Evidence of Horse Exposure to Anaplasma phagocytophilum, Borrelia burgdorferi, and Leishmania infantum in Greece through the Detection of IgG Antibodies in Serum and in an Alternative Diagnostic Sample-The Saliva.

Among the various zoonotic pathogens that infect horses, Anaplasma phagocytophilum, Borrelia spp. and Leishmania spp. have gained scientific interest, and relevant molecular and serological studies in horses have been conducted worldwide. Moreover, human and veterinary medicine have extensively applied alternatives to serum diagnostic samples-such as saliva-for detecting pathogens or antibodies. In this study, we investigated the exposure of horses in Greece to A. phagocytophilum, B. burgdorferi, and L. infantum, and we assessed the diagnostic accuracy of saliva compared to serum in detecting IgG antibodies against the abovementioned pathogens. Paired saliva and serum samples were collected from 317 horses from different regions in Greece. The paired samples were examined using the indirect fluorescent antibody test (IFAT) for detecting IgG antibodies against A. phagocytophilum, B. burgdorferi, and L. infantum. Sensitivity, specificity, positive likelihood ratio (PLR), and negative likelihood ratio (NLR) were determined to assess the validity of saliva as an alternative to serum. The receiver operating characteristic (ROC) curve revealed that the optimal cut-off value for detecting antibodies against all the examined pathogens in saliva was 1/10. Higher seropositivity rates were found for B. burgdorferi (15.14%) and A. phagocytophilum (14.19%) compared to L. infantum (1.26%). The detection of IgG antibodies using IFAT in saliva samples had a good test performance compared to serum. The two sample types had a substantial to almost perfect agreement. Although the sensitivity was moderate (70.83-75.56%) in all cases, the specificity was almost perfect to perfect (99.63-100%). This study provides the first evidence that horses in Greece are exposed to A. phagocytophilum and B. burgdorferi and confirms that the seroprevalence of L. infantum in horses in Greece remains low. Our findings suggest that saliva sampling coupled with IFAT could be successfully applied for detecting IgG antibodies against these important zoonotic pathogens in large-scale epidemiological studies in horses, at the population level, as an alternative to serum.

Seroprevalence of and Risk Factors for Toxoplasma gondii Infection in Cats from Greece.

Toxoplasmosis is one of the most important protozoan diseases with a global impact on the health of domestic cats and with zoonotic significance. The aims of this study were to determine the prevalence of seropositivity for Toxoplasma gondii in different populations of cats in Greece and to assess risk factors for seropositivity. A total of 457 cats were prospectively enrolled, and a commercially available indirect immunofluorescence antibody testing (IFAT) kit was used for the detection of anti-T. gondii immunoglobulin G (IgG) in serum. Overall, 95 (20.8%) of the 457 cats were seropositive for T. gondii. Based on multivariate analysis, factors associated with seropositivity included older age [Odds ratio (OR), 1.33; p < 0.001]; a history of cat-fight trauma (OR, 3.88; p = 0.004); and lack of vaccination against calicivirus, herpesvirus-1, panleukopenia, and rabies (OR, 10; p = 0.002). This study shows a high prevalence of seropositivity for T. gondii in cats in Greece. This implies that toxoplasmosis is still a major public health concern and that optimal strategies for the prevention of infection with T. gondii in cats should be established.

Seroprevalence of and risk factors for feline coronavirus infection in cats from Greece.

Feline coronavirus (FCoV) is a highly contagious and ubiquitous virus of domestic cats and wild felids. Feline infectious peritonitis (FIP) is a fatal, systemic disease caused by FCoV infection when spontaneous mutations of the viral genome take place. The aims of this study were primarily to determine the prevalence of seropositivity for FCoV in different populations of cats in Greece and assess risk factors for seropositivity. A total of 453 cats were prospectively enrolled in the study. A commercially available IFAT kit was used for the detection of FCoV IgG antibodies in serum. Overall, 55 (12.1 %) of the 453 cats were seropositive for FCoV. Based on multivariable analysis, factors associated with FCoV-seropositivity included cats adopted as strays and contact with other cats. This is the first extensive study on the epidemiology of FCoV in cats from Greece and one of the largest worldwide. Feline coronavirus infection is relatively common in Greece. Therefore, it is necessary to establish optimal strategies for the prevention of FCoV infection, considering the high-risk groups of cats identified in this study.

Serial Measurement of Serum Pancreatic Lipase Immunoreactivity, Feline Trypsin-like Immunoreactivity, and Cobalamin Concentrations in Kittens.

Serum concentrations of feline pancreatic lipase immunoreactivity (fPLI), feline trypsin-like immunoreactivity (fTLI), and cobalamin are commonly used for the diagnostic investigation of cats with gastrointestinal signs. No information on these parameters in healthy cats less than 1 year of age exists. We aimed to evaluate serum concentrations of fPLI, fTLI, and cobalamin in healthy cats at different time-points during their first 12 months of life. Fourteen healthy 2-month-old kittens were included. Blood was collected at 2, 3, 4, 6, and 12 months of age, and serum concentrations of fPLI, fTLI, and cobalamin were measured. While there was a statistically significant difference in serum fPLI concentrations over time, there was no statistically significant difference between individual time-points. There was no significant difference in serum fTLI concentrations over time. Serum cobalamin concentrations were below the reference interval in 3/13 cats at 2 months of age and were significantly lower by 3 months, when 13/14 had hypocobalaminemia. By 12 months, serum cobalamin had significantly increased, yet 4/12 cats still had hypocobalaminemia. Serum fPLI and fTLI concentrations did not show any statistically or clinically significant differences in young kittens. In contrast, serum cobalamin concentrations were commonly below the reference interval in kittens. Serum fPLI and fTLI concentrations are not practically affected by age in kittens as young as 2 months of age and could be used for the investigation of pancreatic diseases.

Prevalence and Risk Factors for Bartonella spp. and Haemoplasma Infections in Cats from Greece.

Bartonellosis and haemoplasmosis are vector-borne diseases with global impact on the health of domestic cats and of zoonotic importance. The aim of this study was to describe the epidemiological aspects of various populations of cats infected with Bartonella spp. or haemoplasma species. The populations evaluated included client-owned cats, stray cats and cats that live in breeding catteries in Greece. A total of 452 cats were prospectively enrolled into the study. A commercially available indirect immunofluorescence antibody testkit was used for the detection of Bartonella henselae IgG antibodies in serum. PCRs for the detection of Bartonella spp. and haemoplasma species DNA in the blood were also performed in a subgroup of 242 of the 452 cats. Risk factors for B. henselae seropositivity and infection with the haemoplasma species were determined using multivariable analysis. Overall, 160 (35.4%) of the 452 cats were seropositive for B. henselae. Seven (2.9%) and 46 (19%) of the 242 cats were PCR-positive for Bartonella spp. and haemoplasma species, respectively. The factors associated with B. henselae seropositivity, based on multivariate analysis, included older age, outdoor access, living region and flea infestation. Non-administration of ectoparasiticides was associated with haemoplasma species infection. This study shows a high prevalence of seropositivity for B. henselae and a relatively high prevalence of infection with haemoplasma species. Therefore, it is necessary to establish optimal strategies for the prevention of Bartonella spp. and haemoplasma species infections, considering the high-risk groups of cats identified in this study.

Repeatability and reproducibility of microscopic examination of adhesive tape strip cytology slides for the quantification of Malassezia spp. in canine skin.

BACKGROUND: The optimal microscopic magnification and number of optical fields of adhesive tape strip cytological slides that should be examined when searching for Malassezia yeasts on canine skin are unknown. OBJECTIVES: To determine the optimal magnification and the minimum number of optical fields that should be examined to maximise intraobserver repeatability and interobserver reproducibility. MATERIALS AND METHODS: Seven experienced examiners counted, twice, the number of yeasts in 10, 20, 30, 40 and 50 optical fields of 40 slides at ×400 and ×1000 magnification. RESULTS: The number of yeasts per unit surface area was significantly higher at ×1000 compared to ×400 magnification. Repeatability and reproducibility for counting the yeasts was very poor. CONCLUSIONS AND CLINICAL RELEVANCE: Adhesive tape strip cytological slides should be examined microscopically for Malassezia spp. at ×1000 magnification. The repeatability of this examination for counting the yeasts is poor.

Contexte - Le grossissement microscopique optimal et le nombre de champs optiques des lames cytologiques de bandes adhésives à examiner lors de la recherche de levures Malassezia sur la peau de chien sont inconnus. Objectifs - Déterminer le grossissement optimal et le nombre minimal de champs à examiner pour maximiser la répétabilité intra-observateur et la reproductibilité inter-observateur. Matériels et méthodes - Sept examinateurs expérimentés ont compté, deux fois, le nombre de levures dans 10, 20, 30, 40 et 50 champs de 40 lames aux grossissements ×400 et ×1 000. Résultats - Le nombre de levures par unité de surface était significativement plus élevé au grossissement ×1 000 par rapport au grossissement ×400. La répétabilité et la reproductibilité du comptage des levures étaient très médiocres. Conclusions et pertinence clinique - Les lames cytologiques de bandes adhésives doivent être examinées au microscope pour Malassezia spp. à un grossissement ×1 000. La répétabilité de cet examen de comptage des levures est faible.

Introducción- se desconoce el aumento microscópico óptimo y el número de campos ópticos de los portaobjetos citológicos en tiras de cinta adhesiva que deben examinarse al buscar levaduras Malassezia en la piel canina. Objetivos- determinar el aumento óptimo y el número mínimo de campos ópticos que deben examinarse para maximizar la repetibilidad intraobservador y la reproducibilidad interobservador. Materiales y métodos- siete examinadores experimentados contaron dos veces el número de levaduras en campos ópticos de 10, 20, 30, 40 y 50 de 40 portaobjetos con aumentos de x ×400 y ×1000. Resultados- el número de levaduras por unidad de superficie fue significativamente mayor con un aumento de ×1000 en comparación con un aumento de ×400. La repetibilidad y reproducibilidad para contar las levaduras fue muy pobre. Conclusiones y relevancia clínica - Los portaobjetos citológicos en tiras de cinta adhesiva deben examinarse microscópicamente para detectar Malassezia spp. con un aumento de ×1.000. La repetibilidad de este examen para contar las levaduras es pobre.

Contexto - A ampliação microscópica ideal e o número de campos ópticos das lâminas citológicas de fita adesiva que devem ser examinados nas pesquisas de leveduras do gênero Malassezia em cães são desconhecidos. Objetivos - Determinar a magnificação ideal e o número mínimo de campos ópticos que devem ser examinados para maximizar a repetibilidade intraobservador e a reproducibilidade interobservador. Materiais e métodos - Sete examinadores experientes contaram duas vezes o número de leveduras em 10, 20, 30, 40 e 50 campos ópticos de 40 lâminas nas magnificações de x400 e x1000. Resultados - O número de leveduras por unidade de área de superfície foi significativamente maior em x1000 em comparação com a ampliação de x400. A repetibilidade e a reprodutibilidade para a contagem de leveduras foi muito pobre. Conclusões e relevância clínica - Lâminas de citologia por fica adesiva devem ser examinadas microscopicamente para Malassezia spp a uma magnificação de x1.000. A repetibilidade deste exame para contagem de leveduras foi pobre.

Quantitative and Morphological Blood Cell Findings Associated with the Presence of Antigen and/or Antibodies Against Anaplasma phagocytophilum in Sheep.

Anaplasma phagocytophilum is the causative agent of a disease known as tick borne fever in sheep, although fever is not always present. Due to inconclusive clinical signs, diagnosis is based on the cytological or molecular detection of the microorganism in blood and/or the determination of antibodies against A. phagocytophilum. The aim of the study was to determine the alterations caused by the presence of antibodies and/or the antigen of A. phagocytophilum in the blood cell count and morphology in sheep. Cytology and indirect immunofluorescence assay were performed for detection of antibodies and the antigen of A. phagocytophilum, respectively. The samples were divided into four groups depending on the result of the antigen and antibody detection. The samples that were only positive for antigen detection had mild anemia, leukopenia (lymphopenia), and thrombocytopenia. The samples that were positive in both assays had anemia, leukopenia (neutropenia and lymphopenia), and thrombocytopenia. Samples that were positive only for antibody detection had mild leukopenia. Morphological findings in infected sheep included band neutrophils, toxic neutrophils, reactive lymphocytes, and activated monocytes. The hematological findings along with cytological and serological tests can contribute to the assessment of the stage of the disease. A combination of leukopenia and thrombocytopenia raises a strong suspicion of the disease. When the microorganism and antibodies are simultaneously present, sheep are more susceptible to secondary complications. The first reported morphological findings and the quantitative hematological alterations are indicative of an inflammatory reaction, antigenic stimulation, and stress.