The required blood samples from live or recently deceased (including euthanized) animals are (Waner, 2022): ● For PCR testing, 2–5 ml in an EDTA tube (purple top), ● For serological testing, 2–5 ml in a serum separator tube (yellow top) or plain tube (red or grey/red speckled top). If possible, a 1–2 ml aliquot of clear serum should be obtained. Samples to collect from dead animals are: ● Unclotted heart blood (if available), ● Fresh and formalin-fixed samples of brain, lung, spleen, liver, kidney and ● Submandibular lymph nodes. A result of the presence of E. canis IgG antibodies should not be considered as definitive for an E. canis exposure, because serological cross-reactivity exists between Ehrlichia canis, Ehrlichia. ewingii, Ehrlichia chaffeensis, Anaplasma phagocytophilum, Neorickettsia risticii, and Neorickettsia helminthoec a. (Krämer et. al., 2020) PCR result does not necessarily signify that there was no E. canis DNA in the organ of the dog under examination. On the other hand, the presence of DNA in an organ does not necessary indicate that there are live or viable organisms. (Krämer et. al., 2020)
splenomegaly, dyspnea, anterior uveitis, retinal hemorrhage, and edema. As a result of the effect on the bone marrow, secondary opportunistic infections, such as protozoal infections, viral papillomatosis, and bacterial urinary tract infections, can also develop. (Krämer et. al., 2020) Infected dogs are also more susceptible to secondary infections, and therefore may present with non-healing wounds, dental infections, and apparently resistant infections or treatment failures. Many present with significant and rapid wasting, even if the dog is well fed. This form of the disease is usually fatal and treatment efforts are likely to be futile. (Waner, 2022) Diagnostic tests A dog’s infection status should be interpreted based on laboratory results, clinical signs, and response to previous treatment. E. canis can be detected by real-time polymerase chain reaction (PCR). Serological tests include enzyme-linked immunosorbent assay (ELISA) and an indirect fluorescent antibody test (IFAT). (Waner, 2022) Polymerase chain reaction (PCR) test Polymerase chain reaction (PCR) is a very sensitive and specific method for detection and identification of E. canis DNA in either blood, bone marrow, or splenic aspirates. A negative PCR result might be due to the fact that the concentration of DNA in the collected sample may be below the detection level of the test. Therefore, a negative
Table 1. Diagnosis of Canine Monocytic Ehrlichiosis (Cme) Using a Combination of Serological and PCR Tests SEROLOGY ELISA TEST Positive PCR • Indicates exposure to E. canis rickettsia or an infection with a cross-reacting rickettsial infection. • Acute infection: a dog with a history of tick infestation, thrombocytopenia, and with a positive and increasing antibody titer. • A positive blood PCR result at this stage would confirm an ongoing active specific infection with E. canis and authenticate the diagnosis. Positive ELISA test result
Negative ELISA test result
• Negative blood PCR result: PCR testing of spleen biopsy samples should be considered, as this is regarded as more sensitive than PCR blood results. • In the case of an early infection of E. canis by ticks where antibodies may not yet have developed, a positive PCR result with or without a finding of thrombocytopenia would increase the possibility of an imminent acute disease with E. canis . In this case, detection of E. canis DNA may be useful as early as 3-10 days after infection by ticks, where the incubation period is 8-20 days and antibodies will only appear after 15 days. • A negative PCR result together with negative serology is suggestive of a disease not associated with E. canis. • Since there is always a possibility of a mixed infection involving other types of tick-borne diseases, the veterinarian should always undertake a though blood smear examination in order to eliminate other tick- borne infections and possibly carry out further PCR tests specifically for other tick borne diseases.
Negative PCR
Acute, subacute or subclinical infection
Acute infection
A dog exhibiting suggestive symptoms of CME
Subacute, subclinical or chronic infection, or recovered
Not infected
Notes . (Waner, 2022) (Krämer et. al., 2020)
Differential diagnoses The clinical signs of ehrlichiosis are nonspecific and occur in many systemic diseases of an infectious and/or inflammatory nature, including immune-mediated diseases and neoplasia. Differential diagnoses may include anaplasmosis, babesiosis, lymphoma, multiple myeloma, and immune-mediated thrombocytopenia. (Waner, 2022) Treatment Spontaneous clinical recovery of acutely infected dogs is common; however, dogs at this stage require medical treatment in order to hasten their clinical recovery and to prevent clinical exacerbation or death. An unpredictable
proportion of subclinically infected dogs will eventually develop the chronic, severe form of ehrlichiosis, characterized by aplastic pancytopenia and high mortality. (Australian Government, 2023) The aims of antimicrobial treatment in CME include the achievement of clinical remission, resolution of the clinicopathologic abnormalities, and eradication of the infection, although the latter is not always feasible or diagnostically confirmable. Treatment of dogs with aplastic pancytopenia should be undertaken with the clear understanding that medical management will require long-
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