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.
The Borrelia genus consists of spirochete bacteria known to cause Lyme disease (LD) and relapsing fever in humans. Borrelia pathogens are commonly transmitted via arthropod vectors such as ticks, mites, or lice. Here, we report the molecular screening of LD group Borrelia sp. from ticks (Acari: Ixodidae) collected from rodents trapped in recreational forests and a semiurban residential area in the Selangor state in Malaysia. Of 156 adult ticks surveyed, 72 ticks were determined as positive for Borrelia sp. by polymerase chain reaction (PCR). All Borrelia PCR-positive ticks belonged to the Ixodes granulatus Supino species. Borrelia sp. was not detected in other tick species examined, including Dermacentor sp. and Amblyomma sp. ticks. Thirteen Borrelia PCR-positive tick samples were selected for further sequence analyses. Phylogenetic analyses of partial flaB gene sequences revealed that the Borrelia sp. were closely related to the LD group borreliae, Borrelia yangtzensis; a novel Borrelia genospecies reported in East Asian countries including Japan, Taiwan, and China. To our knowledge, this is the first report of Borrelia sp. related to Borrelia yangtzensis detected in Malaysia and Southeast Asia. The zoonotic potential of the Borrelia sp. reported here merits further investigation, as it may explain the previously reported serological evidence for borrelial infections in Malaysia.
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