This study aimed to direct detection of Mycobacterium avium subsp. paratuberculosis (MAP) in milk by evaluating a multiplex real-time PCR assay targeting IS900 and ISMAV2 sequences including the amplification of PUC19-plasmid as internal control. The sensitivity of the assays was evaluated by testing MAP isolates in broad linear range of DNA (50 ng-5 fg/µl). For the validation of the specificity, 6 MAP isolates and 22 isolates of genus Mycobacteriacea were tested. Results revealed that reproducible detection limit for real-time PCR targeting IS900 and ISMAV2 was 5 fg/µl and 50 fg/µl respectively. By targeting ISMAV2 sequence, 100% specificity was detected. However, a cross reaction with 5 ng/µl of genome of 3 M. avian subspecies avium strains was detected by targeting IS900 and negative in lower genome quantity (5pg/µl). To maximize the assay's detection sensitivity, an efficient strategy for MAP-DNA extraction from spiked milk was assessed. Targeting of IS900 was sensitive and targeting ISMAV2 was very specific. Therefore, a multiplex real-time PCR assay targeting IS900 and ISMAV2 in combination with two commercial DNA extraction kits could be an ideal sensitive and specific protocol for routine large scale analysis of milk samples and other clinical specimens from man and animals.

To understand how the latest dominant bovine leukemia virus (BLV) strains were introduced and spread in the Miyazaki prefecture, we collected blood samples from 3 geographic areas (north, central and south) and carried out sequence analysis of the BLV env gene. Two genotypes, genotype I, and III, were identified and the majority of the strains belonged to genotype I (71/74). To clarify a route of BLV introduction, we divided the strains into 20 subgenotypes based on their nucleotide sequences and performed phylogenetic analysis. Our study indicated that common BLV strains were comparatively evenly distributed even in the area, where the farmers have not introduced cattle from other areas and the cattle have limited exposure to BLV infection in grazing fields.

No abstract

Ehrlichia canis (E. canis) is a tick-borne disease, affect dogs in different areas. Traditional diagnostic techniques including hematology, cytology and isolation are valuable diagnostic tools for E. canis, however a definitive diagnosis of E. canis infection requires molecular investigation techniques. An epidemiological study of E. canis infection among dogs in Egypt was carried out using microscopic examination and molecular tools techniques. A total of 170 canine blood samples were collected from different breed of dogs admitted to veterinary clinics in three governorates (Cairo, Giza and Qalyubia). Microscopic examination revealed that 25 out of 170 (14.7%) was positive for E. canis. Molecular screening by polymerase chain reaction (PCR) was performed using genus-specific primers followed by PCR using E. canis speciesspecific primers. PCR assay can be detected 27 out of 170 dogs for E. canis from examined animals. These results emphasize that serological and molecular studies are needed to clarify the epidemiological feature of the infection in different governorates of Egypt.

Sarcocystis is an intracellular protozoan parasite in the phylum Apicomplexa. It is widely distributed all over the world. There are scarce reports about chicken Sarcocystis. From February 2016 to January 2018, a total number of 630 chicken carcasses, intestines and viscera were collected from different chicken markets in Menoufia and Gharbia Governorates, Middle region of the Nile Delta, Egypt and carefully inspected. Macroscopic and microscopic cysts of Sarcocystis spp. were found in the intestinal wall and mesentery of 5 birds. Histopathological sections revealed the presence of two shapes of the macroscopic cysts (oval and kidney shape). Their wall was striated and characterised by the presence of radial septa. It had compartments mostly of hexagonal shape, containing both bradyzoites and metrocytes in the periphery. The bradyzoites were banana-shaped and measured 20–30 × 8–10 μm with centrally or posteriorly located nuclei. Microscopic cysts of Sarcocystis spp. were detected in-between muscle bundles, with variable shapes (spindle and oval).