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BackgroundAdult fleas are haematophagous ectoparasites of warm-blooded vertebrates, particularly mammals. Among them, the cat flea (Ctenocephalides felis) and the human flea (Pulex irritans) have high veterinary-medical significance, owing to their cosmopolitan distribution and role in the transmission of important vector-borne pathogens. While the taxonomy of Ct. felis has been investigated on a morphological basis during the past decades, its molecular-phylogenetic analyses have been only recently conducted. This study expands the knowledge on Ct. felis from hitherto less studied geographical regions, and includes representatives from additional flea families, less investigated with molecular approaches.MethodsFleas were collected in four countries of the Mediterranean Basin (Croatia, Italy, Malta and Israel), as well as in Hungary, from domestic and wild carnivores, rodents and humans. The DNA extracts of representative fleas (n = 148), belonging to ten species of eight genera, were used for PCR amplification of part of their cytochrome c oxidase subunits 1, 2 (cox1, cox2) and 18S rRNA genes, followed by sequencing and phylogenetic analyses.ResultsThe majority (65.6%) of Ct. felis felis cox2 sequences showed 99.4–100% similarity to each other (haplogroup A), whereas those from Malta and Israel had 98.1–98.7% sequence similarity (haplogroup B), and a third sequence from Israel (haplotype C) had as low as 96.3% sequence similarity in comparison with a reference sequence from group “A”. Except for the shape of the head, no consistent morphological differences (e.g. in chaetotaxy) were found between haplogroups “A” and “C”. Haplotypes of Ct. canis were genetically more homogenous, with 99.6–100% sequence similarity to each other. However, when P. irritans collected from humans was compared to those from three species of wild carnivores, these only had 96.6% cox2 similarity. The mouse flea, Leptopsylla segnis and the northern rat flea, Nosopsyllus fasciatus were both shown to have haplotypes with low intraspecific cox2 similarities (96.2 and 94.4%, respectively). Taken together, differences between mitochondrial lineages within four flea species exceeded that observed between two Chaetopsylla spp. (which had 97.3% cox2 similarity). The topologies of cox1 and cox2 phylogenetic trees were in line with relevant sequence comparisons. Conversely, 18S rRNA gene analyses only resolved differences above the species level.ConclusionsCtenocephalides felis felis, P. irritans, L. segnis and N. fasciatus were shown to have such a high level of mitochondrial gene heterogeneity, that the uniformity of these flea taxa should be reconsidered. Although the present results are limited (especially in the case of L. segnis and N. fasciatus), there appears to be no geographical or host restriction, which could explain the divergence of these genetic lineages.Electronic supplementary materialThe online version of this article (10.1186/s13071-018-2798-4) contains supplementary material, which is available to authorized users.

Increasing amount of data attest that (in the context of vector-borne infections) birds are not only important as hosts of blood-sucking arthropod vectors, but also as reservoirs of vector-borne pathogens. From 2015 to 2019 cadavers of 100 birds (from 45 species, nine orders) were collected in Hungary, and their organs were screened for DNA from a broad range of vector-borne bacteria with PCR and sequencing. Molecular analyses revealed the presence of Anaplasmataceae, and sequencing identified bacteria closely related to Neorickettsia helminthoeca and Ehrlichia chaffeensis in a Eurasian teal (Anas crecca) and a song thrush (Turdus philomelos), respectively. All samples were PCR negative for rickettsiae, borreliae, Francisella and Coxiella spp., as well as for piroplasms. To our knowledge, this is the first report of a Neorickettsia and an Ehrlichia sp., which belong to the phylogenetic groups of N. helminthoeca and E. chaffeensis, respectively, from Europe. The potential presence of these two vector-borne bacteria needs to be taken into account during future studies on the eco-epidemiology of Anaplasmataceae in Europe.

Reports of Sarcocystis rileyi-like protozoa (‘rice breast disease’) from anseriform birds had been rare in Europe until the last two decades, when S. rileyi was identified in northern Europe and the UK. However, despite the economic losses resulting from S. rileyi infection, no recent accounts are available on its presence (which can be suspected) in most parts of central, western, southern and eastern Europe. Between 2014 and 2019, twelve mallards (Anas platyrhynchos) were observed to have rice breast disease in Hungary, and the last one of these 12 cases allowed molecular identification of S. rileyi, as reported here. In addition, S. rileyi was molecularly identified in the faeces of one red fox (Vulpes vulpes). The hunting season for mallards in Hungary lasts from mid-August to January, which in Europe coincides with the wintering migration of anseriform birds towards the south. Based on this, as well as bird ringing data, it is reasonable to suppose that the first S. rileyi-infected mallards arrived in Hungary from the north. on the other hand, red foxes (Vulpes vulpes), which are final hosts of S. rileyi, are ubiquitous in Hungary, and our molecular finding confirms an already established autochthonous life cycle of S. rileyi in the region. Taken together, this is the first evidence for the occurrence of S. rileyi in Hungary and its region.