In the present study, the seroprevalences of some important parasitological and virological agents were investigated in horses and donkeys in Balikesir and surroundings, in Turkey. According to the results, it can be suggested that piroplasmosis agents are generally common in both species except B. caballi, which is not common among donkeys. Nevertheless, T. equi was found that very common in donkeys contrary to B. caballi. Although B. caballi was detected at only one sampling point in both species, T. equi was commonly detected in all sampling points in both species. In contrary to this study, Acici et al. (2008) found that B. caballi were detected more prevalent than T. equi in both horses and donkeys. In parallel with this, Machado et al. (2012) reported that B. caballi and T. equi seroprevalence were 93% and 73% respectively in donkeys. However, Garcia-Bocanegro et al. (2013) reported parallel results with our study; B. caballi in horses 7%, in donkeys 17%, T. equi in horses 48%, in donkeys 47% were detected in Southern Spain. Furthermore, Oduori et al. (2015) reported seroprevalences for T. equi and B. caballi were found 81% and 0% in donkeys respectively. When considered that the chronic form of T. equi is more prevalent in donkeys than horses, and the acute form of the disease is rarely observed in donkeys, it can be suggested that the results of the study are highly compatible with our results (Kumar et al., 2009; Wise et al., 2014). Theileria equi was detected significantly higher than B. caballi in both species and it can be suggested that our results are in agreement with previous reports from endemic countries (Salim et al. 2008; Sigg et al. 2010; Garcia-Bocanegra et al. 2013; Oduouri et al. 2015).
In this study, a prominent difference was found between the T. gondii seroprevalences of both species. In humans and horses, an association has been described between low-income and increased exposure to T. gondii. The high exposure to the parasite may be caused by poor sanitary conditions associated with low-income populations which may increase the chance of contamination in water sources or else. (Li et al. 2020; Rostami et al. 2020). In parallel with this, the donkeys that we collected blood samples had poor sanitary conditions when compared to the horses; it can explain the difference in the seroprevalence of the parasite between both species. The seroprevalence of the parasite in horses show highly variable between countries and it was detected ranges between %1 and %71 in Sweden (Jakubek et al. 2006) and Iran (Hajialilo et al. 2010) respectively. The difference in seroprevalence can result from hygiene conditions, managing, and feeding practices (Li et al. 2020; Tenter et al. 2000). From different countries in the world, different seroprevalence results are reported that the seroprevalence of the parasite is higher in donkeys than in horses; Brazil (Munhoz et al. 2019) 72% in donkeys and 27% in horses and parallel, other studies from Pakistan (Saqib et al. 2015) and Spain (Garcia-Bocanegra et al. 2012) can be suggested. However, because of are fewer studies evaluating both species in the same cohorts, it is difficult to determine that these results are caused by the naturally higher susceptibility of donkeys or poor sanitary conditions. Nevertheless, there are several suggestions that horses are considered to be naturally resistant to T. gondii infection (Dubey 2010) or they develop very low antibody titers to detect by serological tests (Evers et al. 2013). Likewise, Garcia-Bocanegra et al. (2012) reported that the seroprevalence of T. gondii was higher in the animals that are kept outdoor than are kept indoors. Additionally, Alvarado-Esquivel et al. (2017) stated that horses generally received better care about good feeding and drinking healthy water than donkeys. In this study, our results are compatible with the previous results, the higher seroprevalence in donkeys, reported from different countries (Garcia-Bocanegro et al. 2012; Saqib et al. 2015; Munhoz et al. 2019). Several studies were performed by Gazyagci et al. (2011), Karatepe et al. (2010), Akca et al. (2004), Göz et al. (2007) related the seroprevalence of T. gondii in horses in Turkey, and results of the studies were 36%, 7%, 20%, 28% respectively; it can be suggested that the result of this study (9.09%) is in agreement with the previous reports especially Karatepe et al. (2010). Nevertheless, there are not many investigations about the seroprevalence of T. gondii in donkeys from Turkey; Balkaya et al. (2011) was found the prevalence of the parasite 62%, and that is also compatible with our result. However, T. gondii is not only common in equids but also other mammalian such as sheep (98%) (Çiçek et al. 2011) and (31%) (Oncel and Vural 2006), cattle (93%) (Akca and Mor 2010), cat (35%) (Can et al. 2014), dog (62%) (Balkaya et al. 2010), and also human (58%) (Doni et al. 2015) in Turkey. It can be suggested that T. gondii is highly prevalent in Turkey both humans and other mammalians, and this is a very prominent problem according to public health.
Neosporosis is not considered to be zoonotic and its most important effect is the economic loss caused by the disturbance of the reproduction of cattle and small ruminants. (Dubey et al. 2017). However, due to the consumption of donkey and horse meat in some countries and increased interest in general using the animals, several studies were performed. Cong et al. (2018) and Gennari et al. (2016) found the seropositivity of the parasite in donkeys 10% and 2% respectively. Similarly, Waap et al. (2020) and Machacova et al. (2013) reported that the seroprevalence of the parasite is in horses and donkeys were 9% and 11% respectively. Bartova et al. (2017) found the seroprevalence of Neospora sp. was 8% in horses and all donkey was found to be negative. Kligler et al. (2007) found the seroprevalence of Neospora sp. in horses 12% and this result is prominently lower than the seroprevalence of the donkeys who live in similar geography of Israel. Besides the studies which were found low or no seropositivity. Gharekhani et al. (2013) found the seroprevalences of Neospora sp. were 40% and 52% in horses and donkeys, respectively. Also, Tirosh-Levy et al. (2020) found 70% in donkeys. Sevgili et al. (2003) found the seroprevalence of Neospora sp. 8% in Thoroughbred mares Sanliurfa in Turkey. In this study, Neospora sp. seroprevalences were found 10.6% and 23.95% in horses and donkeys respectively; it can be suggested that our results are compatible with previous reports (Machacova et al. 2013; Gennari et al. 2016; Bartova et al. 2017; Cong et al. 2018; Waap et al. 2020) especially those reported from the rural regions of Israel (Kligler et al. 2007). Furthermore, this report is important according to being the first report about the presence of anti-Neospora sp. antibodies in donkeys in Turkey.
Seroprevalence reports of EVA are globally inconsistent which is ranging from 0–20% between countries. It is considered that these changes between the seroprevalence depend on horse population and surveillance of the country (Maclachlan et al. 2006; Bello et al. 2007; Turan et al. 2007). In parallel, Nejat et al. (2015) found EVA antibodies at 4% in Iran, and Cruz et al. (2016b) found 16% in Spain. Similarly, Marenzoni et al. (2013) and Turan et al. (2007) were found the seroprevalence of EVA in horses in Turkey 16% and 14% respectively. Some reports from Turkey indicate relatively low seroprevalence in both horses and donkeys. While Gür et al. (2019) found the seroprevalence of EVA 3% in donkeys, Bulut et al. (2012) found 23% in horses. In this study, contrary to Marenzoni et al. (2013) and Turan et al. (2007), there was no sample determined to be positive for EVA in both of the species, in the same region in Turkey. It can be suggested that the seroprevalence of EVA is highly changeable not just in Turkey, even in the same region.
Many seroprevalence studies were performed regarding EIV in horses and donkeys, and its seropositivity rate is highly changeable. The seropositivity rate of EIV in horses was found at 38%, 11%, and 93% by Blitvich et al. (2010), Sajid et al. (2013), Happold and Rubira (2011) from Mexico, Pakistan, and Australia respectively. On the other hand, Sajid et al. (2013) and Chencev et al. (2011) were found the seropositivity rate of the virus 12% and 65% from Pakistan and Bulgaria in donkeys, respectively. Ataseven and Daly (2007) were performed a large-scale EIV seroprevalence study on more than six hundred equids from five different regions in Turkey. The overall seroprevalence of the virus was found 31%. The highest overall rate for EIV was found in the Marmara region (60%) where this study also performed and the seropositivity of horses (41%) was more than four-fold higher than donkeys (9%). According to Timurkan and Aydin (2019), 26.3% of antibodies were detected in a study conducted in eastern Turkey, in the province of Erzurum, in jereed horses, but they could not detect a virological positivity. Nevertheless, in this study virological prevalence of EIV was found 3% and 1% in horses and donkeys respectively. It can be suggested that our results are prominently low when the results of previous reports are considered. The reason for the seroprevalence difference between previous studies and this study can be the detection method of the virus. While in this study PCR method was used and antigens of the virus were detected, the ELISA method was used, and the virus-specific antibodies were detected in studies discussed before. So, it can be suggested that seroprevalence may be higher in studies conducted with ELISA than in studies conducted with PCR.
Many studies were performed about EHV-1, 2, 4, and 5 in both healthy and unhealthy horses that different and contrary results were found. Ata et al. (2020) were found the seroprevalence of EHV-1 64% in Monufia, Egypt. However, Amer et al. (2011) investigate the seroprevalence of EHV-1,2,4 via polyclonal antibody pool technical ELISA and they found the seroprevalence was 36% in Arabian horses in Egypt. Cruz et al. (2016a) reported that the seroprevalence of EHV-1 and EHV-4 in Spanish purebred horses were 26% and 2% in Spain respectively. In contrary to this report, Aharonson-Raz et al. (2014) reported the seroprevalence of EHV-1 and EHV-4 in horses were 0.7% and 99% in Israel respectively. Although EHV-1 and EHV-4 were detected in many studies that used, no positive samples were detected in this study. In Turkey, Yildirim et al. (2015) found the seroprevalence of EHV-1 and EHV-4 in horses 52% and 83% in Kars, respectively. Ataseven et al. (2009) found the seroprevalences of EHV-1 and EHV-4%4 and 71% in horses respectively from three different regions of Turkey. In another study conducted in Turkey, Dagalp et al. (2018), 3.4%, 58.6%, 58.6% and 75.9% were found positive for EHV-1, EHV-4, EHV-2 and EHV-5, respectively. It is important to note that, all the studies which are reported high seroprevalence were performed using either PCR or ELISA. It can be suggested that the reason for the low seroprevalences of EHV-1 and EHV-4 in our study was the inclusion of random but healthy horses, the use of PCR, and both viral agents caused subclinical infections. Negussie et al. (2017) investigated the seroprevalence of EHV-1,2,4, and 5 in equids with and without clinical symptoms. In horses with clinical symptoms, seroprevalences of both EHV-1 (8.1%) and EHV-4 (7.5%) were found to be low compared to the EHV-2 (25%) and EHV-5 (28%). Furthermore, in horses without clinical symptoms, the seroprevalences of EHV-2 and EHV-5 were 7% and 17% while no horses were positive for EHV-1 and EHV-4. Similarly, Akkutay et al. (2014) investigated the seroprevalences of EHV-2 and EHV-5 in horses with and without clinical signs and they found the results of the seroprevalences 59% and 62% respectively. Krisova et al. (2020) collected peripheral blood samples and ocular swabs in horses suffering from an ocular disease to investigate EHV-2 seroprevalence and the results were found 54% and 51% respectively. In parallel with previous reports, Borchers et al. (1997) investigated the seroprevalence of EHV-2 in horses with and without clinical symptoms, and the overall seroprevalence of the virus was 62% in horses with clinical symptoms. However, the seroprevalence of the virus in horses without clinical symptoms was 42%. According to our literature search, there are fewer studies related seroprevalence of herpesviruses in donkeys. Negussie et al. (2017) found many types of herpes virus in donkeys with clinical signs and their seroprevalences were EHV-1 19%, EHV-2 4%, EHV-4 9%, and EHV-5 7%. However, asymptomatic donkeys were all seronegative. Mekonen et al. (2017) found the seroprevalence of EHV-1/EHV-4 was 74% in donkeys. Similarly, Yildirim et al. (2015) found the seroprevalence of EHV-1 and EHV-4 85% and 64% respectively in donkeys that included in the study randomly. Wondimagegnehu et al. (2021) performed a seroprevalence study about EHV-2 and EHV-5 using PCR donkeys. EHV-2 seroprevalence was found higher in horses (54%) than donkeys (4%). In contrast, EHV-5 seroprevalence was found higher in donkeys (56%) than horses (18%). Ali and Mohammadi (2016) reported that EHV-1 was not detected in any donkeys or mules however, the seroprevalence of EHV-4 was 18% in donkeys and mules. Studies performed in Turkey, Ataaseven et al. (2009) reported that EHV-1 seroprevalence was 24% in donkeys. Additionally, Yildirim et al. (2015) EHV-1 antibodies were found in 85% of investigated donkey samples, and EHV-4 antibodies were found in 64.20% of these samples. EHV-2 positive equids were had clinical signs significantly higher proportion than negative equids. As stated before, the random collection of samples or inclusion of animals with clinical signs into the study, being used ELISA more than PCR can be important factors that increase the seroprevalences of equine herpesviruses in both species.
It has been observed that the results obtained as a result of the hematological examination carried out to evaluate the health status of horses and donkeys are in accordance with the reference values specified for both species (Aiello and Moses 2016; Trimboli et al. 2020).
In this study, the seroprevalence of many parasitological and virological agents was investigated by using different laboratory technics. Most of the samples came from unregistered animals with general use ownership. In Turkey's rural areas, these types of equids are used as transport and pack animals or in traditional gaming. Piroplasmosis can be considered prevalent in the territory. Theileria equi was found highly common in donkeys and this can be important according to be an infection source for horses that show high performance. Toxoplasma gondii was found high prevalence surprisingly in donkeys. Especially donkeys have been observed living in poor managing and health conditions such as their close contact with domestic felids, their free-ranging in the around. Considering the increase in donkey milk consumption and use in recent years, it is important to know the seroprevalence of T. gondii in donkeys. Furthermore, when their meats or carcasses are consumed by wild or domestic felids it can be a serious problem for public health due to the carnivores can be shed oocysts into the environment and water sources via their feces. Similarly, N. caninum can be important especially for ruminant breeding regarding causes abortions and other economic losses. None of the prevalence of viral agents was considered critically high. However, in this study, the seroprevalence of viral agents was determined only by PCR. Besides, it can be argued that the reason why the seroprevalence of viral agents is higher in horses compared to donkeys is that the horses are raised extensively and kept close.
As a result, this study is a comprehensive study investigating both protozoal and virological diseases in horses and donkeys, and similar studies from different regions of Turkey will provide valuable information about the prevalence of the aforementioned infectious agents, which are important for human and animal health. Moreover, this research is important because it is the first in Turkey to confirm the presence of anti-Neospora sp. antibodies in donkeys, as well as seroprevalence of T. gondii in horses and donkeys on the western side of the country.