Phlebotomine sand flies (Diptera: Psychodidae) are the principal vectors of phleboviruses and Leishmania spp., the causative agents of leishmaniases. The Mediterranean sand fly fauna is diverse, and leishmaniasis, mainly caused by Leishmania infantum, is endemic in the Balkan countries. Despite recent entomological surveys, only some districts of Kosovo have been sampled for sand flies, with no proof/confirmation of L. infantum. This study aimed to gain further insights into the species composition of natural sand fly populations in previously unsampled districts and areas in Kosovo without reports of leishmaniasis and to detect Leishmania DNA in sand flies. A sand fly survey was conducted in 2022 in all seven districts of Kosovo. Collected females were screened for Leishmania DNA by PCR. Positive samples were sequenced and subjected to maximum likelihood analysis with reference sequences for further molecular characterization. The trapping activities at 114 different localities resulted in 3272 caught specimens, comprising seven sand fly species of two genera, namely Phlebotomus neglectus, Ph. perfiliewi, Ph. tobbi, Ph. papatasi, Ph. simici, Ph. balcanicus and Sergentomyia minuta. Leishmania infantum DNA was detected in three individual sand flies of Ph. neglectus and Ph. perfiliewi. This study provides the most extensive sand fly survey in Kosovo and reports the first record of L. infantum DNA in sand flies, indicating autochthonous circulation of L. infantum.

Mosquitoes (Diptera: Culicidae) are important vectors of human and animal pathogens. Balkan countries in the southeastern part of Europe exhibit a diverse mosquito fauna and can act as a crossway for both natural and anthropogenic introduction. Some countries however lack data on mosquito fauna distribution, diversity, and pathogen circulation. mosquitoes were trapped in all seven districts of Kosovo in July and August of both 2022 and 2023, then were investigated for the presence of pathogens using PCR-based methods. A total of 144 different locations were surveyed using up to three CDC miniature light traps. Individual trapped mosquitoes were morphologically identified, then pooled, and DNA/RNA was isolated. Identification was confirmed by cytochrome oxidase I (COI) barcoding, and PCR-based pathogen screening was performed, targeting the medically relevant Filarioidea, Plasmodium and trypanosomatid protozoans as well as arboviruses (West Nile, Dengue and Zika virus). Altogether 6491 mosquitoes belonging to 18 species in six genera were trapped, identified, and screened in 1122 pools. Of 986 female pools, 21 pools (2.1%) tested positive for DNA of filarioid helminths (Dirofilaria immitis and D. repens), 52 pools (5.3%) tested positive for avian Plasmodium DNA, and 82 (8.3%) tested positive for trypanosomatid DNA. This study thus presents the most extensive survey of mosquitoes and their parasites in Kosovo to date, providing data on the prevalence of the animal and human pathogens, Filarioidea, Plasmodium and Trypanosoma.

In Europe, phleboviruses are transmitted by phlebotomine sand flies, which are widely distributed in the Mediterranean basin, including the Balkan peninsula. We report the identification and isolation of Mirusha virus (MRSHV), a novel member of the Phlebovirus corfouense species within the Phenuiviridae family, based on a > 95% genetic identity at the aminoacid level for Large segment between MRSHV and Corfou virus.

Crimean–Congo hemorrhagic fever (CCHF) is a severe tick-borne zoonotic disease associated with high case fatality rates in humans and represents a significant public health threat in endemic areas of Kosovo. However, data on viral circulation in ticks in non-endemic regions remain limited. CCHF is caused by Orthonairovirus haemorrhagiae, formerly known as the Crimean–Congo hemorrhagic fever virus (CCHFV). In the present study, 745 ticks were collected from domestic animals and the environment throughout the years 2017–2019 in seven locations of non-endemic regions in the eastern part of Kosovo, respectively Prishtina, Gjilan and Kamenica municipalities. The ticks after species identification were analyzed using RT-PCR for detection of viral RNA of CCHFV. Seven ticks (0.94%) tested positive, including six Rhipicephalus bursa ticks, three of them collected from grazing cattle and sheep in the village Hajvali (Prishtina municipality) and three others in the village Busavat (Kamenica municipality) from grazing cattle, while one positive sample was detected in a Hyalomma marginatum tick collected from a grazing dairy cow in Bresalc village (Gjilan municipality). The virus detected in Rh. bursa is considered to be consistent with the Aigia virus, formerly classified as CCHFV Clade VI/Europe 2 or the AP92-like lineage, which has generally been associated with a lower pathogenic potential and the absence of reported human cases in these areas. The first reported human CCHF case in a non-endemic area occurred in the Bresalc locality (Gjilan Municipality) in June 2018 in a dairy farmer. This occurrence coincided with the detection of Orthonairovirus haemorrhagiae RNA in one Hyalomma marginatum tick among 36 sampled ticks collected from the same farm, following clinical and laboratory confirmation of CCHF in infected patients. Consequently, this finding indicated the circulation of CCHFV in this area. These results demonstrated active circulation of CCHFV in tick populations in regions previously considered non-endemic and highlighted a potential public health risk associated with pathogenic strains. Genomic and tick surveillance, combined with targeted vector control measures and public awareness, are essential for reducing CCHFV transmission and preventing human infections.