Indian Journal of Animal Research

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Indian Journal of Animal Research, volume 56 issue 11 (november 2022) : 1377-1383

Seroprevalence and Spatial Distribution of Toxoplasmosis in Relation to Various Risk Factors in Small Ruminants of Punjab, India

Deeksha Pandit1, Mandeep Singh Bal1, Paramjit Kaur1, L.D. Singla1, Vishal Mahajan2,*, R.K. Setia3
1Department of Veterinary Parasitology, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana-141 001, Punjab, India.
2Animal Disease Research Centre, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana-141 001, Punjab, India.
3Punjab Remote Sensing Centre, The Punjab Agricultural University Campus, Ludhiana-141 001, Punjab, India.
Cite article:- Pandit Deeksha, Bal Singh Mandeep, Kaur Paramjit, Singla L.D., Mahajan Vishal, Setia R.K. (2022). Seroprevalence and Spatial Distribution of Toxoplasmosis in Relation to Various Risk Factors in Small Ruminants of Punjab, India . Indian Journal of Animal Research. 56(11): 1377-1383. doi: 10.18805/IJAR.B-4358.
Background: Toxoplasma gondii is an obligatory apicomplexan, intracellular, ubiquitous, eurykaryotic zoonotic pathogen that infects all warm blooded animals and recognized worldwide as one of major causes of infectious abortions and reproductive failure in small ruminants 

Methods: In the present study, 823 small ruminants (245 sheep, 578 goats) evaluated for T. gondii infection by ELISA to explore seroprevalence and spatial distribution of toxoplasmosis from different agro-climatic zones of Punjab. 

Result: A high seroprevalence (40.58%) of T. gondii recorded in small ruminants being higher in sheep (42.85%) than goat (39.60%). Seropositivity of toxoplasmosis in sheep and goat with history of abortion was found to be 63.80 and 50.00%, respectively. Goats with the history of abortion were 1.72 times more at risk (Cl =1.15-2.56) to T. gondii infection as compared to animals without the history of abortion (Cl =0.39-0.86). Similarly, sheep with the history of abortion were 2.74 times more at risk (Cl =1.31-5.71) to toxoplasmosis as compared to sheep without abortion history (Cl= 0.17-0.76). Goat and sheep farms with frequent access to cats were 2.03 times (CI =1.05-3.90) and 4.04 times (Cl =1.78-9.14) more at risk, as compared to farms with limited cats access. Spatial distribution and predictive seroprevalence analysis indicate possible risk of toxoplasmosis in whole of the state with higher possibility of disease in agroclimatic zones III, IV and V. 
Toxoplasmosis caused by Toxoplasma gondii, is a cosmopolitan zoonosis affecting the warm blooded animals including man as intermediate hosts and cat as a definitive host (Celik et al., 2018). Medical importance of toxoplasmosis was first recognized in 1939 after diagnosed in a congenitally infected baby (Wolf, 1939). Among the three infective stages, tachyzoites are transmitted vertically through placenta, blood transfusion or milk contamination, bradyzoites by ingestion of meat from infected animal and the sporozoites by feed or water contaminated with sporulation of oocysts (Dubey and Thulliez, 1993, Singla et al., 2014).
Toxoplasma gondii is recognized worldwide as one of major causes of infectious abortions and reproductive failure in small ruminants (Dubey et al., 2011; Hussein et al., 2001). Seroprevalence of 25.3 and 30.3% for sheep and goats respectively has been reported from northern India (Chhabra et al., 1985) with scanty reports on exposure of ruminants to T. gondii infection from Punjab state (Sharma et al., 2008; Kalambhe et al., 2017). 
Toxoplasmosis can be diagnosed by direct smear, immuno-histochemistry, sero-molecular techniques, however, for epidemiological survey immunodiagnostic tests are more reliable due to the non-availability of detectable stages in secretions, excretion and blood. Among the immunological tests, ELISA is widely employed to assess the exposure of small ruminants to Toxoplasma infection as it bears high sensitivity, specificity and reliability (Puije et al., 2000).
As toxoplasmosis results in still birth and abortions in small ruminants causing severe economic losses to owners (Hussein et al., 2001), 3.27 and 1.28 lakh of goat and sheep population respectively of Punjab (Anonymous, 2012) is at risk to exposure of toxoplasmosis. Hence the aim of study was to explore the seroprevalence and spatial distribution of toxoplasmosis in small ruminants in the region.
Study area
Study was conducted in five different agro-climatic zones of Punjab state Zone I (sub-mountain undulating zone/Western Himalayas sub humid), Zone II (undulating plain zone/Northern Plain Zone Dry sub humid), Zone III (central plain zone/Northern Plain Zone sub arid), Zone IV (Western Plain Zone) and Zone V (Western Zone) (Gulati et al., 2017). Research work was conducted in Department of Veterinary Parasitology, Animal Disease Research Centre, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana from June 2018 to June 2019.
Sample collection
Sera samples (823) from goat (n=578) and sheep (n=245) collected from different villages selected by using survey tool box software, falling under different districts representing all the agro climatic zones of Punjab state. The study was approved by Institutional Animal Ethics Committee (protocol number GADVASU/2018/IAEC/46/04 (Reference No. IAEC/2018/1153-1188 Dated 28.09.2018).
Serological assay
Serum samples were evaluated by ELISA to detect anti Toxoplasma IgG antibodies using commercial ELISA kit (Prio CHECK Toxoplasma Ab SR Kit) according to manufacturer’s instructions.
Risk factors evaluation
Based on information obtained by animal owners at the time of sample collection, various risk factors (species, agroclimatic zones, age, history of abortion, access of cats to farm, grazing managements and gender) in relation to seroprevalence of T. gondii in small ruminants were evaluated.
Spatial distribution
In order to predict the disease at un-sampled locations, the data of each district was linked with district boundary in GIS and the Inverse Distance Weighted (IDW) method in Arc GIS 10.4 was used for spatial prediction of disease (Sharma et al., 2019). Inverse Distance Weighted is a type of deterministic method for multivariate interpolation with a known scattered set of points. The assigned values to unknown points were calculated with a weighted average of the values available at the known points.
Statistical analysis of data
Statistical analysis of the data was done by chi-square test to evaluate the risk factors associated with the prevalence of toxoplasmosis using SPSS 16.0 software for windows.
Seroprevalence and spatial distribution
Results revealed an overall seropostivity as 40.58% (n=334) for T. gondii antibodies in small ruminants in Punjab state. Spatial distribution of data showed highest prevalence in district Barnala (54.80%) and lowest in S.B.S. Nagar (11.76%) (Table 1, Fig 1). Species wise highest seroprevalence for T. gondii in goats and sheep was seen in districts Sangrur (58.49%) (Table 1, Fig 2) and Fazilka (100%) (Table 1, Fig 3) respectively.

Table 1: Seroprevalence of Toxoplasma gondii in small ruminants of Punjab state.


Fig 1: District wise seroprevalence of T. gondii in small ruminants.


Fig 2: District wise seroprevalence of T. gondii in goats.


Fig 3: District wise seroprevalence of T. gondii in sheep.

Zone-wise highest seroprevalence was recorded in Zone IV (45.78%) followed by Zone III (41.14%), Zone V (39.18%), Zone II (32.40%) and lowest in Zone I (25.00%) (Table 2, Fig 4). Predictive seroprevalence of toxoplasmosis at un-sampled locations based on the spatial distribution indicated small ruminant population in the districts adjoining Haryana and Rajasthan states at more risk, while animals of districts adjoining to the Himalayan region are at lower risk of toxoplasmosis (Fig 5, 6 and 7).

Table 2: Risk factors evaluation in relation to seroprevalence of T. gondii in small ruminants.


Fig 4: Zone wise seroprevalence of T. gondii in small ruminants.


Fig 5: Predictive seroprevalence of T. gondii in small ruminants.


Fig 6: Predictive seroprevalence of T. gondii in goats.


Fig 7: Predictive seroprevalence of T. gondii in sheep.

The present study, a first systematic report on spatial seroepidemiology of toxoplasmosis with overall seroprevalence of 40.58%, indicated wide exposure of small ruminants to T. gondii in Punjab. However, previous single study on seroprevalence of toxoplasmosis in 186 sheep examined from Punjab, showed low level of antibodies of T. gondii (Sharma et al., 2008). High seroprevalence of T. gondii in small ruminants from adjoining Punjab province of Pakistan (Ahmad et al., 2015) is indicative of similar geoclimatic conditions in both neighboring nations. High seroprevalence in sheep and goats, indicated that the meat-producing animals may be a great threat to human population if their meat is consumed undercooked. Further, there is possibility of economic losses to farmers as toxoplasmosis is responsible for abortion, stillbirth and neonatal mortality in sheep and goats (Dubey et al., 2011).
Associated risk factors
Sheep were at apparently higher risk of T. gondii infection (OR=1.14, 95% CI: 0.84-1.54, P=0.431) as compared to goats (OR=0.87, 95% CI: 0.63-1.19, P=0.431) with an overall seropositivity in sheep 42.85% and 39.60% in goats (Table 2). Among zones (I toV) seroprevalence ranged from 25-45.78%, being non significantly highest in zone IV.
Apparently higher seropostivity in goats of > 1 year (39.70%) (OR=1.02, CI=0.65-1.59, P=0.92) was seen as compared to < 1 year of age (OR=0.97, Cl=0.62-1.52, P=0.92). Similarly in sheep animals > 1 year of age were apparently at more risk to toxoplasmosis (OR=1.01, CI=0.57-2.10, P=0.70) as compared to animals < 1 year of age (OR=0.90, Cl=0.46-1.76, P=0.70).
Risk factor analysis indicated that goats with history of abortion were 1.72 times (OR=1.72, CI=1.15-2.56, P=0.008) and sheep were 2.74 times (OR=2.74, CI=1.31-5.71, P=0.006) more at risk to T. gondii as compared to animals with no history of abortion. The seropostivity in goats from farms having frequent access to cats was high (41.40%) to toxoplasmosis as compared to farms having limited access (25.00%) revealing 2.03 times more at risk (OR=2.03, CI=1.05-3.90, P=0.03). Similarly, in sheep higher seropostivity was observed in animals with frequent cat access (48.01%) and were 4 times more at risk to toxoplasmosis (OR=4.04, CI=1.78-9.14, P=0.01) as compared to animals belonging to farm with limited cat access.
Goats with history of grazing only, were 1.65 times higher (OR=1.65, CI=0.87-3.16, P=0.12) at risk to T. gondii infection (40.56%) as compared to animals with grazing coupled with stall feeding (29.16%). Similar pattern of seropositivity in sheep (43.77%) with history of only grazing found 1.40 times (OR=1.40, CI=0.62-3.18, P=0.42) at higher risk of toxoplasmosis as compared to other animals with grazing coupled with stall feeding (35.71%). Gender-wise more seroprevalence was observed in females in both goats (40.03%) and sheep (43.50%) as compared to males as none of the animal was found seropositive.
As in the present study sheep being more susceptible to disease as compared to goats, previous studies from Brazil (Pinheiro et al., 2009), Egypt (Ghazi et al., 2006) and Pakistan (Ahmed et al., 2016) also indicated sheep at higher risk of toxoplasmosis.  It might be due to the fact that samples from sheep were collected from grazing system, while some of sampled goats were reared in the intensive system. Overall higher seroprevalence in sheep could be due to the possibility of picking up infective oocysts from ground as compared to goats as sheep is a ground grazer and goats are browser. On the contrary, Ahmed et al., (2016) reported higher infection rate in goats as compared to sheep. 
Higher seroprevalence in Zone III, IV and V adjoining to Rajasthan and Haryana states as compared to Zone I and II adjoining to Himachal Pradesh may be related to higher temperature, lower precipitation and lower altitude compared to the absence of T. gondii at other locations (Kantzoura et al., 2013). Previous workers reported the toxoplasmosis is more prevalent in humid and damp areas than in dry and hot environmental conditions (Dubey and Thulliez, 1993, Singh and Nautiyal, 1991). It was observed that majority of small ruminant farmers in Zones III-V, were nomads, while majority of animal owners of Zones I and II reared their animals in fixed position due to availability of ample fodder. One possible reason for more seroprevalence in zones 3, 4 and 5 could be the more movement of animals in these zones and more sale and purchase of animals with adjoining states Haryana and Rajasthan. An increased in the T. gondii antibodies level with higher age in both sheep and goats could be possible as older animals are more exposed to pathogens than the younger ones (Li et al., 2016; Othman and Al-Azuheir, 2014; Puije et al., 2000). Previously similar observations were reported by Lashari and Tasawar (2010); Tasawar et al., (2011) and Ramzan et al., (2009).
Significantly higher sero-prevalence observed in animals having history of abortion indicates that T. gondii could be an important abortifacient agent in small ruminants.
Presence and frequent access of cats to small ruminant farms as a significant risk factor for higher seroprevalence may be due to fact that cats being the definitive hosts of the parasite play a vital role in infecting intermediate hosts by shedding oocysts in the environment (Lopes et al., 2010). One study from Poland revealed the presence of free-roaming cats as important risk factor for the transmission of toxoplasmosis in goats (Neto et al., 2008).
More seroprevalence observed in animals reared on grazing system corroborates with the published reports on the assessment of different type of feeding practice revealing animals being raised extensive system to be more prone to the infection (Lopes et al., 2010; Wang et al., 2011).
Higher seroprevalence of toxoplasmosis in females than males is consistent with various reports published globally (Ramzan et al., 2009; Othman and Al-Azuheir, 2014). However, on the other hand few workers (Ntafis et al., 2007; Caballero-Ortega et al., 2008) reported no association between gender and seropositivity.
High seroprevalence of T. gondii in small ruminants and history of abortion, access to cats as major risk factors at ruminant farms of Punjab state, indicate possible risk of zoonotic infection to people in the region. Spatial distribution and predictive seroprevalence data analysis also indicate possible risk of toxoplasmosis in whole of the state with higher chances of disease occurrence in districts adjoining Haryana and Rajasthan states.
Authors are thankful to Director Research, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana for providing necessary funds under RKVY-11 D-2 project to carry out the research work.
Authors declare there is no conflict of interests regarding the publication of this research paper.

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