Protein Profile and Seroprevalence of Fasciola gigantica in Ruminants in the Sudan

Abstract

The current study was designed to study the protein profile (antigenic structure) of adult Fasciola gigantica excecretory-secretory and somatic products from sheep and goats of the same locality (White Nile province) and from three cattle types (White Nile, Mangisto and Niyala types) from different localities in Sudan, using Sodium dodecyle sulphate polyacrylamide gel electrophoresis (SDS-PAGE). This antigenic analysis was applied for detection of immunoreative protein that could be used for early diagnosis and establishment of protective vaccine. Furthermore, indirect ELISA (enzyme-linked immunosorbent assay) was evaluated for its effectiveness in immunodiagnosis of Fasciolosis in sheep and cattle in the area of Rabbak, White Nile Province, Sudan. It was also applied to compare ES Paramphistomum spp. extract versus ES and SO F. gigantica products in both sheep and cattle, to determine the possibility of cross reaction between the two parasite antigens. Generally SDS-PAGE analysis showed the presence of several bands with lower molecular weights ranging from 12-95 KDa in the Fasciola ES products comparing with 14-123KDa in the somatic one.Dominant common bands for both ES and SO extracts were clustered between 27-30 KDa.Major common bands were identified in SO extracts of F.gigantica in sheep, goats and cattle at 45 and 66. Moreover,three common major bands with approximate molecular weight of 27-30, 45 and 66 KDa were found in somatic preparation of the parasite from the three different cattle types. >17, 57, 95 and 110 KDa were only seen in Niyala cattle type, while Mangisto type showed five polypeptide bands with specific bands above 95 and 123 KDa. Only one specific band (<80) was seen in White Nile type.F.gigantica somatic antigens in both sheep and goats revealed 4 common dominant bands 14, 28, 45 and 66 KDa and three goat specific bands 19, 38, 50 KDa. The dominant of F.gigantica ES extracts from sheep, goats and cattle were clustered between 17-24, 27-33 and 40 KDa.The electrophoretic migration showed some similarities and some differences between ES antigen extracts of F.gigantica existing in sheep, goats, and cattle. Twelve bands with molecular weight of 14, 17, 19, 21, 24, 28, 31, 33, 40, 55, 62 and 72 KDa were identified in sheep, whereas eleven bands with molecular weights of >17, 17, 20, 23, 27, 33, 40, 45, 80, 87, and 95 KDa were shown in cattle. On the other hand, nine bands with molecular weights of 12, 14, 17, 19, 21, 24, 40, 55 and 72 were found in goats. Considering this result, we noticed that the highest molecular weights 80, 87 and 95 of F. gigantica ES antigen were recorded only in cattle. One hundred fifty six cattle faecal samples were examined, 30 were found positive with Fasciola gigantica with an overall prevalence of 19.2%. The results of indirect ELISA revealed that higher prevalence was detected by ES Ag (30%) comparing with SO Ag (18.7%). High specificity (81.6%) was recorded using SO antigen in cattle compared with 69% using ES antigen. Meanwhile, low sensitivities of 27.7% and 20% were recorded in both ES and SO antigens, respectively.Twenty one out of ninety two sheep faecal samples examined were found positive with Fasciola gigantica with overall prevalence rate of 22.8%. Moreover, the overall seroprevalence of Fasciola was 22.8% (21/92) and 20.6% (19/92) as assessed by Fasciola gigantica ES and SO antigens, respectively.Higher specificity in sheep was recorded using SO Ag (83%) (59/71) compared with ES Ag (78.9%). However, low sensitivity of 33% and 28.6% was determined when SO and ES antigens were used, respectively. From these results, we concluded that Indirect ELISA using both ES and SO Fasciola antigens in sheep and cattle was not sensitive, although it showed slightly high specificity. A remarkable observation occured in the current study is that the number of sheep and cattle negative to Fasciola ES and SO indirect ELISA and positive to coprological examination was high. In sheep 14 and 15 sera out of 21 positive faecal samples, while in cattle 22 and 24 sera out of 30 positive faecal samples resulted negative, respectively. In cattle by using ES Fasciola gigantica antigen versus Paramphistomum faecal analysis, we observed that 17 sera out of 46 positive faecal Paramphistomum had positive values by Fasciola indirect ELISA. Similarily, in sheep considering both ES and SO Fasciola antigens, 2 sera out of 13 positive Paramphistomum faecal samples were positive to Fasciola gigantica. In addition, indirect ELISA tests using both paramphistomum and Fasciola ES antigens showed that among the 61 Paramphistomum positive sera, 26 sera were found positive for Fasciola. Similarly, the numbers of serologically positive Fasciola samples were 47. Among them 26 were found positive for Paramphistomum. Meanwhile, in sheep the results detected by indirect ELISA showed that among the 20 sera positive for Fasciola using ES, 4 sera were found to be positive for Paramphistomum. Similarly, among the 17 sera positive for SO Fasciola, 8 were found positive for Paramphistomum. So from these results we concluded that cross reaction between Fasciola gigantica and Paramphistomum spp. may exist. ES extract for Paramphistomum was used to evaluate the diagnostic sensitivity and specificity of indirect ELISA for the diagnosis of Paramphistomiasis as well as its prevalence in cattle and sheep.The prevalence of Paramphistomiasis in cattle based on indirect ELISA was higher than that detected by faecal analysis. However, We noted that the prevalence of Paramphistomiasis in sheep was higher by coprological examination (13.8%) than that detected by Indirect ELISA (11.5%).The sensitivity of indirect ELISA in both cattle and sheep was found to be 43.4% and 8.3%, whereas, the specificity was 62.7% and 88%, respectively. This result indicating that indirect ELISA for detection of Paramphistomum spp. is more specific but not sensitive.