The objective of this study was to apply and preliminarily evaluate a High-Resolution Melting (HRM) analysis technique coupled with qPCR, that allows the simultaneous detection of 10 different ruminant abortogenic pathogens, for investigating abortions in sheep and goats throughout Greece. A total of 264 ovine and caprine vaginal swabs were obtained the week following the abortion from aborted females and analyzed using a commercially available kit (ID Gene™ Ruminant Abortion Multiplex HRM, Innovative Diagnostics). Results indicated a high prevalence of Coxiella burnetii and Chlamydophila spp., which were detected in 48.9% and 42.4% of the vaginal swabs, respectively. Results for these most commonly detected pathogens were compared with those of a well-established commercial qPCR kit, with near-perfect agreement. Toxoplasma gondii, Salmonella spp., Brucella spp., Anaplasma phagocytophilum, Campylobacter fetus, and Neospora caninum were also identified, the two latter reported for the first time in the country in small ruminants. Mixed infections occurred in 35.6% of the animals examined. This technique allows for the simultaneous detection of many abortogenic pathogens in an accurate and cost-effective assay. Detection of uncommon or not previously reported pathogens in various cases indicates that their role in ovine and caprine abortions may be underestimated.

The exposure of dairy cows to Coxiella burnetii using molecular and serological techniques was investigated in this study. Bulk tank milk and serum samples were collected from various farms in Greece (mainly northern Greece). DNA extraction was performed on milk samples, and qPCR targeting the IS1111 insertion sequence was performed to detect bacterial pathogens. An ELISA was used to detect specific antibodies in bulk milk and individual serum samples. Data on farms were collected in the field using handheld Global Positioning System Garmin units. The collected data were analyzed using an Ecological Niche Model within the framework of a geographic information system. The results indicated that in more than half of the dairy farms (35/60, 58.3%), C. burnetii is present in milk. Specific antibodies were also detected in almost all milk samples (57/60, 95.0%). At least one seropositive animal was identified using ELISA in the majority of the examined farms (25/28, 89.3%). C. burnetii PCR-positive farms were located in the low-altitude zone with a mean value of 97 m above sea level (range: 2–681). The environmental variable with the highest gain when used in isolation is precipitation in the wettest quarter (28.3% contribution), which therefore appears to have the most useful information by itself. The environmental variable that decreases the gain the most when omitted is the minimal temperature of the coldest month (6.9% contribution). The analysis demonstrated that a mild climate with low precipitation favors a positive status. The exposure of dairy cattle farms to C. burnetii is massive, raising significant concerns regarding livestock production and public health implications.

The role of Neospora caninum in ovine reproductive failures is often neglected. This study documents the first confirmed case of ovine neosporosis in Greece associated with fetal death. The investigation was prompted by reproduction problems on a farm raising Lacaune and Lacaune x Assaf dairy sheep in Northern Greece. A reduction in conception rate from 90 % to 85 % and increased incidence of embryonic losses and pyometras were recorded. During the investigation, an ewe with ultrasound (U/S)-diagnosed fetal death was blood sampled. Serology showed the presence of anti-N. caninum IgG serum antibodies using ELISA. Molecular testing for 10 common abortifacient agents resulted positive only for N. caninum DNA in the embryo's placentomes and brain. Histopathological and immunohistochemical examination of the placentomes further confirmed N. caninum as the causative agent. Our findings suggest that N. caninum infections in sheep may be underdiagnosed, emphasizing the need for veterinarians to include ovine neosporosis in their differential diagnosis of reproductive failures.