The early and accurate diagnosis of brucellosis, a ubiquitous zoonotic infection, is significant in preventing disease transmission. This study aimed to assess the infection rate of Brucella spp. in ruminants and to evaluate the agreement between a serological test and a molecular method for the detection of infected cases. Blood and milk samples of 136 ruminants were analyzed using two laboratory methods: the Rose Bengal plate (RBP) test to detect B. abortus and B. melitensis antibodies and the molecular polymerase chain reaction (PCR) method for the presence of bacterial DNA. The agreement between the methods was assessed using the kappa statistic. Based on the RBP test, there were 12 (8.8%) seropositive animals (10 sheep and 2 cows), while 2 (1.4%) samples were positive on PCR analysis. The positive PCR samples were from seronegative cow samples on RBP testing. There was slight agreement (k = −0.02) between the two methods, which was not statistically significant. Our results indicate that complementary molecular methods are useful to detect the bacteria in infected animals that are seronegative due to an early stage of infection. Therefore, a combination of molecular methods and serological tests can be applied to detect brucellosis in ruminants efficiently.

Brucellosis is one of the most important zoonotic diseases in Greece, causing a significant burden on both human and animal vitality as well as economic loss. The present study was conducted from 2015 to 2022 on 711,415 serum samples by determining the seroepidemiology of Brucellosis among livestock in 24 geographical areas in Greece using the Rose Bengal Test (RBT) and the complement fixation test (CFT) and further performing genetic analysis of Brucella spp. by species-specific real-time PCR and MLVA Brucella analysis. A total of 3086 serum samples from goats, sheep, and cattle showed positive results using the RBT and CFT, and only strongly positive samples (n = 800) were preserved in the Βlood Bank of the Veterinary Laboratory of Brucellosis. From these, 212 sera samples were randomly selected for molecular and genetic analysis. The results indicated that the incidence rate of Brucellosis is higher in cattle herds in comparison with other animal species. Overall, 48 samples tested positive by real-time PCR, of which forty-seven of them were B. abortus and one was B. melitensis. Genetic analysis of two B. abortus samples revealed a common pattern, indicating two Bruce04, two Bruce18, four Bruce07, two Bruce09, three Bruce16, and four Bruce30 for both samples, which, interestingly, were not identical with the known genotypes in the public MLVA Brucella database. Our findings substantiate that animal Brucellosis remains a health issue in Greece, with a stable but apparent incidence rate, and further investigation is needed to fully characterize the newly identified Brucella strains in Greece.

Ticks are temporary ectoparasites that serve as vectors for a wide range of pathogens affecting both wildlife and humans. In Greece, research on the prevalence of tick-borne pathogens in wildlife is limited. This study investigates the presence of pathogens, including Anaplasma spp., Babesia spp., Bartonella spp., Rickettsia spp., as well as tick-borne encephalitis (TBE), and Crimean–Congo hemorrhagic fever (CCHF) viruses, in ticks collected from 28 wild animals across 10 different animal species. Ticks were manually harvested and identified using molecular methods, with sequencing confirming the presence of Hyalomma aegyptium, H. marginatum, H. anatolicum, Ixodes frontalis, and I. ventalloi. Among the pathogens detected, R. aeschlimannii was the most prevalent, particularly in H. aegyptium ticks from tortoises. Additionally, R. africae was identified in H. aegyptium from tortoises, marking the first report of this pathogen in this tick species in Greece. Hemolivia mauritanica, an apicomplexan parasite commonly found in Testudo tortoises, was also detected. No evidence of Babesia spp., Anaplasma spp., Bartonella spp., or viral nucleic acid was found. Phylogenetic analysis revealed close genetic relationships between the detected Rickettsia species and those previously reported in neighboring regions. These findings underscore the role of wildlife in the epidemiology of tick-borne pathogens in Greece and highlight the need for comprehensive surveillance to prevent future outbreaks.

Wild birds are increasingly recognized as contributors to the circulation and environmental dissemination of zoonotic pathogens, yet data from Greece remain limited, particularly for raptors, corvids, and water birds. This study investigated selected parasitic, mycotic, and bacterial pathogens of public health relevance in free-living wild birds originating from various regions of Greece and admitted to two animal health care facilities. Between November 2023 and May 2025, cloacal swabs from 212 injured or sick birds were analyzed using quantitative PCR for Cryptococcus spp., Chlamydia psittaci, Giardia duodenalis, Cryptosporidium spp., and Mycobacterium avium. At least one pathogen was detected in 37 samples (17.5%), with Cryptococcus spp. being the most frequent agent (11.8%), followed by C. psittaci (3.8%), G. duodenalis (0.9%), and Cryptosporidium spp. (0.9%). M. avium was not detected. Pathogen occurrence varied by bird category, genus, and region, and was associated with health status. To the best of our knowledge, this study provides the first molecular evidence worldwide of G. duodenalis in Ardea spp., Cryptosporidium spp. in Asio otus, and C. psittaci in Botaurus stellaris and Plegadis falcinellus. These findings highlight wild birds as potential zoonotic reservoirs and support the implementation of One Health–oriented surveillance programs.