Seroprevalence of Theileria ovis in Goats from M’Sila Region, Central Algeria

Introduction
Theileriosis, caused by the haemoprotozoan Theileria, is one of the most common tick-borne diseases in livestock worldwide (Gharbi et al., 2020). This parasitic disease is considered one of the major constraints of small ruminant production, causing considerable economic losses mainly in Asia, Africa and the southern part of Europe (Schnittger et al., 2004). More than 185 species are known within the genus Theileria in domestic and wild animals (Al-Fahdi et al., 2017). Theileria annulata and Theileria parva are the most important pathogens for cattle (Gharbi et al., 2020; Al-Fahdi et al., 2017), while Theileria lestoquardi (formerly known as Theileria hirci), Theileria luwenshuni and Theileria uilenbergi are pathogenic for small ruminants (Mans et al., 2015; Aydin et al., 2013; Altay et al., 2012). Other species less virulent or non-pathogenic such as Theileria ovis, Theileria separata, Theileria recondita, T. annulata and uncharacterized isolates of Theileria (Theileria sp. OT1, Theileria sp. OT3 and Theileria sp. MK) have been reported in sheep and goats (Aydin et al., 2013; Altay et al., 2012; Zaeemi et al. 2011; Defaye et al. 2022; Stuen, 2020). Theileria species of small ruminants are distributed particularly in the tropical and subtropical regions of Africa, the Middle East, Asia and eastern and southern Europe (Zhang et al., 2015). Various tick species belonging to Hyalomma, Haemaphysalis and Rhipicephalus genera are known as vectors of this protozoan (Mans et al., 2015; Stuen, 2020). Theileria species discrimination is of epidemiological and clinical significance. The species identification on microscopic examination of blood smears is limited. Serological tests are essential and reliable means for carrying out epidemiological surveys. Indirect fluorescent antibody test (IFAT) is the most widely used method for serodiagnosis of the common important species (WOAH, 2018). However, these serological tests have certain drawbacks, such as cross-reactivity of genetically closely related species. Thus, molecular methods remain an essential line of inquiry in determining species and genetic variations (Mans et al., 2015; Nagore et al., 2004; Tahaa et al., 2023).
In Algeria, the epidemiology of theileriosis in livestock is poorly known. Few studies, limited to some geographical regions, have been conducted on the prevalence and molecular characterization of Theileria spp. in ruminants and some tick species. Based on the microscopic examination of Giemsa-stained blood smears, the prevalence of bovine tropical theileriosis has been estimated at 10.4% to 45.5% in central-east of Algeria (Ayadi et al., 2016; Benchikh Elfegoun et al., 2017; Ziam et al., 2020; Foughali et al., 2021a). T. annulata has also been characterized in cattle with a prevalence of 25.4% to 50% (Ayadi et al., 2016; Ziam et al., 2015). Moreover, bovine Theileria isolates closely related to Theileria buffeli have been detected with an infection rate of 6.70% (Ziam et al., 2015). Both T. annulata and T. buffeli were confirmed in cattle and Rhipicephalus annulatus (Sadeddine et al., 2020). In sheep, an infection rate of 2.32% of Theileria spp. has been recorded using microscopic examination (Foughali et al., 2021b). T. ovis was detected in sheep, goats and Rhipicephalus bursa and Rhipicephalus turanicus from northeastern Algeria (Sadeddine et al., 2020; Aouadi et al., 2017). Additionally, T. annulata was detected in sheep from the east of Algeria (Sadeddine et al., 2020). In horses, microscopic examination and competitive ELISA test revealed an overall prevalence of Theileria equi of 15.9% and 29.12%, respectively (Benfenatki et al., 2016). Recently, T. equi was genetically identified in equines and cattle (Sadeddine et al., 2020). Algeria has a large area with climate diversity, which requires exhaustive studies and data to understand the epidemiology of this economically important parasitic disease. To support previously reported data, the present study aimed to investigate the seroprevalence of Theileria spp. in goats from the M’Sila region (central Algeria) using the IFA test.

Materials and Methods

Study area and sampling 
The present study was carried out in the Maâdid locality from M’Sila Province, which is a Steppic region located in the central part of Algeria occupying an area of 18175 km2 (GPS: 35°42’7’’N and 4°32’49’’E) (Figure 1). This region is characterized by a semi-arid climate with very hot and dry summers and relatively cold winters. The average annual temperature ranges from 16.90 -18.40 °C. Breeding of small ruminants represents the main agricultural activity in this region.

Blood samples were collected from 128 goats’ jugular veins from January to September 2016. Goats belong to 19 semi-extensive farms associated or not with sheep breeding. Approximately 10% of each farm’s reared goats were randomly selected for blood sampling. Goats were healthy, and no infection signs were detected during sampling. Data on animal characteristics, including gender, age, breeding system and tick infestation, were recorded during sampling. All samples were collected in EDTA tubes and subsequently centrifuged and the obtained sera were preserved at -20 °C until serological analysis.
Based on the institutional review board statement, the animal study, and the blood sample collection were carried out in accordance with the current Algerian Regulations No. 88-08 of January 26, 1988, related to Veterinary Medicine Activities and the protection of animal health (N°004 JORA of 27-01-1988).

Serological analysis
An indirect fluorescence antibody test (IFAT) was performed to screen the goats as previously described (Papadopoulos et al., 1995). Antibodies against T. lestoquardi were detected using Teflon-coated slides fixed with T. lestoquardi schizont stage antigens. Slides with blood smears infected by T. ovis were used as antigens to detect antibodies against this species. The blood smears were fixed with acetone, as previously described (Ludford, 1969). Positive and negative control sera were included for each reaction.
Sera were diluted in PBS at 1/20 and 1/40 for antibodies IgG detection of T. ovis and T. lestoquardi, respectively. Subsequently, they were deposited in wells of Teflon-coated slides (20 μL/well) and incubated at room temperature for 20 min. After three washes in PBS, specific antibodies for targeted Theileria species could be detected using anti-goat IgG conjugated to fluorescein isothiocyanate diluted at 1/50 in PBS. Finally, sera and conjugates were incubated for 20 min at room temperature. Three washes in PBS were then carried out. The slides were covered with coverslips using glycerol 50% in PBS. The slides were examined under objective 40× (total magnification of 400×) using a fluorescent microscope. Sera were considered positive at titer ≥1/20 and ≥1/40 for T. ovis and T. lestoquardi, respectively. Moreover, positive sera with titer ≥1/20 were tested for other titers (1/100, 1/200, 1/400).

Statistical analyses
Statistical analyses were performed using online MedCalc's free statistical calculators website. The Fisher exact and chi-square tests were exploited to assess the seroprevalence variation according to previously recorded animal attributes (independent variables). The P<0.05 was regarded as statistically significant. 

Results

Seroprevalence and associated risk factors
Out of 128 tested samples, 21 goats (16.40%) were seropositive for T. ovis (Table 1). All tested goats were seronegative for T. lestoquardi. Of the 19 screened farms, 12(63.15%) were seropositive. The seroprevalence of T. ovis in these farms varied from 10% to 30% (Table 1).

Among the positive sera with the titer 1/20 (21 samples; 16.40%), 14(10.93%), 5(3.90%) and 2(1.56%) were positive with titers of 1/100, 1/200 and 1/400, respectively (Table 2).

Furthermore, females revealed slightly higher seroprevalence than males (18.18% vs 13.72%). The highest seroprevalence was recorded in goats over 24 months (20.73%) compared to those aged 12-24 months and less than 12 months, which showed seroprevalence of 8.57% and 9.09%, respectively. The seroprevalence was higher (17.52%) in farms where sheep breeding was absent compared to farms where sheep breeding was present (12.90%). The seroprevalence did not vary significantly with age, sex, or breeding system (presence or absence of sheep) (all results showed P>0.05). Tick-infested goats were significantly more seropositive (40%) than those not infested (12.03%) (Table 3).

Discussion
The present study is based on IFA test reports for the first time on the seroprevalence of T. ovis in goats from the M’Sila region, central Algeria. The IFA test is commonly used as a serological method for detecting specific antibodies of Theileria spp. in small ruminates. This recognition of a specific humoral response benefits the epidemiological surveys compared to the microscopic examination of blood smears, which remains challenging, particularly for chronic infections. However, cross-reactions or weak specific-immune responses represent the main disadvantages of serological methods (WOAH, 2022). Data on the seroprevalence of Theileria spp. in small ruminants are relatively limited worldwide. The seroprevalence of T. ovis in the present study was higher than that in goats from Greece (Papadopoulos et al., 1996). A comparable result has been reported in Turkey (Sayin et al., 2009). These previous studies have documented higher seroprevalences in sheep (Papadopoulos et al., 1996; Sayin et al., 2009). Different serological investigations have also reported variable prevalence rates of Theileria spp. in sheep and goats (Alyasino & Greiner, 1999; Guo et al., 2007; Luo et al., 2017; Li et al., 2017). All seropositive samples were detected with a titer ≥1/20, of which only two were detected with a titer of 1/400. Various factors, such as the study design, sample size, sampling period and tick infestation of animals, as well as the infection status and intensity of the specific humoral response, could influence the seroprevalence variations. The seroprevalence did not vary significantly with age, sex, or breeding system (presence or absence of sheep breeding associated with the examined farms). In goats, no difference in the seroprevalence of Theileria infection was reported between females and males (Luo et al., 2017). In sheep, the molecular prevalence of T. ovis did not vary significantly for age categories, while females were significantly more infected than males (Rjeibi et al., 2014). The effect of associated host factors on the susceptibility to infection is not fully well understood. Tick-infested goats were more seropositive than no infested goats. Different species belonging to Hyalomma, Haemaphysalis and Rhipicephalus, which are distributed in Africa’s tropical and subtropical regions, could transmit Theileria spp. in small ruminants (Stuen, 2020). The presence of T. ovis in sheep and goats has been confirmed in North African countries such as Algeria and Tunisia using molecular approaches (Sadeddine et al., 2020; Aouadi et al., 2017; Rjeibi et al., 2014; M’ghirbi et al., 2013). T. ovis was a non-pathogenic species for infected animals (Aouadi et al., 2017; M’ghirbi et al., 2013).
Similarly to our findings on the seroprevalence of T. lestoquardi (0.0%), all goats and even sheep were tested to be seronegative in Turkey (Sayin et al., 2009). Seroprevalence rates of this species, ranging from 1.2% to 33.8%, were revealed in sheep from Tunisia and Sudan (Rjeibi et al., 2016; Salih et al., 2003; Ahmed et al., 2018; Hassan et al., 2019). The absence of seropositive cases should not be excluded, as it may be related to the detection threshold of the test for a weak specific-immune response. T. lestoquardi, the causative agent of malignant ovine theileriosis (MOT), is distributed particularly in some countries from sub-Saharan Africa (Sudan and Tanzania) and Asia (Iraq, Iran, India, Pakistan, Saudi Arabia, and Oman) (Rjeibi et al., 2016; El Imam & Taha, 2015; Saeed et al., 2015; Awad et al., 2018; Habibi et al., 2020; Rassim Mohammed & Al-Saadi, 2023). The occurrence of this species has been confirmed molecularly in sheep from Tunisia (Rjeibi et al., 2016). Its geographical extent is likely broader than previously reported since this species has occurred, for example, in the Maghreb region.

Conclusion
In conclusion, the current study reports valuable data on the seroprevalence of T. ovis in goats from central Algeria. Further exhaustive studies in different regions throughout the national territory are required to understand better the epidemiology of theileriosis in small ruminants, particularly regarding the prevalence, clinical impact, and molecular characterization of infecting species.

Ethical Considerations

Compliance with ethical guidelines
The animal study protocol was approved by the Institutional Review Board of Research Center in Agropastoralism of Ziane Achour University, Djelfa, Algeria, in accordance Ministry of Higher Education and Scientific Research, for studies involving animals.

Funding
The paper was extracted from the master's thesis of Laadjal Soumia, approved by the Department of Veterinary Sciences, Higher National Veterinary School, Algiers.

Authors' contributions
Conceptualization: Ghalmi Farida, Hafsi Fella, and Cantekin Zafer; Samples collection: Laadjal Soumia, and Reghaissia Nassiba; Experiments: Cantekin Zafer; Statistical analyses: Dahmane Abdeldjalil, and Samari Houssem; Initial draft preparation: Reghaissia Nassiba, Laadjal Soumia, and Laatamna AbdElkarim; Final approval: All authors.

Conflict of interest
The authors declared no conflict of interest.

Acknowledgments
The authors enormously thank the owners of farms for their help in sampling.

References

Ahmed, H. D. M., Ahmed, A. M., Salih, D. A., Hasan, S. K., Masri, M. M., & Etyeb, H. N., et al. (2018). Prevalence, first molecular identification and characterization of Theileria lestoquardi in sheep in Alhuda national sheep research station, Al Gezira state, Sudan. Asian Journal of Biology, 7(2), 1-9. [DOI:10.9734/AJOB/2018/45972]

Al-Fahdi, A., Alqamashoui, B., Al-Hamidhi, S., Kose, O., Tageldin, M. H., & Bobade, P., et al. (2017). Molecular surveillance of Theileria parasites of livestock in Oman. Ticks and Tick-Borne Diseases, 8(5), 741–748. [DOI:10.1016/j.ttbdis.2017.05.008] [PMID]

Altay, K., Dumanli, N., & Aktas, M. (2012). A study on ovine tick-borne hemoprotozoan parasites (Theileria and Babesia) in the East Black Sea region of Turkey. Parasitology Research, 111(1), 149-153.‏ [DOI:10.1007/s00436-011-2811-8] [PMID]

Alyasino, Y., & Greiner, M. (1999). Serosurvey on theileriosis in Awassi sheep in Syria. Veterinary Parasitology, 81(4), 275-280. [DOI:10.1016/S0304-4017(98)00250-7] [PMID]

Aouadi, A., Leulmi, H., Boucheikhchoukh, M., Benakhla, A., Raoult, D., & Parola, P. (2017). Molecular evidence of tick-borne hemoprotozoan-parasites (Theileria ovis and Babesia ovis) and bacteria in ticks and blood from small ruminants in Northern Algeria. Comparative Immunology, Microbiology and Infectious Diseases, 50, 34–39. [DOI:10.1016/j.cimid.2016.11.008] [PMID]

Awad, H., Al-Hamidhi, S., El Hussein, A. M., Yousif, Y. M. Z., Taha, K. M., & Salih, D. A., et al. (2018). Theileria lestoquardi in Sudan is highly diverse and genetically distinct from that in Oman. Infection, Genetics and Evolution, 62, 46-52. [DOI:10.1016/j.meegid.2018.04.014] [PMID]

Ayadi O., Rjeibi, M. R., BenchikhElfegoun, M. C., & Gharbi, M. (2016). Prevalence and risk factors of tropical theileriosis, and sequencing of Theileria annulata, the causative pathogen, in Setif region (Algeria) before and after tick season. Revue d’Elevage et de Médecine Vétérinaire des Pays Tropicaux, 69(4), 161-166.‏ [DOI:10.19182/remvt.31201]

Aydin, M. F., Aktas, M., & Dumanli, N. (2013). Molecular identification of Theileria and Babesia in sheep and goats in the Black Sea region in Turkey. Parasitology Research, 112(8), 2817-2824. [DOI:10.1007/s00436-013-3452-x] [PMID]

Benchikh Elfegoun, M. C., Gharbi, M., Merzekani, Z., & Kohil, K. (2018). Bovine piroplasmosis in the provinces of Skikda and Oum El Bouaghi (Northeastern Algeria): Epidemiological study and estimation of milk yield losses. Revue d’élevage et de Médecine Vétérinaire des Pays Tropicaux, 70(3), 105-110.‏ [DOI:10.19182/remvt.31519]

Benfenatki, A., Younes Bouacida, N. S., Ait Oudhia, K., & Khelef, D. (2016). Prevalence of Theileriaequi infection in Algiers urban area using cELISA and microscopic examination. Asian Journal of Animal and Veterinary Advances, 11(8), 511-515.‏ [DOI:10.3923/ajava.2016.511.515]

Defaye, B., Moutailler, S., Pasqualini, V., & Quilichini, Y. (2022). Distribution of tick-borne pathogens in domestic animals and their ticks in the countries of the Mediterranean basin between 2000 and 2021: A systematic review. Microorganisms, 10(6), 1236. [DOI:10.3390/microorganisms10061236] [PMID] [PMCID]

El Imam A. H., &Taha, K. M. (2015). Malignant ovine theileriosis (Theileria lestoquardi): A review. Jordan Journal of Biological Sciences, 8, 165-174. [DOI:10.12816/0026953]

Foughali, A. A., Jedidi, M., Dhibi, M., Mhadhbi, M., Sassi, L., & Berber, A., et al. (2021b). Infection by haemopathogens and tick infestation of sheep during summer season in Constantine region, Northeast Algeria. Veterinary Medicine and Science, 7(5), 1769-1777.‏ [DOI:10.1002/vms3.551] [PMID] [PMCID]

Foughali, A. A., Ziam, H., Aiza, A., Boulkrout, H., Berber, A., & Bitam, I., et al. (2021a). Cross-sectional survey of cattle haemopathogens in Constantine, Northeast Algeria. Veterinary Medicine and Science, 7(4), 1237-1244. [DOI:10.1002/vms3.459] [PMID] [PMCID]

Gharbi, M., Darghouth, M. A., Elati, K., Al-Hosary, A. A. T., Ayadi, O., & Salih, D. A., et al. (2020). Current status of tropical theileriosis in Northern Africa: A review of recent epidemiological investigations and implications for control. Transboundary and Emerging Diseases, 67(Suppl 1), 8–25. [DOI:10.1111/tbed.13312] [PMID]

Guo, S., Mu, Y., Liu, Z., Ma, D., Yang, S., & Ge, G., et al. (2007). Serological investigation of ovine theileriosis by ELISA in Gannan Tibet region of Gansu province in China. Parasitology Research, 101 (Suppl 2), S197–S200. [DOI:10.1007/s00436-007-0686-5] [PMID]

Habibi, G., Sepahvand-Mohammadi, E., Afshari, A., & Bozorgi, S. (2020). Molecular detection of Theileria spp. and Babesia ovis Infection in Sheep in Baneh, Iran. Archives of Razi Institute, 75(2), 289–296. [PMID]

Hassan, S., Skilton, R. A., Pelle, R., Odongo, D., Bishop, R. P., & Ahmed, J., et al. (2019). Assessment of the prevalence of Theileria lestoquardi in sheep from the Sudan using serological and molecular methods. Preventive Veterinary Medicine, 169, [DOI:10.1016/j.prevetmed.2019.104697] [PMID]

Karaşahi̇n,T., Dursun,S., Hayat Aksoy, N., İpek, H., & Şentürk, G. (2023). Hematological parameters in hair goats during and out of breeding season hair goats seasonal hematological parameters. Iranian Journal of Veterinary Medicine, 17(2), 113-118. [DOI:10.32598/IJVM.17.2.1005334]

Li, Q., Han, Y., Liu, Z., Liu, J., Liu, A., & Li, Y., et al. (2017). Serosurvey of ovine and caprine theileriosis with ELISA in China. Small Ruminant Research, 154, 78-80.‏ [DOI:10.1016/j.smallrumres.2017.07.009]

Ludford, C. G. (1969). Fluorescent antibody staining of four Babesia species. Experimental Parasitology, 24(3), 327-335. [DOI:10.1016/0014-4894(69)90171-4] [PMID]

Luo, H., Li, K., Zhang, H., Lan, Y., Gan, P., & Xiong, X., et al. (2017). Seroprevalence of Theileria infection in goats in Hubei province, China based on circulating antibodies. Pakistan Veterinary Journal, 37(1), 105-107.‏ [Link]

M'ghirbi, Y., Ros-García, A., Iribar, P., Rhaim, A., Hurtado, A., & Bouattour, A. (2013). A molecular study of tick-borne haemoprotozoan parasites (Theileria and Babesia) in small ruminants in Northern Tunisia. Veterinary Parasitology, 198(1-2), 72-77. [DOI:10.1016/j.vetpar.2013.08.005] [PMID]

Mans, B. J., Pienaar, R., & Latif, A. A. (2015). A review of Theileria diagnostics and epidemiology. International Journal for Parasitology. Parasites and Wildlife, 4(1), 104-118.‏ [DOI:10.1016/j.ijppaw.2014.12.006] [PMID] [PMCID]

Nagore, D., García-Sanmartín, J., García-Pérez, A. L., Juste, R. A., & Hurtado, A. (2004). Identification, genetic diversity and prevalence of Theileria and Babesia species in a sheep population from Northern Spain. International Journal for Parasitology, 34(9), 1059-1067.‏ [DOI:10.1016/j.ijpara.2004.05.008] [PMID]

Papadopoulos, B., Brossard, M., & Perié, N. M. (1996a). Piroplasms of domestic animals in the Macedonia region of Greece, 3. Piroplasms of small ruminants. Veterinary Parasitology, 63(1-2), 67-74. [DOI:10.1016/0304-4017(95)00846-2] [PMID]

Papadopoulos, B., Perié, N. M., & Uilenberg, G. (1996b). Piroplasms of domestic animals in the Macedonia region Greece. 1. Serological cross-reactions. Veterinary Parasitology, 63(1-2), 41–56. [DOI:10.1016/0304-4017(95)00878-0] [PMID]

Rassim Mohammed, A., & Al-Saadi, M. (2023). Quantification of acute and chronic theileriosis in sheep by quantitative PCR (qPCR). Archives of Razi Institute, 78(1), 389–396. [PMID]

Rjeibi, M. R., Darghouth, M. A., Rekik, M., Amor, B., Sassi, L., & Gharbi, M. (2016). First molecular identification and genetic characterization of Theileria lestoquardi in sheep of the Maghreb region. Transboundary and Emerging Diseases, 63(3), 278-284.‏ [DOI:10.1111/tbed.12271] [PMID]

Rjeibi, M. R., Gharbi, M., Mhadhbi, M., Mabrouk, W., Ayari, B., Nasfi, I., et al. (2014). Prevalence of piroplasms in small ruminants in North-West Tunisia and the first genetic characterisation of Babesiaovis in Africa. Parasite, 21, [DOI:10.1051/parasite/2014025] [PMID] [PMCID]

Sadeddine, R., Diarra, A. Z., Laroche, M., Mediannikov, O., Righi, S., & Benakhla, A., et al. (2020). Molecular identification of protozoal and bacterial organisms in domestic animals and their infesting ticks from Northeastern Algeria. Ticks and Tick-Borne Diseases, 11(2), 101330.‏ [DOI:10.1016/j.ttbdis.2019.101330] [PMID]

Saeed, S., Jahangir, M., Fatima, M., Shaikh, R. S., Khattak, R. M., Ali, M., & Iqbal, F. (2015). PCR based detection of Theileria lestoquardi in apparently healthy sheep and goats from two districts in Khyber PukhtoonKhwa (Pakistan). Tropical Biomedicine, 32(2), 225–232. [PMID]

Salih, D. A., ElHussein, A. M., Hayat, M., & Taha, K. M. (2003). Survey of Theileria lestoquardi antibodies among Sudanese sheep. Veterinary Parasitology, 111(4), 361–367. [DOI:10.1016/S0304-4017(02)00389-8] [PMID]

Sayin, F., Nalbantoğlu, S., Yukari, B. A., Çakmak, A., & Karaer, Z. (2009). Epidemiological studies on sheep and goat Theileria infection. Ankara ÜniversitesiVeterinerFakültesiDergisi, 56, 127-129. [DOI:10.1501/Vetfak_0000002168]

Schnittger, L., Yin, H., Qi, B., Gubbels, M. J., Beyer, D., & Niemann, S., et al. (2004). Simultaneous detection and differentiation of Theileria and Babesia parasites infecting small ruminants by reverse line blotting. Parasitology Research, 92(3), 189–196. [DOI:10.1007/s00436-003-0980-9] [PMID]

Stuen S. (2020). Haemoparasites-Challenging and wasting infections in small ruminants: A review. Animals, 10(11), 2179.‏ [DOI:10.3390/ani10112179] [PMID] [PMCID]

Taha, K. M., Salih, D. A., Ali, A. M., Omer, R. A., & El Hussein, A. M. (2013). Naturally occurring infections of cattle with Theileria lestoquardi and sheep with Theileria annulata in the Sudan. Veterinary parasitology, 191(1-2), 143–145. [DOI: 10.1016/j.vetpar.2012.08.003] [PMID]

World Organisation of Animal Health. (2018). Theileriosis in cattle (infection with theileria annulata, T. orientalis and T. parva). In: World Organisation of Animal Health (Eds). Manual of Diagnostic Tests and Vaccines for Terrestrial Animals, ( 1-26). World Organisation of Animal Health Publishing. [Link]

World Organisation of Animal Health. (2022). Theileriosis in sheep and goats (Infection with TheileriaLestoguardi, T. luwenshuni and T. uilenbergi). In: World Organisation of Animal Health (Eds). Manual of Diagnostic Tests and Vaccines for Terrestrial Animals, ( 1-14). World Organisation of Animal Health Publishing. [Link]

Zaeemi, M., Haddadzadeh, H., Khazraiinia, P., Kazemi, B., & Bandehpour, M. (2011). Identification of different Theileria species (Theileria lestoquardi, Theileria ovis, and Theileria annulata) in naturally infected sheep using nested PCR-RFLP. Parasitology Research, 108(4), 837–843. [DOI:10.1007/s00436-010-2119-0] [PMID]

Zhang, J., Kelly, P., Li, J., Xu, C., & Wang, C. (2015). Molecular detection of Theileria spp. in livestock on five Caribbean islands. BioMed Research International, 2015, [DOI:10.1155/2015/624728] [PMID] [PMCID]

Ziam, H., Kelanamer, R., Aissi, M., Ababou, A., Berkvens, D., & Geysen, D. (2015). Prevalence of bovine theileriosis in North Central region of Algeria by real-time polymerase chain reaction with a note on its distribution. Tropical Animal Health and Production, 47(5), 787–796. [DOI:10.1007/s11250-015-0772-0] [PMID]

Ziam, H., Kernif, T., Saidani, K., Kelanemer, R., Hammaz, Z., & Geysen, D. (2020). Bovine piroplasmosis-anaplasmosis and clinical signs of tropical theileriosis in the plains of Djurdjura (north Algeria). Veterinary Medicine and Science, 6(4), 720-729.‏ [DOI:10.1002/vms3.305] [PMID] [PMCID]