The fox was first treated with ivermectin for the mange and with amoxicillin combined with clavulanic acid for the suspected urinary infection. Instead of recovering, the animal worsened, growing weaker and paler. Only a month after admission, in September 2024, did the clinical team suspect a blood parasite and run a complete blood count, serum biochemistry, urinalysis and faecal examinations. The haematology revealed a regenerative anaemia marked by anisocytosis, polychromasia, Howell-Jolly bodies, rouleaux formation and increased reticulocytes, alongside leukocytosis with neutrophilia, a left shift and lymphocytosis. Biochemistry showed elevated alanine aminotransferase and alkaline phosphatase, increased total bilirubin, reduced total proteins with hypoalbuminemia, raised cholesterol, amylase and lipase, and a reduced sodium-to-potassium ratio. Urinalysis detected haemoglobinuria at 50 micromoles per litre together with haematuria. A blood smear then delivered the decisive clue: intraerythrocytic inclusions morphologically consistent with small piroplasmids.

Molecular confirmation followed quickly. DNA extracted from whole blood was analysed by conventional PCR targeting the 18S ribosomal DNA gene, and sequencing of the roughly 510-base-pair amplicon, compared against GenBank using BLAST, showed 99.6 per cent nucleotide identity with Babesia banethi. Crucially, the fox screened negative for antibodies against Leishmania infantum, Ehrlichia canis and Anaplasma phagocytophilum/platys, negative for Dirofilaria immitis antigen and negative for Rickettsia species DNA, ruling out common co-infections. B. banethi is a recently described small Babesia species, genetically distinct from the dominant fox piroplasm Babesia vulpes. While B. vulpes belongs to clade I of the piroplasmids, within the B. microti-like group, phylogenetic analyses place B. banethi in clade III, the so-called Western group, well apart from the large Babesia sensu stricto species such as B. canis, B. vogeli and B. gibsoni that infect domestic and wild carnivores. The species had previously been detected in foxes from Israel and Iraq, and its competent tick vector remains unknown, although parasite DNA has been found in Ixodes kaiseri ticks collected from infected animals.

With no guidelines for managing babesiosis in wildlife, the veterinary team turned to standard canine practice. At the end of September 2024 the fox received imidocarb dipropionate at 6.6 milligrams per kilogram subcutaneously, in two doses spaced 28 days apart, the most routinely used regimen for canine babesiosis. Fifteen days after the first dose the fox showed genuine clinical and haematological improvement, with only a persistent lymphocytosis marring the blood picture. Yet the parasitological follow-up told a different story: blood smears and PCR both remained positive, indicating persistent parasitaemia. Two weeks after the second imidocarb dose the animal continued to improve clinically and the parasitaemia and lymphocytosis had disappeared on smear, but PCR was still positive for B. banethi DNA. The drug had suppressed the disease without eradicating the parasite.

The relapse came roughly three months after the second imidocarb dose, in February 2025. The fox began showing mild lethargy and pale mucous membranes again, and laboratory work confirmed the return of anaemia, lymphocytosis, thrombocytosis and microscopically visible parasitaemia. Subsequent examinations at follow-up points roughly 255 and 281 days after the initial treatment recorded a consistent decline in the animal’s condition, with B. banethi detectable by both blood smear and PCR. Almost a full year after the original diagnosis, the team switched strategy. They initiated a combination of atovaquone and proguanil hydrochloride, the antimalarial sold as Malarone, at 20 milligrams per kilogram twice daily by mouth, together with azithromycin at 10 milligrams per kilogram once daily subcutaneously, for ten days, following the general regimen used against small-form Babesia infections in dogs.

The response was striking. At follow-up examinations roughly 331 and 352 days after the initial diagnosis, the fox showed marked clinical and haematological recovery, with a negative blood smear, although PCR positivity persisted. About two months after the end of the Malarone course the animal remained parasitaemic by PCR but was clinically stable, albeit with lingering laboratory abnormalities including lymphocytosis and hypergammaglobulinemia; a repeat SNAP Leish 4Dx test again came back negative. In January 2026 the fox received a second course of the atovaquone-proguanil and azithromycin combination, and one month after that therapy ended the animal was clinically stable, with only persistent lymphocytosis and elevated liver enzymes remaining, and for the first time tested both cytologically and molecularly negative for B. banethi.

The longitudinal record offers several lessons about how small Babesia parasites behave in their hosts and how treatment should be judged. PCR positivity soon after drug administration, the authors note, should not be read as treatment failure, because it may reflect the temporary persistence of parasite DNA rather than active infection; molecular testing is generally recommended no earlier than two months after completing therapy for small Babesia species. When PCR positivity lingers beyond three months, however, clinicians must consider sequestration of the parasite in target organs such as the spleen and in circulating erythrocytes, with the potential for reactivation after immunosuppressive events. For rescued wildlife destined for release, the authors argue that the primary indicator of therapeutic success should be clinical stabilisation and cytological negativity rather than complete clearance of parasite DNA, since foxes should spend as little time as possible in rehabilitation facilities.

The haematological details also distinguish this infection from better-known canine babesiosis. The mild thrombocytosis observed during the acute and relapse phases likely reflects rebound stimulation of the bone marrow in response to haemolytic anaemia, a pattern that contrasts with the thrombocytopenia commonly described in dogs infected with B. vulpes. Most other abnormalities, including anaemia, elevated liver enzymes and increased total bilirubin, mirrored those reported in dogs with small Babesia infections. Recurrent lymphocytosis, hypergammaglobulinemia and hypoalbuminemia pointed to chronic immune stimulation, while the raised ALT and ALP activities with hyperbilirubinaemia and brown urine probably reflected hepatocellular lesions secondary to haemolysis, anoxia and systemic inflammation. No hepatoprotective or additional supportive therapy was given, a limitation the authors acknowledge when interpreting the persistent liver enzyme changes. The team also stresses the complementary value of blood smear examination and molecular testing: smears identify acute, chronic and relapsing phases, while PCR confirms the species and documents persistent infection.

Why did this fox become visibly ill at all, when foxes are generally regarded as asymptomatic reservoirs of piroplasm infections? The severe sarcoptic mange and poor nutritional state may have impaired the animal’s immune response and prolonged the clinical course, but the absence of co-infections with other vector-borne pathogens or gastrointestinal parasites, together with the repeated recurrence of signs and parasitaemia, supports a primary pathogenic role for B. banethi itself. That possibility carries a broader warning. If a parasite can establish a persistent, treatment-refractory infection in a species considered relatively resistant to vector-borne pathogens, the clinical impact on more susceptible hosts, particularly domestic dogs sharing the same environment and exposed to the same potentially competent ticks, could be considerable. The case also highlights practical realities of wildlife medicine: follow-up intervals were stretched to limit repeated handling of the animal, which may have delayed detection of parasitaemia recurrences, and an earlier switch to atovaquone-proguanil with azithromycin as a first-line protocol might have shortened the disease course. Given the off-label use of atovaquone, the authors add that efficacy must be balanced against the risk of fostering antimicrobial resistance, a concern already documented for B. gibsoni in dogs. All clinical assumptions, they caution, now require validation in a larger series of B. banethi cases.

Subject of Research: Clinical course and treatment of Babesia banethi infection in a red fox

Article Title: Clinical Case and Follow‐Up of Babesia banethi Infection in a Red Fox

Article References: Clinical Case and Follow‐Up of Babesia banethi Infection in a Red Fox. (n.d.). https://doi.org/10.1002/vms3.71184

Image Credits: AI Generated

DOI: 10.1002/vms3.71184

Keywords: Babesia banethi, babesiosis, red fox, Vulpes vulpes, tick-borne pathogens, piroplasmids, wildlife medicine, imidocarb dipropionate, atovaquone-proguanil, azithromycin, regenerative anaemia, Italy

Babesia microti belongs to a group of apicomplexan parasites transmitted to animals and humans through the bite of infected hard ticks. In North America, the primary vector is Ixodes scapularis, while Ixodes ricinus serves this role in Europe and Ixodes persulcatus in Asia. Humans are incidental hosts, and human babesiosis has been documented across the United States, Europe and Asia. The parasite primarily infects small rodents, particularly the white-footed mouse, which acts as its principal natural reservoir, although infections have occasionally been reported in domestic animals such as cats and dogs. Because transovarial transmission in ixodid ticks has not been reported, the parasite depends on an enzootic cycle linking ticks and small mammals, with rodents serving as a continuous source of infection for feeding ticks. Beyond tick bites, humans can acquire the parasite through contaminated blood transfusion, organ transplantation and congenital transmission from an infected mother to her foetus.

In most infected people the disease remains asymptomatic, but symptoms typically emerge after an incubation period of one to four weeks and include chills, fever, sweating, fatigue and haemolytic anaemia resulting from the repeated rupture of infected red blood cells. The preferred treatment in humans combines the antiprotozoal drug atovaquone with the antibiotic azithromycin. Rodents are central to the epidemiology of this disease because they are widely distributed, adapt readily to diverse habitats and often live in close proximity to humans. Approximately 200 rodent species are known to harbour more than 60 zoonotic diseases caused by bacteria, viruses and parasites. Pakistan alone hosts 43 documented rodent species, ranging from common mice and rats to porcupines, desert gerbils and giant flying squirrels, yet these animals have rarely been screened for pathogens, with only a handful of reports concerning Toxoplasma gondii and single reports of Hepatozoon and Lankesterella species in local rats and mice.

To address this gap, the research team trapped wild rodents between October 2023 and September 2024 from four districts in Punjab, namely Rajanpur, Dera Ghazi Khan, Multan and Sargodha, and three districts in Khyber Pakhtunkhwa, namely Upper Dir, Mardan and Buner. Using the Thrusfield formula for prevalence studies with an expected prevalence of 10 percent, a 95 percent confidence level and 5 percent precision, they calculated a minimum required sample of 138 animals but nearly doubled this by capturing 284 rodents to improve the reliability of their estimates. Live traps baited with fruit, sweets and biscuits, along with glue traps, were deployed opportunistically at agricultural fields, houses, storage sites and commercial shops. Captured animals were identified to species using standard taxonomic keys, combed for ectoparasites, euthanized under deep isoflurane anaesthesia and sampled by direct cardiac puncture. Blood was collected into tubes containing EDTA anticoagulant, with complete blood counts performed the same day and remaining samples frozen for molecular analysis. All procedures were approved by the ethical review committee of Bahauddin Zakariya University in Multan and conducted in accordance with ARRIVE guidelines.

The trapped animals comprised three species: 137 black rats (Rattus rattus), 80 brown rats (Rattus norvegicus) and 67 house mice (Mus musculus), with males making up 54 percent of the sample. Genomic DNA was extracted from blood using a commercial purification kit and screened by polymerase chain reaction targeting a species-specific region of the parasite’s 18S ribosomal RNA gene, following primers originally described by Persing and colleagues. The assay amplified a 238-base-pair fragment in 35 of the 284 samples, an overall prevalence of 12 percent. Positive controls came from a previously confirmed B. microti-positive dog blood sample from the same laboratory, and distilled water served as the negative control in every run. Representative amplicons from two brown rats, one black rat and one house mouse were confirmed by bidirectional sequencing, submitted to GenBank under accession numbers PQ538588 through PQ538591 and included in phylogenetic analysis.

Prevalence varied modestly among species, with 16 percent of brown rats, 13 percent of black rats and 6 percent of house mice testing positive, though a chi-square comparison found these differences statistically non-significant. District-level patterns were more striking despite lacking statistical significance in binary logistic regression models that used Rajanpur as the reference district. Among brown rats, prevalence peaked at 42 percent in Sargodha; among black rats, at 28 percent in Mardan; and among house mice, at 25 percent in Buner. Notably, every infected house mouse came from Khyber Pakhtunkhwa, while none of the mice from Punjab tested positive, whereas both rat species carried infections in both provinces. Fisher’s exact tests showed that infection was not confined to either sex in any of the three species, and no ticks were detected on any of the trapped rodents, a finding the authors attribute to the temporary nature of tick attachment during blood feeding, seasonal variation in tick activity, host grooming behaviour and the timing of trapping.

Phylogenetic analysis of the partial 18S rRNA sequences, performed in MEGA X using the Maximum Likelihood method with a Kimura 2-parameter model and a discrete gamma distribution, placed all four Pakistani rodent sequences in a single cluster. The sequences showed 100 percent identity with B. microti previously detected in dog and cat blood samples from Pakistan, and 99.49 percent identity with isolates from house mice in Japan and France and from the grey red-backed vole Myodes rufocanus in Russia. They were clearly distinct from 18S rRNA sequences of Babesia leo, Babesia felis, Babesia vulpes and Babesia rodhaini detected in felines and rodents across South Africa, Mozambique, the Czech Republic and Japan, with Cytauxzoon brasiliensis serving as the outgroup. This close relationship between rodent and domestic animal isolates within Pakistan suggests that the same parasite lineage circulates across multiple host species in the country, reinforcing the value of the 18S rRNA gene as a conserved and widely validated marker for sensitive detection, even though its resolution for fine-scale genotypic differentiation remains limited.

The study also documented measurable costs of infection in the rodent hosts. Infected house mice weighed significantly less than uninfected mice, although the observational design prevents any conclusion about causation, since age, nutrition, reproductive condition and concurrent infections could all contribute. In black rats, infection was associated with significantly reduced total white blood cell counts, lymphocyte counts and red blood cell counts. In brown rats, lymphocytes and mean corpuscular volume were significantly decreased while mean corpuscular haemoglobin concentration was significantly elevated. These changes align with the established biology of Babesia infection, in which sporozoites injected by an infected tick invade erythrocytes and undergo asexual replication, with the repeated rupture of infected red blood cells during merozoite release producing haemolytic anaemia. The authors caution that multiple haematological parameters were compared with separate t-tests without adjustment for multiple comparisons, so significant findings should be interpreted carefully and validated in larger samples.

Compared with surveys elsewhere, the Pakistani prevalence of 12 percent sits near the middle of a wide global range, from 0.6 percent in social voles in Turkey and 1.4 percent in Irish wood mice and bank voles to 22 percent among small rodents in Chile, with intermediate figures reported from China, South Korea, Turkey, Lithuania and mainland Southeast Asia. Such variation likely reflects differences in climate, geography, sampling season, host age and immunity, and tick density. The study had limitations: trapping was opportunistic and trap numbers varied by site, potential risk factors such as age, body condition, habitat and season were not evaluated, and the absence of blood smear examination means detection relied entirely on molecular methods, so the reported prevalence reflects molecular detection rather than confirmed active parasitemia and cannot distinguish low-level from heavy infections. No human babesiosis cases have been reported in Pakistan, but limited awareness, inadequate diagnostic facilities and underrecognition of tick-borne diseases, particularly in rural areas, mean undiagnosed infections remain a real possibility. The researchers call for integrated surveillance combining molecular diagnostics with microscopy, investigation of rodent-associated ticks for B. microti and other pathogens, and broader epidemiological studies to evaluate the zoonotic risk that these infected rodents may pose to people and animals across the region.

Subject of Research: First molecular detection and genetic characterization of the tick-borne zoonotic parasite Babesia microti in wild rodents in Pakistan

Article Title: Molecular Detection, Epidemiology and Genetic Diversity of Babesia microti Infecting Wild Rodents

Article References: Munir, H. M., Naeem, M., Ijaz, M., Ullah, S., Usman, M., Nasir, Z. U. R., Khan, A., Almaary, K. S., Bouallegue, A., Mengistie, A. A., & Iqbal, F. (2026). Molecular Detection, Epidemiology and Genetic Diversity of Babesia microti Infecting Wild Rodents. Veterinary Medicine and Science, 12(5), Article e71196. https://doi.org/10.1002/vms3.71196

Image Credits: AI Generated

DOI: 10.1002/vms3.71196

Keywords: Babesia microti, babesiosis, rodents, Pakistan, tick-borne disease, zoonosis, 18S rRNA, phylogenetics, haemolytic anaemia, Rattus norvegicus, Rattus rattus, Mus musculus