Transstadial and intrastadial experimental transmission of Ehrlichia canis by male Rhipicephalus sanguineus

Abstract

The acquisition and transmission of rickettsial pathogens by different tick developmental stages has important epidemiological implications. The purpose of this study was to determine if male Rhipicephalus sanguineus can experimentally acquire and transmit Ehrlichia canis in the absence of female ticks. Two trials were performed where nymphal and male R. sanguineus were simultaneously acquisition fed on the same infected donor hosts, and transstadially or intrastadially exposed male ticks were fed on separate pathogen-free dogs as a test for transmission. A single-step p30-based PCR assay was used to test canine and tick hosts for E. canis infections before and after tick feeding. E. canis was detected after either intrastadial or transstadial passage in male ticks, the organism remained detectable in both tick groups after transmission feeding, and both tick groups transmitted the rickettsia to susceptible dogs. Infection of dogs via tick feeding resulted in milder clinical signs and lower antibody titers than intravenous inoculation of carrier blood, but further investigation is needed to understand the mechanisms responsible for this observation. These results demonstrate that male R. sanguineus can take multiple feedings, and that they can both acquire and transmit E. canis in the absence of female ticks. This tick development stage could be important in transmission of E. canis, and perhaps related pathogens, between vertebrate hosts under natural and experimental conditions.

Tick Acquisition of Ehrlichia canis from Dogs Treated with Doxycycline Hyclate

“…All of the dogs tested negative by PCR and immunofluorescence assays prior to tick feeding, and transmission was confirmed by seroconversion, mild clinical signs, and PCR-positive buffy coats after feeding PCR-positive ticks on these dogs (5). The dog referred to as AXM, which was not exposed to E. canis, served as a negative control for feeding cohorts of the ticks used in this study (5). All of the dogs were cared for in accordance with a protocol on file with The Ohio State University Institutional Laboratory Animal Care and Use Committee.…”

“…Four Beagle dogs were experimentally infected with E. canis (Ebony isolate) by tick transmission that was demonstrated in a previous study (5). All of the dogs tested negative by PCR and immunofluorescence assays prior to tick feeding, and transmission was confirmed by seroconversion, mild clinical signs, and PCR-positive buffy coats after feeding PCR-positive ticks on these dogs (5). The dog referred to as AXM, which was not exposed to E. canis, served as a negative control for feeding cohorts of the ticks used in this study (5).…”

“…Canine blood (5 ml) was collected with EDTA, and buffy coats were separated as previously described (19). Buffy coats and ticks were incubated in protein digestion buffer and subjected to phenol-chloroform extraction and ethanol precipitation as reported by Bremer et al (5), except the digestion buffer contained 2% sodium dodecyl sulfate. PCR assays with primers ECA30-384S and ECA30-583A were exactly as previously described (5).…”

“…For xenodiagnosis, approximately 200 R. sanguineus nymphs were placed on each dog 7 days after treatment and allowed to feed to repletion and to molt into adults (5,19). Cohorts of these nymphs were fed on untreated dogs and an uninfected dog as positive and negative controls, respectively.…”

The glycoprotein TRP36 of Ehrlichia sp. UFMG-EV and related cattle pathogen Ehrlichia sp. UFMT-BV evolved from a highly variable clade of E. canis under adaptive diversifying selection

“…UFMT-BV are closely related to E. canis, however they were associated to new invertebrate and vertebrate hosts, respectively. First, while the common tick vector for E. canis is R. sanguineus [14], Ehrlichia sp. UFMG-EV was isolated from R. microplus hemolymph [2].…”