Giardia duodenalis and Entamoeba spp.: Coprological and antibody-based approaches across farm animals in Egypt.

Giardia duodenalis (syn. G. lamblia, G. intestinalis) and Entamoeba spp. are enteric protozoans causing gastrointestinal illnesses in humans and animals worldwide. This study aimed to determine the prevalence of Giardia and Entamoeba infections in farm animals in Egypt, and to develop an in-house sandwich ELISA for their coproantigen detection. In this study, a total of 2674 fecal samples were collected from buffaloes (N = 875), cattle (N = 770), sheep (N = 520), and goats (N = 509) from different age groups across five governorates. All samples were examined using conventional microscopical methods. Using microscopy, Giardia and Entamoeba infections were detected in 626/2674 (23.4%) and 457/2674 (17.1%) of samples, respectively, and sheep showed the highest infection rates by both parasites (44.6% and 28.8% for Giardia and Entamoeba, respectively). Representative isolates were molecularly characterized, to confirm parasite identity prior to antigen preparation, targeting β-giardin (bg) and small subunit ribosomal RNA (ssu rRNA) gene, confirmed the presence of G. duodenalis (GenBank: OQ388331.1 , OQ388332.1 , OR029977.1 , OR029976.1 , OR029974.1 , OR029975.1 and PP316110.1 ) and E. histolytica (GenBank: OR016432.1 ). Antigens were prepared separately from these isolates, and the polyclonal antibodies (pAbs) were prepared in rabbits and purified using Protein-A-Sepharose affinity columns. Then, Anti‑Giardia IgG pAbs and Anti-Entamoeba IgG pAbs in-house sandwich ELISAs were tested separately on all fecal samples revealing infection rates of 45.2% (1208/2674) for Giardia sp. and 30.1% (805/2674) for Entamoeba sp. with high ability to detect Giardia (χ2 = 281.2, p < 0.001) and Entamoeba (χ2 = 125.6, p < 0.001) species compared to microscopy. In conclusion, the developed sandwich ELISA in our study represents a sensitive and reliable diagnostic tool for the detection of Giardia and Entamoeba infections in farm animals. Its application in routine surveillance along with complementary molecular confirmation could help to improve epidemiological accuracy and assess zoonotic risks.