The objective of this study is to prospectively evaluate the seroepidemiology of maedi-visna (MV) infections in intensively reared dairy sheep. A total of 407 purebred Chios and Lacaune ewes from four farms were surveyed for two consecutive years and were serologically tested semiannually with an indirect ELISA at pre-mating and pre-lambing. The farms’ structure and management practices were similar and animal traits (age, breed, and production stage) were recorded. Based on the serological status, morbidity frequency measures were estimated, and ewes were categorized as constantly seronegative, constantly seropositive, seroconverted, seroreverted, or as animals with an intermittent presence of antibodies. During the study, period seroprevalence, incidence rate, and cumulative incidence were 84.8%, 33.6 new cases per 100 sheep-semesters, and 64.2%. Point-seroprevalence ranged from 48.5% to 96.0% among the studied farms and sampling occasions, and they increased by age. Increased morbidity frequency measures indicate the significance of horizontal transmission in intensive dairy sheep farms. A remarkable percentage of infected animals seroreverted (8.1%) or presented an intermittent presence of antibodies (10.3%) during the study, confirming the risk of misdiagnosis in cross-sectional studies and in the currently implemented testing and elimination programs. The serological patterns observed in our study need to be considered when studying MV epidemiology and for the designing of efficient MV elimination programs.

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 incidence of small ruminant pestivirus infections in Greece remains unknown as they have not been diagnosed in the country since 1974 when the most recent Border Disease Virus (BDV) outbreak was reported. The objective of our study was to explore the possible occurrence of pestiviral infections among sheep and goat farms in Greece and to further determine the variants of major concern. Thus, serum samples were collected from 470 randomly selected animals belonging to 28 different flocks/herds. ELISA on p80 antibody revealed the existence of seropositive animals in four out of the 24 studied sheep flocks, whereas all the goats in the four studied herds were seronegative. Viral RNA and antigens were detected in two sheep out of the four seropositive flocks by RT-PCR and ELISA, respectively. Sequencing and phylogenetic analysis showed that the newly identified Greek variants were closely related to the strains of the BDV-4 genotype. One of the BDV-positive sheep demonstrated the diagnostic profile of a persistently infected (PI) animal, providing additional information regarding the source of the infection. This is the first molecular identification of BDV isolates in Greece. Our findings indicate that BDV infections are likely to remain undiagnosed, highlighting the need for further epidemiological studies and active surveillance programs to determine the prevalence and impact of BDV infections on a countrywide level.

Small ruminant lentiviruses are a group of viruses infecting goat and sheep worldwide. These viruses exhibit an extraordinary degree of genetic and antigenic variability that severely influence in vivo and in vitro features, as well as diagnostic test results. Small ruminant farming is the most important animal farming business in Greece, with a high impact on the Greek primary economy. Although SRLV infection and its impact on animal production are well established in the country, little is known about the circulating SRLV strains and their prevalence. The aim of this study was to characterize SRLVs circulating in Greece with a combined serological and molecular approach, using the bulk milk matrix collected from 60 farms in different municipalities. This study allowed us to estimate a seroprevalence of around 52% at the herd level. The B1, B2 and A3 subtypes and a novel A viral cluster were identified. Moreover, the amplicon sequencing method allowed us to identify more than one viral subtype in a sample. These results again confirm the high variability of these viruses and highlight the importance of the constant monitoring of viral evolution, in particular in antigens of diagnostic interest.

A two-year longitudinal cohort study was conducted on a total of 407 purebred Chios and Lacaune ewes from four intensive dairy sheep farms to assess potential risk factors for small ruminant lentiviruses (SRLVs) seropositivity. Ewes were serologically tested semiannually at pre-mating and pre-lambing, and their age, breed, and body condition score (BCS) were recorded. Εwes were categorized as constantly seronegative, constantly seropositive, seroconverted, seroreverted, or animals with an intermittent presence of antibodies. Mixed binary logistic regression models were used to estimate the adjusted relative risks of the studied risk factors for (i) the individual ewes’ seropositivity, (ii) the manifestation of specific serological patterns, and (iii) the occurrence of seroconversion and seroreversion incidents. Increased age was associated with seropositivity and constantly seropositive status (p < 0.001 in both cases). On the other hand, age was negatively associated with constantly seronegative pattern, seroconversion incident, and the intermittent presence of antibodies (p < 0.05 in all cases). Moreover, breed was recognized as a risk factor: Lacaune ewes demonstrated increased seropositivity, whereas Chios ewes were more likely to demonstrate an intermittent presence of antibodies (p < 0.01 in both cases). Seropositive status (p < 0.001), seropositivity in animals with an intermittent presence of antibodies (p = 0.001), and seroconversion incidents (p < 0.001) were significantly increased at pre-lambing compared to pre-mating. The risk factors recognized in our study contribute to a better understanding of SRLVs epidemiology and the evidence-based designation of SRLVs’ control programs in intensive dairy sheep farms in Greece.

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.