Tad pilus-mediated twitching motility is essential for DNA uptake and survival of Liberibacters

PLoS One. 2021 Oct 13;16(10):e0258583. doi: 10.1371/journal.pone.0258583. eCollection 2021.

Abstract

Axenically cultured Liberibacter crescens (Lcr) is a closely related surrogate for uncultured plant pathogenic species of the genus Liberibacter, including 'Candidatus L. asiaticus' (CLas) and 'Ca. L. solanacearum' (CLso). All Liberibacters encode a completely conserved gene repertoire for both flagella and Tad (Tight Adherence) pili and all are missing genes critical for nucleotide biosynthesis. Both flagellar swimming and Tad pilus-mediated twitching motility in Lcr were demonstrated for the first time. A role for Tad pili in the uptake of extracellular dsDNA for food in Liberibacters was suspected because both twitching and DNA uptake are impossible without repetitive pilus extension and retraction, and no genes encoding other pilus assemblages or mechanisms for DNA uptake were predicted to be even partially present in any of the 35 fully sequenced Liberibacter genomes. Insertional mutations of the Lcr Tad pilus genes cpaA, cpaB, cpaE, cpaF and tadC all displayed such severely reduced growth and viability that none could be complemented. A mutation affecting cpaF (motor ATPase) was further characterized and the strain displayed concomitant loss of twitching, viability and reduced periplasmic uptake of extracellular dsDNA. Mutations of comEC, encoding the inner membrane competence channel, had no effect on either motility or growth but completely abolished natural transformation in Lcr. The comEC mutation was restored by complementation using comEC from Lcr but not from CLas strain psy62 or CLso strain RS100, indicating that unlike Lcr, these pathogens were not naturally competent for transformation. This report provides the first evidence that the Liberibacter Tad pili are dynamic and essential for both motility and DNA uptake, thus extending their role beyond surface adherence.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adenosine Triphosphatases / chemistry
  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / metabolism
  • Amino Acid Sequence
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Citrus / microbiology
  • DNA, Bacterial / metabolism*
  • Fimbriae, Bacterial / genetics
  • Fimbriae, Bacterial / physiology
  • Liberibacter / genetics
  • Liberibacter / growth & development
  • Liberibacter / physiology
  • Mutagenesis, Site-Directed
  • Plant Diseases / microbiology
  • Sequence Alignment

Substances

  • Bacterial Proteins
  • DNA, Bacterial
  • Adenosine Triphosphatases

Supplementary concepts

  • Liberibacter crescens

Grants and funding

This work was supported by the USDA National Institute of Food and Agriculture; Specialty Crops Research Initiative (NIFA-SCRI) grant #2016-70016-24844 to DWG and KMJ. https://nifa.usda.gov/funding-opportunity/specialty-crop-research-initiative-scri. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.