Relationships of Afroablepharus Greer, 1974 skinks from the Gulf of Guinea islands based on mitochondrial and nuclear DNA: patterns of colonization and comments on taxonomy

Partial sequences of three mitochondrial DNA genes, 12S rDNA, 16S rDNA and cytochrome b, and one nuclear gene, c-mos, were used to assess the phylogenetic relationships of species belonging to the genus Afroablepharus from the volcanic islands of the Gulf of Guinea (West Africa) and neighboring continental Africa. Additionally, partial sequences of cytochrome b were used to compare levels of sequence divergence within populations. The three forms from S?o Tomé, Príncipe and Annobon (one per island) are genetically distinct, with high levels of divergence, supporting the recognition of a distinct species in each island. Populations within each island contain very low levels of genetic diversity. These three forms form a monophyletic group suggesting a single initial colonization followed by radiation to the other islands, possibly from S?o Tomé to Príncipe and Annobon. This is contrary to what was found in other reptiles from these islands such as Mabuya (sensu lato) and Hemidactylus, which colonized the islands multiple times. Assuming a molecular clock for cytochrome b of about 2% divergence per million years (usually applied to Sauria), the lineage on Annobon island exceeds the age of the island, thus casting further doubt on this widely used divergence estimate. Partial sequences of c-mos showed no variation within islands. Five to seven sites were variable among islands, which is a high value further supporting the treatment of each island form as a distinct species.     

Sequence evolution and phylogenetic signal in control-region and cytochrome b sequences of rainbow fishes (Melanotaeniidae)

The nucleotide sequences of segments of the cytochrome b gene (351 bp), the tRNA(Pro) gene (49 bp), and the control region (approximately 313 bp) of mitochondrial DNA were obtained from 26 fish representing different populations and species of Melanotaenia and one species of Glossolepis, freshwater rainbow fishes confined to Australia and New Guinea. The purpose was to investigate relative rates and patterns of sequence evolution. Overall levels of divergence were similar for the cytochrome b and tRNA control-region sequences, both ranging from < 1% within subspecies to 15%-19% between genera. However, the patterns of sequence evolution differed. For the cytochrome b gene, transitions consistently exceeded transversions, the bias ranging from 4.2:1 to 2:1, depending on the level of sequence divergence. However, in the control-region sequence, a bias toward transitions (2:1) was observed only in comparisons between very similar sequences, and transversions outnumbered transitions in comparisons of divergent sequences. Graphic comparisons suggested that the control region was saturated for transitions at relatively low levels of sequence divergence but accumulated transversions at a greater rate than did the cytochrome b sequence. These distinct patterns of base substitution are associated with differences in A+T content, which is 70% for the tRNA control- region segment versus 50% for cytochrome b. A test for skewness in the distribution of lengths of random trees indicated that both segments contained phylogenetic signal. Parsimony analyses of the data from the two regions, with or without weighting schemes appropriate to the respective patterns of sequence evolution, identified the same five groupings of sequences, but the relationships among the groups differed. However, in most cases the branches uniting different combinations of groups were poorly supported, and the differences among topologies were insignificant. Considering the observed patterns of base substitution and the results of the phylogenetic analyses, we deduce that both the control region and cytochrome b are appropriate for population genetic studies but that the control region is less effective than cytochrome b for resolving relationships among divergent lineages of rainbow fishes.      

Extreme intraspecific mitochondrial DNA sequence divergence in Galaxias maculatus (Osteichthys: Galaxiidae), one of the world's most widespread freshwater fish

Biogeographic controversies surrounding the widespread freshwater fish, Galaxias maculatus, were addressed with DNA sequence data. Mitochondrial cytochrome b and 16S rRNA sequences were obtained from representatives of six populations of this species. Substantial levels of cytochrome b (maximum 14.6%) and 16S rRNA sequence divergence (maximum 6.0%) were detected between western Pacific (Tasmania-New Zealand) and South American (Chile-Falkland Islands) haplotypes. A considerable level of divergence was also detected between Tasmanian and New Zealand haplotypes (maximum 5.1%) and within and among Chilean and Falkland Island G. maculatus (maximum 3. 8%). The phylogenetic structure of haplotypes conflicts with the accepted pattern of continental fragmentation. Molecular clock calibrations suggest that haplotype divergences postdate the fragmentation of Gondwana. These findings point to marine dispersal rather than ancient vicariance as an explanation for the wide distribution. The phylogenetic structure of South American haplotypes was not consistent with their geographic distribution. We consider factors such as population divergence, population size, dispersal, secondary contact, and philopatry as potential causes of the high level of mtDNA nucleotide diversity in this species.     

Variation of cytochrome b gene in sympatric chars from Kronotsky Lake (Kamchatka oblast)

In previously identified sympatric char forms (species) from Kronotsky Lake (Kamchatka Peninsula) and a number of other chars from the genus Salvelinus, nucleotide sequences of the mtDNA cytochrome b gene were determined and phylogenetic analysis of these forms was performed. The divergence of the cytochrome b gene in the Kronotsky char group (0.48%) coincided with the interpopulation difference estimates in northern Dolly Varden, which indicates a relatively recent divergence of the chars from Kronotsky Lake. Haplotypes of each form of the Kronotsky Lake chars are divided into two groups, which belonged to two different phylogenetic mtDNA lineages of northern Dolly Varden from Chukotka and Kamchatka. The formation of the Kronotsky char forms proved to be unrelated to the observed variability of the mtDNA cytochrome b gene.     

Variation in mitochondrial cytochrome b sequence in natural populations of South American akodontine rodents (Muridae: Sigmodontinae)

A 401-bp fragment of the mitochondrial cytochrome b gene was sequenced from polymerase chain reaction-amplified products for 20 natural populations representing 12 species of South American akodontine rodents (Muridae). Variation among these taxa increased with their hierarchical position, from comparisons within local populations to those among different genera. Two individuals from the same local population differed by less than 1% sequence divergence. Sequence divergence among geographic samples within a species was 0.25%-8%, while that among species was 3%-21%. Comparisons of the akodontine sequences with that for the house mouse show 21%-25% sequence difference. A parsimony-based phylogenetic analysis of the data supports the placement of the taxon Microxus within Akodon (sensu stricto), of Bolomys just outside the Akodon cluster, and of Chroeomys as a separate genus quite distinct from the other members of this group. This phylogenetic hypothesis is identical to that determined from electrophoretic data but is quite divergent from the present taxonomy of the group.     

Genetic divergence does not predict change in ornament expression among populations of stalk-eyed flies

Stalk-eyed flies (Diptera: Diopsidae) possess eyes at the ends of elongated peduncles, and exhibit dramatic variation in eye span, relative to body length, among species. In some sexually dimorphic species, evidence indicates that eye span is under both intra- and intersexual selection. Theory predicts that isolated populations should evolve differences in sexually selected traits due to drift. To determine if eye span changes as a function of divergence time, 1370 flies from 10 populations of the sexually dimorphic species, Cyrtodiopsis dalmanni and Cyrtodiopsis whitei, and one population of the sexually monomorphic congener, Cyrtodiopsis quinqueguttata, were collected from Southeast Asia and measured. Genetic differentiation was used to assess divergence time by comparing mitochondrial (cytochrome oxidase II and 16S ribosomal RNA gene fragments) and nuclear (wingless gene fragment) DNA sequences for c. five individuals per population. Phylogenetic analyses indicate that most populations cluster as monophyletic units with up to 9% nucleotide substitutions between populations within a species. Analyses of molecular variance suggest a high degree of genetic structure within and among the populations; > 97% of the genetic variance occurs between populations and species while < 3% is distributed within populations, indicating that most populations have been isolated for thousands of years. Nevertheless, significant change in the allometric slope of male eye span on body length was detected for only one population of either dimorphic species. These results are not consistent with genetic drift. Rather, relative eye span appears to be under net stabilizing selection in most populations of stalk-eyed flies. Given that one population exhibited dramatic evolutionary change, selection, rather than genetic variation, appears to constrain eye span evolution.     

Variation of cytochrome b gene in sympatric chars from kronotsky lake (Kamchatka Peninsula)

In previously identified sympatric char forms (species) from Kronotsky Lake (Kamchatka Peninsula) and a number of other chars from the genus Salvelinus, nucleotide sequences of the mtDNA cytochrome b gene were determined and phylogenetic analysis of these chars was performed. The divergence estimate of the cytochrome b gene in the Kronotsky char group (0.48%) coincided with the interpopulation difference estimates in northern Dolly Varden, which indicates a relatively recent divergence of the chars from Kronotsky Lake. Haplotypes of each form of the Kronotsky Lake chars are divided into two groups, which belonged to two different phylogenetic mtDNA lineages of northern Dolly Varden from Chukotka and Kamchatka. The formation of the Kronotsky char forms proved to be unrelated to the observed variability of the mtDNA cytochrome b gene.     

The genetic structure of chum salmon (Oncorhynchus keta Walbaum) populations inferred from the nucleotide variation of the mitochondrial DNA cytochrome b gene

The nucleotide sequences of a fragment of the mitochondrial DNA cytochrome b gene were determined in 12 chum salmon populations from the Russian Far East. The level of genetic diversity in the chum salmon populations from the Iturup Island, northern coast of the Sea of Okhotsk, and Anadyr' River was found to be higher than in the populations from Kamchatka and Sakhalin, which may be related to the history of their origin and dispersal. The proportions of intrapopulation genetic variability (F(ct)) and interpopulation genetic variability within the groups (F(sc)) account for 90.87 and 0.9%, respectively, and the intergroup component (F(st)) comprises 8.23%. The predominance of one haplotype, B1, which is common for all populations studied, and a low share of intergroup variability suggest the beginning of colonization by the species of the given region from a common source (group of founders) and a relatively recent time of divergence of the chum salmon populations from the region examined.     

Mitochondrial DNA data unveil highly divergent populations within the genus <Emphasis Type="Italic">Hynobius</Emphasis> (Caudata: Hynobiidae) in South Korea

Korean salamanders of the genus Hynobius are currently classified into 3 species, H. leechii, H. quelpaertensis, and H. yangi. To investigate the phylogenetic relationship of these species, we analyzed the partial sequence of mitochondrial cytochrome b gene (907 bp) of 197 specimens from 43 regions in South Korea. Of these specimens, 93 were additionally examined with 12S rRNA (799 bp). Based on the partial sequence of the mitochondrial cytochrome b gene and 12S rRNA, 89 and 36 haplotypes were defined, respectively, consisting of six subclades (H. leechii, H. quelpaertensis, H. yangi, HC1, HC2, and HC3). Among these subclades, the three subclades (HC1, HC2, and HC3) were clearly separated from the 3 previously reported species in the genus Hynobius. Pairwise sequence divergence between the six subclades ranged from 6.3 to 11.2% in cytochrome b gene and 2.0 to 4.3% in 12S rRNA. These results indicate there may be more divergent populations than the three currently described. Moreover, the estimation of divergence time revealed that the Hynobius species in South Korea diverged during the Miocene epoch, approximately 9 — 5 MYA. In addition, we confirmed the distribution of the three known species (H. leechii, H. quelpaertensis, and H. yangi) and determined the distributions of new, distinct groups (or subclades; HC1, HC1, and HC3). To more accurately establish the taxonomic status and population structure, further genetic, morphological, and ecological studies will be needed.     

Genetic divergence of mykizha (Parasalmo (Oncorhynchus) mykiss) from Kamchatka inferred from restriction analysis and sequencing of mtDNA cytochrome b gene

The populations of mykizha Parasalmo (O.) mykiss from western and eastern coasts of Kamchatka were studied by restriction analysis of a fragment of fish mitochondrial genome that included the control region and the region of the cytochrome b gene (cytb). The restriction patterns obtained with five enzymes (MspI; Tru1I; RsaI; BsuRI; DdeI) were identical in all studied individuals. Sequencing of the cytb gene showed high similarity between all samples (99.6-100%). In general, the geographical group of mykiss from Kamchatka is monophyletic with low genetic divergence at the population level. Shantarian mykiss originates most likely from that native to Kamchatka.     

Molecular phylogenetics of Iberian wall lizards (Podarcis): is Podarcis hispanica a species complex?

Phylogenetic relationships between species and morphotypes of Podarcis wall lizards from the Iberian Peninsula and north Africa were estimated using partial 12S rRNA and cytochrome b mitochondrial DNA sequences. All species except Podarcis hispanica form monophyletic units. P. hispanica is paraphyletic, although all identified morphotypes are monophyletic. These morphotypes represent highly divergent lineages showing 10-15% pairwise sequence divergence with the cytochrome b gene. The data suggest that P. hispanica is a species complex. We recommend using P. hispanica* until additional sampling delimits the number and ranges of species currently referred to P. hispanica. P. carbonelli, which has recently been raised to species status, is confirmed as a genetically distinct form. P. atrata is genetically distinct, but much more closely related to some populations of P. hispanica than previously thought.     

Variability of nucleotide sequences of the mitochondrial DNA cytochrome c gene in dolly varden and taranetz char

Nucleotide sequence of the 307-bp fragment of the mitochondrial DNA cytochrome b gene was determined in representatives of the three species of the Salvelinus genus, specifically, dolly varden char (S. malma), taranetz char (S. taranetzi), and white-spotted char (S. leucomaenis). These results pointed to a high level of mitochondrial DNA (mtDNA) divergence between white-spotted char and dolly varden char, on the one hand, and taranetz char, on the other (the mean d value was 5.45%). However, the divergence between the dolly varden char and taranetz char was only 0.81%, which is comparable with the level of intraspecific divergence in the dolly varden char (d = 0.87%). It was shown that the dolly varden char mitochondrial gene pool contained DNA lineages differing from the main mtDNA pool at least in the taranetz char-specific mitochondrial lineages. One of these dolly varden char mtDNA lineages was characterized by the presence of the restriction endonuclease MspI-D variant of the cytochrome b gene. This lineage was widely distributed in the Chukotka populations but it was not detected in the Yana River (Okhotsk sea) populations. These findings suggest that dolly varden char has a more ancient evolutionary lineage, diverging from the common ancestor earlier than did taranetz char.     

Variation in mitochondrial DNA and population structure of the Taipei treefrog Rhacophorus taipeianus in Taiwan

Mitochondrial DNA (mtDNA) from 40 samples of the Taipei treefrog Rhacophorus taipeianus collected from seven populations in Taiwan were sequenced to document the DNA sequence variation in anuran mtDNA and to elucidate the phylogeographic population structure in the Taipei treefrog. Sequences of 722–764 bases in length, including a 108-bp segment of the cytochrome b gene and a 614–656-bp D-loop segment, were obtained by direct sequencing using polymerase chain reaction (PCR). The variation in length was due to a 40-bp region that tandemly repeated four to five times in the D-loop region. The first repeat is the most conserved one among the five repeats because there are no variable sites in this repeat. Besides the 40-bp length variation, 28 positions in the 764-bp sequences are variable and distributed evenly in the cytochrome b gene fragment and D-loop region. Variation in the D-loop of the Taipei treefrog is comparable to those of other vertebrates. Two well-differentiated lineages (northern and central) differing by mean sequence divergence of 1.7% are identified and concordant with their geographic distributions. The two lineages are inferred to have split from a common ancestral population in the early Pleistocene. However, the interpopulation divergence of the northern lineage (< 0.33%) is apparently lower than that of the central lineage (1.11%), implying that the two lineages evolved independently and had different demographic histories after divergence. This study reveals that anuran D-loop has potential as a genetic marker in phylogenetic and population genetic analyses of anurans.     

伊洛河马口鱼遗传差异及其种群分化研究

研究利用细胞色素b(Cyt b)基因分析了采自于伊洛河的48个马口鱼(Opsariichthys bidens)个体间的遗传距离, 并构建其系统发育关系。分析结果显示, 48个个体聚为两个支持率为100%的分支, 分支间没有共享单倍型。每个分支的样本覆盖了所有的采样点, 分支内个体间的平均遗传距离为0.2%, 而分支间的遗传距离为3.1%。微卫星分析结果显示, 99.88%的遗传差异来自于种群内个体间, 种群间的差异只占了0.12%, 两个分支种群并没有发生显著的遗传分化(F_st=0.0012, P=1)。以δ13C和δ15N构建了两个分支的生态位, 结果显示, 伊洛河马口鱼的两个分支的营养生态位没有发生分离。基于线粒体Cyt b基因的遗传分歧, 伊洛河马口鱼的两个分支可能代表不同的物种。但它们在种群遗传结构上并没有发生显著的种群分化, 个体间亲缘关系树与系统发育树的分歧暗示种群间不存在生殖隔离, 营养生态位也没有分离。研究结果并不符合隐存种的解释, 伊洛河马口鱼两个分支间线粒体DNA的遗传差异可能源自于祖先种群或者种间杂交。     

Genetic diversity of North American populations of Cristatella mucedo, inferred from microsatellite and mitochondrial DNA

Research over the past 20 years has shown, with the help of molecular markers, that the population genetics and distribution patterns of freshwater invertebrates in North America are often more complex than was previously believed. Here we extend this research to an, as yet, unstudied but widespread and common group, the freshwater bryozoans. Colonies of the bryozoan Cristatella mucedo were collected from a number of lakes across central North America, and were characterized genetically by analysis of microsatellite loci and mitochondrial DNA (mtDNA) cytochrome b sequences. The microsatellites illustrate a pattern of generally diverse and highly differentiated populations that contain little evidence of recent gene flow. The mtDNA sequences yielded highly variable levels of divergence, ranging from 0.0 to 8.8% within populations, and 0.0 to 9.8% among populations. The multiple divergent mtDNA lineages within populations provide evidence for repeated colonization events. The lack of clustering of haplotypes by site suggests that there has been widespread dispersal of multiple genetic lineages since the last ice age. While some of the haplotype lineages may have evolved in disjunct glacial refugia, the maximum levels of divergence predate the time since the last glacial-interglacial cycles. It is likely that multiple factors including vicariance events, patterns of dispersal, localized extinction, and an unusual life history, explain the unique phylogeographic patterns evident today in populations of C. mucedo.     

Subpopulations of Cryptocephalus beetles (Coleoptera: Chrysomelidae): geographically close but genetically far

Abstract. The leaf beetles Cryptocephalus coryli, C. decemmaculatus and C. nitidulus are of conservation concern and are included on the UK Biodiversity Action Plan. The distinctiveness of the disjunct remaining populations of these beetles was compared to that of more continuously distributed Cryptocephalus species. This was carried out with a view to defining evolutionary significant units (ESUs) in the rare species. A portion of the cytochrome b gene, an intergenic spacer and partial tRNA was analysed from 93 specimens of Cryptocephalus beetle (Coleoptera: Chrysomelidae). Considerable sequence divergence was apparent in all the species, even at an intersite scale when the distances between sampled localities were very small (< 1 km). Intrapopulation, intersite and interpopulation divergence observed in the rare species was reflected in the species that have a more continuous distribution, implying that dispersal ability in these species is poor and gene flow can be impeded by relatively trivial barriers to dispersal. The evidence suggests that the disjunct populations of the rare Cryptocephalus species can, tentatively, be considered as ESUs. This has important implications for management strategies and reintroductions.     

Comparing the performance of multiple mitochondrial genes in the analysis of Australian freshwater fishes

In this study, four mitochondrial genes (cytochrome oxidase I, ATPase, cytochrome b and control region) were amplified from most of the fish species found in the fresh waters of south-eastern Queensland, Australia. The performance of these different gene regions was compared in terms of their ability to cluster fish families together in a neighbour-joining tree, both individually by gene and in all combinations. The relative divergence rates of each of these genes were also calculated. The three coding genes (cytochrome oxidase I, ATPase and cytochrome b) recovered similar number of families and had broadly similar divergence rates. ATPase diverged a little more quickly than cytochrome oxidase I and cytochrome b slightly more slowly than cytochrome oxidase I. All two-gene combinations recovered the same number of families. Results from the control region were much more variable, and, although generally possessing more diversity than the other regions, were sometimes less variable.     

On the taxonomic rank of ciscaucasicus and its relationships with the pygmy wood mouse Sylvaemus uralensis inferred from the mtDNA cytochrome b gene sequence

To specify the taxonomic rank of form ciscaucasoides (independent species Sylvaemus ciscaucasoides, or intraspecific form of pygmy wood mouse, S. uralensis), a 402-bp the mtDNA cytochrome b gene fragment (402 bp) was examined in S. ciscaucasoides individuals from six geographic localities of the Caucasus and Ciscaucasus, (Krasnodar krai and Adygeya Republic) and 17 S. uralensis individuals from seven localities of the Russian Plai (Saratov oblast, Smolensk oblast, Voronezh oblast, Tula oblast, Moscow oblast, and Tver' oblast). For comparison, the cytochrome b gene was partly sequenced in the samples of yellow necked, S. flavicollis (n = 2, Samara oblast), and Caucasian, S. ponticus (n = 6, Krasnodar krai), wood mice. One Mus musculus specimen from Western Europe, whose nucleotide sequences were deposed in the GenBank, was used as intergeneric outgroup. Phylogenetic trees for the forms examined were constructed based on the mtDNA sequence variation and using the neighbor joining and maximum parsimony methods. The network of the cytochrome b haplotypes was also constructed. The level of genetic divergence was evaluated using Kimura's two-parameter algorithm. Based on the data on the sequence variation in a 402-bp mtDNA cytochrome b gene fragment, the hypothesis on the species status of the ciscaucasicus form was. The mean intergroup distances (d) between the geographic groups of S. uralensis varied from 0.0036 to 0.0152. At the same time, the distances between the pygmy wood mice and the group of S.flavicollis-S. ponticus varies in the range from 0.0860 to 0.0935, and the level of intergeneric genetic differentiation (Sylvaemus-Mus) is higher than the latter index (d = 0.142). Ciscaucasoides should be considered as geographic substitution form of S. uralensis. Furthermore, the Caucasian populations of S. uralensis (= ciscaucasoides) were characterized by a threefold lower value of intergroup genetic divergence (d = 0.0062) than the East European populations (d= 0.0179). This finding pointed to some isolation of Caucasian populations of pygmy wood mouse and depletion of their gene pool. However other molecular genetic data (similarity of nucleotide composition and consistence of the levels of intra- and intergroup distances) suggest the absence of geographic subdivision between Caucasian and East European populations ofS. uralensis relative to the molecular marker examined.     

Mitochondrial genomes and divergence times of crocodile newts: inter-islands distribution of Echinotriton andersoni and the origin of a unique repetitive sequence found in Tylototriton mt genomes

Crocodile newts, which constitute the genera Echinotriton and Tylototriton, are known as living fossils, and these genera comprise many endangered species. To identify mitochondrial (mt) genes suitable for future population genetic analyses for endangered taxa, we determined the complete nucleotide sequences of the mt genomes of the Japanese crocodile newt Echinotriton andersoni and Himalayan crocodile newt Tylototriton verrucosus. Although the control region (CR) is known as the most variable mtDNA region in many animal taxa, the CRs of crocodile newts are highly conservative. Rather, the genes of NADH dehydrogenase subunits and ATPase subunit 6 were found to have high sequence divergences and to be usable for population genetics studies. To estimate the inter-population divergence ages of E. andersoni endemic to the Ryukyu Islands, we performed molecular dating analysis using whole and partial mt genomic data. The estimated divergence ages of the inter-island individuals are older than the paleogeographic segmentation ages of the islands, suggesting that the lineage splits of E. andersoni populations were not caused by vicariant events. Our phylogenetic analysis with partial mt sequence data also suggests the existence of at least two more undescribed species in the genus Tylototriton. We also found unusual repeat sequences containing the 3' region of cytochrome apoenzyme b gene, whole tRNA-Thr gene, and a noncoding region (the T-P noncoding region characteristic in caudate mtDNAs) from T. verrucosus mtDNA. Similar repeat sequences were found in two other Tylototriton species. The Tylototriton taxa with the repeats become a monophyletic group, indicating a single origin of the repeat sequences. The intra-and inter-specific comparisons of the repeat sequences suggest the occurrences of homologous recombination-based concerted evolution among the repeat sequences.     

Mitochondrial DNA signature for range-wide populations of Bicyclus anynana suggests a rapid expansion from recent refugia

This study investigates the genetic diversity, population structure and demographic history of the afrotropical butterfly Bicyclus anynana using mitochondrial DNA (mtDNA). Samples from six wild populations covering most of the species range from Uganda to South Africa were compared for the cytochrome c oxidase subunit gene (COI). Molecular diversity indices show overall high mtDNA diversity for the populations, but low nucleotide divergence between haplotypes. Our results indicate relatively little geographic population structure among the southern populations, especially given the extensive distributional range and an expectation of limited gene flow between populations. We implemented neutrality tests to assess signatures of recent historical demographic events. Tajima's D test and Fu's F(S) test both suggested recent population growth for the populations. The results were only significant for the southernmost populations when applying Tajima's D, but Fu's F(S) indicated significant deviations from neutrality for all populations except the one closest to the equator. Based on our own findings and those from pollen and vegetation studies, we hypothesize that the species range of B. anynana was reduced to equatorial refugia during the last glacial period, and that the species expanded southwards during the past 10.000 years. These results provide crucial background information for studies of phenotypic and molecular adaptation in wild populations of B. anynana.