A Description of a New "Amoebozoan" Isolated from the American Lobster, Homarus americanus

ABSTRACT. Our knowledge of the diversity of amoeboid protists is rapidly expanding as new and old habitats are more fully explored. In 2003, while investigating the cause of an amoeboid disease afflicting lobsters on the East Coast, samples were examined for the presence of amoebae from the carapace washings of the American lobster, Homarus americanus . During this survey a unique community of gymnamoebae was discovered. Among the new taxa discovered was a small Thecamoeba -like organism with a single posteriorly directed pseudopodium. Although resembling Parvamoeba rugata , this amoeba displayed distinctive morphology from that isolate or any other amoebozoan. Phylogenetic analysis shows this amoeba is distantly related to the Thecamoebidae. In this paper we describe the unique morphology of a second species of Parvamoeba and discuss its phylogenetic position with respect to the "Amoebozoa."     

18S ribosomal RNA gene sequences of Cochliopodium (Himatismenida) and the phylogeny of Amoebozoa

Cochliopodium is a very distinctive genus of discoid amoebae covered by a dorsal tectum of carbohydrate microscales. Its phylogenetic position is unclear, since although sharing many features with naked "gymnamoebae", the tectum sets it apart. We sequenced 18S ribosomal RNA genes from three Cochliopodium species (minus, spiniferum and Cochliopodium sp., a new species resembling C. minutum). Phylogenetic analysis shows Cochliopodium as robustly holophyletic and within Amoebozoa, in full accord with morphological data. Cochliopodium is always one of the basal branches within Amoebozoa but its precise position is unstable. In Bayesian analysis it is sister to holophyletic Glycostylida, but distance trees mostly place it between Dermamoeba and a possibly artifactual long-branch cluster including Thecamoeba. These positions are poorly supported and basal amoebozoan branching ill-resolved, making it unclear whether Discosea (Glycostylida, Himatismenida, Dermamoebida) is holophyletic; however, Thecamoeba seems not specifically related to Dermamoeba. We also sequenced the small-subunit rRNA gene of Vannella persistens, which constantly grouped with other Vannella species, and two Hartmannella strains. Our trees suggest that Vexilliferidae, Variosea and Hartmannella are polyphyletic, confirming the existence of two very distinct Hartmannella clades: that comprising H. cantabrigiensis and another divergent species is sister to Glaeseria, whilst Hartmannella vermiformis branches more deeply.     

Ovalopodium desertum n. sp. and the phylogenetic relationships of Cochliopodiidae (Amoebozoa)

An amoeba isolated from a weakly saline semi-desert pond in Kazakhstan (Central Asia) resembles a small Cochliopodium in the light microscope, but has a dorsal fibrous cell coat without scales. Thus it can be identified morphologically as a new species of Ovalopodium Sawyer, 1980, and it is herein named O. desertum. Phylogenetic analysis of the SSU rRNA gene sequences of the new species and four Cochliopodium spp. sequenced additionally shows that Ovalopodium desertum is a sister clade to a robustly monophyletic Cochliopodium. The close relationship between Ovalopodium and Cochliopodium is also confirmed by the analysis of SSU rRNA secondary structure showing the specific helices in the region V5 in all species of both genera. Analysis of actin gene sequences fails to resolve the position of Ovalopodium but demonstrates that Parvamoeba Rogerson, 1993 is probably related to Cochliopodium. The position of Cochliopodiidae within Amoebozoa remains unresolved, despite our efforts to resolve it using broader taxonomic sampling of Amoebozoa, testing alternative tree topologies and removing the fast-evolving sites. Among sequenced genera, Parvamoeba and Endostelium Olive et al., 1984 are probable relatives to Cochliopodiidae. Molecular trees weakly support an inclusion of the family in Flabellinia (Discosea), but more phylogenomic data are necessary to test this hypothesis.     

中国特有果蝇 Drosophila curviceps 种亚组在Drosophila immigrans种组中的种系发生地位

果蝇immigrans种组中的curviceps种亚组是1992年新建立的中国特有果蝇类群。该种亚组中的物种主要分布在中国大陆和台湾。目前除了形态学水平的研究外,还没有其他证据支持建立该种亚组的合理性及其起源和种系发生地位。为了在DNA分子水平上探讨果蝇curviceps种亚组在果蝇immigrans种组中的种系发生地位,从而为今后更深入地研究中国特有果蝇,甚至为果蝇亚属的进化遗传学提供理论依据,测定了immigrans种组5个种亚组（nasuta、immigrans、hypocausta、quadrilineata、curviceps)中12个代表物种的rDNA的ITS1和部分Adh基因的序列。其中ITS1序列的长度为513－587bp,共有191个信息位点;Adh基因片段的长度在714－747bp之间,共99个信息位点。考虑到单个分子提供的信息较少,将两个分子的序列综合起来,组成一个较长的复合序列。分别根据ITS1,Adh和两个分子的复合序列排比（Alignment)结果,和最大简约法和邻接法构建分子系统树,其中根据复合序列构建的系统树与形态学研究结果最为一致。分子树显示curviceps种亚组的特种确定单独形成一个分枝,为种亚组级的分类阶元,支持了形态学将其建立为一个新种亚组。根据Kimura距离,估算了复合分子的替换速率约为每百万年1．48％,进而计算出5个种亚组的分 歧年代。结合各物种的地理分布,推测了immigrans种组的进化历史：curviceps种亚组与quadrilineata种亚组的亲缘关系最近,主要分布在中国南部的温带地区。它们之间的分歧时间大约为3．4百万年,是最年轻的两个种亚组。主要分布在苏门答腊及附近的热带地区的hypocausta种亚组的物种是最早分化出来的,与其他种亚组的分歧时间约为9．2百万年。该结果与形态学和生物地理学研究相吻合。值得一提是的,目前归属仍存在争议的物种D.neohypocausta,在分子系统树中与hypocausta种亚组的物种相距较远,而与immiagrasn种亚组的关系较近,但分枝置信度较低（＜50％）。由于还缺乏其他方面的证据,因此D.neohypocausta的归属有待今后的研究来作定论。     

Phylogenetic position of Eulipotyphla inferred from the cDNA sequences of pepsinogens A and C

Although to date the phylogenetic position of the provisional order Eulipotyphla has been assessed by various molecular markers, it has not been conclusively clarified due to low statistical supporting values and inconsistent results. To clarify the phylogenetic position of Eulipotyphla, we cloned cDNAs for pepsinogens A and C from five mammalian species belonging to four different orders and determined their nucleotide sequences. Molecular phylogenetic analysis based on the 1st and 2nd codon positions of the protein-coding region of cDNA sequences strongly supported the close relationship between Eulipotyphla and Chiroptera. Carnivora was found to be a sister group to these two orders. The monophyly of the order Rodentia and that of the cohort Glires (Rodentia and Lagomorpha) was also shown by the present phylogenetic trees of pepsinogens.     

EXTINCTION RATES SHOULD NOT BE ESTIMATED FROM MOLECULAR PHYLOGENIES

Molecular phylogenies contain information about the tempo and mode of species diversification through time. Because extinction leaves a characteristic signature in the shape of molecular phylogenetic trees, many studies have used data from extant taxa only to infer extinction rates. This is a promising approach for the large number of taxa for which extinction rates cannot be estimated from the fossil record. Here, I explore the consequences of violating a common assumption made by studies of extinction from phylogenetic data. I show that when diversification rates vary among lineages, simple estimators based on the birth–death process are unable to recover true extinction rates. This is problematic for phylogenetic trees with complete taxon sampling as well as for the simpler case of clades with known age and species richness. Given the ubiquity of variation in diversification rates among lineages and clades, these results suggest that extinction rates should not be estimated in the absence of fossil data.     

Species tree estimation for a deep phylogenetic divergence in the New World monkeys (Primates: Platyrrhini)

The estimation of a robust phylogeny is a necessary first step in understanding the biological diversification of the platyrrhines. Although the most recent phylogenies are generally robust, they differ from one another in the relationship between Aotus and other genera as well as in the relationship between Pitheciidae and other families. Here, we used coding and non-coding sequences to infer the species tree and embedded gene trees of the platyrrhine genera using the Bayesian Markov chain Monte Carlo method for the multispecies coalescent (?BEAST) for the first time and to compared the results with those of a Bayesian concatenated phylogenetic analysis. Our species tree, based on all available sequences, shows a closer phylogenetic relationship between Atelidae and Cebidae and a closer relationship between Aotus and the Cebidae clade. The posterior probabilities are lower for these conflictive tree nodes compared to those in the concatenated analysis; this finding could be explained by some gene trees showing no concordant topologies between Aotus and the other genera. Moreover, the topology of our species tree also differs from the findings of previous molecular and morphological studies regarding the position of Aotus. The existence of discrepancies between morphological data, gene trees and the species tree is widely reported and can be related to processes such as incomplete lineage sorting or selection. Although these processes are common in species trees with low divergence, they can also occur in species trees with deep and rapid divergence. The sources of the inconsistency of morphological and molecular traits with the species tree could be a main focus of further research on platyrrhines.     

Phylogeny of Acomys spinosissimus (Rodentia, Muridae) from north Malawi and Tanzania: evidence from morphological and molecular analysis

New specimens of Acomys spinosissimus from northern Malawi and southern Tanzania were subjected to morphological, molecular (cytochrome b mitochondrial gene) and cytogenetical analysis in order to establish the phylogenetic position of these populations among A. spinosissimus, as well as their relationships with other species (A. subspinosus and A. wilsoni). The monophyly of the spinosissimus clade is supported by morphological and molecular phylogenies. Cytogenetics and morphology, but not the cytochrome b phylogeny show the possible existence of two sibling species within spinosissimus. Incongruences are noted between molecular and morphological trees, especially concerning the position of the Transvaal specimens. Two different biogeographic scenarios for the origin and dispersal of A. spinosissimus are discussed in the light of available palaeontological data.     

Molecular phylogeny of Sinocyclocheilus (Cypriniformes: Cyprinidae) inferred from mitochondrial DNA sequences

More than 10 species within the freshwater fish genus Sinoncyclocheilus adapt to caves and show different degrees of degeneration of eyes and pigmentation. Therefore, this genus can be useful for studying evolutionary developmental mechanisms, role of natural selection and adaptation in cave animals. To better understand these processes, it is indispensable to have background knowledge about phylogenetic relationships of surface and cave species within this genus. To investigate phylogenetic relationships among species within this genus, we determined nucleotide sequences of complete mitochondrial cytochrome b gene (1140 bp) and partial ND4 gene (1032 bp) of 31 recognized ingroup species and one outgroup species Barbodes laticeps. Phylogenetic trees were reconstructed using maximum parsimony, Bayesian, and maximum likelihood analyses. Our phylogenetic results showed that all species except for two surface species S. jii and S. macrolepis clustered as five major monophyletic clades (I, II, III, IV, and V) with strong supports. S. jii was the most basal species in all analyses, but the position of S. macrolepis was not resolved. The cave species were polyphyletic and occurred in these five major clades. Our results indicate that adaptation to cave environments has occurred multiple times during the evolutionary history of Sinocyclocheilus. The branching orders among the clades I, II, III, and IV were not resolved, and this might be due to early rapid radiation in Sinocyclocheilus. All species distributed in Yunnan except for S. rhinocerous and S. hyalinus formed a strongly supported monophyletic group (clade V), probably reflecting their common origins. This result suggested that the diversification of Sinocyclocheilus in Yunnan may correlate with the uplifting of Yunnan Plateau.     

Phylogeny of the genus Chironomus (Diptera) inferred from DNA sequences of mitochondrial cytochrome b and cytochrome oxidase I

Two mitochondrial genes, Cytochrome b (Cytb) and Cytochrome c oxidase subunit I (COI), have been used as phylogenetic markers in Chironomids. The nucleotide sequences of 685 bp from Cytb and 596 bp from COI have been determined for 36 Chironomus species from the Palearctic, or Holarctic, and Australasia. The concatenated sequence of 1281 bp from both genes was used to investigate the phylogenetic relationships among these species. The nucleotide sequence alignments were used for construction of phylogenetic trees based on maximum-parsimony and neighbor-joining methods. Both techniques produced similar phylogenies. Monophyly of the genus Chironomus is supported by a bootstrap value of 100% at the basal branch. Six clusters of species have been revealed with high bootstrap values supporting both monophyly of each cluster and the validity of the branching order within each cluster. Four species, C. circumdatus, C. nepeanensis, C. dorsalis, and C. crassiforceps, cannot be placed into any cluster. Cytological phylogenies were constructed using the same set of species, except for C. biwaprimus. These trees showed many similarities to that obtained from the mitochondrial (mt) sequence analysis, but also a number of significant differences. When compared with the tree constructed from the sequence of 23 species available for one of the globin genes, globin 2b (gb2b), there was better support for the mt tree than for the cytological trees. An intron, which varies in its occurrence and position in gb2b, was also investigated and the distribution of the introns supports the phylogenetic history of the genus Chironomus obtained with mt data. The differences observed in the cytological trees seem to be attributable more to the retention of the same chromosome banding sequence across several species, rather than convergent evolutionary events. An important question is the determination of the position of the subgenus Camptochironomus in relation to the representatives of the nominal subgenus Chironomus, since it has been suggested that this is a separate genus. The Camptochironomus species are internal to the trees and have arisen more recently than some of the species of the subgenus Chironomus, indicating that they are not sufficiently differentiated to be considered more than a subgenus.     

IS A NEW AND GENERAL THEORY OF MOLECULAR SYSTEMATICS EMERGING?

The advent and maturation of algorithms for estimating species trees—phylogenetic trees that allow gene tree heterogeneity and whose tips represent lineages, populations and species, as opposed to genes—represent an exciting confluence of phylogenetics, phylogeography, and population genetics, and ushers in a new generation of concepts and challenges for the molecular systematist. In this essay I argue that to better deal with the large multilocus datasets brought on by phylogenomics, and to better align the fields of phylogeography and phylogenetics, we should embrace the primacy of species trees, not only as a new and useful practical tool for systematics, but also as a long‐standing conceptual goal of systematics that, largely due to the lack of appropriate computational tools, has been eclipsed in the past few decades. I suggest that phylogenies as gene trees are a “local optimum” for systematics, and review recent advances that will bring us to the broader optimum inherent in species trees. In addition to adopting new methods of phylogenetic analysis (and ideally reserving the term “phylogeny” for species trees rather than gene trees), the new paradigm suggests shifts in a number of practices, such as sampling data to maximize not only the number of accumulated sites but also the number of independently segregating genes; routinely using coalescent or other models in computer simulations to allow gene tree heterogeneity; and understanding better the role of concatenation in influencing topologies and confidence in phylogenies. By building on the foundation laid by concepts of gene trees and coalescent theory, and by taking cues from recent trends in multilocus phylogeography, molecular systematics stands to be enriched. Many of the challenges and lessons learned for estimating gene trees will carry over to the challenge of estimating species trees, although adopting the species tree paradigm will clarify many issues (such as the nature of polytomies and the star tree paradox), raise conceptually new challenges, or provide new answers to old questions.     

A molecular phylogeny of New Zealand's Petroica (Aves: Petroicidae) species based on mitochondrial DNA sequences

The New Zealand robin (Petroica australis), tomtit (P. macrocephala), and Chatham Island black robin (P. traversi) are members of the Petroicidae family of Australo-Papuan robins, found throughout Australasia and the western Pacific. In the nearly 200 years since the New Zealand members of Petroicidae were first described, the division of species, subspecies, and even genera has undergone many changes. In this study, we investigate whether molecular phylogenies based on mitochondrial DNA sequences support current taxonomic classifications based on morphology. Petroica traversi, P. australis, and P. macrocephala form distinct clades in phylogenetic trees constructed from Cytochrome b and control region sequences, however the position of the black robin is at odds with the morphological and behavioral data. The black robin does not appear to be a derivative of the New Zealand robin, instead it groups strongly with the tomtit, indicating that lineage sorting and/or introgressive hybridization may have occurred. There is some evidence to support the hypothesis that two invasions of Petroica from Australia have occurred, however additional data from Australian Petroica taxa are required to confirm this. Control region sequences confirm a deep split between the North and South Island P. australis lineages, but suggest a recent radiation of P. macrocephala.     

Evolution of diversity: the Cape flora

Although the environmental correlates of plant species richness have long received attention, research into the genesis of this diversity is in its infancy. The recent development of molecular dating methods and the inference of robust phylogenetic hypotheses have made it possible to investigate this problem. I use the Cape flora as a model to show that much modern diversity could be the result of recruiting diverse lineages over the entire Cenozoic into this flora, followed by in situ diversification (thus increasing species richness) of at least some of these lineages over a long period in an environmentally heterogeneous area.     

Genetic identification and molecular phylogeny of Pseudodiaptomus species (Calanoida, Pseudodiaptomidae) in Korean waters

Five species of the genus Pseudodiaptomus, P. inopinus, P. poplesia, P. marinus, P. nihonkaiensis, and P. sp. occur in Korea nearshore waters. Although the four species except for P. sp. have been classified into Lobus and Ramosus groups, two for each group, based on morphological characters, this classification had yet to be confirmed by molecular characters. Here, we determined molecular characters and phylogenetic relationships of the five species in order to evaluate the morphology-based groupings and the species identifications. For this, a 625-bp DNA region of the mitochondrial gene cytochrome oxidase subunit I (mtCOI) was sequenced and compared among the species. Intraspecific variation of the sequences is less than 0.6%, while interspecific variation ranges from 17.6-26.7%, indicating every species, including P. sp., is a genetically distinct, valid species. Phylogenetic trees of the mtCOI DNA reveal that the Lobus-group species including P. inopinus and P. poplesia form a well-supported clade and that P. sp. belongs to this group. On the other hand, the Ramosus group clade consisting of P. marinus and P. nihonkaiensis is not well supported by bootstrap analyses, suggesting that further evaluation of the validity of this group assignment is needed.     

Congruency of phylogenies derived from different proteins

Summary This communication examines the question of phylogenetic congruency- i.e., whether or not the branching order of evolutionary trees is independent of the protein studied. It was found that trees constructed for birds on the basis of immunological comparison of their transferrins, albumins, and ovalbumins agree approximately with a published tree based on the amino acid sequences of their lysozymesc. This congruency is especially noteworthy with respect to the phylogenetic position of the chachalaca, a Mexican bird classified on morphological grounds in the family Cracidae of the order Galliformes. At the protein level, this species differs as much from non-cracid galliform birds as does the duck, which belongs to another order. Despite the organismal similarity between cracid and non-cracid galliform birds, the molecular relationship is remote. If this contrast between organismal and molecular results had been based on comparative studies with only lysozyme, one could have ascribed the contrast to the possibility that chachalaca lysozyme was paralogous, rather than orthologous, to the other bird lysozymesc. Examination of several proteins is thus desirable in cases of possible paralogy.This work was supported in part by grants GB-42028X from NSF and GM-21509 from NIH     

应用非序列联配方法对哺乳动物系统发育关系的探讨

非序列联配的序列分析方法,将序列中特定寡聚核苷酸的kmer统计频率作为特征,在序列间按特征进行比较和分析。这种方法综合考虑了所有变异类型对序列整体特征的影响,因而在组学数据分析上有独特的优势。但是,这类方法在复杂多细胞生物基因组系统发育中的适用性仍然有待检验。在本文中,我们使用基于非序列联配方法的CVTree软件,以45种哺乳动物的蛋白质组数据建立了系统发育关系NJ树,并据此探讨了哺乳动物系统发育的若干问题。在广受关注的真兽下纲四个总目的关系问题上,CVTree支持形态学的普遍结论即上兽类(Epitheria)假说。这与基于序列联配方法支持的外非洲胎盘类(Exafro-placentalia )假说不同。在哺乳动物内部目的层次上,CVTree树的结论与分子和形态所普遍接受的系统发育关系基本一致。但是在目的内部,CVTree树会有较多的差异。研究结果初步显示非序列联配方法在使用复杂多细胞生物的组学数据进行系统发育关系分析中的可行性。对非序列联配方法自身的改进及其与传统基于取代的序列联配方法之间的比较仍有待深入研究。     

5S rRNA sequences of 12 species of flatworms: implications for the phylogeny of the Platyhelminthes

5S rRNAs from 12 species of free living and parasitic platyhelminthes were sequenced. In the phylogenetic analysis, attention was focused on the statistical estimates of the trees corresponding to existing phylogenetic hypotheses. The available 5S rRNA data agree well with widely accepted views on the relationships between the Acoela, Polycladida, Tricladida, and Neorhabdocoela; our analysis of the published 18S rRNA sequences also demonstrated good correspondence between these views and molecular data. With available 5S rRNA data the hypothesis that the dalyellioid turbellarians is the sister group of the Neodermata is less convincing than the hypotheses proposing the Neodermata as the sister group of the Neorhabdocoela, or of the Seriata, or of the branch uniting them. A relatively low rate of base replacement in parasitic flatworms, probably, accounts for the uncertain position of the Neodermata, while a relatively high rate in planarians may explain a relatively too early divergence of the Tricladida in several published phylogenetic trees constructed from various rRNA data.     

Evolutionary history of the Tricladida and the Platyhelminthes: an up-to-date phylogenetic and systematic account

Within the free-living platyhelminths, the triclads, or planarians, are the best-known group, largely as a result of long-standing and intensive research on regeneration, pattern formation and Hox gene expression. However, the group's evolutionary history has been long debated, with controversies ranging from their phyletic structure and position within the Metazoa to the relationships among species within the Tricladida. Over the the last decade, with the advent of molecular phylogenies, some of these issues have begun to be resolved. Here, we present an up-to-date summary of the main phylogenetic changes and novelties with some comments on their evolutionary implications. The phylum has been split into two groups, and the position of the main group (the Rhabdithophora and the Catenulida), close to the Annelida and the Mollusca within the Lophotrochozoa, is now clear. Their internal relationships, although not totally resolved, have been clarified. Tricladida systematics has also experienced a revolution since the implementation of molecular data. The terrestrial planarians have been demonstrated to have emerged from one of the freshwater families, giving a different view of their evolution and greatly altering their classification. The use of molecular data is also facilitating the identification of Tricladida species by DNA barcoding, allowing better knowledge of their distribution and genetic diversity. Finally, molecular phylogenetic and phylogeographical analyses, taking advantage of recent data, are beginning to give a clear picture of the recent history of the Dugesia and Schmidtea species in the Mediterranean.     

Molecular analysis of populations of Ctenomys (Caviomorpha,Rodentia) with high karyotypic variability

The tuco-tucos (Ctenomys) are South American subterranean rodents that are some of the most chromosomally variable of all mammals. In this study we focus on Ctenomys of the "Corrientes species group" from that Argentine province and consisting of C. dorbignyi, C. perrensi, C. roigi and unnamed populations (Ctenomys sp.). A diploid range of 41-70 has been demonstrated for these taxa with multiple Robertsonian changes, pericentric inversions, heterochromatic modifications and other chromosomal rearrangements. To analyse the molecular variation in the Corrientes group, a 402 base pair fragment of mitochondrial cytochrome b was sequenced in 75 individuals from 26 populations. This generated 15 different haplotypes which were subjected to phylogenetic analysis. The different species within the Corrientes group failed to form monophyletic groups in the phylogenetic trees we generated and the divergences between haplotypes were low. Therefore, these molecular data demonstrate the recent subdivision of the Corrientes group with a substantial accumulation of chromosomal differences. Through our new chromosomal data and cytochrome b studies, we are able to subdivide the Ctenomys sp. populations into two groups and hypothesize on their relationships with the described Corrientes group species. Further studies are needed to establish the basis of the phenomenal chromosomal variation in this group.