Ehrlichia canis infection in the cerebrospinal fluid of a dog characterized by morulae within monocytes and neutrophils

An 8-year-old neutered female English Pointer was referred to a veterinary referral center (southwest of England) with a 4-5-month history of fecal incontinence and no evidence of urinary incontinence. Blood and free-catch urine samples were collected and sent to an off-site laboratory. Further investigations were postponed until laboratory results were available. Blood results showed a mild leukopenia, mild nonregenerative anemia, moderate to marked thrombocytopenia, and a mild increase in ALT and ALP activities. The primary veterinarian and client did not proceed with any further investigations for thrombocytopenia. Three weeks after the initial presentation, there was considerable clinical deterioration and progression of neurologic signs. Thoracic radiographs and an abdominal ultrasonographic examination were unremarkable. Magnetic resonance imaging (MRI) of the brain and spinal cord revealed an intramedullary lesion at the level of the C7 vertebra, a cystic lesion in the forebrain, and a bilateral lesion in the thalamus. A lumbar cerebrospinal fluid (CSF) was collected. CSF analysis showed a robustly increased protein concentration and marked pleocytosis. The cytologic evaluation revealed a mixed cellular population. Occasional neutrophils and monocytoid cells showed purple spherical intracellular inclusions, resembling Ehrlichia morulae. An aliquot of CSF was used off-label with a dot ELISA test, which showed a strong positive result for antibodies against Ehrlichia canis/Ehrlichia ewingii. PCR identified these morulae to be E canis. To best of the authors’ knowledge, this is the first case of ehrlichial infection in canine CSF where Ehrlichia sub-species morulae present within neutrophils were confirmed to be Ehrlichia canis using PCR.     

Suppurative spinal meningomyelitis in a dog with intra-neutrophilic cerebrospinal fluid cells Ehrlichia canis morulae

A 2-year-old castrated male Creole Shepherd mixed dog was presented for non-ambulatory paraparesis of the pelvic limbs. The magnetic resonance imaging and cerebrospinal fluid (CSF) analysis were consistent with meningomyelitis. Positive serology for Ehrlichia canis/Ehrlichia ewingii suggested exposure to a pathogen; qPCR on the serum and the CSF confirmed active infection. Ehrlichial morulae were observed within CSF and peripheral blood polymorphonuclear neutrophils; a species-specific PCR confirmed E. canis infection. This is an interesting report of E. canis infection in a dog with morulae observed in neutrophils both in the peripheral blood and CSF.     

Mixed Ehrlichia canis, Hepatozoon canis, and presumptive Anaplasma phagocytophilum infection in a dog

A 5-month-old, female, mongrel dog was admitted to the Clinic of Companion Animal Medicine, Aristotle University of Thessaloniki, Greece, with depression, anorexia, fever, peripheral lymphadenopathy, splenomegaly, oculonasal discharge, nonregenerative anemia, and mild thrombocytopenia. Cytology of Giemsa-stained buffy coat, bone marrow, and lymph node aspiration smears revealed numerous morulae in mononuclear leukocytes and in neutrophils, and Hepatozoon canis gamonts in neutrophils. The dog was seropositive to Ehrlichia canis (immunofluorescence assay [IFA]) and Hepatozoon canis (ELISA) but not to Anaplasma phagocytophilum (IFA). A nested polymerase chain reaction performed on bone marrow aspirates was positive for E canis. This method was not applied for the detection of A phagocytophilum. Treatment with doxycycline and imidocarb dipropionate resulted in both clinical and parasitologic cure. This is the first reported case of a mixed infection with E canis, H canis, and presumptive A phagocytophilum. The findings emphasize the value of cytology in offering a quick and inexpensive diagnosis in mixed tick-borne infections of dogs.     

Doxycycline clearance of experimentally induced chronic Ehrlichia canis infection in dogs

BACKGROUND: Ineffective clearance of Ehrlichia canis after doxycycline administration has been reported despite the fact that the recommended treatment for canine ehrlichiosis is doxycycline. The effectiveness of doxycycline in clearing E canis infection from the blood and tissues of dogs requires additional evaluation. HYPOTHESIS: Doxycycline (5 mg/kg PO q12h), administered for 4 weeks, will eliminate E canis infection from the blood and tissues of experimentally infected dogs. ANIMALS: Fifteen Walker hound-mixed breed dogs were inoculated subcutaneously with E canis-infected canine histiocytic cells 4 months before doxycycline treatment. METHODS: Four dogs were treated with doxycycline (5 mg/kg PO q12h for 3 weeks), 5 dogs were treated with doxycycline at the same dosage for 4 weeks, and 5 control dogs were not treated. Dexamethasone (0.4 mg/kg i.v.) was given after treatment to precipitate recrudescence of any remaining E canis organisms. Platelet counts, anti-E canis immunofluorescent antibodies, and polymerase chain reaction (PCR) detection of E canis deoxyribonucleic acid (DNA) in blood and tissues were evaluated. RESULTS: E canis DNA was not detected in the blood and tissues of doxycycline-treated dogs after treatment. Platelet counts were within reference intervals, and E canis antibodies decreased. Spontaneous clearance of E canis infection occurred in 2 of 5 control dogs. Three control dogs had E canis DNA detected in blood and tissues, platelet counts remained low or within the reference interval, and E canis antibodies remained high. CONCLUSIONS AND CLINICAL IMPORTANCE: As administered in this study, doxycycline cleared E canis from the blood and tissues of experimentally infected dogs.     

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.     

Prevalence of Ehrlichia canis infection in thrombocytopenic dogs from Rio de Janeiro, Brazil

BACKGROUND: Infection with Ehrlichia canis causes a highly variable, multisystemic disease in dogs. Nevertheless, many clinicians in Rio de Janeiro, Brazil, use the presence of only thrombocytopenia to make a presumptive diagnosis of E canis infection. OBJECTIVE: The objective of this study was to determine the prevalence of E canis in thrombocytopenic dogs from Rio de Janeiro, Brazil, using polymerase chain reaction (PCR). METHODS: Following DNA extraction of whole blood samples from 226 dogs, PCR assays were done using primers for rickettsial DNA (including Ehrlichia spp, Anaplasma platys and A phagocytophilum) and using E canis-specific primers (16S rRNA gene). Dogs were grouped as thrombocytopenic and nonthrombocytopenic based on platelet counts. The null hypothesis that there was no difference in the prevalence of E canis in these groups was rejected at P<.05. RESULTS: Thirty-six (32.1%) of the thrombocytopenic dogs and 4 (3.5%) of the nonthrombocytopenic dogs were positive for rickettsial gene sequences (P<.0001). Further, 30 (26.8%) of thrombocytopenic dogs and 4 (3.5%) nonthrombocytopenic dogs were positive for E canis-specific gene sequences (P<.0001). CONCLUSIONS: Although the prevalence of E canis infection was higher in thrombocytopenic dogs, less than one third of these dogs had demonstrable E canis infection. Thus, thrombocytopenia is not specific for the detection of E canis infection and should not be used solely to establish a diagnosis of canine ehrlichiosis, even in a geographic area with relatively high disease prevalence.     

Cultivation and molecular identification of Ehrlichia canis and Ehrlichia chaffeensis from a naturally co-infected dog in Venezuela

Background: Diagnosis of canine ehrlichiosis in Venezuela is normally performed by examination of buffy coat smears (BCS). Characteristic inclusion bodies are frequently observed in leukocytes and platelets from dogs with clinical signs of the disease. Objective: The purpose of this study was to investigate the co-infection of a dog with Ehrlichia canis and E hrlichia chaffeensis using microbiological and molecular techniques. Methods: Primary cultures of monocytes from a dog showing signs of ehrlichiosis were performed. Ehrlichial inclusions in blood cells were demonstrated by BCS and in cultured cell smears with direct immunofluorescence and Dip Quick staining. Nested PCR analysis was performed with DNA from blood samples and cultures, using primers specific for E. canis and E. chaffeensis. The amplified DNA fragments were sequenced to confirm the specificity of the amplifications. Results: The BCS of the naturally infected dog contained intracellular morulae. Ehrlichial inclusions were observed 9 days after inoculation of the primary cultures. After 3 passages with monocytes from a healthy dog, 65% of infected cells, and cells with >60 morulae were observed. A healthy female German Shepherd dog, seronegative for E. canis and E. chaffeensis antigens and without contact to ticks, was inoculated with an infected culture. The animal developed signs of canine monocytic ehrlichiosis and became seropositive. Nested PCR results and sequencing of amplified DNA fragments demonstrated the simultaneous presence of E. canis and E. chaffeensis in both dogs. Conclusions: This is the first report of E. chaffeensis in dogs in South America. This organism was previously identified in dogs by PCR only in the United States.     

Disseminated infection with Neospora caninum in a ten-year-old dog

Naturally-occurring neosporosis with multiple organ involvement was identified in a 10-year-old neutered male Basset hound dog. Clinical signs were first noticed 3 weeks prior to referral and consisted of crouched stance and mild pelvic limb ataxia. Dexamethasone administration had provided transient improvement. On presentation to the teaching hospital, clinical signs included depression, pelvic limb ataxia, inability to stand without assistance, and pain on deep palpation of the cervical and lumbar vertebral column. Lesions were found in the myocardium, liver, spleen, adrenal glands, brain, and spinal cord. Tachyzoites of Neospora caninum were found in the myocardium and adrenal glands. Organisms stained with anti-Neospora caninum, but not to anti-Toxoplasma gondii serum in an immunohistochemical test.     

Ocular manifestations of natural canine monocytic ehrlichiosis (Ehrlichia canis): a retrospective study of 90 cases

OBJECTIVE: To investigate the spectrum, prevalence and treatment response rate of ocular manifestations associated with natural canine monocytic ehrlichiosis (CME) due to Ehrlichia canis. METHODS: The medical records of 90 dogs that were admitted for primary ocular problems and reacted positive for E. canis serum antibodies were reviewed. All the dogs were treated with oral doxycycline. In 79 dogs, a combination of systemic corticosteroids and/or topical anti-inflammatory, mydriatics/cycloplegics and antibacterial agents were applied. RESULTS: The dogs ranged from 0.5 to 15 years in age; 55 were male and 35 female. A total of 19 breeds were represented. In the majority of dogs, ocular abnormalities were noticed in conjunction with other CME-induced clinical signs. In 30/90 dogs, ophthalmic signs were the sole presenting complaint. Unilateral (22/90, 24.5%) and bilateral (68/90, 75.5%) uveitis was the most common ophthalmic diagnosis and was classified as anterior, posterior or panuveitis in 58 (64.5%), 8 (8.9%) and 24 (26.6%) of the dogs, respectively. Bilateral uveitis was significantly more common than unilateral uveitis (P < 0.0001), and anterior significantly more common than posterior uveitis (P < 0.0001) or panuveitis (P < 0.0001). In addition, corneal ulceration (12/90, 13.3%), necrotic scleritis (10/90, 11.1%), low tear production (8/90, 8.9%) and orbital cellulitis (3/90, 3.3%) were seen. Of the 45 (50%) dogs with a satisfactory follow-up, 25 (55.5%), 11 (24.5%), and 9 (20%) showed complete, partial and poor response to treatment, respectively. The number of dogs exhibiting complete resolution of the ocular manifestations was significantly higher than those with partial (P < 0.0001) or poor (P < 0.0001) response. CONCLUSIONS: CME should be considered a major differential for a wide range of ocular manifestations exhibited by dogs residing in the endemic areas of the disease. Anterior bilateral uveitis appears to be the most prevalent ocular lesion and a favorable outcome to systemic and topical treatment may be expected in the majority of the affected dogs.     

Host surveys, ixodid tick biology and transmission scenarios as related to the tick-borne pathogen, Ehrlichia canis

The ehrlichioses have been subject to increasing interest from veterinary and public health perspectives, but experimental studies of these diseases and their etiologic agents can be challenging. Ehrlichia canis, the primary etiologic agent of canine monocytic ehrlichiosis, is relatively well characterized and offers unique advantages and opportunities to study interactions between a monocytotropic pathogen and both its vertebrate and invertebrate hosts. Historically, advances in tick-borne disease control strategies have typically followed explication of tick-pathogen-vertebrate interactions, thus it is reasonable to expect novel, more sustainable approaches to control of these diseases as the transmission of their associated infections are investigated at the molecular through ecological levels. Better understanding of the interactions between E. canis and its canine and tick hosts would also elucidate similar interactions for other Ehrlichia species as well as the potential roles of canine sentinels, reservoirs and models of tick-borne zoonoses. This article summarizes natural exposure studies and experimental investigations of E. canis in the context of what is understood about biological vectors of tick-borne Anaplasmataceae.     

Serum hyperviscosity syndrome associated with Ehrlichia canis infection in a dog

A 10-year-old male Lhasa Apso was examined because of pyrexia, vomiting, weight loss, lethargy, and disorientation. The clinical and laboratory findings were supportive of a diagnosis of serum hyperviscosity syndrome associated with an Ehrlichia canis infection. After tetracycline therapy the serum hyperviscosity syndrome and E. canis infection were successfully resolved.     

Diagnosis and treatment of Neospora caninum infection in a dog

Neospora caninum, a protozoan organism, caused extensor rigidity of the pelvic limbs in a 12-week-old dog. Diagnosis was based on results of muscle biopsy, neuroelectrodiagnostics, serotesting, and cell culture. Indirect fluorescent antibody (IFA) titer to N caninum was 1:800 at time of admission and 1:3,200 after 4 and 6 weeks. A reciprocal IFA titer of 50 to N caninum was also found in the CSF. Serotesting for T gondii was negative. Treatment with clindamycin followed by sulfadiazine and trimethoprim did not change the pelvic limb extensor rigidity, but other signs of minor neurologic dysfunction improved.     

Babesia canis vogeli,Ehrlichia canis,and Anaplasma platys infection in a dog

A 12‐month‐old male neutered mixed breed dog was presented with a history of diarrhea, lethargy, emaciation, polydypsia, and sniffling. Physical examination findings included pale mucous membranes, increased heart and respiratory rates, and normal rectal temperature (38°C). Hematologic abnormalities included anemia and thrombocytopenia. Biochemical abnormalities included hypoalbuminemia, hyperbilirubinemia, and elevated ALP and ALT activities. A SNAP 4Dx test result was positive for Ehrlichia canis. Babesia canis vogeli organisms were found in the peripheral blood films, while morulae of E canis were not seen. Real‐time polymerase chain reaction testing confirmed the presence of both B c vogeli and E canis organisms, and also was positive for Anaplasma platys infection. The dog recovered following treatment with doxycycline and imidocarb dipropionate, with normal hematology and biochemical profiles.     

青海省格尔木市乳牛新孢子虫病的血清学调查

用重组蛋白GST-NcSAG1t作为ELISA诊断抗原,对格尔木市乳牛犬新孢子虫病进行了调查.结果,从 35份乳牛血清样品中检出犬新孢子虫抗体阳性血清4份,阳性率为11.42%.检测结果说明,青海省格尔木市饲养的乳牛牛群中存在犬新孢子虫感染.     

Molecular assessment of the transplacental transmission of Toxoplasma gondii,Neospora caninum,Brucella canis and Ehrlichia canis in dogs

Given the fact that numerous microbial species can be detected in pregnant female dogs, the objective of this study was to assess the transplacental transmission of Brucella canis, Ehrlichia canis, Neospora caninum and Toxoplasma gondii in stillborn puppies. This study involved 41 stillborn puppies, 78.6% of which were positive for T. gondii, 52.4% for N. caninum and 59.5% for B. canis. E. canis was not detected in any of the analyzed puppies. Pregnancy is an important physiological condition for the transmission of infectious agents to puppies and transplacental transmission may be epidemiologically relevant in the spread of these opportunistic agents.     

Molecular identification of Ehrlichia ewingii in a polyarthritic Texas dog

An 8‐year‐old, neutered male, Golden Retriever presented for bilateral carpal joint effusion. A complete blood count revealed mild leukopenia and marked thrombocytopenia. Samples were sent to the Texas A&M Veterinary Medical Diagnostic Laboratory for blood smear review and serologic testing for tick‐borne diseases. Numerous morulae were observed within neutrophils, and antibodies against Ehrlichia canis were detected at a 1:512 dilution via the indirect fluorescent antibody (IFA) test. As neutrophilic morulae are morphologically indistinguishable between Ehrlichia ewingii and Anaplasma phagocytophilum, and genus‐wide cross‐reactivity is possible with serologic testing, additional molecular testing was performed. Quantitative real‐time polymerase chain reaction (qPCR) followed by conventional PCR and Sanger sequencing were performed on serum identified with E ewingii as the sole disease‐causing agent. Three months after diagnosis and treatment, no morulae were found, molecular testing for E ewingii detected no DNA, and convalescent IFA testing demonstrated a continued detection of antibodies for E canis at a 1:512 dilution. To the authors’ knowledge, this is the first reported case of E ewingii confirmed with molecular diagnostics in a Texas dog. The zoonotic transmission potential of E ewingii should be noted as Texas supports competent tick vectors, and dogs represent effective sentinels for human ehrlichiosis. This report also highlights the utility of molecular diagnostics when serologic and microscopic evaluations are not sufficient in providing the species‐level identity of a causative agent.     

Outbreak of canine monocytic ehrlichiosis in Saudi Arabia

BACKGROUND: Canine monocytic ehrlichiosis (CME) is a widespread tickborne infection of canids caused by Ehrlichia canis, a gram-negative obligatory intracellular bacteria belonging to the family Anaplasmataceae. CME is reported to have worldwide distribution, but its presence in a region requires the presence of the vector, the brown dog tick Rhipicephalus sanguineus. OBJECTIVE: This purpose of this report was to describe an outbreak of CME in a colony of dogs resident in the eastern region of Saudi Arabia. METHODS: History, presenting clinical signs, and the results of a CBC, biochemical panel, and serology (using a commercial test for E canis antibodies) were evaluated in 9 male Labrador Retrievers between October and December 2006. RESULTS: The majority of dogs presented with severe lethargy (7/9) and acute anorexia (5/9), and had fever (7/9) and generalized lymphadenopathy (7/9). The most common laboratory abnormalities were anemia (8/9), leukopenia (7/9), and hypoalbuminemia (6/9). Thrombocytopenia was found in only 2 dogs, 1 of which had increased bleeding tendency. Ehrlichia morulae were identified in blood films from 4/9 dogs and serologic test results were positive in 7/9 dogs. Confirmation of Ehrlichia sp infection was obtained in 1 dog by using a genus-specific real-time PCR assay. Four dogs had tick infestation; the ticks on 1 dog were identified as R sanguineus. All of the dogs had a rapid clinical response to doxycycline hyclate. CONCLUSIONS: This report, to our knowledge, is the first to describe the presence of a pathogenic Ehrlichia organism affecting dogs in Saudi Arabia. Additional molecular studies are necessary to confirm E canis infection, and to identify the strain of the organism.