Molecular detection and phylogenetic analysis of Neospora caninum in various hosts from Iran

Different N. caninum strains with distinct biological characteristics and pathogenicity have been introduced from various areas of the world. This study aimed to detect Neospora-infection in different hosts from Iran using the nested-Polymerase Chain Reaction (PCR) technique and to perform the phylogenetic analysis of N. caninum isolates using the Nc-5 gene. Different species of parasite hosts (cattle, dogs, rodents, cats, and birds) from the dairy farms with history of abortion with intensive breeding systems in Hamedan Province, Iran were included for sampling. The stillbirth fetuses of cattle, sheep, and goats were also collected from the slaughterhouse. The nested-PCR technique was used to identify the Neospora-DNA in the aborted fetal materials. The PCR products of the positive samples were sequenced and compared with other isolates in GenBank. Antibodies to N. caninum was found in 19.1% of dogs. Also, 11.4% of dogs were positive for Neospora-like oocysts using microscopy method. The molecular prevalence of N. caninum was 10.6% in dogs, 13.6% in cattle, 35.4% in the aborted fetuses of cows, 31.9% in rodents, 7.6% in pigeons, 1.9% in sparrows, 5.1% in cats, and 7% in the semen sample of bulls. A sampled coyote was positive while the stillbirth fetuses of sheep and goats were negative. The multiple sequence alignments indicated that 34 sequences of N. caninum bore a 94.9-99.3% similarity to each other. Hence, these sequences were classified into two major clusters (clusters A and B) and seven sub-groups (isolates 1-7). The sequence of coyote had a high similarity to the sequences derived from dogs and cows. Also, root diagram showed the isolates of current study and previous reported from Iran have a similar origin. This study provided a phylogenetic analysis of Neospora-infection in different hosts, and for the first time, explored Neospora-infection in coyotes from Iran. Different isolates were detected in the studied population in the region. Comparison of the sequences showed the farm dogs play an important role in the life cycle and distribution of the parasite; so the relationship of dogs with the herd and their access to aborted materials should be restricted. Also, regular control of rodents' population and culling of seropositive cows with a history of abortion is essential for cutting the chain of transmission.