Brucellosis is a common zoonotic disease in Egypt. However, there are limited data available on the genetic diversity of brucellae circulating in Egypt and other Mediterranean areas. One hundred and nine Brucella (B.) strains were isolated from different animal species in thirteen Egyptian governorates. Multi-locus variable number tandem repeats (VNTRs) analysis (MLVA-16) was employed to determine the geographical relatedness and the genetic diversity of a panel of selected Egyptian strains (n = 69), with strains originating from Italy (n = 49), Portugal (n = 52), Greece (n = 63), and Tunisia (n = 4). Egyptian B. melitensis strains clustered into two main clusters containing 21 genotypes. Egyptian B. abortus strains clustered into three main clusters containing nine genotypes. The genotypes were irregularly distributed over time and space in the study area. Egyptian strains of B. melitensis showed MLVA-16 patterns closer to that of Italian strains. Egyptian B. abortus strains isolated from cattle share the same genotype with strains from Portugal and similar to strains from Italy with low genetic diversity. Strains with similar MLVA patterns isolated from different governorates highlight the movement of the pathogen among governorates. Hence, it may also reflect the long endemicity of brucellosis in Egypt with earlier dispersal of types and great local genetic diversity. Open markets may contribute to cross-species transmission and dissemination of the new types nationwide. The presence of West Mediterranean lineages of B. melitensis and relatedness of B. abortus strains from the studied countries is a result of the socio-historical connections among the Mediterranean countries. Transnational eradication of brucellosis in the Mediterranean basin is highly demanded.

Swine brucellosis caused by B. suis biovar 2 is an emergent disease in domestic pigs in Europe. The emergence of this pathogen has been linked to the increase of extensive pig farms and the high density of infected wild boars (Sus scrofa). In Portugal and Spain, the majority of strains share specific molecular characteristics, which allowed establishing an Iberian clonal lineage. However, several strains isolated from wild boars in the North-East region of Spain are similar to strains isolated in different Central European countries. Comparative analysis of five newly fully sequenced B. suis biovar 2 strains belonging to the main circulating clones in Iberian Peninsula, with publicly available Brucella spp. genomes, revealed that strains from Iberian clonal lineage share 74% similarity with those reference genomes. Besides the 210 kb translocation event present in all biovar 2 strains, an inversion with 944 kb was presented in chromosome I of strains from the Iberian clone. At left and right crossover points, the inversion disrupted a TRAP dicarboxylate transporter, DctM subunit, and an integral membrane protein TerC. The gene dctM is well conserved in Brucella spp. except in strains from the Iberian clonal lineage. Intraspecies comparative analysis also exposed a number of biovar-, haplotype- and strain-specific insertion-deletion (INDELs) events and single nucleotide polymorphisms (SNPs) that could explain differences in virulence and host specificities. Most discriminative mutations were associated to membrane related molecules (29%) and enzymes involved in catabolism processes (20%). Molecular identification of both B. suis biovar 2 clonal lineages could be easily achieved using the target-PCR procedures established in this work for the evaluated INDELs. Whole-genome analyses supports that the B. suis biovar 2 Iberian clonal lineage evolved from the Central-European lineage and suggests that the genomic specialization of this pathogen in the Iberian Peninsula is independent of a specific genomic event(s), but instead driven by allopatric speciation, resulting in the establishment of a new ecovar.

The first official case of bluetongue virus serotype 3 (BTV-3) in Portugal was confirmed in sheep from the district of Évora in September 2024. Notably, mortality was observed in pregnant sheepdogs within the affected sheep flocks. This study presents four cases of pregnant dogs infected with BTV-3 in mid-September 2024, all of which aborted prior to death. BTV-3 was identified by RT-qPCR following initial positive results from pan-BTV RT-qPCR. The virus was subsequently isolated from the blood of one of the dogs in BHK-21 cells, and a partial sequence of the vp2 gene was obtained. This sequence showed 100% similarity to sheep BTV3/3234/PT2024, identified in Portugal in September 2024, as well as to BTV-3/NET2023, first reported in the Netherlands in 2023. These findings suggest that the viruses may be related or share a common origin. Co-infection with common canine viruses and pathogenic bacteria was ruled out, confirming that the fatalities were due to BTV-3 infection, probably by ingestion of sheep placenta after lambing. Our results confirm the potential for the transmission of BTV-3 to non-ruminant species, particularly carnivores, and, therefore, the wider ecological implications of this virus. In addition, the identification of transplacental transmission of BTV-3 in one of the dogs provides new evidence highlighting the complexity of the virus’ transmission mechanisms.

This study investigates Brucella ceti infection in marine mammals stranded along the Lisbon and Tagus Valley coast between 2022 and mid-2024, marking the first report of Brucella presence in Portuguese waters. Out of 59 examined marine mammals, B. ceti was isolated in three common dolphins (5.1%), a prevalence rate consistent with previous studies from other coastlines. PCR-based detection indicated a higher infection rate (23.7%), suggesting an underestimation of the prevalence of B. ceti infection in this population. Multi-locus Sequence Typing (MLST) and Multiple-Locus Variable-Number Tandem-Repeat Analysis (MLVA) revealed distinct genetic profiles and close relationships to B. ceti strains from the Atlantic, supporting the hypothesis of specific host-adapted lineages in dolphins. Virulence genes, including those for host interaction (bspE, btpB) and intracellular survival (virB7, vceA), were consistent across isolates, highlighting the pathogenic potential. Additionally, antimicrobial resistance (AMR) genes, such as mprF and efflux proteins (bepC-G), were also identified. These findings underscore the need for further research and surveillance to understand B. ceti transmission, host range, and impacts on Atlantic cetaceans, as well as to develop effective diagnostic and management strategies to mitigate infection risks in marine environments.