A tick cell line as a powerful tool to screen the antimicrobial susceptibility of the tick-borne pathogen Anaplasma marginale.

Anaplasma marginale is the causative agent of the severe bovine anaplasmosis. The tick Rhipicephalus microplus is one of the main vectors of A. marginale in tropical and subtropical regions of the world. After the tick bite, the bacterium invades and proliferates within the bovine erythrocytes leading to anemia, impairment of milk production and weight loss. In addition, infection can cause abortion and high mortality in areas of enzootic instability. Immunization with live and inactivated vaccines are employed to control acute bovine anaplasmosis. However, they do not prevent persistent infection. Consequently, infected animals, even if immunized, are still reservoirs of the bacterium and contribute to its dissemination. Antimicrobials are largely employed for the prophylaxis of bovine anaplasmosis. However, they are often used in sublethal doses which may select pre-existing resistant bacteria and induce genetic or phenotypic variations. Therefore, we propose a new standardized in vitro assay to evaluate the susceptibility of A. marginale strains to different antimicrobials. This tool will help health professionals to choose the more adequate treatment for each herd which will prevent the selection and spread of resistant strains. For that, we initially evaluated the antimicrobial susceptibility of two field isolates of A. marginale (Jaboticabal and Palmeira) infecting bovines. The least susceptible strain (Jaboticabal) was used for the standardization of an antimicrobial assay using a culture of Ixodes scapularis-derived tick cell line, ISE6. Results showed that enrofloxacin (ENRO) at 0.25, 1 or 4 µg/mL and oxytetracycline (OTC) at 4 or 16 µg/mL are the most efficient treatments, followed by OTC at 1 µg/mL and imidocarb dipropionate (IMD) at 1 or 4 µg/mL. In addition, this proposed tool has technical advantages compared to the previously established bovine erythrocyte culture. Thereby, it may be used to guide cattle farmers to the correct use of antimicrobials. The choice of the most suitable antimicrobial is essential to eliminate persistent infections, prevent the spread of resistant strains and help controlling of bovine anaplasmosis.

Parasitological and molecular diagnostic of a clinical Babesia caballi outbreak in Southern Romania.

Equine piroplasmosis (EP) is a tick-borne disease of equids caused by Babesia caballi and/or Theileria equi, which is endemic in many tropical and temperate areas of the world. However, clinical outbreaks of EP in Romania during the last decades have not been reported Therefore, the aim of this paper is (i) to describe a clinical B. caballi outbreak in horses on several farms in Southern Romania using a diagnostic and therapeutic approach and (ii) the molecular diagnostic of EP in an endemic area of Romania. In the first case, a 10-month-old stallion male was presented with lethargy, anorexia, fever (40.9 °C), pale mucosal/mucous/membranes and a marked anemia. In the subsequent weeks, three horses from other farms located in the same area, displayed similar clinical signs. B. caballi was diagnosed in all the horses based on Giemsa-stained blood smears and the diagnosis was further confirmed by polymerase chain reaction (PCR), using a single-round and multiplex PCR and sequencing. All four horses were treated with imidocarb dipropionate, at a dose rate of 2.2 mg/kg body weight (two injections at 48 h apart), and all horses clinically recovered within 24-48 h, post-treatment. This report presents the first molecularly characterized B. caballi outbreak in Romania in clinically affected horses, confirmed by DNA sequencing.

First clinical case report of Cytauxzoon sp. infection in a domestic cat in France.

BACKGROUND: Feline cytauxzoonosis is an emerging infection caused by tick-transmitted apicomplexan parasites of the genus Cytauxzoon. The association of clinical disease with Cytauxzoon infection appears to be limited to C. felis infections in the Americas. Sporadic infections of wild and domestic felids with Cytauxzoon sp. were recently described in European countries but clinical reports of the infection are rare and incomplete. This case report brings new interesting information on cytauxzoonosis expression in Europe. CASE PRESENTATION: A 9-years-old castrated European shorthair cat living in rural area of north-eastern France (Saint Sauveur, Bourgogne-Franche-Comté region), without any travel history was presented for consultation due to hyperthermia, anorexia, depression and prolonged fever that didn't respond to antibiotic therapy. The cat had outdoor access with a history of vagrancy and was adequately vaccinated (core vaccines and FeLV vaccine). During biological investigations, intraerythrocytic inclusions were observed on blood smear and were further investigated by PCR analysis and sequencing. Molecular analyses confirmed Cytauxzoon sp. infection. The cat was treated with a subcutaneous injection of imidocarb dipropionate (3.5 mg/kg). One week after treatment, the cat improved clinically, although parasitic inclusions within erythrocytes persisted, and only a mild lymphocytosis was found. Two weeks after treatment, the cat appeared in excellent health, appetite was normal and parasitemia was negative. However, one month after treatment the cat relapsed with hyperthermia, anorexia, and depression. Blood smears and PCR were once again positive. Subsequently, the cat received an additional dose of imidocarb dipropionate (3.5 mg/kg SC) and recovered rapidly without other clinical signs. Two weeks after the second imidocarb injection, the cat was hit by a car and died. CONCLUSION: This case provides the first clinical description of infection by Cytauxzoon sp. in a domestic cat in France. These findings support the fact that cytauxzoonosis should be considered in the differential diagnosis of acute febrile illness which does not respond to antibiotic in cats with outdoor access especially in areas where populations of wild felids are present.

Possibilities to control Ichthyophthirius multifiliis infestation with medicated feed in rainbow trout (Oncorhynchus mykiss) and chub (Leuciscus cephalus).

In the present study, the treatment of ichthyophthiriasis with medicated feed was investigated in rainbow trout, Oncorhynchus mykiss, and chub, Leuciscus cephalus. The anti-parasitics toltrazuril and imidocarb; the antibiotics doxycycline, erythromycin and sulphadiazine and the anti-inflammatory acetylsalicylic acid were tested. In vitro experiment revealed that all tested anti-parasitics and antibiotics were effective in killing the isolated trophonts and theronts. Minimum doses for killing 100 % of the viable trophonts and for inhibiting the development of theronts were 3 mg/L for doxycycline, 30 mg/L for erythromycin, 2 mg/L for imidocarb dipropionate, 30 mg/L for sulphadiazine and 20 mg/L for toltrazuril. Acetylsalicylic acid (40 mg/kg fish/day), doxycycline (3 and 6 mg/kg/day), erythromycin (40 mg/kg/day), imidocarb dipropionate (5.0 mg/kg/day), sulphadiazine (40 mg/kg/day), toltrazuril (20 and 40 mg/kg/day) and combinations of doxycycline and toltrazuril (3 + 20 mg/kg/day, 6 + 40 mg/kg/day) were tested as medicated feed. When administered as medicated feed, only doxycycline, toltrazuril and combinations of doxycycline and toltrazuril reduced the fish mortality and infestation level. Best results were obtained by feeding a combination of 6 mg/kg/day doxycycline and 40 mg/kg/day toltrazuril. In O. mykiss, this treatment reduced the mortality rate from 100 to 50 ± 14 % after 10 days and the infestation level from grade 4 (≥100 trophonts per skin mucus sample) to 3.5 (50-100 trophonts). In L. cephalus, the mortality rate was decreased from 100 to 39 ± 5 % and the infestation level from grades 4 to 2 (ten to 50 trophonts) after 10 days.

Molecular detection and treatment of tick-borne pathogens in domestic dogs in Khon Kaen, northeastern Thailand.

We determined the prevalence of tick-borne pathogens in domestic dogs using microscopy and polymerase chain reaction (PCR) techniques. A total of 303 EDTA blood samples were collected from domestic dogs in Khon Kaen Province, Thailand, in May 2013. Microscopic observation of Giemsa-stained smears and molecular diagnosis using conventional PCR were performed. Infected dogs were treated with imidocarb dipropionate, a combination of imidocarb dipropionate and doxycycline, or doxycycline alone. Seventy-one (23.4%) out of 303 dogs were positive for DNA of tick-borne pathogens. Of the 303 animals, 13.2% and 1.3% were positive for a single infection with Babesia spp or Ehrlichia canis, respec- tively using microscopy; whereas 19.5% and 3.0% were positive using the PCR technique. Co-infection with Babesia spp and E. canis was observed in 0.7%, and coinfection with Hepatozoon canis and E. canis in 0.3%. Infected dogs were treated with the assigned drugs, and elimination of the pathogens was demonstrated by microscopy and PCR. The results indicated that while both microscopic and PCR diagnostic techniques were useful for tick-borne pathogen detection, PCR was more effective. Imidocarb dipropionate and doxycycline were found to be effective for treatment of babesiosis and ehrlichiosis, respectively. The present study suggests that the PCR technique has high sensitivity and specificity for Babesia and Ehrlichia diagnosis as well as for detection of Babesia spp, E. canis and H. canis DNA in EDTA blood specimens.

Association between decreased blood pressure and azotaemia in canine babesiosis.

Acute tubular necrosis (ATN) was described in canine babesiosis. Hypotension is considered as one of the factors which influence the development of hypoxic renal damage. In this study hypotension defined as mean arterial pressure (MAP) < 80 mmHg was detected in 7 out of 48 dogs (14.6%) infected with Babesia canis. Lower systolic arterial pressure (SAP), diastolic arterial pressure (DAP) and MAP were detected in azotaemic dogs infected with B. canis. Statistically significant negative correlations between blood pressures (SAP, DAP and MAP) and serum creatinine and urea concentrations showed the influence of decreased blood pressure on the development of azotaemia and is probably also associated with ATN in canine babesiosis.

Repeated imidocarb treatment failure suggesting emerging resistance of Babesia canis in a new endemic area in north-eastern Germany.

Canine babesiosis has been increasingly diagnosed in various regions of Germany such as north-eastern Germany in recent years. A dog with several relapses of Babesia canis infection after treatment with imidocarb is described. A 9-year-old male Magyar Viszla with B. canis infection was referred after two treatments with imidocarb (dosage 2.1 mg/kg SC) because of lethargy, fever and pancytopenia (additional treatments with prednisolone and doxycycline). Merozoites were detected in the blood smear and imidocarb treatment was repeated. Clinical signs, pancytopenia and a positive B. canis PCR occurred after the 3rd (6 mg/kg SC), 4th (7.7 mg/kg SC) and 5th (7.5 mg/kg SC and doxycycline for 4 weeks in addition) imidocarb injection and thorough tick prevention with isoxazoline and permethrin products. 12 days after the 5th injection, the PCR was negative for the first time. The dog was again presented with fever 35 days after the 5th injection. The B. canis PCR was positive and laboratory examination revealed pancytopenia. Treatment with atovaquone/azithromycin for 18 days was performed and no further relapse occurred for 32 weeks. In the case of suspected imidocarb resistance in B. canis infection, treatment with atovaquone/azithromycin can be an alternative.

Diminazene aceturate and imidocarb dipropionate-based combination therapy for babesiosis - A new paradigm.

In the present study, the effect of a combination therapy consisting of diminazene aceturate (DA) and imidocarb dipropionate (ID) on the in vitro growth of several parasitic piroplasmids, and on Babesia microti in BALB/c mice was evaluated using a fluorescence-based SYBR Green I test. We evaluated the structural similarities between the regularly used antibabesial medications, DA and ID, and the recently found antibabesial drugs, pyronaridine tetraphosphate, atovaquone, and clofazimine, using atom pair fingerprints (APfp). The Chou-Talalay approach was used to determine the interactions between the two drugs. A Celltac MEK-6450 computerized hematology analyzer was used to detect hemolytic anemia every 96 hours in mice infected with B. microti and in those treated with either mono- or combination therapy. According to the APfp results, DA and ID have the most structural similarities (MSS). DA and ID had synergistic and additive interactions against the in vitro growth of Babesia bigemina and Babesia bovis, respectively. Low dosages of DA (6.25 mg kg-1) and ID (8.5 mg kg-1) in conjunction with each other inhibited B. microti growth by 16.5 %, 32 %, and 4.5 % more than 25 mg kg-1 DA, 6.25 mg kg-1 DA, and 8.5 mg kg-1 ID monotherapies, respectively. In the blood, kidney, heart, and lung tissues of mice treated with DA/ID, the B. microti small subunit rRNA gene was not detected. The obtained findings suggest that DA/ID could be a promising combination therapy for treating bovine babesiosis. Also, such combination may overcome the potential problems of Babesia resistance and host toxicity induced by utilizing full doses of DA and ID.

Autochthonous Babesia canis infections in 49 dogs in Germany.

BACKGROUND: Vector-borne diseases are of increasing importance in Germany. Since 2015, autochthonous cases have been increasingly documented in Berlin/Brandenburg. OBJECTIVES: Describe autochthonous Babesia canis infection in the Berlin/Brandenburg region. ANIMALS: Forty-nine dogs with autochthonous B. canis infection. METHODS: Evaluation of history, clinical signs, laboratory abnormalities, treatment, and outcome. RESULTS: Dogs were presented between March and August (9) and September and January (40) in the years 2015-2021. Historical and clinical findings were lethargy (100%), pale mucous membranes (63%), fever (50%), and pigmenturia (52%). Common clinicopathological findings were thrombocytopenia (100%), anemia (85%), intravascular hemolysis (52%), pancytopenia (41%), and systemic inflammatory response syndrome (SIRS; 37%). Babesia detection was based on blood smear evaluation (n = 40) and PCR targeting the 18S rRNA gene of piroplasms (n = 49). Sequencing indicated 99.47% to 100% identity to B. canis sequences from GenBank. All dogs were treated with imidocarb (2.4-6.3 mg/kg; median, 5 mg/kg); 8 dogs received 1, 35 received 2, and 1 dog each received 3, 4, or 5 injections, respectively. Continued PCR-positive results were detected in 7 dogs after the 1st, in 5 after the 2nd, in 2 after the 3rd, and in 1 28 days after the 4th injection. Four dogs were euthanized and 3 dogs died. CONCLUSIONS AND CLINICAL IMPORTANCE: Autochthonous B. canis infections in Berlin/Brandenburg were associated with severe clinicopathological changes, SIRS, and multiorgan involvement. Testing by PCR during and after treatment is advisable to monitor treatment success. Screening of blood donors in high-risk areas and year-round tick protection is strongly recommended.

Severe anemia caused by babesiosis in a maned wolf (Chrysocyon brachyurus).

An 8-yr-old, captive, spayed, female maned wolf (Chrysocyon brachyurus) developed progressive lethargy and weakness over a 24-hr period. Clinical signs included vomiting, recumbency, horizontal nystagmus, possible blindness, pale icteric mucus membranes, and port-wine colored urine. A complete blood cell count revealed severe anemia (packed cell volume [PCV], 6%) and intraerythrocytic piroplasms consistent with a Babesia species. Polymerase chain reaction testing later confirmed babesiosis. The wolf was treated with imidocarb dipropionate, antibiotics, and fluid therapy. A whole-blood transfusion from a sibling maned wolf also was performed. Despite aggressive treatment, the wolf failed to improve and was euthanized. To the authors' knowledge, this is the first documented case of babesiosis in a captive maned wolf in North America. Surveillance of infectious diseases in captive and wild maned wolf populations should be expanded to include screening for Babesia species. Tick control also should be implemented to prevent and decrease transmission of the disease to this endangered species.

Efficacy of long-acting oxytetracycline and imidocarb dipropionate for the chemosterilization of Anaplasma marginale in experimentally infected carrier cattle in Argentina.

The expansion of anaplasmosis to non-endemic areas in Argentina has created the need for specific treatments to eliminate Anaplasma marginale from carriers. The most recent studies have failed to chemosterilize A. marginale infections. In this work, we compare the efficacy of long-acting oxytetracycline (OTC) and imidocarb dipropionate (IMD) to chemosterilize the A. marginale infection. For this purpose, twenty steers were randomly clustered into two groups of ten animals each 78 days after A. marginale experimental infection (day 0). Cattle from group 1 (G1) were treated with three doses of 20 mg kg-1 of OTC (Terramycin® LA, 200 mg/ml) 7 days apart by intramuscular injection. Cattle from G2 were treated with two doses of 5 mg kg-1 of IMD (Imizol®, 120 mg/ml) 14 days apart by intramuscular injection. The efficacy of sterilizing treatments was evaluated by detection of DNA by nested PCR, anti-MSP5 antibodies by ELISA and by inoculation of splenectomized calves with blood from the steers 104 days post-treatment (dpt). The results showed 50% efficacy of the OTC treatment to chemosterilize persistent A. marginale infections in cattle and the failure of the IMD treatment under the evaluated conditions. The persistence of specific antibody levels in the sterilized animals (56 dpt) was shorter than the period of DNA detection. The ELISA was the test of choice to confirm the sterilizing outcome after 60 dpt. In spite of its limitations, the sterilization of A. marginale carrier status using OTC, could be useful for high-value bovines in non-endemic areas.

[Theileria equi-infection in 2 German horses returning from a trail ride in southern France]. / Theileria equi-Infektion bei 2 Pferden nach einem Wanderritt in Südfrankreich.

Equine piroplasmosis, caused by the protozoan pathogens Theileria equi or Babesia caballi, is endemic in many (sub-)tropical regions worldwide, including Southern Europe. In Germany, it mainly plays a role as an imported parasitosis. Nevetherless, tick species with known vector potential for equine piroplasms occur in Germany. Thus, a long-term establishment of autochthonous cycles after introduction via infected equids or ticks cannot be excluded. In the present case, 2 horses developed clinical T. equi infection after returning from a trail riding trip to southern France. During treatment, a feeding tick was discovered on one of the horses, which was identified as a Rhipicephalus bursa specimen. This tick species cannot establish in Germany under the current climatic conditions; nevertheless, this discovery illustrates the potential of introducing infected ticks to Germany. The 2 horses were treated with imidocarb (3 mg/kg i. m.; 3 treatments in 3-day intervals) and no adverse drug effects were recorded. To prevent establishment of autochthonous infection cycles in Germany as well as a chronic manifestation of piroplasmosis, treatment should aim at eliminating the parasites despite possible adverse effects of imidocarb.

Ehrlichia canis in dogs experimentally infected, treated, and then immune suppressed during the acute or subclinical phases.

BACKGROUND: Concerns for recrudescence of Ehrlichia canis infection arise when immunosuppressive drugs are used to treat immune-mediated diseases in dogs previously infected with E. canis. OBJECTIVES: Determine whether administration of prednisolone and cyclosporine would reactivate E. canis infection in dogs previously treated with doxycycline during the acute or subclinical phases. ANIMALS: Seven beagles previously experimentally infected with E. canis and administered doxycycline for 4 weeks were included. Three of the 7 dogs were incidentally concurrently infected with Anaplasma platys and Babesia vogeli and were administered 2 doses of imidocarb 2 weeks apart before enrollment in the current study. METHODS: Experimental study. Each dog was administered prednisolone and cyclosporine for 6 weeks. Clinical signs, complete blood cell count (CBC), polymerase chain reaction (PCR) assays for E. canis, A. platys, and B. vogeli DNA in blood, E. canis indirect fluorescent antibodies (IFA) titers, and flow cytometry for antiplatelet antibodies were monitored. RESULTS: All dogs completed the immunosuppressive protocol. No evidence for recrudescence of E. canis, A. platys, or B. vogeli were detected based on clinical signs or results of CBC, PCR, IFA, and flow cytometry for antiplatelet antibodies. E. canis IFA titers were negative in 5/7 dogs at the end of immunosuppressive protocol and were negative 6 months after the protocol in 5/5 dogs available for testing. CONCLUSIONS AND CLINICAL IMPORTANCE: Dogs administered with a 4-week course of doxycycline with or without imidocarb failed to show evidence of activation of E. canis infection after administration of a commonly used immune suppressive protocol.

Imidocarb dipropionate clears persistent Babesia caballi infection with elimination of transmission potential.

Antimicrobial treatment of persistent infection to eliminate transmission risk represents a specific challenge requiring compelling evidence of complete pathogen clearance. The limited repertoire of antimicrobial agents targeted at protozoal parasites magnifies this challenge. Using Babesia caballi as both a model and a specific apicomplexan pathogen for which evidence of the elimination of transmission risk is required for international animal movement, we tested whether a high-dose regimen of imidocarb dipropionate cleared infection from persistently infected asymptomatic horses and/or eliminated transmission risk. Clearance with elimination of transmission risk was supported by the following four specific lines of evidence: (i) inability to detect parasites by quantitative PCR and nested PCR amplification, (ii) conversion from seropositive to seronegative status, (iii) inability to transmit infection by direct inoculation of blood into susceptible recipient horses, and (iv) inability to transmit infection by ticks acquisition fed on the treated horses and subsequently transmission fed on susceptible horses. In contrast, untreated horses remained infected and capable of transmitting B. caballi using the same criteria. These findings establish that imidocarb dipropionate treatment clears B. caballi infection with confirmation of lack of transmission risk either by direct blood transfer or a high tick burden. Importantly, the treated horses revert to seronegative status according to the international standard for serologic testing and would be permitted to move between countries where the pathogen is endemic and countries that are free of the pathogen.

Detection of a large unnamed Babesia piroplasm originally identified in dogs in North Carolina in a dog with no history of travel to that state.

CASE DESCRIPTION: A 12-year-old 46-kg (101.2-lb) sexually intact male Labrador Retriever was evaluated because of lymphadenomegaly. The dog resided in Texas, and its travel history included many southeastern and eastern shore states but not North Carolina. CLINICAL FINDINGS: Following evaluation of the dog, a diagnosis of stage IVa intermediate- to large-cell lymphoma was made. A cyclophosphamide-hydroxydaunorubicin (doxorubicin)-vincristine-prednisone chemotherapy protocol was initiated. One week after the first chemotherapeutic treatment, a routine blood smear evaluation revealed single and paired intraerythrocytic large piroplasms that resembled Babesia canis. Via molecular testing, the organism was identified as a Babesia sp that had been detected previously in dogs in North Carolina. TREATMENT AND OUTCOME: The dog was administered imidocarb diproprionate (7 mg/kg [3.2 mg/lb], IM) on 2 occasions (3-week interval). At 1, 4, 15, and 50 weeks after the second treatment, blood samples were analyzed specifically for the North Carolina Babesia sp via PCR assay; the result of each assay was positive. CLINICAL RELEVANCE: Because of the morphologic similarity of the large piroplasm detected in dogs in North Carolina to B canis, molecular testing of large piroplasms detected in dogs is needed to definitively identify the infective Babesia sp. In the dog of this report, the infection was not eliminated following treatment with imidocarb diproprionate, which may have been a result of the immunocompromised state of the dog or the drug's ineffectiveness against this parasite. If imidocarb diproprionate is ineffective against the North Carolina Babesia sp, treated dogs may act as reservoirs of infection.

Alterations in some blood coagulation parameters in naturally occurring cases of canine babesiosis.

Changes in coagulation parameters were studied in dogs naturally infected with Babesia canis canis (n = 30), and haemostasis was evaluated and compared to values obtained from healthy dogs (n = 29). To date, there have not been any studies examining the dynamics of thrombin-antithrombin complex formation in cases of canine babesiosis. Coagulation parameters evaluated before (day 0) and on days 1, 2, and 3 after treatment with imidocarb (6 mg/kg inj. s.c.) included the determination of platelet counts, the formation of thrombin-antithrombin complexes (TAT), prothrombin time (PT), activated partial thromboplastin time (APTT) and antithrombin III (AT III) activity. TAT complexes were significantly elevated in animals with babesiosis on days 0 and 2 (mean 49.7 and 87.7 microg/L vs. control, 7.2 microg/L). AT III activity was significantly decreased at all time-points examined. There were no differences in PT. On days 2 and 3 the APTT was significantly shortened in the infected dogs when compared to control animals (means of 21.3 and 19.2 s vs. control, 30.0 s). Our analysis demonstrated that infected dogs had significant thrombocytopenia during the course of the study (mean day 0 - 29 x 10(9) /L, day 1 - 48 x 10(9) /L, day 2 - 47 x 10(9) /L and day 3 - 87 x 10(9) /L, vs. control -259 x 10(9) /L). These data suggest that babesiosis in dogs compromise primary and secondary haemostasis and that induction of disseminated intravascular coagulation (DIC) occurs in canine babesiosis.

Putative clinical piroplasmosis in a Burchell's zebra (Equus quagga burchelli).

A 10-year-old tame zebra gelding was presented after suffering from lethargy, nervousness, reported anaemia and icterus as well as a decreased appetite. These symptoms were seen over some months, with changing severity. The animal was immobilised, treated, and blood specimens were submitted for haematology and biochemistry. This report describes molecular characterisation of Theileria equi recovered from this animal, as well as the clinical findings, treatment and historical relevance of piroplasmosis in zebra in southern Africa.

Diagnosis and treatment of Babesia odocoilei in captive reindeer (Rangifer tarandus tarandus) and recognition of three novel host species.

Two captive reindeer (Rangifer tarandus tarandus) at a New York zoological institution were diagnosed with Babesia odocoilei. Clinical signs consistent with acute babesiosis included fever, hemoglobinuria, and hemolytic anemia. Both episodes were precipitated by stressful events that may have compromised their immunocompetence. The diagnosis was confirmed by visualization of intraerythrocytic parasites on stained blood smears, polymerase chain reaction, and speciation of the Babesia by sequencing a hypervariable region of the 18S rRNA gene. One reindeer died with gross and histopathologic lesions, including pigmentary nephrosis with severe acute tubular degeneration and necrosis secondary to intravascular hemolysis. A second reindeer was successfully treated with supportive care and an antiprotozoal, imidocarb dipropionate (Imizol, 12%, Schering-Plough Animal Health, Union, New Jersey 07083, USA) at 3 mg/kg s.c. or i.m. s.i.d. on days 1, 2, 6, 9, and 21. Two other reindeer in the exhibit tested negative for Babesia by polymerase chain reaction but were treated with imidocarb dipropionate as prophylaxis while final testing results were pending. Additionally, B. odocoilei was identified in three novel asymptomatic host species within the collection: yak (Bos grunniens), muntjac (Muntiacus reevesi), and markhor goat (Caprafalconeri). Due to the high morbidity and mortality associated with acute babesiosis, captive reindeer should receive tick prevention, be tested for subclinical infections in endemic areas, and receive aggressive treatment for acute infections when clinical babesiosis is suspected.

The clinical course of babesiosis in 76 dogs infected with protozoan parasites Babesia canis canis.

The aim of the study was to trace the clinical course of babesiosis in 76 dogs infected with Babesia canis protozoa and to assess the usefulness of PCR method in the routine diagnosis of the disease. The investigations were conducted in three successive seasons of the biological activity of ticks on dogs displaying possible clinical signs of babesiosis, the latter assigned individual numbers from 001 to 076. All the animals underwent routine clinical examinations and blood was collected for haematological, biochemical, parasitological and molecular tests for babesiosis. The most frequent clinical signs observed in the course of the disease were changes in urine colour and xanthosis or paleness of mucous membranes, whereas in the haematological and biochemical examinations, the most frequent laboratory findings were thrombocytopenia, leucopoenia, anaemia and an increase in levels of urea and bilirubin. In all blood smears stained with the May-Grunwald and Giemsa methods, from the 76 dogs, the presence of Babesia canis protozoa was observed in erythrocytes, and their DNA was detected in 69 blood samples by means of PCR technique. The course of the disease and the results of molecular examinations suggested the haemolytic form of babesiosis. The previous genetic analysis of isolates of Babesia canis canis from the eastern areas of Poland helped to distinguish two specific groups, A and B, within the species (Adaszek and Winiarczyk 2008a). The present study revealed a certain interrelation between the intensification of thrombocytopenia and the fact that protozoa belong to either group A or B. The mean number of thrombocytes in dogs infected with protozoa from group A was 61.11 thousand/mm3, whereas the mean number of thrombocytes in the blood of dogs infected with protozoa from group B was 27.47 thousand/mm3. A strong correlation was also observed between the low level of thrombocytes and the increase in the internal body temperature (p = 0.02), accelerated pulse rate (p = 0.01) and discoloration of urine (p = 0.04). As a result of the treatment of dogs with imidocarb, recovery was observed in 73 out of the 76 dogs brought to the clinic.

Fatal acute babesiosis associated with Babesia venatorum infection (Babesia sp. EU1) in a captive reindeer calf in Switzerland.

Babesia venatorum was isolated from a captive reindeer calf in Switzerland. The clinical signs consistent with acute babesiosis included hemolytic anemia and hemoglobinuria. The diagnosis was made based on visualization of intraerythrocytic parasites in the stained blood smears and confirmed by PCR analysis of the 18S rRNA gene, with subsequent species identification within Babesia confirmed by sequencing. The reindeer calf was initially treated with supportive care and an antiprotozoal drug (imidocarb dipropionate) but died a few days after hospitalization. Babesia venatorum is also known as Babesia sp. EU1 and can infect different mammalian species, including humans. The current case report aims to increase awareness among veterinarians and reindeer owners about the presence and the associated risk of this zoonotic pathogen. Considering the high morbidity and possible mortality associated with acute babesiosis, captive reindeer should receive tick prevention measures and be tested for subclinical infections in endemic area.