Hyperbetaglobulinemia, anaemia and thrombocytopenia in a domestic ferret (Mustela putorius furo) associated to Leishmania infantum.

Leishmaniosis in domestic ferrets is a vector-borne disease caused in Europe by the protozoan parasite Leishmania infantum. There is limited information on clinical signs and laboratory abnormalities in ferrets due to leishmaniosis. This clinical case report described a domestic ferret (Mustela putorius furo) with severe hyperbetaglobulinemia, anaemia, thrombocytopenia, and abnormal renal parameters. A good clinical response following an anti-Leishmania infantum treatment protocol was achieved. However, the presence of pain at the site of injection was the main side effect due to meglumine antimoniate administration. Xanthine crystalluria was not observed in urine sediment with no other urine alterations detected by urine analysis during the follow-up. Initially, clinical signs noted in this ferret could not initially be attributed to leishmaniosis. However, no causes were found that could have caused the hyperglobulinemia in this patient. A reduction of the levels of anti-L. infantum serum antibodies and the concentrations of beta-globulin fraction was detected in this patient after anti-Leishmania treatment administered as well as the disappearance of thrombocytopenia. To extent of the knowledge of leishmaniosis in ferrets, this is the fourth case report of leishmaniosis documented in this species.

First report on natural infection by Dirofilaria repens in a cat in Spain: case report and literature review of feline subcutaneous dirofilariosis in Europe.

Dirofilaria repens is an endemic mosquito-borne pathogen widely spread throughout Europe and other non-Europen regions. Infection by D. repens has been reported in dogs, although little is known about the occurrence and epidemiological features of this nematode in cats. During the surgical procedure within the framework of a trap, neuter and release sterilization program, two female filarial nematodes with spontaneous movement were removed from the internal part of the spermatic cord from asymptomatic stray cat in Spain (Grao de Castellón, Castellon de la Plana, province of Spain). Moreover, the presence of microfilariae was detected by using three different methods, including direct blood smear technique, the microhematocrit tube test, and Knott´s modified test. PCR performed from the nematodes extracted from the spermatic cord and from microfilariae in EDTA-blood sample tested both positive for D. repens. To the authors' knowledge, this is the first report that demonstrates the presence of D. repens in cats in Spain. In this sense, a review of the current scientific status of feline subcutaneous dirofilariosis in Europe was also performed. Future investigations should analyze the epidemiological role of cats in D. repens infection including the prevalence of infection. Veterinarians working in endemic areas should be aware of this infection in cats and their susceptibility.

The dynamics of neutralizing antibodies against SARS-CoV-2 in cats naturally exposed to virus reveals an increase in antibody activity after re-infection.

Severe Acute Respiratory Syndrome Coronavirus 2 is the causative agent of Coronavirus Disease 2019 in humans. To date, little is known about the persistence of antibodies against SARS-CoV-2 in animals under natural conditions, in particular susceptible pets such as cat. This study reports the detection and monitoring of the humoral response against SARS-CoV-2 including the detection of immunoglobulins G specific for receptor binding domain of SARS-CoV-2 spike protein by an enzyme-linked immunosorbent assay and neutralizing antibodies by virus neutralization assay. Results showed that these antibodies last longer than 16 months in two naturally apparently healthy infected cats with the absence of clinicopathological findings during the follow-up. Moreover, re-infection is also possible with an important increase in virus neutralization test titers in both animals with no evident systemic signs found during each physical examination and with values of hematologic and biochemical parameters inside the normal reference intervals. Our results confirm a slow but progressive decrease of the kinetics and immunity of neutralizing antibodies in cats after the infection. Furthermore, similar to humans SARS-CoV-2 reinfection can stimulate an increase of the neutralizing antibodies determined by these two serological techniques in domestic cats.

Evaluation of an immunochromatographic serologic test to detect the presence of anti-Toxoplasma gondii antibodies in cats.

BACKGROUND: Toxoplasmosis is a protozoan disease caused by Toxoplasma gondii. Different T. gondii confirmatory techniques, including serologic methods, are available to detect the presence of the parasite. Among serology techniques, immunochromatographic rapid testing could be a reliable alternative to serologic laboratory techniques. OBJECTIVE: This study evaluated a commercial immunochromatographic test (FASTest TOXOPLASMA g) in seronegative and seropositive cats. METHODS: Two indirect immunofluorescence antibody reference tests, an in-house technique, and a commercial test were used to classify 292 feline serum samples. The rapid test was evaluated in different groups of cats, including healthy seronegative cats (n = 121), seropositive cats with variable anti-Toxoplasma antibodies (n = 146), and cats with positive serologic results for other pathogens (n = 25). The sensitivity, specificity, accuracy, receiver operating characteristic curves, and kappa statistics were analyzed as performance measures. RESULTS: Of the 292 samples, 146 were classified as T. gondii seropositive and 146 as T. gondii seronegative. Concordant results were obtained for all samples using immunofluorescence antibody tests. The diagnostic measures of this rapid test showed 98.63% sensitivity and 100% specificity, and 99.32% accuracy. The kappa statistics value was 0.986, and the area under the receiver operating characteristic curve was 0.993. CONCLUSIONS: This rapid test showed diagnostic measurements similar to those of traditional quantitative serologic methods. In situations where laboratory techniques are not available, this test, under clinical conditions, could be a useful alternative to obtain accurate results rapidly.

A cross-sectional serosurvey of SARS-CoV-2 and co-infections in stray cats from the second wave to the sixth wave of COVID-19 outbreaks in Spain.

Severe Acute Respiratory Syndrome Coronavirus 2 is the causative agent of Coronavirus Disease 2019 in humans. Among domestic animals, cats are more susceptible to SARS-CoV-2 than dogs. The detection of anti-SARS-CoV-2 antibodies in seemingly healthy cats and/or infected cats which are in close contact with infected humans has been described. The presence of animals that tested positive by serology or molecular techniques could represent a potential transmission pathway of SARS-CoV-2 that can spill over into urban wildlife. This study analyses the seroprevalence variation of SARS-CoV-2 in stray cats from different waves of outbreaks in a geographical area where previous seroepidemiological information of SARS-CoV-2 was available and investigate if SARS-CoV-2-seropositive cats were exposed to other co-infections causing an immunosuppressive status and/or a chronic disease that could lead to a SARS-CoV-2 susceptibility. For this purpose, a total of 254 stray cats from Zaragoza (Spain) were included. This analysis was carried out by the enzyme-linked immunosorbent assay using the receptor binding domain of Spike antigen and confirmed by serum virus neutralization assay. The presence of co-infections including Toxoplasma gondii, Leishmania infantum, Dirofilaria immitis, feline calicivirus, feline herpesvirus type 1, feline leukemia virus and feline immunodeficiency virus, was evaluated using different serological methods. A seropositivity of 1.57% was observed for SARS-CoV-2 including the presence of neutralizing antibodies in three cats. None of the seropositive to SARS-CoV-2 cats were positive to feline coronavirus, however, four SARS-CoV-2-seropositive cats were also seropositive to other pathogens such as L. infantum, D. immitis and FIV (n = 1), L. infantum and D. immitis (n = 1) and L. infantum alone (n = 1).Considering other pathogens, a seroprevalence of 16.54% was detected for L. infantum, 30.31% for D. immitis, 13.78%, for T. gondii, 83.86% for feline calicivirus, 42.52% for feline herpesvirus type 1, 3.15% for FeLV and 7.87% for FIV.Our findings suggest that the epidemiological role of stray cats in SARS-CoV-2 transmission is scarce, and there is no increase in seropositivity during the different waves of COVID-19 outbreaks in this group of animals. Further epidemiological surveillances are necessary to determine the risk that other animals might possess even though stray cats do not seem to play a role in transmission.

Anaemia in Sheep Caused by Babesia and Theileria Haemoparasites.

Piroplasmoses in sheep are caused by vector-borne apicomplexan protozoa, Babesia and Theileria. Different species are responsible for the disease; some species are more pathogenic than others and have a worldwide distribution. In this sense, these causative agents can cause anaemia in flocks. In general, these vector-borne diseases infect small ruminants and cause host-mediated pathology. In the case of Babesia species, a combination of different mechanisms is involved: red blood cell lysis due to intracellular parasite multiplication, activation of biogenic amines and the coagulation system with the possibility of disseminated intravascular coagulation. By contrast, less information is available on the different immunopathogenic mechanisms involved in the development of anaemia in sheep with theileriosis. However, the mechanisms of pathogenic action in theileriosis are similar to those studied in babesiosis. Diagnosis is based on compatible clinical signs, laboratory findings, specific diagnostic tests and the presence of the tick vector. Some of these tests detect the causative agent itself, such as direct identification by light microscopy and molecular analysis. In contrast, other tests detect the sheep's immune response to the organism by serology. Both diseases pose a significant diagnostic challenge for veterinary practitioners around the world. This review presents the most frequent clinical signs, pathogenesis and clinicopathological findings, diagnosis.

First serological study of Dirofilaria immitis antibodies in household domestic ferrets (Mustela putorius furo) in southern Spain - Short communication.

Dirofilaria immitis is an endemic mosquito-borne pathogen widely spread throughout Europe as well as North and South America. Infection by D. immitis has been reported in domestic ferrets, although little is known about the occurrence and the epidemiological features of this nematode in this species. The aim of the present retrospective study was to assess the prevalence of D. immitis antibodies using an in-house enzyme-linked immunosorbent assay specifically developed for use in ferrets. One hundred and eighty-six serum samples were obtained from the Province of Valencia (Spain), an area endemic for dirofilariosis. Of the 186 serum samples included in the study, 27 (14.51%) were classified as D. immitis seropositive and 159 samples as D. immitis seronegative. The results provide valuable information on the seroprevalence of D. immitis infection in domestic ferrets in an area endemic for this vector-borne pathogen. The presence of seropositive ferrets should be taken into account and preventive measures should be implemented, including the possibility of serological screening for the early detection of Dirofilaria antibodies as a serological marker of exposure. This is the first study that demonstrates the presence of D. immitis exposure in ferrets in Spain. Veterinarians working in endemic areas should be aware of this infection in ferrets and their susceptibility.

Serum protein electrophoretogram profile detected in apparently healthy cats infected with Leishmania infantum - Short communication.

The information about the clinical features of Leishmania infantum infection in cats is scarce. In this study, we evaluated the serum protein electrophoresis of samples from 19 infected but apparently healthy cats. To detect L. infantum infection, two serological tests, i.e. western blot (WB) and enzyme-linked immunosorbent assay (ELISA) as well as quantitative polymerase chain reaction (qPCR) on the blood samples were performed. Eventual infection by several selected bacterial and viral pathogens was also tested. All but one of the cats were found positive with WB. The WB-negative cat was positive by ELISA only. From the 18 WB-positive cats, only three were positive also by ELISA and eight with qPCR, including the only animal which was positive in all the three tests. No concomitant infections were detected in any of the cats. The main alteration of the proteinogram was characterised by an increase of the α-2 fraction. In the five cats with hypergammaglobulinaemia, the pattern detected was polyclonal. None of the cats were seropositive to any other pathogens tested. The presence of polyclonal gammopathy and elevation of the α-2 fraction could suggest the presence of active infection. In contrast, the only detection of an increase of the α-2 fraction alone with the presence of positive serological result could be associated by immune response activation against L. infantum.

Evaluation of five different rapid immunochromatographic tests for canine leishmaniosis in Spain.

Canine leishmaniosis is a vector-borne disease caused by Leishmania parasites. Serological methods are the most common tests used for the diagnosis. This study aimed to evaluate and compare different serological commercial immunochromatographic rapid tests available in Spain to detect anti-Leishmania canine antibodies. The immunochromatographic tests were evaluated in different groups of dogs (healthy seronegative dogs (n = 21), naturally-sick dogs with moderate anti-Leishmania antibodies (n = 39), naturally-sick dogs with high anti-Leishmania antibodies (n = 37), dogs with the serological result of other pathogens infection (n = 20) and exposed dogs (n = 33)) admitted to the Veterinary Teaching Hospital of the University of Zaragoza (Spain) according to the clinical information sent with the sample to the laboratory for diagnostic purposes. The serology status was also routinely recorded through an in-house enzyme-linked immunosorbent assay (ELISA) and an in-house indirect immunofluorescence test (IFAT). The qualitative commercial serological immunochromatographic tests used were: FASTest LEISH, Uranotest Leishmania, Uranotest Leishmania 2.0, Speed Leish K, Witness Leishmania, and DFV Test Leishmania. Performance measures analyzed for each test were: sensitivity, specificity, and area under the receiver-operating (ROC) curve. The maximum specificity (1.00) was attained for Uranotest Leishmania and DFT Test Leishmania, followed by FASTest LEISH (0.98), Uranotest Leishmania 2.0 (0.98), Speed Leish K (0.98), and Witness Leishmania (0.95). The maximum sensitivity was attained for FASTest LEISH (1.00), followed by Uranotest 2.0 (0.97), Speed Leish K (0.97), Uranotest (0.96), and the lowest results with Witness (0.84) and DFV Test (0.59). Regarding the ROC curve, the maximum value was attained with the FASTest LEISH (0.99), followed by Uranotest (0.98), Uranotest 2.0 (0.97), Speed Leish K (0.97), Witness (0.90), and the lowest result with DFV Test (0.79). Efforts in the field of diagnosis should focus on establishing a commercial immunochromatographic test with high sensitivity and specificity with a reasonable cost-benefit balance.

Leishmaniosis caused by Leishmania infantum in ferrets: Update review.

Leishmaniosis in domestic ferrets (Mustela putorius furo) is a disease caused by Leishmania infantum, a parasite transmitted through the bite of an infected female phlebotomine sand fly. Among vertebrates, the dog is the primary domestic reservoir of the parasite; however, other domestic animals can be implicated such as cats. The first description of a clinical case of leishmaniosis in domestic ferrets was reported recently. As a result, new knowledge has been published including empirically based treatment protocols, confirmatory techniques to detect the presence of the parasite infection and seasonal variation in the antibodies against Leishmania in apparently healthy domestic ferrets. The most common clinical signs observed are enlargement of peripheral lymph nodes and skin lesions such as papular and/or ulcerative dermatitis. Additionally, the most frequent laboratory alterations seen are hyperproteinaemia with hyperglobulinaemia and biochemical analytes alterations depending on the affected tissue. Two different therapeutic protocols have been described to treat domestic ferrets with leishmaniosis: meglumine antimoniate plus allopurinol protocol or miltefosine plus allopurinol protocol. These treatment protocols seemed to be able to control the Leishmania infection, although the presence of xanthinuria could be detected. The susceptibility of domestic ferrets to Leishmania infantum, the clinical picture, treatment of infected animals and prevention are poorly understood, due to the scarcity of recent description in the literature. Different proposed diagnostic algorithms have been included for domestic ferrets with suspected leishmaniosis, clinically healthy domestic ferrets and animals as blood donors. In this sense, the present review provides updated data on scientific knowledge of leishmaniosis in ferrets.

Serological evidence of SARS-CoV-2 and co-infections in stray cats in Spain.

A new coronavirus known as SARS-CoV-2 emerged in Wuhan in 2019 and spread rapidly to the rest of the world causing the pandemic disease named coronavirus disease of 2019 (COVID-19). Little information is known about the impact this virus can cause upon domestic and stray animals. The potential impact of SARS-CoV-2 has become of great interest in cats due to transmission among domestic cats and the severe phenotypes described recently in a domestic cat. In this context, there is a public health warning that needs to be investigated in relation with the epidemiological role of this virus in stray cats. Consequently, in order to know the impact of the possible transmission chain, blood samples were obtained from 114 stray cats in the city of Zaragoza (Spain) and tested for SARS-CoV-2 and other selected pathogens susceptible to immunosuppression including Toxoplasma gondii, Leishmania infantum, feline leukaemia virus (FeLV) and feline immunodeficiency virus (FIV) from January to October 2020. Four cats (3.51%), based on enzyme-linked immunosorbent assay (ELISA) using the receptor binding domain (RBD) of Spike antigen, were seroreactive to SARS-CoV-2. T. gondii, L. infantum, FeLV and FIV seroprevalence was 12.28%, 16.67%, 4.39% and 19.30%, respectively. Among seropositive cats to SARS-CoV-2, three cats were also seropositive to other pathogens including antibodies detected against T. gondii and FIV (n = 1); T. gondii (n = 1); and FIV and L. infantum (n = 1). The subjects giving positive for SARS-CoV-2 were captured in urban areas of the city in different months: January 2020 (2/4), February 2020 (1/4) and July 2020 (1/4). This study revealed, for the first time, the exposure of stray cats to SARS-CoV-2 in Spain and the existence of concomitant infections with other pathogens including T. gondii, L. infantum and FIV, suggesting that immunosuppressed animals might be especially susceptible to SARS-CoV-2 infection.

Detection of Anti-Leishmania infantum Antibodies in Wild European and American Mink (Mustela lutreola and Neovison vison) from Northern Spain, 2014-20.

The European mink (Mustela lutreola) is listed as a critically endangered species because of ongoing population reduction from habitat degradation and the effects of introduced species, such as American mink (Neovison vison). This small, fragmented population becomes vulnerable to many other threats, including diseases. Leishmaniosis is a zoonotic disease caused by the protozoan parasite Leishmania infantum found in the Mediterranean area, which affects many mammals, including wild small mammals. Furthermore, clinical disease caused by L. infantum has recently been described in other mustelids. To assess the exposure to Leishmania sp. infection in mink species in northern Spain, blood samples from 139 feral American mink and 42 native European mink from north Spain were evaluated for Leishmania sp. infection using enzyme-linked immunosorbent assays against Leishmania spp. antibodies, with 52.4% of American mink and 45.3% of European mink being found seropositive. This finding raises questions regarding how the disease may affect these species and the potential repercussions for conservation efforts. Despite a high seroprevalence being observed in wild mink of both species in this study, association with clinical or pathologic signs of disease has yet to be elucidated.

Clinical leishmaniosis in a domestic ferret (Mustela putorius furo) treated with miltefosine plus allopurinol: Serological and clinical follow-up.

The published information on the treatment of mustelid leishmaniosis is extremely scarce because there are only two case reports available. In one case, a domestic ferret (Mustela putorius furo) was treated with a combination of meglumine antimoniate plus allopurinol and, in the other case, a therapeutic regimen with allopurinol was administrated to a Eurasian otter (Lutra lutra). This article describes for the first time a combined therapeutic protocol with miltefosine (2 mg/kg once a day during 28 days per os), and allopurinol (10 mg/kg twice a day PO sine die) in a domestic ferret with splenomegaly, lymphadenomegaly and a facial pyogranulomatous dermatitis, with a moderate level of antibodies to Leishmania infantum.

Antibodies to Leishmania in naturally exposed domestic ferrets (Mustela putorius furo) in Spain.

Zoonotic leishmaniosis due to Leishmania infantum is a vector-borne disease endemic in southern Europe and dogs are the main reservoir for this infection. Seasonal variations in antibody titers in this species in areas where phlebotomine vectors have seasonal patterns of activity are important for epidemiological, preventive and clinical studies related with canine leishmaniosis. It has been suggested that cats, rabbits and ferrets may act as peridomestic reservoirs and not only as accidental hosts. The aim of this study was to determine if seropositive ferrets (Mustela putorius furo) to Leishmania could be affected by seasonal variations of anti-Leishmania antibodies. A group of seropositive clinically healthy ferrets (n = 21) were included in this study. A significant reduction in anti-Leishmania infantum antibodies was detected during non-transmission period (December 2020-February 2021) in comparison to transmission period (April-October 2020). This study describes for the first time a seasonal variation in the anti-Leishmania antibodies detected in domestic ferrets following natural exposure during sand fly transmission period and the following non-sand fly transmission period in a Mediterranean area considered as an area where L. infantum is endemic.

SARS-CoV-2 Seroprevalence in Household Domestic Ferrets (Mustela putorius furo).

Animal infections with SARS-CoV-2 have been reported in different countries and several animal species have been proven to be susceptible to infection with SARS-CoV-2 both naturally and by experimental infection. Moreover, infections under natural conditions in more than 20 mink farms have been reported where humans could have been the source of infection for minks. However, little information is available about the susceptibility of pet animals under natural conditions and currently there is no SARS-CoV-2 epidemiological assessment occurrence in household ferrets. In this study, the presence of SARS-CoV-2 antibodies was evaluated in serum samples obtained from 127 household ferrets (Mustela putorius furo) in the Province of Valencia (Spain). Two ferrets tested positive to SARS-CoV-2 (1.57%) by in-house enzyme-linked immunosorbent assay based on receptor binding domain (RBD) of Spike antigen. Furthermore, anti-RBD SARS-CoV-2 antibodies persisted at detectable levels in a seropositive SARS-CoV-2 domestic ferret beyond 129 days since the first time antibodies were detected. This study reports for the first time the evidence of household pet ferrets exposure to SARS-CoV-2 in Spain to date.

A cross-sectional study of Leishmania infantum infection in stray cats in the city of Zaragoza (Spain) using serology and PCR.

BACKGROUND: Feline leishmaniosis is a vector-borne parasitic disease caused by Leishmania spp. Leishmania infection in dogs is prevalent in the Mediterranean basin, but in other animals, such as cats, it could also play a role in the epidemiology of the disease. Information on the geographical distribution and epidemiological features of L. infantum infection in cats is scarce, particularly in urban stray cats living in regions where canine leishmaniosis is endemic. As diagnosis can be challenging, combining different serological and molecular methods is a useful approach. Our aim was to investigate the prevalence of infection of L. infantum in apparently healthy stray cats in an endemic region of Spain (Zaragoza city) using serological and molecular methods, and to compare the results of the different techniques. METHODS: The prevalence of Leishmania infection was studied in stray cats captured in urban and peri-urban areas of Zaragoza. Blood was collected from each animal for serology and molecular analysis. Three serological methods, namely the immunofluorescent antibody test (IFAT), enzyme-linked immunosorbent assay (ELISA) and western blot (WB), were used to detect L. infantum antibodies and a real-time PCR (qPCR) assay was used to detect L. infantum DNA. The results were analyzed by Fisher's exact test and Cohen's kappa statistic (κ) to assess the level of agreement between the diagnostic techniques. RESULTS: Serological analysis of blood samples from 180 stray cats revealed 2.2% (4/179) Leishmania infection positivity by IFAT, 2.8% (5/179) by ELISA and 14.5% (26/179) by WB. Leishmania DNA was detected by qPCR in 5.6% (10/179) of the cats. Sixteen cats (8.9%) tested positive by only one serological technique and four tested positive by all three serological methods used. The overall rate of infected cats (calculated as the number of cats seropositive and/or qPCR positive) was 15.6%, and only two cats tested positive by all the diagnostic methods. A significant association was found between male cats and a positive qPCR result. Comparison of the techniques revealed a fair agreement in seropositivity between blood qPCR and IFAT (κ = 0.26), blood qPCR and ELISA (κ = 0.24), WB and ELISA (κ = 0.37) and WB and IFAT (κ = 0.40). The highest agreement between seropositive results was between IFAT and ELISA (κ = 0.89), and the lowest was between blood qPCR and WB (κ = 0.19). The prevalence of the feline leukemia virus antigen was 4.49% (8/178 cats) and that of the feline immunodeficiency virus (FIV) antibody was 6.74% (12/178), while co-infection with both retroviruses was observed in one female cat (1/178). Leishmania ELISA and IFAT seropositivity were statistically associated with FIV status by the chi-square test. CONCLUSIONS: The results obtained in this study, using serological tests and qPCR, indicate the existence of L. infantum asymptomatic infection in apparently healthy stray cats in the city of Zaragoza, an endemic area in Spain.

Prevalence of microfilariae, antigen and antibodies of feline dirofilariosis infection (Dirofilaria immitis) in the Zaragoza metropolitan area, Spain.

Feline heartworm disease is a vector-borne parasitical disease caused by Dirofilaria immitis. Heartworm infection in dogs is prevalent in the Mediterranean countries. Information about the geographical distribution and epidemiological features of D. immitis infection in cats is scarce, particularly in urban stray cats that live within endemic regions for canine heartworm disease. The aim of the current study was to determine the seroprevalence of antigen and antibodies to D. immitis in feral cats in Zaragoza city, an endemic region of Spain. For this purpose, blood samples were examined for microfilariae using a direct blood smear technique and the modified Knott test. Two serological techniques for anti-D. immitis antibody detection (Solo Step® FH and in-house ELISA) and three different commercial antigen tests (DiroChek®, MegaELISA® DIRO Antigen and FASTest® HW) were performed. Blood samples from 250 stray cats were tested: 61 cats (24.40%) tested positive by the in-house ELISA, and 9 cats gave positive (3.6%) results with Solo Step® FH. The global seroprevalence of D. immitis in the feline population of the studied area of Zaragoza was 25.20% (63/250) including Solo Step® FH result and in-house ELISA. The blood exam for all samples was negative when evaluating for microfilariae and not a single cat was positive for antigen testing. This study demonstrates the presence of D. immitis infection in Zaragoza city. Veterinarians working in endemic areas should be aware of this infection in cats at risk and their susceptibility.

Evaluation of the performance of three serological tests for diagnosis of Leishmania infantum infection in dogs using latent class analysis.

Canine leishmaniasis (CanL) is a disease caused by Leishmania infantum. Serological methods are the most common diagnostic techniques used for the diagnosis of the CanL. The objective of our study was to estimate the sensitivity and specificity of one in-house ELISA kit (ELISA UNIZAR) and three commercially available serological tests (MEGACOR Diagnostik GmbH) including an immunochromatographic rapid test (FASTest LEISH®), an immunofluorescent antibody test (MegaFLUO LEISH®) and an enzyme-linked immunosorbent assay (MegaELISA LEISH®), using latent class models in a Bayesian analysis. Two hundred fifteen serum samples were included. The highest sensitivity was achieved for FASTest LEISH® (99.38%), ELISA UNIZAR (99.37%), MegaFLUO LEISH® (99.36%) followed by MegaELISA LEISH® (98.49%). The best specificity was obtained by FASTest LEISH® (98.43%), followed by ELISA UNIZAR (97.50%), whilst MegaFLUO LEISH® and MegaELISA LEISH® obtained the lower specificity (91.94% and 91.93%, respectively). The results of present study indicate that the immunochromatographic rapid test evaluated FASTest LEISH® show similar levels of sensitivity and specificity to the quantitative commercial tests. Among quantitative serological tests, sensitivity and specificity were similar considering ELISA or IFAT techniques.

Treatment and follow-up of a domestic ferret (Mustela putorius furo) with clinical leishmaniosis caused by Leishmania infantum.

Leishmania infantum infection including treatment and follow up in domestic animals other than dogs and cats has not been described at this moment. This article describes the anti-Leishmania treatment and follow-up of a ferret (Mustela putorius furo) with leishmaniosis. A combined therapeutic protocol established for the patient, not yet approved for ferrets, was a combination of meglumine antimoniate plus allopurinol. A follow-up was established monthly during the first year in order to monitor the health condition of the patient. Six months after commencing allopurinol therapy, xanthine crystalluria was observed in urine sediment with no other urine alterations detected by urine analysis. The ferret worsened progressively with diarrhoea and weight loss after cohabiting with another ferret diagnosed with cryptosporidiosis. Cryptosporidium parvum was isolated in faecal samples from the patient detected by three different methods including Ziehl-Neelsen staining, a qualitative test to detection of C. parvum antigens and finally a specific molecular analysis to characterize the species. To the best of the authors´ knowledge, this is the first report providing information about anti-Leishmania protocol therapy used and follow-up in a domestic ferret with clinical leishmaniosis. Veterinarians practicing in endemic areas should be aware of this infection in ferrets at risk and their susceptibility especially when immunosuppressive conditions are present.

First report on natural infection with Leishmania infantum in a domestic ferret (Mustela putorius furo) in Spain.

A pet domestic ferret (Mustela putorius furo) with a papular lesion involving the right pinna was diagnosed with chronic pyogranulomatous dermatitis by histopathologic examination. Intralesional, intracytoplasmic oval microorganisms compatible with Leishmania spp. or Histoplasma spp. were observed in macrophages and multinucleate giant cells. Leishmania infantum (L. infantum) infection was diagnosed by PCR, culture in Novy-MacNeal-Nicolle medium, and immunohistochemistry. Abnormal clinicopathological results included increased alanine transferase, alkaline phosphatase, serum gamma glutamyl transferase and polyclonal gammpathy. Anti-Leishmania antibodies were detected by enzyme-linked immunosorbent assay, immunofluorescence antibody test and western blot using L. infantum antigen. Immunoreactivity against the 16 kDa specific L. infantum antigen fraction was observed by western blot. PCR performed in blood samples obtained from this patient after positive parasite isolation detected L. infantum DNA. To the authors' knowledge, this is the first diagnosis and isolation of L. infantum in a domestic ferret naturally infected in an endemic region (Spain) where canine and feline leishmaniosis is frequently detected. According to these findings, ferrets should be included as potential reservoir hosts of L. infantum. Future investigations should analyze the epidemiological role of ferrets in L. infantum infection including the prevalence of infection.