requirement for metal cofactors of redox proteins or as a possible electron shuttling mechanism.
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Affiliation(s)
- Brian J. Eddie
- Naval Research Laboratory, 4555 Overlook Ave., SW, Washington, DC, 20375, USA
| | | | | | - Daniel A. Phillips
- Oak Ridge Institute for Science and Education / US Army DEVCOM Chemical Biological Center, Biochemistry Branch, Aberdeen Proving Grounds, MD, 21010 USA
| | - Sarah M. Glaven
- Naval Research Laboratory, 4555 Overlook Ave., SW, Washington, DC, 20375, USA
- Corresponding author. 4555 Overlook Ave, Washington, DC, 20375.
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Wang T, Du X, Ji L, Han Y, Dang J, Wen J, Wang Y, Pu Q, Wu M, Liang H. Pseudomonas aeruginosa T6SS-mediated molybdate transport contributes to bacterial competition during anaerobiosis. Cell Rep 2021; 35:108957. [PMID: 33852869 DOI: 10.1016/j.celrep.2021.108957] [Citation(s) in RCA: 38] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2020] [Revised: 01/06/2021] [Accepted: 03/16/2021] [Indexed: 12/20/2022] Open
Abstract
Type VI secretion system (T6SS) is widely distributed in Gram-negative bacteria and functions as a versatile protein export machinery that translocates effectors into eukaryotic or prokaryotic target cells. Growing evidence indicates that T6SS can deliver several effectors to promote bacterial survival in harmful environments through metal ion acquisition. Here, we report that the Pseudomonas aeruginosa H2-T6SS mediates molybdate (MoO42-) acquisition by secretion of a molybdate-binding protein, ModA. The expression of H2-T6SS genes is activated by the master regulator Anr and anaerobiosis. We also identified a ModA-binding protein, IcmP, an insulin-cleaving metalloproteinase outer membrane protein. The T6SS-ModA-IcmP system provides P. aeruginosa with a growth advantage in bacterial competition under anaerobic conditions and plays an important role in bacterial virulence. Overall, this study clarifies the role of T6SS in secretion of an anion-binding protein, emphasizing the fundamental importance of this bacterium using T6SS-mediated molybdate uptake to adapt to complex environmental conditions.
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Affiliation(s)
- Tietao Wang
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China
| | - Xiao Du
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China
| | - Linxuan Ji
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China
| | - Yuying Han
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China
| | - Jing Dang
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China
| | - Jing Wen
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China
| | - Yarong Wang
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China
| | - Qinqin Pu
- Department of Basic Science, School of Medicine and Health Science, University of North Dakota, Grand Forks, ND 58203, USA
| | - Min Wu
- Department of Basic Science, School of Medicine and Health Science, University of North Dakota, Grand Forks, ND 58203, USA
| | - Haihua Liang
- Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi'an, ShaanXi 710069, China.
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Durán D, Bernal P, Vazquez-Arias D, Blanco-Romero E, Garrido-Sanz D, Redondo-Nieto M, Rivilla R, Martín M. Pseudomonas fluorescens F113 type VI secretion systems mediate bacterial killing and adaption to the rhizosphere microbiome. Sci Rep 2021; 11:5772. [PMID: 33707614 PMCID: PMC7970981 DOI: 10.1038/s41598-021-85218-1] [Citation(s) in RCA: 29] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2020] [Accepted: 02/26/2021] [Indexed: 02/06/2023] Open
Abstract
The genome of Pseudomonas fluorescens F113, a model rhizobacterium and a plant growth-promoting agent, encodes three putative type VI secretion systems (T6SSs); F1-, F2- and F3-T6SS. Bioinformatic analysis of the F113 T6SSs has revealed that they belong to group 3, group 1.1, and group 4a, respectively, similar to those previously described in Pseudomonas aeruginosa. In addition, in silico analyses allowed us to identify genes encoding a total of five orphan VgrG proteins and eight putative effectors (Tfe), some with their cognate immunity protein (Tfi) pairs. Genes encoding Tfe and Tfi are found in the proximity of P. fluorescens F113 vgrG, hcp, eagR and tap genes. RNA-Seq analyses in liquid culture and rhizosphere have revealed that F1- and F3-T6SS are expressed under all conditions, indicating that they are active systems, while F2-T6SS did not show any relevant expression under the tested conditions. The analysis of structural mutants in the three T6SSs has shown that the active F1- and F3-T6SSs are involved in interbacterial killing while F2 is not active in these conditions and its role is still unknown.. A rhizosphere colonization analysis of the double mutant affected in the F1- and F3-T6SS clusters showed that the double mutant was severely impaired in persistence in the rhizosphere microbiome, revealing the importance of these two systems for rhizosphere adaption.
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Affiliation(s)
- David Durán
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain
| | - Patricia Bernal
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain.,Departamento de Microbiología, Facultad de Biología, Universidad de Sevilla, Avenida de la Reina Mercedes, 6, 41012, Sevilla, Spain
| | - David Vazquez-Arias
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain
| | - Esther Blanco-Romero
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain
| | - Daniel Garrido-Sanz
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain
| | - Miguel Redondo-Nieto
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain
| | - Rafael Rivilla
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain
| | - Marta Martín
- Departamento de Biología, Facultad de Ciencias, Universidad Autónoma de Madrid, Darwin, 2, 28049, Madrid, Spain.
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T6SS Mediated Stress Responses for Bacterial Environmental Survival and Host Adaptation. Int J Mol Sci 2021; 22:ijms22020478. [PMID: 33418898 PMCID: PMC7825059 DOI: 10.3390/ijms22020478] [Citation(s) in RCA: 30] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2020] [Revised: 12/25/2020] [Accepted: 01/01/2021] [Indexed: 02/07/2023] Open
Abstract
The bacterial type VI secretion system (T6SS) is a protein secretion apparatus widely distributed in Gram-negative bacterial species. Many bacterial pathogens employ T6SS to compete with the host and to coordinate the invasion process. The T6SS apparatus consists of a membrane complex and an inner tail tube-like structure that is surrounded by a contractile sheath and capped with a spike complex. A series of antibacterial or antieukaryotic effectors is delivered by the puncturing device consisting of a Hcp tube decorated by the VgrG/PAAR complex into the target following the contraction of the TssB/C sheath, which often leads to damage and death of the competitor and/or host cells. As a tool for protein secretion and interspecies interactions, T6SS can be triggered by many different mechanisms to respond to various physiological conditions. This review summarizes our current knowledge of T6SS in coordinating bacterial stress responses against the unfavorable environmental and host conditions.
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Steinbach G, Crisan C, Ng SL, Hammer BK, Yunker PJ. Accumulation of dead cells from contact killing facilitates coexistence in bacterial biofilms. J R Soc Interface 2020; 17:20200486. [PMID: 33292099 PMCID: PMC7811593 DOI: 10.1098/rsif.2020.0486] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2020] [Accepted: 11/12/2020] [Indexed: 02/06/2023] Open
Abstract
Bacterial communities are governed by a wide variety of social interactions, some of which are antagonistic with potential significance for bacterial warfare. Several antagonistic mechanisms, such as killing via the type VI secretion system (T6SS), require killer cells to directly contact target cells. The T6SS is hypothesized to be a highly potent weapon, capable of facilitating the invasion and defence of bacterial populations. However, we find that the efficacy of contact killing is severely limited by the material consequences of cell death. Through experiments with Vibrio cholerae strains that kill via the T6SS, we show that dead cell debris quickly accumulates at the interface that forms between competing strains, preventing physical contact and thus preventing killing. While previous experiments have shown that T6SS killing can reduce a population of target cells by as much as 106-fold, we find that, as a result of the formation of dead cell debris barriers, the impact of contact killing depends sensitively on the initial concentration of killer cells. Killer cells are incapable of invading or eliminating competitors on a community level. Instead, bacterial warfare itself can facilitate coexistence between nominally antagonistic strains. While a variety of defensive strategies against microbial warfare exist, the material consequences of cell death provide target cells with their first line of defence.
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Affiliation(s)
- Gabi Steinbach
- School of Physics, Georgia Institute of Technology, Atlanta, GA, USA
- Center for Microbial Dynamics and Infection, Georgia Institute of Technology, Atlanta, GA, USA
| | - Cristian Crisan
- Center for Microbial Dynamics and Infection, Georgia Institute of Technology, Atlanta, GA, USA
- School of Biological Sciences, Georgia Institute of Technology, Atlanta, GA, USA
- Institute for Bioengineering and Biosciences, Georgia Institute of Technology, Atlanta, GA, USA
| | - Siu Lung Ng
- Center for Microbial Dynamics and Infection, Georgia Institute of Technology, Atlanta, GA, USA
- School of Biological Sciences, Georgia Institute of Technology, Atlanta, GA, USA
- Institute for Bioengineering and Biosciences, Georgia Institute of Technology, Atlanta, GA, USA
| | - Brian K. Hammer
- Center for Microbial Dynamics and Infection, Georgia Institute of Technology, Atlanta, GA, USA
- School of Biological Sciences, Georgia Institute of Technology, Atlanta, GA, USA
- Institute for Bioengineering and Biosciences, Georgia Institute of Technology, Atlanta, GA, USA
| | - Peter J. Yunker
- School of Physics, Georgia Institute of Technology, Atlanta, GA, USA
- Center for Microbial Dynamics and Infection, Georgia Institute of Technology, Atlanta, GA, USA
- Institute for Bioengineering and Biosciences, Georgia Institute of Technology, Atlanta, GA, USA
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González-Magaña A, Sainz-Polo MÁ, Pretre G, Çapuni R, Lucas M, Altuna J, Montánchez I, Fucini P, Albesa-Jové D. Structural insights into Pseudomonas aeruginosaType six secretion system exported effector 8. J Struct Biol 2020; 212:107651. [PMID: 33096229 DOI: 10.1016/j.jsb.2020.107651] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2020] [Revised: 10/13/2020] [Accepted: 10/15/2020] [Indexed: 12/24/2022]
Abstract
Recent reports indicate that the Type six secretion system exported effector 8 (Tse8) is a cytoactive effector secreted by the Type VI secretion system (T6SS) of the human pathogen Pseudomonas aeruginosa. The T6SS is a nanomachine that assembles inside of the bacteria and injects effectors/toxins into target cells, providing a fitness advantage over competing bacteria and facilitating host colonisation. Here we present the first crystal structure of Tse8 revealing that it conserves the architecture of the catalytic triad Lys84-transSer162-Ser186 that characterises members of the Amidase Signature superfamily. Furthermore, using binding affinity experiments, we show that the interaction of phenylmethylsulfonyl fluoride (PMSF) to Tse8 is dependent on the putative catalytic residue Ser186, providing support for its nucleophilic reactivity. This work thus demonstrates that Tse8 belongs to the Amidase Signature (AS) superfamily. Furthermore, it highlights Tse8 similarity to two family members: the Stenotrophomonas maltophilia Peptide Amidase and the Glutamyl-tRNAGln amidotransferase subunit A from Staphylococcus aureus.
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Affiliation(s)
- Amaia González-Magaña
- Instituto Biofisika (UPV/EHU, CSIC), Fundación Biofísica Bizkaia/Biofisika Bizkaia Fundazioa (FBB) and Departamento de Bioquímica y Biología Molecular, University of the Basque Country, 48940 Leioa, Spain
| | - M Ángela Sainz-Polo
- Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Bizkaia Technology Park, 48160 Derio, Spain
| | - Gabriela Pretre
- Instituto Biofisika (UPV/EHU, CSIC), Fundación Biofísica Bizkaia/Biofisika Bizkaia Fundazioa (FBB) and Departamento de Bioquímica y Biología Molecular, University of the Basque Country, 48940 Leioa, Spain
| | - Retina Çapuni
- Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Bizkaia Technology Park, 48160 Derio, Spain
| | - María Lucas
- Instituto de Biomedicina y Biotecnología de Cantabria (IBBTEC), Consejo Superior de Investigaciones Científicas (CSIC)-Universidad de Cantabria. Santander, 39011 Cantabria, Spain
| | - Jon Altuna
- Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Bizkaia Technology Park, 48160 Derio, Spain
| | - Itxaso Montánchez
- Departamento de Inmunología, Microbiología y Parasitología, University of the Basque Country, 48940 Leioa, Spain
| | - Paola Fucini
- Ikerbasque, Basque Foundation for Science, 48013 Bilbao, Spain
| | - David Albesa-Jové
- Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Bizkaia Technology Park, 48160 Derio, Spain; Ikerbasque, Basque Foundation for Science, 48013 Bilbao, Spain.
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Denet E, Triadou S, Michalet S, Nazaret S, Favre-Bonté S. Growth of Stenotrophomonas maltophilia and expression of Sme efflux pumps encoding genes in the presence of supernatants from amoebal and bacterial co-cultures: towards the role of amoebal secondary metabolites. ENVIRONMENTAL MICROBIOLOGY REPORTS 2020; 12:702-711. [PMID: 32902135 DOI: 10.1111/1758-2229.12884] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/31/2020] [Revised: 09/02/2020] [Accepted: 09/07/2020] [Indexed: 06/11/2023]
Abstract
Resistance-Nodulation-Division (RND) efflux pumps are relevant determinants of Stenotrophomonas maltophilia multidrug resistance as they can extrude a broad range of antibiotics and compounds involved in virulence and physiological functions. S. maltophilia, an environmental bacterium, was shown to be associated with amoebae and able to multiply inside them. To explore whether S. maltophilia RND efflux pumps play a role when interacting with amoebae, we evaluated the effect of amoebal culture and co-culture supernatants on the growth of S. maltophilia and the expression of sme efflux pump genes. Acanthamoeba castellanii and Willaertia magna were used as amoebal models and strain S. maltophilia BurE1 as bacterial one. Our data showed that both bacterial growth and sme gene expression were not modified by amoebal culture supernatants. On the contrary, co-culture supernatants negatively impacted the growth of BurE1 and induced the expression of three out of eight efflux pump genes, i.e. smeE, smeN and smeZ. Finally, we evidenced the production of A. castellanii secondary metabolites, putatively belonging to the diterpene family, in the amoebal supernatant and in the co-culture supernatant of A. castellanii and BurE1. Whether these compounds act directly as substrates of the efflux pumps and/or inducers of the sme genes need further investigations.
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Affiliation(s)
- Elodie Denet
- Université Lyon 1, Research Group on Environmental Multi-Resistance and Bacterial Efflux, UMR CNRS 5557/ UMR INRAe 1418 Ecologie Microbienne, 43 Boulevard du 11 Novembre 1918, Villeurbanne Cedex, 69622, France
| | - Sylvain Triadou
- Université Lyon 1, Research Group on Environmental Multi-Resistance and Bacterial Efflux, UMR CNRS 5557/ UMR INRAe 1418 Ecologie Microbienne, 43 Boulevard du 11 Novembre 1918, Villeurbanne Cedex, 69622, France
| | - Serge Michalet
- Université Lyon 1, Research Group on Environmental Multi-Resistance and Bacterial Efflux, UMR CNRS 5557/ UMR INRAe 1418 Ecologie Microbienne, 43 Boulevard du 11 Novembre 1918, Villeurbanne Cedex, 69622, France
| | - Sylvie Nazaret
- Université Lyon 1, Research Group on Environmental Multi-Resistance and Bacterial Efflux, UMR CNRS 5557/ UMR INRAe 1418 Ecologie Microbienne, 43 Boulevard du 11 Novembre 1918, Villeurbanne Cedex, 69622, France
| | - Sabine Favre-Bonté
- Université Lyon 1, Research Group on Environmental Multi-Resistance and Bacterial Efflux, UMR CNRS 5557/ UMR INRAe 1418 Ecologie Microbienne, 43 Boulevard du 11 Novembre 1918, Villeurbanne Cedex, 69622, France
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Varadarajan AR, Allan RN, Valentin JDP, Castañeda Ocampo OE, Somerville V, Pietsch F, Buhmann MT, West J, Skipp PJ, van der Mei HC, Ren Q, Schreiber F, Webb JS, Ahrens CH. An integrated model system to gain mechanistic insights into biofilm-associated antimicrobial resistance in Pseudomonas aeruginosa MPAO1. NPJ Biofilms Microbiomes 2020; 6:46. [PMID: 33127897 PMCID: PMC7603352 DOI: 10.1038/s41522-020-00154-8] [Citation(s) in RCA: 28] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/11/2020] [Accepted: 10/07/2020] [Indexed: 12/11/2022] Open
Abstract
Pseudomonas aeruginosa MPAO1 is the parental strain of the widely utilized transposon mutant collection for this important clinical pathogen. Here, we validate a model system to identify genes involved in biofilm growth and biofilm-associated antibiotic resistance. Our model employs a genomics-driven workflow to assemble the complete MPAO1 genome, identify unique and conserved genes by comparative genomics with the PAO1 reference strain and genes missed within existing assemblies by proteogenomics. Among over 200 unique MPAO1 genes, we identified six general essential genes that were overlooked when mapping public Tn-seq data sets against PAO1, including an antitoxin. Genomic data were integrated with phenotypic data from an experimental workflow using a user-friendly, soft lithography-based microfluidic flow chamber for biofilm growth and a screen with the Tn-mutant library in microtiter plates. The screen identified hitherto unknown genes involved in biofilm growth and antibiotic resistance. Experiments conducted with the flow chamber across three laboratories delivered reproducible data on P. aeruginosa biofilms and validated the function of both known genes and genes identified in the Tn-mutant screens. Differential protein abundance data from planktonic cells versus biofilm confirmed the upregulation of candidates known to affect biofilm formation, of structural and secreted proteins of type VI secretion systems, and provided proteogenomic evidence for some missed MPAO1 genes. This integrated, broadly applicable model promises to improve the mechanistic understanding of biofilm formation, antimicrobial tolerance, and resistance evolution in biofilms.
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Affiliation(s)
- Adithi R Varadarajan
- Research Group Molecular Diagnostics Genomics & Bioinformatics, Agroscope and SIB Swiss Institute of Bioinformatics, Wädenswil, Switzerland.
| | - Raymond N Allan
- School of Biological Sciences and Institute for Life Sciences, University of Southampton, Southampton, SO17 1BJ, UK
- National Biofilms Innovation Centre, University of Southampton, Southampton, SO17 1BJ, UK
- School of Pharmacy, Faculty of Health and Life Sciences, De Montfort University, Leicester, LE1 9BH, UK
| | - Jules D P Valentin
- Laboratory for Biointerfaces, Empa, Swiss Federal Laboratories for Materials Science and Technology, St. Gallen, Switzerland
- Department of BioMedical Engineering, University of Groningen and University Medical Center Groningen, Groningen, Netherlands
| | - Olga E Castañeda Ocampo
- Department of BioMedical Engineering, University of Groningen and University Medical Center Groningen, Groningen, Netherlands
| | - Vincent Somerville
- Research Group Molecular Diagnostics Genomics & Bioinformatics, Agroscope and SIB Swiss Institute of Bioinformatics, Wädenswil, Switzerland
| | - Franziska Pietsch
- Division of Biodeterioration and Reference Organisms, Federal Institute for Materials Research and Testing (BAM), Berlin, Germany
| | - Matthias T Buhmann
- Laboratory for Biointerfaces, Empa, Swiss Federal Laboratories for Materials Science and Technology, St. Gallen, Switzerland
| | - Jonathan West
- Faculty of Medicine, University of Southampton, Southampton, SO17 1BJ, UK
- Centre for Hybrid Biodevices, University of Southampton, Southampton, SO17 1BJ, UK
| | - Paul J Skipp
- Centre for Proteomics Research, University of Southampton, Southampton, SO17 1BJ, UK
| | - Henny C van der Mei
- Department of BioMedical Engineering, University of Groningen and University Medical Center Groningen, Groningen, Netherlands
| | - Qun Ren
- Laboratory for Biointerfaces, Empa, Swiss Federal Laboratories for Materials Science and Technology, St. Gallen, Switzerland
| | - Frank Schreiber
- Division of Biodeterioration and Reference Organisms, Federal Institute for Materials Research and Testing (BAM), Berlin, Germany
| | - Jeremy S Webb
- School of Biological Sciences and Institute for Life Sciences, University of Southampton, Southampton, SO17 1BJ, UK
- National Biofilms Innovation Centre, University of Southampton, Southampton, SO17 1BJ, UK
| | - Christian H Ahrens
- Research Group Molecular Diagnostics Genomics & Bioinformatics, Agroscope and SIB Swiss Institute of Bioinformatics, Wädenswil, Switzerland.
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Yuan S, Qi M, Peng Q, Huang G, Liu J, Xu Z, Gong X, Zhang G. Adaptive behaviors of planktonic Pseudomonas aeruginosa in response to the surface-deposited dead siblings. Colloids Surf B Biointerfaces 2020; 197:111408. [PMID: 33099147 DOI: 10.1016/j.colsurfb.2020.111408] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2020] [Revised: 09/23/2020] [Accepted: 10/08/2020] [Indexed: 11/27/2022]
Abstract
In this study, the 3D motion behaviors and the underlying adaptation mechanism of planktonic Pseudomonas aeruginosa (PAO1) in response to the deposited dead siblings nearby were explored. Utilizing a real-time 3D tracking technique, digital holographic microscopy (DHM), we demonstrate that planktonic cells near the surface covered with dead siblings have a lower density and a reduced 3D velocity compared with those upon viable ones. As a sign of chemosensory responses, bacteria swimming near the dead siblings exhibit increase in frequency of the 'flick' motion. Transcriptomic analysis by RNA-seq reveals an upregulated expression of dgcM and dgcE inhibited the movement of PAO1, accompanied by increased transcriptional levels of the virulence factor-related genes hcp1, clpV1, and vgrG1. Moreover, the decrease in l-glutamate and the increase in succinic acid in the metabolites of the dead bacteria layer promote the dispersion of planktonic bacteria. As a result, the dead siblings on a surface inhibit the bacterial accumulation and activate the adaptive defensive responses of planktonic PAO1 in the vicinity.
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Affiliation(s)
- Shuo Yuan
- Faculty of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, PR China
| | - Meng Qi
- Faculty of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, PR China
| | - Qingmei Peng
- Faculty of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, PR China
| | - Gui Huang
- Faculty of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, PR China
| | - Jun Liu
- Faculty of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, PR China
| | - Zhenbo Xu
- School of Food Science and Engineering, South China University of Technology, Guangzhou 510640, PR China
| | - Xiangjun Gong
- Faculty of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, PR China; Guangdong Provincial Key Laboratory of Luminescence from Molecular Aggregates (South China University of Technology), PR China.
| | - Guangzhao Zhang
- Faculty of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, PR China
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Shahbaz MU, Qian S, Yun F, Zhang J, Yu C, Tian F, Yang F, Chen H. Identification of the Regulatory Components Mediated by the Cyclic di-GMP Receptor Filp and Its Interactor PilZX3 and Functioning in Virulence of Xanthomonas oryzae pv. oryzae. MOLECULAR PLANT-MICROBE INTERACTIONS : MPMI 2020; 33:1196-1208. [PMID: 32720873 DOI: 10.1094/mpmi-04-20-0088-r] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/11/2023]
Abstract
The degenerate GGDEF/EAL domain protein Filp was previously shown to function as a cyclic di-GMP (c-di-GMP) signal receptor through its specific interaction with an atypical PilZ domain protein PilZX3 (formerly PXO_02715) and that this interaction is involved in regulating virulence in Xanthomonas oryzae pv. oryzae. As a step toward understanding the regulatory role of Filp/PilZX3-mediated c-di-GMP signaling in the virulence of X. oryzae pv. oryzae, differentially expressed proteins (DEPs) downstream of Filp/PilZX3 were identified by isobaric tagging for relative and absolute quantitation (iTRAQ). A total of 2,346 proteins were identified, of which 157 displayed significant differential expression in different strains. Western blot and quantitative reverse transcription-PCR analyses showed that the expression of HrrP (histidine kinase-response regulator hybrid protein), PhrP (PhoPQ-regulated protein), ProP (prophage Lp2 protein 6) were increased in the ∆filp, ∆pilZX3, and ∆filp∆pilZX3 mutant strains, while expression of CheW1 (chemotaxis protein CheW1), EdpX2 (the second EAL domain protein identified in X. oryzae pv. oryzae), HGdpX2 (the second HD-GYP domain protein identified in X. oryzae pv. oryzae) was decreased in all mutant strains compared with that in the wild type, which was consistent with the iTRAQ data. Deletion of the hrrP and proP genes resulted in significant increases in virulence, whereas deletion of the cheW1, hGdpX2, or tdrX2 genes resulted in decreased virulence. Enzyme assays indicated that EdpX2 and HGdpX2 were active phosphodiesterases (PDEs). This study provides a proteomic description of putative regulatory pathway of Filp and PilZX3 and characterized novel factors that contributed to the virulence of X. oryzae pv. oryzae regulated by c-di-GMP signaling.
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Affiliation(s)
- Muhammad Umar Shahbaz
- State Key Laboratory for Biology Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China
- Plant Pathology Section, Plant Pathology Research Institute, AARI, Faisalabad 38850, Pakistan
| | - Shanshan Qian
- State Key Laboratory for Biology Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Fei Yun
- National Tobacco Cultivation and Physiology and Biochemistry Research Centre/Key Laboratory for Tobacco Cultivation of Tobacco Industry, Henan Agricultural University, Zhengzhou 450002, China
| | - Jie Zhang
- State Key Laboratory for Biology Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Chao Yu
- State Key Laboratory for Biology Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Fang Tian
- State Key Laboratory for Biology Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Fenghuan Yang
- State Key Laboratory for Biology Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Huamin Chen
- State Key Laboratory for Biology Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China
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Using proteomics to identify host cell interaction partners for VgrG and IglJ. Sci Rep 2020; 10:14612. [PMID: 32884055 PMCID: PMC7471685 DOI: 10.1038/s41598-020-71641-3] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2020] [Accepted: 07/23/2020] [Indexed: 11/17/2022] Open
Abstract
Francisella tularensis is a highly virulent intracellular bacterium and the causative agent of tularemia. The disease is characterized by the suboptimal innate immune response and consequently by the impaired adaptive immunity. The virulence of this pathogen depends on proteins encoded by a genomic island termed the Francisella Pathogenicity Island (FPI). However, the precise biological roles of most of the FPI-encoded proteins remain to be clarified. In this study, we employed stable isotope labeling by amino acids in cell culture (SILAC) in combination with affinity protein purification coupled with liquid chromatography–mass spectrometry to identify potential protein-effector binding pairs for two FPI virulence effectors IglJ and VgrG. Our results may indicate that while the IglJ protein interactions primarily affect mitochondria, the VgrG interactions affect phagosome and/or autophagosome biogenesis via targeting components of the host’s exocyst complex.
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Li Y, Chen L, Zhang P, Bhagirath AY, Duan K. ClpV3 of the H3-Type VI Secretion System (H3-T6SS) Affects Multiple Virulence Factors in Pseudomonas aeruginosa. Front Microbiol 2020; 11:1096. [PMID: 32547522 PMCID: PMC7273116 DOI: 10.3389/fmicb.2020.01096] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2020] [Accepted: 05/01/2020] [Indexed: 11/13/2022] Open
Abstract
The type VI secretion system (T6SS) is a toxic effector delivery apparatus widely distributed in Gram-negative bacteria. The opportunistic pathogen Pseudomonas aeruginosa encodes three T6SSs, namely H1-, H2-, and H3-T6SS. Each T6SS possesses its own effectors and their roles are not yet fully understood. Here, we report that an H3-T6SS deletion mutant PAO1(ΔclpV3) significantly affected the virulence-related phenotypes including pyocyanin production, biofilm formation, proteolytic activity, and motilities. Most interestingly, the expression of T3SS genes was markedly affected, indicating a link between H3-T6SS and T3SS. RNA-Sequencing was performed to globally identify the genes differentially expressed when H3-T6SS was inactivated and the results obtained correlated well with the observed phenotypes. Interestingly, the expressions of T2SS, T3SS, H2-T6SS, and H3-T6SS were all significantly decreased, while H1-T6SS was increased in the PAO1(ΔclpV3) strain. We also observed that the intracellular concentration of secondary messenger cAMP was reduced in PAO1(ΔclpV3), and the c-di-GMP level was also decreased as indicated by the decreased cdrA reporter activity. Finally, by using a Galleria mellonella infection model, we show that H3-T6SS plays a key role in the pathogenicity of P. aeruginosa in vivo. Overall, our study highlights the unique connection of H3-T6SS in P. aeruginosa with T3SS, pyocyanin production, biofilm formation and in vivo pathogenicity.
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Affiliation(s)
- Yanqi Li
- Department of Oral Biology, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB, Canada
| | - Lin Chen
- College of Life Sciences, Northwest University, Xi’an, China
| | - Pansong Zhang
- College of Life Sciences, Northwest University, Xi’an, China
| | - Anjali Y. Bhagirath
- Department of Oral Biology, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB, Canada
| | - Kangmin Duan
- Department of Oral Biology, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB, Canada
- Department of Medical Microbiology & Infectious Diseases, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB, Canada
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Song H, Kang Y, Qian A, Shan X, Li Y, Zhang L, Zhang H, Sun W. Inactivation of the T6SS inner membrane protein DotU results in severe attenuation and decreased pathogenicity of Aeromonas veronii TH0426. BMC Microbiol 2020; 20:76. [PMID: 32245412 PMCID: PMC7119292 DOI: 10.1186/s12866-020-01743-5] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/27/2019] [Accepted: 03/04/2020] [Indexed: 11/19/2022] Open
Abstract
Background The inner membrane protein DotU of Aeromonas veronii is an important component of the minimal core conserved membrane proteome required for the formation of an envelope-transmembrane complex. This protein functions in a type VI secretion system (T6SS), and the role of this T6SS during the pathogenic process has not been clearly described. Results A recombinant A. veronii with a partial disruption of the dotU gene (720 bp of the in-frame sequence) (defined as ∆dotU) was constructed by two conjugate exchanges. We found that the mutant ∆dotU allele can be stably inherited for more than 50 generations. Inactivation of the A. veronii dotU gene resulted in no significant changes in growth or resistance to various environmental changes. However, compared with the wild-type strain colony, the mutant ∆dotU colony had a rough surface morphology. In addition, the biofilm formation ability of the mutant ∆dotU was significantly enhanced by 2.1-fold. Conversely, the deletion of the dotU gene resulted in a significant decrease in pathogenicity and infectivity compared to those of the A. veronii wild-type strain. Conclusions Our findings indicated that the dotU gene was an essential participant in the pathogenicity and invasiveness of A. veronii TH0426, which provides a novel perspective on the pathogenesis of TH0426 and lays the foundation for discovering potential T6SS effectors.
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Affiliation(s)
- Haichao Song
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China
| | - Yuanhuan Kang
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China
| | - Aidong Qian
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China
| | - Xiaofeng Shan
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China
| | - Ying Li
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China
| | - Lei Zhang
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China
| | - Haipeng Zhang
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China
| | - Wuwen Sun
- College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, 130118, China.
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Liu J, Adrian L, Häggblom MM. Transcriptomic and Proteomic Responses of the Organohalide-Respiring Bacterium Desulfoluna spongiiphila to Growth with 2,6-Dibromophenol as the Electron Acceptor. Appl Environ Microbiol 2020; 86:e02146-19. [PMID: 31836581 PMCID: PMC7028966 DOI: 10.1128/aem.02146-19] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2019] [Accepted: 12/10/2019] [Indexed: 01/31/2023] Open
Abstract
Organohalide respiration is an important process in the global halogen cycle and for bioremediation. In this study, we compared the global transcriptomic and proteomic analyses of Desulfoluna spongiiphila strain AA1, an organohalide-respiring member of the Desulfobacterota isolated from a marine sponge, with 2,6-dibromophenol or with sulfate as an electron acceptor. The most significant difference of the transcriptomic analysis was the expression of one reductive dehalogenase gene cluster (rdh16), which was significantly upregulated with the addition of 2,6-dibromophenol. The corresponding protein, reductive dehalogenase RdhA16032, was detected in the proteome under treatment with 2,6-dibromophenol but not with sulfate only. There was no significant difference in corrinoid biosynthesis gene expression levels between the two treatments, indicating that the production of corrinoid in D. spongiiphila is constitutive or not specific for organohalide versus sulfate respiration. Electron-transporting proteins or mediators unique for reductive dehalogenation were not revealed in our analysis, and we hypothesize that reductive dehalogenation may share an electron-transporting system with sulfate reduction. The metabolism of D. spongiiphila, predicted from transcriptomic and proteomic results, demonstrates high metabolic versatility and provides insights into the survival strategies of a marine sponge symbiont in an environment rich in organohalide compounds and other secondary metabolites.IMPORTANCE Respiratory reductive dehalogenation is an important process in the overall cycling of both anthropogenic and natural organohalide compounds. Marine sponges produce a vast array of bioactive compounds as secondary metabolites, including diverse halogenated compounds that may enrich for dehalogenating bacteria. Desulfoluna spongiiphila strain AA1 was originally enriched and isolated from the marine sponge Aplysina aerophoba and can grow with both brominated compounds and sulfate as electron acceptors for respiration. An understanding of the overall gene expression and the protein production profile in response to organohalides is needed to identify the full complement of genes or enzymes involved in organohalide respiration. Elucidating the metabolic capacity of this sponge-associated bacterium lays the foundation for understanding how dehalogenating bacteria may control the fate of organohalide compounds in sponges and their role in a symbiotic organobromine cycle.
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Affiliation(s)
- Jie Liu
- Department of Biochemistry and Microbiology, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, New Jersey, USA
| | - Lorenz Adrian
- Department of Isotope Biogeochemistry, Helmholtz Centre for Environmental Research-UFZ, Leipzig, Germany
- Fachgebiet Geobiotechnologie, Technische Universität Berlin, Berlin, Germany
| | - Max M Häggblom
- Department of Biochemistry and Microbiology, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, New Jersey, USA
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44
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Cui J, Hu J, Du X, Yan C, Xue G, Li S, Cui Z, Huang H, Yuan J. Genomic Analysis of Putative Virulence Factors Affecting Cytotoxicity of Cronobacter. Front Microbiol 2020; 10:3104. [PMID: 32117082 PMCID: PMC7019382 DOI: 10.3389/fmicb.2019.03104] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2019] [Accepted: 12/23/2019] [Indexed: 01/07/2023] Open
Abstract
Cronobacter spp. can cause systemic infections, such as meningitis, sepsis, and necrotizing enterocolitis, in immunocompromised patients, especially neonates. Although some virulence factors have been reported previously, the pathogenesis of Cronobacter remains unclear. In this study, we compared genome sequences from different Cronobacter species, sequence types, and sources, with the virulence genes in the virulence factor database. The results showed that Cronobacter has species specificity for these virulence genes. Additionally, two gene clusters, including sfp encoding fimbriae and hly encoding hemolysin, were discovered. Through cell adhesion, cytotoxicity, and hemolysis assays, we found that the isolates possessing the two gene clusters had higher cytotoxicity and stronger hemolysis capacity than those of other isolates in this study. Moreover, analysis of type VI secretion system (T6SS) cluster and putative fimbria gene clusters of Cronobacter revealed that T6SS have species specificity and isolates with high cytotoxicity possessed more complete T6SS cluster construction than that of the rest. In conclusion, the two novel gene clusters and T6SS cluster were involved in the mechanism underlying the cytotoxicity of Cronobacter.
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Affiliation(s)
- Jinghua Cui
- Department of Bacteriology, Capital Institute of Pediatrics, Beijing, China.,State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Jinrui Hu
- State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Xiaoli Du
- State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Chao Yan
- Department of Bacteriology, Capital Institute of Pediatrics, Beijing, China
| | - Guanhua Xue
- Department of Bacteriology, Capital Institute of Pediatrics, Beijing, China
| | - Shaoli Li
- Department of Bacteriology, Capital Institute of Pediatrics, Beijing, China
| | - Zhigang Cui
- State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
| | - Hua Huang
- Beijing Products Quality Supervision and Inspection Institute, Beijing, China
| | - Jing Yuan
- Department of Bacteriology, Capital Institute of Pediatrics, Beijing, China
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45
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Hsieh PF, Lu YR, Lin TL, Lai LY, Wang JT. Klebsiella pneumoniae Type VI Secretion System Contributes to Bacterial Competition, Cell Invasion, Type-1 Fimbriae Expression, and In Vivo Colonization. J Infect Dis 2019; 219:637-647. [PMID: 30202982 PMCID: PMC6350951 DOI: 10.1093/infdis/jiy534] [Citation(s) in RCA: 64] [Impact Index Per Article: 10.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2018] [Accepted: 09/03/2018] [Indexed: 01/25/2023] Open
Abstract
Background We previously isolated a Klebsiella pneumoniae strain, NTUH-K2044, from a community-acquired pyogenic liver abscess (PLA) patient. Analysis of the NTUH-K2044 genome revealed that this strain harbors 2 putative type VI secretion system (T6SS)-encoding gene clusters. Methods The distribution of T6SS genes in the PLA and intestinal-colonizing K pneumoniae clinical isolates was examined. icmF1-, icmF2-, icmF1/icmF2-, and hcp-deficient K pneumoniae strains were constructed using an unmarked deletion method. The roles of T6SSs in antibacterial activity, type-1 fimbriae expression, cell adhesion, and invasion and intestinal colonization were determined. Results The prevalence of T6SSs is higher in the PLA strains than in the intestinal-colonizing strains (37 of 42 vs 54 of 130). Deletion of icmF1/icmF2 and hcp genes significantly reduced interbacterial and intrabacterial killing. Strain deleted for icmF1 and icmF2 exhibited decreased transcriptional expression of type-1 fimbriae and reduced adherence to and invasion of human colorectal epithelial cells and was attenuated for in vivo competition to enable colonization of the host gut. Finally, Hcp expression in K pneumoniae was silenced by the histone-like nucleoid structuring protein via direct binding. Conclusions These results provide new insights into T6SS-mediated bacterial competition and attachment in K pneumoniae and could facilitate the prevention of K pneumoniae infection.
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Affiliation(s)
- Pei-Fang Hsieh
- Department of Microbiology, National Taiwan University College of Medicine, Taipei
| | - Yi-Rou Lu
- Department of Microbiology, National Taiwan University College of Medicine, Taipei
| | - Tzu-Lung Lin
- Department of Microbiology, National Taiwan University College of Medicine, Taipei
| | - Li-Yin Lai
- Department of Microbiology, National Taiwan University College of Medicine, Taipei
| | - Jin-Town Wang
- Department of Microbiology, National Taiwan University College of Medicine, Taipei.,Department of Internal Medicine, National Taiwan University Hospital, Taipei
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46
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Tipping MJ, Gibbs KA. Peer pressure from a Proteus mirabilis self-recognition system controls participation in cooperative swarm motility. PLoS Pathog 2019; 15:e1007885. [PMID: 31323074 PMCID: PMC6682164 DOI: 10.1371/journal.ppat.1007885] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2019] [Revised: 08/05/2019] [Accepted: 06/03/2019] [Indexed: 11/25/2022] Open
Abstract
Colonies of the opportunistic pathogen Proteus mirabilis can distinguish self from non-self: in swarming colonies of two different strains, one strain excludes the other from the expanding colony edge. Predominant models characterize bacterial kin discrimination as immediate antagonism towards non-kin cells, typically through delivery of toxin effector molecules from one cell into its neighbor. Upon effector delivery, receiving cells must either neutralize it by presenting a cognate anti-toxin as would a clonal sibling, or suffer cell death or irreversible growth inhibition as would a non-kin cell. Here we expand this paradigm to explain the non-lethal Ids self-recognition system, which stops access to a social behavior in P. mirabilis by selectively and transiently inducing non-self cells into a growth-arrested lifestyle incompatible with cooperative swarming. This state is characterized by reduced expression of genes associated with protein synthesis, virulence, and motility, and also causes non-self cells to tolerate previously lethal concentrations of antibiotics. We show that temporary activation of the stringent response is necessary for entry into this state, ultimately resulting in the iterative exclusion of non-self cells as a swarm colony migrates outwards. These data clarify the intricate connection between non-lethal recognition and the lifecycle of P. mirabilis swarm colonies. A resident of animal intestines, Proteus mirabilis is a major cause of catheter-associated urinary tract infections and can cause recurrent, persistent infections. Swarming, which is a collective behavior that promotes centimeter-scale population migration, is implicated in colonization of bladders and kidneys. A regulatory factor of swarming is kin recognition, which involves the transfer of a self-identity protein from one cell into a physically adjacent neighboring cell. However, how kin recognition regulates swarming was previously unclear. We have now shown a mechanism linking kin recognition, swarm migration, and antibiotics tolerance: cells induce a transient antibiotics-tolerant, persister-like state in adjacent non-identical cells which in turn prevents non-identical cells from continuing to participate in collective swarming. These affected non-identical cells continue to exhibit large-scale gene expression suggesting an active shift into a different expression state. These data provide two key insights for the field. First, kin recognition can be a regulatory mechanism that acts with spatial and temporal precision. Second, induction into an antibiotics-tolerant state, instead of occurring stochastically, can be physically and spatially regulated by neighboring cells. These insights highlight the importance of further developing four-dimensional (time and X-, Y-, Z-axes) model systems for interrogating cell-cell signaling and control in microbial populations.
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Affiliation(s)
- Murray J. Tipping
- Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts, United States of America
| | - Karine A. Gibbs
- Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts, United States of America
- * E-mail:
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47
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Liebl D, Robert-Genthon M, Job V, Cogoni V, Attrée I. Baseplate Component TssK and Spatio-Temporal Assembly of T6SS in Pseudomonas aeruginosa. Front Microbiol 2019; 10:1615. [PMID: 31379775 PMCID: PMC6657622 DOI: 10.3389/fmicb.2019.01615] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2019] [Accepted: 06/28/2019] [Indexed: 11/22/2022] Open
Abstract
The Gram-negative bacteria use the contractile multi-molecular structure, called the Type VI Secretion System (T6SS) to inject toxic products into eukaryotic and prokaryotic cells. In this study, we use fluorescent protein fusions and time-lapse microscopy imaging to study the assembly dynamics of the baseplate protein TssK in Pseudomonas aeruginosa T6SS. TssK formed transient higher-order structures that correlated with dynamics of sheath component TssB. Assembly of peri-membrane TssK structures occurred de novo upon contact with competing bacteria. We show that this assembly required presence of TagQ-TagR envelope sensors, activity of PpkA kinase and anchoring to the inner membrane via TssM. Disassembly and repositioning of TssK component was dependent on PppA phosphatase and indispensable for repositioning and deployment of the entire contractile apparatus toward a new target cell. We also show that TssE is necessary for correct elongation and stability of TssB-sheath, but not for TssK assembly. Therefore, in P. aeruginosa, assembly of the TssK-containing structure relays on the post-translational regulatory envelope module and acts as spatio-temporal marker for further recruitment and subsequent assembly of the contractile apparatus.
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Affiliation(s)
- David Liebl
- Univ. Grenoble Alpes, CNRS, Bacterial Pathogenesis and Cellular Responses, ERL 5261, INSERM, UMR-S 1036, CEA, Grenoble, France
| | - Mylène Robert-Genthon
- Univ. Grenoble Alpes, CNRS, Bacterial Pathogenesis and Cellular Responses, ERL 5261, INSERM, UMR-S 1036, CEA, Grenoble, France
| | - Viviana Job
- Univ. Grenoble Alpes, CNRS, Bacterial Pathogenesis and Cellular Responses, ERL 5261, INSERM, UMR-S 1036, CEA, Grenoble, France.,Univ. Grenoble Alpes, CEA, CNRS, Institut de Biologie Structurale (IBS), Grenoble, France
| | - Valentina Cogoni
- Univ. Grenoble Alpes, CNRS, Bacterial Pathogenesis and Cellular Responses, ERL 5261, INSERM, UMR-S 1036, CEA, Grenoble, France
| | - Ina Attrée
- Univ. Grenoble Alpes, CNRS, Bacterial Pathogenesis and Cellular Responses, ERL 5261, INSERM, UMR-S 1036, CEA, Grenoble, France
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48
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Li L, Yuan L, Shi Y, Xie X, Chai A, Wang Q, Li B. Comparative genomic analysis of Pectobacterium carotovorum subsp. brasiliense SX309 provides novel insights into its genetic and phenotypic features. BMC Genomics 2019; 20:486. [PMID: 31195968 PMCID: PMC6567464 DOI: 10.1186/s12864-019-5831-x] [Citation(s) in RCA: 26] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2018] [Accepted: 05/23/2019] [Indexed: 12/20/2022] Open
Abstract
Background Pectobacterium carotovorum subsp. brasiliense is a broad host range bacterial pathogen, which causes blackleg of potatoes and bacterial soft rot of vegetables worldwide. Production of plant cell wall degrading enzymes is usually critical for Pectobacterium infection. However, other virulence factors and the mechanisms of genetic adaptation still need to be studied in detail. Results In this study, the complete genome of P. carotovorum subsp. brasiliense strain SX309 isolated from cucumber was compared with eight other pathogenic bacteria belonging to the Pectobacterium genus, which were isolated from various host plants. Genome comparison revealed that most virulence genes are highly conserved in the Pectobacterium strains, especially for the key virulence determinants involved in the biosynthesis of extracellular enzymes and others including the type II and III secretion systems, quorum sensing system, flagellar and chemotactic genes. Nevertheless, some variable regions of the T6SS and the CRISP-Cas immune system are unique for P. carotovorum subsp. brasiliense. Conclusions The extensive comparative genomics analysis revealed highly conserved virulence genes in the Pectobacterium strains. However, several variable regions of type VI secretion system and two subtype Cas mechanism-Cas immune systems possibly contribute to the process of Pectobacterium infection and adaptive immunity. Electronic supplementary material The online version of this article (10.1186/s12864-019-5831-x) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Lei Li
- Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, 100081, China
| | - Lifang Yuan
- Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.,Department of Plant Pathology, College of Plant Protection, China Agricultural University, Beijing, 100193, China
| | - Yanxia Shi
- Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, 100081, China
| | - Xuewen Xie
- Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, 100081, China
| | - Ali Chai
- Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, 100081, China
| | - Qi Wang
- Department of Plant Pathology, College of Plant Protection, China Agricultural University, Beijing, 100193, China
| | - Baoju Li
- Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
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Xu Z, Xie J, Soteyome T, Peters BM, Shirtliff ME, Liu J, Harro JM. Polymicrobial interaction and biofilms between Staphylococcus aureus and Pseudomonas aeruginosa: an underestimated concern in food safety. Curr Opin Food Sci 2019. [DOI: 10.1016/j.cofs.2019.03.006] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/27/2022]
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50
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Vacheron J, Péchy-Tarr M, Brochet S, Heiman CM, Stojiljkovic M, Maurhofer M, Keel C. T6SS contributes to gut microbiome invasion and killing of an herbivorous pest insect by plant-beneficial Pseudomonas protegens. ISME JOURNAL 2019; 13:1318-1329. [PMID: 30683920 DOI: 10.1038/s41396-019-0353-8] [Citation(s) in RCA: 62] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/25/2018] [Revised: 01/09/2019] [Accepted: 01/09/2019] [Indexed: 12/31/2022]
Abstract
Pseudomonas protegens are multi-talented plant-colonizing bacteria that suppress plant pathogens and stimulate plant defenses. In addition, they are capable of invading and killing agriculturally important plant pest insects that makes them promising candidates for biocontrol applications. Here we assessed the role of type VI secretion system (T6SS) components of type strain CHA0 during interaction with larvae of the cabbage pest Pieris brassicae. We show that the T6SS core apparatus and two VgrG modules, encompassing the respective T6SS spikes (VgrG1a and VgrG1b) and associated effectors (RhsA and Ghh1), contribute significantly to insect pathogenicity of P. protegens in oral infection assays but not when bacteria are injected directly into the hemolymph. Monitoring of the colonization levels of P. protegens in the gut, hemolymph, and excrements of the insect larvae revealed that the invader relies on T6SS and VgrG1a module function to promote hemocoel invasion. A 16S metagenomic analysis demonstrated that T6SS-supported invasion by P. protegens induces significant changes in the insect gut microbiome affecting notably Enterobacteriaceae, a dominant group of the commensal gut bacteria. Our study supports the concept that pathogens deploy T6SS-based strategies to disrupt the commensal microbiota in order to promote host colonization and pathogenesis.
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Affiliation(s)
- Jordan Vacheron
- Department of Fundamental Microbiology, University of Lausanne, Lausanne, Switzerland
| | - Maria Péchy-Tarr
- Department of Fundamental Microbiology, University of Lausanne, Lausanne, Switzerland
| | - Silvia Brochet
- Department of Fundamental Microbiology, University of Lausanne, Lausanne, Switzerland
| | - Clara Margot Heiman
- Department of Fundamental Microbiology, University of Lausanne, Lausanne, Switzerland
| | - Marina Stojiljkovic
- Department of Fundamental Microbiology, University of Lausanne, Lausanne, Switzerland
| | - Monika Maurhofer
- Plant Pathology, Institute of Integrative Biology, Swiss Federal Institute of Technology (ETH) Zurich, Zurich, Switzerland.
| | - Christoph Keel
- Department of Fundamental Microbiology, University of Lausanne, Lausanne, Switzerland.
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