Canine Ehrlichiosis in Six Dogs with Persistently Increased Antibody Titers

Chronic ehrlichiosis was diagnosed in six dogs on the basis of increased immunofluorescent antibody titers to Ehrlichia canis. Although clinical signs varied, all six dogs were anemic, hyperglobulinemic, and an IgG monoclonal gammopathy was documented in five dogs in which serum protein electrophoreses were performed. All dogs were treated with tetracycline for at least 14 days; four dogs also received immunosuppressive drugs. Clinical signs resolved within 1 week, hematologic abnormalities resolved in 1 to 5 months, and increased globulin concentrations normalized in 1 to 15 months; however, E. canis antibody titers remained increased for 15 to 31 months after initiation of treatment. Results of this study show that increased E. canis titers can persist in dogs with ehrlichiosis for many months after clinical recovery.     

Molecular identification of Ehrlichia ewingii infection in dogs: 15 cases (1997-2001)

OBJECTIVE: To determine historical, physical examination, hematologic, and serologic findings in dogs with Ehrlichia ewingii infection. DESIGN: Retrospective study. ANIMALS: 15 dogs. PROCEDURE: In all dogs, infection with E ewingii was confirmed with a polymerase chain reaction (PCR) assay. Follow-up information and clarification of information recorded in the medical records was obtained by telephone interviews and facsimile correspondence with referring veterinarians and owners. RESULTS: Fever and lameness were the most common findings with each occurring in 8 dogs. Five dogs had neurologic abnormalities including ataxia, paresis, proprioceptive deficits, anisocoria, intention tremor, and head tilt. Neutrophilic polyarthritis was identified in 4 dogs. No clinical signs were reported in 3 dogs. The predominant hematologic abnormality was thrombocytopenia, which was identified in all 12 dogs for which a platelet count was available. Reactive lymphocytes were seen in 5 of 13 dogs. Concurrent infection with another rickettsial organism was identified in 4 dogs. Of the 13 dogs tested, 7 were seroreactive to E canis antigens. Morulae consistent with E ewingii infection were identified in neutrophils in 8 dogs. Treatment with doxycycline, with or without prednisone, resulted in a rapid, favorable clinical response in the 9 dogs for which follow-up information was available. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that PCR testing for E ewingii infection should be considered in dogs with fever, neutrophilic polyarthritis, unexplained ataxia or paresis, thrombocytopenia, or unexplained reactive lymphocytes, and in dogs with clinical signs suggestive of ehrlichiosis that are seronegative for E canis. Following treatment with doxycycline, the prognosis for recovery is good.     

Use of plasmapheresis and chemotherapy for treatment of monoclonal gammopathy associated with Ehrlichia canis infection in a dog

Monoclonal gammopathy associated with Ehrlichia canis infection was diagnosed in a German Shepherd Dog. The dog was treated by use of chemotherapy and tetracycline and by plasmapheresis. The dog tolerated plasmapheresis and long-term drug therapy well, and clinical signs resolved over a 90-day period. The monoclonal gammopathy resolved after treatment, but specific antibody to E canis indicated suppression followed by a rebound to the initial high titer.     

Evaluation of neutrophil oxidative metabolism in canine monocytic ehrlichiosis

BACKGROUND: Canine monocytic ehrlichiosis (CME) is a tick-borne disease caused by Ehrlichia canis, a rickettsia that infects the monocytes of dogs. This infection can result in a chronic and life-threatening disease. Thrombocytopenia, mild anemia, and leukopenia are the most common hematologic findings in CME. OBJECTIVE: To investigate the role of peripheral blood neutrophils in CME, an evaluation was conducted of their functional state during the acute phase of the disease in dogs experimentally infected by E canis. METHODS: Seven dogs were inoculated with E canis, and 3 remained as uninfected controls. All dogs had physical exams and hematologic tests (CBC and nitroblue tetrazolium [NBT] reduction) during a 6-week period. RESULTS: There was no difference (P > .05) in spontaneous NBT reduction results between the 2 groups of dogs throughout the 6-week period of observation. Nevertheless, when stimulated, the neutrophils showed higher activity in the infected group (P = .01) on weeks 4 and 5 after infection. CONCLUSION: Infection by E canis has no influence on neutrophil oxidative metabolism even though during the remission period of the acute phase of the disease, the neutrophils seem to be more reactive under stimulation.     

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.     

Canine monocytic ehrlichiosis presenting as acute blindness 36 months after importation into the UK

A four-year-old Labrador retriever developed sudden-onset blindness, associated with bilateral uveitis, intraocular haemorrhage and retinal detachment. It had been imported into the UK from Sardinia 36 months before presentation. Haematological abnormalities included non-regenerative anaemia, thrombocytopenia and neutropenia. Serum and urine protein electrophoresis demonstrated a monoclonal gammopathy. An immunofluorescent antibody test for Ehrlichia canis was positive, with a titre of 1:320, confirming a diagnosis of chronic monocytic ehrlichiosis. This case highlights how the prolonged subclinical phase of monocytic ehrlichiosis could enable infected dogs to enter the UK without signs of disease. Chronic monocytic ehrlichiosis should be considered in dogs which have been imported from E canis-endemic countries and present with bleeding disorders and gammopathy, even if signs develop many years after importation.     

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.     

Evaluation of granulocytic ehrlichiosis in dogs of Missouri, including serologic status to Ehrlichia canis, Ehrlichia equi and Borrelia burgdorferi.

Canine granulocytic ehrlichiosis was diagnosed in 37 dogs by finding ehrlichial morulae in 0.1 to 26.2% of their blood neutrophils and eosinophils. All 37 dogs had clinical signs of arthritis or muscular stiffness. Titer to Ehrlichia canis was determined in sera from 31 of the 37 dogs; 25 dogs had titer ranging from 1:20 to 1:5,120. In the other 6 dogs, titer to E canis was less than 1:10. The most common hematologic abnormality in these dogs, other than rickettsiemia, was thrombocytopenia. Granulocytes infected with ehrlichial organisms were not found in another 10 dogs that had clinical signs of arthritis or muscular stiffness. Of these 10 dogs, 3 had titer to E canis ranging from 1:40 to 1:320. Titer in the other 7 dogs was less than 1:10. Ehrlichial morulae were not found in the granulocytes of 18 healthy dogs. Of these 18 dogs, 9 had titer to E canis ranging from 1:20 to 1:5,120. Titer in the other 9 dogs was less than 1:10 Titer to Borrelia burgdorferi was determined in dogs with granulocytic ehrlichiosis, arthritic dogs without detected rickettsiemia, and in healthy dogs. Low titer determined by 2 laboratories was considered to be nonspecific reaction in all 3 groups of dogs and, thus, did not indicate that the arthritic disorders were attributable to canine borreliosis.     

Multiple myeloma in a dog with multiple concurrent infectious diseases and persistent polyclonal gammopathy

A 12‐year‐old, spayed female, mixed‐breed dog was presented for acute hematuria, stranguria, polyuria, and polydipsia, as well as lameness for 8 days. Previous medical history included treatment for infection with Ehrlichia canis, Anaplasma phagocytophilum, Leishmania infantum, and Dirofilaria immitis 6.5 years prior to presentation. Besides persistently increased antibody titers to E canis and A phagocytophilum, polyclonal gammopathy with a monoclonal spike and moderate hypercalcemia were observed. There was marked hematuria, and Staphylococcus aureus was cultured from urine. Two weeks after successful treatment of the urinary tract infection, radiographs showed an extensive destructive monostotic lesion of the right humerus. Cytologic examination of fine‐needle aspirates of this lesion revealed a neoplastic round cell population suggestive of multiple myeloma. The dog was treated with melphalan and prednisolone for suspected multiple myeloma and doxycycline for suspected ehrlichiosis and anaplasmosis. Treatments lead to resolution of the clinical signs, hypercalcemia, and monoclonal gammopathy, and there was radiographic improvement of bone lesions; polyclonal gammopathy persisted. About one year after presentation the dog was still in clinical remission. This is a rare report of a dog with suspected multiple myeloma and a history of multiple chronic infectious diseases, suggesting that chronic infection and uncontrolled long‐term stimulation of the immune system could contribute to the pathogenesis of multiple myeloma.     

Molecular detection and characterization of Ehrlichia canis from dogs in Turkey

Seroprevalence of Ehrlichia canis antibodies among dogs in Turkey were previously reported, however, the ehrlichial organism has never been characterized in this region. The current study examined dogs from Ankara with febrile illness for E. canis infection with E. canis-specific PCR. Three of the 12 blood specimens from dogs showing clinical signs compatible with canine ehrlichiosis were found to be positive by PCR using E. canis-specific primers. E. canis detected in one of the blood specimens was designated as Kutahya strain. The representative E. canis strain was characterized by 16S rRNA gene sequencing and Western blot analysis of the plasma sample from the dog infected with E. canis. The 16S rRNA sequence (1,388 bp) of the E. canis Kutahya was identical to that of Ehrlichia ovina from a sheep in Turkey and Venezuelan Dog Ehrlichia (VDE) and was closely related (99.9%) to that of type strain of E. canis, Oklahoma. The plasma of the dog infected with E. canis Kutahya was analyzed by Western blotting using the purified E. canis Oklahoma strain as antigen. The reactive antibody profiles of the dog infected with E. canis Kutahya was found to be similar to those of dogs infected with E. canis Oklahoma and VDE, suggesting the antigenic similarities among these strains. The findings in this study would help for a better understanding of epidemiology of canine ehrlichiosis. This is the first report of molecular detection and characterization of an ehrlichial agent in Turkey.     

Ehrlichia canis infection in two dogs that emigrated from endemic areas

Two dogs, emigrated from Zambia and China to Japan, were diagnosed with Ehrlichia canis infection. Both cases had thrombocytopenia, non-regenerative anemia, and hypergloblinemia with polyclonal gammopathy. Case 1 had ataxia of the hind limbs. Severe meningitis was revealed by magnetic resonance imaging examination. Intracytoplasmic inclusions were observed in mononuclear cells of cerebrospinal fluid. Case 2 had a history of bilateral epistaxis, and severe pancytopenia was noticed in complete blood count. Diagnosis was finally achieved by nested polymerase chain reaction and sequence analysis. Thus, even in non-endemic areas, E. canis infection should be included in the differential diagnosis of clinically ill dogs that emigrated from endemic areas.     

Clinical and hematologic findings in canine ehrlichiosis

The clinical and hematologic findings for 56 dogs with ehrlichiosis were studied retrospectively. All dogs had a high serum antibody titer to Ehrlichia canis. The frequency of bleeding disorders, thrombocytopenia, and pancytopenia in these dogs was lower than previously described for dogs so affected. A bleeding disorder caused by a suspected qualitative platelet defect was found in some dogs that did not have thrombocytopenia. Megakaryocytic hyperplasia and high numbers of plasma cells were observed frequently on marrow aspirate smears. The clinical and laboratory findings of dogs were variable and were considered nonspecific for ehrlichiosis.     

Elevated fructosamine concentrations caused by IgA paraproteinemia in two dogs

An 8-year-old male Austrian Pinscher and a 14-year-old male Golden Retriever were presented for evaluation due to unexplainable high fructosamine values despite euglycemia and epistaxis in combination with polydipsia/polyuria, respectively. Blood analysis revealed severe hyperglobulinemia, hypoalbuminemia and markedly elevated fructosamine concentrations in both dogs. Multiple myeloma with IgA-monoclonal gammopathy was diagnosed by serum and urine electrophoresis including immunodetection with an anti-dog IgA antibody and bone marrow aspirations. Diabetes mellitus was excluded by repeated plasma and urine glucose measurements. Fructosamine values were positively correlated with globulin, but negatively correlated with albumin concentrations. These cases suggest that, as in human patients, monoclonal IgA gammopathy should be considered as a possible differential diagnosis for dogs with high fructosamine concentrations.     

Serosurvey of Ehrlichia canis and Hepatozoon canis infection in dogs in Yamaguchi Prefecture, Japan

Antibodies to Ehrlichia canis and Hepatozoon canis in dogs at the Animal Hospital in Yamaguchi University were surveyed and potential risk factors for both pathogens were evaluated. Among 430 dogs examined, 20 (4.7%) and 18 (4.2%) dogs showed positive findings for E. canis and H. canis, respectively. Neither, sex nor age was associated with the seropositivity of either pathogen, but the positive rate in dogs kept outside was slightly higher than that in dogs kept inside for both pathogens. A higher seropositive reaction to E. canis and H. canis was observed in dogs that lived in certain cities and towns. Beagles, golden retrievers and pointers had higher seropositivity than other breeds in E. canis, whereas shibas, akitas, beagles, pointers and mongrels had higher positive rates than other breeds in H. canis.     

Seroprevalence of Ehrlichia canis, Ehrlichia equi, and Ehrlichia risticii in sick dogs from North Carolina and Virginia

Ehrlichia canis, E. equi, and E. risticii seroprevalence was determined by microimmunofluorescent antibody testing (IFA) in a sequential population of 1,845 sick dogs admitted during a 1-year period to the North Carolina State University Veterinary Teaching Hospital. A seroreactor was defined by a reciprocal IFA titer of > or =80 to E. canis, E. equi, or E. risticii antigens. Of the 48 IFA seroreactors, 44 dogs were seroreactive to E. canis, 21 to E. equi, and 0 to E. risticii. Seventeen dogs reacted to both E. canis and E. equi antigens. There was concordance of E. canis IFA and western immunoblot (WI) test results for 36/44 dogs. Because of cross-reactivity of E. canis sera with E. equi antigens, WI was of less utility to confirm E. equi exposure. After elimination of E. canis seroreactors, there was concordance of 2/4 E. equi IFA and WI test results. Based upon a retrospective review of medical records, ehrlichiosis was diagnosed in 10/48 (21%) IFA seroreactive dogs, 9 of which were confirmed positive by WI. Of the remaining 38 IFA seroreactors, 29 also were confirmed by E. canis or E. equi WI. These results indicate that (1) ehrlichiosis was not diagnosed in the majority of serologically confirmed cases, (2) based upon E. canis and E. equi WI analysis, IFA testing was not specific (21% false positive), (3) E. canis sera cross-react with E. equi antigens, and (4) serologic evidence of E. risticii infection was lacking in the dog population studied.     

Concurrent babesiosis and ehrlichiosis in the dog: blood smear examination supplemented by the indirect fluorescent antibody test, using Cowdria ruminantium as antigen

Giemsa-stained, peripheral blood smears of 67 dogs, showing clinical signs typical of babesiosis or reminiscent of concurrent babesiosis and ehrlichiosis, were examined for the presence of Babesia canis and Ehrlichia canis. Since Cowdria ruminantium cross-reacts with Ehrlichia, the sera of these dogs were also subjected to the indirect fluorescent antibody (IFA) test in which C. ruminantium was used as antigen. Fifty-five per cent of these dogs had mixed infections of B. canis and E. canis, as judged by blood smear examination and serology. The serum of 32% of these dogs with mixed infections reacted positively in the IFA test. Six out of 9 dogs, the blood smears of which were negative for both B. canis and E. canis, were serologically positive for E. canis. Furthermore, sero-conversion from a negative in the initial serum sample to titres of up to 1:160 in a subsequent sample was recorded in 9 out of 13 dogs with suspected mixed infection on blood smear.     

Serological survey for Ehrlichia canis in urban dogs from the major population centres of northern Australia

OBJECTIVE: To detect evidence of Ehrlichia canis infection of dogs from the major population centres of northern Australia, if present. DESIGN: Serological investigation for E. canis. PROCEDURE: The sera of 316 domestic dogs, collected from the northern Australian population centres of Townsville, Cairns, Darwin, Kununurra and Broome from May 1997 to August 1999, were investigated for evidence of infection with E. canis. Samples were tested for antibodies to E. canis using an indirect fluorescent antibody (IFA) test. The buffy coats from blood of dogs whose serum reacted in the IFA test were subsequently tested with a nested PCR to detect E. canis DNA. When available, blood from these dogs was injected into suckling mice, which were then examined for clinical disease and tested for the presence of E. canis antibodies. RESULTS: Of the 316 samples tested seven reacted in the IFA test for E. canis. None of the dogs from which these samples were obtained exhibited clinical signs of acute or chronic ehrlichiosis. The six positive samples available for testing were negative when tested with the nested PCR. Suckling mice inoculated with blood from three of the dogs whose serum was positive by IFA test showed no signs of clinical disease nor did their give positive reactions in the IFA test. CONCLUSIONS: No evidence of E. canis infection was confirmed in any of the dogs examined. Northern Australia would appear to remain free of this obligate parasite.     

Detection of Ehrlichia canis by PCR in different tissues obtained during necropsy from dogs surveyed for naturally occurring canine monocytic ehrlichiosis

A molecular study for the detection of Ehrlichia canis was carried out on tissues obtained at necropsy from randomly selected dogs with the intention of investigating naturally-occurring canine ehrlichiosis. The tissues evaluated for the presence of E. canis included lymph nodes, spleen, liver, bone marrow, and blood. Eight of the 18 dogs included were found to be positive for E. canis by polymerase chain reaction (PCR) and sequencing of the 16S rRNA gene. Two dogs were positive for Anaplasma platys of which one dog was co-infected with E. canis and A. platys. Blood (5/8) and lymph nodes (5/8) were the tissues found to yield the highest number of positive E. canis PCR results with 7/8 dogs positive in the blood or lymph node. E. canis and A. platys DNA could be amplified by PCR when tissue samples were obtained 72h after the time of death.     

Investigation of renal protein loss in dogs with acute experimentally induced Ehrlichia canis infection.

Urinary protein-to-creatinine ratios and serum albumin concentrations were measured in 8 adult male dogs experimentally inoculated with Ehrlichia canis. Urinary protein concentration increased significantly, but transiently, during the acute phase of infection. Urinary protein-to-creatinine ratios were highest (mean, 8.6) during the third and fourth weeks after infection, and decreased to less than 0.5 by 6 weeks after infection. Correspondingly, albumin concentration decreased significantly during the acute phase. Serum albumin concentrations were lowest (mean, 2.1 g/dl) the fourth week after infection and increased to greater than 3.0 g/dl by 11 weeks after infection. There was an inverse linear correlation between urinary protein-to-creatinine ratio and serum albumin concentration. The magnitude of proteinuria and its inverse relationship with serum albumin concentration suggested that hypoalbuminemia associated with acute E canis infection may be attributable primarily to increased renal loss of protein, rather than decreased hepatic synthesis as previously suggested. Another dog was subsequently inoculated with E canis from 1 of the experimentally infected dogs and a renal biopsy was performed during peak proteinuria (urinary protein-to-creatinine ratio = 22 and serum albumin = 1.1 g/dl). Immunofluorescent staining revealed mild to moderate deposits of anti-canine IgM, and to a lesser extent, anti-canine IgG and complement factor C3 in the glomerular tufts and mesangium. Ultrastructural evaluation revealed distortion and fusion of podocyte foot processes and increased microvilli on podocytes. These morphologic changes were consistent with transient glomerular leakage of protein of a magnitude that would significantly contribute to hypoalbuminemia during acute E canis infection. An underlying immunologic mechanism was suggested by positive glomerular immunofluorescence and previously described histologic findings.     

Efficacy of a modified polymerase chain reaction assay for detection of Ehrlichia canis infection.

Detection of Ehrlichia canis in acutely infected and convalescent dogs is important for effective treatment and control. However, accurate detection has been difficult to achieve, in part because dogs that have been treated therapeutically often remain seropositive for extended periods. A new method, polymerase chain reaction (PCR) assay using biotinylated E. canis-specific primers (PCR-BP), was developed for detection of E. canis. Four dogs experimentally infected with E. canis by intravenous inoculation of whole blood from carrier dogs and 2 naturally infected convalescent carriers were used to compare the specificity and sensitivity of the new method with that of microscopy/blood smear evaluation, serologic test, and conventional PCR assay using E. canis-specific primers. In experimentally infected animals, infection was detected as early as 7 days post-exposure using PCR-BP. Although the 2 naturally infected dogs were positive by serologic test and PCR-BP, both were negative by conventional PCR. Results suggest that the new method is a sensitive assay for detection of E. canis infection. In addition, results were obtained more rapidly than with other PCR-based assays.