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

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.     

Detection and molecular characterization of Babesia canis vogeli from naturally infected dogs and Rhipicephalus sanguineus ticks in Tunisia

Canine babesiosis, caused by intra-erythrocytic Babesia, is a tick-borne disease of worldwide importance. No information on canine babesiosis has been documented in Tunisia. Detection and analysis of Babesia species from naturally infected dogs and ticks recovered from dogs were attempted by reverse line blot hybridization and nucleotide sequence analysis based on 18S rRNA gene. Out of 180 blood samples collected from domestic dogs in 4 villages situated in different bioclimatic zones, 12 were positive for Babesia canis vogeli. In addition, a total of 160 Rhipicephalus sanguineus were analysed; only one male was infected by B. canis vogeli. This is the first report on the detection of DNA belonging to B. canis vogeli in domestic dogs and in R. sanguineus in Tunisia.     

The brown dog tick Rhipicephalus sanguineus and the dog as experimental hosts of Ehrlichia canis

Adult Rhipicephalus sanguineus ticks efficiently transmitted Ehrlichia canis to susceptible dogs for 155 days after detachment as engorged nymphs from a dog in the acute phase of ehrlichiosis. Adult ticks that had similarly engorged on a dog in the chronic phase of ehrlichiosis failed to transmit E canis to susceptible dogs. Infected but unfed adult ticks may be of greater importance than the chronically infected carrier dog as a natural reservoir of E canis.     

Epidemiological survey of Anaplasma platys and Ehrlichia canis using ticks collected from dogs in Japan

Detection of Anaplasma platys and Ehrlichia canis in ticks recovered from dogs in Japan was attempted using a species-specific nested PCR based on the 16S rRNA gene. A total of 1211 ticks recovered from 1211 dogs from all over Japan were examined for A. platys and E. canis. Four tick samples from Fukushima, Miyazaki and Kagoshima Prefectures recovered from four different dogs showed a positive reaction for A. platys. Although the four dogs did not show any clinical signs and no blood examination data were available, it is possible that A. platys has already been spread widely in Japan. No positive reactions were observed in any ticks examined for E. canis.     

Molecular detection of tick-borne haemopathogens in shelter dogs and Rhipicephalus sanguineus (sensu lato) ticks from Peninsular Malaysia

Ticks are important vectors in transmitting various pathogens and they could jeopardize the health and welfare of humans and animals worldwide. The present study aimed to investigate the presence of important tick-borne haemopathogens (TBH) in dogs and ticks via polymerase chain reaction (PCR) assays. A total of 220 blood samples and 140 ticks were collected from 10 animal shelters in Peninsular Malaysia. Of 220 blood samples, 77 (35 %) were positive to TBH, of which 20 % were E. canis, 12 % were A. platys, 7 % were B. gibsoni and 7 % were B. vogeli. All ticks were identified as Rhipicephalus sanguineus with five samples (3.57 %) positive with TBH. Co-infections of TBH (0.45–9.55 %) in dogs were also observed in this study.     

Apparent tick paralysis by Rhipicephalus sanguineus (Acari: Ixodidae) in dogs

Certain tick species including Ixodes holocyclus can inoculate neurotoxins that induce a rapid, ascending flaccid paralysis in animals. Rhipicephalus sanguineus, the most widespread tick of dogs, is recognized as a vector of several pathogens causing diseases in dogs and humans. A single report suggests its role as cause of paralysis in dogs. This study presents the clinical history of 14 young dogs heavily infested by R. sanguineus (intensity of infestation, 63-328) in an endemic area of southern Italy. During May to June of 2011, dogs were presented at the clinical examination with neurological signs of different degrees (e.g., hind limb ataxia, generalized lethargy, and difficulty in movements). All animals were treated with acaricides and by manual tick removal but ten of them died within a day, displaying neurological signs. The other 4 dogs recovered within 3 days with acaricidal and supportive treatment. Twelve dogs were positive by blood smear examination for Hepatozoon canis with a high parasitemia, two also for Babesia vogeli and two were negative for hemoparasites. Low-grade thrombocytopenia, hypoalbuminemia, and pancytopenia were the haematological alterations most frequently recorded. Other causes of neurological disease in dogs were excluded and the diagnosis of tick paralysis by R. sanguineus was confirmed (ex juvantibus) by early and complete recovery of 4 dogs following acaricidal treatment and tick removal.     

Evaluation of Rhipicephalus sanguineus as a potential biologic vector of Ehrlichia platys

The brown dog tick, Rhipicephalus sanguineus (Acari: Ixodidae), transmits several diseases among dogs including Ehrlichia canis infection. The role of Rhipicephalus sanguineus as a biologic vector for E platys, the rickettsial agent of infectious canine cyclic thrombocytopenia, was studied in dogs. Laboratory-cultured, pathogen-free nymph ticks were fed to repletion on dogs acutely infected with E platys. Tick engorgement coincided with the development of initial parasitemia and thrombocytopenia in the infected dogs. Following repletion, nymph ticks were allowed to molt under controlled conditions. One-month-old E platys-exposed adult ticks failed to infect naive dogs in animal transmission studies. The presence of E platys was not detected in midguts or salivary glands of similarly exposed adult ticks by use of light and transmission electron microscopy. These studies indicate that R sanguineus may not transmit E platys infection.     

Duration of antibodies against 24 kd protein of Rhipicephalus sanguineus extract in dogs infested with the adult ticks

A 24 kd protein from Rhipicephalus sanguineus (Rs24p) which was common to larvae, nymphs, male and female whole body and salivary gland extract of males and female was detected specifically in the serum from dogs after repeated infestation with adult R. sanguineus. The duration of antibodies against Rs24p in dogs infested with adults was examined by Western blotting analysis. Anti-Rs24p antibody was detected in two of 4 dogs during the period of 40 days in the first infestation. In the second infestation, all dogs showed positive reaction against Rs24p, but the duration of the antibodies varied greatly among the animals.     

First evidence and molecular characterization of Babesia vogeli in naturally infected dogs and Rhipicephalus sanguineus ticks in southern France

Babesiosis is an emerging tick-borne disease of animals and humans caused by intraerythrocytic protozoa of the genera Babesia and Theileria. In France canine babesiosis has a high prevalence with Babesia canis thought to be the main aetiological agent of the disease. Babesia vogeli has already been reported to occur in Europe and in other countries around the Mediterranean Sea. The tick Rhipicephalus sanguineus, the main known vector of B. vogeli, occurs in southern France. However, only one case of a B. vogeli infected dog has been reported to date in France. To gain further insight into the prevalence of Babesia and Theileria infections in dogs and ticks of the R. sanguineus complex, a study was conducted in a veterinary practice in the south of France from January to September 2010. Twelve bloods from dogs and 36 R. sanguineus ticks were analyzed using PCR and sequencing. For the analysis of ticks, a new primer was designed to specifically amplify the B. vogeli 18S rRNA gene. Four dogs (33.3%) and 8 ticks (22.2%) were found to be infected with B. vogeli. This approach has thus revealed for the first time a cluster of cases of canine babesiosis caused by B. vogeli in France and highlights the need to systematically screen for pathogens potentially responsible for canine babesiosis at the species level using suitable molecular tools.     

Polymyositis associated with Ehrlichia canis infection in two dogs

Clinical, haematological, biochemical, electrophysiological and pathological features of two dogs infected with Ehrlichia canis and with concurrent signs of polymyositis are presented. Both dogs had a history of relatively acute onset, progressive tetraparesis, hyporeflexia and generalised muscle wasting. Skeletal muscles were atrophic and characterised histologically by plasmocytic, lymphocytic and immature lymph-oreticular cellular infiltrates with accompanying areas of necrosis. Histopathological similarities between ehrlichiosis and polymyositis are noted and a probable aetiological relationship is inferred.     

Detection of Ehrlichia canis by polymerase chain reaction in dogs from Portugal

Antibodies against Ehrlichia canis, the cause of canine monocytic ehrlichiosis, have been reported previously in clinically ill and stray dogs from Portugal. In this study, the 16S rRNA gene of E. canis was detected by the polymerase chain reaction (PCR) in 12/55 (22%) of dogs with suspected tick-borne disease in the Algarve region in Portugal.     

Movement of Rhipicephalus sanguineus adults between co-housed dogs during active feeding

Adult male ticks have been shown capable of experimental acquisition and transmission of tick-borne pathogens without requiring a molt. To determine the ecological relevance of this intrastadial transmission route, we evaluated the extent to which actively feeding male Rhipicephalus sanguineus move naturally between co-housed infested dogs. Dogs (n=4) were infested with single color-coded ticks, individually housed in tick-confinement cages for 48 h while infestations established, and then each dog examined and the ticks present counted. Dogs were then co-housed in a large, group pen for an additional 7 (study 1) or 5 (study 2) days. In the first study, sex ratios were adjusted to encourage migration, with two dogs receiving predominantly female ticks and two dogs receiving all male ticks. In the second study, each dog received a ratio of ticks that parallels that found in natural infestations (4:1 male to female). Results showed that ticks readily migrated between infested, co-housed dogs. Rates of immigration, defined as the percentage of ticks previously attached to one dog that moved onto another dog, ranged from 0 to 46% (mean=31.1% study 1; 9.4% study 2). Emigration rates, defined as the number of ticks initially infesting one dog that moved to another dog, averaged 35.2% in study 1 and 10.8% in study 2 (3.6-67.6%). Movement of adult ticks between dogs represents a naturally occurring form of interrupted feeding, a strategy which has been shown to shorten the feeding time necessary to allow transmission of pathogens. In ticks that readily detach from one host and reattach to a second host to resume feeding, replication of any pathogens present has already been initiated and therefore the same delay in transmission seen in ticks attached to a host for the first time may not occur.     

Experimental transmission of field Anaplasma marginale and the A. centrale vaccine strain by Hyalomma excavatum, Rhipicephalus sanguineus and Rhipicephalus (Boophilus) annulatus ticks

The cattle rickettsia Anaplasma marginale is distributed worldwide and is transmitted by about 20 tick species, but only Rhipicephalus simus, a strictly African tick species, has been shown to transmit the vaccine strain of A. centrale. The aim of the present study was to examine transmission of field strains of A. marginale and of the vaccine strain of A. centrale by three tick species -Hyalomma excavatum, Rhipicephalus sanguineus and Rhipicephalus (Boophilus) annulatus - to susceptible calves. Two genetically distinct Israeli field strains of A. marginale, tailed and non-tailed (AmIsT and AmIsNT, respectively), were efficiently transmitted by R. sanguineus, whereas H. excavatum transmitted only the tailed isolate, and R. (Boophilus) annulatus did not transmit A. marginale. None of the three tick species transmitted A. centrale. By means of msp1a primers in PCR assays, amplicons of similar sizes were obtained from either A. marginale-infected calves that were used for acquisition feeding, from R. sanguineus fed on the infected calves, or from calves to which anaplasmosis had been successfully transmitted by these ticks. Although an A. centrale-specific fragment was amplified from salivary glands of R. sanguineus, no transmission to susceptible cattle occurred during 3 months of observation, and anaplasmosis was not induced in splenectomized calves that were subinoculated with blood from calves on which R. sanguineus had fed.     

Failure of imidocarb dipropionate to clear experimentally induced Ehrlichia canis infection in dogs

The recommended treatment for canine ehrlichiosis is tetracycline or its analog doxycycline, although recent reports have documented ineffective clearing of Erchlichia canis after doxycycline administration. Imidocarb dipropionate is used as an alternative treatment to tetracycline or is used in conjunction with doxycycline. The effectiveness of imidocarb dipropionate in clearing Ehrlichia species from the blood and tissues of dogs with E. canis infection has not been thoroughly evaluated. Fifteen dogs were experimentally infected with E. canis. Ten dogs were treated with imidocarb dipropionate (6.6 mg/kg, IM, 2 injections given 2 weeks apart). Five infected control dogs were not treated. Blood samples from all 15 dogs were E. canis DNA positive by PCR assay by 3 weeks after inoculation (PI), and E. canis antibodies were detected by IFA assay by 1 week PI. Blood platelet counts in all dogs were below the reference interval by 4 weeks PI. E. canis DNA was detected in bone marrow and splenic aspirates by PCR assay 4 weeks PI but not before infection. Bone marrow aspirates were E. canis DNA positive by PCR assay in 14/15 dogs, and splenic aspirates were E. canis DNA positive by PCR assay in 13/15 dogs. Blood samples from all treated and control dogs remained positive for E. canis DNA by PCR assay, and platelet counts remained below preinoculation values 13 weeks PI (6 weeks after 2nd treatment). As administered in this study, imidocarb dipropionate did not clear experimental E. canis infection in dogs.     

Renal amyloidosis caused by Ehrlichia canis

An 8 year old, intact female, Old English Sheepdog was presented with lethargy and chronic weight loss of one year duration. The dog suffered from recurrent fever and a mild peripheral lymphadenopathy. Mild thrombocytopenia, monoclonal hyperglobulinemia and positive Ehrlichia canis antibody titer were indicative of Ehrlichiosis. The patient developed proteinuria and renal failure, which suggested the presence of glomerulopathy. Immunohistochemical staining of renal biopsy specimen demonstrated AA amyloidosis. Chronic ehrlichiosis was suspected to be the cause of the dog's renal amyloidosis.