Pulmonate phylogeny based on 28S rRNA gene sequences: a framework for discussing habitat transitions and character transformation.

Pulmonate snails occupy a wide range of marine, estuarine, freshwater and terrestrial environments. Non-terrestrial forms are supposed to be basal in pulmonate evolution but the group's phylogeny is not well resolved either morphologically or on the basis of available DNA sequence data. The lack of a robust phylogeny makes it difficult to understand character polarization and habitat transformation in pulmonates. We have investigated pulmonate relationships using 27 new sequences of 28S rRNA from pulmonates and outgroups, augmented with data from GenBank. The complete alignments comprised about 3.8kb. Maximum parsimony, maximum likelihood and Bayesian analyses of alignments generated under different assumptions are reported. Complete alignments appear to have a degree of substitution saturation so where there is conflict between hypothesised relationships more weight is given to analyses where regions of random similarity are excluded and which are not affected by this complication. Monophyly of the five main pulmonate groups was robustly supported in almost all analyses. The marine group Amphiboloidea and the freshwater Glacidorbidae are the most basal. The remaining pulmonates (Siphonariidae, Hygrophila and Eupulmonata) form a moderately-supported monophyletic group in all analyses bar one probably affected by saturation of substitutions. Siphonariidae, a predominantly marine and intertidal family, and Eupulmonata (mainly terrestrial with marine, estuarine and freshwater species) form a strongly supported clade that is the sister group to Hygrophila (freshwater). Multiple colonizations of freshwater and terrestrial habitats by pulmonate snails are suggested. No analyses strongly support the possibility of habitat reversions. The colonizations of freshwater by Hygrophila and of land by Stylommatophora were apparently phylogenetically independent although it cannot yet be excluded that there were transient terrestrial phases in the history of the former group or freshwater phases in the latter.

Phylogeographic analysis of the threatened and endangered superconglutinate-producing mussels of the genus Lampsilis (Bivalvia: Unionidae).

Several species of freshwater unionid mussels in the genus Lampsilis exhibit a remarkable reproductive strategy. Female mussels of these species enclose their larvae in a minnow-like lure, called a 'superconglutinate', to attract piscivorous fishes. When a fish attempts to ingest the superconglutinate the lure ruptures and the larvae are released to parasitize the fish. Of the four species of mussel which exhibit this strategy and are endemic to the Gulf Coast drainages of the southeastern United States, three are protected under the Endangered Species Act, and one is recognized as imperiled. Phylogenetic analysis of nucleotide sequences of the mitochondrial 16S ribosomal RNA and the first subunit of the cytochrome oxidase c genes was conducted on 18 individual specimens representing these four species and six outgroup taxa. Phylogenetic analyses of these data support the monophyly of the superconglutinate-producing mussels, and indicates a strong geographical component to the data. The zoogeographic patterns of the four taxa included in the study are congruent with those seen in freshwater vertebrates, and are consistent with a vicariant pattern resulting from fluctuations in sea level during the Pleistocene. Despite the strong geographical structuring of the data, only one species, Lampsilis subangulata, was recovered as monophyletic. The authors attribute the lack of support for the monophyly of the remaining species to insufficient sequence variation and the recent origin of the ancestor of these taxa. Based on these data, any future captive breeding projects aimed at augmenting or re-establishing populations should do so only from the appropriate source populations so as to maintain the genetic integrity of these nascent species.

Phylogenetic analysis of molluscan mitochondrial LSU rDNA sequences and secondary structures.

Mollusks are an extraordinarily diverse group of animals with an estimated 200,000 species, second only to the phylum Arthropoda. We conducted a comparative analysis of complete mitochondrial ribosomal large subunit sequences (LSU) of a chiton, two bivalves, six gastropods, and a cephalopod. In addition, we determined secondary structure models for each of them. Comparative analyses of nucleotide variation revealed substantial length variation among the taxa, with stylommatophoran gastropods possessing the shortest lengths. Phylogenetic analyses of the nucleotide sequence data supported the monophyly of Albinaria, Euhadra herklotsi + Cepaea nemoralis, Stylommatophora, Cerithioidea, and when only transversions are included, the Bivalvia. The phylogenetic limits of the mitochondrial LSU rRNA gene within mollusks appear to be up to 400 million years, although this estimate will have to be tested further with additional taxa. Our most novel finding was the discovery of phylogenetic signal in the secondary structure of rRNA of mollusks. The absence of entire stem/loop structures in Domains II, III, and V can be viewed as three shared derived characters uniting the stylommatophoran gastropods. The absence of the aforementioned stem/loop structure explains much of the observed length variation of the mitochondrial LSU rRNA found within mollusks. The distribution of these unique secondary structure characters within mollusks should be examined.

Phylogenetic utility of the tyrosine kinase gene X-src for assessing relationships among representative cichlid fishes.

The nuclear gene X-src is a member of the tyrosine-kinase class of proto-oncogenes whose normal product is localized within the cytoplasm of the cell. The X-src gene has been used in only a few phylogenetic studies, each focusing on systematics of killifishes (Cyprinodontiformes). The present study is an attempt to examine the phylogenetic utility of X-src for uncovering relationships of representative cichlid fishes, especially the cichlids of Middle America. The family Cichlidae is a species-rich group of tropical freshwater fish made up of more than 1000 species which show a Gondwanan pattern of distribution. Cichlid fish have been the focus of numerous studies ranging from behavioral to biogeographical to systematic in nature. Particular emphasis has focused on the cichlids of the African Great Lakes and the explosive adaptive radiation of this group. However, Neotropical cichlids have received considerably less attention than their African counterparts. Our findings regarding the utility of X-src concur with those of previous phylogenetic analyses showing the exons of X-src to be highly conserved and useful mostly for revealing deep relationships among taxa. Like previous X-src studies, we also found the intron sequences of the gene to be variable in length and difficult to align across distantly related taxa but they provided useful information for resolving relationships among more closely related taxa. The X-src phylogeny supports the monophyly of Neotropical cichlids and cichlasomines groups A (=heroines) + B (=cichlasomines). A highly resolved tree is obtained within the heroines but little support is evident for most nodes based on the low number of unambiguous substitutions. The X-src gene is likely to be quite useful for resolving deep phylogenetic relationships such as those among major groups of actinopterygian fishes.

Molecular systematics of Middle American cichlid fishes and the evolution of trophic-types in 'Cichlasoma (Amphilophus)' and 'C. (Thorichthys)'.

The majority of Middle American cichlids are placed in the informal assemblage 'Cichlasoma.' The group is divided into eight sections which appear to be based primarily on trophic morphology. Although several members of 'Cichlasoma' have been used in ecomorphological, behavioral, and biogeographic studies, no phylogenetic hypotheses for the group exist. In an attempt to develop a better understanding of the phylogenetic relationships of 'cichlasomine' cichlids, we examined the evolution of the trophic specialization, substratum-sifting, in two sections, 'Cichlasoma (Thorichthys)' and 'C. (Amphilophus),' to determine whether the trait reflects common ancestry. We sequenced the complete mitochondrial cytochrome b gene for 19 cichlids representing six sections of 'Cichlasoma,' and representatives of other Neotropical Cichlidae. Additional cichlid, and noncichlid outgroup sequences were included for a total of 22 taxa. The molecular phylogeny supports the recognition of the section 'C. (Thoricthys)' as a natural group, and we place those cichlids in the genus Thorichthys. The phylogeny also depicts 'C. (Amphilophus)' as paraphyletic, with substratum-sifters and generalized predators forming separate nonsister clades. We recommend that the substratum-sifting clade of the section 'C. (Amphilophus)' be placed in the resurrected genus Astatheros. The generalized predator clade of 'C. (Amphilophus)' contains only two species, 'C. (A.) citrinellum' and 'C. (A.) labiatum,' which we place in the genus Amphilophus. The phylogenetic hypotheses generated indicate that the substratum-sifting genera Thorichthys and Amphilophus do not share a common ancestor. Reconstruction of the evolution of substratum-sifting is equivocal, requiring either the independent evolution of the trait on two separate occasions or its presence in a more inclusive clade and subsequent loss in nonsubstratum sifting species.

A molecular phylogeny of Mobile River drainage basin pleurocerid snails (Caenogastropoda: Cerithioidea).

Sequences from the mitochondrial 16S rRNA gene were obtained to construct a molecular phylogeny for Mobile River drainage basin pleurocerid snails. Data from 876 aligned positions generated a single most-parsimonious tree for each of three analytical approaches: (1) equal weighting, (2) transversions weighted 2 x transitions; and (3) transversions weighted 4 x transitions. Identical topologies for the resulting trees depict the genera Elimia and Pleurocera as monophyletic sister taxa. The genus Leptoxis is paraphyletic with Leptoxis plicata sister to the Elimia + Pleurocera clade. L. taeniata and L. ampla are sister taxa and L. picta is the most basal pleurocerid examined. When transversions were weighted 10x transitions a single most-parsimonious tree was obtained with the only topological difference being L. picta depicted as sister to L. taeniata and L. ampla and L. plicata is now the most basal pleurocerid examined. Many of the Elimia species are closely related, but we await further data before making any taxonomic recommendations. L. picta and L. plicata are quite distinct from each other and all other pleurocerid species examined. These data serves as an important foundation for future studies examining conservation genetics and systematics of this diverse and imperiled family.

Molecular systematics and evolution of reproductive traits of North American freshwater unionacean mussels (Mollusca: Bivalvia) as inferred from 16S rRNA gene sequences.

North American freshwater unionacean bivalves are a diverse group of nearly 300 species. Unionaceans exhibit an array of conchological, anatomical, life history, and reproductive characteristics that have figured prominently in proposed classification schemes. Recently, two very different classifications of North American unionaceans have been proposed. Depending on the classification system utilized, a very different evolutionary trajectory of anatomical and reproductive features is obtained. The lack of a robust, well corroborated phylogeny of North American unionacean bivalves hinders the progress of evolutionary and ecological studies involving these species. Here we present a mitochondrial DNA (mtDNA) based phylogeny for North American unionacean mussels and compare it to previously proposed classifications. In addition, we present a 'total evidence' phylogeny which incorporates both the mtDNA sequence data and available morphological data. The molecular and total evidence phylogenies agree largely with the conclusions of a previous study based largely on immunoelectrophoretic data. North American unionaceans can be divided into two families: the Unionidae, which is comprised of most of the species and the Margaritiferidae. Within the Uniondae are two subfamilies, the Anodontinae and Ambleminae. The resultant phylogeny was used to examine the evolution of several key anatomical features including the number of gills (demibranchs) used by females to brood developing embryos, incubation length (bradytictic vs tachytictic), larval (glochidial) tooth structures, and shell texture. Both molecular and total evidence phylogenies indicate several of the aforementioned characters evolved independently or were subsequently lost or gained in several lineages.

A molecular phylogeny of the gopher tortoises, with comments on familial relationships within the Testudinoidea.

Sequences from the mitochondrial cytochrome b gene were obtained to examine molecular phylogenetic relationships among the North American gopher tortoises. Data from 352 aligned positions generated a single most-parsimonious tree for each of three analytical approaches: (1) equal weighting, all substitutions; (2) equal weighting, third position changes limited to transversions; and (3) transversions weighted 10 times transitions. Identical topologies for the resulting trees depict the gopher tortoises as a monophyletic group comprising two well-defined clades. Observed sequence divergence (7.0%) between the agassizii (Gopherus agassizii; G. berlandieri) and the polyphemus (G. flavomarginatus; G. polyphemus) clades suggests an Early Miocene separation of these lineages. The cytochrome b phylogeny complements some previous systematic interpretations, including formal taxonomic recognition of the two distinct groups, but is at odds with the most recent morphological analysis. Additional sequence comparisons of selected testudinoid (batagurid, emydid, and testudinid) taxa yielded a phylogeny consistent with a morphologically based hypothesis demonstrating close phylogenetic affinities between the Testudinidae and the "Bataguridae."

The evolution of copulatory organs, internal fertilization, placentae and viviparity in killifishes (Cyprinodontiformes) inferred from a DNA phylogeny of the tyrosine kinase gene X-src.

Cyprinodontiforms are a diverse group of approximately 900 pantropical and temperate fishes, mostly found in freshwater. Whereas the vast majority of fishes lay eggs (i.e. are oviparous), this group is unusual in that four groups of cyprinodont fishes give birth to living young (i.e. are viviparous). A molecular phylogenetic hypothesis was based on partial DNA sequences of the tyrosine kinase gene X-src. The study included the major lineages of fishes of the suborder Cyprinodontoidei, order Cyprinodontiformes. Our phylogeny agrees with some but not all of the conclusions of a previous morphological cladistic analysis (Parenti (Bull. Am. Mus. nat. Hist. 168, 335 (1981)). The differences are: (i) the Profundulidae are the sister group to the Goodeidae, not the sister group to all other cyprinodontoids; (ii) Fundulidae are the sister group to the Profundulidae and Goodeidae; (iii) Cubanichthys and the Cyprinodontinae might not be sister taxa; (iv) Cubanichthys, and not the Profundulidae, might be the most basal member of the cyprinodontoids; and (v) the Anablepinae and Poeciliinae might be sister groups. The molecular phylogeny was used to reconstruct the evolution of major life-history traits such as internal fertilization, copulatory organs, livebearing and placentas. Internal fertilization, modifications of the male's anal fin to form a copulatory organ, and viviparity probably evolved independently three times in cyprinodontiform fishes: in the subfamilies Goodeinae, Anablepinae and Poeciliinae (sensu Parenti 1981). The evolution of bundled sperm, spermatozeugmata, is probably not a prerequisite for internal fertilization because at least one species with internal fertilization has free spermatozoa. Livebearing (viviparity), which takes the form of ovoviviparity (where embryos are nourished by their yolk sac only), evolved only in the subfamily Poeciliinae. Advanced forms of viviparity, in which the mother provides additional nourishment to the embryos through placenta-like structures, apparently evolved at least three times from egg-laying ancestors: in the subfamilies Goodeinae, Anablepinae, and more than once in the Poeciliinae.

Parasitic and phoretic arthropods of the elephant-eared and the Santa Cruz kangaroo rats.

The parasitic and phoretic arthropods of the elephant-eared kangaroo rat (Dipodomys elephantinus) and Santa Cruz kangaroo rat (D. venustus) are reported; most of these represent new host records. Thirteen of 14 (93%) of D. elephantinus and 11 of 12 (92%) of D. venustus had 11 and nine arthropod species, respectively. Larval and nymphal stages of the tick Dermacentor occidentalis were the most prevalent parasite (77%) on D. elephantinus whereas the mesostigmatid mite Androlaelaps fahrenholzi was the most prevalent (56%) on D. venustus. The arthropod fauna of these two closely related rodents were similar with seven of the 14 arthropod species occurring on both host species. Two species of the host specific listrophorid mite of the genus Geomylichus were found on both hosts.

COMPARATIVE POPULATION GENETIC STRUCTURE OF A PARASITE (FASCIOLOIDES MAGNA) AND ITS DEFINITIVE HOST.

The population genetic structure of the American liver fluke, Fascioloides magna, and its definitive host the white-tailed deer, Odocoileus virginianus, was examined in South Carolina. Flukes were significantly more common in deer from river-swamp habitat than upland areas and prevalence increased with host age. The distribution of flukes among deer occurred as a negative binomial with the mean dispersion parameter, k, equal to 0.17 and the range from 0.10 to 1.11 within local areas. Significant spatial genetic differentiation was observed for flukes and deer. Patterns of genetic distance in flukes were not concordant with those of the definitive host nor were they related to geographic distance between sample locations. Spatial genetic differentiation among flukes reflected the tendency for individual hosts to harbor multiple individuals from a limited number of parasite clones. The large population size of the parasite and movements of the definitive host tend to counteract factors that lead to spatial differentiation.