Piroplasmosis and anaplasmosis are serious tick-borne diseases (TBDs) that are concerning for the public and animal health. This study aimed to detect the molecular prevalence and epidemiological risk factors of Piroplasma and Anaplasma species in animal hosts and their associated ticks in Egypt. A total of 234 blood samples and 95 adult ticks were collected from animal hosts (112 cattle, 38 sheep, 28 goats, 26 buffaloes, 22 donkeys, and 8 horses) from six provinces of Egypt (AL-Faiyum, AL-Giza, Beni-Suef, Al-Minufia, Al-Beheira, and Matruh). Blood and tick samples were investigated by polymerase chain reaction coupled with sequencing targeting 18S and 16S RNA genes for Piroplasma and anaplasmataceae, respectively. Statistical analysis was conducted on the potential epidemiological factors. Of the 234 animals examined, 54 (23.08%) were positive for pathogens DNA distributed among the six provinces, where 10 (4.27%) were positive for Piroplasma, 44 (18.80%) for anaplasmataceae, and 5 (2.14%) were co-infected. Co-infections were observed only in cattle as Theileria annulata and Anaplasma marginale plus Babesia bigemina, A. marginale plus B. bigemina, and T. annulata plus B. bigemina. Piroplasmosis was recorded in cattle, with significant differences between their prevalence in their tick infestation factors. Animal species, age, and tick infestation were the potential risk factors for anaplasmosis. All ticks were free from piroplasms, but they revealed high prevalence rates of 72.63% (69/95) with anaplasmataceae. We identified T. annulata, B. bigemina, and A. marginale in cattle; A. platys in buffaloes; A. marginale and A. ovis in sheep; for the first time, A. ovis in goats; and Ehrlichia sp. in Rhipicephalus annulatus ticks. Our findings confirm the significant prevalence of piroplasmosis and anaplasmosis among subclinical and carrier animals in Egypt, highlighting the importance of the government developing policies to improve animal and public health security.

Cryptosporidium and Giardia are globally significant protozoan parasites responsible for severe foodborne and waterborne outbreaks, posing substantial zoonotic and environmental risks. This study aimed to comprehensively assess the prevalence of cryptosporidiosis, giardiasis, and co-infections in Beni-Suef Governorate, Egypt, using an integrated diagnostic approach combining microscopy and molecular techniques. Additionally, it was sought to identify associated risk factors in cattle fecal samples. Microscopical examination of 970 cattle fecal samples revealed an overall infection rate of 67.42% (654/970), with Cryptosporidium detected in 42.68% (414/970), Giardia in 11.96% (116/970), and co-infections in 12.78% (124/970) of cases. In irrigation water, Cryptosporidium oocysts and Giardia cysts were detected in 2/24 (8.33%) and 1/24 (4.16%) of samples, respectively. Molecular and phylogenetic analyses identified Cryptosporidium hominis in cattle and, for the first time in Egypt, Cryptosporidium ubiquitum and Cryptosporidium ryanae in irrigation water, while also proving the presence of Cryptosporidium bovis and Giardia assemblage A in cattle. Risk factors, including sex, age, season, and fecal consistency, significantly influenced infection rates, with higher prevalence in females, calves under two months, spring season, and diarrheic feces. These findings underscore the urgent need for One Health-based control strategies, integrating targeted interventions to mitigate the burden of Cryptosporidium and Giardia infections and environmental contamination.

Background and Aim: Cryptosporidiosis is a leading cause of diarrheal disease worldwide and is an animal and public health burden. This study aimed to evaluate the protective potential of affinity-purified Cryptosporidium parvum oocyst antigen as a vaccine candidate according to fecal oocyst shedding, humoral and cellular immune responses, histopathological changes, and the number of parasite developmental stages in ileal and hepatic tissues. Materials and Methods: We isolated oocysts from naturally infected buffalo calves and identified them molecularly as C. parvum isolates (GenBank: ON730707 and ON730708) by targeting the Cryptosporidium oocyst wall protein gene. We propagated the C. parvum oocysts in mice. In addition, we prepared crude antigen from the isolated oocysts by purification using cyanogen bromide-activated Sepharose-4B affinity chromatography coupled with rabbit hyperimmune serum. Then, we divided 81 parasite-free mice into three groups: (1) non-vaccinated non-infected mice, (2) mice orally infected with 1 × 105 C. parvum oocysts on week 4 of the experiment, and (3) mice immunized twice with 40 μg/kg of the purified fraction at 2-week intervals. Then, we challenged the vaccinated group with C. parvum oocysts after 2 weeks, and the positive control group was infected at the same time. Results: We observed a prolonged prepatent period and decreased oocyst shedding in the vaccinated infected mice compared with the non-vaccinated infected mice (t < 0.001). The vaccinated mice had significantly higher immunoglobulin G levels than those in the other two groups at all examined weeks. In addition, the production of cytokines interferon-gamma, interleukin (IL)-10, IL-12, and IL-15 was activated post-vaccination. After the challenge, all tested cytokines were significantly increased (p < 0.001) in the two infected groups compared with the non-vaccinated non-infected group, with the highest levels in the vaccinated infected group. Vaccinated infected mice exhibited significantly fewer pathological lesions in the ileum and liver than non-vaccinated infected mice, which showed prominent histopathological lesions. Endogenous developmental stages of C. parvum indicated that the ileum was more parasitized than the liver and that vaccination resulted in a lower number of oocysts in ileal and hepatic tissues (p < 0.05). Conclusion: Our prepared affinity-purified vaccine candidate could be promising in protecting against cryptosporidiosis.