Comparison of LFCI, LAT and ELISA for the diagnosis of toxoplasmosis in goats 

Toxoplasma gondii is an obligate intracellular protozoan parasite, belongs to phylum Apicomplexa, class Sprozoa, order Eucoccida, suborder Emmerinae and family Sarcocystidaev, which causes toxoplasmosis. This disease is zoonotic which is cosmopolitan in distribution. The main clinical sign of the disease in Goats, sheep and humans is premature and stillbirth. Cats are definitive host whereas other vertibrates are intermediate hosts of Toxoplasma (Abu-Dalbou et al., 2010; Dubey et al., 2008). Toxoplasmosis causes heavy financial harms to livestock industry worldwide (Lashari et al., 2010). Goats are economically important animals for production of meat and milk (Jittapalapong et al., 2005). In most countries, toxoplasmosis arises as the second in prevalence after chlamydial abortion (Radostits et al., 1994). There is a little Information about the seroprevalence of T. gondii in sheep in District Dir (Lower), Khyber Pakhtunkhwa, Pakistan. The aim of this study was, therefore, to investigate the prevalence of T. gondii.

Study design
 
District Dir Lower was divided into seven different areas such as Lal Qilla, Munda, Samarbagh, Balambat, Timergara, Khall and Adenzai. One hundred and seventy five blood samples were collected from goats C for All the samples was analyzed by Lateral Flow Chromatographic Immunoassay (LFCI), Latex Agglutination Test (LAT) and screened for specific IgM antibody against T. gondii using ELISA. For this purpose PrioCHECK Toxoplasma Ab SR (ELISA) kits were used with coated micro well plates having purified Toxoplasma antigen.
 
Lateral flow Chromatographic Immunoassay Technique
 
To perform this test, one drop (30.5 - 45.5 µl) of serum followed by same volume of diluent was poured into the sample well. Using this test, the coloured positive line develop after 10-15 min for both the antibodies viz. IgM and IgG separately.
 
Qualitative Technique of Latex Agglutination Test
 
The test was performed by comparing with the positive and negative controls. To perform this test, 50 µl of serum and 40 µl of Toxo latex chemical were placed on one portion of the slide and mixed for 4-5 seconds to obtain uniform suspension. Then the slide was shaken at 100 rpm and the shaking was continued for 5 minutes. Thereafter, the slide was observed under microscope for the presence or absence of agglutination.
 
Enzyme-Linked Immunosorbent Assay (ELISA) Technique
 
The samples tested by Lateral Flow Chromatographic Immunoassay and Latex Agglutination Test (LAT) were confirmed by ELISA using Prio CHECK Toxoplasma Ab SR kit to detect the specific antibodies against Toxoplasma antigen.
 
Statistical analysis
 
Bio-statistical analysis was performed by Chi-square (c2) test (Graph Pad Prism 6.01 Version).

The prevalence of toxoplasmosis was highest at Balambat followed by Khall, Samarbagh areas using LFCI technique (Table 1). There was no significant difference in the seroprevalence of toxoplasmosis among different areas of District Dir Lower, Pakistan. Moreover, the prevalence was significantly higher (P<0.05) in older goats compared to the younger ones (25.3%). However, the prevalence was non-significantly higher (P>0.05) in females compared to the males. Similar pattern was recorded by LAT and ELISA. However, the highest seroprevalence was recorded at Samarbagh followed by Balambat area using ELISA technique. The detailed results are shown in (Table 1).



Overall, 33.14%, 37.14% and 40% prevalence of toxoplasmosis was recorded in goats by LFCI, LAT and ELISA, respectively. The detailed comparative results are shown in (Table 1).

Out of 175 samples of goats tested, 58 (33.14%), 65 (37.14%) and 70 (40 %) were found positive by LFCI, LAT and ELISA, respectively.

The present study exhibited a higher seroprevalance of toxoplasmosis among goats in the region of District Dir (Lower), Pakistan. Present study reports prevalence of toxoplasmosis lower (40%) than 42% from District Mardan and 53% from Mohmand agency (Shah et al., 2013) and higher than 14% reported in Pothwar Region, Northern Punjab, Pakistan (Ahmad et al., 2015).

Various studies reported 4-81% prevalence of toxoplasmosis from other countries (Alvarado-Esquivel et al., 2011; Chandrawathani et al., 2008; Iovu et al., 2012; Jittapalapong et al., 2005; Kamani et al., 2010 and Neto et al., 2008) and supports the findings of present study.

A high seroprevalence rate of T. gondii infection was observed in females compared to the males which are corroborated with the previous reports (Etheredge et al., 2004). T. gondii infection has been reported higher in areas where the people eat raw meat, unwashed vegetables and fruits and in the people who have close contacts with cats and dogs or other domestic animals or have direct contact with the soil (Ertug et al., 2005) Toxoplasmosis is more common in those regions where people drink municipal water (Shah et al., 2013).

Present study reports serodiagnosis of toxoplasmosis with 89.74%, 95.45% and 100% specificity by LFCI, LAT and ELISA, respectively, whereas a sensitivity of 100% was recorded by all three diagnostic techniques. The seroprevalence of T. gondii infection was highest in Samarbagh (48%) area and lowest (36%) in LalQilla, Adenzai and Timergara areas of Dir district. Moreover, the prevalence was recorded higher (48.91%) in the animals above the age of one year compared to 30.12% in animals below one year. Further, the prevalence of T. gondii infection was higher (45%) in female goats compared to the males (29%).

The present study reported that the toxoplasmosis was prevalent in all age groups and in both the sexes of goats from District Dir Lower Pakistan. However, the variation was recorded with higher prevalence in females and elder goats compared to the males and younger animals. 

The authors are thankful to the Deanship of Scientific Research, King Saud University, Riyadh, Saudi Arabia, for funding the work through the research Group project No. RGP- 210.