Research Article

A Case Report of Equine Piroplasmosis in a Postpartum Indigenous Pony Mare in Malaysia

Nur Zul Izzati Mohd Rajdi1, Rumaizi Shaari1*, Mimi Armiladiana Mohamad1, Tan Li Peng2, Mohd Farhan Hanif Reduan3, Mohammad Sabri Abdul Rahman1, Mohammad Dauda Goni1

1Equine Health and Medicine Research Group, Department of Veterinary Clinical Studies, Faculty of Veterinary Medicine, Universiti Malaysia Kelantan, 16100 Kota Bharu, Kelantan; 2Department of Veterinary Preclinical Studies, Faculty of Veterinary Medicine, Universiti Malaysia Kelantan, 16100 Kota Bharu, Kelantan; 3Department of Veterinary Paraclinical Studies, Faculty of Veterinary Medicine, Universiti Malaysia Kelantan, 16100 Kota Bharu, Kelantan.

Received | September 13, 2020; Accepted | May 17, 2025; Published | July 31, 2025

*Correspondence | Rumaizi Shaari, Equine Health and Medicine Research Group, Department of Veterinary Clinical Studies, Faculty of Veterinary Medicine, Universiti Malaysia Kelantan, 16100 Kota Bharu, Kelantan; Email: [email protected]

Citation | Mohd Rajdi NZI, Shaari R, Mohamad MA, Tan LP, Reduan MFH, Rahman MSA, Goni MD (2025). A case report of equine piroplasmosis in a postpartum indigenous pony mare in Malaysia. Adv. Anim. Vet. Sci. 13(8): 1834-1843.

DOI | https://dx.doi.org/10.17582/journal.aavs/2025/13.8.1834.1843

ISSN (Online) | 2307-8316; ISSN (Print) | 2309-3331

Copyright: 2025 by the authors. Licensee ResearchersLinks Ltd, England, UK.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

INTRODUCTION

Equine Piroplasmosis (EP) is a tick-borne disease caused by the protozoa from the phylum of Apicomplexa, class of Aconoidasida, and order of Piroplasmida. It is listed as one of the notifiable diseases by the World Organization for Animal Health (OIE). It consists of two main families that are responsible for EP which are Babesiidae and Theileridae. The most common species infecting horses are Babesia caballi and Theileria equi. They are transmitted by 14 Ixodid tick species, including Hyalomma, Rhipicephalus, Dermacentor, Amblyomma, Haemaphysalis and Ixodes (Onyiche et al., 2019; Romiti et al., 2020; Scoles and Ueti, 2013; Rocafort-Ferrer et al., 2022).

Tick transmission can occur via intrastadial, transstadial, or transovarial, with transovarial transmission being most common in B. caballi infection (Tamzali, 2013). The parasite also can be transmitted through iatrogenic transmission where it occurs when an uninfected blood is accidentally mixed with infected blood through contaminated needles or blood transfusions (Wise et al., 2013). Other than that, transplacental transmission of T. equi from a carrier mare to fetuses had been reported, causing neonatal piroplasmosis or abortion in horses and mules (Allsopp et al., 2007; Chhabra et al., 2012; Georges et al., 2011; Sudan et al., 2015; Sant et al., 2016; Françoso et al., 2018). Transplacental transmission has been conclusively demonstrated for T. equi, supported by serological, microscopic, and molecular findings in foals across several countries, whereas evidence for B. caballi remains inconclusive despite occasional DNA detection (Phipps and Otter, 2004; Allsopp et al., 2007; Kumar et al., 2008; Georges et al., 2011; Chhabra et al., 2012; Sudan et al., 2015; Sant et al., 2016; de Sousa et al., 2017; Oliveira et al., 2019; Françoso et al., 2018; Bartolomé del Pino et al., 2023; Hermans et al., 2025). Additionally, the recent identification of T. equi DNA in colostrum suggested a possible alternative transmission route beyond placental transfer (Hermans et al., 2025).

Equine Piroplasmosis (EP) mostly infects equidae family groups such as horses, zebras, donkeys and mules (Rothschild, 2013). Other than that, current epidemiological studies also revealed infection of T. equi and B. caballi in camels (Qablan et al., 2012; Jasim et al., 2015; Bahrami et al., 2017), dogs (Rosa et al., 2014; Fritz, 2010; Hegab et al., 2023), tapir (Da Silveira et al., 2017; de Souza Gonçalves et al., 2020) and subclinically detected in Black Rhinoceros (Zimmermann et al., 2021).The capacity of these protozoa to infect multiple host species is of particular concern in Malaysia, where multispecies rearing is a common practice among horse owners. Additionally, the proximity of equine facilities to forested areas increases the risk of exposure to wildlife that may serve as reservoirs for vector-borne transmission (Naama et al., 2021).

Equine piroplasmosis imposes considerable economic burdens on horse owners due to increased treatment expenses, diminished reproductive efficiency, and pregnancy losses. Infections with T. equi of greater concern, as affected horses can remain lifelong carriers, thereby restricting their participation in international events where seropositive animals are prohibited (Aziz and Al-Barwary, 2018). Clinically, the disease may manifest in peracute, acute, subacute, or chronic forms, with the severity and presentation varying depending on the progression of infection. Peracute EP is rare and causes sudden death in adults and neonatal foals, although some infected foals are born without any sign of illness (Allsopp et al., 2007). Acute EP has a poorer prognosis and is more severe in stress-induced and immunocompromised horses. The symptoms include fatigue, anorexia, peripheral edema, high fevers, tachypnea, tachycardia, petechial, and ecchymosis hemorrhage of mucous membrane (Tamzali, 2013). In severe cases, the digestive, hepato-biliary and cardiovascular system may get involved, leading to signs such as colic, impaction, constipation, diarrhea (Wise et al., 2013; Tamzali, 2013), hemoglobinuria, bilirubinuria, jaundice (Rothschild, 2013) weak and arrhythmic arterial pulse and diastolic murmur (Diana et al., 2007; Aida et al., 2023).Subacute EP infection has a similar presentation but may be less severe (Zobba et al., 2008). Chronic EP infection is difficult to suspect due to non-specific signs, such as loss of appetite, performance reduction, weight loss, and poor body condition (Rothschild, 2013).

Equine piroplasmosis (EP) has been reported in 41 countries, predominantly in South and North America, South and West Africa, Europe, and Asia (WAHIS, 2025). The prevalence of EP varies considerably across regions, corresponding closely to the distribution of competent tick vectors (Onyiche et al., 2019). In Asia, low molecular prevalence rates were observed in Indonesia, with 8.5% for T. equi and 2.1% for B. caballi (Nugraha et al., 2018), while serological surveys in Thailand reported prevalence rates of 5.4% and 2.5% for T. equi and B. caballi, respectively (Kamyingkird et al., 2014). In contrast, substantially higher prevalence rates were recorded in Mongolia (78.9%) and India (62.7%) (Otgensuren et al., 2024; Mavadiya et al., 2023).

In Malaysia, several reports published by the Department of Veterinary Services (DVS) based on their national surveillance program, have documented a low prevalence of equine hemoprotozoan infections in Malaysia (Naama et al., 2021; Zawida et al., 2018; Chandrawathani et al., 1998). However, the surveillance relied solely on microscopic examination of blood smears, a method with limited sensitivity, which may underestimate the true extent of the disease in the country. In addition, a study on equine piroplasmosis using microscopic, molecular, and serological methods was conducted by Al-Sarhan (2017), but it was limited to Kelantan, a state in Malaysia and therefore did not represent the national situation. The first clinical case of EP in Malaysia was reported by Al-obaidi et al. (2015), highlighting the emergence of the disease as a potential threat to the Malaysian equine industry. Lack of study has been published on the phylogenetic analysis of equine piroplasmosis (EP) in Malaysia, leaving the origin and geographical distribution of the disease uncharacterized. Phylogenetic analysis of piroplasmosis agents is essential for tracing the source of infection, thereby facilitating the development of targeted preventive strategies to control its spread. This study reported the phylogenetic characterization of T. equi isolated from naturally infected indigenous pony horses in Malaysia.

Clinical Report

Case history: A 15-year-old Malaysian Indigenous Pony mare was presented to University Malaysia Kelantan Veterinary Teaching Hospital (HPV UMK) with complaints of persistent diarrhea and loss of weight for the past 2 months. History taking revealed the mare was purchased from the local seller last 5 months earlier and it was in good health. However, the mare started to have diarrhea and decrease in appetite in the last 2 months after giving birth to a full-term foal. The foal died 24 hours postpartum after showing signs of weakness, lethargic, unable to stand and refused to suckle despite getting assistance from the owner. The mare was previously treated symptomatically by a local private veterinarian with Marbofloxacin, Oxytetracycline, Flunixin Meglumine and multiple fluid therapy sessions with 0.9% Sodium Chloride (0.9% NaCl) also done within this 2 months duration, however the clinical signs persist. No diagnostic procedures were conducted initially; thus, the condition was managed according to its symptoms. The mare was not vaccinated against any disease before and after being purchased. However, it was dewormed with Ivermectin after being purchased. There was no tick present on the mare and other horses in the same stable.

Clinical presentation: Upon presentation at HPVUMK, the mare appeared dull but was still responsive to the stimuli. General observation revealed a rough and dull hair coat with a body score of 1/5, characterized by a prominent rib cage and pelvic bone indicating emaciation (Figure 1A). No ticks were observed on the skin, but there was evidence of persistent watery diarrhea characterized by the presence of fecal stain around its anus and tail region (Figure 1B). The feces were observed and scored as 4/4 based on its malformation and the presence of watery diarrhea. The physical examination revealed a heart rate of 42 beats/minute (normal range: 28 - 40 beats/minutes), a respiratory rate of 26 breaths/minute (normal range: 10 - 14 breaths/minutes), and a rectal temperature of 35.7 °C (normal range: 37.3 °C - 38.2 °C). The mare was hypothermic with tachypnea and slight tachycardia. The mare’s capillary refill time (CRT) was at 3 seconds (normal CRT: <2 seconds) while the dehydration status was 10% (normal dehydration status: < 5%) using indicators from the delay in the skin tent, dryness in mucous membrane and sunken of eyeballs. The oral mucous membrane appeared pale (Figure 1C), while the vulvar mucosa indicated the presence of petechial hemorrhage on its mucosa (Figure 1D). Muscle weakness in both hindlimbs was detected upon gait examination. There was also the presence of a reddish color of urine observed when the horse urinated. Blood was taken from the jugular vein in both plain and EDTA tubes for thin blood smear examination, complete blood count, serum biochemistry analysis, and Polymerase Chain Reaction (PCR). The feces were taken for bacteriology and parasitology, and urine was sent for urinalysis. At this point in time, the differential diagnosis for this case was highly suggestive of hemo- and endoparasite infestation based on the indication from the mucous membrane and urine which showed evidence of RBC lysis in the blood circulation. It was supported with evidence of poor body condition and gastrointestinal problem which is commonly seen in both hemo- and endoparasite infestation.

 

Diagnosis

Clinical pathology: The blood samples were collected by jugular venipuncture at presentation. The blood samples were placed in two 10-mL tubes. One tube contained Ethylenediaminetetraacetic acid (EDTA) at a concentration of 1 mg/mL as an anticoagulant; the second tube contained no anticoagulant. Sera were separated from the second tube and stored at -20°C until required for examination. The hematology and serum biochemistry results were listed in Table 1.

The hematological results demonstrate severe macrocytic hypochromic anemia characterized by low hematocrit (HCT), hemoglobin (Hb), mean corpuscular hemoglobin (MCH), and mean corpuscular hemoglobin concentration (MCHC), while high in mean corpuscular volume (MCV). Other than that, there was also the presence of moderate leukocytosis and lymphocytosis, and monocytosis, results of increased inflammatory responses and enhanced mononuclear phagocytic activities to engulf the senescent and dead erythrocytes due to extensive extravascular and intravascular hemolysis.

 

Table 1: Hematologic and serum biochemistry result of the mare infected with Theileria equi.

PARAMETER	RESULT	REFERENCE
Hb (g/dl)	7.0	11.0-17.0
HCT (%)	15.6	35.0-55.0
MCV (µm³)	140.0	34.0-58.0
MCH (pg)	18.6	26.0-34.0
MCHC (g/dl)	16.1	31.0-35.5
RDW (%)	14.2	10.0-16.0
WBC (10³/µL)	31.9	4.0-12.0
Lymphocyte (10³/µL)	17.0	1.5-5.5
Monocyte (10³/µL)	1.0	0.1-0.8
Granulocyte (10³/µL)	14.2	2.0-8.0
Albumin (g/dL)	2.7	1.9-3.2
Alanine transaminase (µ/l)	148	5-50
Blood urea nitrogen (mg/dL)	45	10-25
Creatinine (mg/dL)	5.6	0.8-2.2
Phosphate (mg/dL)	10.5	1.8-5.6
Total protein (g/dL)	9.7	5.6-7.9
Globulin (g/dL)	7.0	2.4-4.7

 

Serum biochemistry results demonstrate moderate elevation of alanine transaminase (ALT) suggestive liver injury secondary to oxidative stress from the immune response against the parasite. Other than that, a moderate increase in blood urea nitrogen (BUN) and creatinine indicated azotemia, suggestive of kidney dysfunction. Furthermore, the phosphate (PHOS), total protein (TP), globulin (GLOB) and albumin (ALB) levels were also increased, indicating presence of active infection and inflammation together with dehydration occurred due to diarrhea and lack of appetite.

Parasitology: Fecal and blood samples were collected to screen for endo and hemoparasites. Ten grams of fecal samples were sent to Parasitology Lab and examined using a modified McMaster and sedimentation method to count for the nematodes and trematodes eggs, respectively. Test results were negative for both tests. Five (5) mL blood aliquot with EDTA was also submitted for hemoparasite examination. A thin blood smear was performed and stained with 10% Giemsa stain. The blood smear revealed the presence of trophozoites appearing as oval, round, and spindle shapes, measuring approximately 2-3 µm in the erythrocytes, suspected as T. equi. (Figure 2A). The trophozoites can also be seen in the neutrophil (Figure 2B). Both fecal and blood samples were processed according to the guidelines provided by Manual on Parasitology (Chandrawathani et al., 2019).

 

Urinalysis

Fifty (50) ml urine was collected by manual catch during urination into a urine specimen container and submitted to Clinical Pathology Lab for physical, chemical and microscopic urinalysis. Physical urinalysis revealed reddish to brownish colour with cloudy turbidity. Chemical urinalysis using Dipstick test revealed positive in nitrite and leukocyte esterase, suggestive of urinary tract infection. There was also the presence of glucose and urobilinogen indicating kidney and liver injury. Erythrocytes and hemoglobin were also detected in the urine, indicating intravascular hemolysis caused by the T. equi. The result of urinalysis was presented in Table 2.

 

Table 2: Urinalysis result of the mare infected with Theileria equi.

PARAMETER	RESULT	REFERENCE
pH	8	5.5-8.0
Nitrites	Positive (+2)	Negative
Protein	Negative	Negative
Glucose	Positive (+4)	Negative
Ketone	Negative	Negative
Urobilinogen	Positive (+4)	Negative
Bilirubin	Negative	Negative
Erythrocyte	Positive (+4)	Negative
Hemoglobin	Positive (+4)	Negative

 

Bacteriology

Rectal swabs were submitted to the Bacteriology Lab for bacterial isolation and identification. It was cultured in blood and Mac Conkey agar at 37°C for 24 hours. The swab culture showed growth of mucoid, large, grayish colony on blood agar, but none was growth in Mac Conkey agar. The organism was identified as Enterobacter sp. based on the biochemical test. Enterobacter sp. was considered as opportunistic bacteria in horses and is mostly found in the intestine and environment such as from soil and sewage (Uchida-Fujii et al., 2020).

 

Table 3: The protocols for each primer used to detect T. evansi, B. caballi and T. equi for this study.

Agent	Primer name	Fragment size (bp)	Initial denaturation	Denaturation and annealing extension	Cycles	Final extension	Target gene
T. evansi	Kin 1	540	95°C / 3 minutes	95°C / 45 seconds	35	72°C / 10 minutes	ITSI rDNA
	Kin 2
B. caballi	TMB-F	650					SSU rRNA
	BC_R
T. equi	TMB-F	360
	EQUI_R

 

Molecular Biology

A 5 ml of whole blood coated in an EDTA tube was processed for Polymerase Chain Reaction (PCR) to test for Trypanosoma evansi, Babesia caballi and Theileria equi using different sets of primer as mentioned in Table 3. The DNA was extracted from 300 µl of whole blood using PrimeWay Genomic DNA Extraction Kit according to manufacturer instructions. A total of 25 µl mixture of PCR reaction was prepared, which included 12.5 µl of commercial Master mix (PCRBIO Taq Mix Red), 5.5 µl of nuclease free water, 1 µl each of forward and reverse primer and 5 µl of genomic DNA. Each mixture was submitted for amplification following the protocol outlined in Table 3. The product of amplification was visualized on 1.5% agarose gel stained with Midori green dye using Gel DocTM EZ imager (BIO RAD, USA). The mare was tested PCR positive for T. equi but negative for B. caballi and T. evansi. Thus, the definitive diagnosis for this case is Equine Piroplasmosis caused by T. equi. The amplified product of PCR mixture was sequenced in both directions. The result from the sequence was compared with the available sequence in the GenBank of the National Centre for Biotechnological Information (NCBI - www.ncbi.nim.nih.gov) using the Basic Local Alignment Search Tool (BLAST) programme. The nucleotide sequence showed 100% identity and query cover for Theileria equi. The sequence was deposited in GenBank with accession number PP195844. A phylogenetic analysis was performed with the Maximum Likelihood technique using the MEGA X program (Kumar et al., 2018) based on the model General Time Reversible. It contained eleven nucleotide sequences: the one acquired for this work, nine sequences of T. equi (known genotype) received from GenBank®, and one outgroup sequence of T. annulata. A discrete MUSCLE technique was used to align the sequences. The cut off point was set at 50% to further investigate the significance of the divergence.

 

As indicated in Figure 3, all the sequences were clustered within the T. equi clade, except for the outgroup sequence which in T. annulata clade, indicating genetic uniformity in the population. It showed 100% similarity with isolate from South Korea (HM229407), 94% with isolate from South Africa (EU642507) and 83% with isolate from China (KF559357). Other than that, three isolates from Malaysia, including one from this case showed genetic variation among isolates indicating genetic diversity of T. equi in Malaysia. Difference in breeds, location and years of sample collected may influence the diversity in T. equi isolates in Malaysia.

Treatment

On the day of the presentation, only Kaolin Pectin, 4 ml/kg, twice a day, per orally, was given to the mare to encounter the diarrhea. It used to harden the stool and absorb any toxin release from the chronic diarrhea. The mare was already treated by a private vet a day before our visit. The vet had administered 0.9% sodium chloride (INFUSOL®, NS) dosed at 20 ml kg−1 h−1, intravenously to correct hydration status. It also had been injected with two types of antibiotics, which were Marbofloxacin, 2 mg/kg, and Oxytetracycline, 6.6mg/kg. Marbofloxacin was given as a broad-spectrum, bactericidal antibiotic aimed to treat both gram-positive and gram- negative bacterial infections, while Oxytetracycline was given to treat hemoparasite infestation, which was suspected based on the clinical presentation. Other than that, 20 ml of multivitamins (Vitavet) was given as a vitamin supplement to support the body system and stimulate appetite, and ivermectin, 0.2mg/kg was injected intramuscularly, and at the same time, to stabilize the mare’s condition. Once the confirmatory diagnosis of Equine Piroplasmosis was reached, Diminazene Aceturate (Barenil®) was planned to be administered (3.5 mg/kg), intramuscularly. However, the mare was found dead by the owner on the day of the planned second visit. No post-mortem was conducted, and the carcass was disposed of by the owner. Equine Piroplasmosis is a notifiable disease in Malaysia; thus, veterinary authorities were promptly informed of this case.

RESULTS AND DISCUSSION

In this case, EP was diagnosed based on the clinical signs and laboratory findings. Pale mucous membrane and petechial hemorrhage in the vulva accompanied with hemoglobinuria seen in this horse that put blood parasite infestation as one of the differential diagnoses. T. equi and B. caballi will invade the bloodstream and replicate within the erythrocytes, leading to the accumulation of oxidative ions (Gopalakrishnan et al., 2015), which will further alter the cellular membrane by disrupting its osmotic fragility, causing hemolysis (Harvey, 1997). Hemolysis of RBC will reduce the number of circulating RBC in the bloodstream which contributes to the pale appearance of mucous membrane, as seen in this case. It was also reflected in a blood analysis which showed severe macrocytic hypochromic anemia similar to what was described by Maiti et al. (2021), Mahmoud et al. (2016), Al-Obaidi et al. (2016).

The lysis of RBC releases hemoglobin into the bloodstream, which will be processed to form its byproducts such as bilirubin. It will be sent to the liver to be eliminated from the body through feces, while the excess hemoglobin will be filtered by the kidneys. In severe cases where massive hemolysis occurs, the overflow of hemoglobin in the bloodstream will burden the kidney to filter the hemoglobin effectively, leading to kidney dysfunction. A marked increase in kidney parameters such as BUN and creatinine in this case was evidence of kidney dysfunction. This condition will lead to the escape of hemoglobin into the urine, known as hemoglobinuria, which can be confirmed by urinalysis. Other than that, massive hemolysis will also cause the excessive accumulation of bilirubin in the liver, which will then develop signs such as jaundice. In this case, jaundice was not seen yet; however, the liver parameter such as ALT had been significantly increased, indicating the early signs of liver injury.

Lysis of RBC is also speculated to cause coagulopathy in horses. The mechanism is still, inconclusive; however, it may be related to overproduction of inflammatory mediators that triggers the host defense mechanism and inflammatory reaction by promoting the coagulation causing hypercoagulability, systemic inflammatory response syndrome, and subsequent multiorgan system dysfunction as seen in canine babesiosis (Barić Rafaj et al., 2013). It is supported by the blood analysis which shows marked increase in leukocyte including neutrophils and lymphocyte count indicating severe infection which stimulated high amounts of inflammatory mediators and causing coagulopathy which was reflected in low amounts of platelets in blood analysis. Petechiation of mucous membrane as seen in this case was one of the signs seen as a result of coagulation disorder caused by the parasites.

Based on the clinical signs and laboratory diagnosis, it was consistent with the diagnosis of equine piroplasmosis. Pale mucous membranes, petechial hemorrhages, and hemoglobinuria were attributed to severe hemolysis caused by intraerythrocytic replication of T. equi, as confirmed by macrocytic hypochromic anemia on blood analysis. Other than that, elevated kidney and liver parameters indicated secondary renal and early hepatic dysfunction due to the accumulation of hemoglobin and bilirubin. Leukocytosis, thrombocytopenia, and mucosal petechiation shown in this case suggested the presence of inflammatory-mediated coagulopathy. The signs and laboratory analysis reflected a systemic parasitic infection leading to multisystem involvement, highlighting the need for early diagnosis and intervention.

All these signs of EP were seen more obviously after the mare delivered its foal that died 24 hours after the parturition. Thus, it is speculated that the mare was a carrier for the EP and the parasites flare up in response to the parturition. A carrier horse may have EP without any obvious clinical signs, but a flare-up of the infection and impairment of the immune system may relapse following stressful events (Ali et al., 1996). It has the risk of turning overt infection into clinical disease following a stressful event (Hailat et al., 1997). Pregnancy in carrier mares can result in abortion, miscarriage, stillbirth, or neonatal EP via intrauterine infection (De Waal, 1992; Lewis et al., 1999; Allsopp et al., 2007). In this case, the foal was successfully delivered at full term; however, its condition deteriorated after delivery, and it died 24 hours postpartum. No postmortem had been conducted on the foal; however, it can be speculated that the foal died due to neonatal piroplasmosis based on the mare’s diagnosis and information from the owner regarding the signs shown by the foal before its death. The signs of neonatal piroplasmosis are similar to the infection of EP in adult horses, but it is accompanied by other diverse and nonspecific signs such as weakness at delivery, which make the foal unable to stand properly for suckling (Rothschild, 2013), although some infected foals are born without any signs of illness (Allsopp et al., 2007). The amount of parasite load across the placenta during pregnancy, the virulence profile of the strain, and the immunological status of the foal or mare may contribute to the severity of the infection (Levi et al., 2018).

Equine Piroplasmosis can be diagnosed by microscopic examination of thin blood smear stained with 10% Giemsa stain. This method had low sensitivity in carrier horses as it was only detected in a high burden of parasitemia. B. caballi and T. equi can be distinguished through blood smear by observing the shape of the merozoites. Merozoites of B. caballi often form pairs and join at their posterior end in two pyriform shaped, while T. equi form “Maltese cross” formation from four pyriform shaped (de Waal, 1992). It is difficult to differentiate both parasites from trophozoites appearance as both of them appear similar as round, oval or elliptical in up to 3 μm in diameter (Rothchild, 2013) inside the erythrocytes for both parasites, while inside lymphocytes for T. equi (Mehlhorn and Schein, 1998). Although the presence of trophozoites in both erythrocytes and lymphocytes in this case suggested a T. equi infection, it is not entirely ruled out that there may be a mixed infection with B. caballi, necessitating further molecular identification using PCR. Polymerase Chain Reaction (PCR) was more reliable for clinical diagnosis as it will detect the antigen which reflects current exposure compared to other serological diagnosis which detected antibodies resulting from current or previous exposure. In this case, PCR analysis targeting SSU rRNA confirmed that the causative agent was T. equi.

The phylogenetic analysis showed 100% similarity with isolate found in South Korea which was previously deposited in GenBank under accession numbers HM229407. The epidemiological link may not be 100% accurate due to usage of partial sequence from this study, however; we can conclude that there is a similarity between the isolates from Malaysia and South Korea, potentially; due to shared ancestral genes of T. equi. The animal movement inter-country due to the importation of live horses Seo et al. (2013) and participation in international equestrian competitions may be the possible reasons for the migration of piroplasm transmission. In 2023, Aida et al. (2023) reported a clinical case of equine piroplasmosis from participating horses in equestrian competition that shows the possibilities of infection even though the horse had been screened to be sero-negative for EP upon arrival. Thus, close monitoring of animal health during quarantine is very crucial to control disease transmission worldwide.

In summary, this case reflects the hidden risks posed by carrier horses, where EP can silently persist and then flare into severe, life-threatening disease under stressful conditions such as parturition. The mare’s progression from subclinical infection to systemic illness, and the suspected neonatal loss, highlight the profound impact EP can have not only on individual animals but also on breeding outcomes. The confirmation of T. equi infection through PCR and the genetic link to strains from South Korea emphasize the role of international horse movement to silently facilitate disease transmission. This case serves as a reminder that even clinically healthy horses may harbor significant pathogens, reinforcing the need for vigilant screening, long-term monitoring, and a deeper understanding of stress-induced disease reactivation to ensure the health of horses was protected.

CONCLUSIONS AND RECOMMENDATIONS

Equine Piroplasmosis can be a life-threatening event in immunosuppressed horses. Screening for the disease before mating and parturition is necessary to prevent the disease transmission and promote an early detection of the disease before they are exposed to the immunosuppressed condition. Treatment of piroplasmosis in chronically infected horses is not promising, thus early detection and prompt treatment are necessary to improve the prognosis of the infected horses.

ACKNOWLEDGEMENTS

This work was supported by the Fundamental Research Grant Scheme (FRGS) funded by the Ministry of Higher Education (KPT), Malaysia (Project No. FRGS/1/2022/WAB04/UMK/03/1) We are thankful to Ms. C.W Salma from Molecular Biology Laboratory, University Veterinary Diagnostic Centre (UVDC), UMK for technical help in the molecular work and Mr. Fakrul Nizam from HPV UMK for his help in field investigation.

NOVELTY STATEMENTS

This report documents the first molecular and phylogenetic confirmation of Theileria equi infection in a postpartum mare in Kelantan, Malaysia. The isolate demonstrated 100% genetic similarity to a South Korean strain, suggesting a possible transboundary epidemiological connection. The case highlights the clinical impact of T. equi infection in immunocompromised horses and emphasizes the need for routine pre-breeding and pre-parturition screening to prevent disease-associated reproductive losses. These findings contribute novel insights into regional transmission dynamics and support improved control strategies for equine piroplasmosis in Southeast Asia.

AUTHOR’S CONTRIBUTIONS

Nur Zul Izzati Mohd Rajdi: Methodology, Investigation, Writing‐original draft.

Mimi Armiladiana Mohamad: Investigation, Resources.

Rumaizi Shaari: Investigation, Writing‐review and editing.

Mohd Farhan Hanif Reduan: Formal analysis, Writing‐original draft.

Mohammad Sabri Abdul Rahman: Methodology, Formal analysis, Writing‐review and editing. Mohammad Dauda Goni: Writing‐review and editing.

Li Peng Tan: Formal analysis, Writing‐original draft, Methodology.

Conflict of Interest

The authors declare that the research was conducted in the absence of any potential conflict of interest.

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