Actinarctus doryphorus (Tanarctidae) DNA barcodes and phylogenetic reinvestigation of Arthrotardigrada with new A. doryphorus and Echiniscoididae sequences.

Little is still known about the diversity and evolution of marine arthrotardigrades, as they are generally difficult to sample, resulting in a limited amount of molecular data for barcoding and phylogenetic studies. With the current study, we provide the first investigation into COI haplotype diversity in a marine tanarctid and at the same time readdress arthrotardigrade phylogeny. Specifically, we provide COI mtDNA, 18S and 28S rDNA sequences from a population of Actinarctus doryphorus (Tanarctidae) sampled off the coast of Roscoff, France and further provide new 18S sequences from two marine echiniscoidids. A. doryphorus COI sequences confirmed the presence of a single species and further revealed five haplotypes shared among nine sequenced individuals. Our 18S and 28S rDNA datasets were individually and combined analysed with Bayesian inference and Maximum Likelihood. Actinarctus doryphorus was placed together with Tanarctus sequences within a maximally supported Tanarctidae, confirming previous interpretations that the clade is distinct from Halechiniscidae. Although several studies in recent decades have concluded that the marine arthrotardigrades are paraphyletic, recent studies have argued that the clade may not be paraphyletic. Our phylogenetic analyses consistently inferred Arthrotardigrada as paraphyletic, as the clade includes the monophyletic Echiniscoidea. Accordingly, we propose that it is time to suppress the order Arthrotardigrada as it clearly does not reflect tardigrade phylogeny.

Extreme freeze-tolerance in cryophilic tardigrades relies on controlled ice formation but does not involve significant change in transcription.

Subzero temperatures are among the most significant factors defining the distribution of organisms, yet, certain taxa have evolved to overcome this barrier. The microscopic tardigrades are among the most freeze-tolerant animals, with selected species reported to survive milli-Kelvin temperatures. Here, we estimate survival of fully hydrated eutardigrades of the species Ramazzottius varieornatus following exposures to -20 °C and  -80 °C as well as -196 °C with or without initial cooling to -80 °C. The tardigrades easily survive these temperatures, yet with a significant decrease in viability following rapid cooling by direct exposure to -196 °C. Hence, post-freeze recovery of R. varieornatus seems to rely on cooling rate and thus controlled ice formation. Cryophilic organisms are renowned for having cold-active enzymes that secure appropriate reaction rates at low temperatures. Hence, extreme freeze-tolerance in R. varieornatus could potentially involve syntheses of cryoprotectants and de novo transcription. We therefore generated a reference transcriptome for this cryophilic R. varieornatus population and explored for differential gene expression patterns following cooling to -80 °C as compared to active 5 °C controls. Specifically, we tested for fast transcription potentially occurring within 25 min of cooling from room temperature to a supercooling point of ca. -20 °C, at which the tardigrades presumably freeze and enter into the ametabolic state of cryobiosis. Our analyses revealed no evidence for differential gene expression. We, therefore, conclude that extreme freeze-tolerance in R. varieornatus relies on controlled extracellular freezing with any freeze-tolerance related genes being constitutively expressed.

20 years of Zootaxa: Tardigrada (Ecdysozoa: Panarthropoda).

Over the last two decades, Zootaxa has hosted nearly 200 papers concerning tardigrade taxonomy, systematics, phylogeny, and evolution. A total of 160 researchers from all continents (except the Antarctic) published descriptions of almost 200 new taxa, mostly species, but also genera and higher taxonomic ranks, such as families and superfamilies. This editorial is dedicated to the late Professor Clark W. Beasley who was the first tardigrade Associate Editor for Zootaxa.

Comparative myoanatomy of Tardigrada: new insights from the heterotardigrades Actinarctus doryphorus (Tanarctidae) and Echiniscoides sigismundi (Echiniscoididae).

BACKGROUND: Tardigrada is a group of microscopic invertebrates distributed worldwide in permanent and temporal aquatic habitats. Famous for their extreme stress tolerance, tardigrades are also of interest due to their close relationship with Arthropoda and Cycloneuralia. Despite recent efforts in analyzing the musculature of a number of tardigrade species, data on the class Heterotardigrada remain scarce. Aiming to expand the current morphological framework, and to promote the use of muscular body plans in elucidating tardigrade phylogeny, the myoanatomy of two heterotardigrades, Actinarctus doryphorus and Echiniscoides sigismundi, was analyzed by cytochemistry, scanning electron and confocal laser scanning microscopy and 3D imaging. We discuss our findings with reference to other tardigrades and internal phylogenetic relationships of the phylum. RESULTS: We focus our analyses on the somatic musculature, which in tardigrades includes muscle groups spanning dorsal, ventral, and lateral body regions, with the legs being musculated by fibers belonging to all three groups. A pronounced reduction of the trunk musculature is seen in the dorsoventrally compressed A. doryphorus, a species that generally has fewer cuticle attachment sites as compared to E. sigismundi and members of the class Eutardigrada. Interestingly, F-actin positive signals were found in the head appendages of A. doryphorus. Our analyses further indicate that cross-striation is a feature common to the somatic muscles of heterotardigrades and that E. sigismundi-as previously proposed for other echiniscoidean heterotardigrades-has relatively thick somatic muscle fibers. CONCLUSIONS: We provide new insights into the myoanatomical differences that characterize distinct evolutionary lineages within Tardigrada, highlighting characters that potentially can be informative in future phylogenetic analyses. We focus our current analyses on the ventral trunk musculature. Our observations suggest that seven paired ventromedian attachment sites anchoring a large number of muscles can be regarded as part of the ground pattern of Tardigrada and that fusion and reduction of cuticular attachment sites is a derived condition. Specifically, the pattern of these sites differs in particular details between tardigrade taxa. In the future, a deeper understanding of the tardigrade myoanatomical ground pattern will require more investigations in order to include all major tardigrade lineages.

Comparative transcriptomics suggest unique molecular adaptations within tardigrade lineages.

BACKGROUND: Tardigrades are renowned for their ability to enter cryptobiosis (latent life) and endure extreme stress, including desiccation and freezing. Increased focus is on revealing molecular mechanisms underlying this tolerance. Here, we provide the first transcriptomes from the heterotardigrade Echiniscoides cf. sigismundi and the eutardigrade Richtersius cf. coronifer, and compare these with data from other tardigrades and six eukaryote models. Investigating 107 genes/gene families, our study provides a thorough analysis of tardigrade gene content with focus on stress tolerance. RESULTS: E. cf. sigismundi, a strong cryptobiont, apparently lacks expression of a number of stress related genes. Most conspicuous is the lack of transcripts from genes involved in classical Non-Homologous End Joining. Our analyses suggest that post-cryptobiotic survival in tardigrades could rely on high fidelity transcription-coupled DNA repair. Tardigrades seem to lack many peroxins, but they all have a comprehensive number of genes encoding proteins involved in antioxidant defense. The "tardigrade unique proteins" (CAHS, SAHS, MAHS, RvLEAM), seem to be missing in the heterotardigrade lineage, revealing that cryptobiosis in general cannot be attributed solely to these proteins. Our investigation further reveals a unique and highly expressed cold shock domain. We hypothesize that the cold shock protein acts as a RNA-chaperone involved in regulation of translation following freezing. CONCLUSIONS: Our results show common gene family contractions and expansions within stress related gene pathways in tardigrades, but also indicate that evolutionary lineages have a high degree of divergence. Different taxa and lineages may exhibit unique physiological adaptations towards stress conditions involving possible unknown functional homologues and/or novel physiological and biochemical mechanisms. To further substantiate the current results genome assemblies coupled with transcriptome data and experimental investigations are needed from tardigrades belonging to different evolutionary lineages.

Modelling extreme desiccation tolerance in a marine tardigrade.

It has recently been argued that the enigmatic tardigrades (water bears) will endure until the sun dies, surviving any astrophysical calamities in Earth's oceans. Yet, our knowledge of stress tolerance among marine tardigrade species is very limited and most investigations revolve around species living in moist habitats on land. Here, we investigate desiccation tolerance in the cosmopolitan marine tidal tardigrade, Echiniscoides sigismundi, providing the first thorough analysis on recovery upon desiccation from seawater. We test the influence on survival of desiccation surface, time spent desiccated (up to 1 year) and initial water volume. We propose analysis methods for survival estimates, which can be used as a future platform for evaluating and analysing recovery rates in organisms subjected to extreme stress. Our data reveal that marine tidal tardigrades tolerate extremely rapid and extended periods of desiccation from seawater supporting the argument that these animals are among the toughest organisms on Earth.

Comparative Investigation of Copper Tolerance and Identification of Putative Tolerance Related Genes in Tardigrades.

Tardigrades are microscopic aquatic animals renowned for their tolerance toward extreme environmental conditions. The current study is the first to investigate their tolerance toward heavy metals and we present a novel tardigrade toxicant tolerance assay based on activity assessments as a measure of survival. Specifically, we compare tolerance toward copper in four species representing different evolutionary lineages, habitats and adaptation strategies, i.e., a marine heterotardigrade, Echiniscoides sigismundi, a limno-terrestrial heterotardigrade, Echiniscus testudo, a limno-terrestrial eutardigrade, Ramazzottius oberhaeuseri, and a marine eutardigrade, Halobiotus crispae. The latter was sampled at a time of year, when the population is predominantly represented by aberrant P1 cysts, while the other species were in normal active states prior to exposure. Based on volume measurements and a general relation between body mass and copper tolerance, expected tardigrade EC50 values were estimated at 0.5-2 µg l-1. Following 24 h of exposure, tolerance was high with no apparent link to lineage or habitat. EC50s (95% CI), 24 h after exposure, were estimated at 178 (168-186) and 310 (295-328) µg l-1, respectively, for E. sigismundi and R. oberhaeuseri, whereas E. testudo and H. crispae were less affected. Highest tolerance was observed in H. crispae with a mean ± s.e.m. activity of 77 ± 2% (n = 3) 24 h after removal from ~3 mg l-1 copper, suggesting that tardigrade cysts have increased tolerance toward toxicants. In order to identify putative tolerance related genes, an E. sigismundi transcriptome was searched for key enzymes involved in osmoregulation, antioxidant defense and copper metabolism. We found high expression of Na/K ATPase and carbonic anhydrase, known targets for copper. Our transcriptome, furthermore, revealed high expression of antioxidant enzymes, copper transporters, ATOX1, and a Cu-ATPase. In summary, our results indicate that tardigrades express well-known key osmoregulatory enzymes, supporting the hypothesis that copper inhibits sodium turnover as demonstrated for other aquatic organisms. Tardigrades, nevertheless, have high tolerance toward the toxicant, which is likely linked to high expression of antioxidant enzymes and an ability to enter dormant states. Tardigrades, furthermore, seem to have a well-developed battery of cuproproteins involved in copper homeostasis, providing basis for active copper sequestering and excretion.

The tardigrade fauna of Australian marine caves: with descriptions of nine new species of Arthrotardigrada.

Marine caves are known to support a rich macrofauna; however, few studies have focused on meiofauna. Marine cave meiofaunal tardigrades have been reported from Japan and the Mediterranean Sea and a preliminary list of species including a redescription of Actinarctus neretinus Grimaldi de Zio, D'Addabbo Gallo, Morone De Lucia, Vaccarella and Grimaldi, 1982 was reported from Fish Rock Cave and Jim's Cave on the coast of Australia. This study is the fourth in a series describing the unique meiofauna in two Australian submarine caves located off the coast of New South Wales, describing nine new species.        Only 67 tardigrades were collected from the two caves, yet these contained a high diversity of at least 16 different species which are quite different in the two caves. The fauna includes nine arthrotardigrade genera: Actinarctus, Batillipes, Dipodarctus, Halechiniscus, Raiarctus, Styraconyx, Tanarctus, Tholoarctus, and Wingstrandarctus. This fauna is different from that reported for the high energy beaches along the East Coast of Australia.        We describe nine new species comprising a single batillipedid and eight halechiniscids: Batillipes solitarius nov. sp., Dipodarctus australiensis nov. sp., Dipodarctus susannae nov. sp., Raiarctus jesperi nov. sp., Raiarctus katrinae nov. sp., Tanarctus hirsutospinosus nov. sp., Tholoarctus oleseni nov. sp., Wingstrandarctus stinae nov. sp. and Wingstrandarctus unsculptus nov. sp.

Brain anatomy of the marine tardigrade Actinarctus doryphorus (Arthrotardigrada).

Knowledge of tardigrade brain structure is important for resolving the phylogenetic relationships of Tardigrada. Here, we present new insight into the morphology of the brain in a marine arthrotardigrade, Actinarctus doryphorus, based on transmission electron microscopy, supported by scanning electron microscopy, conventional light microscopy as well as confocal laser scanning microscopy. Arthrotardigrades contain a large number of plesiomorphic characters and likely represent ancestral tardigrades. They often have segmented body outlines and each trunk segment, with its paired set of legs, may have up to five sensory appendages. Noticeably, the head carries numerous cephalic appendages that are structurally equivalent to the sensory appendages of the trunk segments. Our data reveal that the brain of A. doryphorus is partitioned into three paired lobes, and that these lobes exhibit a more pronounced separation as compared to that of eutardigrades. The first brain lobe in A. doryphorus is located anteriodorsally, with the second lobe just below it in an anterioventral position. Both of these two paired lobes are located anterior to the buccal tube. The third pair of brain lobes are situated posterioventrally to the first two lobes, and flank the buccal tube. In addition, A. doryphorus possesses a subpharyngeal ganglion, which is connected with the first of the four ventral trunk ganglia. The first and second brain lobes in A. doryphorus innervate the clavae and cirri of the head. The innervations of these structures indicate a homology between, respectively, the clavae and cirri of A. doryphorus and the temporalia and papilla cephalica of eutardigrades. The third brain lobes innervate the buccal lamella and the stylets as described for eutardigrades. Collectively, these findings suggest that the head region of extant tardigrades is the result of cephalization of multiple segments. Our results on the brain anatomy of Actinarctus doryphorus support the monophyly of Panarthropoda.

Bulinus globosus (Planorbidae; Gastropoda) populations in the Lake Victoria basin and coastal Kenya show extreme nuclear genetic differentiation.

Bulinus globosus, a key intermediate host for Schistosoma haematobium that causes urinary schistosomiasis, is a hermaphroditic freshwater Planorbid snail species that inhabits patchy and transient water bodies prone to large seasonal variations in water availability. Although capable of self-fertilizing, this species has been reported to be preferentially out crossing. In this study, we characterized the population genetic structure of 19 B. globosus populations sampled across the Lake Victoria basin and coastal Kenya using four polymorphic microsatellite loci. Population genetic structure was characterized and quantified using FST statistics and Bayesian clustering algorithms. The four loci used in this study contained sufficient statistical power to detect low levels of population genetic differentiation and were highly polymorphic with the number of alleles per locus across populations ranging from 16 to 22. Average observed and expected heterozygosities across loci in each population ranged from 0.13 to 0.69 and from 0.39 to 0.79, respectively. Twenty-five of the seventy-six possible population-locus comparisons significantly deviated from Hardy-Weinberg equilibrium proportions after Bonferroni corrections, mostly due to the deficiency of heterozygotes. Significant genetic differentiation was observed between populations and Bayesian inferences identified 15 genetic clusters. The excess homozygosity, significant inbreeding and population genetic differentiation observed in B. globosus populations are likely to be due to the habitat patchiness, mating system and the proneness to cyclic extinction and recolonization in transient habitats.

The ITS2 of the genus Bulinus: novel secondary structure among freshwater snails and potential new taxonomic markers.

The freshwater snail genus Bulinus has been intensively investigated due to its role as intermediate host for trematode blood flukes that cause the debilitating disease schistosomiasis in man and livestock. Owing to taxonomic ambiguities within Bulinus, attention has often focused upon species delineation and several molecular methods have recently been used for identification and characterization purposes. Inspection of compensatory base changes (CBCs) in the secondary structure of the nuclear ribosomal internal transcribed spacer (ITS) has been used to differentiate species in other genera, and here we present a study investigating the presence of CBCs between species in the species groups within Bulinus. CBCs were present within B. forskalii and B. globosus indicating that these widely distributed taxa might constitute cryptic species complexes. However, other currently recognized species could not be distinguished by CBC analysis. The putative secondary structure of the very long ITS2 sequence of the B. reticulatus species group had an additional helix (DIIa) between DII and DIII not seen in other species groups of Bulinus. The accumulation and inspection of further ITS2 sequences will no doubt reveal additional variation between Bulinus populations, and CBCs should be incorporated in future taxonomic work in this group.

Surface enhanced Raman scattering on Tardigrada--towards monitoring and imaging molecular structures in live cryptobiotic organisms.

Tardigrades are microscopic metazoans which are able to survive extreme physical and chemical conditions by entering a stress tolerant state called cryptobiosis. At present, the molecular mechanisms behind cryptobiosis are still poorly understood. We show that surface enhanced Raman scattering supported by plasmonic gold nanoparticles can measure molecular constituents and their local distribution in live tardigrades. Surface enhanced Raman signatures allow to differentiate between two species and indicate molecular structural differences between tardigrades in water and in a dry state. This opens new avenues for exploring cryptobiosis by studying molecular changes in live cryptobiotic organisms.

Inorganic ion composition in Tardigrada: cryptobionts contain a large fraction of unidentified organic solutes.

Many species of tardigrades are known to tolerate extreme environmental stress, yet detailed knowledge of the mechanisms underlying the remarkable adaptations of tardigrades is still lacking, as are answers to many questions regarding their basic biology. Here, we present data on the inorganic ion composition and total osmotic concentration of five different species of tardigrades (Echiniscus testudo, Milnesium tardigradum, Richtersius coronifer, Macrobiotus cf. hufelandi and Halobiotus crispae) using high-performance liquid chromatography and nanoliter osmometry. Quantification of the ionic content indicates that Na(+) and Cl(-) are the principal inorganic ions in tardigrade fluids, albeit other ions, i.e. K(+), NH4(+), Ca(2+), Mg(2+), F(-), SO4(2-) and PO4(3-) were also detected. In limno-terrestrial tardigrades, the respective ions are concentrated by a large factor compared with that of the external medium (Na(+), ×70-800; K(+), ×20-90; Ca(2+) and Mg(2+), ×30-200; F(-), ×160-1040, Cl(-), ×20-50; PO4(3-), ×700-2800; SO4(2-), ×30-150). In contrast, in the marine species H. crispae, Na(+), Cl(-) and SO4(2-) are almost in ionic equilibrium with (brackish) salt water, while K(+), Ca(2+), Mg(2+) and F(-) are only slightly concentrated (×2-10). An anion deficit of ~120 mEq l(-1) in M. tardigradum and H. crispae indicates the presence of unidentified ionic components in these species. Body fluid osmolality ranges from 361±49 mOsm kg(-1) in R. coronifer to 961±43 mOsm kg(-1) in H. crispae. Concentrations of most inorganic ions are largely identical between active and dehydrated groups of R. coronifer, suggesting that this tardigrade does not lose large quantities of inorganic ions during dehydration. The large osmotic and ionic gradients maintained by both limno-terrestrial and marine species are indicative of a powerful ion-retentive mechanism in Tardigrada. Moreover, our data indicate that cryptobiotic tardigrades contain a large fraction of unidentified organic osmolytes, the identification of which is expected to provide increased insight into the phenomenon of cryptobiosis.

Desiccation tolerance in the tardigrade Richtersius coronifer relies on muscle mediated structural reorganization.

Life unfolds within a framework of constraining abiotic factors, yet some organisms are adapted to handle large fluctuations in physical and chemical parameters. Tardigrades are microscopic ecdysozoans well known for their ability to endure hostile conditions, such as complete desiccation--a phenomenon called anhydrobiosis. During dehydration, anhydrobiotic animals undergo a series of anatomical changes. Whether this reorganization is an essential regulated event mediated by active controlled processes, or merely a passive result of the dehydration process, has not been clearly determined. Here, we investigate parameters pivotal to the formation of the so-called "tun", a state that in tardigrades and rotifers marks the entrance into anhydrobiosis. Estimation of body volume in the eutardigrade Richtersius coronifer reveals an 87 % reduction in volume from the hydrated active state to the dehydrated tun state, underlining the structural stress associated with entering anhydrobiosis. Survival experiments with pharmacological inhibitors of mitochondrial energy production and muscle contractions show that i) mitochondrial energy production is a prerequisite for surviving desiccation, ii) uncoupling the mitochondria abolishes tun formation, and iii) inhibiting the musculature impairs the ability to form viable tuns. We moreover provide a comparative analysis of the structural changes involved in tun formation, using a combination of cytochemistry, confocal laser scanning microscopy and 3D reconstructions as well as scanning electron microscopy. Our data reveal that the musculature mediates a structural reorganization vital for anhydrobiotic survival, and furthermore that maintaining structural integrity is essential for resumption of life following rehydration.

Neuroanatomy of Halobiotus crispae (Eutardigrada: Hypsibiidae): Tardigrade brain structure supports the clade Panarthropoda.

The position of Tardigrada in the animal tree of life is a subject that has received much attention, but still remains controversial. Whereas some think tardigrades should be categorized as cycloneuralians, most authors argue in favor of a phylogenetic position within Panarthropoda as a sister group to Arthropoda or Arthropoda + Onychophora. Thus far, neither molecular nor morphological investigations have provided conclusive results as to the tardigrade sister group relationships. In this article, we present a detailed description of the nervous system of the eutardigrade Halobiotus crispae, using immunostainings, confocal laser scanning microscopy, and computer-aided three-dimensional reconstructions supported by transmission electron microscopy. We report details regarding the structure of the brain as well as the ganglia of the ventral nerve cord. In contrast to the newest investigation, we find transverse commissures in the ventral ganglia, and our data suggest that the brain is partitioned into at least three lobes. Additionally, we can confirm the existence of a subpharyngeal ganglion previously called subesophagal ganglion. According to our results, the original suggestion of a brain comprised of at least three parts cannot be rejected, and the data presented supports a sister group relationship of Tardigrada to 1) Arthropoda or 2) Onychophora or 3) Arthropoda + Onychophora.

Molecular phylogeny of Arthrotardigrada (Tardigrada).

Tardigrades are microscopic ecdysozoans with a worldwide distribution covering marine, limnic and terrestrial habitats. They are regarded as a neglected phylum with regard to studies of their phylogeny. During the last decade molecular data have been included in the investigation of tardigrades. However, the marine arthrotardigrades are still poorly sampled due to their relative rarity, difficult identification and minute size even for tardigrades. In the present study, we have sampled various arthrotardigrades and sequenced the 18S and partial 28S ribosomal subunits. The phylogenetic analyses based on Bayesian inference and maximum parsimony inferred Heterotardigrada (Arthrotardigrada+Echiniscoidea) and Eutardigrada to be monophyletic. Arthrotardigrada was inferred to be paraphyletic as the monophyletic Echiniscoidea is included within the arthrotardigrades. The phylogenetic positions of Stygarctidae and Batillipedidae are poorly resolved with low branch support. The Halechiniscidae is inferred to be polyphyletic as the currently recognized Styraconyxinae is not part of the family. Archechiniscus is the sister-group to the Halechiniscidae and Orzeliscus is placed as one of the basal halechiniscids. The phylogeny of the included eutardigrade taxa resembles the current molecular phylogenies. The genetic diversity within Arthrotardigrada is much larger (18S 15.1-26.5%, 28S 7.2-20.7%) than within Eutardigrada (18S 1.0-12.6%, 28S 1.3-8.2%). This can be explained by higher substitution rates in the arthrotardigrades or by a much younger evolutionary age of the sampled eutardigrades.

Genetic diversity of schistosomes and snails: implications for control.

Molecular approaches are providing new insights into the genetic diversity of schistosomes and their intermediate snail hosts. For instance, molecular tools based on the polymerase chain reaction are being developed for the diagnosis of schistosomiasis and the detection of prepatent schistosome infections in snails at transmission sites. Robust phylogenies of the different species of Schistosoma, Bulinus and Biomphalaria have been determined and novel methods are available to identify the different and cryptic taxa involved. Microsatellite analyses and mitochondrial DNA sequencing methods have been developed and are contributing to a better understanding of the genetic structure of both schistosome and snail populations. New sampling procedures to capture DNA of eggs and larval stages of schistosomes in field situations are facilitating more detailed and ethically advantageous studies on parasite heterogeneity. Knowledge of the genetic diversity of schistosome and snail populations adds a further dimension to the monitoring and surveillance of disease, and the implementation of new molecular-based approaches will be of increasing importance in helping to assess the impact of schistosomiasis control strategies.

Molecular phylogenetic investigations of the Viviparidae (Gastropoda: Caenogastropoda) in the lakes of the Rift Valley area of Africa.

The freshwater gastropod family Viviparidae is nearly cosmopolitan, but absent from South America. On the African continent, two genera are recognized; the widespread Bellamya and the monotypic Neothauma, which is confined to Lake Tanganyika. Most of the African Bellamya species are confined to the major lakes of the Rift Valley area in Africa, i.e. Lake Albert, Lake Malawi, Lake Mweru, and Lake Victoria. The phylogenetic analyses of mitochondrial (COI and 16S) and nuclear (H3, 18S and 28S) DNA inferred three major lake-clades; i.e. Lake Victoria/Kyoga/Albert, Lake Malawi and Lake Mweru/Bangweulu. The endemic B. rubicunda from Lake Albert and B. unicolor from Lake Kyoga were inferred to be part of the Lake Victoria clade. Bellamya capillata as identified by shell characters was polyphyletic in gene trees. The monophyletic Bellamya species radiation in Lake Malawi was most nearly related to the Lake Victoria/Kyoga/Albert-clade. Taxa from the Zambian lakes, Mweru and Bangweulu, were inferred together and placed ancestral to the other lakes. Neothauma tanganyicense was inferred as the sister-group to the Zambian Bellamya. Within the lake-clades the endemic radiations show very low genetic diversities (0-4.1% in COI), suggesting much faster morphological divergence than molecular divergence. Alternatively, Bellamya in Africa constitutes only a few species with several sub-species or eco-phenotypic morphs. The African viviparids were inferred to be the sister-group to a clade comprising Asian species, and the relatively low genetic diversity between the clades (12.6-15.5% in COI) makes a recent Miocene dispersal event from Asia to Africa much more likely than an ancient Gondwana vicarience distribution.

An investigation of the "Ancyloplanorbidae" (Gastropoda, Pulmonata, Hygrophila): preliminary evidence from DNA sequence data.

The Planorbidae is the largest family of freshwater pulmonate snails, yet an understanding of their intrafamily phylogenetic relationships is lacking and existing inferences are tentative. Moreover, it has been suggested that the Ancylidae, limpet-like freshwater pulmonates, should be merged with Planorbidae according to analysis of internal organ morphology. The present study explicitly tests this hypothesis by phylogenetic inference from partial DNA sequences of three molecular markers, nuclear ribosomal small subunit 18S and the mitochondrial cytochrome oxidase, and large subunit 16S. A molecular phylogeny was inferred based upon 22 taxa representing 12 ancylid and planorbid genera; additional taxa were included from the authors' database and from available sequences from GenBank, to further explore this basic data set. Taxa from Acroloxidae, Lymnaeidae, and Physidae were used as outgroups. Ancylidae and Planorbidae were found to be paraphyletic, with Planorbidae including some members of Ancylidae. "Ancyloplanorbidae" was also found to be paraphyletic because Acroloxus (Acroloxidae) surprisingly was included. Burnupia was found to be ancestral to "Ancyloplanorbidae" (including Acroloxus). The following clades of Planorbidae were supported: Bulininae and Planorbinae, Biomphalarini (including Helisoma and Planorbarius), and Planorbini and Segmentini.