This paper presents the 2024 research results from the Danube Delta Biosphere Reserve (DDBR), including reed bed mapping and typology for fire management, assessment of sustainable use of vegetal resources, conservation status of invertebrates, fish, reptiles, amphibians, colonial waterbirds, and mammals of Community interest, evaluation of species invasiveness under climate change, and studies on human well-being and ecotourism within a sustainable development framework.

West Nile virus (WNV) is a zoonotic flavivirus whose transmission cycle in nature includes wild birds as amplifying hosts and ornithophilic mosquito vectors. Bridge vectors can transmit WNV to mammal species potentially causing West Nile Fever. Wild bird migration is a mode of WNV introduction into new areas. The Danube Delta Biosphere Reserve (DDBR) is a major stopover of wild birds migrating between Europe and Africa. The aim of this study was to investigate the presence of WNV in the DDBR during the 2016 transmission season in wild birds and mosquitoes. Blood from 68 wild birds (nine different species) trapped at four different locations was analyzed by competitive ELISA and Virus Neutralization Test (VNT), revealing positive results in 8/68 (11.8%) of the wild birds by ELISA of which six samples (three from juvenile birds) were confirmed seropositive by VNT. Mosquitoes (n = 6523, 5 genera) were trapped with CDC Mini Light traps at two locations and in one location resting mosquitoes were caught. The presence of WNV RNA was tested in 134 pools by reverse transcription quantitative PCR (RT-qPCR). None of the pools was positive for WNV-specific RNA. Based on the obtained results, WNV was circulating in the DDBR during 2016.

In the context of climate change, globalization, and enhanced human traveling, arboviruses continue to represent a threat to public health. West Nile and Usutu viruses are mosquito-borne viruses belonging to the Flaviviridae family (Flavivirus genus) and members of the Japanese encephalitis virus serocomplex. Included in the Togaviridae family (Alphavirus genus), the Sindbis virus is also vectored by mosquitoes. In the present study, we aimed to analyze the presence of antibodies concerning the abovementioned viruses in migratory and resident birds in the South-Eastern region of Romania, as avian hosts represent the main reservoir for human infection. Blood samples were collected from wild birds between May 2018 and October 2019 in nine locations from three counties. All the samples were serologically tested by ELISA and a serum neutralization test. Overall, a seroprevalence of 8.72% was registered for the West Nile virus, 2.71% for the Usutu virus, and 0% for the Sindbis virus. To our best knowledge, this is the first large-scale comprehensive study to assess the West Nile virus seropositivity in wild birds and the first serological confirmation of the Usutu virus in wild birds in Romania. Moreover, this is the only follow-up study reviewing the current seroprevalence of the Sindbis virus in Romania since 1975.

Background: In a countrywide study aiming to update the knowledge on diversity of sand fly species in Romania, a sand fly population was observed in an isolated system of cave microhabitats. The caves are located in the protected area of Canaraua Fetii, Dobrogea region, southeastern Romania. The highest sand fly diversity was recorded in this area between 1968 and 1970. This work presents a study conducted to estimate the seasonal variation of the sand fly species in correlation with the particular environmental factors of the isolated system of cave microhabitats. Methods: Sand flies were collected between May and October 2020 from one trapping site of interest in Canaraua Fetii. The trapping site consisted of a cave entrance. CDC miniature light traps and sticky traps were used to collect insects from the exterior walls of the cave entrance. Species identification of collected sand flies was done using morphological keys. Statistical analysis of the trapping and climatic data was performed. Results: From all collected sand flies, 99.7% (818/822) were Phlebotomus neglectus, 0.1% (1/822) Ph. balcanicus and 0.2% (2/822) Sergentomyia minuta. Sand fly activity was first observed on 2 July and last on 24 September. A monomodal abundance trend was present, with the peak activity between 16 and 17 July. The analysis of the climatic data showed correlations between the total number of captured sand flies and both average temperature and average relative humidity. The total number of collected specimens was statistically higher when CDC miniature light traps were used compared to sticky traps. The number of females on the sticky traps was significantly higher than the number of males on the same trap type. Compared with the sticky traps, significantly more males were collected by CDC miniature light traps. This is the first record of Se. minuta in Romania after 50 years of no records (despite the trapping effort of the last 5 years in the country). Also, Ph. sergenti, previously present in this location, was not found. Conclusions: In the investigated natural habitat, the diversity of the sand fly species appears to have changed, with the predominance of Ph. neglectus instead of Ph. balcanicus and Se. minuta (recorded as the two predominant species in 1968–1970). A monomodal abundance trend was observed as in other regions of the country. The sand fly activity in this particular cave microhabitat appears to be longer than in other regions in Romania. Longer sand fly activity increases the zoonotic risk of various pathogenic species’ transmission, with an impact on public health, as sand flies are important insect vectors. Graphical

The European mink Mustela lutreola is one of the most threatened carnivores in Europe, having suffered dramatic range contractions and severe population fragmentation. Accurate knowledge of its genetic diversity is crucial for conservation planning, yet earlier studies based on partial mitochondrial markers offered limited resolution and often underestimated haplotype richness. In this study, complete mitochondrial genomes from four extant populations (Russia, n = 11; Romania, n = 16; Germany, n = 24; France–Spain, n = 15) were analysed using a suite of non-parametric and asymptotic estimators (Fisher’s α, ACE, Jackknife1, Bootstrap, Chao1-based iNEXT) together with negative binomial modelling. A total of 41 haplotypes were detected, but extrapolated estimates indicated substantially higher richness, particularly in populations dominated by singletons. Rarefaction and extrapolation analyses revealed that sample sizes of 70–130 individuals per population are needed to approach complete haplotype detection. The France–Spain and Romania populations harboured the highest predicted richness, whereas Germany and Russia, both represented by ex situ stocks, showed lower diversity. These results refine earlier assumptions of extreme homogeneity in the Western population and demonstrate that significant mitochondrial variation persists at the continental scale. The study provides quantitative benchmarks for sampling design and genetic management, supporting preservation of evolutionary potential in this critically endangered species.

Angiostrongylus vasorum, commonly known as the French heartworm, is a metastrongyloid nematode infecting domestic and wild carnivores. While red foxes (Vulpes vulpes) are known as the primary reservoir hosts in Europe, recent studies suggest that golden jackals (Canis aureus) could also be considered as important reservoir hosts. Due to the rapid expansion of golden jackals in Europe, their role in the epidemiology of A. vasorum remains insufficiently explored. This study aimed to assess the occurrence of A. vasorum in golden jackals from Romania. Between October 2013 and November 2024, a total of 137 golden jackals were collected from 21 counties in Romania through legal hunting or roadkill. A full parasitological necropsy was performed (other data presented elsewhere), with the cardiorespiratory tract dissected for adult nematodes. First-stage larvae (L1) were isolated using the Baermann method. Morphological examination of nematodes and larvae was conducted using standard microscopy. Molecular identification of adult nematodes was carried out using PCR targeting the ITS-2 region, followed by sequencing and phylogenetic analysis. Additionally, histopathological analysis was performed on lung tissue. A. vasorum was identified in the pulmonary arteries of 2 out of 137 golden jackals (1.45 %). The presence of first-stage larvae in lung tissue confirmed their role as definitive hosts. Molecular sequencing revealed 100 % identity with A. vasorum isolated from Europe. Histopathological examination showed granulomatous pneumonia with parasitic eggs, larvae, and vascular changes associated with the infection. This study confirms golden jackals as suitable hosts for A. vasorum in Romania, indicating their potential role as reservoir hosts. Given the increasing overlap between jackals, domestic dogs, and red foxes, further surveillance is needed to understand the epidemiological implications of A. vasorum in wildlife and domestic animal health.