No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Fewer species of Argosarchus and Clitarchus stick insects (Phasmida, Phasmatinae): Evidence from nuclear and mitochondrial DNA sequence data
Abstract
The systematics of three genera of New Zealand stick insect in the subfamily Phasmatinae were investigated in light of inconsistencies in morphological variability within and among species. We sequenced a region of the mitochondrial genome, cytochrome oxidase (COI & COII; 1448 bp), and a nuclear marker, the internal transcribed spacers (ITS1 & ITS2; 1804 bp) from 49 stick insects. Mitochondrial DNA sequence divergences among the three genera (Argosarchus, Clitarchus and Acanthoxyla) were relatively high (∼12%) but the current taxonomy within genera was not supported. Within the three genera, low levels of genetic divergence were observed at both nuclear and mitochondrial loci, and phylogenetic analyses failed to support reciprocal monophyly of the two species in Argosarchus and Clitarchus. Sympatric individuals of Argosarchus spiniger and A. horridus were more closely related to each other than to members of their respective morphospecies from elsewhere. No males were found in the Chatham Island population of Argosarchus and although this population has been referred to as A. schauinslandi, genetic and morphological evidence does not support its distinction from mainland Argosarchus. Likewise, individuals identified as Clitarchus tuberculatus were genetically identical, or most similar to, C. hookeri from the same or adjacent sites rather then grouping with the stick insects they were morphologically most similar to. Lack of spatial, behavioural or ecological evidence concordant with the described species A. spiniger, A. schauinslandi and C. tuberculatus leads us to infer that these species are synonymies of A. horridus and C. hookeri respectively. We conclude that Argosarchus and Clitarchus have each been over-split and actually consist of a single morphologically polymorphic, facultative parthenogenetic species. The genus Acanthoxyla with eight described species also has low levels of genetic divergence, similar to those found in Argosarchus and Clitarchus. A possible hybrid origin of Acanthoxyla involving its sister genus Clitarchus is implied by sharing of ITS sequence variants, but further sampling is needed before the species status of these obligate parthenogenetic lineages can be resolved. In contrast to some New Zealand Orthoptera, the Phasmatinae show little genetic variation suggesting coalescence in recent times, possibly reflecting lineage sorting in the Pleistocene.
... Most recently, Salmon (1991) described the New Zealand species C. tuberculatus Salmon for female specimens of Clitarchus that were brown coloured with abundant tubercles on the body. Subsequent genetic studies Buckley et al. , 2010a showed that individual females matching the description of C. tuberculatus could not be differentiated genetically from C. hookeri suggesting that C. tuberculatus is a synonym of C. hookeri (see, also, Jewell & Brock 2002), and it was treated as such by Trewick et al. (2005), Buckley et al. (2008Buckley et al. ( , 2010a, but not formalised. Clitarchus is placed in the Phasmatidae, Phasmatinae, Acanthoxylini by Günther (1953) and subsequent cataloguers (Otte and Brock 2005). ...
... The occurrence of two undescribed species and the presence of likely synonymies indicate that a revision of the genus is required (Jewell and Brock 2002;Buckley et al. 2010a). Many New Zealand stick insects demonstrate extreme variation in colouration and the size and distribution of spines and tubercles (Salmon 1991;Trewick et al. 2005;Buckley et al. 2010a). For these reasons, attention to the structures of the terminalia is likely to yield more robust information on species limits. ...
... Salmon (1991) also believed the species to be parthenogenetic and he apparently found no males associated with the C. tuberculatus females or males that matched the morphology of the female. However, our own research and that of Trewick et al. (2005) has shown populations of what are clearly C. hookeri containing females matching the type of C. tuberculatus. We have also observed many males that are brown coloured and with many tubercles, like the females of C. tuberculatus, and we therefore conclude that these different morphs represent a single species. ...
We describe two new species of Clitarchus Stål from Northland, New Zealand. Clitarchus rakauwhakanekeneke sp. nov. is described from the Poor Knights Islands and Clitarchus tepaki sp. nov. is described from the Te Paki / North Cape area and the Karikari Peninsula at the northernmost tip of New Zealand. Two new synonymies are proposed including Cli-tarchus multidentatus Brunner (syn. nov.) and Clitarchus tuberculatus Salmon (syn. nov.) as synonyms of Clitarchus hookeri (White). Clitarchus magnus Brunner, recorded from Thailand, is transferred to Ramulus Saussure and given the replacement name Ramulus changmaiense nom. nov. The holotype of C. multidentatus was recorded as being collected from New Caledonia; however we believe this is a labelling error and the specimen was collected from New Zealand. These taxonomic changes render Clitarchus endemic to New Zealand and consisting of three species; C. hookeri, C. rakauwhakanekeneke and C. tepaki. Keys to the adult males and females of Clitarchus species are given in addition to notes on host plants, ecology and geographic distributions.
... O'Neill et al. 2009), A. horridus is an ecological generalist. A further interesting aspect of A. horridus is that not all populations have been observed to possess males (Table 1; see also Trewick et al. 2005) and therefore like many stick insects (Salmon 1955; Bergerard 1962; Law Populations where males and females were collected are indicated by ''M ⁄ F'' and populations where only females were collected are indicated by '' & Crespi 2002; Scali et al. 2003), A. horridus appears to be capable of parthenogenetic reproduction. Understanding geographic parthenogenesis (Vandel 1928) is viewed as important to the broader question of the prevalence of sexual reproduction in nature (Bengtsson 2003). ...
... For this reason our sampling within populations is low (Table 1 ). Although the genus Argosarchus contains several available names, these are all regarded as junior synonyms of A. horridus (Hutton 1899; Salmon 1991; Jewell & Brock 2002; Trewick et al. 2005). Our study of the male and female terminalia of Argosarchus, including type specimens, has revealed no significant variation (T. ...
... The Chatham Islands sample of A. horridus is genetically identical to individuals from the eastern coast of South Island (clade F), and a close relationship between the Chatham and mainland populations was also observed by Trewick et al. (2005). The phylogenetic grouping of the Chatham Islands clades to either east coast of mainland New Zealand or widespread mainland New Zealand clades was noted by Skelley & Leschen (2007) for other New Zealand taxa, and this pattern is observed in A. horridus. ...
We have used phylogeographic analysis of mitochondrial DNA (COI and COII genes) and ecological niche modelling (ENM) to reconstruct the population history of Argosarchus horridus (White), a widespread species of New Zealand stick insect. These data were used to address outstanding questions on the role of glacial refugia in determining the distribution and genetic structure of New Zealand species. Phylogeographic analysis shows a general pattern of high diversity in upper North Island and reduced diversity in lower North Island and South Island. The ENM indicates that during the last glacial maximum, A. horridus was largely restricted to refugia around coastal areas of North Island. The ENM also suggests refugia on the northeast coast of South Island and southeast coast of North Island and this prediction is verified by phylogeographic analysis, which shows a clade restricted to this region. Argosarchus horridus is also most likely a geographic parthenogen where males are much rarer at higher latitudes. The higher levels of genetic variation in northern, bisexual populations suggest southern and largely unisexual populations originated from southwardly expanding parthenogenetic lineages. Bayesian skyline analysis also provides support for a recent population size increase consistent with a large increase in geographic distribution in the late Pleistocene. These results exemplify the utility of integrating ENM and phylogeographic analysis in testing hypotheses on the origin of geographic parthenogenesis and effects of Pleistocene environmental change on biodiversity.
... Doch eine verlässliche Hypo- these zu dieser Phylogenie existiert bei den Phasmatodea bislang nicht. In molekular-phylogenetischen Analysen wurde bislang lediglich die Evolution nahe verwandter Arten eingehend untersucht, so z.B. bei der ursprüngli- chen kalifornischen Gattung Timema ( Crespi & Sandoval 2000;Law & Crespi 2002a, b), bei den europäischen Taxa Bacillus, Clonopsis und Lepty- nia ( Mantovani et al. 2001;Scali et al. 2003;Passamonti et al. 2004) und bei einigen nahe miteinander verwandten neuseeländischen Stabschrecken (Morgan- Trewick et al. 2005Trewick et al. , 2008Buckley et al. 2008). ...
... Dies gilt ebenso für die Pachymorphinae, von denen die australische Pachymor- pha mit großer Sicherheit zu den Lanceocercata gehört (Bradler 1999a). Die Lanceocercata umfassen auch die neuseeländischen Stabschrecken, die ursprünglich zu den Pachymorphinae (Micrarchus) und Phasmatinae (Argosar- chus, Clitarchus) gezählt wurden und die nach bisheriger Ansicht keine engen phylogenetischen Beziehungen zu australischen Formen haben sollen ("the wingless New Zealand fauna are not closely related to Australian phas- mids […] which include many winged species", Jewell & Brock 2002: 189;Trewick et al. 2005). Die Annahme, dass die neuseeländischen Taxa ein Monophylum bilden Trewick et al. 2008), lässt sich hier nicht bestätigen. ...
... The reason for this peculiar distribution is probably derived from range expansion after the Last Glacial Maximum (LGM) [8,10], because asexual reproduction is under positive selection when populations are growing, such as during range expansion [11] and when densities are low and gene flow inhibits local adaptation [12]. No ecological divergence is known to distinguish the parthenogenetic and sexual lineages, which are both silent, nocturnal, and feed on the same host plants (Leptospermum scoparium-m¯ anuka and Kunzea ericoides-k¯ anuka), where they also reproduce [13]. Taking advantage of this unusual system, the aim of this study was to explore morphological and chemical trait variation in Clitarchus hookeri to identify differences between males and females and between sexual and parthenogenetic females. ...
The New Zealand stick insect Clitarchus hookeri has both sexual and parthenogenetic (all-female) populations. Sexual populations exhibit a scramble competition mating system with distinctive sex roles, where females are signalers and males are searchers, which may lead to differences in the chemical and morphological traits between sexes. Evidence from a range of insect species has shown a decay of sexual traits is common in parthenogenetic lineages, especially those traits related to mate attraction and location, presumably due to their high cost. However, in some cases, sexual traits remain functional, either due to the recent evolution of the parthenogenetic lineage, low cost of maintenance, or because there might be an advantage in maintaining them. We measured morphological and chemical traits of C. hookeri to identify differences between males and females and between females from sexual and parthenogenetic populations. We also tested the ability of males to discriminate between sexual and parthenogenetic females in a laboratory bioassay. Our results show that male C. hookeri has morphological traits that facilitate mobility (smaller body with disproportionately longer legs) and mate detection (disproportionately longer antennae), and adult females release significantly higher amounts of volatile organic compounds than males when this species is sexually active, in accordance with their distinctive sex roles. Although some differences were detected between sexual and parthenogenetic females, the latter appear to maintain copulatory behaviors and chemical signaling. Males were unable to distinguish between sexual and parthenogenetic females, suggesting that there has been little decay in the sexual traits in the parthenogenetic lineage of C. hookeri.
... Clitarchus and another New Zealand genus, Argosarchus (Phasmatidae), are also found to be facultatively thelytokous (Salmon 1955;Trewick et al. 2005;Buckley et al. 2009;Morgan-Richards et al. 2010). The genera mentioned above are iconic examples of geographical parthenogenesis in animals. ...
In hexapods, unlike the majority of animals, development without fertilization is a common phenomenon. They evolved a striking diversity of unisexual reproductive types that include a variety of modes starting from spontaneous parthenogenesis in females to the production of impaternate males with different variants in between. Many reports about parthenogenetic species have accumulated over time. Here, we present a review of various parthenogenetic hexapod groups with a particular focus on their chromosome systems and ploidy level. We show that conclusions about the reproductive mode often lack solid evidence and sometimes inefficiently demonstrate how parthenogenesis is maintained in corresponding groups. In this review, basal hexapods (Protura, Collembola, Diplura), primarily wingless insect groups (‘Apterygota’) and non-holometabolous insects are listed with references to a variety of their unisexual reproductive modes.
... It has been suggested that during the Last Glacial Maximum, this species was restricted to several refugia located on the northern half of North Island, followed by subsequent population expansion across the country [22]. Molecular resources from C. hookeri are lacking, restricted to a few neutral gene markers that were sequenced and used for phylogenetic construction [4, 19, 20, 22, 25] and a head and prothorax transcriptome that was constructed for the study of cold temperature adaptation [15]. In order to examine digestive enzymes, olfaction-associated proteins and accessory gland proteins in a New Zealand stick insect species, we carried out an RNA-Seq experiment by sequencing the transcriptomes of C. hookeri female antennae, midgut and male terminalia. ...
Phasmatodea, more commonly known as stick insects, have been poorly studied at the molecular level for several key traits, such as components of the sensory system and regulators of reproduction and development, impeding a deeper understanding of their functional biology. Here, we employ de novo transcriptome analysis to identify genes with primary functions related to female odour reception, digestion, and male sexual traits in the New Zealand common stick insect Clitarchus hookeri (White). The female olfactory gene repertoire revealed ten odorant binding proteins with three recently duplicated, 12 chemosensory proteins, 16 odorant receptors, and 17 ionotropic receptors. The majority of these olfactory genes were over-expressed in female antennae and have the inferred function of odorant reception. Others that were predominantly expressed in male terminalia (n = 3) and female midgut (n = 1) suggest they have a role in sexual reproduction and digestion, respectively. Over-represented transcripts in the midgut were enriched with digestive enzyme gene families. Clitarchus hookeri is likely to harbour nine members of an endogenous cellulase family (glycoside hydrolase family 9), two of which appear to be specific to the C. hookeri lineage. All of these cellulase sequences fall into four main phasmid clades and show gene duplication events occurred early in the diversification of Phasmatodea. In addition, C. hookeri genome is likely to express γ-proteobacteria pectinase transcripts that have recently been shown to be the result of horizontal transfer. We also predicted 711 male terminalia-enriched transcripts that are candidate accessory gland proteins, 28 of which were annotated to have molecular functions of peptidase activity and peptidase inhibitor activity, two groups being widely reported to regulate female reproduction through proteolytic cascades. Our study has yielded new insights into the genetic basis of odour detection, nutrient digestion, and male sexual traits in stick insects. The C. hookeri reference transcriptome, together with identified gene families, provides a comprehensive resource for studying the evolution of sensory perception, digestive systems, and reproductive success in phasmids.
... DNA barcoding has its disadvantages (Taylor & Harris 2012), as it is of limited use to identify certain species and is not efficiently amplified by PCR in all animal taxa, but it may be used as an additional, and apparently very powerful, method in taxonomy (Schlick-Steiner et al. 2010). Despite the wide use of DNA barcodes in the current taxonomy and biodiversity studies, the method has only scarcely been used in taxonomic studies except for the Australian and New Zealand stick insect (e.g., Trewick et al. 2005Trewick et al. , 2008Buckley et al. 2009Buckley et al. , 2010Morgan-Richards et al. 2010;Schwander et al. 2011;Nosil et al. 2012). The COII mitochondrial gene has been sequenced over a wide variety of taxa and has proven to be useful for phylogenetic research, especially in Phasmida (Mantovani et al. 2001;Passamonti et al. 2004;Ghiselli et al. 2007;Scali 2009;Scali et al. 2012Scali et al. , 2013. ...
Male, female and egg of Pijnackeria recondita sp. n. are described from specimens collected at about 2,000 m in Sierra Nevada (Spain) feeding on Cytisus scoparius. The number of antennae segments in males, the smooth thorax in females and the different sculpturing of the egg capsule are the main differences from the other species of the genus. In addition, DNA barcode sequences (COI and COII) clearly differ from the other Iberian species of the genus. For COI, K2P minimum distance between the new species and the most morphological related species, Pijnackeria hispanica (Bolivar, 1878), showed a mean of 8%. In the case of COII, comparison with the other species of Pijnackeria, showed a K2P minimum distance range from 8 to 10.5% (mean 9.2%); and comparison with the species of the related genus Leptynia, showed a K2P minimum distance range from 7.1 to 10.5%.
... Such results are in keeping with previous studies of taxa that incorporate New Zealand and Chatham Island sampling, where divergence estimates range from populations with identical cpDNA haplotypes in the fern Asplenium hookerianum [52], to 2% mtDNA sequence divergence in a skink Oligosoma nigriplantare nigriplantare [53], and plants with up to 6.4% sequence divergence [54]. Among these examples are spiders [55], insects (cicada [56]; cockroach [51]; damselfly [57]; stick insect [58]), amphipods [59], isopods [60], and birds (rails [61]; robin [62]; parakeet [63,64]; pigeon [18]). Given the wide taxonomic and ecological diversity represented in these studies, perhaps this wide range in divergence estimates reflects differences in biology rather than history, or a combination of the two. ...
A prediction in phylogeographic studies is that patterns of lineage diversity and timing will be similar within the same landscape under the assumption that these lineages have responded to past environmental changes in comparable ways. Eight invertebrate taxa from four different orders were included in this study of mainland New Zealand and Chatham Islands lineages to explore outcomes of island colonization. These comprised two orthopteran genera, one an endemic forest-dwelling genus of cave weta (Rhaphidophoridae, Talitropsis) and the other a grasshopper (Acrididae, Phaulacridum) that inhabits open grassland; four genera of Coleoptera including carabid beetles (Mecodema), stag beetles (Geodorcus), weevils (Hadramphus) and clickbeetles (Amychus); the widespread earwig genus Anisolabis (Dermaptera) that is common on beaches in New Zealand and the Chatham Islands, and an endemic and widespread cockroach genus Celatoblatta (Blattodea). Mitochondrial DNA data were used to reconstruct phylogeographic hypotheses to compare among these taxa. Strikingly, despite a maximum age of the Chathams of ~4 million years there is no concordance among these taxa, in the extent of genetic divergence and partitioning between Chatham and Mainland populations. Some Chatham lineages are represented by insular endemics and others by haplotypes shared with mainland populations. These diverse patterns suggest that combinations of OPEN ACCESS Insects 2011, 2 370 intrinsic (taxon ecology) and extrinsic (extinction and dispersal) factors can result in apparently very different biogeographic outcomes.
... For each specimen, a leg was removed for DNA extraction using a salting-out method (Sunnucks & Hales, 1996;Trewick et al., 2005). For 16 Acanthoxyla, partial mitochondrial cytochrome oxidase subunit I and II were amplified and sequenced (Morgan-Richards & using primers C1-J-2195, L2-N-3014, TL2-J-3034, and TK-N-3785 (Simon et al., 1994). ...
Although hybridisation is common in animals, it rarely results in speciation. Yet, many examples of hybrid species have been documented in one animal group, the stick insects (Phasmida).The New Zealand stick insect Acanthoxyla is of particular interest as the entire genus is of hybrid origin and consists of eight morphological forms recognised as species, all of which are obligate parthenogens.Using five complementary techniques on the same individuals, our study confirms that both triploids and diploids are present in Acanthoxyla populations, and further, that some individuals contain both diploid and triploid cells.Chromosome spreads and estimates of relative DNA content from flow cytometry provided contrasting information about the ploidy of this unusual parthenogenetic genus.Analysis of morphometric variation showed no correlation with ploidy level in Acanthoxyla, and also mtDNA sequence networks failed to distinguish morphospecies or ploidy level.Unexpectedly, cloned sequences of a putatively single‐copy nuclear marker were also unhelpful in distinguishing ploidy, instead indicating that phosphoglucose isomerase is likely to be a multiple copy gene.We propose a mechanism for the evolution of the Acanthoxyla lineage and suggest that interpretation may be complicated by the presence of individuals that are diploid and triploid mosaics.
... The phylogenetic analyses of Trewick, Morgan-Richards & Collins, (2008) and Buckley et al., (2010b) showed that Pseudoclitarchus was sister group to the widespread genus Clitarchus. Clitarchus contains one recognized described species (Trewick, Goldberg & Morgan-Richards, 2005;Buckley et al., 2010a) and two undescribed species (Watt, 1982;Buckley et al., 2010a) in New Zealand. Clitarchus hookeri (White) is found throughout the North Island and northern/coastal regions of the South Island, whereas Clitarchus sp. ...
We have used comparative phylogenetic analysis to infer the age and biogeographical origins of the Three Kings Islands insect fauna, an archipelago only 56 km off the northern tip of New Zealand. We densely sampled six insect lineages (five Coleoptera, Brachynopus latus, Brachynopus scutellaris, Tarphiomimus spp., Epistranus lawsoni, and Syrphetodes spp., and one Phasmatodea, Pseudoclitarchus sentus) throughout New Zealand and sequenced mitochondrial DNA to assess phylogenetic relationships and determine ages of haplotype lineages on the Three Kings Islands. We recovered two biogeographical patterns. The first pattern was seen in three taxa, B. latus, Syrphetodes spp., and E. lawsoni, which had sister group relationships between the Three Kings and the adjacent North Cape region at the very northern tip of New Zealand. The second pattern, inferred in P. sentus, B. scutellaris, and Tarphiomimus spp., was where Three Kings lineages had sister groups that were widespread throughout most or all of New Zealand. The divergence dates, estimated using a range of previously estimated substitution rates, ranged from as old as 24 Mya in B. scutellaris to as young as 2.24 Mya in Tarphiomimus. These results are consistent with continual emergent land on the Three Kings Ridge since at least the Miocene and a lack of land connections between the Three Kings Islands and mainland New Zealand during Pleistocene sea‐level lowering. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 108, 361–377.
... Not surprisingly, the cladistic analyses of morphological data by Tilgner (2002) and Bradler (2009) did not corroborate the classification proposed by Günther (1953) and successive authors (Bradley & Galil, 1977;Kevan, 1982;Key, 1991;Zompro, 2004a). Molecular-based phylogenetic studies have mainly focussed on biogeographically restricted taxa such as New Zealand stick insects (Trewick et al., 2005(Trewick et al., , 2008Buckley et al., 2008O'Neill et al., 2009;Buckley & Leschen, 2013), circum-mediterranean taxa (Scali et al., 2003(Scali et al., , 2012(Scali et al., , 2013Ghiselli et al., 2007;Scali, 2009) and the genus Timema in Southern North America (Law & Crespi, 2002a,b;Schwander et al., 2011). ...
The phasmatodeans or stick and leaf insects are considered to be a mesodiverse insect order with more than 3000 species reported mainly from the tropics. The stick insect subfamily Necrosciinae comprises approximately 700 described species in more than 60 genera from the Oriental and Australian region, forming the most species-rich subfamily traditionally recognized within Phasmatodea. However, the monophyly of this taxon has never been thoroughly tested and the evolutionary relationships among its members are unknown. We analyse three nuclear (18S and 28S rDNA, histone 3) and three mitochondrial (CO II, 12S and 16S rDNA) genes to infer the phylogeny of 60 species of stick insects that represent all recognized families and major subfamilies sensu Günther and the remarkable diversity within Necrosciinae. Maximum parsimony, maximum likelihood and Bayesian techniques largely recover the same substantial clades, albeit with highly discordant relationships between them. Most members of the subfamily Necrosciinae form a clade. However, the genus Neohirasea – currently classified within Lonchodinae – is strongly supported as subordinate to Necrosciinae, whereas Baculofractum, currently classified within Necrosciinae, is strongly supported within Lonchodinae. Accordingly, we formally transfer Neohirasea and allied taxa (namely Neohiraseini) to Necrosciinae sensu nova (s.n.) and Baculofractum to Lonchodinae s.n. We also provide further evidence that Leprocaulinus, until recently recognized as Necrosciinae, belongs to Lonchodinae, and forms the sister taxon of Baculofractum. Furthermore, Lonchodinae is paraphyletic under exclusion of Eurycantha and Neopromachus. We reinstate the traditional view that Neopromachus and related taxa (Neopromachini sensu Günther) are a subgroup of Lonchodinae and transfer those taxa + the New Guinean Eurycanthinae accordingly. Morphological evidence largely corroborates our molecular-based findings and also reveals that Menexenus fruhstorferi is a member of the genus Neohirasea and is thus transferred from Menexenus (Lonchodinae) to Neohirasea, as Neohirasea fruhstorferi comb.n. (Necrosciinae s.n.). Other phylogenetic results include Areolatae and Anareolatae each supported as polyphyletic, Heteropteryginae and Lanceocercata (Bayesian analysis) are monophyletic, albeit with low support, and Necrosciinae s.n. and Lonchodinae s.n. are recovered as sister taxa (Bayesian analysis).
... The Chatham Islands have long been assumed to be ancient land because this fitted with a preconception about the vicariant origin of biotas (Craw, 1988). It is now evident from geological (Campbell et al., 1994Campbell et al., , 2006 Campbell, 1998), taxonomic (Emberson, 1998) and molecular data (Trewick, 2000; Trewick et al., 2005; Paterson et al., 2006) that the extant Chathams emerged from the sea 1– 3 Ma. This implies that the ancestors of all of the extensive Chatham's flora and fauna have arrived in a geologically short period across a significant water gap (> 800 km). ...
Islands of the Pacific Ocean have long fascinated evolutionists. Oceanic islands, generally the products of volcanic activity, provide natural experiments as biological populations are well delimited and the age of islands can be determined using radiometric dating. ‘Continental islands’, including New Caledonia and New Zealand, provide equally valuable opportunities for evolutionary study. For students of New Zealand biogeography, the peculiar composition of the biota coupled with a limited interpretation of geology has resulted in the widespread acceptance that the flora and fauna is primarily ancient and of vicariant Gondwanan origin. There is increasing evidence from molecular data that much of this biodiversity is the product of evolution following relatively recent colonization. Such data have prompted biologists to consider geological information on New Zealand in more detail. At the heart of the issue is the question of whether modern New Zealand has a terrestrial link through time with the continent Zealandia that split from Gondwanaland some 80 Ma. Zealandia, which includes New Caledonia, Lord Howe Island and several of the subantarctic islands, is now largely submerged, and New Zealand's present terrestrial existence is the product of tectonic activity initiated around 26 Ma. We argue that for the purposes of biogeographical interpretation, New Zealand can be treated as an oceanic island.
... Hybridization is common in stick insects (Scali et al., 2003; Buckley et al., 2008; Schwander & Crespi, 2009b), and will increase the incidence of species that are non-monophyletic for mtDNA. Furthermore, lineage sorting and subsequent non-monophyly of diverging species is expected to be common when the ancestral species is highly diverse genetically and has strong phylogeographic structure, as is the case with all New Zealand stick insects examined to date ( Trewick et al., 2005; Buckley et al., 2008; O'Neill et al., 2009). Therefore, the lack of mtDNA monophyly for C. hookeri presents no problem for its recognition as a separate species. ...
Aim Increasing our understanding of the effects of the Last Glacial Maximum (LGM) and determining the location of refugia requires studies on widely distributed species with dense sampling of populations. We have reconstructed the biogeographic history of Clitarchus hookeri (White), a widespread species of New Zealand stick insect that exhibits geographic parthenogenesis, using phylogeographic analysis and ecological niche modelling.Location New Zealand.Methods We used DNA sequence data from the mitochondrial cytochrome c oxidase subunit I gene to reconstruct phylogenetic relationships among haplotypes from C. hookeri and two undescribed Clitarchus species. We also used distribution data from our own field surveys and museum records to reconstruct the geographic distribution of C. hookeri during the present and the LGM, using ecological niche modelling.Results The ecological niche models showed that the geographic distribution of C. hookeri has expanded dramatically since the LGM. Our model predicted large areas of suitable LGM habitat in upper North Island, and small patches along the east coast of South Island. The phylogeographic analysis shows that populations in the northern half of North Island contain much higher levels of genetic variation than those from southern North Island and South Island, and is congruent with the ecological niche model. The distribution of bisexual populations is also non-random, with males completely absent from South Island and very rare in southern North Island.Main conclusions During the LGM C. hookeri was most likely restricted to several refugia in upper North Island and one or more smaller refugia along the east coast of South Island. The unisexual populations predominate in post-glacial landscapes and are clearly favoured in the recolonization of such areas. Our study exemplifies the utility of integrating ecological niche modelling and phylogeographic analysis.
... The New Zealand stick insects are all polyphagous and are especially common on species from the families Cunoniaceae (Weinmannia), Polygonaceae (Muehlenbeckia), Rosaceae (Rubus, Acaenea), Malvaceae (Hoheria), Ericales (Dracophyllum, Cyathodes), Asteliaceae (Astelia) and especially Myrtaceae (e.g. Kunzea, Leptospermum, Metrosideros, Lophomyrtus) (Salmon, 1991; Trewick et al., 2005; Buckley et al., 2008 Buckley et al., , 2009b, c; O'Neill et al., 2009). Many genera from these families that are favoured by the New Zealand species, and other host genera, are present in New Caledonia (e.g. ...
The Lanceocercata are a clade of stick insects (Phasmatodea) that have undergone an impressive evolutionary radiation in Australia, New Caledonia, the Mascarene Islands and areas of the Pacific. Previous research showed that this clade also contained at least two of the nine New Zealand stick insect genera. We have constructed a phylogeny of the Lanceocercata using 2277 bp of mitochondrial and nuclear DNA sequence data to determine whether all nine New Zealand genera are indeed Lanceocercata and whether the New Zealand fauna is monophyletic. DNA sequence data were obtained from mitochondrial cytochrome oxidase subunits I and II and the nuclear large subunit ribosomal RNA and histone subunit 3. These data were subjected to Bayesian phylogenetic inference under a partitioned model and maximum parsimony. The resulting trees show that all the New Zealand genera are nested within a large New Caledonian radiation. The New Zealand genera do not form a monophyletic group, with the genus Spinotectarchus Salmon forming an independent lineage from the remaining eight genera. We analysed Lanceocercata apomorphies to confirm the molecular placement of the New Zealand genera and to identify characters that confirm the polyphyly of the fauna. Molecular dating analyses under a relaxed clock coupled with a Bayesian extension to dispersal-vicariance analysis was used to reconstruct the biogeographical history for the Lanceocercata. These analyses show that Lanceocercata and their sister group, the Stephanacridini, probably diverged from their South American relatives, the Cladomorphinae, as a result of the separation of Australia, Antarctica and South America. The radiation of the New Caledonian and New Zealand clade began 41.06 million years ago (mya, 29.05–55.40 mya), which corresponds to a period of uplift in New Caledonia. The main New Zealand lineage and Spinotectarchus split from their New Caledonian sister groups 33.72 (23.9–45.62 mya) and 29.9 mya (19.79–41.16 mya) and began to radiate during the late Oligocene and early Miocene, probably in response to a reduction in land area and subsequent uplift in the late Oligocene and early Miocene. We discuss briefly shared host plant patterns between New Zealand and New Caledonia. Because Acrophylla sensu Brock & Hasenpusch is polyphyletic, we have removed Vetilia Stål from synonymy with Acrophylla Gray.
... cichlid fishes: Schluter and Rambaut, 1996; Galis and Metz, 1998; stickleback fishes: Rundle et al., 2000; Fucus macroalgae: Pereyra et al., 2009) that reproductive isolation can evolve over relatively short (e.g., postglacial) timeframes. In biogeographic terms, the close genetic relationship observed between Chatham Island and mainland New Zealand populations of Durvillaea certainly supports the recent origins proposed for Chatham Islands biota (Trewick, 2000; Trewick et al., 2005; Paterson et al., 2006; Heenan et al., 2010). ...
Durvillaea (southern bull-kelp) is an economically and ecologically important brown algal genus that dominates many exposed, rocky coasts in the cold-temperate Southern Hemisphere. Of its five currently-recognized species, four are non-buoyant and restricted to the south-western Pacific, whereas one is both buoyant and widely distributed. Durvillaea has had an unsettled taxonomic history. Although its position within the brown algae (Phaeophyceae) has now been largely resolved through the use of molecular techniques, the taxonomic status of several Durvillaea species/morphotypes remains unresolved. Previous molecular phylogenetic studies of phaeophycean taxa have included few Durvillaea samples, and have consequently paid little or no attention to variation within this genus. The current study presents phylogenetic analyses of four genetic markers (mitchondrial: COI; chloroplast: rbcL; and nuclear: 18S and 28S) to resolve phylogenetic relationships within Durvillaea. Results support the monophyly of solid-bladed taxa D. willana, D. potatorum, and D. sp. A (an undescribed species from the Antipodes Islands), whereas the widespread, buoyant D. antarctica is paraphyletic, with solid-bladed D.chathamensis placed sister to a D. antarctica clade from northern NZ but within D. antarctica sensu lato. The phylogenetic and ecological diversity detected within D. antarctica indicate that it is a species complex of five deeply divergent clades. Under a phylogenetic species concept, Durvillaea can be interpreted as a complex of nine distinct evolutionary lineages, only one of which has an intercontinental distribution ('subantarctic'D. antarctica).
... The most recent revision (Salmon, 1991) of the genus Clitarchus recognized two species within New Zealand, C. hookeri and C. tuberculatus Salmon. However, we can find no diagnostic morphological characters to dif-ferentiate these two species and therefore we only recognize C. hookeri (see also Trewick et al., 2005). We sampled 13 individuals of this species from throughout its geographic range (Table 1 andFig. ...
The New Zealand stick insect genus Acanthoxyla Uvarov is extremely unusual among higher taxa of animals in that all known species are obligate parthenogens. We have used a combination of the mitochondrial DNA genes cytochrome oxidase subunits I and II, 28S nuclear ribosomal RNA, and the two single-copy nuclear genes elongation factor 1alpha and phosphoglucose isomerase to test hypotheses on the role of hybridization in the evolution of this genus. Alleles at the single-copy nuclear loci in three sampled species of Acanthoxyla were resolved by cloning the PCR products. Analysis of multilocus genotypes shows that most sampled individuals of Acanthoxyla possess three alleles at the single-copy nuclear loci, which we have interpreted to indicate triploidy. Because most of the alleles from Acanthoxyla form a monophyletic group, including sets of alleles possessed by the putative triploids, we have inferred that the extant parthenogenetic lineages formed via hybridization between species of Acanthoxyla, at least one of which must have been sexual. More recently, there have been multiple introgression events from the related species Clitarchus hookeri White, although C. hookeri does not appear to be involved with the origin of parthenogenesis in Acanthoxyla. Our study demonstrates the utility of cloning alleles from multiple single-copy nuclear genes for resolving the origins of parthenogenetic lineages.
The hybrid stick insect genus Acanthoxyla Uvarov 1944 is unusual for an obligate parthenogen, in the extreme morphological diversity it exhibits that has led to eight species being recognised. The New Zealand sexual species Clitarchus hookeri [White, A. 1846. The zoology of the Voyage of H.M.S. Erebus and Terror. In: 1 Insects of New Zealand. E.W. Janson, London.] is the putative parental species in the hybridization that gave rise to the hybrid lineage Acanthoxyla. In an effort to identify the maternal ancestor of Acanthoxyla we sequenced nuclear 28S rDNA and/or mtDNA COI & COII of all nine endemic New Zealand stick insect genera, representing 17 of the 22 described species. We also sequenced 28S from eight non-New Zealand stick insects to supplement published 28S sequence data that provided a taxonomically and geographically broad sampling of the phasmids. We applied a novel search algorithm (SeqSSi=Sequence Similarity Sieve) to assist in selection of outgroup taxa for phylogenetic analysis prior to alignment. Phylogenetic reconstructions resolved an exclusively New Zealand clade to which the maternal lineage of Acanthoxyla belonged, but did not support existing higher level taxonomy of stick insects. We did not find a sexual maternal species for Acanthoxyla but phylogenetic relationships indicate that this species lived in New Zealand and could be classified among the New Zealand Phasmatinae. Among the available taxa, the nearest evolutionary neighbours to the New Zealand phasmid fauna as a whole were predominantly from the New Zealand region (Fiji, Australia, New Guinea, New Caledonia and South America). As it appears to be an orphan, it is interesting to speculate that a combination of parthenogenetic reproduction and/or hybrid vigour in Acanthoxyla may have contributed to the extinction of its mother.
Worldwide, parthenogenetic reproduction has evolved many times in the stick insects (Phasmatidae). Many parthenogenetic stick insects show the distribution pattern known as geographic parthenogenesis, in that they occupy habitats that are at higher altitude or latitude compared with their sexual relatives. Although it is often assumed that, in the short term, parthenogenetic populations will have a reproductive advantage over sexual populations; this is not necessarily the case. We present data on the distribution and evolutionary relationships of sexual and asexual populations of the New Zealand stick insect, Clitarchus hookeri. Males are common in the northern half of the species' range but rare or absent elsewhere, and we found that most C. hookeri from putative-parthenogenetic populations share a common ancestor. Female stick insects from bisexual populations of Clitarchus hookeri are capable of parthenogenetic reproduction, but those insects from putative-parthenogenetic populations produced few offspring via sexual reproduction when males were available. We found similar fertility (hatching success) in mated and virgin females. Mated females produce equal numbers of male and female offspring, with most hatching about 9-16 weeks after laying. In contrast, most eggs from unmated females took longer to hatch (21-23 weeks), and most offspring were female. It appears that all C. hookeri females are capable of parthenogenetic reproduction, and thus could benefit from the numerical advantage this yields. Nevertheless, our phylogeographic evidence shows that the majority of all-female populations over a wide geographic area originate from a single loss of sexual reproduction.
New Zealand biogeography has been dominated by the knowledge that its geophysical history is continental in nature. The continental crust (Zealandia) from which New Zealand is formed broke from Gondwanaland ca 80 Ma, and there has existed a pervading view that the native biota is primarily a product of this long isolation. However, molecular studies of terrestrial animals and plants in New Zealand indicate that many taxa arrived since isolation of the land, and that diversification in most groups is relatively recent. This is consistent with evidence for species turnover from the fossil record, taxonomic affinity, tectonic evidence and observations of biological composition and interactions. Extinction, colonization and speciation have yielded a biota in New Zealand which is, in most respects, more like that of an oceanic archipelago than a continent.
The application of new molecular technologies is central to the search for
causal mechanisms capable of explaining the modern-day biogeography of the
southern continents. Projects have previously focused on marine mammals and
birds, but in recent years they have begun to expand in scope. We now describe
the results from three studies carried out recently on parakeets (genus
Cyanoramphus), cicadas (genus
Maoricicada) and geckos (genera
Hoplodactylus and Naultinus) in
the context of the Gondwanan affinities of the New Zealand biota. The work
described here has been the subject of independent reports (see text for
individual references) and their findings have been brought together for the
first time here in a more general synthesis.
▪ Abstract Many uses of gene trees implicitly assume that nominal species are monophyletic in their alleles at the study locus. However, in well-sampled gene trees, certain alleles in one species may appear more closely related to alleles from different species than to other conspecific alleles. Such deviations from species-level monophyly have a variety of causes and may lead to erroneous evolutionary interpretations if undetected. The present paper describes the causes and consequences of these paraphyletic and polyphyletic patterns. It also provides a detailed literature survey of mitochondrial DNA studies on low-level animal phylogeny and phylogeography, results from which reveal the frequency of nonmonophyly and patterns of interpretation and sampling. This survey detected species-level paraphyly or polyphyly in 23% of 2319 assayed species, demonstrating this phenomenon to be statistically supported, taxonomically widespread, and far more common than generally recognized. Our findings call for increa...
The evolutionary relationships of the New Zealand representatives of the family Anostostomatidae were examined using DNA sequence data. All species of giant weta (Deinacrida), tree weta (Hemideina) and tusked weta (Anisoura, Motuweta) were included in the study plus 4 taxa from the large genus of New Zealand ground weta (Hemiandrus). Sequence data from 2 mitochondrial genes (COI and 12S) were analysed to obtain a hypothesis of the evolution of these species. The 3 New Zealand tusked weta species formed a monophyletic clade with respect to the ground weta and to the giant and tree weta clad0e. We found no support for the placement of Anisoura nicobarica Ander within Deinacridinae as has previously been suggested. The giant and tree weta (Deinacridinae) consistently formed a monophyletic clade with respect to the tusked and ground weta. However, we found little support in our data for the reciprocal monophyly of the tree and giant weta genera. The tree weta Hemideina broughi (Buller) appears to be more closely related to Deinacrida pluvialis Gibbs and D. talpa Gibbs than to any other Hemideina species. The deinacridine radiation of leaf-eating weta comprises at least 6 comparatively ancient lineages (Hemideina and Deinacrida). Habitat-specialisation in South Island appears to have evolved in response to habitat diversification associated with Pliocene mountain building.
Brachaspis robustus is an endangeredgrasshopper endemic to South Island, NewZealand. It is both rare and localised;occupying low altitude floodplain terraces andbraided riverbeds of the Mackenzie Basin. Thisis in stark contrast to the two other speciesin this genus (B. nivalis and B.collinus) which occupy montane habitats.Mitochondrial and nuclear sequence data wereemployed to explore genetic diversity andphylogenetic relationships of populations ofBrachaspis with a view to establishingthe status of B. robustus. Molecularevidence indicates that Brachaspisprobably radiated during the Pliocene and thatdivisions within the genus relate more tospatial distribution developed during thePleistocene than to ecology. The mitochondrial(Cytochrome oxidase I) and nuclear (ITS)sequence data indicate that Brachaspisnivalis is divided into northern and southernpopulations. The northern clade is furthersubdivided geographically. The southern cladecomprises alpine populations of B.nivalis and includes the lowland B.robustus. Additionally, it is observed thatsome morphological features previously thoughtto be specific to B. robustus also occurin members of the southern B. nivalisclade. It is suggested that the taxon B.robustus should include all of the southernBrachaspis populations. But it is arguedthat the absence of genetic evidencedistinguishing the endangered population doesnot preclude it from conservation effort. Acombination of morphological and habitatpeculiarities indicate that the survival ofB. robustus (sensu lato) isimportant to the maintenance of diversity.
We inferred a phylogeny for the walking-stick genus Timema (Insecta: Phasmatoptera) using mitochondrial DNA sequence, and we used the phylogeny to infer temporal patterns of speciation and the evolutionary history of parthenogenesis. Maximum parsimony, neighbour-joining and maximum-likelihood analyses of 660 base pairs (bp) of cytochrome oxidase I (COI) yielded phylogenies that were well resolved and topologically identical or very similar. Application of an insect molecular clock for COI suggests that the genus originated in southern California, northern Mexico or Arizona about 20 million years ago and underwent a burst of speciation 1.5 to 3 million years ago during the uplifts of the Sierra Nevada, Coast, and Transverse Ranges. The phylogeny indicates that the three parthenogenetic lineages of Timema have arisen independently and are each closely related to morphologically indistinguishable or similar sexual species. Each of the three lineages exhibits an allopatric or parapatric, and more northerly, distribution with regard to their closest sexual relative. COI divergence levels between each of the three parthenogens and their closest sexual relative suggest ancient origins of parthenogenesis, 1.5 to 3 million years ago, that may coincide with the extensive glaciation that formed the North American ice sheets.
Polymerase chain reaction (PCR) products corresponding to 803 bp of the cytochrome oxidase subunits I and II region of mitochondrial DNA (mtDNA COI-II) were deduced to consist of multiple haplotypes in three Sitobion species. We investigated the molecular basis of these observations. PCR products were cloned, and six clones from one individual per species were sequenced. In each individual, one sequence was found commonly, but also two or three divergent sequences were seen. The divergent sequences were shown to be nonmitochondrial by sequencing from purified mtDNA and Southern blotting experiments. All seven nonmitochondrial clones sequenced to completion were unique. Nonmitochondrial sequences have a high proportion of unique sites, and very few characters are shared between nonmitochondrial clones to the exclusion of mtDNA. From these data, we infer that fragments of mtDNA have been transposed separately (probably into aphid chromosomes), at a frequency only known to be equalled in humans. The transposition phenomenon appears to occur infrequently or not at all in closely related genera and other aphids investigated. Patterns of nucleotide substitution in mtDNA inferred over a parsimony tree are very different from those in transposed sequences. Compared with mtDNA, nonmitochondrial sequences have less codon position bias, more even exchanges between A, G, C and T, and a higher proportion of nonsynonymous replacements. Although these data are consistent with the transposed sequences being under less constraint than mtDNA, changes in the nonmitochondrial sequences are not random: there remains significant position bias, and probable excesses of synonymous replacements and of conservative inferred amino acid replacements. We conclude that a proportion of the inferred change in the nonmitochondrial sequences occurred before transposition. We believe that Sitobion aphids (and other species exhibiting mtDNA transposition) may be important for studying the molecular evolution of mtDNA and pseudogenes. However, our data highlight the need to establish the true evolutionary relationships between sequences in comparative investigations.
If a gene tree is to be judiciously used for inferring the histories of closely related taxa, (1) its topology must be sufficiently resolved and robust that noteworthy phylogenetic patterns can be confidently documented, and (2) sampling of species, populations, and pertinent biological variation must be sufficiently broad that otherwise misleading sources of genetic variation can be detected. These principles are illustrated by the complex gene tree of Neochlamisus leaf beetles that I reconstructed using 90,000 bp of cytochrome oxidase I (COI) and 16S mitochondrial DNA (mtDNA) sequences from over 100 specimens. Cytochrome oxidase I haplotypes varied up to 25.1% within Neochlamisus and up to 11.1% within the gibbosus species group, while exhibiting very low A + T bias for insect mtDNA (63%), low transition saturation, and conservative patterns of amino acid variation. 16S exhibited lower sequence divergences and greater A + T bias and transition saturation than COI, and substitutions were more constrained in stems than in loops. Comparisons with an earlier study of Ophraella leaf beetles highlighted conservative and labile elements of molecular evolution across genes and taxa. Cytochrome oxidase I parsimony and neighbor-joining analyses strongly supported a robust mtDNA genealogy that revealed the monophyly of Neochlamisus and of the gibbosus species group. Phylogeographic relationships suggested that the eastern U.S. gibbosus group derives from southwestern velutinus group ancestors. Haplotypes from individual velutinus group species clustered monophyletically, as expected. However, haplotypes from each of several gibbosus group taxa were polyphyletically distributed, appearing in divergent parts of the tree. 16S provided a less-resolved gibbosus group topology that was congruent with the COI tree and corroborated patterns of mitochondrial polyphyly. By subsampling haplotypes corresponding to particular species, populations, and ecological variants of gibbosus group taxa, I demonstrate that recovered topologies and genetic distances vary egregiously according to sampling regime. This study thus documents the potentially dire consequences of inadequate sampling when inferring the evolutionary history of closely related and mitochondrially polyphyletic taxa.
Most research on the biological effects of Pleistocene glaciation and refugia has been undertaken in the northern hemisphere and focuses on lowland taxa. Using single-strand conformation polymorphism (SSCP) analysis and sequencing of mitochondrial cytochrome oxidase I, we explored the intraspecific phylogeography of a flightless orthopteran (the alpine scree weta, Deinacrida connectens) that is adapted to the alpine zone of South Island, New Zealand. We found that several mountain ranges and regions had their own reciprocally monophyletic, deeply differentiated lineages. Corrected genetic distance among lineages was 8.4% (Kimura 2-parameter [K2P]) / 13% (GTR + I + Gamma), whereas within-lineage distances were only 2.8% (K2P) / 3.2% (GTR + I + Gamma). We propose a model to explain this phylogeographical structure, which links the radiation of D. connectens to Pliocene mountain building, and maintenance of this structure through the combined effects of mountain-top isolation during Pleistocene interglacials and ice barriers to dispersal during glacials.
New Zealand's isolation, its well-studied rapidly changing landscape, and its many examples of rampant speciation make it an excellent location for studying the process of genetic differentiation. Using 1520 base pairs of mitochondrial DNA from the cytochrome oxidase subunit I, ATPase subunits 6 and 8 and tRNA(Asp) genes, we detected two well-differentiated, parapatrically distributed clades within the widespread New Zealand cicada species Maoricicada campbelli that may prove to represent two species. The situation that we uncovered is unusual in that an ancient lineage with low genetic diversity is surrounded on three sides by two recently diverged lineages. Using a relaxed molecular clock model coupled with Bayesian statistics, we dated the earliest divergence within M. campbelli at 2.3 +/- 0.55 million years. Our data suggest that geological and climatological events of the late Pliocene divided a once-widespread species into northern and southern components and that near the middle of the Pleistocene the northern lineage began moving south eventually reaching the southern clade. The southern clade seems to have moved northward to only a limited extent. We discovered five potential zones of secondary contact through mountain passes that will be examined in future work. We predict that, as in North American periodical cicadas, contact between these highly differentiated lineages will exist but will not involve gene flow.
The performance of 14 different recombination detection methods was evaluated by analyzing several empirical data sets where
the presence of recombination has been suggested or where recombination is assumed to be absent. In general, recombination
methods seem to be more powerful with increasing levels of divergence, but different methods showed distinct performance.
Substitution methods using summary statistics gave more accurate inferences than most phylogenetic methods. However, definitive
conclusions about the presence of recombination should not be derived on the basis of a single method. Performance patterns
observed from the analysis of real data sets coincided very well with previous computer simulation results. Previous recombination
inferences from some of the data sets analyzed here should be reconsidered. In particular, recombination in HIV-1 seems to
be much more widespread than previously thought. This finding might have serious implications on vaccine development and on
the reliability of previous inferences of HIV-1 evolutionary history and dynamics.
Parallel evolution of similar traits in independent populations that inhabit ecologically similar environments strongly implicates natural selection as the cause of evolution. Parallel speciation is a special form of parallel evolution where traits that determine reproductive isolation evolve repeatedly, in closely related populations, as by-products of adaptation to ecological conditions. The outcome of such parallel evolution is that ecologically divergent pairs of populations exhibit greater levels of reproductive isolation than ecologically similar pairs of populations of a similar or younger age. The parallel evolution of reproductive isolation provides strong evidence for natural selection in the process of speciation, but only one conclusive example from nature is known. Populations of the walking-stick insect Timema cristinae that use different host-plant species have diverged in body size and shape, host preference, behaviour and the relative frequency of two highly cryptic colour-pattern morphs. Here we report that divergent selection for host adaptation, and not genetic drift, has promoted the parallel evolution of sexual isolation in this species. Our findings represent a clear demonstration that host-plant adaptation can play a crucial and repeatable role in the early stages of speciation.
Determining the evolutionary age of asexual lineages should help in inferring the temporal scale under which asexuality and sex evolve and assessing selective factors involved in the evolution of asexuality. We used 416 bp of the mitochondrial COI gene to infer phylogenetic relationships of virtually all known Timema walkingstick species, including extensive intraspecific sampling for all five of the asexuals and their close sexual relatives. The asexuals T. douglasi and T. shepardii were very closely related to each other and evolutionarily young (less than 0.5 million years old). For the asexuals T. monikensis and T. tahoe, evidence for antiquity was weak since only one population of each was sampled, intraspecific divergences were low, and genetic distances to related sexuals were high: maximum-likelihood molecular-clock age estimates ranged from 0.26 to 2.39 million years in T. monikensis and from 0.29-1.06 million years in T. tahoe. By contrast, T. genevieve was inferred to be an ancient asexual, with an age of 0.81 to 1.42 million years. The main correlate of the age of asexual lineages was their geographic position, with younger asexuals being found further north.
— We studied sequence variation in 16S rDNA in 204 individuals from 37 populations of the land snail Candidula unifasciata (Poiret 1801) across the core species range in France, Switzerland, and Germany. Phylogeographic, nested clade, and coalescence analyses were used to elucidate the species evolutionary history. The study revealed the presence of two major evolutionary lineages that evolved in separate refuges in southeast France as result of previous fragmentation during the Pleistocene. Applying a recent extension of the nested clade analysis (Templeton 2001), we inferred that range expansions along river valleys in independent corridors to the north led eventually to a secondary contact zone of the major clades around the Geneva Basin. There is evidence supporting the idea that the formation of the secondary contact zone and the colonization of Germany might be postglacial events. The phylogeographic history inferred for C. unifasciata differs from general biogeographic patterns of postglacial colonization previously identified for other taxa, and it might represent a common model for species with restricted dispersal.
Evolution, weighting, and phylogenetic utility of mitochondrial gene sequences and a compilation of conserved polymerase chain reaction primers. C SIMON, F FRATI, A BECKENBACH, B CRESPI, H LIU, P
Species-oriented conservation programs attempt to analyze and maintain intra-specific variation in order to maximally preserve biological diversity. The “evolutionarily significant unit” has become an operational term for a group of organisms that should be the minimial unit for conservation management. No generally accepted definition for this term exists that would be the basis for the evaluation of these units in practical conservation situations. Currently, taxonomic decisions in species conservation are mostly based on the biological species concept. But the universal application of criteria of reproductive isolation or phenetic similarity to delimit conservation units is problematical. We favor a definition for evolutionarily significant units based on patterns of variation. In the theoretical framework of the phylogenetic species concept, conservation units are delimited by characters that diagnose clusters of individuals or populations to the exclusion of other such clusters. Characters are used for cladistic analysis to infer hypotheses of the phylogenetic relationships of individuals, and differentiated populations are diagnosed using population aggregation analysis. Characters can be based on genetic, morphological, ecological, or behavioral information, provided they are inferred to be heritable. The use of cladistics and population aggregation analysis has the potential to make the evaluation of evoluntionarily significant units objective and testable, an important consideration in politically controversial cases. Our cladistic approach is demonstrated by the evaluation of potential conservation units in the endangered tiger beetles Cicindela dorsalis and C. puritana.
Systematic biologists have directed much attention to species concepts because they realize that the origin of taxonomic diversity is the fundamental problem of evolutionary biology. Questions such as, What are the units of evolution? and, How do these units originate? thus continually capture the attention of many. It is probably no exaggeration to say that most believe the “systematic” aspects of the problem have been solved to a greater or lesser extent, whereas the task before us now is to understand the “genetic” and “ecologic” components of differentiation, i. e., those aspects often perceived to constitute the “real mechanisms” of speciation: A study of speciation is, to a considerable extent, a study of the genetics and evolution of reproductive isolating mechanisms (Bush, 1975, p. 339). ... a new mechanistic taxonomy of speciation is needed before population genetics, which deals with evolutionary mechanisms, can be properly integrated with speciation theory; that is, the various modes of speciation should be characterized according to the various forces and genetic mechanisms that underly [sic] the evolution of isolating barriers (Templeton 1980, p. 720).
This catalogue lists all genera and species of stick insects recorded or described from New Zealand. Genera are listed in alphabetical order within established subfamilies and tribes. Some taxonomic changes are made, with major changes as follows: 2 new genera in the subfamily Pachymorphinae are erected -Niveaphasma (type species -Pachymorpha annulata Hutton 1898) and Asteliaphasma (type species - Spinotectarchus jucundus Salmon 1991). Mimarchus tarsatus Carl 1913 is reduced to synonymy under Argosarchus horridus (White 1846), resulting in Mimarchus Carl 1913 becoming a synonym of Argosarchus Hutton 1898. Lectotypes are designated for Argosarchus schauinslandi Brunner 1907, Clitarchus interruptelineatus Brunner 1907, Clitarchus laeviusculus Stål 1875, Micrarchus parvulus Carl 1913, Micrarchus tarsatus Carl 1913 and Pachymorpha bouvieri Brunner 1907. Keys to adults and eggs of genera are given. The bibliography includes all references containing descriptions of species recorded or described from New Zealand.
The sexually reproducing stick insects Bacillus rossius and B. grandii are sharply differentiated in terms of allozyme gene alleles; B. atticus is a polyclonal automictic parthenogen sister to B. grandii grandii . Although well differen-tiated for coding genes, these hybridize to produce diploid (B. whitei = rossius/grandii) or triploid (B. lynceorum = rossius/grandii/atticus) clonal forms which reproduce apomictically. Allozyme analyses of unisexual Bacillus clearly establish their relationships from bisexual ancestor species as does the existence in all of them of several clones (especially in B. atticus) whose egg maturation allows regular recombination to occur. Bacillus taxa share the Bag320 satellite DNA family within different reproductive frameworks, allowing satellite variant homogenization to be uncoupled from fixation. The nested analysis of monomers reveals different patterns of sequence diversity: sexual reproduction includes both homogenization and variant fixation, whereas the slowing of molecular turnover pro-cesses and the absence of syngamy in the parthenogens realizes a similar range of sequence diversity at the level of the individual and supra-individual, but with no fixation. On the other hand, the actual values of sequence diversity appear mostly linked to species traits – range size, copy number of repeats, number of hybrid crosses – and possibly transposon activity, rather than to the reproductive mode. In addition, the mitochondrial genome reveals a compa-rable level of cox2 sequence variability in sexual and parthenogenetic taxa, thus adding to clonal variability. From Bacillus and other stick insect complexes, an overall picture of genomic diversification of parthenogens is therefore beginning to emerge. To define those animals that reproduce by non-canonical sexual modes (i.e. parthenogenesis, hybridogenesis), but make use of egg and meiotic mechanisms, the term meta-sexual is proposed.
The nucleotide sequence of the mitochondrial ND2, COI, COll, ATPase8, srRNA and nine tRNA genes have been sequenced from two individuals of the meadow grasshopper Chorthippus parallelus. Comparisons are made to other insects for which the same regions are completely sequenced. Percentage A + T is found to be relatively low in C. parallelus though consistent with that of the other Orthopteran, Locusta migratoria. The relative number of substitutions observed in the different protein-coding genes was analysed between pairs of insect species sharing different levels of relatedness. A clear change in this rate was observed between the within-genus and between-genera comparisons. This change is interpreted in terms of the functional constraints acting on these four different genes. The patterns seem to result from an early saturation of COl and COll genes with synonymous substitutions, and a tolerance of ND2 and ATPase8 function to high levels of amino acid replacements. This analysis highlights a need for further sequence studies and comparisons between taxa of different levels of divergence in order to understand the patterns of mtDNA evolution on which many evolutionary investigations are based.
— New Zealand's isolation, its well-studied rapidly changing landscape, and its many examples of rampant speciation make it an excellent location for studying the process of genetic differentiation. Using 1520 base pairs of mitochondrial DNA from the cytochrome oxidase subunit I, ATPase subunits 6 and 8 and tRNAAsp genes, we detected two well-differentiated, parapatrically distributed clades within the widespread New Zealand cicada species Maoricicada campbelli that may prove to represent two species. The situation that we uncovered is unusual in that an ancient lineage with low genetic diversity is surrounded on three sides by two recently diverged lineages. Using a relaxed molecular clock model coupled with Bayesian statistics, we dated the earliest divergence within M. campbelli at 2.3 ± 0.55 million years. Our data suggest that geological and climatological events of the late Pliocene divided a oncewidespread species into northern and southern components and that near the middle of the Pleistocene the northern lineage began moving south eventually reaching the southern clade. The southern clade seems to have moved northward to only a limited extent. We discovered five potential zones of secondary contact through mountain passes that will be examined in future work. We predict that, as in North American periodical cicadas, contact between these highly differentiated lineages will exist but will not involve gene flow.
w riting in the first issue of TREE, Ryder' brought the term 'Evolution- arily Significant Unit' (ESU) to the atten- tion of a broad audience of ecologists and evolutionary biologists. The ESU concept was developed to provide a rational basis for prioritizing taxa for conservation effort (e.g. captive breeding), given that resources are limited and that existing taxonomy may not adequately reflect underlying genetic diversity*. With the explicit recognition of the genetic com- ponent of biodiversity in conservation legislation of many countries and in the Convention on Biological Diversity, the ESU concept is set to become increas- ingly significant for conservation of natural as well as captive populations. However, the ESU remains poorly de- fined, both conceptually and operation- ally. Most definitions suggest than an ESU should be geographically discrete, but genetic criteria range from significant di- vergence of allele frequencies3 through some level of genetic distance' to congru- ently structured phylogenies among gene+. Several authors have argued that an ESU should display concordant divergence for both molecular and non- molecular traitG6. Although all are try- ing to achieve the same end, it seems that the operational definitions vary according to the biological and legislat- ive context. The purpose of this essay is to revisit the ESU concept in relation to recent developments in molecular popu- lation genetics. The suggested defi- nitions and criteria are not supposed to be proscriptive - rather, the intention is to promote debate on the purpose and practice of using genetic information to define conservation units. Conservation goals: what do we mean by 'significant'?
Restricted Item. Print thesis available in the University of Auckland Library or available through Inter-Library Loan. Although the Phasmidae are a fairly large family of the Orthoptera and are well known for their expert mimicry of plant forms, and colour change in response to their surroundings, comparitively little is known of their reproductive biology. Some species are known to be exclusively parthenogenic (for example BacillusjCarauaiua) and 'males' when they occur, are intersexes. Other normally bisexual members of this family that have been investigated are facultative parthenogens; this is probably the usual condition. A massive literature that is mainly taxonomic exists on the external genitalia, and the gross morphology of a number of species is known from scattered systematic and miscellaneous papers. Detailed structure and cytology of the reproductive organs are, however, few.
The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.
The nucleotide sequence of the mitochondrial ND2, COI, COII, ATPase8, srRNA and nine tRNA genes have been sequenced from two individual of the meadow grasshopper Chorthippus parallelus. Comparisons are made to other insects for which the same regions are completely sequenced. Percentage A + T is found to be relatively low in C. parallelus though consistent with that of the other Orthopteran, Locusta migratoria. The relative number of substitutions observed in the different protein-coding genes was analysed between pairs of insect species sharing different levels of relatedness. A clear change in this rate was observed between the within-genus and between-genera comparisons. This change is interpreted in terms of the functional constraints acting on these four different genes. The patterns seem to result from an early saturation of COI and COII genes with synonymous substitutions, and a tolerance of ND2 and ATPase8 function to high levels of amino acid replacements. This analysis highlights a need for further sequence studies and comparisons between taxa of different levels of divergence in order to understand the patterns of mtDNA evolution on which many evolutionary investigations are based.
Complete nucleotide sequences of the mitochondrial cytochrome b, tRNA(prolime), and tRNA(threonine) genes were described for 166 brown bears (Ursus arctos) from 10 geographic regions of Alaska to describe natural genetic variation, construct a molecular phylogeny, and evaluate classical taxonomies. DNA sequences of brown bears were compared to homologous sequences of the polar bear (maritimus) and of the sun bear (Helarctos malayanus), which was used as an outgroup. Parsimony and neighbor-joining methods each produced essentially identical phylogenetic trees that suggest two distinct clades of mtDNA for brown bears in Alaska: one composed only of bears that now reside on some of the islands of southeastern Alaska and the other which includes bears from all other regions of Alaska. The very close relationship of the polar bear to brown bears of the islands of southeastern Alaska as previously reported by us and the paraphyletic association of polar bears to brown bears reported by others have been reaffirmed with this much larger data set. A weak correlation is suggested between types of mtDNA and habitat preference by brown bears in Alaska. Our mtDNA data support some, but not all, of the currently designated subspecies of brown bears whose descriptions have been based essentially on morphology.
A combination of single-strand conformation polymorphism analysis (SSCP) and sequencing were used to survey cytochrome oxidase I (COI) mitochondrial DNA (mtDNA) diversity among New Zealand ovoviviparous Onychophora. Most of the sites and individuals had previously been analysed using allozyme electrophoresis. A total of 157 peripatus collected at 54 sites throughout New Zealand were screened yielding 62 different haplotypes. Comparison of 540-bp COI sequences from Peripatoides revealed mean among-clade genetic distances of up to 11. 4% using Kimura 2-parameter (K2P) analysis or 17.5% using general time-reversible (GTR + I + Gamma) analysis. Phylogenetic analysis revealed eight well-supported clades that were consistent with the allozyme analysis. Five of the six cryptic peripatus species distinguished by allozymes were confirmed by mtDNA analysis. The sixth taxon appeared to be paraphyletic, but genetic and geographical evidence suggested recent speciation. Two additional taxa were evident from the mtDNA data but neither occurred within the areas surveyed using allozymes. Among the peripatus surveyed with both mtDNA and allozymes, only one clear instance of recent introgression was evident, even though several taxa occurred in sympatry. This suggests well-developed mate recognition despite minimal morphological variation and low overall genetic diversity.
Sequencing of a cytochrome oxidase II (COII) gene fragment in Bacillus taxa provided evidence that the bisexual B. rossius is the maternal ancestor of the hybridogenetic B. rossius-grandii strains and revealed the same ancestry for both parthenogenetic hybrids: the diploid B. whitei (B. rossius/grandii grandii) and the triploid B. lynceorum (B. rossius/grandii grandii/atticus). Present data clearly demonstrate that all Bacillus unisexuals arose through asymmetrical hybridization events and realized a paraphyletic derivation from the B. rossius redtenbacheri subspecies. The invention of B. rossius mitochondrial DNA haplotypes in specimens with B. grandii grandii nuclear genomes revealed the occurrence of androgenesis in nature. Natural androgens represent a peculiar escape from hybridity and can help maintain the hybridogenetic system through the production of the fathering taxon via hybrid females. Results from the COII gene support the phyletic relationships among taxa suggested by previous taxonomical approaches, but also indicate a departure of B. grandii subspecies from the established taxonomy. Assuming the existence of a molecular clock, the evaluated substitution rate brings the splitting between B. rossius and B. grandii/B. atticus back to 22.79 +/- 2.65 myr before present, while the origin of hybrids appears to be much more recent (1.06 +/- 0.53 myr).
There are eight distinct chromosomal races of the New Zealand weta Hemideina thoracica. We used mtDNA sequence data to test the hypothesis that these races originated on islands during the early Pliocene (7--4 million years ago). Nine major mitochondrial lineages were identified from 65 cytochrome oxidase I sequences. Phylogenetic analysis of these lineages suggests that they arose at approximately the same time. The geographical distribution of some lineages coincides with areas that were islands during the Pliocene. Overall, hierarchical AMOVA analysis shows that chromosomal races and Pliocene islands describe only 28% and 24%, respectively, of the total current mtDNA variation. However, removing one widespread (A) and one putatively introgressed (F) lineage increases these estimates to 65% and 80%, respectively. Intraspecific sequence divergence was very high, reaching a maximum of 9.5% (uncorrected distance) and GC content was high compared to other insect mtDNA sequences. Average corrected distance among mtDNA lineages supports the Pliocene origins of this level of genetic diversity. In the southern part of the species range there is reduced mtDNA variation, probably related to local extinction of H. thoracica populations from recent volcanic activity and subsequent re-colonization from a leading edge. In contrast, in this southern part there are five chromosome races, suggesting that chromosome races here may be younger than those in the north.
The existence of areas of lower endemism and disjunction of New Zealand biota is typified by Nothofagus beech trees (hence "beech-gap") and have been attributed to a variety of causes ranging from ancient fault-mediated displacement (20-25 million years ago) to Pleistocene glacial extirpation (< 1.8 million years ago). We used cytochrome oxidase I and 12S mtDNA sequence data from a suite of endemic invertebrates to explore phylogeographic depth and patterns in South Island, New Zealand, where the "beech-gap" occurs. Phylogeographic structure and genetic distance data are not consistent with ancient vicariant processes as a source of observed pattern. However, we also find that phylogeographic patterns are not entirely congruent and appear to reflect disparate responses to fragmentation, which we term "gap," "colonization," and "regional." Radiations among congenerics, and in at least one instance within a species, probably took place in the Pliocene (2-7 million years ago), possibly under the influence of the onset of mountain building. This orogenic phase may have had a considerable impact on the development of the biota generally. Some of the taxa that we studied do not appear to have suffered range reduction during Pleistocene glaciation, consistent with their survival throughout that epoch in alpine habitats to which they are adapted. Other taxa have colonized the beech-gap recently (i.e., after glaciation), whereas few among our sample retain evidence of extirpation in the most heavily glaciated zone.
Phylogenetic studies of asexual lineages and their sexual progenitors are useful for inferring the causes of geographical parthenogenesis and testing hypotheses regarding the evolution of sex. With five known parthenogens and well-studied ecology, Timema walking-sticks are a useful system for studying these questions. Timema are mainly endemic to California and they exhibit the common pattern of geographical parthenogenesis, with asexuals exhibiting more-northerly distributions. Neighbour-joining and maximum-parsimony analyses of 416 bp of mitochondrial cytochrome oxidase I (COI) from 168 individuals were used to infer general phylogenetic relationships, resulting in three major phylogeographical subdivisions: a Northern clade; a Santa Barbara clade; and a Southern clade. A nested cladistic analysis, comparing intra- and interspecific haplotypic variation on a geographical scale, revealed that the overall pattern of geographical parthenogenesis in Timema could be attributed to historical range expansion. These results suggest that geographical parthenogenesis is the result of more-extensive northerly dispersal of asexuals than sexuals.
Determining the evolutionary age of asexual lineages should help in inferring the temporal scale under which asexuality and sex evolve and assessing selective factors involved in the evolution of asexuality. We used 416 bp of the mitochondrial COI gene to infer phylogenetic relationships of virtually all known Timema walkingstick species, including extensive intraspecific sampling for all five of the asexuals and their close sexual relatives. The asexuals T. douglasi and T. shepardii were very closely related to each other and evolutionarily young (less than 0.5 million years old). For the asexuals T. monikensis and T. tahoe, evidence for antiquity was weak since only one population of each was sampled, intraspecific divergences were low, and genetic distances to related sexuals were high: maximum-likelihood molecular-clock age estimates ranged from 0.26 to 2.39 million years in T. monikensis and from 0.29-1.06 million years in T. tahoe. By contrast, T. genevieve was inferred to be an ancient asexual, with an age of 0.81 to 1.42 million years. The main correlate of the age of asexual lineages was their geographic position, with younger asexuals being found further north.
Spectronet is a package that uses various methods for exploring and visualising complex evolutionary signals. Given an alignment in NEXUS format, the package works by computing a collection of weighted splits or bipartitions of the taxa and then allows the user to interactively analyse the resulting collection using tools such as Lento-plots and median networks. The package is highly interactive and available for PCs.
The origin of the obligate-parthenogenetic New Zealand stick insect genus Acanthoxyla was investigated using cytogenetics and sequencing of nuclear and mitochondrial DNA. Little mitochondrial DNA sequence variation (COI-II) was found among seven species of the genus Acanthoxyla and we found no evidence for monophyly of the morphologically distinguished lineages. In contrast, two distinct clades of nuclear sequence (ITS) were obtained, one is restricted to the genus Acanthoxyla, while the other includes sequences obtained from its sister genus Clitarchus. Although Acanthoxyla appears to be diploid (2n = 36-38), it has two ill-matched chromosome pairs. We hypothesize that two or more hybridization events involving the parental sexual species Clitarchus hookeri and an unknown taxon probably resulted in the formation of the parthenogenetic genus Acanthoxyla. However, the karyotype of Acanthoxyla bears little resemblance to the karyotype of the putative paternal species C. hookeri so the exact nature of Acanthoxyla remains in question.
An in-depth analysis of the Leptynia attenuata species complex has been performed by cytochrome oxidase subunit 2 (cox2) gene sequencing as well as karyotype and allozyme analysis. The whole set of data allows to largely resolve the taxonomy of the group and suggests an overall trend of chromosomal repatternings through a progressive reduction of the chromosome number. A previously suggested new species has been also confirmed on a genetic basis. Data are discussed in order to depict a phylogenetic and phylogeographic scenario fitting the observed genetic relationships between the different species of the group. Chromosome rearrangements are proposed as the major speciation driving force within the group and androgenetic reproduction is suggested as a shortcut to overcome the problem of fixing chromosomal rearrangements that are strongly underdominant in heterozygotes.
The recognition concept of species Species and Speciation (pp. 21–29) Pretoria: Transvaal Museum Monograph No Evaluation of methods for detecting recombina-tion from DNA sequences: Empirical data
- H E H Paterson
Paterson, H. E. H. (1985). The recognition concept of species. In E. S. Vrba (Ed.) Species and Speciation (pp. 21–29). Pretoria: Transvaal Museum Monograph No. 4. Posada, D. (2002). Evaluation of methods for detecting recombina-tion from DNA sequences: Empirical data. Molecular Biology and Evolution, 19, 708 –717.
The Zoology of the Voyage of H. M. S. Erebus and Terror, 1. Insects of New Zealand
- A White
White, A. (1846). The Zoology of the Voyage of H. M. S. Erebus and Terror, 1. Insects of New Zealand. London: E. W. Janson.
Revision of the New Zealand Phasmidae. Transactions of the New Zealand Institute
- F W Hutton
Hutton, F. W. (1899). Revision of the New Zealand Phasmidae.
Transactions of the New Zealand Institute, 30, 50 -59.
New Zealand Vegetation History (pp. 557– 602) Dordrecht: Kluwer Academic Press Hybrid origin of a parthenogenetic genus
- M S Mcglone
McGlone, M. S. (1988). New Zealand. In B. Huntley & T. Webb (Eds) Vegetation History (pp. 557– 602). Dordrecht: Kluwer Academic Press. Morgan-Richards, M. & Trewick, S. A. (2005). Hybrid origin of a parthenogenetic genus? Molecular Ecology, 14, 2133 –2142.
The eggs of three Acanthoxyla species
- B Mantovani
- V Scali
Mantovani, B. & Scali, V. (1987). The eggs of three Acanthoxyla
species. In M. Mazzini & V. Scali (Eds) Stick Insects: Phylogeny and
Reproduction -Proceedings of the First International Symposium on
Stick Insects. Italy: University of Siena.
Se-Al: Sequence Alignment Editor
- A Rambaut
Rambaut, A. (1996). Se-Al: Sequence Alignment Editor. Available via
http://evolve.zoo.ox.ac.uk /.