Acute Canine Monocytic Ehrlichiosis Research Articles

Novel renal injury markers in dogs with ehrlichiosis

Canine monocytic ehrlichiosis (CME) has been observed to impact renal function. Currently, the recognition of acute kidney injury is through the nonspecific biomarker serum creatinine (sCr). Novel markers of renal injury such as urinary clusterin (uClust) and urinary cystatin B (uCysB) may increase our understanding of the relationship between ehrlichiosis and renal cellular injury. The aim of this study was to evaluate novel renal injury biomarkers in dogs with acute CME. Twenty healthy dogs were enrolled in the control group (CG), and 16 dogs naturally infected with Ehrlichia canis were included in the Ehrlichia Group (EG). All dogs were followed for 45 days. EG dogs were treated with doxycycline twice daily for the first 30 days. Urine and serum were collected at: 0, 0.5, 1, 15, 30, and 45 days after start of treatment. Urine concentrations of uClust and uCysB were determined using a research ELISA immunoassay. A linear mixed model was used to estimate population mean of renal injury markers with patient as the random effect, and day and treatment as fixed effects. EG was observed to have higher uClust values compared to CG (estimated population mean EG: 213 ng/dL vs. CG: 84 ng/dL, P < 0.001). EG was observed to have higher uCysB values compared to CG (estimated population mean EG: 248 ng/dL vs. CG: 38 ng/dL, P < 0.001). Increases in uCysB and uClust suggest the presence of renal injury and a possible mechanism for the observed predisposition to chronic kidney disease in dogs with ehrlichiosis.

Clinical and Clinicopathologic Discriminators Between Canine Acute Monocytic Ehrlichiosis and Primary Immune Thrombocytopenia

Acute canine monocytic ehrlichiosis due to Ehrlichia canis (aCME), and primary immune thrombocytopenia (pITP) are major differentials for dogs presented with thrombocytopenia, and the two diseases may clinically overlap. The aim of this study was to compare dogs diagnosed with naturally occurring aCME and pITP, to establish potentially useful clinical and clinicopathologic discriminators. A clinical record-based retrospective study was performed in 35 dogs diagnosed with aCME and 29 dogs with pITP. Dogs with aCME were significantly younger, and were more likely to experience depression or lethargy, anorexia, body weight loss, fever, lymphadenomegaly, tick infestation, and ocular discharge on admission, compared to dogs with pITP. In contrast, dogs with pITP presented more frequently with overt bleeding and had a significantly higher bleeding score compared to dogs with aCME. Dogs with aCME were more likely to be anemic and hypoalbuminemic on presentation compared to dogs with pITP. Dogs with pITP had higher white blood cell and neutrophil counts as well as lower platelet counts than dogs with aCME and were more likely to present with leukocytosis, neutrophilia and monocytosis. These clinical, hematological, and biochemical findings may be helpful discriminators between aCME and pITP, on the understanding that they will be interpreted in the context of disease-specific testing.

Acute Canine Monocytic Ehrlichiosis and Sertoli Cell Tumour in an Eleven-‎Year-Old Dachshund Cross-Bred Dog

An eleven-year-old Dachshund cross-bred dog was presented to the Small Animal Clinic of Ahmadu Bello University Veterinary Teaching Hospital, Zaria, with complaints of reduced appetite noticed 2 days earlier, and an enlarged testis was noticed three months prior to presentation. Clinical diagnosis was based on history of reduced appetite and enlarged testis; physical findings such as pyrexia, tick infestation, congested ocular mucous membranes, generalized enlarged superficial lymph nodes, enlargement of the mammary glands, alopecia, atrophy of the right testis and discoloration of the scrotum with large, firm palpable mass; complete blood count revealed slight anaemia (PCV 34 %, neutrophilia (12.88 x 109/L) and presence of band cells (0.32 x 109/L); buffy coat smear showed the presence of intracytoplasmic morulae in just one monocyte in the peripheral blood smear; Ehrlichia canis antibodies was detected in serum using Ehrlichia test kit (immunochromatography); serum oestradiol-17β (42.05 pg/mL) and anti- Müllerian hormone (AMH; 25.50 μg/mL) were employed as serum tumour markers; transverse and longitudinal scrotal sonographs revealed diffuse non-homogenous tissues with increase in the echogenicity of the testicles; radiographic findings of thoraco-abdomino-pelvic regions revealed normal organ opacity indicating absence of metastases. Tissue samples were taken from the tumor following surgical excision and submitted for histopathology. The findings were large cytoplasmic vacuoles with slightly irregular nucleoli. It was confirmed to be sertoli cell tumor. The conditions were managed medically, and by surgical excision of the tumour and scrotal ablation. Outcome was successful, and patient started feeding 24 hours after the surgery.

Detection of specific IgM and IgG antibodies in acute canine monocytic ehrlichiosis that recognize recombinant gp36 antigens

The efficacy of antibody detection tools for all stages of Ehrlichia canis infections and for various genotypes remains unclear. We produced recombinant gp36 (rgp36) antigens from different isolates of Thai E. canis to confirm the immunoreactivities to these recombinant proteins from naturally infected dogs. Sera and blood samples were taken from 21 dogs naturally infected with E. canis and in the clinical stages of acute phase ehrlichiosis. The expression vectors and competent E. coli produced two isolates of rgp36. These two major rgp36s were recognized by the dogs' sera in Western blotting, with both anti-dog IgM and IgG used as secondary antibodies. The two different genotypes of these local recombinant immunoreactive proteins were gp36 subgroup A (isolate 1055) and subgroup B (isolate 533). The Western blot analyses successfully identified both specific IgM and IgG from the dogs' sera. Of all 21 cases, five dogs presented specific IgM, twenty dogs presented specific IgG, and the commercial test used found fifteen seropositive dogs. There were four dogs that presented both specific IgM and IgG. Only one dog presented specific IgM only. This report is the first identification of a specific IgM in dogs in response to acute infections with E. canis. The recombinant gp36 isolates may be useful as potential antigenic material for subsequent serological tests that have a high possibility for differentiating between acute, chronic, primary, and nonprimary infections with E. canis.

Conventional and Holter Electrocardiographic Assessment of Dogs Infected Naturally With Acute Canine Monocytic Ehrlichiosis

Canine Monocytic Ehrlichiosis (CME) is a disease of worldwide distribution caused by the bacteria Ehrlichia canis, appearing primarily in hot climates due to the massive prevalence of its vector, the tick Rhipicephalus sanguineus. Previous studies have shown that dogs afflicted by CME in the chronic phase can develop infectious myocarditis, arrhythmias, and alterations in heart rate variability (HRV), but there are few studies correlating cardiac diseases with the acute phase of CME. This study aims at assessing electric cardiac alterations and HRV in the time and frequency domains during the acute phase of CME. This study assessed 22 animals divided into 2 distinct groups: the control group, comprised by 10 healthy dogs, and the sick group, comprised of 12 dogs infected naturally with ehrlichiosis which presented clinical and hematological signs compatible with the acute phase of the disease. The animals underwent conventional and Holter electrocardiographic evaluations, systolic blood pressure measurement, complete blood count and biochemical assays (urea, creatinine, alanine aminotransferase (ALT), alkaline phosphatase (AP), and gamma glutamyl transferase (GGT)). The sick group presented higher activity in the sympathetic nervous system than in the parasympathetic nervous system, manifest as a significant increase in mean heart rate and a reduction in the HRV indexes for the time and frequency domains. The frequency-domain HRV indexes presented sympathetic prevalence during the sleep and vigilance states. Sinus tachycardia was the predominant heart rhythm in 58.33% of the animals. The mean systolic blood pressure diverged between the groups and no significant arrhythmias were observed during monitoring. The serum concentrations of urea, creatinine, AP, ALT, and GGT were within the established reference values for the species. We observed no indication that there was enough time during the acute phase for the disease to evolve in a way that resulted in arrhythmias, as is common in the chronic phase, but we observed that animals in the acute phase already present reduced HRV indexes.

Η κλινική χρησιμότητα των ορολογικών δοκιμών στη μονοκυτταρική ερλιχίωση (Ehrlichia canis) του σκύλου

Canine monocytic ehrlichiosis (CME) is primarily caused by Ehrlichia canis, a member of the genus Ehrlichia (family: Anaplasmataceae). Serology is the most frequently used diagnostic modality in CME. Indirect fluorescent antibody (IFA) testing remains the “gold standard” for the detection and titration of the E. canis-specific antibodies; in addition, several user-friendly, in-clinic assays have recently gained tremendous popularity due to their practical and cost-effective nature. The latter assays provide mostly qualitative results and have a high diagnostic specificity; however, their sensitivity usually declines when the IFA-titers are lower than 1/320, suggesting a suboptimal performance in acutely infected dogs.Overall, a positive IgG antibody titer indicates past exposure to and infection by Ehrlichia canis or a closely related agent (e.g. E. chaffeensis, E. ewingii), while rarely, true false positives may adversely affect the interpretation. As a rule, a negative IgG antibody titer rules out exposure to E. canis, with the notable exception of the acute CME in which clinical and clinicopathological manifestations may preceed seroconversion which occurs 7-35 days post-infection. The latter problem may be overcome by testing paired serum samples obtained 2-3 weeks apart. Unlike IgG antibodies, IgM titers develop inconsistently in the course of the infection, thus, not justifying their routine measurement for diagnostic purposes. Importantly, IgG antibody titers do not reliably correlate with the duration of infection, the current carrier status, or the presence and severity of clinical disease. Because of the prolonged latent period and the persistent seropositivity following therapy or self-eradication of the infection, clinicians should be well aware that seroreactivity to E. canis, especially in an endemic area, does not unequivocally confirm that the clinical manifestations and the clinicopathological abnormalities are due to E. canis infection. The decision to treat a clinically healthy, seropositive dog may be particularly challenging, especially in endemic areas.A positive or negative polymerase chain reaction (PCR) result dictates towards or against treatment, respectively. If PCR is not available, the proper course of action should be decided on a case-by-case basis. The authors suggest treating these dogs if they are thrombocytopenic and/or hyperglobulinemic and no other potential causes of these abnormalities (e.g. pseudothrombocytopenia and/or comorbid conditions) can be demonstrated. The antibody kinetics is quite unpredictable, frequently persisting several months to years following eradication of the organism, which limits the value of serology as a post-treatment monitoring tool. This review article addresses a series of questions pertaining to the interpretation of E. canisspecific serology in the context of the clinical phase of the disease, the antibody kinetics, the sensitivity and specificity of the assays and the prior treatment status. The ultimate goal is to facilitate the clinical decision-making towards the diagnosis and post-treatment monitoring of the CME.

Quantitative and qualitative leukocyte abnormalities in dogs with experimental and naturally occurring acute canine monocytic ehrlichiosis.

Canine monocytic ehrlichiosis (CME) is one of the most important tick-borne diseases worldwide. Cytopenias have been observed in both acute (nonmyelosuppressive) and chronic (myelosuppressive) CME; however, leukocyte abnormalities and indices have been incompletely described in dogs with acute CME. The aims of this study were to analyze temporal changes in differential leukocyte counts, leukocyte morphology, myeloperoxidase index (MPXI), and lobularity index (LI) in dogs with experimental and naturally occurring acute CME. Differential leukocyte counts and morphology were evaluated in archived blood smears from 13 Beagle dogs experimentally infected with Ehrlichia canis and evaluated weekly for 42days postinfection (DPI); 20 dogs with naturally occurring acute CME also were evaluated. MPXI and LI were obtained from ADVIA reports. Wilcoxon tests were used to assess changes over time; leukogram results in natural cases were assessed in comparison with reference intervals. In experimental dogs, significant decreases in neutrophil, monocyte, lymphocyte, and eosinophil counts, and a mild left shift occurred within 14 DPI. The MPXI decreased significantly between 14 and 21 DPI and remained low, while LI increased from 14 to 35 DPI. Lymphocyte counts rebounded at 21 DPI, normalizing total WBC counts. Neutrophil toxicity was seen rarely, but reactive lymphocytes were observed frequently. Dogs with natural infection had variable patterns of leukocyte changes. Acute CME is associated with several discrete quantitative and qualitative leukogram changes indicative of concurrent inflammation, antigenic stimulation, and stress. Changes in MPXI and LI warrant further investigation in dogs with CME and other diseases.

Acute phase protein and antioxidant responses in dogs with experimental acute monocytic ehrlichiosis treated with rifampicin

There is currently lack of information on the changes of acute phase proteins (APP) and antioxidant markers and their clinical relevance as treatment response indicators in canine monocytic ehrlichiosis (CME). The objective of this study was to investigate the patterns of C-reactive protein (CRP), haptoglobin (Hp), ferritin and paraoxonase-1 (PON-1) during treatment of dogs with acute CME with rifampicin. Blood serum samples from ten Beagle dogs with experimental acute CME were retrospectively examined. Five dogs (Group A) were treated with rifampicin (10mg/Kg/24h), per os, for 3 weeks and 5 dogs (Group B) received no treatment (infected controls). Two Beagle dogs served as uninfected controls. Blood serum samples were serially examined prior to Ehrlichia canis inoculation and on post-inoculation days 14, 21, 28, 35 and 42. Significant changes of CRP, Hp, ferritin and PON-1 values were found in the majority of infected dogs. However, their concentrations did not differ between the two groups during the treatment observation period. The results of this study indicate that although several APP and PON-1 tend to significantly change in the majority of dogs with acute CME, they were of limited clinical relevance as treatment response indicators in this experimental setting.

Synovial fluid cytology in experimental acute canine monocytic ehrlichiosis (Ehrlichia canis)

Evidence-based information of a cause-and-effect relationship between Ehrlichia canis infection and polyarthritis in naturally- or experimentally-infected dogs is currently lacking. The aim of this prospective study was to investigate whether synovial fluid cytological evidence of arthritis could be documented in dogs with acute monocytic ehrlichiosis. Direct synovial fluid cytology smears from eight Beagle dogs experimentally infected with E. canis were examined prior to, and on 21, 35 and 63 days post-inoculation. The cytological variables assessed included cellularity, percentages of mononuclear cells and neutrophils, macrophage reactivity and evidence of E. canis morulae. The median cellularity and percentages of mononuclear cells and neutrophils prior to inoculation did not differ when compared to post-inoculation cytological evaluation. Increased cellularity, E. canis morulae or cytological evidence of arthritis or macrophage reactivity were not observed throughout the course of the study. In the present study, no cytological evidence of arthritis was found in dogs with experimental acute canine monocytic ehrlichiosis, suggesting that E. canis infection should be considered a rather uncommon cause of arthritis in dogs.

Evaluation of a commercial in-clinic point-of-care polymerase chain reaction test for Ehrlichia canis DNA in artificially infected dogs

A novel in-clinic point-of-care (ICPOC) polymerase chain reaction (PCR) test was evaluated for its ability to detect Ehrlichia canis DNA in artificially infected dogs compared to a real-time PCR assay. Six Beagle dogs negative for E. canis antibodies and PCR negative were artificially infected with an Israeli E. canis strain (611). All dogs developed IgG antibodies 8 days post infection (PI), and clinical and hematological abnormalities on day 10 PI.Only the real-time PCR detected E. canis DNA in the blood of five dogs at days 3 and 5 PI. At day 12 PI during the acute phase of the disease, 1 day after the initiation of doxycycline treatment, the ICPOC PCR assay detected E. canis DNA in all infected dogs, which were also positive by the real-time PCR. Two days later the ICPOC PCR assay was able to detect only 3/6 infected dogs, which were all positive by the real-time PCR. At days 17 and 19 PI, the ICPOC PCR assay did not detect E. canis DNA in the dogs while the real-time PCR detected all dogs as positive on day 17 PI and two dogs on day 19 PI. In conclusion, the sensitivity of the ICPOC PCR assay was 75% for the acute phase of the disease and 30% for the whole study, suggesting that this ICPOC assay has a potential utility for the diagnosis of acute canine monocytic ehrlichiosis.

Efficacy of rifampicin in the treatment of experimental acute canine monocytic ehrlichiosis

To assess the efficacy of rifampicin in achieving clinical and haematological recovery and clearing infection in dogs with experimentally induced acute monocytic ehrlichiosis. Five Ehrlichia canis-infected Beagle dogs were treated with rifampicin (10 mg/kg/24 h orally for 3 weeks), nine E. canis-infected dogs received no treatment (infected untreated dogs) and two dogs served as uninfected controls. Clinical score, platelet counts, immunofluorescent antibody titres and PCR detection of E. canis-specific DNA in blood, bone marrow and spleen aspirates were evaluated on post-inoculation days 21 (start of rifampicin), 42 (end of rifampicin) and 98 (end of the study). By day 21 post-inoculation, all infected dogs became clinically ill and thrombocytopenic, seroconverted and were PCR positive in at least one tissue. Clinical scores and antibody titres did not differ between the treated and infected untreated dogs throughout the study. The rifampicin-treated dogs experienced an earlier resolution of their thrombocytopenia (Kaplan-Meier survival plot, P=0.048), and the median platelet counts were significantly higher in the treated compared with the infected untreated dogs on post-inoculation days 42 (P=0.0233) and 98 (P=0.0195). At the end of the study, three treated and six untreated infected dogs remained PCR positive in one tissue each. The rifampicin treatment regimen applied in this study hastened haematological recovery, but was inconsistent in eliminating the acute E. canis infection.

Monocytic Ehrlichiosis in dogs

Ehrlichiosis is the multiorgan infectious disease caused by small, intracellular rickettsias from the genus Ehrlichia. These microorganisms are known as an etiologic factor of infections world wide in humans and in different species of animals. Dog ehrlichiosis can be caused by several species of Ehrlichia attacking different groups of blood cells, but most often an infection by Ehrlichia canis is diagnosed with special relation to monocytes. A vector for E. canis are Rhipicephalus sanguineus and Ixodes ricinus, commonly occurring in Poland. Disease caused by E. canis is known as Canine Monocytic Ehrlichiosis (CME). The disease most often has an asymptomatic course which can, in favourable circumstances, run into acute or chronic forms. The acute form of CME proceeds usually with fever, apathy, weakness and accompanying respiratory symptoms, lameness and disturbances in blood coagulation. In laboratory examinations thrombocytopenia, anemia and leucopenia are ascertained. The chronic form of CME proceeds among gentle, unspecific symptoms which may last even 5 years. The CME diagnosis is difficult and often demands parallel different diagnostic methods. A medicines of choice in the ehrlichiosis treatment are antibiotics from the group of tetracyclines, given at least for 28 days. They are largely efficient during treatment of the acute CME, causing the quick improvement. Instead, in the case of chronic form, answer for treatment can be weak, and cases of resistance to antibiotics ave known.

Comparison of Simultaneous Splenic Sample PCR with Blood Sample PCR for Diagnosis and Treatment of Experimental Ehrlichia canis Infection

This report presents evidence that dogs recover from acute canine monocytic ehrlichiosis (CME) after 16 days of doxycycline treatment (10 mg/kg of body weight every 24 h). Blood PCR was as valuable as splenic aspirate PCR for early diagnosis of acute CME. Splenic aspirate PCR was, however, superior to blood PCR for the evaluation of ehrlichial elimination.

Evaluation of cytology in the diagnosis of acute canine monocytic ehrlichiosis ( Ehrlichia canis): a comparison between five methods

The purpose of this study was the comparison of the diagnostic sensitivity between buffy coat (BC), peripheral blood (PB), lymph node (LN), bone marrow (BM) and short-term culture (P-D) cytology that has been based on the detection of Ehrlichia canis morulae, in the acute phase of canine monocytic ehrlichiosis (CME). Their cellular localization, total numbers and microscopic differentials were also investigated. The highest sensitivities were achieved after evaluating 1000 oil immersion fields (OIFs) in BC (66%) and an equal number in LN (60.9%) smears, separately or together (74%). The morulae were more often detected into lymphocytes than monocytes. The highest total number of morulae ( n=143) were found in P-D smears. Finally, to avoid false positive diagnoses, platelets, lymphocytic azurophilic granules, lymphoglandular bodies and phagocytosed nuclear material should not be confused with the morulae.