A canine case of Ehrlichia canis infection without a history of being in an endemic area in Japan.

A mixed-breed, 8-year-old male dog developed neutropenia, thrombocytopenia, and hyperglobulinemia. Bone marrow hyperplasia and splenic plasmacytosis were cytologically observed. The dog had never been outside of Tokyo or Shizuoka Prefecture. Splenectomy was performed to confirm and remove the cause of splenic plasmacytosis. A histopathological diagnosis of splenic plasmacytoma was made; however, serum protein electrophoresis showed polyclonal gammopathy. Further screening was performed, and Ehrlichia canis infection was confirmed. The dog was treated with doxycycline for 5 weeks. After the antibiotic therapy, no relapse of neutropenia, thrombocytopenia, hyperglobulinemia, or positive polymerase chain reaction result of E. canis infection was observed for 3 years. Careful attention should be given to ehrlichiosis when exploring the cause of pancytopenia or hyperglobulinemia, regardless of the travel history.

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.

Correlation between canine biochemical analytes and TRP36 ELISA seropositivity for Ehrlichia canis in Brazil.

BACKGROUND: Synthetic peptides of tandem repeat proteins (TRPs) have been employed in the serologic analysis of canine monocytic ehrlichiosis (CME) and used in epidemiological studies in Brazil. Based on molecular studies of TRPs, different genotypes of Ehrlichia canis have been described, but data on their pathogenicity remain unknown. OBJECTIVES: To correlate hepatic, renal, and muscular alterations in relation to different genotypes of E. canis in naturally exposed dogs using enzyme-linked immunosorbent assay (ELISA) with TRP19 and TRP36 synthetic protein antigens. METHODS: Two hundred serum samples were subjected to ELISA with the antigens of TRP19 and three genotypes (US, Br, and CR) of TRP36 of E. canis circulating in Brazil. Positive sera were evaluated through eight biochemical parameters, and the results were evaluated by principal component analysis and canonical correlation. RESULTS: ELISA revealed that 47 (23.5%) serum samples reacted to the BrTRP36 peptide, 36 (18%) reacted to the TRP19 peptide, and 8 (4%) reacted to the USTRP36 and CRTRP36 peptides separately. The most frequent biochemical alterations observed were for CK (59.4%), ALB (31.8%), GLO (28.9%), TP (28.9%), ALP (26%), urea (24.6%), creatinine (14.4%), and ALT (14.4%). The most prominent diagnostic method in canonical correlation analysis was BrTRP36, followed by TRP19, which correlated with hyperglobulinemia and hypoalbuminemia. CONCLUSIONS: Antibodies that reacted against the Brazilian genotype of E. canis correlated positively with hyperglobulinemia and increases in serum urea and creatinine. According to our results, the Brazilian genotype of E. canis is related to the chronic phase of CME.

Detection of vector-borne pathogens in owned dogs with cranial cruciate ligament rupture living in the Mediterranean area.

BACKGROUND: Cranial cruciate ligament rupture (CCLR) results from a multifactorial degenerative process that leads to rupture of the ligament. Vector-borne pathogens (VBP) in dogs can induce joint disease but their role in CCLR has not been previously investigated. The aim of the present work is to evaluate the prevalence of VBP in dogs with CCLR. METHODS: This was a prospective study that included 46 dogs presented for CCLR surgical treatment and 16 control dogs euthanized for diseases unrelated to the joints. Specimens collected included blood, synovial fluid, and synovial membrane biopsy. Pathogen testing consisted of serology for Leishmania infantum (quantitative ELISA), Ehrlichia canis/ewingii, Borrelia burgdorferi, Anaplasma phagocytophilum/platys, and Dirofilaria immitis (4DX IDEXX test), and PCR for L. infantum, Ehrlichia/Anaplasma spp., Bartonella spp., piroplasms (Babesia spp. and Theileria spp.), and filariae (D. immitis, Dirofilaria repens, Acanthocheilonema dracunculoides, Acanthocheilonema reconditum, and Cercopithifilaria spp.) on both EDTA-whole blood (EB) and synovial fluid (SF) samples. SF cytology and histopathological evaluation of synovial membrane were also performed. RESULTS: The prevalence of VBP was 19.6% in the CCLR group and 18.8% in the control group, with no statistical difference among them. The presence of synovitis was not more frequent in CCLR dogs (45.6%) than in control dogs (43.7%). Lymphoplasmacytic infiltration was the most common inflammatory pattern detected in the joints of both groups of dogs. CONCLUSIONS: This study failed to demonstrate a role of canine VBP in CCLR or the presence or different pattern of joint inflammation in pathogen-positive dogs.

Elective ovariohysterectomy or castration can proceed in shelter dogs despite mild to moderate presurgical leukocytosis.

OBJECTIVE: To determine whether shelter dogs presenting for elective ovariohysterectomy or castration have leukocytosis, whether leukocytes are associated with age and infection, and whether leukocytosis precludes progression to surgery. ANIMALS: 138 dogs (from 13 regional shelters) presented for ovariohysterectomy or castration between October 7 and December 6, 2019. PROCEDURES: For this prospective study, each dog underwent presurgical physical examination, CBC, and tests for Dirofilaria immitis antigen and Anaplasma phagocytophilum, Borrelia burgdorferi, and Ehrlichia canis antibodies, with additional tests performed as needed. Dogs were aged by dentition as juvenile (< 3 or ≥ 3 to ≤ 6 months) or adult (> 6 months). Leukogram results were compared across age groups with recognized infections and parasitism and with dogs' progression to surgery. RESULTS: There were 34 dogs < 3 months old, 22 dogs ≥ 3 to ≤ 6 months old, and 82 > 6 months old. Sixty-three of 138 (45.6%) dogs had leukocytosis (median, 16,500 cells/µL; range, 13,700 to 28,300 cells/µL). Dogs < 3 months of age had higher median leukocyte and lymphocyte counts (14,550 cells/µL and 3,700 cells/µL, respectively) than dogs > 6 months of age (12,500 cells/µL and 2,400 cells/µL, respectively). Only 1 dog had a stress leukogram. Forty-seven dogs had recognized infection, but there was no association with leukocytosis. Surgery proceeded successfully for all dogs with leukocytosis. CLINICAL RELEVANCE: Mild to moderate leukocytosis is common before elective surgery in shelter dogs, but surgery can proceed safely. A CBC should be reserved for ill-appearing dogs rather than as a screening test, and age-specific reference intervals should be considered.

Seroprevalence and hematological abnormalities associated with Ehrlichia canis in dogs referred to a veterinary teaching hospital in central-western Brazil / Soroprevalência e alterações hematológicas associadas com Ehrlichia canis em cães encaminhados a um hospital veterinário-escola no centro-oeste do Brasil

Ticks are significant parasites of dogs in the tropics, where tick-borne pathogens are highly prevalent, especially in areas where tick control measures are frequently neglected. This study investigated the seroprevalence and hematological abnormalities associated with Ehrlichia canis in dogs referred to a veterinary teaching hospital in Central-western Brazil. Out of 264 dogs tested for anti-Ehrlichia canis antibodies by an indirect immunofluorescence assay (IFA), 59.1% (156/264) were positive. Seropositivity was significantly associated to anemia and thrombocytopenia, alone or in combination, and to leukopenia. Conversely, there were no differences in terms of seroprevalence according to sex, breed and age. This study demonstrated that dogs referred to a veterinary teaching hospital in Central-western Brazil are highly exposed to E. canis and that seropositive dogs are more likely to present hematological abnormalities, particularly anemia, thrombocytopenia and leukopenia. To our knowledge, this is the first study on detection of anti-E. canis antibodies by means of IFA among dogs in the state of Goiás. These findings highlighted the need for increasing awareness among dog owners regarding tick control measures in Central-western Brazil, ultimately to reduce the risk of exposure to E. canis and other tick-borne pathogens.

Carrapatos são importantes parasitos de cães nos trópicos, onde patógenos transmitidos por carrapatos são altamente prevalentes, especialmente em áreas onde as medidas de controle de carrapatos são frequentemente negligenciadas. O estudo investigou a soroprevalência e as anormalidades hematológicas associadas à Ehrlichia canis em cães encaminhados para um hospital veterinário-escola no Centro-oeste do Brasil. Dos 264 cães testados para anticorpos anti-Ehrlichia canis por meio da reação de imunofluorescência indireta (RIFI), 59.1% (156/264) foram positivos. A soropositividade foi associada significativamente à anemia e trombocitopenia, isoladamente ou em combinação, e à leucopenia. Por outro lado, não houve diferenças quanto à soroprevalência segundo sexo, raça e idade. Este estudo demonstrou que cães encaminhados a um hospital veterinário-escola na região Centro-oeste do Brasil são altamente expostos à E. canis, e que cães soropositivos têm maior probabilidade de apresentar alterações hematológicas, principalmente anemia, trombocitopenia e leucopenia. Para o nosso conhecimento, este é o primeiro estudo sobre a detecção de anticorpos anti-E. canis por meio da RIFI em cães do estado de Goiás. Essas descobertas destacam a necessidade de aumentar a conscientização entre os proprietários de cães em relação às medidas de controle do carrapato no Centro-oeste do Brasil, em última análise, para reduzir o risco de exposição ao E. canis e outros patógenos transmitidos por carrapatos.

Glomerulotubular pathology in dogs with subclinical ehrlichiosis.

Subclinical stage of ehrlichiosis is characterized by absence of clinical or laboratory alterations; however, it could lead to silent glomerular/tubular changes and contribute significantly to renal failure in humans and animals. The aim of this study was to evaluate glomerular and tubular alterations in dogs with subclinical ehrlichiosis. We evaluated renal biopsies of 14 bitches with subclinical ehrlichiosis and 11 control dogs. Samples were obtained from the left kidney, and the tissue obtained was divided for light microscopy, immunofluorescence, and transmission electron microscopy. Abnormalities were identified by light microscopy in 92.9% of dogs with ehrlichiosis, but not in any of the dogs of the control group. Mesangial cell proliferation and synechiae (46.1%) were the most common findings, but focal segmental glomerulosclerosis and ischemic glomeruli (38.4%), focal glomerular mesangial matrix expansion (30.7%), mild to moderate interstitial fibrosis and tubular atrophy (23%), and glomerular basement membrane spikes (23%) were also frequent in dogs with ehrlichiosis. All animals with ehrlichiosis exhibited positive immunofluorescence staining for immunoglobulins. Transmission electron microscopy from dogs with ehrlichiosis revealed slight changes such as sparse surface projections and basement membrane double contour. The subclinical phase of ehrlichiosis poses a higher risk of development of kidney damage due to the deposition of immune complexes.

Comparisons of hematologic results for juvenile versus adult shelter dogs presented for ovariohysterectomy or castration.

OBJECTIVE: To compare hematologic results for juvenile versus adult dogs from shelters that outwardly appeared healthy and were presented for ovariohysterectomy or castration. ANIMALS: 138 dogs from 13 regional shelters. PROCEDURES: Each dog underwent a physical examination (including use of a flea comb), age estimation by dental eruption characteristics, PCV, CBC, and tests for Dirofilaria immitis antigen and Anaplasma phagocytophilum, Borrelia burgdorferi, and Ehrlichia canis antibodies. Additional diagnostic tests were performed as needed. Dogs were grouped by age as < 3, ≥ 3 to ≤ 6, or > 6 months of age, with dogs ≤ 6 months of age considered juveniles and dogs > 6 months of age considered adults. Hematologic results were compared across groups. RESULTS: There were 138 dogs, of which 56 were juveniles (34 dogs < 3 months old; 22 dogs ≥ 3 to ≤ 6 months old) and 82 were adults. Juvenile (vs adult) dogs had lower mean calculated Hct and mean PCV whether dogs with infectious agents or parasites were included or excluded. The mean PCV and mean cell hemoglobin concentration were lower and the reticulocyte count higher for juvenile dogs < 3 months old (35.8%, 33.1 g/dL, and 135,000 reticulocytes/µL) versus adults (44.9%, 34.7 g/dL, and 68,500 reticulocytes/µL). Most (98.6%) dogs underwent surgery as scheduled; 2 dogs had surgery postponed because of thrombocytopenia or parvovirus infection. CONCLUSIONS AND CLINICAL RELEVANCE: Our findings indicated that outwardly healthy-appearing juvenile shelter dogs often have results for PCV and calculated Hct that are lower than those for adult shelter dogs and adult dog reference intervals but rarely require postponement of ovariohysterectomy or castration.

Determination of Serum Oxidative Stress, Antioxidant Capacity and Protein Profiles in Dogs Naturally Infected with Ehrlichia canis.

PURPOSE: Canine ehrlichiosis is an important tick-borne disease of dogs worldwide. In the present study, we aimed to determine the serum total antioxidant capacity (TAC), malondialdehyde (MDA), nitric oxide (NO), superoxide dismutase, (SOD), glutathione peroxidase (GSH-Px), adenosine deaminase (ADA) activity and serum protein profiles in dogs affected with naturally acquired ehrlichiosis. METHODS: The animal materials had been consisted of ten dogs naturally infected with Ehrlichia canis, and ten controls negative for Ehrlichia canis. TAC, MDA, NO, SOD, GSH-Px, ADA activity and TP, ALB, GLOB levels were measured in sera of the animals. The serum protein concentrations were measured by autoanalyzer. The electrophoretic profiles of serum total protein were determined by native polyacrylamide gel electrophoresis (Native-PAGE). RESULTS: In dogs with ehrlichiosis, decreased TAC (P < 0.05) and GSH-Px (P > 0.05) levels were determined. However, NO (P > 0.05), SOD (P < 0.05), ADA (P > 0.05), MDA (P > 0.05), TP (P < 0.05) and GLO (P < 0.05) levels were found as increased in the Ehrlichia positive dogs. ALB levels were decreased without a statistical significance (P > 0.05). ALB, α1 and ß2 globulin strip densities were found as decreased in native-PAGE, while ß1 and γ globulin strip densities were significantly increased in the E. canis positive group when compared to the control. CONCLUSION: It was determined that the oxidative stress decreased high antioxidant activity in dogs naturally infected with E. canis, and consequently, pro-oxidant and antioxidant defense and serum protein profiles were affected. It was thought that antioxidant supplementation could be beneficial to the treatment of the disease.

Comparative Analysis of Genome of Ehrlichia sp. HF, a Model Bacterium to Study Fatal Human Ehrlichiosis.

BACKGROUND: The genus Ehrlichia consists of tick-borne obligatory intracellular bacteria that can cause deadly diseases of medical and agricultural importance. Ehrlichia sp. HF, isolated from Ixodes ovatus ticks in Japan [also referred to as I. ovatus Ehrlichia (IOE) agent], causes acute fatal infection in laboratory mice that resembles acute fatal human monocytic ehrlichiosis caused by Ehrlichia chaffeensis. As there is no small laboratory animal model to study fatal human ehrlichiosis, Ehrlichia sp. HF provides a needed disease model. However, the inability to culture Ehrlichia sp. HF and the lack of genomic information have been a barrier to advance this animal model. In addition, Ehrlichia sp. HF has several designations in the literature as it lacks a taxonomically recognized name. RESULTS: We stably cultured Ehrlichia sp. HF in canine histiocytic leukemia DH82 cells from the HF strain-infected mice, and determined its complete genome sequence. Ehrlichia sp. HF has a single double-stranded circular chromosome of 1,148,904 bp, which encodes 866 proteins with a similar metabolic potential as E. chaffeensis. Ehrlichia sp. HF encodes homologs of all virulence factors identified in E. chaffeensis, including 23 paralogs of P28/OMP-1 family outer membrane proteins, type IV secretion system apparatus and effector proteins, two-component systems, ankyrin-repeat proteins, and tandem repeat proteins. Ehrlichia sp. HF is a novel species in the genus Ehrlichia, as demonstrated through whole genome comparisons with six representative Ehrlichia species, subspecies, and strains, using average nucleotide identity, digital DNA-DNA hybridization, and core genome alignment sequence identity. CONCLUSIONS: The genome of Ehrlichia sp. HF encodes all known virulence factors found in E. chaffeensis, substantiating it as a model Ehrlichia species to study fatal human ehrlichiosis. Comparisons between Ehrlichia sp. HF and E. chaffeensis will enable identification of in vivo virulence factors that are related to host specificity, disease severity, and host inflammatory responses. We propose to name Ehrlichia sp. HF as Ehrlichia japonica sp. nov. (type strain HF), to denote the geographic region where this bacterium was initially isolated.

Dal-induced red blood cell incompatibilities in a Doberman Pinscher with von Willebrand factor deficiency and ehrlichiosis.

OBJECTIVE: To describe a complex case involving the management of a dog with von Willebrand disease (vWD), active ehrlichiosis infection, nonregenerative anemia, and blood type incompatibility related to the Dal antigen. CASE SUMMARY: A 13-week-oldintact male Doberman Pinscher weighing 7.2 kg was presented to the emergency service for a previous hemorrhaging event and progressive nonregenerative anemia. The dog had received a fresh whole blood transfusion 8 days prior to presentation due to severe anemia. Upon presentation, the puppy was tachycardic, and his mucous membranes were pale. A CBC revealed a nonregenerative anemia with a PCV of 0.11 L/L (11%). von Willebrand factor deficiency was suspected and later confirmed. The dog's blood type was dog erythrocyte antigen (DEA) 1 positive, but cross-matching to 4 RBC units, both DEA 1 positive and negative, failed to yield any compatible units. Antibody against a possible Dal RBC antigen was suspected, and 11 blood donors (Dalmatians and Dobermans) were cross-matched to find 2 compatible donors. After an uneventful fresh whole blood transfusion, a bone marrow biopsy revealed a hypocellular bone marrow and erythroid hypoplasia. A SNAP4DxPlus test and subsequent polymerase chain reaction (PCR) testing were positive for Ehrlichia ewingii and E. canis. Treatment with doxycycline was started, and the PCV was 0.17 L/L (17%) at discharge. At the 1-week follow-up, the PCV was 0.24 L/L (24%), and the puppy was doing well. NEW OR UNIQUE INFORMATION PROVIDED: This is a unique case of a dog presenting with several challenging disorders, including vWD resulting in hemorrhage, ehrlichiosis potentially contributing to a nonregenerative anemia, and a blood type incompatibility due to the Dal antigen. Doberman Pinschers have a high prevalence of vWD- and Dal-negative phenotype, which emphasizes the value of cross-matching and the recognition of antigen prevalence in specific breeds.

Comparison of Anaplasma and Ehrlichia species-specific peptide ELISAs with whole organism-based immunofluorescent assays for serologic diagnosis of anaplasmosis and ehrlichiosis in dogs.

OBJECTIVE: To compare the performance of 5 synthetic peptide-based ELISAs with that of 3 commercially available immunofluorescent assays (IFAs) for serologic diagnosis of anaplasmosis and ehrlichiosis in dogs. SAMPLE: A convenience set of 109 serum samples obtained before and at various times after inoculation for 23 dogs that were experimentally infected with Anaplasma phagocytophilum, Anaplasma platys, Ehrlichia canis, Ehrlichia chaffeensis, or Ehrlichia ewingii and 1 uninfected control dog in previous studies. PROCEDURES: All serum samples were assessed with 5 synthetic peptide-based ELISAs designed to detect antibodies against A phagocytophilum, A platys, E canis, E chaffeensis, and E ewingii and 3 whole organism-based IFAs designed to detect antibodies against A phagocytophilum, E canis, and E chaffeensis. The species-specific seroreactivity, cross-reactivity with the other tick-borne pathogens (TBPs), and diagnostic sensitivity and specificity were calculated for each assay and compared among assays. RESULTS: All serum samples obtained from dogs experimentally infected with a TBP yielded positive results on a serologic assay specific for that pathogen. In general, sensitivity was comparable between ELISAs and IFAs and tended to increase with duration after inoculation. Compared with the IFAs, the corresponding ELISAs were highly specific and rarely cross-reacted with antibodies against other TBPs. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that peptide-based ELISAs had enhanced specificity relative to whole organism-based IFAs for detection of antibodies against Anaplasma and Ehrlichia spp, which should facilitate accurate diagnosis and may help detect dogs coinfected with multiple TBPs.

Perfil clínico e laboratorial de cadelas sororeativas para erliquiose tratadas em um Hospital Veterinário Universitário em Niterói, Estado do Rio de Janeiro, Brasil / Clinical and laboratory profile of dogs seroreactive to ehrlichiosis treated at the Veterinary Medical Teaching Hospital in Niterói, State of Rio de Janeiro, Brazil

Ehrlichiosis is a tick-borne disease highly prevalent in Brazil, and is relevant in canine clinical practice due to its high morbidity and mortality. Its clinical signs are nonspecific and its phases are acute, lasting 2 to 4 weeks; subclinical, i.e., asymptomatic; and chronic, resembling an autoimmune disease. The purpose of this study was to identify the occurrence of reactivity to Ehrlichia canis of bitches treated at the Veterinary Medical Teaching Hospital of the Universidade Federal Fluminense (UFF) - Niterói, RJ, Brazil, based on serological examination by iELISA, and to compare the hematological, biochemical, urinary protein-creatinine and urinary density profiles of reactive and non-reactive animals. This study involved solely bitches, regardless of breed, starting at 1 year of age. One hundred and thirty bitches, 1 to 16 year-old (mean age 7.02 ± 4.00), weighing 1.5 to 50 kg (mean weight 12.12 ± 10.65) were subjected to clinical examination and abdominal ultrasound. Complete blood count, biochemical measurements, urinalysis and serology for E. canis were also performed. The serum was used in the iELISA to identify immunoglobulin G (IgG), using a canine Ehrlichia Imunotest® diagnostic kit (Imunodot®, Jaboticabal, SP, Brazil) according to the manufacturer's instructions. Sixty animals (46.20%) were reactive to E. canis. According to their owners, only 5 (8.3%) of the 60 seroreactive animals had a history of tick-borne disease. The most common profile was that of mixed breed animals living with their owners, older than 7 years, who had not been treated preventatively with specific drugs against ectoparasites. Laboratory tests showed significant differences between groups in terms of total protein (TP), and calcium and urinary protein-creatinine ratio (UPC). TP and UPC were elevated in the non-reactive group, while the only significant change in the reactive group was mild hypocalcemia. In this study, 30% (18/60) of the bitches were seroreactive to E. canis and had hypocalcemia. Of these, 50% (9/18) had a UPC above 0.5. Furthermore, 66.7% (12/18) of this group with hypocalcemia also showed urine density (UD) of less than 1024. Among these 18 bitches, 5 had both alterations, i.e., UPC > 0.5 and UD < 1024. In this study, a high prevalence of bitches seroreactive to Ehrlichia canis was observed, despite the absence of clinical and/or laboratory signs indicative of the disease. In the investigation of IgG class antibodies, it is not possible to determine the exact time of infection, and titers may remain high for a period of more than 11 months, even after treatment and elimination of the bacterium. The fact that most seroreactive bitches showed no symptoms compatible with the disease either before or during the study suggests that they were in the subclinical phase of ehrlichiosis. The main reason for calcium metabolism disorders is a phosphorus imbalance, a condition that occurs in kidney diseases. Isosthenuria reflects the kidney's inability to concentrate urine. This finding may be one of the first clinical manifestations of chronic kidney disease (CKD), especially in dogs. On the other hand, the UPC ratio may increase with the progression of CKD. The presence of hypocalcemia, isosthenuria and increased UPC associated with seroreactivity suggests that infection by E. canis may be associated with the onset of CKD. Veterinarians should keep in mind the complexity of the pathophysiology of ehrlichiosis to ensure the disease is not underdiagnosed in any of its phases, thereby ensuring the correct treatment is provided. Such awareness is expected to reduce the chronicity of the disease and underlying sequelae among dogs.(AU)

Canonical correlative analyses among an enzyme-linked immunosorbent assay using synthetic peptides, an indirect fluorescent antibody test, and hematologic measurements in dogs infected with Ehrlichia canis.

BACKGROUND: Immunoreactive tandem repeat proteins (TRPs) in amino acid sequences were identified and employed in the serologic diagnosis of canine monocytic ehrlichiosis (CME). OBJECTIVES: This study evaluated using TRP19 and TRP36 synthetic protein antigens with enzyme-linked immunosorbent assays (ELISAs) and compared the results with an indirect fluorescent antibody test (IFAT) to diagnose CME in the serum of dogs with suspected CME. METHODS: The sera of 243 dogs that exhibited clinical and hematologic signs suggestive of CME had IFATs performed. An ELISA with synthetic TRP19 and TRP36 antigens from two E canis genotypes (USTRP36 and BrTRP36) that circulate in Brazil were also performed. Canonical correlations and chi-square tests were evaluated on the results of these tests. RESULTS: Among the 243 dogs, 179 (73.6%) were reactive at an IFAT ≥ 40, 167 (68.72%) reacted at the cut-off ≥160, and 149 (61.31%) reacted at ≥640. The ELISA revealed that 172 (70.8%) dogs reacted to TRP19 peptide, 163 (67.1%) reacted to USTRP36, and 114 (46.9%) reacted to BrTRP36. The canonical correlation analyses showed that the TRP19 ELISA was the most effective diagnostic method. Serum total protein levels showed strong positive correlations with the USTRP36 peptide. Qualitative analyses revealed an association between the TRP19 peptide and the presence of anemia, thrombocytopenia, and hyperproteinemia. The IFAT results at titers of ≥160 and ≥640 were associated with thrombocytopenia. CONCLUSIONS: The results of this study indicated that the TRP19 peptide was an excellent antigen and that the IFAT was diagnostically important in confirming the diagnosis of CME in serum samples.

Demonstrating the presence of Ehrlichia canis DNA from different tissues of dogs with suspected subclinical ehrlichiosis.

BACKGROUND: Nowadays, Ehrlichia canis receives increasing attention because of its great morbidity and mortality in animals. Dogs in the subclinical and chronic phases can be asymptomatic, and serological tests show cross-reactivity and fail to differentiate between current and past infections. Moreover, there could be low parasitaemia, and E. canis might be found only in target organs, hence causing results to be negative by polymerase chain reaction (PCR) on blood samples. METHODS: We evaluated by PCR the prevalence of E. canis in blood, liver, spleen, lymph node and bone marrow samples of 59 recently euthanised dogs that had ticks but were clinically healthy. RESULTS: In total, 52.55% of the blood PCRs for E. canis were negative, yet 61.30% yielded positive results from tissue biopsies and were as follows: 63.15% from bone marrow; 52.63% from liver; 47.36% from spleen; and 15.78% from lymph node. In addition, 33% had infection in three tissues (spleen, liver and bone marrow). CONCLUSIONS: Our results show the prevalence of E. canis from tissues of dogs that were negative by blood PCR. Ehrlichia canis DNA in tissue was 30% lower in dogs that tested negative in PCR of blood samples compared to those that were positive. However, it must be taken into account that some dogs with negative results were positive for E. canis in other tissues.

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.

Differential diagnoses in 83 dogs with icterus / Diagnósticos diferenciais em 83 cães com icterícia

Icterus (jaundice) is a yellowish pigmentation resulting from the depositing of bilirubin in tissues due to its high plasmatic concentration. The pathogenesis of icterus includes metabolic changes or obstructed bilirubin excretion and it is classified as pre-hepatic, hepatic and post-hepatic. This study aimed to evaluate and classify different causes of icterus in dogs during post mortem examination. These dogs were examined from 2014 to 2017, using macroscopic and histologic exams as well as ancillary tests. Eighty-three dogs were examined macroscopically and microscopically. They were separated into groups of icterus types: 24 (28.9%) dogs had pre-hepatic icterus, 45 (54.2%) had hepatic, 13 (15.7%) pre-hepatic and hepatic and one (1.2%) had post-hepatic icterus. Many factors were identified as a cause of icterus, including infectious agents (51/83), neoplasms (13/83), hepatic degeneration (11/83), chronic hepatic diseases (6/83), and obstructive causes (1/87). Among the infectious causes, leptospirosis, ehrlichiosis and disorders suggestive of septicemia were diagnosed. Neoplasms associated with icterus were cholangiocarcinoma, hemangiosarcoma and lymphoma. Other causes of icterus included degenerative diseases, such as lipidosis and glycogen degeneration. Hepatic fibrosis (cirrhosis) as a chronic disease and cholelithiasis also produced icterus. PCR was performed to confirm leptospirosis and ehrlichiosis. Samples of total DNA were used to amplify a fragment of a gene from Leptospira interrogans and Ehrlichia canis. In some dogs, co-infection of these agents was detected. The classification and identification of icterus etiologies in dogs is very important due to the number of diseases with this alteration, where ante mortem diagnosis is not always easily performed when some of these conditions are present.(AU)

Icterícia é a pigmentação amarelada decorrente da deposição de bilirrubina em tecidos devido à elevada concentração plasmática. A patogênese da icterícia inclui alterações no metabolismo ou na excreção de bilirrubina, sendo classificada em pré-hepática, hepática ou pós-hepática. O objetivo desse estudo foi identificar, avaliar e classificar as causas de icterícia em cães necropsiados de 2014 a 2017, associando as lesões macroscópicas, histológicas e exames complementares. Foram avaliados macro- e microscopicamente 83 cães com diferentes intensidades de icterícia. Os cães foram separados em grupos de acordo com o tipo de icterícia: 24 (28,9%) cães com icterícia pré-hepática, 45 (54,2%) cães com icterícia hepática, 13 (15,7%) com icterícia pré-hepática e hepática e um (1,2%) com icterícia pós-hepática. Foram identificadas várias etiologias associadas à icterícia, dentre elas pode-se destacar, agentes infecciosos (51/83), neoplasmas (13/83), processos degenerativos (11/83), crônicos (6/83) e obstrutivos (1/83). Dentre as causas infecciosas, destacam-se a leptospirose, a erliquiose e as lesões sugestivas de septicemia. Entre os neoplasmas associados com icterícia destacaram-se o colangiocarcinoma, hemangiossarcoma e linfoma. Outras causas de icterícia incluiriam os processos degenerativos como as degenerações gordurosa e glicogênica. Fibrose hepática (cirrose) e colelitíase foram também diagnosticados como causa de icterícia. A PCR foi utilizada para o diagnóstico confirmatório de leptospirose e erliquiose. Amostras de DNA total foram utilizadas para amplificar um fragmento dos genes de Leptospira interrogans e de Ehrlichia canis. Em alguns cães foi detectada co-infecção por estes agentes. A classificação e a identificação das causas de icterícia em cães são relevantes devido ao grande número de doenças que apresentam essa alteração, muitas vezes sem diagnóstico ante mortem.(AU)

Anaplasma phagocytophilum infection in cats: A literature review to raise clinical awareness.

PRACTICAL RELEVANCE: Granulocytic anaplasmosis is a disease in humans and animals caused by the Gram-negative bacterium Anaplasma phagocytophilum within the family Anaplasmataceae. The pathogen is transmitted by ticks of the Ixodes species. Infections with A phagocytophilum have often been described in dogs but reports on natural infections in cats are rare. An infection with A phagocytophilum should be considered as a differential diagnosis in cats if the history reveals tick infestation and/or outdoor access in combination with the relevant clinical signs. GLOBAL IMPORTANCE: A phagocytophilum is also important in human medicine because of its zoonotic potential. Due to the risk of vector-borne infections for both feline and public health, cats should be protected with ectoparasiticides, especially in endemic areas. AIM: The aim of this review is to give an overview of the published data and summarise the epidemiology, pathogenesis, diagnosis, clinical signs and therapy of feline granulocytic anaplasmosis. As clinical signs are vague and non-specific, this review aims to raise awareness of A phagocytophilum infection, both among clinicians, so that they consider testing potentially exposed cats, and scientists, in order to prompt further research. EVIDENCE BASE: Sixteen publications describing 55 cats have been reviewed. Thirty-four cats were well diagnosed based on guidelines of the European Advisory Board on Cat Diseases and blood analyses were performed to varying extents for these cats. Because of the limited number of studies and a lack of knowledge in cats, clinical signs and blood analyses are compared with available data in dogs.

Tick-borne pathogens in dogs, wild small mammals and their ectoparasites in the semi-arid Caatinga biome, northeastern Brazil.

Caatinga is a biome exclusive to the semiarid zone of Brazil, where studies on ticks and tick-borne diseases are scarce. Herein, we investigated the occurrence of Rickettsia, Ehrlichia, and Coxiella in wild mammals, domestic dogs and their ectoparasites using molecular and serological techniques. During 2014-2016, blood samples and ectoparasites were collected from 70 small mammals (51 rodents, 18 marsupials, 1 wild canid) and 147 domestic dogs in three areas of the Caatinga. Through serological analyses of domestic dogs of the three areas, 8 to 11 % were seropositive for Rickettsia rickettsii, 9 to 37 % for Rickettsia amblyommatis, 61 to 75 % for Ehrlichia canis, and 0-5% for Coxiella burnetii. All wild mammals were seronegative for Rickettsia spp. and C. burnetii, except for one rodent (Wiedomys pyrrhorhinos) and one marsupial (Didelphis albiventris) that were seroreactive to C. burnetii, one wild canid (Cerdocyon thous) for R. amblyommatis, and two Rattus rattus for Rickettsia spp. Through PCR targeting DNA of Rickettsia, Ehrlichia or Coxiella, all blood samples were negative, except for the presence of Ehrlichia canis DNA in 8.8 % of the domestic dogs, and a recently reported novel agent, Ehrlichia sp. strain Natal, in one marsupial (Gracilinanus agilis). A total of 222 ticks, 84 fleas, and six lice were collected. Ticks were mostly Rhipicephalus sanguineus sensu lato, some Ixodes loricatus, Ornithodoros rietcorreai, Haemaphysalis sp., and Amblyomma spp.; fleas were Ctenocephalides felis felis, Pulex sp. and Polygenis (Polygenis) bohlsi jordani; and lice were Polyplax sp. and Gyropus sp. Through molecular detection of microorganisms, 9% of C. felis felis contained Rickettsia felis, 20 % of A. auricularium contained R. amblyommatis and 13 % of A. parvum contained 'Candidatus Rickettsia andeanae', whereas Ehrlichia canis DNA was detected in at least 6% of the R. sanguineus s.l. from one area. We report a variety of ectoparasites infesting small mammals and domestic dogs in the Caatinga biome, where these ectoparasites probably act as vectors of rickettsiae, ehrlichial agents (E. canis and Ehrlichia sp. strain Natal) and C. burnetii. Our results highlight to the potential risks of human infection by these tick-borne agents in the Caatinga biome.

Seroprevalence of viral and vector-borne bacterial pathogens in domestic dogs (Canis familiaris) in northern Botswana.

BACKGROUND: Domestic dogs (Canis familiaris) have the potential to act as disease reservoirs for wildlife and are important sentinels for common circulating pathogens. Therefore, the infectious disease seroprevalence among domestic dogs in northern Botswana may be indicative of pathogen exposure of various wildlife species. The objective of this study was to assess the seroprevalence of Ehrlichia spp., Borrelia burgdorferi, Anaplasma spp., Dirofilaria immitis, canine adenovirus, canine parvovirus, and canine distemper virus in domestic dogs as proxies of disease prevalence in the local wildlife in the Okavango Delta region of Botswana. Statistical analysis assessed crude and factor-specific seroprevalence proportions in relation to age, sex, and geographical location as predictors of seropositivity. Logistic regression was used to identify adjusted predictors of seropositivity for each of the pathogens of interest. RESULTS: Samples from 233 dogs in a total of seven locations in Maun, Botswana, and surrounding villages were collected and serologically analyzed. No dogs were seropositive for B. burgdorferi, while low seroprevalence proportions were observed for Anaplasma spp. (2.2%) and D. immitis (0.9%). Higher seroprevalence proportions were observed for the tick-borne pathogen Ehrlichia spp. (21.0%), and 19.7% were seropositive for canine adenovirus (hepatitis). The highest seroprevalence proportions were for canine parvovirus (70.0%) and canine distemper virus (44.8%). The predictors of seropositivity revealed that adults were more likely to be seropositive for canine adenovirus, canine distemper virus, and canine parvovirus than juveniles, and location was a risk factor for canine adenovirus, canine distemper virus, canine parvovirus, and Ehrlichia spp. CONCLUSIONS: Results indicate that increasing tick control and vaccination campaigns for domestic dogs may improve the health of domestic animals, and potentially wildlife and humans in the Okavango Delta since viral and vector-borne bacterial pathogens can be transmitted between them.