Phylogenetic study of Theileria ovis and Theileria lestoquardi in sheep from Egypt: Molecular evidence and genetic characterization.

BACKGROUND AND AIM: Ovine theileriosis caused by Theileria ovis and Theileria lestoquardi is an important infectious disease affecting small ruminants in regions of the tropic and subtropic zones. There is limited studies about ovine theileriosis in Egypt; so the present study aims to assess the occurrence of ovine theileriosis in Egypt at the molecular level. MATERIALS AND METHODS: Blood samples were collected from 115 randomly selected sheep, which were apparently healthy; the ages of the sampled sheep ranged from 1 to 5 years old, from a local breed (barkae and balade), and showed no symptoms indicating infection with Theileria spp. The study was conducted in three governorates representing Lower Egypt (Menoufia and Beheira) and Upper Egypt (El-Wady El-Geded). All blood samples were subjected to polymerase chain reaction (PCR) and semi-nested PCR to target Theileria spp. 18S rRNA genes. Positive samples were sequenced, and these sequences were analyzed using nucleotidebasic local alignment search tool (BLAST). RESULTS: Six animals (5.22%) were PCR-positive carriers for ovine theileriosis. Nucleotide BLAST and phylogenetic analyses of the six obtained sequences showed that T. ovis was present in five animals (4.37%) in Menoufia (n=2) and El-Wady El-Geded (n=3), whereas T. lestoquardi was detected in 1 animal (0.87%) in El-Wady El-Geded. CONCLUSION: This study is the first to provide molecular evidence, genetic characterization, and phylogenetic analysis of ovine Theileria spp. in Egypt. Specifically, T. lestoquardi and T. ovis carrier statuses of sheep were confirmed. These results highlight the importance of developing an effective control strategy against ovine theileriosis carriers that might develop and/or spread theileriosis.

New approach to molecular characterization of Paramphistomum cervi and Carmyerius gregarius and comparative analyses with selected trematodes.

The two ruminant parasites, Paramphistomum cervi and Carmyerius gregarius, were collected from fresh-slaughtered native cattle at local abattoirs in Sadat district, Menoufia province and identified morphologically, then molecularly by sequencing the nucleotides of 18S ribosomal RNA gene (18S rRNA). The nucleotide sequences of the two isolates were 456 (P. cervi) and 401 bases for (C. gregarius). The data were used along with those of several other helminth species from the GenBank to identify these two species genetically. The nucleotide sequences were aligned using multiple sequence alignments of nucleotides by Clustal W 12.1 V and construct their relationship. Neighbor-joining analytical method was used showing sister relationship between C. gregarius from Sadat district and Gastrodiscoides hominis (EF027096) with relative identity of (98%) due to the presence of single nucleotides polymorphisms (SNPs) in the form of indels as nine nucleotides positions. But when clustering of P. cervi Sadat isolate with Paramphistomoidea sp. S4 isolate P5 (GU735643), this relationship shows complete identity (99%) between them. The homology and diversity was done using Bayesian analyses in MrBayes v3.1. This work will give a useful guide for other researchers for the molecular taxonomic position of Paramphistomatidae spp. in Sadat district among the different species around the world.

An epidemiological survey of bovine Babesia and Theileria parasites in cattle, buffaloes, and sheep in Egypt.

Cattle, buffaloes, and sheep are the main sources of meat and milk in Egypt, but their productivity is thought to be greatly reduced by hemoprotozoan parasitic diseases. In this study, we analyzed the infection rates of Babesia bovis, Babesia bigemina, Theileria annulata, and Theileria orientalis, using parasite-specific PCR assays in blood-DNA samples sourced from cattle (n=439), buffaloes (n=50), and sheep (n=105) reared in Menoufia, Behera, Giza, and Sohag provinces of Egypt. In cattle, the positive rates of B. bovis, B. bigemina, T. annulata, and T. orientalis were 3.18%, 7.97%, 9.56%, and 0.68%, respectively. On the other hand, B. bovis and T. orientalis were the only parasites detected in buffaloes and each of these parasites was only found in two individual DNA samples (both 2%), while one (0.95%) and two (1.90%) of the sheep samples were positive for B. bovis and B. bigemina, respectively. Sequence analysis showed that the B. bovis Rhoptry Associated Protein-1 and the B. bigemina Apical Membrane Antigen-1 genes were highly conserved among the samples, with 99.3-100% and 95.3-100% sequence identity values, respectively. In contrast, the Egyptian T. annulata merozoite surface antigen-1 gene sequences were relatively diverse (87.8-100% identity values), dispersing themselves across several clades in the phylogenetic tree containing sequences from other countries. Additionally, the T. orientalis Major Piroplasm Surface Protein (MPSP) gene sequences were classified as types 1 and 2. This is the first report of T. orientalis in Egypt, and of type 2 MPSP in buffaloes. Detection of MPSP type 2, which is considered a relatively virulent genotype, suggests that T. orientalis infection may have veterinary and economic significance in Egypt. In conclusion, the present study, which analyzed multiple species of Babesia and Theileria parasites in different livestock animals, may shed an additional light on the epidemiology of hemoprotozoan parasites in Egypt.