Isolation and propagation of laboratory strains and a novel flea-derived field strain of Wolbachia in tick cell lines

Jing Jing Khoo, Timothy J. Kurtti, Nurul Aini Husin, Alexandra Beliavskaia, Fang Shiang Lim, Mulya Mustika Sari Zulkifli, Alaa M. Al-Khafaji, Catherine Hartley, Alistair C. Darby, Grant L. Hughes, Sazaly Abubakar, Benjamin L. Makepeace, Lesley Bell-Sakyi

Research output: Contribution to journalArticlepeer-review

3 Scopus citations


Wolbachia are intracellular endosymbionts of several invertebrate taxa, including insects and nematodes. Although Wolbachia DNA has been detected in ticks, its presence is generally associated with parasitism by insects. To determine whether or not Wolbachia can infect and grow in tick cells, cell lines from three tick species, Ixodes scapularis, Ixodes ricinus and Rhipicephalus microplus, were inoculated with Wolbachia strains wStri and wAlbB isolated from mosquito cell lines. Homogenates prepared from fleas collected from cats in Malaysia were inoculated into an I. scapularis cell line. Bacterial growth and identity were monitored by microscopy and PCR amplification and sequencing of fragments of Wolbachia genes. The wStri strain infected Ixodes spp. cells and was maintained through 29 passages. The wAlbB strain successfully infected Ixodes spp. and R. microplus cells and was maintained through 2–5 passages. A novel strain of Wolbachia belonging to the supergroup F, designated wCfeF, was isolated in I. scapularis cells from a pool of Ctenocephalides sp. cat fleas and maintained in vitro through two passages over nine months. This is the first confirmed isolation of a Wolbachia strain from a flea and the first isolation of any Wolbachia strain outside the “pandemic” A and B supergroups. The study demonstrates that tick cells can host multiple Wolbachia strains, and can be added to panels of insect cell lines to improve success rates in isolation of field strains of Wolbachia.

Original languageEnglish (US)
Article number988
Pages (from-to)1-19
Number of pages19
Issue number7
StatePublished - Jul 2020

Bibliographical note

Funding Information:
Funding: This research was supported by an Institutional Links grant, ID 332192305, under the Newton-Ungku Omar Fund partnership. The grant is funded by the UK Department of Business, Energy and Industrial Strategy (BEIS) and the Energy and Industrial Strategy and Malaysian Industry-Government Group for High Technology (MIGHT), and delivered by the British Council. Support was also provided by the UK BBSRC GCRF grants BB/P024270/1 and BB/P024378/1, and the Higher Institution Centre of Excellence (HICoE) program (Project MO002-2019), Ministry of Education, Malaysia for niche area research. TJK was supported by the University of Minnesota Agricultural Experiment Station. GLH was supported by UK BBSRC grant BB/T001240/1, a Royal Society Wolfson Fellowship (RSWF\R1\180013), US NIH grants R21AI138074 and R21AI129507 and a UK NIHR grant NIHR2000907.

Publisher Copyright:
© 2020 by the authors. Licensee MDPI, Basel, Switzerland.


  • Ctenocephalides
  • Flea
  • In vitro culture
  • Malaysia
  • Phylogeny
  • Tick cell line
  • Wolbachia

PubMed: MeSH publication types

  • Journal Article


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