1887

Abstract

Type IV pili are involved in adhesion, twitching motility, aggregation, biofilm formation and virulence in a variety of Gram-negative bacteria. the causative agent of melioidosis and a Tier 1 biological select agent, is a Gram-negative bacterium with eight type IV pili-associated loci (TFP1 to TFP8). Most have not been fully characterized. In this study, we investigated , an uncharacterized TFP8 gene that encodes a type IVB pilus protein subunit. Using genetic deletion and complementation analysis in JW270, we demonstrate that plays an important role in twitching motility and adhesion to A549 human alveolar epithelial cells. Compared to JW270, the JW270 mutant failed to display twitching motility and did not adhere to the epithelial cells. These phenotypes were partially reversed by the complementation of in the mutant strain. The study also shows that is expressed only during the onset of mature biofilm formation and at the dispersal of a biofilm, suggesting that the motility characteristic is required to form a biofilm. Our study is the first to suggest that the gene in TFP8 contributes to twitching motility, adhesion and biofilm formation, indicating that the gene may contribute to virulence.

Funding
This study was supported by the:
  • Defense Threat Reduction Agency (Award CB10207)
    • Principle Award Recipient: DavidDeShazer
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License.
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.001150
2022-03-16
2022-05-18
Loading full text...

Full text loading...

/deliver/fulltext/micro/168/3/mic001150.html?itemId=/content/journal/micro/10.1099/mic.0.001150&mimeType=html&fmt=ahah

References

  1. Wiersinga WJ, Virk HS, Torres AG, Currie BJ, Peacock SJ et al. Melioidosis. Nat Rev Dis Primers 2018; 4:17107 [View Article] [PubMed]
    [Google Scholar]
  2. Limmathurotsakul D, Golding N, Dance DAB, Messina JP, Pigott DM et al. Predicted global distribution of Burkholderia pseudomallei and burden of melioidosis. Nat Microbiol 2016; 1:15008 [View Article] [PubMed]
    [Google Scholar]
  3. Limmathurotsakul D, Wongratanacheewin S, Teerawattanasook N, Wongsuvan G, Chaisuksant S et al. Increasing incidence of human melioidosis in Northeast Thailand. Am J Trop Med Hyg 2010; 82:1113–1117 [View Article] [PubMed]
    [Google Scholar]
  4. Hahn HP. The type-4 pilus is the major virulence-associated adhesin of Pseudomonas aeruginosa-a review. Gene 1997; 192:99–108 [View Article] [PubMed]
    [Google Scholar]
  5. Pizarro-Cerdá J, Cossart P. Bacterial adhesion and entry into host cells. Cell 2006; 124:715–727 [View Article] [PubMed]
    [Google Scholar]
  6. Lukaszczyk M, Pradhan B, Remaut H. The biosynthesis and structures of bacterial pili. Subcell Biochem 2019; 92:369–413 [View Article] [PubMed]
    [Google Scholar]
  7. Craig L, Forest KT, Maier B. Type IV pili: dynamics, biophysics and functional consequences. Nat Rev Microbiol 2019; 17:429–440 [View Article] [PubMed]
    [Google Scholar]
  8. Strom MS, Lory S. Structure-function and biogenesis of the type IV pili. Annu Rev Microbiol 1993; 47:565–596 [View Article] [PubMed]
    [Google Scholar]
  9. Essex-Lopresti AE, Boddey JA, Thomas R, Smith MP, Hartley MG et al. A type IV pilin, PilA, Contributes To Adherence of Burkholderia pseudomallei and virulence in vivo. Infect Immun 2005; 73:1260–1264 [View Article] [PubMed]
    [Google Scholar]
  10. Boddey JA, Flegg CP, Day CJ, Beacham IR, Peak IR. Temperature-regulated microcolony formation by Burkholderia pseudomallei requires pilA and enhances association with cultured human cells. Infect Immun 2006; 74:5374–5381 [View Article] [PubMed]
    [Google Scholar]
  11. Nandi T, Ong C, Singh AP, Boddey J, Atkins T et al. A genomic survey of positive selection in Burkholderia pseudomallei provides insights into the evolution of accidental virulence. PLoS Pathog 2010; 6:e1000845 [View Article] [PubMed]
    [Google Scholar]
  12. Sangdee K, Waropastrakul S, Wongratanachewin S, Homchampa P. Heterologously type IV pilus expressed protein of Burkholderia pseudomallei is immunogenic but fails to induce protective immunity in mice. Southeast Asian J Trop Med Public Health 2011; 42:1190–1196 [PubMed]
    [Google Scholar]
  13. Nandi T, Holden MTG, Holden MTG, Didelot X, Mehershahi K et al. Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles. Genome Res 2015; 25:129–141 [View Article] [PubMed]
    [Google Scholar]
  14. Krachler AM, Orth K. Targeting the bacteria-host interface: strategies in anti-adhesion therapy. Virulence 2013; 4:284–294 [View Article] [PubMed]
    [Google Scholar]
  15. Okaro U, Green R, Mohapatra S, Anderson B. The trimeric autotransporter adhesin BadA is required for in vitro biofilm formation by Bartonella henselae. NPJ Biofilms Microbiomes 2019; 5:10 [View Article] [PubMed]
    [Google Scholar]
  16. Locht C, Bertin P, Menozzi FD, Renauld G. The filamentous haemagglutinin, a multifaceted adhesion produced by virulent Bordetella spp. Mol Microbiol 1993; 9:653–660 [View Article] [PubMed]
    [Google Scholar]
  17. Guo H, Yi W, Song JK, Wang PG. Current understanding on biosynthesis of microbial polysaccharides. Curr Top Med Chem 2008; 8:141–151 [View Article] [PubMed]
    [Google Scholar]
  18. Berne C, Ducret A, Hardy GG, Brun YV. Adhesins involved in attachment to abiotic surfaces by gram-negative bacteria. Microbiol Spectr 2015; 3: [View Article] [PubMed]
    [Google Scholar]
  19. Flemming HC, Wingender J. The biofilm matrix. Nat Rev Microbiol 2010; 8:623–633 [View Article] [PubMed]
    [Google Scholar]
  20. Ruhal R, Kataria R. Biofilm patterns in gram-positive and gram-negative bacteria. Microbiol Res 2021; 251:126829 [View Article] [PubMed]
    [Google Scholar]
  21. Sharma V, von Ossowski I, Krishnan V. Exploiting pilus-mediated bacteria-host interactions for health benefits. Mol Aspects Med 2021; 81:100998 [View Article] [PubMed]
    [Google Scholar]
  22. Das T, Sehar S, Koop L, Wong YK, Ahmed S et al. Influence of calcium in extracellular DNA mediated bacterial aggregation and biofilm formation. PLoS One 2014; 9:e91935 [View Article] [PubMed]
    [Google Scholar]
  23. Whitchurch CB, Tolker-Nielsen T, Ragas PC, Mattick JS. Extracellular DNA required for bacterial biofilm formation. Science 2002; 295:1487 [View Article] [PubMed]
    [Google Scholar]
  24. Ma L, Jackson KD, Landry RM, Parsek MR, Wozniak DJ. Analysis of Pseudomonas aeruginosa conditional psl variants reveals roles for the psl polysaccharide in adhesion and maintaining biofilm structure postattachment. J Bacteriol 2006; 188:8213–8221 [View Article] [PubMed]
    [Google Scholar]
  25. Begun J, Gaiani JM, Rohde H, Mack D, Calderwood SB et al. Staphylococcal biofilm exopolysaccharide protects against Caenorhabditis elegans immune defenses. PLoS Pathog 2007; 3:e57 [View Article] [PubMed]
    [Google Scholar]
  26. Chin CY, Hara Y, Ghazali AK, Yap SJ, Kong C et al. Global transcriptional analysis of Burkholderia pseudomallei high and low biofilm producers reveals insights into biofilm production and virulence. BMC Genomics 2015; 16:471 [View Article] [PubMed]
    [Google Scholar]
  27. Joshua GWP, Karlyshev AV, Smith MP, Isherwood KE, Titball RW et al. A Caenorhabditis elegans model of Yersinia infection: biofilm formation on a biotic surface. Microbiology (Reading) 2003; 149:3221–3229 [View Article] [PubMed]
    [Google Scholar]
  28. O’Toole GA, Kolter R. Flagellar and twitching motility are necessary for Pseudomonas aeruginosa biofilm development. Mol Microbiol 1998; 30:295–304 [View Article] [PubMed]
    [Google Scholar]
  29. Otton LM, da Silva Campos M, Meneghetti KL, Corção G. Influence of twitching and swarming motilities on biofilm formation in Pseudomonas strains. Arch Microbiol 2017; 199:677–682 [View Article] [PubMed]
    [Google Scholar]
  30. Wang T, Guan W, Huang Q, Yang Y, Yan W et al. Quorum-sensing contributes to virulence, twitching motility, seed attachment and biofilm formation in the wild type strain Aac-5 of Acidovorax citrulli. Microb Pathog 2016; 100:133–140 [View Article] [PubMed]
    [Google Scholar]
  31. Pu M, Storms E, Chodur DM, Rowe-Magnus DA. Calcium-dependent site-switching regulates expression of the atypical iam pilus locus in Vibrio vulnificus. Environ Microbiol 2020; 22:4167–4182 [View Article] [PubMed]
    [Google Scholar]
  32. Rose SJ, Bermudez LE. Identification of bicarbonate as a trigger and genes involved with extracellular DNA export in mycobacterial biofilms. mBio 2016; 7:e01597-16 [View Article] [PubMed]
    [Google Scholar]
  33. Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 2014; 30:2114–2120 [View Article] [PubMed]
    [Google Scholar]
  34. Hamad MA, Zajdowicz SL, Holmes RK, Voskuil MI. An allelic exchange system for compliant genetic manipulation of the select agents Burkholderia pseudomallei and Burkholderia mallei. Gene 2009; 430:123–131 [View Article] [PubMed]
    [Google Scholar]
  35. Schell MA, Ulrich RL, Ribot WJ, Brueggemann EE, Hines HB et al. Type VI secretion is a major virulence determinant in Burkholderia mallei. Mol Microbiol 2007; 64:1466–1485 [View Article] [PubMed]
    [Google Scholar]
  36. O’Toole GA. Microtiter dish biofilm formation assay. J Vis Exp 2011; 2011:47 [View Article] [PubMed]
    [Google Scholar]
  37. Rosenberg M, Azevedo NF, Ivask A. Propidium iodide staining underestimates viability of adherent bacterial cells. Sci Rep 2019; 9:6483 [View Article] [PubMed]
    [Google Scholar]
  38. Ribet D, Cossart P. How bacterial pathogens colonize their hosts and invade deeper tissues. Microbes Infect 2015; 17:173–183 [View Article] [PubMed]
    [Google Scholar]
  39. Pakkulnan R, Anutrakunchai C, Kanthawong S, Taweechaisupapong S, Chareonsudjai P et al. Extracellular DNA facilitates bacterial adhesion during Burkholderia pseudomallei biofilm formation. PLoS One 2019; 14:e0213288 [View Article] [PubMed]
    [Google Scholar]
  40. Ofek I, Hasty DL, Sharon N. Anti-adhesion therapy of bacterial diseases: prospects and problems. FEMS Immunol Med Microbiol 2003; 38:181–191 [View Article] [PubMed]
    [Google Scholar]
  41. Qvortrup K, Hultqvist LD, Nilsson M, Jakobsen TH, Jansen CU et al. Small molecule anti-biofilm agents developed on the basis of mechanistic understanding of biofilm formation. Front Chem 2019; 7:742 [View Article] [PubMed]
    [Google Scholar]
  42. Kunyanee C, Kamjumphol W, Taweechaisupapong S, Kanthawong S, Wongwajana S et al. Burkholderia pseudomallei biofilm promotes adhesion, internalization and stimulates proinflammatory cytokines in human epithelial A549 cells. PLoS One 2016; 11:e0160741 [View Article] [PubMed]
    [Google Scholar]
  43. Mattick JS. Type IV pili and twitching motility. Annu Rev Microbiol 2002; 56:289–314 [View Article] [PubMed]
    [Google Scholar]
  44. Rashid MH, Kornberg A. Inorganic polyphosphate is needed for swimming, swarming, and twitching motilities of Pseudomonas aeruginosa. Proc Natl Acad Sci U S A 2000; 97:4885–4890 [View Article] [PubMed]
    [Google Scholar]
  45. Mattingly AE, Weaver AA, Dimkovikj A, Shrout JD. Assessing travel conditions: environmental and host influences on bacterial surface motility. J Bacteriol 2018; 200:e00014–18 [View Article] [PubMed]
    [Google Scholar]
  46. Han X, Kennan RM, Davies JK, Reddacliff LA, Dhungyel OP et al. Twitching motility is essential for virulence in Dichelobacter nodosus. J Bacteriol 2008; 190:3323–3335 [View Article] [PubMed]
    [Google Scholar]
  47. Chiang P, Burrows LL. Biofilm formation by hyperpiliated mutants of Pseudomonas aeruginosa. J Bacteriol 2003; 185:2374–2378 [View Article] [PubMed]
    [Google Scholar]
  48. Willcocks SJ, Denman C, Cia F, McCarthy E, Cuccui J et al. Virulence of the emerging pathogen, Burkholderia pseudomallei, depends upon the O-linked oligosaccharyltransferase, PglL. Future Microbiol 2020; 15:241–257 [View Article] [PubMed]
    [Google Scholar]
  49. Schreiner HC, Sinatra K, Kaplan JB, Furgang D, Kachlany SC et al. Tight-adherence genes of Actinobacillus actinomycetemcomitans are required for virulence in a rat model. Proc Natl Acad Sci U S A 2003; 100:7295–7300 [View Article] [PubMed]
    [Google Scholar]
  50. Fine DH, Furgang D, Kaplan J, Charlesworth J, Figurski DH. Tenacious adhesion of Actinobacillus actinomycetemcomitans strain CU1000 to salivary-coated hydroxyapatite. Arch Oral Biol 1999; 44:1063–1076 [View Article] [PubMed]
    [Google Scholar]
  51. Simon R, Priefer U, Pühler A. A Broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in gram negative bacteria. Nat Biotechnol 1983; 1:784–791 [View Article]
    [Google Scholar]
  52. Warawa JM, Long D, Rosenke R, Gardner D, Gherardini FC. Role for the Burkholderia pseudomallei capsular polysaccharide encoded by the wcb operon in acute disseminated melioidosis. Infect Immun 2009; 77:5252–5261 [View Article] [PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.001150
Loading
/content/journal/micro/10.1099/mic.0.001150
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error