Evolution and characterization of tetraonine endogenous retrovirus: a new virus related to avian sarcoma and leukosis viruses.

In a previous study, we found avian sarcoma and leukosis virus (ASLV) gag genes in 19 species of birds in the order Galliformes including all grouse and ptarmigan (Tetraoninae) surveyed. Our data suggested that retroviruses had been transmitted horizontally among some host species. To further investigate these elements, we sequenced a replication-defective retrovirus, here named tetraonine endogenous retrovirus (TERV), from Bonasa umbellus (ruffed grouse). This is the first report of a complete, replication-defective ASLV provirus sequence from any bird other than the domestic chicken. We found a replication-defective proviral sequence consisting of putative Gag and Env proteins flanked by long terminal repeats. Reverse transcription-PCR analysis showed that retroviral gag sequences closely related to TERV are transcribed, supporting the hypothesis that TERV is an active endogenous retrovirus. Phylogenetic analyses suggest that TERV may have arisen via recombination between different retroviral lineages infecting birds. Southern blotting using gag probes showed that TERV occurs in tetraonines but not in chickens or ducks, suggesting that integration occurred after the earliest phasianid divergences but prior to the radiation of tetraonine birds.

Cospeciation and horizontal transmission of avian sarcoma and leukosis virus gag genes in galliform birds.

In a study of the evolution and distribution of avian retroviruses, we found avian sarcoma and leukosis virus (ASLV) gag genes in 26 species of galliform birds from North America, Central America, eastern Europe, Asia, and Africa. Nineteen of the 26 host species from whom ASLVs were sequenced were not previously known to contain ASLVs. We assessed congruence between ASLV phylogenies based on a total of 110 gag gene sequences and ASLV-host phylogenies based on mitochondrial 12S ribosomal DNA and ND2 sequences to infer coevolutionary history for ASLVs and their hosts. Widespread distribution of ASLVs among diverse, endemic galliform host species suggests an ancient association. Congruent ASLV and host phylogenies for two species of Perdix, two species of Gallus, and Lagopus lagopus and L. mutus also indicate an old association with vertical transmission and cospeciation for these ASLVs and hosts. An inference of horizontal transmission of ASLVs among some members of the Tetraoninae subfamily (grouse and ptarmigan) is supported by ASLV monophyletic groups reflecting geographic distribution and proximity of hosts rather than host species phylogeny. We provide a preliminary phylogenetic taxonomy for the new ASLVs, in which named taxa denote monophyletic groups.

Primers for a PCR-based approach to mitochondrial genome sequencing in birds and other vertebrates.

A PCR-based approach to sequencing complete mitochondrial genomes is described along with a set of 86 primers designed primarily for avian mitochondrial DNA (mtDNA). This PCR-based approach allows an accurate determination of complete mtDNA sequences that is faster than sequencing cloned mtDNA. The primers are spaced at about 500-base intervals along both DNA strands. Many of the primers incorporate degenerate positions to accommodate variation in mtDNA sequence among avian taxa and to reduce the potential for preferential amplification of nuclear pseudogenes. Comparison with published vertebrate mtDNA sequences suggests that many of the primers will have broad taxonomic utility. In addition, these primers should make available a wider variety of mitochondrial genes for studies based on smaller data sets.

Interordinal relationships of birds and other reptiles based on whole mitochondrial genomes.

Several different groups of birds have been proposed as being the oldest or earliest diverging extant lineage within the avian phylogenetic tree, particularly ratites (Struthioniformes), waterfowl (Anseriformes), and shorebirds (Charadriiformes). Difficulty in resolving this issue stems from several factors, including the relatively rapid radiation of primary (ordinal) bird lineages and the lack of characters from an extant outgroup for birds that is closely related to them by measure of time. To help resolve this question, we have sequenced entire mitochondrial genomes for five birds (a rhea, a duck, a falcon, and two perching birds), one crocodilian, and one turtle. Maximum parsimony and maximum likelihood analyses of these new sequences together with published sequences (18 taxa total) yield the same optimal tree topology, in which a perching bird (Passeriformes) is sister to all the other bird taxa. A basal position for waterfowl among the bird study taxa is rejected by maximum likelihood analyses. However, neither the conventional view, in which ratites (including rhea) are basal to other birds, nor tree topologies with falcon or chicken basal among birds could be rejected in the same manner. In likelihood analyses of a subset of seven birds, alligator, and turtle (9 taxa total), we find that increasing the number of parameters in the model shifts the optimal topology from one with a perching bird basal among birds to the conventional view with ratites diverging basally; moreover, likelihood scores for the two trees are not significantly different. Thus, although our largest set of taxa and characters supports a tree with perching birds diverging basally among birds, the position of this earliest divergence among birds appears unstable. Our analyses indicate a sister relationship between a waterfowl/chicken clade and ratites, relative to perching birds and falcon. We find support for a sister relationship between turtles and a bird/crocodilian clade, and for rejecting both the Haemothermia hypothesis (birds and mammals as sister taxa) and the placement of turtles as basal within the phylogenetic tree for amniote animals.

Multiple independent origins of mitochondrial gene order in birds.

Mitochondrial genomes of all vertebrate animals analyzed to date have the same 37 genes, whose arrangement in the circular DNA molecule varies only in the relative position of a few genes. This relative conservation suggests that mitochondrial gene order characters have potential utility as phylogenetic markers for higher-level vertebrate taxa. We report discovery of a mitochondrial gene order that has had multiple independent originations within birds, based on sampling of 137 species representing 13 traditionally recognized orders. This provides evidence of parallel evolution in mitochondrial gene order for animals. Our results indicate operation of physical constraints on mitochondrial gene order changes and support models for gene order change based on replication error. Bird mitochondria have a displaced OL (origin of light-strand replication site) as do various other Reptilia taxa prone to gene order changes. Our findings point to the need for broad taxonomic sampling in using mitochondrial gene order for phylogenetic analyses. We found, however, that the alternative mitochondrial gene orders distinguish the two primary groups of songbirds (order Passeriformes), oscines and suboscines, in agreement with other molecular as well as morphological data sets. Thus, although mitochondrial gene order characters appear susceptible to some parallel evolution because of mechanistic constraints, they do hold promise for phylogenetic studies.