Patterns of polymorphism and gene flow of gender-associated mitochondrial DNA lineages in European mussel populations

Mussels of the genus Mytilus have two types of mitochondrial DNA (mtDNA). The M type is transmitted paternally and the F type is transmitted maternally. RFLP analysis is used to assess phylogenetic relationships and nucleotide diversity and divergence for both mtDNA genomes in European populations of M. edulis and Atlantic and Mediterranean forms of M. galloprovincialis. Ten restriction endonucleases were used to assay variation in regions of the ND2 and COIII genes for a total of 77 individuals. F and M genomes show a concordant phylogenetic split into two major divergent clades, one specific to Mediterranean M. galloprovincialis and the other containing haplotypes from the three taxa. For both genomes, the geographical distribution of mtDNA variation suggests: (i) extensive levels of mtDNA introgression; (ii) asymmetric mtDNA gene flow from Atlantic to Mediterranean populations; and (iii) recurrent historical hybridization events. Significantly higher mtDNA diversity and divergence are observed for the M than F genome in all three Mytilus taxa, although the evolutionary forces responsible for these differences cannot be resolved. The extensive mtDNA gene flow between European Mytilus taxa conflicts with the restricted mtDNA introgression observed in American mussels , implying geographical variation in the nature of nuclear/mtDNA interactions regulating biparental inheritance.     

Interspecies transfer of female mitochondrial DNA is coupled with role-reversals and departure from neutrality in the mussel Mytilus trossulus.

Mussels of the genus Mytilus have distinct and highly diverged male and female mitochondrial DNA (mtDNA) genomes with separate routes of inheritance. Previous studies of European populations of Mytilus trossulus demonstrated that 33% of males are heteroplasmic for a second mtDNA genome of increased length and that hybridization with Mytilus edulis does not block mtDNA introgression, in contrast to reports for American populations. Here, we demonstrate that the female mtDNA type of M. edulis has replaced the resident female mtDNA type of European M. trossulus. This is supported by COIII sequence data indicating that the female mtDNA of European M. trossulus is very similar to that of M. edulis and that in phylogenetic trees, the mtDNAs of these two species cluster together but separately from American M. trossulus sequences, the latter not being disturbed by introgressive hybridization. We also provide evidence that the mtDNA genome of increased length found in heteroplasmic males of European M. trossulus derives from a recent partition of an introgressed M. edulis female type into the male route of transmission. Neutrality tests reveal that European populations of M. trossulus display an excess of replacement polymorphism within the female mtDNA type with respect to conspecific American populations, as well as a significant excess of rare variants, of a similar magnitude to those previously reported for the invading European M. edulis mtDNA. Results are consistent with a nearly neutral model of molecular evolution and suggest that selection acting on European M. trossulus mtDNA is largely independent of the nuclear genetic background.     

Positive selection drives the evolution of the Acp29AB accessory gland protein in Drosophila.

Nucleotide sequence variation at the Acp29AB gene region has been surveyed in Drosophila melanogaster from Spain (12 lines), Ivory Coast (14 lines), and Malawi (13 lines) and in one line of D. simulans. The approximately 1.7-kb region studied encompasses the Acp29AB gene that codes for a male accessory gland protein and its flanking regions. Seventy-seven nucleotide and 8 length polymorphisms were detected. Nonsynonymous polymorphism was an order of magnitude lower than synonymous polymorphism, but still high relative to other non-sex-related genes. In D. melanogaster variation at this region revealed no major genetic differentiation between East and West African populations, while differentiation was highly significant between the European and the two African populations. Comparison of polymorphism and divergence at synonymous and nonsynonymous sites showed an excess of fixed nonsynonymous changes, which indicates that the evolution of the Acp29AB protein has been driven by directional selection at least after the split of the D. melanogaster and D. simulans lineages. The pattern of variation in extant populations of D. melanogaster favors a scenario where the fixation of advantageous replacement substitutions occurred in the early stages of speciation and balancing selection is maintaining variation in this species.     

Comparative analysis of gender-associated complete mitochondrial genomes in marine mussels (Mytilus spp.)

Marine mussels of the genus Mytilus have an unusual mode of mitochondrial DNA (mtDNA) transmission termed doubly uniparental inheritance (DUI). Female mussels are homoplasmic for the F mitotype, which is inherited maternally, while males are usually heteroplasmic, carrying a mixture of the maternal F mitotype and the paternally inherited M genome. Two classes of M genomes have been observed: "standard" M genomes and "recently masculinized" M genomes. The latter are more similar to F genomes at the sequence level but are transmitted paternally like standard M genomes. In this study we report the complete sequences of two standard male M. edulis and one recently masculinized male M. trossulus mitochondrial genome. A comparative analysis, including the previously sequenced M. edulis F and M. galloprovincialis F and M mtDNAs, reveals that these genomes are identical in gene order, but highly divergent in nucleotide and amino acid sequence. The large amount (>20%) of nucleotide substitutions that fall in coding regions implies that there are several amino acid replacements between the F and M genomes, which likely have an impact on the structural and functional properties of the mitochondrial proteome. Correlation of the divergence rate of different protein-coding genes indicates that mtDNA-encoded proteins of the M genome are still under selective constraints, although less highly than genes of the F genome. The mosaic F/M control region of the masculinized F genome provides evidence for lineage-specific sequences that may be responsible for the different mode of transmission genetics. This analysis shows the value of comparative genomics to better understand the mechanisms of maintenance and segregation of mtDNA sequence variants in mytilid mussels.     

Molecular population genetics of the male and female mitochondrial DNA molecules of the California sea mussel, Mytilus californianus

The presence of two gender-associated mitochondrial genomes in marine mussels provides a unique opportunity to investigate the dynamics of mtDNA evolution without complications inherent in interspecific comparisons. Here, we assess the relative importance of selection, mutation, and differential constraint in shaping the patterns of polymorphism within and divergence between the male (M) and female (F) mitochondrial genomes of the California sea mussel, Mytilus californianus. Partial sequences were obtained from homologous regions of four genes (nad2, cox1, atp6, and nad5) totaling 2307 bp in length. The M and F mtDNA molecules of M. californianus exhibited extensive levels of nucleotide polymorphism and were more highly diverged than observed in other mytilids (overall Tamura-Nei distances >40%). Consistent with previous studies, the M molecule had significantly higher levels of silent and replacement polymorphism relative to F. Both genomes possessed large numbers of singleton and low-frequency mutations that gave rise to significantly negative Tajima's D values. Mutation-rate scalars estimated for silent and replacement mutations were elevated in the M genome but were not sufficient to account for its higher level of polymorphism. McDonald-Kreitman tests were highly significant at all loci due to excess numbers of fixed replacement mutations between molecules. Strong purifying selection was evident in both genomes in keeping the majority of replacement mutations at low population frequencies but appeared to be slightly relaxed in M. Our results suggest that a reduction in selective constraint acting on the M genome remains the best explanation for its greater levels of polymorphism and faster rate of evolution.     

Mitochondrial DNA variation in a species with two mitochondrial genomes: the case of Mytilus galloprovincialis from the Atlantic, the Mediterranean and the Black Sea

We have examined mitochondrial DNA (mtDNA) variation in samples of the mussel Mytilus galloprovincialis from the Black Sea, the Mediterranean and the Spanish Atlantic coast by scoring for presence or absence of cleavage at 20 restriction sites of a fragment of the COIII gene and at four restriction sites of the 16S RNA gene. This species contains two types of mtDNA genomes, one that is transmitted maternally (the F type) and one that is transmitted paternally (the M type). The M genome evolves at a higher rate than the F genome. Normally, females are homoplasmic for an F type and males are heteroplasmic for an F and an M type. Occasionally molecules from the F lineage invade the paternal transmission route, resulting in males that carry two F-type mtDNA genomes. These features of the mussel mtDNA system give rise to a new set of questions when using mtDNA variation in population studies and phylogeny. We show here that the two mtDNA types provide different information with regard to amounts of variation and genetic distances among populations. The F genome exhibits higher degrees of diversity within populations, while the M genome produces higher degrees of differentiation among populations. There is a strong differentiation between the Atlantic and the Black Sea. The Mediterranean samples have intermediate haplotype frequencies, yet are much closer to the Black Sea than to the Atlantic. We conclude that in this species gene flow among the three Seas is restricted and not enough to erase the combined effect of mutation and random drift. In one sample, that from the Black Sea, the majority of males did not contain an M mtDNA type. This suggests that a molecule of the maternal lineage has recently invaded the paternal route and has increased its frequency in the population to the point that the present pool of paternally transmitted mtDNA molecules is highly heterogeneous and cannot be used to read the population's history. This liability of the paternal route means that in species with doubly uniparental inheritance, the maternal lineage provides more reliable information for population and phylogenetic studies.     

Lineage-specific selection in human mtDNA: lack of polymorphisms in a segment of MTND5 gene in haplogroup J

Human mitochondrial DNA (mtDNA) is a nonrecombining genome that codes for 13 subunits of the mitochondrial oxidative phosphorylation system, 2 rRNAs, and 22 tRNAs. Mutations have accumulated sequentially in mtDNA lineages that diverged tens of thousands of years ago. The genes in mtDNA are subject to different functional constraints and are therefore expected to evolve at different rates, but the rank order of these rates should be the same in all lineages of a phylogeny. Previous studies have indicated, however, that specific regions of mtDNA may have experienced different histories of selection in different lineages, possibly because of lineage-specific interactions or environmental factors such as climate. We report here on a survey for lineage-specific patterns of nucleotide polymorphism in human mtDNA. We calculated molecular polymorphism indices and neutrality tests for classes of functional sites and genes in 837 human mtDNA sequences, compared the results between continent-specific mtDNA lineages, and used two sliding window methods to identify differences in the patterns of polymorphism between haplogroups. A general correlation between nucleotide position and the level of nucleotide polymorphism was identified in the coding region of the mitochondrial genome. Nucleotide diversity in the protein-coding sequence of mtDNA was generally not much higher than that found for many genes in nuclear DNA. A comparison of nonsynonymous/synonymous rate ratios in the 13 protein-coding genes suggested differences in the relative levels of selection between haplogroups, including the European haplogroup clusters. Interestingly, a segment of the MTND5 gene was found to be almost void of segregating sites and nonsynonymous mutations in haplogroup J, which has been associated with susceptibility to certain complex diseases. Our results suggest that there are haplogroup-specific differences in the intensity of selection against particular regions of the mitochondrial genome, indicating that some mutations may be non-neutral within specific phylogenetic lineages but neutral within others.     

Incorporation of mitochondrial fragments in the nuclear genome (Numts) of the longhorned beetle Monochamus galloprovincialis (Coleoptera, Cerambycidae)

A mitochondrial DNA (mtDNA) study, based on 43 European populations (33 of them sampled in France) of Monochamus galloprovincialis , vector of the pinewood nematode, and 14 populations of its sister species Monochamus sutor was realized. Sequencing of 792 bp of the cytochrome oxidase I (COI) and 521 bp of the COII genes revealed numerous ambiguities on multiple nucleotide sites for half of M. galloprovincialis specimens studied (44.8%). Hypotheses of heteroplasmy and pseudogenes ( Numts ) were examined. The mtDNA isolation by alkaline lysis and cloning (for three successfully used individuals) both support the hypothesis that the ambiguous sequences amplified were not of mtDNA nature and validate the presence of Numts in the nuclear genome of M. galloprovincialis . Multiple copies of mtDNA-like sequences were found paralogous to COI, tRNA leucin and COII regions. Phenetic analysis placed different recently diverged mtDNA-like sequences as a close relative of mtDNA sequences, and grouped 10 closely related mtDNA-like sequences as a more basal clade, closer to ancestral states and to M. sutor . This result supports that this nuclear family of pseudogenes arose independently of the other events and may represent mitochondrial haplotypes sampled from M. galloprovincialis ancestral populations. This is the first time that Numts are proved for a longhorned beetle, whereas no Numts were found within its sister species M. sutor. The incorporation mechanism of Numts in unknown for M. galloprovincialis , however, excess of ambiguous sites corresponding to synonymous mutations placed on third codon position as well as the absence of Numts in M. sutor , conducted to the hypothesis of a recent transfer of these Numts in the nuclear genome of M. galloprovincialis .     

Positive selection on an acrosomal sperm protein,M7 lysin,in three species of the mussel genus Mytilus

Marine invertebrate sperm proteins are particularly interesting because they are characterized by positive selection and are likely to be involved in prezyogotic isolation and, thus, speciation. Here, we present the first survey of interspecific and intraspecific variation of a bivalve sperm protein among a group of species that regularly hybridize in nature. M7 lysin is found in sperm acrosomes of mussels and dissolves the egg vitelline coat, permitting fertilization. We sequenced multiple alleles of the mature protein-coding region of M7 lysin from allopatric populations of mussels in the Mytilus edulis species group (M. edulis, M. galloprovincialis, and M. trossulus). A significant McDonald-Kreitman test showed an excess of fixed amino acid replacing substitutions between species, consistent with positive selection. In addition, Kolmogorov-Smirnov tests showed significant heterogeneity in polymorphism to divergence ratios for both synonymous variation and combined synonymous and nonsynonymous variation within M. galloprovincialis. These results indicate that there has been adaptive evolution at M7 lysin and, furthermore, show that positive selection on sperm proteins can occur even when postzygotic reproductive isolation is incomplete.     

Degree of Selective Constraint as an Explanation of the Different Rates of Evolution of Gender-Specific Mitochondrial DNA Lineages in the Mussel Mytilus

Mussels of the genus Mytilus segregate for a maternally transmitted F lineage and a paternally transmitted M lineage of mitochondrial DNA. Previous studies demonstrated that these lineages are older than the species of the M. edulis complex and that the M lineage evolves faster than the F lineage. Here we show that the latter observation also applies to a region of the molecule with no assigned function. Sequence data for the mitochondrial COIII gene and the ``unassigned' region of the F and M lineages of M. edulis and M. trossulus are used to evaluate various hypotheses that may account for the faster rate of evolution of the M lineage. Tests based on the proportion of synonymous and nonsynonymous substitutions suggest that the M lineage experiences relatively relaxed selection. Further support for this hypothesis comes from an examination of COIII amino acid substitutions at sites defined as either conserved or variable based on the pattern of variation in other mollusks and Drosophila. Most substitutions in the M lineage occur in regions that are also variable among non-Mytilus taxa. We suggest that these differences in selection pressure are a consequence of doubly uniparental mitochondrial DNA transmission in Mytilus.     

The role of selection in the evolution of human mitochondrial genomes

High mutation rate in mammalian mitochondrial DNA generates a highly divergent pool of alleles even within species that have dispersed and expanded in size recently. Phylogenetic analysis of 277 human mitochondrial genomes revealed a significant (P < 0.01) excess of rRNA and nonsynonymous base substitutions among hotspots of recurrent mutation. Most hotspots involved transitions from guanine to adenine that, with thymine-to-cytosine transitions, illustrate the asymmetric bias in codon usage at synonymous sites on the heavy-strand DNA. The mitochondrion-encoded tRNAThr varied significantly more than any other tRNA gene. Threonine and valine codons were involved in 259 of the 414 amino acid replacements observed. The ratio of nonsynonymous changes from and to threonine and valine differed significantly (P = 0.003) between populations with neutral (22/58) and populations with significantly negative Tajima's D values (70/76), independent of their geographic location. In contrast to a recent suggestion that the excess of nonsilent mutations is characteristic of Arctic populations, implying their role in cold adaptation, we demonstrate that the surplus of nonsynonymous mutations is a general feature of the young branches of the phylogenetic tree, affecting also those that are found only in Africa. We introduce a new calibration method of the mutation rate of synonymous transitions to estimate the coalescent times of mtDNA haplogroups.     

Contrasting patterns of synonymous and nonsynonymous sequence evolution in asexual and sexual freshwater snail lineages

In asexual lineages, both synonymous and nonsynonymous sequence polymorphism may be reduced due to severe founder effects when asexual lineages originate. However, mildly deleterious (nonsynonymous) mutations may accumulate after asexual lineages are formed, because the efficiency of purifying selection is reduced even in the nonrecombining mitochondrial genome. Here we examine patterns of synonymous and nonsynonymous mitochondrial sequence polymorphism in asexual and sexual lineages of the freshwater snail Campeloma. Using clade-specific estimates, we found that synonymous sequence polymorphism was significantly reduced by 75% in asexuals relative to sexuals, whereas nonsynonymous sequence polymorphism did not differ significantly between sexuals and asexuals. Two asexual clades had high negative values for Tajima's D statistic. Coalescent simulations confirmed that various bottleneck scenarios can account for this result. We also used branch-specific estimates of the ratio of amino acid to silent substitutions, K(a)/K(s). Our study revealed that K(a)/K(s) ratios are six times higher in terminal branches of independent asexual lineages compared to sexuals. Coalescent-based reconstruction of gene networks for all sexual and asexual clades indicated that nonsynonymous mutations occurred at a higher frequency in recently derived asexual haplotypes. These findings suggest that patterns of synonymous and nonsynonymous nucleotide polymorphism in asexual snail lineages may be shaped by both severe founder effect and relaxed purifying selection.     

Mitochondrial phylogenomics of the Bivalvia (Mollusca): searching for the origin and mitogenomic correlates of doubly uniparental inheritance of mtDNA

Background  

Doubly uniparental inheritance (DUI) is an atypical system of animal mtDNA inheritance found only in some bivalves. Under DUI, maternally (F genome) and paternally (M genome) transmitted mtDNAs yield two distinct gender-associated mtDNA lineages. The oldest distinct M and F genomes are found in freshwater mussels (order Unionoida). Comparative analyses of unionoid mitochondrial genomes and a robust phylogenetic framework are necessary to elucidate the origin, function and molecular evolutionary consequences of DUI. Herein, F and M genomes from three unionoid species, Venustaconcha ellipsiformis, Pyganodon grandis and Quadrula quadrula have been sequenced. Comparative genomic analyses were carried out on these six genomes along with two F and one M unionoid genomes from GenBank (F and M genomes of Inversidens japanensis and F genome of Lampsilis ornata).     

Elevated rates of nonsynonymous substitution in island birds

Slightly deleterious mutations are expected to fix at relatively higher rates in small populations than in large populations. Support for this prediction of the nearly-neutral theory of molecular evolution comes from many cases in which lineages inferred to differ in long-term average population size have different rates of nonsynonymous substitution. However, in most of these cases, the lineages differ in many other ways as well, leaving open the possibility that some factor other than population size might have caused the difference in substitution rates. We compared synonymous and nonsynonymous substitutions in the mitochondrial cyt b and ND2 genes of nine closely related island and mainland lineages of ducks and doves. We assumed that island taxa had smaller average population sizes than those of their mainland sister taxa for most of the time since they were established. In all nine cases, more nonsynonymous substitutions occurred on the island branch, but synonymous substitutions showed no significant bias. As in previous comparisons of this kind, the lineages with smaller populations might differ in other respects that tend to increase rates of nonsynonymous substitution, but here such differences are expected to be slight owing to the relatively recent origins of the island taxa. An examination of changes to apparently "preferred" and "unpreferred" synonymous codons revealed no consistent difference between island and mainland lineages.     

Hotspots of Biased Nucleotide Substitutions in Human Genes

Genes that have experienced accelerated evolutionary rates on the human lineage during recent evolution are candidates for involvement in human-specific adaptations. To determine the forces that cause increased evolutionary rates in certain genes, we analyzed alignments of 10,238 human genes to their orthologues in chimpanzee and macaque. Using a likelihood ratio test, we identified protein-coding sequences with an accelerated rate of base substitutions along the human lineage. Exons evolving at a fast rate in humans have a significant tendency to contain clusters of AT-to-GC (weak-to-strong) biased substitutions. This pattern is also observed in noncoding sequence flanking rapidly evolving exons. Accelerated exons occur in regions with elevated male recombination rates and exhibit an excess of nonsynonymous substitutions relative to the genomic average. We next analyzed genes with significantly elevated ratios of nonsynonymous to synonymous rates of base substitution (d_N/d_S) along the human lineage, and those with an excess of amino acid replacement substitutions relative to human polymorphism. These genes also show evidence of clusters of weak-to-strong biased substitutions. These findings indicate that a recombination-associated process, such as biased gene conversion (BGC), is driving fixation of GC alleles in the human genome. This process can lead to accelerated evolution in coding sequences and excess amino acid replacement substitutions, thereby generating significant results for tests of positive selection.     

Comparative Genomics of Marine Mussels (Mytilus spp.) Gender Associated mtDNA: Rapidly Evolving atp8

The unusual mode of mitochondrial DNA inheritance, with two separate: maternal (F) and paternal (M) lineages, gives unique opportunities to study the evolution of the mitochondrial genome. This system was first discovered in the marine mussels Mytilus. The three related species: Mytilus edulis, Mytilus galloprovincialis and Mytilus trossulus form a complex in which the divergence of M and F lineages pre-dates the speciation. The complete mitochondrial genomes of both lineages were known for all species except Pacific M. trossulus. Here we report, for the first time, the complete sequences of both mitochondrial genomes of Pacific M. trossulus, filling the gap. While the reported M and F genomes are highly diverged (26%), they have similar organisation. The only difference is the translocation of one tRNA gene into the long, mosaic control region of the F genome. Consistent presence of an ORF which most likely represents the atp8 gene was confirmed in both genomes. The predicted protein has characteristics expected of the functional atp8 even though the M and F versions are markedly different in length. Comparative analysis involving all three species led to the conclusion that the cause of a faster evolution of atp8 and Mytilus mtDNA in general is most likely the Compensation-Draft Feedback process coupled with relatively relaxed selection in the M lineage. Thus, we postulate that the adaptive changes may have played a role in the emergence of highly diverged, barely recognizable atp8 in Mytilus mussels.     

Evidence for recombination of mtDNA in the marine mussel Mytilus trossulus from the Baltic

A number of studies have claimed that recombination occurs in animal mtDNA, although this evidence is controversial. Ladoukakis and Zouros (2001) provided strong evidence for mtDNA recombination in the COIII gene in gonadal tissue in the marine mussel Mytilus galloprovincialis from the Black Sea. The recombinant molecules they reported had not however become established in the population from which experimental animals were sampled. In the present study, we provide further evidence of the generality of mtDNA recombination in Mytilus by reporting recombinant mtDNA molecules in a related mussel species, Mytilus trossulus, from the Baltic. The mtDNA region studied begins in the 16S rRNA gene and terminates in the cytochrome b gene and includes a major noncoding region that may be analogous to the D-loop region observed in other animals. Many bivalve species, including some Mytilus species, are unusual in that they have two mtDNA genomes, one of which is inherited maternally (F genome) the other inherited paternally (M genome). Two recombinant variants reported in the present study have population frequencies of 5% and 36% and appear to be mosaic for F-like and M-like sequences. However, both variants have the noncoding region from the M genome, and both are transmitted to sperm like the M genome. We speculate that acquisition of the noncoding region by the recombinant molecules has conferred a paternal role on mtDNA genomes that otherwise resemble the F genome in sequence.     

Pyganodon (Bivalvia: Unionoida: Unionidae) phylogenetics: a male- and female-transmitted mitochondrial DNA perspective

Species boundaries, evolutionary relationships and geographic distributions of many unionoid bivalve species, like those in the genus Pyganodon, remain unresolved in Eastern North America. Because unionoid bivalves are one of the most imperiled groups of animals in the world, understanding the genetic variation within and among populations as well as among species is crucial for effective conservation planning. Conservation of unionoid species is indispensable from a freshwater habitat perspective but also because they possess a unique mitochondrial inheritance system where distinct gender-associated mitochondrial DNA lineages coexist: a female-transmitted (F) mt genome and a male-transmitted (M) mt genome that are involved in the maintenance of separate sexes (=dioecy). In this study, 42 populations of Pyganodon sp. were sampled across a large geographical range and fragments of two mitochondrial genes (cox1 and cox2) were sequenced from both the M- and F-transmitted mtDNA genomes. Our results support the recency of the divergence between P. cataracta and P. fragilis. We also found two relatively divergent F and M lineages within P. grandis. Surprisingly, the relationships among the P. grandis specimens in the F and M sequence trees are not congruent. We found that a single haplotype in P. lacustris has recently swept throughout the M genotype space leading to an unexpectedly low diversity in the M lineage in that species. Our survey put forward some challenging results that force us to rethink hybridization and species boundaries in the genus Pyganodon. As the M and F genomes do not always display the same phylogeographic story in each species, we also discuss the importance of being careful in the interpretation of molecular data based solely on maternal transmitted mtDNA genomes. The involvement of F and M genomes in unionoid bivalve sex determination likely played a role in the genesis of the unorthodox phylogeographic patterns reported herein.     

Likelihood ratio tests for detecting positive selection and application to primate lysozyme evolution

An excess of nonsynonymous substitutions over synonymous ones is an important indicator of positive selection at the molecular level. A lineage that underwent Darwinian selection may have a nonsynonymous/synonymous rate ratio (dN/dS) that is different from those of other lineages or greater than one. In this paper, several codon-based likelihood models that allow for variable dN/dS ratios among lineages were developed. They were then used to construct likelihood ratio tests to examine whether the dN/dS ratio is variable among evolutionary lineages, whether the ratio for a few lineages of interest is different from the background ratio for other lineages in the phylogeny, and whether the dN/dS ratio for the lineages of interest is greater than one. The tests were applied to the lysozyme genes of 24 primate species. The dN/dS ratios were found to differ significantly among lineages, indicating that the evolution of primate lysozymes is episodic, which is incompatible with the neutral theory. Maximum- likelihood estimates of parameters suggested that about nine nonsynonymous and zero synonymous nucleotide substitutions occurred in the lineage leading to hominoids, and the dN/dS ratio for that lineage is significantly greater than one. The corresponding estimates for the lineage ancestral to colobine monkeys were nine and one, and the dN/dS ratio for the lineage is not significantly greater than one, although it is significantly higher than the background ratio. The likelihood analysis thus confirmed most, but not all, conclusions Messier and Stewart reached using reconstructed ancestral sequences to estimate synonymous and nonsynonymous rates for different lineages.      

Comparative Genomics of Mitochondrial DNA in Drosophila simulans

The current study compares the nucleotide variation among 22 complete mitochondrial genomes of the three distinct Drosophila simulans haplotypes with intron 1 of the alcohol dehydrogenase-related locus. This is the first study to investigate the sequence variation of multiple complete mitochondrial genomes within distinct mitochondrial haplotypes of a single species. Patterns of variation suggest distinct forces are influencing the evolution of mitochondrial DNA (mtDNA) and autosomal DNA in D. simulans. First, there is little variation within each mtDNA haplotype but strong differentiation among them. In contrast, there is no support for differentiation of the mitochondrial haplotypes at the autosomal locus. Second, there is a significant deficiency of mitochondrial variation in each haplotype relative to the autosomal locus. Third, the ratio of nonsynonymous to synonymous substitutions is not equal in all branches of the well-resolved phylogeny. There is an excess of nonsynonymous substitutions relative to synonymous substitutions within each D. simulans haplotype. This result is similar to that previously observed within the mtDNA of distinct species. A single evolutionary force may be causally linked to the observed patterns of mtDNA variation—a rickettsia-like microorganism, Wolbachia pipientis, which is known to directly influence mitochondrial evolution but have a less direct influence on autosomal loci. Received: 16 September 1999 / Accepted: 14 March 2000