Difference in cytological findings of healthy and conjunctivitis/keratoconjunctivitis affected canine eyes between variably experienced evaluators.

Background and Aim: Cytology investigations are a frequent part of ophthalmological examination. We aimed to assess whether the cytological findings of healthy and conjunctivitis/keratoconjunctivitis samples differed based on the evaluator's experience. Materials and Methods: A study evaluated healthy eyes (n = 40) and eyes affected with keratoconjunctivitis and/or conjunctivitis (n = 28) in dogs. An ophthalmological examination was performed before sampling the eyes using a sterile cotton swab. An evaluator with theoretical experience and one with undergone clinical pathology residency training performed cytology blinded to the clinical findings. Results: In the healthy eyes group, the agreement between the evaluators for cellularity was nonexistent, while that for cell preservation and mucus content was fair. In the affected eyes group, the agreement for cellularity and mucus content was moderate, while that for cell preservation was fair. The inadequate sample rate differed significantly between the two evaluators in the healthy eyes group (p = 0.006) but not in the affected eyes group (p = 0.083). Bacterial presence was detected by both evaluators, and the findings did not differ statistically from the bacteriology results (p = 0.05). Significant variations were noted in the differential cell count; the mean count of the superficial epithelial cells and goblet cells of the healthy eyes group (p < 0.05) and that of the basal/intermediate cells and neutrophils of the affected eyes (p < 0.05) showed significant differences. Conclusion: The evaluator's experience significantly affected the differential cell count in both the healthy and affected eyes groups. Neutrophil degeneration was not observed by the less experienced evaluator, whereas bacteria were detected equally well by both the evaluators.

Haemotropic Mycoplasma species in pet cats in Latvia: a study, phylogenetic analysis and clinical case report.

OBJECTIVES: The aim of this study was to evaluate whether haemotropic Mycoplasma species are detected in pet cats in Latvia, to perform a phylogenetic analysis of the detected pathogens and to report a clinical case of feline infectious anaemia. METHODS: Peripheral blood samples (n = 125) from pet cats were submitted; 99 samples were adequate to test for the presence of Mycoplasma species DNA by nested PCR. A clinical case was added in the later stages of the study. Positive isolates were subjected to phylogenetic analysis. RESULTS: The prevalence of 'Candidatus Mycoplasma haemominutum' was 15% (n = 15/99), that of Mycoplasma haemofelis was 5% (5/99) and that of 'Candidatus Mycoplasma turicensis' was 2% (n = 2/99). Cases of coinfection included 'Candidatus M haemominutum' + M haemofelis (4%; n = 4/99) and 'Candidatus M haemominutum' + 'Candidatus M turicensis' (1%; n = 1/99). This is the first published report of M haemofelis infection in the Baltic states. Two different 'Candidatus M turicensis' isolates were discovered after phylogenetic analysis. CONCLUSIONS AND RELEVANCE: This report is the first of an autochthonous feline infectious anaemia case in the Baltic region. The prevalence of Mycoplasma species was similar to that in other northern European countries. Phylogenetic analysis revealed variability of the isolates; one of the 'Candidatus M turicensis' genotypes was detected for the first time in Europe.

Tick-borne pathogens in ticks collected from dogs, Latvia, 2011-2016.

BACKGROUND: Different tick species are able to transmit different pathogens, and tick-borne diseases are of substantial concern worldwide for both humans and animals. Environmental changes and changes in the range of tick species, including Dermacentor reticulatus in Europe, can affect the spread of zoonotic pathogens. The aim of this study was to investigate the prevalence of the tick-borne pathogens in ticks removed from dogs in Latvia, and to explore possible changes between years 2011 and 2016. RESULTS: In 2011, only Ixodes ticks (221 Ixodes ricinus and 22 Ixodes persulcatus) were collected from dogs, while in 2016 tick samples belonged to Ixodes ricinus (360), Ixodes persulcatus (2) and Dermacentor reticulatus (27) species. In total, 35.8 and 40.0% of adult ticks were pathogen-positive in 2011 and 2016, respectively; the difference was not statistically significant (P > 0.05). The molecular analysis indicated the presence of 13 tick-borne microorganisms; the most prevalent pathogen was Rickettsia, followed by Borrelia burgdorferi sensu lato group spirochetes, Anaplasma phagocytophilum and Babesia species. Borrelia miyamotoi was also present. A co-infection with two and three tick-borne pathogens was detected in 7.9 and 7.4% of Ixodes ricinus and Dermacentor reticulatus, respectively. The results of this study confirmed that the spread of novel vectors could bring additional risk of exposure to novel emerging pathogens to pets and their owners, as both Babesia canis and Rickettsia raoultii were shown to be highly associated with Dermacentor reticulatus ticks in Latvia. CONCLUSIONS: This study demonstrates the potential danger from the inadvertent introduction of novel disease pathogens and vectors. Awareness of co-infections and Dermacentor reticulatus-related pathogens needs to be increased.

Fatal Babesia canis canis infection in a splenectomized Estonian dog.

A previously splenectomized dog from Estonia was presented with a sudden lack of appetite and discoloration of the urine. Despite supportive therapy, its condition deteriorated dramatically during 1 day. Severe thrombocytopenia and high numbers of protozoan hemoparasites were evident in blood smears, and the hematocrit dropped from 46 to 33 %. The dog was euthanized before specific antibabesial treatment was initiated. Blood samples from the dog and from two other dogs in the same household tested positive for Babesia using molecular methods, and the sequences of partial 18S rRNA gene confirmed the causative species as Babesia canis canis. The risk of severe, rapidly progressing babesiosis in splenectomized dogs merits awareness.

Prevalence and phylogenetic analysis of Babesia spp. in Ixodes ricinus and Ixodes persulcatus ticks in Latvia.

Babesia spp. are tick-borne protozoan parasites that have been reported in many European countries and are considered to be emerging pathogens. Several Babesia spp. have been identified in ticks in Latvia. Recently, canine babesiosis cases were diagnosed for the first time in Latvia; therefore, continued studies on the prevalence and occurrence of new species are warranted. In the present study, questing tick samples collected in 2005-2007 were screened for the presence of Babesia spp.; in total, 432 Ixodes ricinus and 693 Ixodes persulcatus ticks were analyzed. Babesia spp. were detected in 1.4% of the I. ricinus ticks and in 1.9% of I. persulcatus ticks. Sequencing revealed that ixodid ticks in Latvia contained Babesia microti, Babesia capreoli, and Babesia venatorum. Babesia microti was the most prevalent species, accounting for 58% of all positive samples; moreover, two distinct B. microti genotypes were identified. Phylogenetic analysis of the full-length 18S rRNA gene of two B. capreoli/B. divergens isolates indicated a closer relationship to the B. capreoli clade than B. divergens. This is the first report of B. venatorum in I. persulcatus ticks in Latvia. Our results suggest that both I. ricinus and I. persulcatus ticks play important roles in the epidemiology of these zoonotic pathogens in Latvia.

Anaplasma phagocytophilum DNA amplified from lesional skin of seropositive dogs.

Canine granulocytic anaplasmosis (CGA) is caused by the rickettsial microorganism Anaplasma phagocytophilum. CGA is typically characterized by fever, thrombocytopenia, lethargy, anorexia, arthropy, and other nonspecific clinical signs. Skin lesions have been described in naturally infected lambs and humans. The pathophysiology of CGA is not entirely clear, and the persistence of the organism after the resolution of clinical signs has been described. The aim of the study was to investigate if A. phagocytophilum can be detected in canine lesional skin biopsies from A. phagocytophilum-seropositive dogs with etiologically unclear skin lesions that improved after the treatment with doxycycline. Paraffin-embedded lesional skin biopsies were allocated into separate groups: biopsies from A. phagocytophilum-seropositive dogs responsive to treatment with doxycycline (n=12), biopsies from A. phagocytophilum-seronegative dogs (n=2), and biopsies in which skin lesions histopathologically resembled a tick bite (n=10). The serological status of the latter group was unknown. Histology of the seropositive and seronegative dog skin lesions did not indicate an etiology. DNA was extracted, and a conventional PCR for partial 16S rRNA gene was performed. Anaplasma phagocytophilum DNA was amplified from 4/12 seropositive dogs' skin biopsies. All sequences were 100% identical to the prototype A. phagocytophilum human strain (GenBank accession number U02521). Anaplasma phagocytophilum was not amplified from the 2 seronegative and 10 suspected tick bite dogs. Serum antibody titers of the PCR-positive dogs ranged from 1:200 to 1:2048. Histopathologically, a mild-to-moderate perivascular to interstitial dermatitis composed of a mixed cellular infiltrate and mild-to-moderate edema was seen in all seropositive dogs. In 8/12 seropositive dogs, vascular changes as vasculopathy, fibrinoid necrosis of the vessel walls, and leukocytoclastic changes were observed. In summary, our results support the hypothesis that the persistence of A. phagocytophilum in the skin may be causative for otherwise unexplained skin lesions in seropositive dogs.

Seroprevalence against Borrelia burgdorferi sensu lato and occurence of antibody co-expression with Anaplasma phagocytophilum in dogs in Latvia.

BACKGROUND: Lyme disease is commonly diagnosed in humans in Latvia, but up to date no studies have been performed to investigate its prevalence in dogs. The aim of this study was to evaluate if seroprevalence against B. burgdorferi sensu lato (B. burgdorferi s.l.) and co-expression of antibodies against B.burgdorferi s.l. and A. phagocytophilum is higher in dogs with clinical suspicion of tick-borne diseases compared to healthy dogs. FINDINGS: Venous blood was taken from healthy dogs (n=441) and dogs suspected to have borreliosis and/ or canine granulocytic anaplasmosis (n=29). The presence of antibodies was detected with SNAP 4Dx test (IDEXX, Westbrook, Maine, USA). The seroprevalence against B. burgdorferi s.l. in healthy dogs was 2.49% (11/441) and 36% (4/11) of seropositive dogs had antibodies against both of investigated bacteria. None of the dogs in sick dog group had detectable antibodies against B. burgdorferi s.l. CONCLUSIONS: We conclude that seroprevalence to B. burgdorferi s.l. in dogs in Latvia is low and that dogs with suspicion of tick-borne disease do not have higher B. burgdorferi s.l. seroprevalence than healthy dogs. Dogs that express antibodies against B. burgdorferi s.l. frequently co-express antibodies against A. phagocytophilum.

Autochthonous canine babesiosis caused by Babesia canis canis in Latvia.

This is the first report of confirmed canine babesiosis in Latvia supporting the observed geographical expansion of this disease. Between 2009 and 2011 three dogs which have not traveled outside of Latvia were diagnosed with babesiosis. Hematological analysis and serological tests for granulocytic anaplasmosis, ehrlichiosis and borreliosis were negative (Idexx SNAP 4Dx test). Peripheral blood erythrocytes of the three dogs contained large Babesia that were identified as Babesia canis canis by PCR. Sequences of partial 18S rRNA gene were 98-100% similar to the sequences of B. canis canis isolated from dogs in other European countries. We conclude that these are the first autochthonous canine babesiosis cases reported from Latvia.

Rapid, effective and user-friendly immunophenotyping of canine lymphoma using a personal flow cytometer.

BACKGROUND: Widespread use of flow cytometry for immunophenotyping in clinical veterinary medicine is limited by cost and requirement for considerable laboratory space, staff time, and expertise. The Guava EasyCyte Plus (Guava Technologies, Hayward, CA, US) is the first, personal, bench-top flow cytometer designed to address these limitations. OBJECTIVE: The aim of this study was to adapt the immunohistochemical protocol used for immunophenotyping of canine lymphoma to the personal flow cytometer for rapid, effective and user-friendly application to the diagnosis and prognosis of canine lymphoma and to demonstrate its practicality for widespread veterinary application. Performance of the personal flow cytometer for immunophenotyping T and B lymphocytes in blood and lymph nodes from normal dogs and dogs with lymphoproliferative disease, was assessed using only two monoclonal antibodies (against CD3 and CD21), and by comparison with analysis using two conventional flow cytometers. METHODS: 26 dogs with lymphoproliferative disease (23 with lymphoma, 3 with lymphocytic leukaemia) were studied along with 15 controls (2 non-lymphoma lymph nodes and 13 non-leukemic bloods. Lymphocytes were immunostained with fluorescent-labeled, monoclonal antibodies against CD3 and CD21. To assess the effectiveness of the personal flow cytometer in discrimination between T and B cell immunophenotypes, T and B cell counts for half the samples (14 blood and 11 lymph node) were also determined using the same method and conventional flow cytometers (FACSCalibur, Cyan Dako). To assess the effectiveness of the personal flow cytometer in discriminating between leukocyte types, lymphocyte differential counts were determined for 21 blood samples and compared with those from automated hematology analyzers (CELL-DYN 3500, n=11 and ADVIA 2120, n=10). Quality and sub-cellular distribution of immunostaining was assessed using fluorescence microscopy. RESULTS: The protocol for immunophenotyping took 2 to 3 hours to complete from the point of receipt of sample to reporting of immunophenotype. The personal flow cytometer differential lymphocyte counts correlated highly (n=20; r=0.97, p<0.0001) with those of automated haematology analyzers. The personal flow cytometer counts consistently, but mildly, underestimated the percentages of lymphocytes in the samples (mean bias of -5.3%.). The personal flow cytometer immunophenotype counts were indistinguishable from those of conventional flow cytometers for both peripheral blood samples (n=13; r=0.95; p<0.0001; bias of -1.1%) and lymph node aspirates (n=11,r=0.98; p<0.001; bias of 1%). All but one leukemic and one lymphomatous lymph node sample, out of 26 samples of dogs with lymphoproliferative disease analyzed, could be immunophenotyped as either B or T cells. CONCLUSIONS: We conclude that use of only 2 monoclonal antibodies is sufficient for immunophenotyping most cases of canine lymphoma by flow cytometry and enables rapid immunophenotyping. The personal flow cytometer may be as effectively used for immunophenotyping canine lymphoma as conventional flow cytometers. However, the personal flow cytometer is more accessible and user-friendly, and requires lower sample volumes.

Association between Anaplasma phagocytophilum seroprevalence in dogs and distribution of Ixodes ricinus and Ixodes persulcatus ticks in Latvia.

Anaplasma phagocytophilum has been detected in ticks in Latvia; however, this is the first study to investigate this pathogen in dogs in Latvia. The aims of this study were: (i) to determine A. phagocytophilum seroprevalence in dogs, (ii) to correlate A. phagocytophilum seroprevalence in dogs with the geographic distribution of the tick species Ixodes ricinus and Ixodes persulcatus, and (iii) to determine if seroprevalence for A. phagocytophilum is higher in dogs with clinical signs suggestive of canine granulocytic anaplasmosis (CGA). Peripheral venous blood samples were collected from 3 dog groups: (i) clinically healthy dogs (HD, n=400), (ii) clinically healthy hunting dogs (HHD, n=41), and (iii) dogs with a clinical suspicion of anaplasmosis (SD, n=29). Sampling was carried out in regions inhabited by I. ricinus (IR), I. persulcatus (IP), and in regions where both tick species were present (M). SNAP 4Dx test (IDEXX) was used to detect antibodies against A. phagocytophilum in the blood of all dogs; nested PCR was performed in selected dogs of the SD group. Seroprevalence for A. phagocytophilum was calculated and correlated with the prevalent tick species in the region. A. phagocytophilum seroprevalence was 11.0% in HD, 12% in HHD, and 17% in SD with no significant differences among groups. In the IR region, seroprevalence was 12.5% (34/272) while seroprevalence in the M region was 17% (13/76), and both were significantly higher than the seroprevalence of 2% in the IP region (2/93; p<0.0005). One CGA case was diagnosed. We conclude that A. phagocytophilum seroprevalence in Latvia is within the range reported from other European countries. CGA should be included in the differential list in Latvian dogs with appropriate clinical signs and laboratory abnormalities, especially in I. ricinus habitat areas.

Subconjunctival mycetoma caused by Scedosporium apiospermum infection in a horse.

An 11-year-old American Saddlebred gelding was presented for evaluation of a nonpainful subconjunctival mass involving the lateral canthus of the left eye. Other findings included a central corneal scar and a small central cataract of the lens in the left eye. Fine-needle aspiration of the mass was performed and cytologic examination revealed marked pyogranulomatous inflammation with intralesional fungal hyphae, consistent with mycetoma. The fungal structures were elongated and characterized by nonstaining walls; several bulbous yeast-like structures were also observed. The mycetoma was surgically removed and submitted for histopathologic examination and fungal culture. The histopathologic diagnosis was subconjunctival phaeohyphomycosis. Scedosporium apiospermum was identified based on macroscopic and microscopic features of the organism in culture. Scedosporium spp. have been reported as causes of mycetomatous and nonmycetomatous infections in both immunocompromised and immunocompetent people and animals. S. apiospermum and Pseudallescheria boydii, which is its teleomorphic counterpart, have been implicated as potentially emerging human and veterinary pathogens. Timely diagnosis is essential as the organism is often resistant to commonly used antifungal drugs. This report provides a detailed cytologic description of the organism and recent information on the taxonomy of this fungus and the diagnostic peculiarities of this particular infection.

Evaluation of serum cardiac troponin I concentration in dogs with renal failure.

OBJECTIVE: To determine whether dogs with renal failure have higher serum cardiac troponin I (cTnI) concentrations than healthy dogs. DESIGN: Case-control study. ANIMALS: 31 dogs with renal failure and 51 healthy dogs. PROCEDURES: Serum concentrations of creatinine and cardiac troponin I, urine specific gravity, and systolic arterial blood pressure were measured for all dogs. Dogs underwent a standardized physical examination, and any dog with evidence of cardiovascular disease or other nonrenal disease was excluded from final analyses. Dogs were considered to be in renal failure when the serum creatinine concentration was >or= 3.0 mg/dL, urine specific gravity was between 1.007 and 1.030, and renal failure had been clinically diagnosed. RESULTS: Dogs with renal failure had significantly higher serum cTnI concentrations (median, 0.35 ng/mL) than did healthy dogs (0.20 ng/mL). The renal failure group also had a significantly higher median systolic blood pressure (156 mm Hg) than did healthy dogs (138 mm Hg), although serum cTnI concentration was not correlated with systolic blood pressure in dogs with renal failure. There was no significant difference in age between dogs with renal failure and healthy dogs, but dogs with renal failure had significantly higher serum creatinine concentration and lower urine specific gravity. CONCLUSIONS AND CLINICAL RELEVANCE: Although dogs with renal failure did not have overt clinical signs of cardiac disease, they had high serum cTnI concentrations, which may have been associated with subclinical cardiovascular disease. The cause of the high serum cTnI concentration in these dogs requires additional investigation.

Correlation between cytologic and histopathologic diagnoses of bone lesions in dogs: a study of the diagnostic accuracy of bone cytology.

BACKGROUND: The diagnostic value of cytology compared with histopathology varies by tissue, but there is little information regarding this comparison involving canine bone. OBJECTIVES: The objective of this retrospective study was to compare primary pathologic processes for cytology and histopathology of canine bone lesions. We adopted a proposed standardized format for reporting studies of diagnostic accuracy. METHODS: A computer search of canine medical records at the University of Minnesota Veterinary Medical Center from September 2002 through October 2006 identified 52 bone cytology samples that had incisional (IncB) and/or excisional (ExcB) biopsy performed. The primary pathologic process was determined by evaluation of original reports. Cytologic vs IncB and cytologic vs ExcB were compared pairwise for agreement. Agreement was compared for neoplastic and non-neoplastic processes using the combined IncB/ExcB data, which included all ExcB (n=21) and IncB when that was the only biopsy available (n=31). Combined data were used to determine the effect of cytology cellularity on the diagnostic correlation. RESULTS: The correlation in primary process between cytology and IncB was 71%, and for ExcB was 71%. For lesions with a cytologic diagnosis of neoplasia compared with the combined IncB/ExcB data set, cytology and histopathology agreed in 92% of cases, which was significantly greater (P<.0001, chi2) than the 27% for non-neoplastic processes. Cytology cellularity significantly affected rates of correlation (P=.026), with high, moderate, and poor cellularity samples having concordant primary processes in 88%, 77%, and 47% of cases, respectively. CONCLUSIONS: Cytologic diagnosis of neoplasia for samples collected from canine bone correlates better with histopathology than cytologic diagnosis of non-neoplastic proliferative processes or inflammation. Cytologic diagnoses from highly cellular samples are more likely to correlate with histopathology than those from less cellular samples.