Clinicopathological findings and risk factors associated with Cytauxzoon spp. infection in cats: A case-control study (2008-2021).

In Europe, Cytauxzoon spp. infection was documented in domestic and wild felids. Cats often develop a subclinical infection, while fatal disease is rare. Currently, information on the epidemiology, risk factors and clinicopathological findings of Cytauxzoon spp. infection remains limited and obtained by a single subject or small groups of cats. The objective of this case-control study was to evaluate clinicopathological findings and to describe risk factors associated with Cytauxzoon spp. infection in domestic cats. Infected cats (n = 39) and non-infected (n = 190) cats were selected from the database of the referral San Marco Veterinary Laboratory between 2008 and 2021. Demographic information, a preset questionnaire considering lifestyle, environment, and clinical status, and a CBC performed contextually with the PCR analysis were recorded for all cats. Data on the biochemical profile and serum protein electrophoresis were also evaluated when available. Compared to the control group, infection was more likely to occur in stray cats (24/39, 61.5%, P < 0.001), living totally/partially outdoors (36/39, 92.3%, P < 0.001), in an urban context (37/39, 94.9%, P = 0.002), taken or recently adopted from colonies (34/35, 97.1, P < 0.001), with irregular or absent parasite preventive treatments (39/39, 100%, p = 0.005), without fleas (28/35, 80%, P = 0.047) and without clinical signs (22/39, 56.4%, p = 0.026) at the time of medical evaluation. Anemia was not associated with infection, but in cats without clinical signs, the percentage of anemic-infected cats (7/22, 31.8%, P = 0.009) was higher compared to non-infected cats (5/65, 7.7%). Furthermore, a decrease in total iron serum concentration approximating the lowest reference interval [median values (IQR): 79 µg/dL (52.25) vs. 50.5 µg/dL (34), P = 0.007] was likely in infected cats. No other laboratory findings were associated with infection. Interestingly, a partial/total outdoor lifestyle was a risk factor for infection (OR: 8.58, 95% CI: 2.90-37.0, P < 0.001). In conclusion, the present study revealed that Cytauxzoon spp. infection manifests itself prevalently as a subclinical infection, based on physical examination and laboratory findings, in domestic European cats. However, subclinical infected cats were more likely to be anemic compared to non-infected.

Babesia gibsoni infection in Italy: a cross sectional study of 607 blood samples belonging to dogs that needed a molecular analysis investigation (2016-2019).

Babesia gibsoni (B. gibsoni) infection was sporadically described in Europe. In Italy, in particular, it was detected by PCR analysis only in one dog with unclear source of infection, while it has never been found in molecular studies involving more subjects. The aim of this retrospective, cross-sectional study was to determine the presence and the proportion of B. gibsoni infected dogs in a convenient Italian canine population. Babesia spp PCR analysis reports of canine blood samples collected from January 2016 to December 2019 were selected from the San Marco veterinary laboratory database and evaluated. Piroplasms PCR analysis was performed on 18S rRNA gene and positive samples were sequenced. Available data about signalment, living area and season of diagnosis were evaluated and statistically analyzed. The studied canine population had a median age of 62 months (interquartile range, IQR: 72.5) and was composed of male/female purebred and mixed breed dogs. Through sequencing, B. canis (26/40, 65%), B. vogeli (7/40, 17.5%), B. gibsoni (6/40, 15%) and B. vulpes (1/40, 2.5%) infections were reported. The proportion of B. gibsoni infected dogs in the described population was of 0.99%. The infection resulted more likely in male (6/6, 100%), purebred (5/6, 83.3%) dogs with median age of 40.5 months (IQR:32.3). In one case, the clinical signs and parasitemia appeared after a dogfight with a cohabitant infected dog. No other plausible sources of infection were detected. Likely, this could be considered the first autochthonous B. gibsoni infection case documented in Italy. Our results underline the need of considering B. gibsoni not only an exotic, random and rare cause of disease for dogs in Italy, but a possible emerging pathogen to take into account during diagnostic and epidemiological investigations, in symptomatic as well as in asymptomatic patients.

Prevalence and molecular characterization of canine and feline hemotropic mycoplasmas (hemoplasmas) in northern Italy.

BACKGROUND: Hemotropic mycoplasmas (hemoplasmas), the agents of infectious anemia, have been reported in dogs and cats. Little data are available on hemoplasma infections in Italy. The aim of this study was to evaluate the species of hemoplasmas and their prevalence in dogs and cats of northern Italy. METHODS: Blood samples were obtained from 117 candidate blood donor dogs, 278 free-roaming dogs and 227 free-roaming cats in 2014 and 2015. Samples were first screened for hemoplasmas with a SYBR green real time PCR. The positive samples were confirmed by a second SYBR green real time PCR and sequencing. Co-infections were detected using species-specific SYBR green real time PCR. RESULTS: The overall prevalence in dogs was 4.5% (18/395). Among the donors only one dog was positive for Mycoplasma haemocanis (0.8%). The overall prevalence of infection in free-roaming dogs was 6.1% (17/278), which was significantly higher than in candidate donors (P < 0.05). Both M. haemocanis (13/278; 4.7%) and "Candidatus M. haematoparvum" (4/278; 1.4%) were identified. In dogs, no significant association was found between hemoplasma infection and gender, age or origin. The overall prevalence in cats was 13.2% (30/227). All three feline hemoplasma species were detected, i.e. "Candidatus Mycoplasma haemominutum" (28; 12.3%), "Candidatus Mycoplasma turicensis" (11; 4.8%) and Mycoplasma haemofelis (9; 4.0%). Half of the infected cats were co-infected (15; 6.6%) with different species of hemoplasmas. Risk factor analysis confirmed that older age, male gender and FIV positivity are predisposing factors for hemoplasma infection in cats. CONCLUSION: This study found that candidate blood donor dogs in northern Italy show a negligible risk for hemoplasma infection, confirming the appropriateness of the candidate selection criteria and the low prevalence in the study area. Accordingly, testing for hemoplasma should be considered optional for canine blood donor screening. Hemoplasma infection was instead common in free-roaming cats, and is expected to be non-negligible in owned cats with outdoor access. Feline candidates for blood donation will therefore need to be carefully selected.

Exposure to vector-borne pathogens in candidate blood donor and free-roaming dogs of northeast Italy.

BACKGROUND: Many vector-borne pathogens including viruses, bacteria, protozoa and nematodes occur in northeast Italy, representing a potential threat to animal and human populations. Little information is available on the circulation of the above vector-borne pathogens in dogs. This work aims to (i) assess exposure to and circulation of pathogens transmitted to dogs in northeast Italy by ticks, sandflies, and mosquitoes, and (ii) drive blood donor screening at the newly established canine blood bank of the Istituto Zooprofilattico Sperimentale delle Venezie. METHODS: Blood samples from 150 privately-owned canine candidate blood donors and 338 free-roaming dogs were screened by serology (IFA for Leishmania infantum, Ehrlichia canis, Anaplasma phagocythophilum, Babesia canis, Rickettsia conorii, R. rickettsii), microscopic blood smear examination, and blood filtration for Dirofilaria spp. All candidate donors and seropositive free-roaming dogs were tested by PCR for L. infantum, E. canis, A. phagocythophilum, Babesia/Theileria and Rickettsia spp. The dogs had no clinical signs at the time of sampling. RESULTS: Overall, 40 candidate donors (26.7 %) and 108 free-roaming dogs (32 %) were seroreactive to at least one vector-borne pathogen. Seroprevalence in candidate donors vs free-roaming dogs was: Leishmania infantum 6.7 vs 7.1 %; Anaplasma phagocytophilum 4.7 vs 3.3 %; Babesia canis 1.3 vs 2.7 %; Ehrlichia canis none vs 0.9 %; Rickettsia conorii 16 vs 21.3 % and R. rickettsii 11 vs 14.3 %. Seroreactivity to R. rickettsii, which is not reported in Italy, is likely a cross-reaction with other rickettsiae. Filariae, as Dirofilaria immitis (n = 19) and D. repens (n = 2), were identified in free-roaming dogs only. No significant differences were observed between candidate donors and free-roaming dogs either in the overall seroprevalence of vector-borne pathogens or for each individual pathogen. All PCRs and smears performed on blood were negative. CONCLUSIONS: This study demonstrated that dogs are considerably exposed to vector-borne pathogens in northeast Italy. Although the dog owners reported regularly using ectoparasiticides against fleas and ticks, their dogs had similar exposure to vector-borne pathogens as free-roaming dogs. This prompts the need to improve owner education on the use of insecticidal and repellent compounds in order to reduce the risk of arthropod bites and exposure to vector-borne pathogens. Based on the absence of pathogens circulating in the blood of healthy dogs, the risk of transmission of these pathogens by blood transfusion seems to be low, depending also on the sensitivity of the tests used for screening.

First detection of Cytauxzoon spp. infection in European wildcats (Felis silvestris silvestris) of Italy.

Cytauxzoonosis is an emerging, tick-transmitted, protozoan disease affecting domestic and wild felids and caused by Cytauxzoon felis, Cytauxzoon manul and Cytauxzoon spp. This study aimed to determine the presence of infection with Cytauxzoon spp. in Felis silvestris silvestris in Italy, in order to enhance the comprehension of its pattern distribution among domestic cat populations. In addition, wildcats were tested for other endemic vector-borne pathogens in Italy. The carcasses of 21 F. s. silvestris were collected from central and northern regions of Italy. All the animals were submitted to necropsy and samples of the spleens were collected. Cytauxzoon infection was surveyed by a conventional PCR amplifying a portion of the SSU-rDNA of species of Piroplasmida. The samples were also screened for Anaplasma spp., Ehrlichia spp., Rickettsia spp., Babesia spp., Theileria spp., and Leishmania spp. using SYBR Green Real-Time PCR (rPCR) assays. Four animals (19%) were positive for Piroplasmida-PCR assay and three sequenced amplicons were obtained (14.3%), clustering with the Italian, Spanish, French and Romanian Cytauxzoon spp. isolates and with C. manul found in Mongolia. The samples were negative for the other pathogens screened. The present results showed that Cytauxzoon spp. may infect both F. s. silvestris and F. s. catus.

Cytauxzoon sp. infection in two free ranging young cats: clinicopathological findings, therapy and follow up.

Two young brother male free-ranging domestic shorthair cats were evaluated for diarrhea. They presented with intraerythrocytic piroplasms on blood smear evaluation. Only the first cat was anemic (mild non-regenerative anemia). A partial segment of the 18S rRNA was amplified and sequenced, revealing a homology of 99% with Cytauxzoon sp. and of 93% with Cytauxzoon felis. The first cat was treated with doxycycline and imidocarb dipropionate and monitored by serial laboratory exams, resulting negative for Cytauxzoon sp. infection after the end of the therapy (follow-up period of 175 days). The second cat received the same therapy, but doxycycline was discontinued by the owner after 1 week. He was monitored for 130 days, remaining erythroparasitemic and asymptomatic. We described cases of Cytauxzoon sp. infection in domestic cats with detailed clinical data, description of two therapeutic protocols, and follow-up after treatment with opposite parasitological responses (parasitological cure versus persistence of infection).

Serum acute phase protein concentrations in dogs with hyperadrenocorticism with and without concurrent inflammatory conditions.

BACKGROUND: Acute phase proteins (APPs) are promising markers of inflammation in dogs, because they are more sensitive than WBC counts in detecting clinical and subclinical inflammation. Endogenous corticosteroids can mask an acute phase response and make it more difficult to identify underlying inflammatory disease. OBJECTIVE: The purpose of this study was to evaluate the acute phase protein response in dogs with spontaneous hyperadrenocorticism (HAC) with and without concurrent inflammatory conditions. METHODS: Serum concentrations of C-reactive protein (CRP), haptoglobin (Hp), fibrinogen, and albumin were measured in 44 healthy adult dogs and 39 dogs with HAC; the HAC group was further divided into dogs with and without concurrent infection/inflammation. A fourth group of dogs with severe sepsis and without HAC was compared with the dogs with HAC and severe sepsis. RESULTS: Dogs with uncomplicated HAC had significantly higher Hp and fibrinogen concentrations compared with healthy control dogs (P<.001). Dogs with HAC and severe inflammatory disease also had significantly higher CRP and lower albumin concentrations than control dogs and dogs with HAC without concurrent inflammation. Dogs with sepsis but without HAC had significantly higher CRP concentrations than dogs with HAC and sepsis. CONCLUSIONS: Dogs with HAC had increases in the moderate APPs (Hp and fibrinogen), and no significant changes in CRP and albumin compared with healthy dogs. Although concurrent HAC appeared to blunt the CRP response in dogs with sepsis, increased serum CRP concentration in dogs with HAC is likely indicative of severe concurrent inflammation.

Hematologic abnormalities and flow cytometric immunophenotyping results in dogs with hematopoietic neoplasia: 210 cases (2002-2006).

BACKGROUND: Growing interest in veterinary oncohematology has facilitated the recent development and advancement of new techniques, such as flow cytometry, for immunophenotyping hematopoietic neoplasia in animals. OBJECTIVE: The aim of this retrospective study was to characterize hematologic abnormalities and flow cytometric immunophenotyping (FCI) results in cases of hematopoietic neoplasia in dogs. METHODS: Signalment, CBC data, and FCI results were obtained for 210 dogs with blood samples submitted to our laboratory. Immunophenotyping was carried out using an Epics XL-MCL flow cytometer and a panel of 10 antibodies (CD45, CD3, CD4, CD8, CD79, CD21, CD14, CD34, CD41/61, CD61). The prevalence and severity of hematologic abnormalities was determined for the different types of hematopoietic neoplasms. RESULTS: Based on cell morphology and phenotype, cases were classified as: acute lymphoblastic leukemia (ALL, n=51), acute myeloid leukemia (AML, n=33), chronic lymphocytic leukemia (CLL, n=61), and leukemic high-grade lymphoma (L-HGL, n=65). Most cases of ALL (47/51) and L-HGL (41/65) had a B-cell phenotype, while most cases of CLL (54/61) had a T-cell phenotype, with a high prevalence of the large granular lymphocyte subtype (49/61). Anemia was found in 85% of all cases and was significantly more severe in ALL and AML compared with CLL and L-HGL. Neutropenia was seen in 64-78% of acute leukemias (AML and ALL) in contrast to no cases of CLL and 11% of L-HGL. Thrombocytopenia was seen in 88-90% of acute leukemias in contrast to 15% of CLL and 40% of L-HGL. Thrombocytopenia was more prevalent (71% vs 22%) and significantly more severe in T-cell vs B-cell L-HGL. CONCLUSION: A standard CBC is useful in suggesting the type of hemoproliferative disorder and may also help to predict the phenotype, especially in cases of L-HGL.

Evaluation of EDTA hematology tubes for collection of blood samples for tests of secondary hemostasis in dogs.

OBJECTIVE: To evaluate the use of EDTA tubes for collection of blood samples for assays of secondary hemostasis in dogs. ANIMALS: 108 dogs of various ages, breeds, and sexes (19 healthy and 89 with abnormalities of secondary hemostasis). PROCEDURES: Blood samples were collected via cephalic venipuncture and transferred to sodium citrate tubes and EDTA tubes. Plasma was harvested from each type of tube for assays of concentrations of fibrinogen and D-dimer as well as prothrombin time, activated partial thromboplastin time, and antithrombin activity. Intra-assay and interassay precision and correlation coefficients for all hemostatic tests were calculated for each type of plasma sample. The effect of storage conditions on assay results for the 2 types of plasma samples was also evaluated. RESULTS: Results of hemostatic tests were highly correlated between citrated and EDTA-treated plasma samples. Intra-assay imprecision for all hemostatic tests with the exception of D-dimer concentration was < 10% for both citrated and EDTA-treated plasma samples; interassay imprecision was higher for EDTA-treated versus citrated plasma samples. Storage of plasma samples for 1 hour did not result in significantly different assay results for either type of plasma sample, but storage for 2 hours significantly affected values for EDTA-treated plasma samples. CONCLUSIONS AND CLINICAL RELEVANCE: Although evaluation of the sensitivity and specificity of hemostatic tests that use EDTA-treated plasma samples is required, EDTA may be a suitable alternative to sodium citrate as an anticoagulant for use in hemostatic testing in conditions in which tests could be performed within 1 hour after sample collection.

Vertebral polyostotic lymphoma in a young dog.

An unusual clinical presentation of lymphoma with vertebral involvement in a dog is reported. A 20-month-old intact female Golden Retriever presented with progressive paraparesis and anorexia. Complete blood count and serum biochemistry profile demonstrated pancytopenia and hypercalcemia. Ventral fusion of the lumbar vertebrae by new bony tissue deposition was evident on X-ray and CT scan. Fine needle aspiration revealed neoplastic lymphoid cells in lymph nodes and bone marrow. Histologically, vertebral bone and osteophytes, liver, bone marrow, kidney, and lymph nodes were diffusely infiltrated by neoplastic, lymphoid cells, with scant cytoplasm and round hyperchromatic nuclei. Polyostotic and medullary T-cell lymphoma with spondylosis was diagnosed. Lymphoma mainly affecting bone is uncommon in the dog. The present case differs from previously described polyostotic lymphomas in clinical signs of the disease, mainly attributable to spondylarthrosis. In addition, lymphomatous proliferation was associated with osteoproductive lesions of the vertebrae.

Artifactual changes in canine blood following storage, detected using the ADVIA 120 hematology analyzer.

BACKGROUND: Artifactual changes in blood may occur as a consequence of delayed analysis and may complicate interpretation of CBC data. OBJECTIVE: The aim of this study was to characterize artifactual changes in canine blood, due to storage, using the ADVIA 120 hematology analyzer. METHODS: Blood samples were collected into EDTA from 5 clinically healthy dogs. Within 1 hour after blood sample collection and at 12, 24, 36 and 48 hours after storage of the samples at either 4 degrees C or room temperature (approximately 24 degrees C), a CBC was done using the ADVIA 120 and multispecies software. A linear mixed model was used to statistically evaluate significant differences in values over time, compared with initial values. RESULTS: The HCT and MCV were increased significantly after 12 hours of collection at both 4 degrees C and 24 degrees C, and continued to increase through 48 hours. The MCHC initially decreased significantly at 12-24 hours and then continued to decrease through 48 hours at both temperatures. Changes in HCT, MCV, and MCHC were greater at 24 degrees C than at 4 degrees C at all time points. A significant increase in MPV and a decrease in mean platelet component concentration were observed at all time points at 24 degrees C. Samples stored at 24 degrees C for 48 hours had significantly higher percentages of normocytic-hypochromic RBCs, and macrocytic-normochromic RBCs, and lower platelet and total WBC counts. CONCLUSIONS: Delayed analysis of canine blood samples produces artifactual changes in CBC results, mainly in RBC morphology and platelet parameters, that are readily detected using the ADVIA 120. Refrigeration of specimens, even after 24 hours of storage at room temperature, is recommended to improve the accuracy of CBC results for canine blood samples.

A retrospective study of 60 cases of eccentrocytosis in the dog.

BACKGROUND: Eccentrocytes are RBCs that appear in a peripheral blood smear to have their hemoglobin shifted to one side of the cell. This abnormality, which is confined to the RBC membrane and cytoskeleton, is induced by oxidative damage. Eccentrocytes have been reported rarely in dogs and are associated with onion and garlic ingestion and the administration of oxidant drugs. OBJECTIVE: The purpose of this study was to describe the occurrence and severity of eccentrocytosis in dogs and the diseases or disorders associated with eccentrocytes. METHODS: Detailed history, and results of physical examination, CBC, biochemical and coagulation profiles, and urinalysis from all canine patients admitted during a 2.5-year period were evaluated. Eccentrocytes, when observed, were graded 1+ (few) to 4+ (many). The severity of eccentrocytosis was compared with that of anemia and reticulocytosis. RESULTS: Eccentrocytes were found in blood smears from 60 of 4251 dogs (1.4%) and were associated with mild to moderate anemia in 40 (66.6%) of the cases. Eccentrocytosis was found in 16 (26.6%) dogs with drug administration, 11 (18.3%) with presumptive onion and garlic ingestion, 8 (13.3%) with vitamin K antagonist intoxication, 7 (11.6%) with ketoacidotic diabetes, 5 (8.3%) with T-cell lymphoma, 4 (6.6%) with severe infections, 1 (1.6%) with compensated diabetes mellitus, and 8 (13.3%) with other conditions. Certain dog breeds, such as Whippet, Boxer, and English Setter, and young dogs seemed to be overrepresented. CONCLUSIONS: We describe, for the first time, associations between eccentrocytes and diabetes mellitus, T-cell lymphoma and vitamin K antagonist intoxication in dogs. The significance of eccentrocytes should not be underestimated, because they can be a signal of an oxidative process.

Use of fine needle aspirates and flow cytometry for the diagnosis, classification, and immunophenotyping of canine lymphomas.

Fifty canine lymphomas were classified cytomorphologically using the updated Kiel classification scheme. Aspirates of lymph nodes from dogs with lymphoma were stained using 5 canine-specific antibodies and 3 human-specific antibodies that cross-react with canine lymphocytes. The antibody-stained aspirates were analyzed by flow cytometry. A total of 32 (64%) of the 50 lymphomas were characterized as B-cell origin and 18 (36%) were of T-cell origin. B-cell lymphomas were identified in 12 females and 20 males with a mean age of 8.35 years. T-cell lymphomas were identified in 8 females and 10 males with a mean age of 7.9 years. A minority of the lymphomas were low-grade B-cell and T-cell lymphomas (6/50, 12% and 4/50, 8%, respectively). The most common morphologic types were high-grade centroblastic and unclassifiable plasmacytoid for B- and T-cell lymphomas (18/50, 36% and 7/50, 14%, respectively).