Male-biased recombination at chromosome ends in a songbird revealed by precisely mapping crossover positions.

Recombination plays a crucial role in evolution by generating novel haplotypes and disrupting linkage between genes, thereby enhancing the efficiency of selection. Here, we analyze the genomes of 12 great reed warblers (Acrocephalus arundinaceus) in a 3-generation pedigree to identify precise crossover positions along the chromosomes. We located more than 200 crossovers and found that these were highly concentrated toward the telomeric ends of the chromosomes. Apart from this major pattern in the recombination landscape, we found significantly higher frequencies of crossovers in genic compared with intergenic regions, and in exons compared with introns. Moreover, while the number of recombination events was similar between the sexes, the crossovers were located significantly closer to the ends of paternal compared with maternal chromosomes. In conclusion, our study of the great reed warbler revealed substantial variation in crossover frequencies within chromosomes, with a distinct bias toward the sub-telomeric regions, particularly on the paternal side. These findings emphasize the importance of thoroughly screening the entire length of chromosomes to characterize the recombination landscape and uncover potential sex-biases in recombination.

Eggshell colour differences in a classic example of coevolved eggshell mimicry.

Avian brood parasitism is a model system for understanding coevolutionary arms races, and the great reed warbler (Acrocephalus arundinaceus, hereafter 'warbler') and its parasite the common cuckoo (Cuculus canorus, hereafter 'cuckoo') are prime examples of this coevolutionary struggle. Here, warblers select for egg colour mimicry by rejecting poorly matched cuckoo eggs. Contrary to long-held assumptions, recent work showed that warblers tend to reject lighter and browner eggs but tended to accept darker and bluer eggs rather than basing rejection decisions solely on perceived colour differences (i.e. the degree of mimicry). This counterintuitive, colour-biased rejection behaviour would select for bluer and darker cuckoo eggs, but would only be adaptive if cuckoos were consistently lighter and browner than warbler eggs. Therefore, we tested whether warbler eggs were consistently bluer and darker than cuckoo eggs. To do so, we re-analysed eggshell reflectance spectra of warblers and the cuckoos that parasitized them in the Czech Republic. As expected, we found that warbler eggs were significantly bluer and darker than the cuckoo eggs at the population level. Thus, we demonstrate imperfect mimicry in a long-coevolved cuckoo host-race and provide insights for exploring the coevolutionary interactions among hosts and their brood parasites.

The structure of songbird MHC class I reveals antigen binding that is flexible at the N-terminus and static at the C-terminus.

Long-distance migratory animals such as birds and bats have evolved to withstand selection imposed by pathogens across the globe, and pathogen richness is known to be particularly high in tropical regions. Immune genes, so-called Major Histocompatibility Complex (MHC) genes, are highly duplicated in songbirds compared to other vertebrates, and this high MHC diversity has been hypothesised to result in a unique adaptive immunity. To understand the rationale behind the evolution of the high MHC genetic diversity in songbirds, we determined the structural properties of an MHC class I protein, Acar3, from a long-distance migratory songbird, the great reed warbler Acrocephalus arundinaceus (in short: Acar). The structure of Acar3 was studied in complex with pathogen-derived antigens and shows an overall antigen presentation similar to human MHC class I. However, the peptides bound to Acar3 display an unusual conformation: Whereas the N-terminal ends of the peptides display enhanced flexibility, the conformation of their C-terminal halves is rather static. This uncommon peptide-binding mode in Acar3 is facilitated by a central Arg residue within the peptide-binding groove that fixes the backbone of the peptide at its central position, and potentially permits successful interactions between MHC class I and innate immune receptors. Our study highlights the importance of investigating the immune system of wild animals, such as birds and bats, to uncover unique immune mechanisms which may neither exist in humans nor in model organisms.

Contrasting mtDNA and microsatellite data of great reed warbler Acrocephalus arundinaceus breeding populations on a small geographic scale.

The great reed warbler has two genetically distinguishable haplogroups: "Clade A" occurs in higher proportions in Western Europe and Kazakhstan, and colonised Europe and Asia from a refugium in South-West Europe; and "Clade B", which is more common in Eastern Europe, and colonised parts of Europe from a refugium in the Middle East. Our aims were (i) to analyse the rate of differentiation in Hungarian breeding populations in order to see whether European-scale pattern is visible or not on as a small scale as the territory of Hungary and (ii) to compare the results obtained with mtDNA sequencing and microsatellite markers. To analyse the genetic differentiation, the mtDNA control region II was sequenced in 68 adult breeding birds, and 51 were fingerprinted at 11 microsatellite loci, while both analyses were performed on 36 birds (a total of 83 birds). The microsatellite data gave a better resolution and represented the fine-scale pattern of the suspected recolonisation. The lack of genetic differentiation among the breeding populations based on mitochondrial data seems to support this finding, because the admixture of the clades in this particular geographic region obliterates differentiation. Accordingly, the Fst values from different branches are significantly based on microsatellite data only. The mtDNA methods only give reliable results when a geographic and ecological factor plays a role in the population subdivision, but in the case of an intermixing population larger-scale studies are needed.

Extreme variation in recombination rate and genetic diversity along the Sylvioidea neo-sex chromosome.

Recombination strongly impacts sequence evolution by affecting the extent of linkage and the efficiency of selection. Here, we study recombination over the Z chromosome in great reed warblers (Acrocephalus arundinaceus) using pedigree-based linkage mapping. This species has extended Z and W chromosomes ("neo-sex chromosomes") formed by a fusion between a part of chromosome 4A and the ancestral sex chromosomes, which provides a unique opportunity to assess recombination and sequence evolution in sex-linked regions of different ages. We assembled an 87.54 Mbp and 90.19 cM large Z with a small pseudoautosomal region (0.89 Mbp) at one end and the fused Chr4A-part at the other end of the chromosome. A prominent feature in our data was an extreme variation in male recombination rate along Z with high values at both chromosome ends, but an apparent lack of recombination over a substantial central section, covering 78% of the chromosome. The nonrecombining region showed a drastic loss of genetic diversity and accumulation of repeats compared to the recombining parts. Thus, our data emphasize a key role of recombination in affecting local levels of polymorphism. Nonetheless, the evolutionary rate of genes (dN/dS) did not differ between high and low recombining regions, suggesting that the efficiency of selection on protein-coding sequences can be maintained also at very low levels of recombination. Finally, the Chr4A-derived part showed a similar recombination rate as the part of the ancestral Z that did recombine, but its sequence characteristics reflected both its previous autosomal, and current Z-linked, recombination patterns.

Effects of blood parasite infections on spatiotemporal migration patterns and activity budgets in a long-distance migratory passerine.

How blood parasite infections influence the migration of hosts remains a lively debated issue as past studies found negative, positive, or no response to infections. This particularly applies to small birds, for which monitoring of detailed migration behavior over a whole annual cycle has been technically unachievable so far. Here, we investigate how bird migration is influenced by parasite infections. To this end, we tracked great reed warblers (Acrocephalus arundinaceus) with multisensor loggers, characterized general migration patterns as well as detailed flight bout durations, resting times and flight heights, and related these to the genus and intensity of their avian haemosporidian infections. We found migration distances to be shorter and the onset of autumn migration to be delayed with increasing intensity of blood parasite infection, in particular for birds with Plasmodium and mixed-genus infections. Additionally, the durations of migratory flight bout were prolonged for infected compared to uninfected birds. But since severely infected birds and particularly birds with mixed-genus infections had shorter resting times, initial delays seemed to be compensated for and the timing in other periods of the annual cycle was not compromised by infection. Overall, our multisensor logger approach revealed that avian blood parasites have mostly subtle effects on migratory performance and that effects can occur in specific periods of the year only.

Gut microbiota in a host-brood parasite system: insights from common cuckoos raised by two warbler species.

An animal's gut microbiota (GM) is shaped by a range of environmental factors affecting the bacterial sources invading the host. At the same time, animal hosts are equipped with intrinsic mechanisms enabling regulation of GM. However, there is limited knowledge on the relative importance of these forces. To assess the significance of host-intrinsic vs environmental factors, we studied GM in nestlings of an obligate brood parasite, the common cuckoo (Cuculus canorus), raised by two foster species, great reed warblers (Acrocephalus arundinaceus) and Eurasian reed warblers (A. scirpaceus), and compared these with GM of the fosterers' own nestlings. We show that fecal GM varied between cuckoo and warbler nestlings when accounting for the effect of foster/parent species, highlighting the importance of host-intrinsic regulatory mechanisms. In addition to feces, cuckoos also expel a deterrent secretion, which provides protection against olfactory predators. We observed an increased abundance of bacterial genera capable of producing repulsive volatile molecules in the deterrent secretion. Consequently, our results support the hypothesis that microbiota play a role in this antipredator mechanism. Interestingly, fosterer/parent identity affected only cuckoo deterrent secretion and warbler feces microbiota, but not that of cuckoo feces, suggesting a strong selection of bacterial strains in the GM by cuckoo nestlings.

Miniaturized multi-sensor loggers provide new insight into year-round flight behaviour of small trans-Sahara avian migrants.

BACKGROUND: Over the past decade, the miniaturisation of animal borne tags such as geolocators and GPS-transmitters has revolutionized our knowledge of the whereabouts of migratory species. Novel light-weight multi-sensor loggers (1.4 g), which harbour sensors for measuring ambient light intensity, atmospheric pressure, temperature and acceleration, were fixed to two long-distance migrant bird species - eurasian hoopoe (Upupa epops) and great reed warbler (Acrocephalus arundinaceus). Using acceleration and atmospheric pressure data recorded every 5 and 30 min, respectively, we aimed at reconstructing individual diurnal and seasonal patterns of flight activity and flight altitude and thereby, at describing basic, yet hitherto unknown characteristics of migratory flight behaviour. Furthermore, we wanted to characterise the variability in these migration characteristics between individuals, species and migration periods. RESULTS: The flight duration from breeding to sub-Saharan African non-breeding sites and back was more variable within than between the species. Great reed warblers were airborne for a total of 252 flight hours and thus, only slightly longer than eurasian hoopoes with 232 h. With a few exceptions, both species migrated predominantly nocturnally - departure around dusk and landing before dawn. Mean flight altitudes were higher during pre- than during post-breeding migration (median 1100 to 1600 m a.s.l.) and flight above 3000 m occurred regularly with a few great reed warblers exceeding 6000 m a.s.l. (max. 6458 m a.s.l.). Individuals changed flight altitudes repeatedly during a flight bout, indicating a continuous search for (more) favourable flight conditions. CONCLUSIONS: We found high variation between individuals in the flight behaviour parameters measured - a variation that surprisingly even exceeded the variation between the species. More importantly, our results have shown that multi-sensor loggers have the potential to provide detailed insights into many fundamental aspects of individual behaviour in small aerial migrants. Combining the data recorded on the multiple sensors with, e.g., remote sensing data like weather and habitat quality on the spatial and temporal scale will be a great step forward to explore individual decisions during migration and their consequences.

Evidence for sexual conflict over major histocompatibility complex diversity in a wild songbird.

Sex differences in parasite load and immune responses are found across a wide range of animals, with females generally having lower parasite loads and stronger immune responses than males. Intrigued by these general patterns, we investigated if there was any sign of sex-specific selection on an essential component of adaptive immunity that is known to affect fitness, the major histocompatibility complex class I (MHC-I) genes, in a 20-year study of great reed warblers. Our analyses on fitness related to MHC-I diversity showed a highly significant interaction between MHC-I diversity and sex, where males with higher, and females with lower, MHC-I diversity were more successful in recruiting offspring. Importantly, mean MHC-I diversity did not differ between males and females, and consequently neither sex reached its MHC-I fitness optimum. Thus, there is an unresolved genetic sexual conflict over MHC-I diversity in great reed warblers. Selection from pathogens is known to maintain MHC diversity, but previous theory ignores that the immune environments are considerably different in males and females. Our results suggest that sexually antagonistic selection is an important, previously neglected, force in the evolution of vertebrate adaptive immunity, and have implications for evolutionary understanding of costs of immune responses and autoimmune diseases.

A spatiotemporal analysis of acoustic interactions between great reed warblers (Acrocephalus arundinaceus) using microphone arrays and robot audition software HARK.

Acoustic interactions are important for understanding intra- and interspecific communication in songbird communities from the viewpoint of soundscape ecology. It has been suggested that birds may divide up sound space to increase communication efficiency in such a manner that they tend to avoid overlap with other birds when they sing. We are interested in clarifying the dynamics underlying the process as an example of complex systems based on short-term behavioral plasticity. However, it is very problematic to manually collect spatiotemporal patterns of acoustic events in natural habitats using data derived from a standard single-channel recording of several species singing simultaneously. Our purpose here was to investigate fine-scale spatiotemporal acoustic interactions of the great reed warbler. We surveyed spatial and temporal patterns of several vocalizing color-banded great reed warblers (Acrocephalus arundinaceus) using an open-source software for robot audition HARK (Honda Research Institute Japan Audition for Robots with Kyoto University) and three new 16-channel, stand-alone, and water-resistant microphone arrays, named DACHO spread out in the bird's habitat. We first show that our system estimated the location of two color-banded individuals' song posts with mean error distance of 5.5 ± 4.5 m from the location of observed song posts. We then evaluated the temporal localization accuracy of the songs by comparing the duration of localized songs around the song posts with those annotated by human observers, with an accuracy score of average 0.89 for one bird that stayed at one song post. We further found significant temporal overlap avoidance and an asymmetric relationship between songs of the two singing individuals, using transfer entropy. We believe that our system and analytical approach contribute to a better understanding of fine-scale acoustic interactions in time and space in bird communities.

Why Do Migratory Birds Sing on Their Tropical Wintering Grounds?

Many long-distance migratory birds sing extensively on their tropical African wintering grounds, but the function of this costly behavior remains unknown. In this study, we carry out a first empirical test of three competing hypotheses, combining a field study of great reed warblers (Acrocephalus arundinaceus) wintering in Africa with a comparative analysis across Palearctic-African migratory songbird species. We asked whether winter song (i) functions to defend nonbreeding territories, (ii) functions as practice to improve complex songs for subsequent breeding, or (iii) is a nonadaptive consequence of elevated testosterone carryover. We found support for neither the long-assumed territory-defense hypothesis (great reed warblers had widely overlapping home ranges and showed no conspecific aggression) nor the testosterone-carryover hypothesis (winter singing in great reed warblers was unrelated to plasma testosterone concentration). Instead, we found strongest support for the song-improvement hypothesis, since great reed warblers sang a mate attraction song type rather than a territorial song type in Africa, and species that sing most intensely in Africa were those in which sexual selection acts most strongly on song characteristics; they had more complex songs and were more likely to be sexually monochromatic. This study underlines how sexual selection can have far-reaching effects on animal ecology throughout the annual cycle.

Brood parasites lay eggs matching the appearance of host clutches.

Interspecific brood parasitism represents a prime example of the coevolutionary arms race where each party has evolved strategies in response to the other. Here, we investigated whether common cuckoos (Cuculus canorus) actively select nests within a host population to match the egg appearance of a particular host clutch. To achieve this goal, we quantified the degree of egg matching using the avian vision modelling approach. Randomization tests revealed that cuckoo eggs in naturally parasitized nests showed lower chromatic contrast to host eggs than those assigned randomly to other nests with egg-laying date similar to naturally parasitized clutches. Moreover, egg matching in terms of chromaticity was better in naturally parasitized nests than it would be in the nests of the nearest active non-parasitized neighbour. However, there was no indication of matching in achromatic spectral characteristics whatsoever. Thus, our results clearly indicate that cuckoos select certain host nests to increase matching of their own eggs with host clutches, but only in chromatic characteristics. Our results suggest that the ability of cuckoos to actively choose host nests based on the eggshell appearance imposes a strong selection pressure on host egg recognition.

GENETIC SIMILARITY BETWEEN PARENTS PREDICTS HATCHING FAILURE: NONINCESTUOUS INBREEDING IN THE GREAT REED WARBLER?

The DNA-fingerprinting technique was used to find the true pedigrees and to detect the overall genetic similarity between mates of great reed warblers (Acrocephalus arundinaceus) at an isolated breeding site in Sweden. The study covered 4 yr preceded by 3 yr when almost all adults and nestlings in the study area had been banded. DNA fingerprinting revealed that the putative father had sired 97% of the young (N = 455). The mate's genetic similarity, revealed as the proportion of bands shared in restriction fragment length polymorphism (RFLP) patterns, was high compared with other species of wild birds. Also, band sharing was higher between mates native to the area than between pairs in which the female was experimentally introduced from a distant breeding site. Hatching success of eggs was negatively correlated with the degree of genetic similarity between the mates, whereas pedigree data, up to the level of great-grandparents, clearly demonstrated an absence of close inbreeding. These are the first data showing a significant fitness cost associated with the choice of a mate that has high genetic similarity, even if it is not a close kin. This cost might be caused by generalized negative consequences of genomewide inbreeding in the present study, possibly accentuated by recent population bottlenecks.