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Motility characteristics are altered for Rickettsia bellii transformed to overexpress a heterologous rickA gene.

ABSTRACT: The rickettsial protein RickA activates host cell factors associated with the eukaryotic actin cytoskeleton and is likely involved with rickettsial host cell binding and infection and the actin-based motility of spotted fever group rickettsiae. The rickA gene sequence and protein vary substantially between Rickettsia species, as do observed motility-associated phenotypes. To help elucidate the function of RickA and determine the effects of species-specific RickA variations, we compared extracellular binding, intracellular motility, and intercellular spread phenotypes of three Rickettsia bellii variants. These included two shuttle vector-transformed R. bellii strains and the wild-type isolate from which they were derived, R. bellii RML 369C. Both plasmid shuttle vectors carried spectinomycin resistance and a GFPuv reporter; one contained Rickettsia monacensis-derived rickA, and the other lacked the rickA gene. Rickettsia bellii transformed to express R. monacensis rickA highly overexpressed this transcript in comparison to its native rickA. These rickettsiae also moved at higher velocities and followed a more curved path than the negative-control transformants. A lower proportion of R. monacensis rickA-expressing bacteria ever became motile, however, and they formed smaller plaques.

SUBMITTER: Oliver JD

PROVIDER: S-EPMC3911190 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Motility characteristics are altered for Rickettsia bellii transformed to overexpress a heterologous rickA gene.

Oliver Jonathan D JD Burkhardt Nicole Y NY Felsheim Roderick F RF Kurtti Timothy J TJ Munderloh Ulrike G UG

Applied and environmental microbiology 20131202 3

The rickettsial protein RickA activates host cell factors associated with the eukaryotic actin cytoskeleton and is likely involved with rickettsial host cell binding and infection and the actin-based motility of spotted fever group rickettsiae. The rickA gene sequence and protein vary substantially between Rickettsia species, as do observed motility-associated phenotypes. To help elucidate the function of RickA and determine the effects of species-specific RickA variations, we compared extracell ...[more]

PMID: 24296498

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Project description:BACKGROUND: The purpose of this study was to identify the presence of rickettsia and hantavirus in wild rodents and arthropods in response to an outbreak of acute unidentified febrile illness among Indians in the Halataikwa Indian Reserve, northwest of the Mato Grosso state, in the Brazilian Amazon. Where previously surveillance data showed serologic evidence of rickettsia and hantavirus human infection. METHODS: The arthropods were collected from the healthy Indian population and by flagging vegetation in grassland or woodland along the peridomestic environment of the Indian reserve. Wild rodents were live-trapped in an area bordering the reserve limits, due the impossibility of capturing wild animals in the Indian reserve. The wild rodents were identified based on external and cranial morphology and karyotype. DNA was extracted from spleen or liver samples of rodents and from invertebrate (tick and louse) pools, and the molecular characterization of the rickettsia was through PCR and DNA sequencing of fragments of two rickettsial genes (gltA and ompA). In relation to hantavirus, rodent serum samples were serologically screened by IgG ELISA using the Araraquara-N antigen and total RNA was extracted from lung samples of IgG-positive rodents. The amplification of the complete S segment was performed. RESULTS: A total of 153 wild rodents, 121 louse, and 36 tick specimens were collected in 2010. Laguna Negra hantavirus was identified in Calomys callidus rodents and Rickettsia bellii, Rickettsia amblyommii were identified in Amblyomma cajennense ticks. CONCLUSIONS: Zoonotic diseases such as HCPS and spotted fever rickettsiosis are a public health threat and should be considered in outbreaks and acute febrile illnesses among Indian populations. The presence of the genome of rickettsias and hantavirus in animals in this Indian reserve reinforces the need to include these infectious agents in outbreak investigations of febrile cases in Indian populations.

Project description:Members of the genus Rickettsia range from nonpathogenic endosymbionts to virulent pathogens such as Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever. Many rickettsiae are considered nonpathogenic because they have been isolated from ticks but not vertebrate hosts. We assessed the ability of three presumed endosymbionts: Rickettsia amblyommatis, Rickettsia bellii, and Rickettsia montanensis, to infect a guinea pig animal model. These species were chosen because of their high prevalence in respective tick vectors or published reports suggestive of human or animal pathogenicity. Following intraperitoneal (IP) inoculation of cell culture suspensions of R. rickettsii, R. amblyommatis, R. bellii, or R. montanensis into guinea pigs, animals were monitored for signs of clinical illness for 13 days. Ear biopsies and blood samples were taken at 2- to 3-day intervals for detection of rickettsial DNA by PCR. Animals were necropsied and internal organ samples were also tested using PCR assays. Among the six guinea pigs inoculated with R. amblyommatis, fever, orchitis, and dermatitis were observed in one, one, and three animals respectively. In R. bellii-exposed animals, we noted fever in one of six animals, orchitis in one, and dermatitis in two. No PCR-positive tissues were present in either the R. amblyommatis- or R. bellii-exposed groups. In the R. montanensis-exposed group, two of six animals became febrile, two had orchitis, and three developed dermatitis in ears or footpads. R. montanensis DNA was detected in ear skin biopsies collected on multiple days from three animals. Also, a liver specimen from one animal and spleen specimens of two animals were PCR positive. The course and severity of disease in the three experimental groups were significantly milder than that of R. rickettsii. This study suggests that the three rickettsiae considered nonpathogenic can cause either subclinical or mild infections in guinea pigs when introduced via IP inoculation.

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Motility characteristics are altered for Rickettsia bellii transformed to overexpress a heterologous rickA gene.

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Motility characteristics are altered for Rickettsia bellii transformed to overexpress a heterologous rickA gene.

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