DNA barcoding (molecular characterisation) is a useful tool for describing the taxonomy and systematics of organisms. Over 250 species of avian haemosporidian parasites have been described using morphological characters, yet molecular techniques based on polymerase chain reaction (PCR) suggest this diversity is underestimated. Moreover, molecular techniques are particularly useful for the detection of chronic infections and tissue stages of these parasites. Species delimitation is problematic among haemosporidians, and many questions about the mechanisms and patterns of speciation, host specificity and pathogenicity are still unresolved. Accumulation of additional genetic and morphological information is needed to approach these questions. Here, we combine microscopic examination with PCR-based methods to develop molecular characterisation of Haemoproteus (Parahaemoproteus) manwelli Bennett, 1978 and Haemoproteus (Parahaemoproteus) gavrilovi Valkiūnas & Iezhova, 1990, both of which parasitise the bee-eater Merops apiaster L. We also describe a new species, Haemoproteus (Parahaemoproteus) palloris n. sp., from the blood of the willow warbler Phylloscopus trochilus (L.). We performed phylogenetic analyses with a set of mitochondrial cytochrome b (cyt b) gene lineages, which have been linked to parasite morphospecies and are available in the MalAvi database. Our findings show that morphological characters, which have been traditionally used in the description of haemosporidians, exhibit phylogenetic congruence. This study contributes to a better understanding of avian haemosporidian diversity and provides new molecular markers (cyt b and apicoplast gene sequences) for the diagnostics of inadequately investigated haemosporidian infections.

Across their ranges, different populations of migratory species often use separate routes to migrate between breeding and non‐breeding grounds. Recent changes in climate and land‐use have led to breeding range expansions in many species but it is unclear whether these populations also establish new migratory routes, non‐breeding sites and migration phenology. Thus, we compared the migration patterns of European Bee‐eaters Merops apiaster from two established western (n = 5) and eastern (n = 6) breeding populations in Europe, with those from a newly founded northern population (n = 19). We aimed to relate the breeding populations to the two known non‐breeding clusters in Africa, and to test for similarities of migration routes and timing between the old and new populations. Western Bee‐eaters used the western flyway to destinations in West Africa; the eastern birds uniformly headed south to southern African non‐breeding sites, confirming a complete separation in time and space between these long‐established populations. The recently founded northern population, however, also used a western corridor, but crossed the Mediterranean further east than the western population and overwintered mainly in a new non‐breeding area in southern Congo/northern Angola. The migration routes and the new non‐breeding range overlapped only slightly with the western, but not with the eastern, population. In contrast, migration phenology appeared to differ between the western and both the northern and the eastern populations, with tracked birds from the western population migrating 2–4 weeks earlier. The northern population thus shares some spatial traits with western Bee‐eaters, but similar phenology only with eastern population. This divergence highlights the adjustments in the timing of migration to local environmental conditions in newly founded populations, and a parallel establishment of new breeding and non‐breeding sites.

Immune response to pathogens such as avian malaria (Plasmodium spp.) and related haemosporidian parasites (Haemoproteus spp. and Leucocytozoon spp.) can lead to increases in energy demands of birds. Migrations are energetically costly for birds and require resources that could be otherwise invested in immune defence against pathogens. Here, we examined the prevalence and intensity of haemosporidian parasites (Apicomplexa, Haemosporida) in rosy starlings (Pastor roseus), an irruptive migrant on the Balkans, during their pre-migratory period. We further evaluated if these infections affected the stress levels of birds brought into overnight captivity, using leukocyte profiles (ratios of heterophils to lymphocytes, or H/L ratios). Haemosporidian parasites were detected based on microscopic examination, PCR and sequencing. We found high prevalence of haemosporidian parasites (83.9%) in rosy starlings (n=31), with predominant infections of Haemoproteus pastoris (cytochrome b lineage LAMPUR01) (80.1%). All birds had high heterophil/lymphocyte ratios (mean = 3.86), indicating the transition to captivity was stressful. However, there was no added stress from blood parasite infection during captivity, since infected birds did not have significantly higher heterophil/lymphocyte ratios than uninfected birds, despite all birds facing the same stress conditions. There was a negative correlation between intensity of H. pastoris and monocyte proportions, which may indicate an association with infection, or an artefact of the stress-induced shift in leukocyte profile following captivity. Our findings suggest that blood parasites have minimal additional impact on rosy starlings during an acute stress scenario (overnight captivity), arguing that they carry little cost to the energetically challenging migratory flight.