Population Genomics and Structure of the Critically Endangered Mariana Crow (Corvus kubaryi).

The Mariana Crow, or Åga (Corvus kubaryi), is a critically endangered species (IUCN -International Union for Conservation of Nature), endemic to the islands of Guam and Rota in the Mariana Archipelago. It is locally extinct on Guam, and numbers have declined dramatically on Rota to a historical low of less than 55 breeding pairs throughout the island in 2013. Because of its extirpation on Guam and population decline on Rota, it is of critical importance to assess the genetic variation among individuals to assist ongoing recovery efforts. We conducted a population genomics analysis comparing the Guam and Rota populations and studied the genetic structure of the Rota population. We used blood samples from five birds from Guam and 78 birds from Rota. We identified 145,552 candidate single nucleotide variants (SNVs) from a genome sequence of an individual from Rota and selected a subset of these to develop an oligonucleotide in-solution capture assay. The Guam and Rota populations were genetically differentiated from each other. Crow populations sampled broadly across their range on Rota showed significant genetic structuring ⁻ a surprising result given the small size of this island and the good flight capabilities of the species. Knowledge of its genetic structure will help improve management strategies to help with its recovery.

Sequencing of the complete mitochondrial genome of the common raven Corvus corax (Aves: Corvidae) confirms mitogenome-wide deep lineages and a paraphyletic relationship with the Chihuahuan raven C. cryptoleucus.

Previous studies based on single mitochondrial markers have shown that the common raven (Corvus corax) consists of two highly diverged lineages that are hypothesised to have undergone speciation reversal upon secondary contact. Furthermore, common ravens are paraphyletic with respect to the Chihuahuan raven (C. cryptoleucus) based on mitochondrial DNA (mtDNA). Here we explore the causes of mtDNA paraphyly by sequencing whole mitochondrial genomes of 12 common ravens from across the Northern Hemisphere, in addition to three Chihuahuan ravens and one closely related brown-necked raven (C. ruficollis) using a long-range PCR protocol. Our raven mitogenomes ranged between 16925-16928 bp in length. GC content varied from 43.3% to 43.8% and the 13 protein coding genes, two rRNAs and 22 tRNAs followed a standard avian mitochondrial arrangement. The overall divergence between the two common raven clades was 3% (range 0.3-5.8% in 16 regions including the protein coding genes, rRNAs and the control region). Phylogenies constructed from whole mitogenomes recovered the previously found mitochondrial sister relationship between the common raven California clade and the Chihuahuan raven (overall divergence 1.1%), which strengthens the hypothesis that mtDNA paraphyly in the common raven results from speciation reversal of previously distinct Holarctic and California lineages.

The origin and phylogenetic relationships of the New Zealand ravens.

The relationships of the extinct New Zealand ravens (Corvus spp.) are poorly understood. We sequenced the mitogenomes of the two currently recognised species and found they were sister-taxa to a clade comprising the Australian raven, little raven, and forest raven (C.coronoides, C. mellori and C. tasmanicus respectively). The divergence between the New Zealand ravens and Australian raven clade occurred in the latest Pliocene, which coincides with the onset of glacial deforestation. We also found that the divergence between the two putative New Zealand species C. antipodum and C. moriorum probably occurred in the late Pleistocene making their separation as species untenable. Consequently, we consider Corax antipodum (Forbes, 1893) to be a subspecies of Corvus moriorum Forbes, 1892. We re-examine the osteological evidence that led 19th century researchers to assign the New Zealand taxa to a separate genus, and re-assess these features in light of our new phylogenetic hypotheses. Like previous researchers, we conclude that the morphology of the palate of C. moriorum is unique among the genus Corvus, and suggest this may be an adaptation for a specialist diet.

Discovery of species-wide tool use in the Hawaiian crow.

Only a handful of bird species are known to use foraging tools in the wild. Amongst them, the New Caledonian crow (Corvus moneduloides) stands out with its sophisticated tool-making skills. Despite considerable speculation, the evolutionary origins of this species' remarkable tool behaviour remain largely unknown, not least because no naturally tool-using congeners have yet been identified that would enable informative comparisons. Here we show that another tropical corvid, the 'Alala (C. hawaiiensis; Hawaiian crow), is a highly dexterous tool user. Although the 'Alala became extinct in the wild in the early 2000s, and currently survives only in captivity, at least two lines of evidence suggest that tool use is part of the species' natural behavioural repertoire: juveniles develop functional tool use without training, or social input from adults; and proficient tool use is a species-wide capacity. 'Alala and New Caledonian crows evolved in similar environments on remote tropical islands, yet are only distantly related, suggesting that their technical abilities arose convergently. This supports the idea that avian foraging tool use is facilitated by ecological conditions typical of islands, such as reduced competition for embedded prey and low predation risk. Our discovery creates exciting opportunities for comparative research on multiple tool-using and non-tool-using corvid species. Such work will in turn pave the way for replicated cross-taxonomic comparisons with the primate lineage, enabling valuable insights into the evolutionary origins of tool-using behaviour.

Development and characterization of microsatellite loci for common raven (Corvus corax) and cross species amplification in other Corvidae.

BACKGROUND: A priority for conservation is the identification of endemic populations. We developed microsatellite markers for common raven (Corvus corax), a bird species with a Holarctic distribution, to identify and assess endemic populations in Alaska. RESULTS: From a total of 50 microsatellite loci, we isolated and characterized 15 loci. Eight of these loci were polymorphic and readily scoreable. Eighteen to 20 common ravens from Fairbanks, Alaska were genotyped showing the following variability: 3-8 alleles per locus, 0.25-0.80 observed heterozygosity (Ho), and 0.30-0.80 expected heterozygosity (He). All loci were in Hardy-Weinberg equilibrium and linkage equilibrium and many loci amplified and were polymorphic in related taxa. CONCLUSIONS: These loci will be used to identify endemic populations of common raven and assess their genetic diversity and connectivity.

An extensive candidate gene approach to speciation: diversity, divergence and linkage disequilibrium in candidate pigmentation genes across the European crow hybrid zone.

Colouration patterns have an important role in adaptation and speciation. The European crow system, in which all-black carrion crows and grey-coated hooded crows meet in a narrow hybrid zone, is a prominent example. The marked phenotypic difference is maintained by assortative mating in the absence of neutral genetic divergence, suggesting the presence of few pigmentation genes of major effect. We made use of the rich phenotypic and genetic resources in mammals and identified a comprehensive panel of 95 candidate pigmentation genes for birds. Based on functional annotation, we chose a subset of the most promising 37 candidates, for which we developed a marker system that demonstrably works across the avian phylogeny. In total, we sequenced 107 amplicons (∼3 loci per gene, totalling 60 kb) in population samples of crows (n=23 for each taxon). Tajima's D, Fu's FS, DHEW and HKA (Hudson-Kreitman-Aguade) statistics revealed several amplicons that deviated from neutrality; however, none of these showed significantly elevated differentiation between the two taxa. Hence, colour divergence in this system may be mediated by uncharacterized pigmentation genes or regulatory regions outside genes. Alternatively, the observed high population recombination rate (4Ner∼0.03), with overall linkage disequilibrium dropping rapidly within the order of few 100 bp, may compromise the power to detect causal loci with nearby markers. Our results add to the debate as to the utility of candidate gene approaches in relation to genomic features and the genetic architecture of the phenotypic trait in question.

Comparative phylogeography of two crow species: jungle crow Corvus macrorhynchos and carrion crow Corvus corone.

The jungle crow Corvus macrorhynchos Wagler, 1827, and the carrion crow Corvus corone L., 1758, are two closely related species with similar ecological requirements that occupy wide distribution ranges in the Palearctic. We studied patterns of their genetic variation by using sequences of the mitochondrial cytochrome b gene. Corvus macrorhynchos demonstrates a low level of variation and differentiation throughout its range, except for a highly diverged population of Cheju Island (Korea). The haplotype network shows two haplogroups. The island group comprises populations of Sakhalin, Hokkaido, Honshu, and Kyushu, while the haplotypes of Taiwan and Ryukyu Islands proved to be closer to the mainland group, which also includes populations from the Primorye, Khabarovsk, Amur, and Magadan regions in the Russian Far East. This pattern allowed us to develop a phylogeographic hypothesis regarding the two modes of settling of the island populations. Concerning C. corone, the presence of two distinct haplogroups was confirmed within the range of C. c. orientalis. Both haplogroups are found within the same populations in Kamchatka and North Sakhalin, which implies secondary contacts there. Populations of C. corone are found to be rather stable in the western parts of its range, while in the Far East populations experienced recent growth, as was observed for C. macrorhynchos in general. The two species appear to have passed through different evolutionary scenarios.

Restricted gene flow and fine-scale population structuring in tool using New Caledonian crows.

New Caledonian crows Corvus moneduloides are the most prolific avian tool users. It has been suggested that some aspects of their complex tool use behaviour are under the influence of cultural processes, involving the social transmission-and perhaps even progressive refinement-of tool designs. Using microsatellite and mt-haplotype profiling of crows from three distinct habitats (dry forest, farmland and beachside habitat), we show that New Caledonian crow populations can exhibit significant fine-scale genetic structuring. Our finding that some sites of <10 km apart were highly differentiated demonstrates considerable potential for genetic and/or cultural isolation of crow groups. Restricted movement of birds between local populations at such small spatial scales, especially across habitat boundaries, illustrates how specific tool designs could be preserved over time, and how tool technologies of different crow groups could diverge due to drift and local selection pressures. Young New Caledonian crows have an unusually long juvenile dependency period, during which they acquire complex tool-related foraging skills. We suggest that the resulting delayed natal dispersal drives population-divergence patterns in this species. Our work provides essential context for future studies that examine the genetic makeup of crow populations across larger geographic areas, including localities with suspected cultural differences in crow tool technologies.

Specific chicken repeat 1 (CR1) retrotransposon insertion suggests phylogenetic affinity of rockfowls (genus Picathartes) to crows and ravens (Corvidae).

While the monophyly of the order Passeriformes as well as its suborders suboscines (Tyranni) and oscines (Passeri) is well established, both on morphological and molecular grounds, lower phylogenetic relationships have been a continuous matter of debate, especially within oscines. This is particularly true for the rockfowls (genus Picathartes), which phylogenetic classification has been an ongoing puzzle. Sequence-based molecular studies failed in deriving unambiguously resolved and supported hypotheses. We present here a novel approach: use of retrotransposon insertions as phylogenetic markers in passerine birds. Chicken repeat 1 (CR1) is the most important non-LTR retrotransposon in birds. We present two truncated CR1 loci in passerine birds, not only found in representatives of Corvinae (jays, crows and allies), but also in the West-African Picathartes species which provide new evidence for a closer relationship of these species to Corvidae than has previously been thought. Additionally, we show that not only the absence/presence pattern of a CR1 insertion, but also the CR1 sequences themselves contain phylogenetic information.