The systematic position of some nippostrongyline nematodes (Trichostrongylina: Heligmosomoidea) parasitic in Argentinean sigmodontine rodents

The systematic position of two nippostrongyline nematodes described from Argentinean sigmodontine rodents is clarified. The first species, Hassalstrongylus multiovatus Suriano & Navone, 1992, parasitic in Akodon simulator Thomas from the province of Tucumán, was studied on the basis of type and voucher material. H. multiovatus is proposed as a junior synonym of Trichofreitasia lenti Sutton & Durette-Desset, 1991, a parasite described from Oligoryzomys flavescens (Waterhouse) in the province of Buenos Aires. The holotype and three of seven paratypes deposited as H. multiovatus were identified as T. lenti. One male paratype was identified as Guerrerostrongylus uruguayensis Sutton & Durette-Desset, 1991, a parasite described from O. flavescens in Uruguay. Three female paratypes were identified as Guerrerostrongylus sp. The second species, Stilestrongylus scapteromys Suriano & Navone, 1996, parasitic in Scapteromys aquaticus Thomas from the province of Buenos Aires, was studied on voucher material. Stilestrongylus scapteromys and Malvinema frederici Digiani, Sutton & Durette-Desset, 2003, the type-species of Malvinema Digiani, Sutton & Durette-Desset, 2003, were described from the same host and geographical region. As they are considered to refer to one and the same taxon, the new combination Malvinema scapteromys n. comb. is proposed for this species.     

Two new species of Nippostrongylinae (Nematoda: Trichostrongylina: Heligmonellidae) from the grey leaf-eared mouse Graomys griseoflavus (Sigmodontinae) in Argentina

Two new species of Nippostrongylinae, Hassalstrongylus puntanus n. sp., and Stilestrongylus franciscanus n. sp. are described from the intestine of the grey leaf-eared mouse Graomys griseoflavus (Waterhouse, 1837) (Sigmodontinae) from the Province of San Luis, Argentina. Hassalstrongylus puntanus n. sp. is distinguished from the most closely related species H. dollfusi (Díaz-Ungría, 1963), a parasite of Mus musculus from Venezuela by longer rays 2, shorter rays 4, proximal half of the dorsal ray non-enlarged and a non-retractile female tail. Stilestrongylus franciscanus n. sp. is distinguished from the most closely related species S. flavescens Sutton & Durette-Desset, 1991, a parasite of Oryzomys flavescens from Argentina, by rays 2 and 3 diverging separately from common trunk of rays 2 to 6, and by right ray 3 arising from this trunk more distally than ray 6. Stilestrongylus azarai Durette-Desset & Sutton, 1985, a parasite of Akodon azarae from Buenos Aires, Argentina, was also found parasitizing G. griseoflavus, representing new host and locality records. Some additional morphological data for this species are also provided.     

A new Heligmonellid (Nematoda: Heligmonellidae) from Oecomys mamorae (Rodentia: Sigmodontinae) in the pantanal and new data on the synlophe of Guerrerostrongylus zetta from the Atlantic Forest, Brazil

A new species of Heligmonellidae nematode, Guerrerostrongylus gomesae n. sp., is described from specimens collected from the small intestine of the rodent Oecomys mamorae Thomas, 1906 in the Brazilian Pantanal. It differs from the 2 other species of the genus by the number of ridges in the synlophe 35-46 and 40-48 at the midbody in the male and female, respectively, by rays 8 arising at midlength of the dorsal trunk, ending near the margin of the caudal bursa, and the dorsal ray divided at the second-third part into 2 branches, each branch divided into 2 subequal subbranches. In addition, specimens of Guerrerostrongylus zetta Travassos, 1937 collected in Oligoryzomys nigripes Olfers, 1818 contain synlophe with 36-42 cuticular ridges in males and 38-42 in females at the midbody. By the characteristics of the synlophe and caudal bursa, Gerrerostrongylus gomesae n. sp. is considered a new species. We suggest that the number of cuticular ridges for this genus broadens the range of the cuticular ridges to at least 35 at midbody.     

Trichostrongylina (Nematoda) parasitic in Phyllotis sp. (Rodentia: Sigmodontinae) from Argentina, with description of three new species

The Trichostrongylina parasitic in the leaf-eared mouse Phyllotis sp. (Sigmodontinae) from the Province of Catamarca, Argentina are studied. Three new species of Stilestrongylus Freitas, Lent and Almeida, 1937 (Heligmosomoidea: Heligmonellidae: Nippostrongylinae) are described. Stilestrongylus andalgala n. sp. is distinguished from the most closely related species S. moreli Diaw, 1976 parasitic in Auliscomys boliviensis from Bolivia, by the absence of a common trunk of right rays 2 and 3, by left ray 4 shorter than ray 5, and by the shape of the female tail, without a mucron. Stilestrongylus catamarca n. sp. is distinguished from all the other species in the genus by hypertrophied rays 2, larger than rays 3, and from the closely related species Stilestrongylus barusi Durette-Desset, 1971, parasitic in Sigmodontomys alfari from Colombia, by right ray 4 longer than ray 5 and left ray 4 shorter than ray 5, by a larger spicule length/body length ratio (22.6% vs. 7.2%), and by the presence of a mucron on the female tail. Stilestrongylus gracielae n. sp. most closely resembles Stilestrongylus azarai Durette-Desset and Sutton, 1985, parasitic in Akodon azarae and in Graomys griseoflavus, and Stilestrongylus franciscanus Digiani and Durette-Desset, 2003, also parasitic in G. griseoflavus, both from Argentina. These latter differ from the new species by the right lobe of the caudal bursa: in S. azarai rays 4, 5 and 6 arise at same level from their common trunk and in S. franciscanus right rays 4 and 5 diverge at their distal extremity. Lamanema chavezi Becklund, 1963 (Molineoidea: Molineinae), a parasite of South American camelids, is reported in rodents for the second time.     

Morphology of some tricostrongilinae (Strongylida) from the National Helminth Collection, Institute of Biology, UNAM, Mexico

The present study analyses the taxonomic status of eleven species of trichostrongylins that parasitize rodents and lagomorphs deposited in the Colección Nacional de Helmintos, Instituto de Biología. UNAM. Mexico. This analysis is based on the morphology of the synlophe, characteristic that had not been studied for most of these nematode species and at present, it has a very important taxonomic value. As a result of this study, the identity of five species is ratified (Trichostrongylus calcaratus, Obeliscoides cuniculi, Carolinensis huehuetlana. Stilestrongylus peromysci and Nippostrongylus brasiliensis), the transference suggested previously for two more (Vexillata convoluta and Vexillata vexillata) is confirmed, Trichostrongylus chiapensis is synonymized with Boehmiella willsoni, and finally Lamothiella romerolagi is re-determined as Teporingonema cerropeladoensis and Stilestrongylus atlatilpinensis as Stilestrongylus hidalguensis.     

A new genus of Nippostrongylinae (Nematoda: Heligmonellidae) from the water rat Scapteromys aquaticus (Sigmodontinae) in Argentina

A new genus of Nippostrongylinae, Malvinema n. gen., with 3 coparasitic species M. frederici n. sp., M. carolinae n. sp., and M. victoriae n. sp. from the intestine of the water rat, Scapteromys aquaticus Thomas (Rodentia: Muridae), from the northeast of Buenos Aires Province, Argentina, is proposed in this study. The new genus shows similarities to 2 Neotropical Nippostrongylinae: Carolinensis (Travassos, 1937) by some characters of the synlophe and Stilestrongylus Freitas, Lent and Almeida, 1937, by the pattern of the caudal bursa. It is characterized by a synlophe with triple or quadruple gradient of size of the ridges, lateromedian, decreasing from the largest left and right ridges. The gradient situated in the right ventral quadrant is always present. The caudal bursa shows a pattern of type 1-4. Malvinema frederici possesses a synlophe with 17 ridges and an axis of orientation inclined at 45 degrees from the sagittal axis; M. carolinae possesses a synlophe with 22-24 ridges and an axis of orientation almost merged with the sagittal axis. Both species have a caudal bursa with the right lobe enlarged transversally. Malvinema victoriae possesses a synlophe with 22-24 ridges, an axis of orientation inclined at 45 degrees from the sagittal axis, and a caudal bursa with the right lobe enlarged vertically.     

Systematic position of some nearctic Heligmosomoidea (Nematoda: Trichostrongylina) from the U.S. National Parasite collection and their description

The systematic position of some heligmosomoid nematodes from rodents, deposited in the U.S. National Parasite Collection (USNPC), is revised, mainly through the study of their synlophe, which in all cases was unknown or insufficiently described. The material was registered as different species of Longistriata Schulz, 1926, a genus whose representatives are only parasitic in Holarctic insectivores. Longistriata norvegica Dikmans, 1935, parasitic in Rattus sp. becomes a synonym of Hassalstrongylus aduncus (Chandler, 1932). Specimens registered as Longistriata dalrymplei Dikmans, 1935, from Ondatra zibethicus, are confirmed to belong to Carolinensis (Travassos, 1937). Specimens registered as Longistriata noviberiae Dikmans, 1935, parasitic in Sylvilagus floridanus alacer, were found to belong to Vexillata, and Vexillata noviberiae n. comb. is here proposed. This is the first record of a species of Vexillata in a lagomorph. Other specimens registered as Longistriata norvegica, parasitic in Geomys floridanus austrinus, were also found to be an undescribed species of Vexillata (Hall, 1916), which is named Vexillata chitwoodi n. sp. This is similar to Vexillata chabaudi Yoyotte-Vado, 1972, Vexillata petteri Durette-Desset, 1970, Vexillata scorzai Guerrero, 1984 and Vexillata tejerai Guerrero, 1984, all having the same number of cuticular ridges (4 dorsal, 5 ventral) and the division of the dorsal ray at its apex. The most related species is V. chabaudi, which is differentiated from the new species by rays 4 not curved distally, by thick rays 8, and by a dorsal ray enlarged at the level of the arising of rays 8.     

2 new species of Stilestrongylus (Nematoda: Heligmonellidae) parasites of Peromyscus (Rodentia: Cricetidae) from Mexico

Stilestrongylus peromysci n. sp. collected from Peromyscus difficilis (Hidalgo state, México), differs from other species in the genus in number of the spines (30) in the synlophe (both sexes) and because the eighth ray arises from the root of the ninth ray; S. hidalguensis n. sp. parasitised Peromyscus sp. and differs from all other congeneric species in the presence of 24 spines in the male synlophe and in the arrangement of the bursal rays (2-2-1 in the right lobe and 2-3 in the left lobe). A key to the species of Stilestrongylus is provided.     

Multilocus molecular systematics and evolution in time and space of Calligrapha (Coleoptera: Chrysomelidae,Chrysomelinae)

Calligrapha (Coleoptera: Chrysomelidae) is a genus with species present in most of the American continent, from the Arctic polar circle to the Pampas in Argentina. In its current concept, the genus comprises some 80 species, but the diagnosis of the genus is problematic, based on a combination of potentially symplesiomorphic character states. In this study, we investigate the largest taxonomic sample of Calligrapha diversity to date (43 species) using a phylogenetic perspective based on more than 6000 molecular characters from eight genes (four mitochondrial and four nuclear) for a systematic evaluation of the genus. The analyses also include thirteen species in the closely related Zygospila (currently a subgenus of Zygogramma) to assist the systematic delimitation of Calligrapha. Partitioned and total evidence phylogenetic trees were additionally used for molecular clock analyses and dating based on standard mtDNA evolutionary rates, and for likelihood‐based inference of ancestral areas. Calligrapha and Zygospila are reciprocally paraphyletic, and our interpretation of taxonomic stability merges both taxa into a larger genus Calligrapha which plausibly originated in the dry steppes of southern North America in the Late Miocene. The genus includes a minimum of five strongly supported lineages which initially diversified in the Pliocene, fully congruent with expectations from morphology, but of uncertain mutual relationships. Only two of these lineages dispersed to South America: the group of C. polyspila right at the time of the final closure of the Isthmus of Panama in the Early Pliocene and the group of C. argus only in recent times, well in the Pleistocene. The most species‐rich lineage of Calligrapha, associated to trees and shrubs typical of riverine and lacustrine environments (as opposed to herbaceous steppe plants, generally Malvaceae and Asteraceae, for most other groups) diversified and spread in North America in the Late Pliocene. The ecological shift to a stable habitat spreading in the continent due to climate change is hypothesized as one possible explanation for the evolutionary success of this group.     

Morphological variability of potato cyst nematodes (Globodera spp.) in the Canary Islands

Thirty populations of potato cyst nematode (Globodera spp.) from the Island of Tenerife and two populations from the UK were assessed for several morphometric and non-morphometric characters thought to discriminate between the species G. rostochiensis and G. pallida. Also 200 cysts from each population were analysed by isoelectric focusing of soluble proteins. Correlation analysis, analysis of variance and principal component analysis were used to investigate relationships between the morphometric characters, how the relationships varied between species and between populations, and which characters were most useful for discriminating between species. The two species differed significantly for each of the four morphometric characters: stylet length, fenestra length, anus-fenestra distance and the number of ridges. The stylet length and fenestra length also showed differences between populations of G. rostochiensis while stylet length and number of ridges showed differences between populations of C. pallida. In general, populations of G. pallida showed greater variation than populations of C. rostochiensis. Principal component analysis of the population means indicated that over 73% of the variation in the characters could be explained by the contrast of stylet and fenestra lengths against the anus-fenestra distance and number of ridges. A plot of the first two principal components separated the two species. Stepwise discriminant analysis provided a linear combination of these four variables which discriminated between the species. Stylet length was found to be the most useful characteristic for distinguishing the species whilst anus-fenestra distance was the least useful.     

Phylogenomics,biogeography, and evolution of morphology and ecological niche of the eastern Asian–eastern North American Nyssa (Nyssaceae)

Nyssa (Nyssaceae, Cornales) represents a classical example of the well‐known eastern Asian–eastern North American floristic disjunction. The genus consists of three species in eastern Asia, four species in eastern North America, and one species in Central America. Species of the genus are ecologically important trees in eastern North American and eastern Asian forests. The distribution of living species and a rich fossil record of the genus make it an excellent model for understanding the origin and evolution of the eastern Asian–eastern North American floristic disjunction. However, despite the small number of species, relationships within the genus have remained unclear and have not been elucidated using a molecular approach. Here, we integrate data from 48 nuclear genes, fossils, morphology, and ecological niche to resolve species relationships, elucidate its biogeographical history, and investigate the evolution of morphology and ecological niches, aiming at a better understanding of the well‐known EA–ENA floristic disjunction. Results showed that the Central American (CAM) Nyssa talamancana was sister to the remaining species, which were divided among three, rapidly diversified subclades. Estimated divergence times and biogeographical history suggested that Nyssa had an ancestral range in Eurasia and western North America in the late Paleocene. The rapid diversification occurred in the early Eocene, followed by multiple dispersals between and within the Erasian and North American continents. The genus experienced two major episodes of extinction in the early Oligocene and end of Neogene, respectively. The Central American N. talamancana represents a relic lineage of the boreotropical flora in the Paleocene/Eocene boundary that once diversified in western North America. The results supported the importance of both the North Atlantic land bridge and the Bering land bridge (BLB) for the Paleogene dispersals of Nyssa and the Neogene dispersals, respectively, as well as the role of Central America as refugia of the Paleogene flora. The total‐evidence‐based dated phylogeny suggested that the pattern of macroevolution of Nyssa coincided with paleoclimatic changes. We found a number of evolutionary changes in morphology (including wood anatomy and leaf traits) and ecological niches (precipitation and temperature) between the EA–ENA disjunct, supporting the ecological selection driving trait evolutions after geographic isolation. We also demonstrated challenges in phylogenomic studies of lineages with rapid diversification histories. The concatenation of gene data can lead to inference of strongly supported relationships incongruent with the species tree. However, conflicts in gene genealogies did not seem to impose a strong effect on divergence time dating in our case. Furthermore, we demonstrated that rapid diversification events may not be recovered in the divergence time dating analysis using BEAST if critical fossil constraints of the relevant nodes are not available. Our study provides an example of complex bidirectional exchanges of plants between Eurasia and North America in the Paleogene, but “out of Asia” migrations in the Neogene, to explain the present disjunct distribution of Nyssa in EA and ENA.     

The systematic significance of seed morphology in Sagina (Caryophyllaceae) under scanning electron microscopy

The seed morphology of 15 species ofSagina from North America, Europe, and eastern Asia was studied, utilizing the scanning electron microscope, to determine the significance of seed coat features as taxonomic characters. There is variation in particular surface features within some members of the genus. However, superficial ridges and tubercules exhibit patterns very helpful in determining relationships among some species. Two basic seed types occur in the genus and are diagnostic at the sectional level. The saginoid seed, characteristic of sectionSagina, is obliquely triangular in outline, is grooved along the two dorsal ridges, and has slightly concave lateral surfaces. The crassuloid seed, characteristic of sectionMaxima, is obliquely reniform in outline, lacks dorsal grooves, and has shallowly convex lateral surfaces.     

Karyotype and mitochondrial 16S gene characterizations in seven South American Cichlasomatini species (Perciformes, Cichlidae)

The family Cichlidae constitutes most of the freshwater fish fauna of South America; its taxonomy is at present mainly based on morphological characters. Here, relationships among seven Cichlasomatini species have been investigated by studying their karyotype structure and by sequencing a 520 bp fragment of the mitochondrial 16S gene. Molecular data sets point to a high affinity of Cichlasoma amazonarum with Aequidens sensu stricto group, in particular with Aequidens tetramerus . Aequidens never form a single monophyletic clade: molecular trees group together ' Aequidens pulcher ' and ' Aequidens rivulatus ' and suggest their close relationship with Bujurquina and Laetacara , rather than with the A. sensu stricto group. Both molecular and karyotypic data confirm that Cleithracara maronii belongs to a distinct clade, thus supporting its generic differentiation based on morphological characters. Chromosome number, karyotype structure and molecular data suggest that Laetacara dorsigera is related to Bujurquina vittata and confirm their generic level of differentiation. From a cytotaxonomic point of view, a karyotype of 2 n  = 48 with most acrocentric or subacrocentric chromosomes could be the ancestral one from which the others might have derived.     

The geography of diversification in the mormoopids (Chiroptera: Mormoopidae)

The traditional explanation of the distribution of the Mormoopidae is that this family originated in southern Central America or northern South America, later expanding its range north to Mexico and the West Indies, and differentiating into eight species. An alternative fossil-based hypothesis argues that the family originated in the northern Neotropics, reached the Caribbean early in its history, and dispersed to South America after the completion of the Isthmus of Panama. The present study analyses new and previously published sequence data from the mitochondrial 12S, tRNA^val, 16S, and cytochrome b , and the nuclear Rag 2, to evaluate species boundaries and infer relationships among extant taxa. Fixed differences in cytochrome b often coincide with published morphological characters and show that the family contains at least 13 species. Two additional, morphologically indistinct, lineages are restricted to Suriname and French Guiana. Phylogeny-based inferences of ancestral area are equivocal on the geographical origin of mormoopids, in part because several internal nodes are not resolved with the available data. Divergences between Middle American and Antillean populations are greater than those between Mexico/Central America and South America. This suggests that mormoopids diversified in northern Neotropics before entering South America. A northern neotropical origin for mormoopids is congruent with both the Tertiary fossil record and recent phylogenetic hypotheses for the sister family to the Mormoopidae, the Phyllostomidae.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 88 , 101–118.     

A cladistic analysis of Asarum (Aristolochiaceae) and implications for the evolution of herkogamy

A cladistic analysis of Asarum was conducted to examine relationships among species within the genus and to test the monophyly of several groups of taxa that have often been treated as segregate genera. Thirty-two species were drawn from throughout the range of the genus, representing a broad sample of sections and all segregate genera. The data matrix included 37 characters derived from various aspects of vegetative and floral morphology. A strict consensus of all most parsimonious trees suggests that Asarum s.l. is monophyletic and consists of two main clades: an Asarum clade, which is characterized by connate styles and inferior ovaries, and an Asiasarum-Hexastylis-Heterotropa clade, which is characterized by ridges on the inner perianth surface, dorsal stigmas, and bifid style extensions. The latter is a large and morphologically diverse clade that includes the North American segregate Hexastylis and two Asiatic segregates. Examination of pollination mechanisms in the context of this phylogeny supports the conclusion that herkogamy, and thus obligate insect pollination, is derived from a plesiomorphic condition of autonomous self-pollination. Associated with herkogamy are characters such as glandular trichomes and other ornamentation of the surface of the calyx that probably represent increased specialization to attract insect pollinators. This study also indicates that chromosomal evolution has occurred via aneuploid decrease from an ancestral chromosome number of 2n = 26 to 2n = 24 in Heterotropa. The recognition of two subgenera, subgenus Asarum and subgenus Heterotropa, corresponding to the two clades in the cladistic analysis, is recommended.     

Morphological phylogenetics of Puya subgenus Puya (Bromeliaceae)

Puya , a large genus mostly from South America, has been taxonomically divided into two subgenera: Puyopsis and Puya. The latter includes only eight species distributed mainly in Chile, extending to Argentina, Bolivia, and Peru. The species of subgenus Puya are recognized by the presence of a sterile apex of the inflorescence branches, whereas those of subgenus Puyopsis have fertile flowers all along the branches. The objectives of this article were to determine whether this diagnostic character was synapomorphic for subgenus Puya , and to explore the relationships between its species. Parsimony analyses were performed for 43 taxa and 93 morphological characters, 87 of which were discrete and six continuous. In the analysis that included all characters, a single most parsimonious tree was found that supported subgenus Puya by two synapomorphic character states, including the diagnostic character of a sterile inflorescence branch apex and a blooming pattern in which flowers open gradually from base to apex. The trees were better supported when the continuous characters were included. Further studies are suggested to resolve the infrageneric classification of Puya and the relationships of the species belonging to subgenus Puya .  © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society , 2008, 156 , 93–110.     

基于部分18S rDNA, 28S rDNA和COI基因序列的索科线虫亲缘关系

通过PCR扩增获得我国常见昆虫病原索科线虫6属10种18S rDNA、28S rDNA(D3区)和COI基因序列,结合来自GenBank中6属10种索科线虫的18S rDNA同源序列,用邻接法和最大简约法构建系统进化树。结果显示:12属索科线虫分为三大类群,第一大类群是三种罗索属线虫(Romanomermis)先聚在一起,再与两索属(Amphimermis)和蛛索属(Aranimermis)线虫聚为一支;在第二大类群中,六索属(Hexamermis)、卵索属线虫(Ovomermis)和多索属(Agamermis)亲缘关系最近,先聚在一起,再与八腱索属(Octomyomermis)和Thaumamermis线虫聚为一支。第三大类群由索属(Mermis)和异索属(Allomermis)线虫以显著水平的置信度先聚在一起,再与蠓索属(Heleidomermis)和施特克尔霍夫索属(Strelkovimermis)线虫聚为一支。从遗传距离看,基于3个基因的数据集均显示索科线虫属内种间差异明显小于属间差异,武昌罗索线虫(R.wuchangensis)和食蚊罗索线虫(R.culicivorax)同属蚊幼寄生罗索属线虫,其种间的遗传距离最小。     

Phylogenetic relationships of North American garter snakes (Thamnophis) based on four mitochondrial genes: how much DNA sequence is enough?

The clade of garter snakes (Thamnophis) includes some of the most abundant and well-studied snakes in North America. However, phylogenetic relationships within this group have been little studied. We used DNA sequences of four mitochondrial genes (cytochrome b and NADH dehydrogenase subunits 1, 2, and 4) to estimate relationships among 29 of the 31 recognized species of Thamnophis plus the related species Adelophis foxi. Both maximum parsimony (MP) and maximum-likelihood (ML) analyses of all these genes combined produced well-resolved trees with moderate (70-89%) to strong (90-100%) bootstrap support for most clades. MP and ML trees were very similar, with no strongly supported conflict between the two analyses. These analyses identify a clade of 12 species largely restricted to México (the "Mexican clade"), and a clade containing 15 species that collectively range from Central America to southern Canada (the "widespread clade"). These two groups are identified as sister taxa in both MP and ML analyses. A clade consisting of the ribbon snakes (T. sauritus and T. proximus) and the common garter snake (T. sirtalis) is placed as the sister group to all other Thamnophis (i.e., the Mexican + widespread clades) in our analyses. High bootstrap proportions at several levels in the tree support the inclusion of both Thamnophis validus, which has traditionally been placed in the genus Nerodia, and the poorly known species Adelophis foxi within Thamnophis. We used randomly sampled characters (i.e., standard bootstrapping) and randomly sampled contiguous blocks of characters to examine the effect of number of characters on resolution of and support for relationships within Thamnophis using MP. In general, these analyses indicate that we have reached a point of strongly diminishing returns with respect to the effect of adding mtDNA sequence characters for the current set of taxa; our sample of 3809 mtDNA characters is apparently "enough." The next steps to improve the phylogenetic estimate may be to add nuclear DNA sequences, morphology, or behavior, or to sequence additional mtDNA lineages within species.