IDENTITY OF THE ENDOSYMBIONT OF PERIDINIUM FOLIACEUM (PYRROPHYTA): ANALYSIS OF THE rbcLS OPERON1

Extant chromophytic algae have been suggested to have originated via the engulfment of a photo synthetic alga by a colorless protist. The dinoflagellate Peridinium foliaceum (Stein) Biecheler contains a reduced chlorophyll c–containing endosymbiont and, thus, represents an evolutionary intermediate stage in the establishment of chloroplasts. Although the exact phylogenetic relationship of the symbiont to extant algal species is unknown, it had been suggested that the P. foliaceum symbiont was either a diatom or a chrysophyte. Identification of the closest living relative of the P. foliaceum symbiont would provide a free-living model system with which the photosynthetic symbiont could be compared. Nucleotide sequence analysis of rbcL and rbcS (encoding the large and small subunits ofribulose-1,5-bisphosphate carboxylase/oxygenase) by the P. foliaceum symbiont was performed to provide insights into its identity. Cloned restriction fragments from a chloroplast DNA library were screened, and clones encoding the rbcLS operon were sequenced. Parsimony phylogenetic analysis was performed for each gene. Our data strongly suggest that the symbiont originated from a photosynthetic diatom.     

IDENTIFICATION OF CULTURED PSEUDO-NITZSCHIA (BACILLARIOPHYCEAE) USING SPECIES-SPECIFIC LSU rRNA-TARGETED FLUORESCENT PROBES1

Some, but not all, marine pennate diatoms of the genus Pseudo-nitzschia H. Peragallo are associated with the production of domoic acid, a naturally occurring amino acid responsible for amnesic shellfish poisoning. Distinguishing between potentially toxic and nontoxic representatives of this genus is time-consuming and difficult because it demands scanning electron microscopy of cleaned frustules. The objective of this work is to speed and ease identification of these organisms by using whole-cell (in situ) hybridization and species-specific large-subunit ribosomal RNA (LSU rRNA)-targeted oligonucleotide probes. Toward that end, cultures of P. australis Frenguelli, P. pungens (Grunow) Hasle, P. multiseries (Hasle) Hasle, P. fraudulenta (P. T. Cleve) Heiden, P. heimii Manguin, P. delicatissima (P. T. Cleve) Heiden, P. pseudo-delicatissima (Hasle) Hasle, and P. americana (Hasle) Fryxell were screened with a suite of 15 putative species-specific probes. Of those, a subset of eight probes was found that distinguished each species tested. In addition, Pseudo-nitzschia chloroplasts were labeled with a probe directed against a eubacterial-conserved sequence. Identification of new cultures based on their reactivity toward a set of probes agreed with species designations as defined by morphological criteria. Whole-cell hybridization is a rapid, simple, and cost-effective technique for discriminating among cultured Pseudo-nitzschia species.     

Phylogenetic Position and In Situ Identification of Ectosymbiotic Spirochetes on Protists in the Termite Gut

Phylogenetic relationships, diversity, and in situ identification of spirochetes in the gut of the termite Neotermes koshunensis were examined without cultivation, with an emphasis on ectosymbionts attached to flagellated protists. Spirochetes in the gut microbial community investigated so far are related to the genus Treponema and divided into two phylogenetic clusters. In situ hybridizations with a 16S rRNA-targeting consensus oligonucleotide probe for one cluster (known as termite Treponema cluster I) detected both the ectosymbiotic spirochetes on gut protists and the free-swimming spirochetes in the gut fluid of N. koshunensis. The probe for the other cluster (cluster II), which has been identified as ectosymbionts on gut protists of two other termite species, Reticulitermes speratus and Hodotermopsis sjoestedti, failed to detect any spirochete population. The absence of cluster II spirochetes in N. koshunensis was confirmed by intensive 16S ribosomal DNA (rDNA) clone analysis, in which remarkably diverse spirochetes of 45 phylotypes were identified, almost all belonging to cluster I. Ectosymbiotic spirochetes of the three gut protist species Devescovina sp., Stephanonympha sp., and Oxymonas sp. in N. koshunensis were identified by their 16S rDNA and by in situ hybridizations using specific probes. The probes specific for these ectosymbionts did not receive a signal from the free-swimming spirochetes. The ectosymbionts were dispersed in cluster I of the phylogeny, and they formed distinct phylogenetic lineages, suggesting multiple origins of the spirochete attachment. Each single protist cell harbored multiple spirochete species, and some of the spirochetes were common among protist species. The results indicate complex relationships of the ectosymbiotic spirochetes with the gut protists.     

Molecular phylogeny and evolution of phenotype in silica‐scaled chrysophyte genus Mallomonas

The evolution of phenotypes is highly understudied in protists, due to the dearth of morphological characters, missing fossil record, and/or unresolved phylogeny in the majority of taxa. The chrysophyte genus Mallomonas (Stramenopiles) forms species‐specific silica scales with extraordinary diversity of their ornamentation. In this paper, we molecularly characterized three additional species to provide an updated phylogeny of 43 species, and combined this with evaluations of 24 morphological traits. Using phylogenetic comparative methods, we evaluated phylogenetic signal in traits, reconstructed the trait evolution, and compared the overall phylogenetic and morphological diversity. The majority of traits showed strong phylogenetic signal and mostly dynamic evolution. Phylogenetic relatedness was often reflected by the phenotypic similarity. Both V‐rib and dome are very conservative structures that are presumably involved in precise scale overlap and bristle attachment, respectively. Based on modern species, it seems the dome firstly appeared on apical and/or caudal scales, and only later emerged on body scales. Bristle was presumably present in the common ancestor and gradually elongated ever since. However, most other morphological traits readily changed during the evolution of Mallomonas.     

Altering the balance between bacterial production and protistan bacterivory triggers shifts in freshwater bacterial community composition

Bacterivorous protists are known to induce changes in bacterial community composition (BCC). We hypothesized that changes in BCC could be related quantitatively to a measure of grazing: the ratio of bacterial mortality to growth rate. To test this hypothesis, we analyzed time-course changes in BCC, protistan grazing rate, and bacterial production from 3 in situ studies conducted in a freshwater reservoir and three laboratory studies. In the field experiments, samples were manipulated to yield different levels of protistan bacterivory and incubated in dialysis bags. Laboratory investigations were continuous cultivation studies in which different bacterivorous protists were added to bacterial communities. BCC was assessed using 4–6 different rRNA-targeted oligonucleotide probes for community analysis. Change in BCC (Δ BCC) was estimated as the sum of changes in the proportions of the two phylogenetic groups that showed the largest shifts. Analysis of a set of 22 estimates of shifts in the ratio of grazing to production rate over periods of 48–72 h and Δ BCC showed that Δ BCC was positively and tightly correlated (r ² = 0.784) with shifts in the ratio of grazing mortality to cell production. While the nature of a shift in BCC is unpredictable, the magnitude of the change can be related to changes in the balance between bacterial production and protistan grazing. This revised version was published online in August 2006 with corrections to the Cover Date.     

Scale evolution in Paraphysomonadida (Chrysophyceae): Sequence phylogeny and revised taxonomy of Paraphysomonas,new genus Clathromonas,and 25 new species

Heterotrophic chrysomonads of the genus Paraphysomonas are ubiquitous phagotrophs with diverse silica scale morphology. Over 50 named species have been described by electron microscopy from uncultured environmental samples. Sequence data exist for very few, but the literature reveals misidentification or lumping of most previously sequenced. For critically integrating scale and sequence data, 59 clonal cultures were studied light microscopically, by sequencing 18S ribosomal DNA, and recording scale morphology by transmission electron microscopy. We found strong congruence between variations in scale morphology and rDNA sequences, and unexpectedly deep genetic diversity. We now restrict Paraphysomonas to species with nail-like spine scales, establishing 23 new species and eight subspecies (Paraphysomonadidae). Species having base-plates with dense margins form three distinct subclades; those with a simple margin only two. We move 29 former Paraphysomonas species with basket scales into a new genus, Clathromonas, and describe two new species. Clathromonas belongs to a very distinct rDNA clade (Clathromonadidae fam. n.), possibly distantly sister to Paraphysomonas. Molecular and morphological data are mutually reinforcing; both are needed for evaluating paraphysomonad diversity and confirm excessive past lumping. Former Paraphysomonas species with neither nail-like nor basket scales are here excluded from Paraphysomonas and will be assigned to new genera elsewhere.     

ACCELERATED EVOLUTION OF LAND SNAILS MANDARINA IN THE OCEANIC BONIN ISLANDS: EVIDENCE FROM MITOCHONDRIAL DNA SEQUENCES

An endemic land snail genus Mandarina of the oceanic Bonin (Ogasawara) Islands shows exceptionally rapid evolution not only of morphological and ecological traits, but of DNA sequence. A phylogenetic relationship based on mitochondrial DNA (mtDNA) sequences suggests that morphological differences equivalent to the differences between families were produced between Mandarina and its ancestor during the Pleistocene. The inferred phylogeny shows that species with similar morphologies and life habitats appeared repeatedly and independently in different lineages and islands at different times. Sequential adaptive radiations occurred in different islands of the Bonin Islands and species occupying arboreal, semiarboreal, and terrestrial habitat arose independently in each island. Because of a close relationship between shell morphology and life habitat, independent evolution of the same life habitat in different islands created species possesing the same shell morphology in different islands and lineages. This rapid evolution produced some incongruences between phylogenetic relationship and species taxonomy. Levels of sequence divergence of mtDNA among the species of Mandarina is extremely high. The maximum level of sequence divergence at 16S and 12S ribosomal RNA sequence within Mandarina are 18.7% and 17.7%, respectively, and this suggests that evolution of mtDNA of Mandarina is extremely rapid, more than 20 times faster than the standard rate in other animals. The present examination reveals that evolution of morphological and ecological traits occurs at extremely high rates in the time of adaptive radiation, especially in fragmented environments.     

Identification and characterization of a bacteria-like sequence in the genome of some Silene species

The aim of this work was to characterize a nucleotide sequence MK14 that originated from a plasmid library obtained via degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR) amplification of laser microdissected Y-chromosomes of Silene latifolia. This sequence showed significant similarity to parts of two adjoining genes from bacterial representatives of the genus Ralstonia. MK14 sequence contains a part of a conserved domain, and phylogenetic analysis based on this region confirmed its relationship to Ralstonia-derived sequences. Genomic Southern blot analysis proved the presence of this fragment in the genome of S. latifolia. We hypothesize that this insertion is of bacterial origin, and was probably gained via horizontal gene transfer. Moreover, MK14 insertion is shared by some closely related Silene species, suggesting an ancient spontaneous transformation by an ancestor of bacteria from the genus Ralstonia.     

A TAXONOMIC STUDY OF THE GENUS PADINA (DICTYOTALES,PHAEOPHYCEAE) INCLUDING THE DESCRIPTIONS OF FOUR NEW SPECIES FROM JAPAN,HAWAII, AND THE ANDAMAN SEA1

A taxonomic study of the genus Padina from Japan, Southeast Asia, and Hawaii based on morphology and gene sequence data (rbcL and cox3) resulted in the recognition of four new species, that is, Padina macrophylla and Padina ishigakiensis from Ryukyu Islands, Japan; Padina maroensis from Hawaii; and Padina usoehtunii from Myanmar and Thailand. All species are bistratose and morphologically different from one another as well as from any known taxa by a combination of characters relating to degree of calcification; the structure, position, and arrangement of hairlines (HLs) and reproductive sori; and the presence or absence of rhizoid‐like groups of hairs and an indusium. Molecular phylogenetic analyses demonstrated a close relationship between P. ishigakiensis, P. macrophylla, P. maroensis, and Padina australis Hauck. The position of P. usoehtunii, however, was not fully resolved, being either sister to a clade comprising the other three new species and P. australis in the rbcL tree or more closely related to a clade comprising several other recently described species in the cox3 tree. The finding of the four new species demonstrates high species diversity particularly in southern Japan. The following characters were first recognized here to be useful for species delimitation: the presence or absence of small rhizoid‐like groups of hairs on the thallus surface, structure and arrangement of HLs on both surfaces either alternate or irregular, and arrangement of the alternating HLs between both surfaces in equal or unequal distance. The evolutionary trajectory of these and six other morphological characters used in species delineation was traced on the phylogenetic tree.     

A Bacterium Belonging to the Rickettsiaceae Family Inhabits the Cytoplasm of the Marine Ciliate Diophrys appendiculata (Ciliophora, Hypotrichia)

Bacteria of the family Rickettsiaceae (order Rickettsiales, α-Proteobacteria) are mainly known to be endosymbionts of arthropods with the capability to infect also vertebrate cells. Recently, they have also been found as leech endocytobionts. In the present paper, we report the first finding of a bacterium belonging to the family Rickettsiaceae in a natural population of a marine ciliate protozoan, namely Diophrys appendiculata, collected in the Baltic Sea. Bacteria were unambiguously identified through morphological characterization and the “full-cycle rRNA approach” (i.e., 16S rRNA gene characterization and use of specifically designed oligonucleotide probes for in situ detection). Symbionts are rod-shaped bacteria that grow freely in the cytoplasm of the host cell. They present two different morphotypes, similar in size, but different in cytoplasmic density. These are typical morphological features of members of the family Rickettsiaceae. 16S rRNA gene sequence showed that Diophrys symbionts share a high similarity value (>92%) with bacteria belonging to the genus Rickettsia. Phylogenetic analysis revealed that these new endosymbionts are clearly included in the clade of the family Rickettsiaceae, but they occupy an independent phylogenetic position with respect to members of the genus Rickettsia. This is the first report of a member of this family from a host protozoan and from a marine habitat. This result shows that this bacterial group is more diversified and widespread than supposed so far, and that its ecological relevance could until now have been underestimated. In light of these considerations, the two 16S rRNA oligonucleotide probes here presented, specific for members of the Rickettsiaceae, can represent useful tools for further researches on the presence and the spread of these microorganisms in the natural environment.     

MOLECULAR PHYLOGENY OF THE GREEN ALGAL ORDER PRASIOLALES (TREBOUXIOPHYCEAE,CHLOROPHYTA)1

The systematics of the Prasiolales was investigated by phylogenetic inference based on analyses of the rbcL and 18S rRNA genes for representatives of all four genera currently attributed to this order (Prasiococcus, Prasiola, Prasiolopsis, Rosenvingiella), including all type species. The rbcL gene had higher sequence divergence than the 18S rRNA gene and was more useful for phylogenetic inference at the ranks of genus and species. In the rbcL gene phylogeny, three main clades were observed, corresponding to Prasiola, Prasiolopsis, and Rosenvingiella. Prasiococcus was nested among species of Prasiola occurring in subaerial and supralittoral habitats. Trichophilus welckeri Weber Bosse, a subaerial alga occurring in the fur of sloths in Amazonia, was closely related to Prasiolopsis ramosa Vischer. The species of Prasiola were grouped into three well‐supported clades comprising (i) marine species, (ii) freshwater and terrestrial species with linear blades, and (iii) terrestrial species with rounded or fan‐shaped blades. Sequence divergence was unexpectedly low in the marine group, which included species with different morphologies. For the 18S rRNA gene, the phylogenetic analyses produced several clades observed for the rbcL gene sequence analysis, but, due to very little sequence variation, it showed considerably lower resolution for inference at the species and genus levels. Due to the low support of some internal branches, the results of the analyses did not allow an unambiguous clarification of the origin and the early evolution of the Prasiolales.     

高望界土壤放线菌抗菌活性筛选及生物学特征

为了从土壤环境中筛选产生抗菌活性物质的放线菌,采用琼脂扩散法对分离自湖南省高望界自然保护区森林土壤的169株放线菌进行抗菌活性检测,采用16S RNA基因序列分析法进行系统发育分析,采用常规方法进行生物学特性研究。结果表明,有47株受试菌株的发酵产物显示抗菌活性阳性(27.8%),其中6株具有较强抗菌活性。形态观察结果表明,这6株菌株均为典型的产丝产孢放线菌,在多数培养基上生长良好,气生菌丝和孢子丝丰富。系统发育分析表明,6株菌株均属于链霉菌属(Streptomyces),归为5个物种。菌株JSM 147612、JSM 147786、JSM 147831和JSM 147846分别与链霉菌属的已知物种S.violascens(序列相似性为99.93%)、S.malachitospinus(99.65%)、S.anulatus(99.72%)和S.aureus(99.38%)系统发育关系最为密切;菌株JSM 147823和JSM 147842之间的序列相似性为99.86%,应该属于同一物种,与它们系统发育关系最为密切的是S.albidoflavus(相似性分别为99.72%和99.58%)。     

Molecular biogeography: using the Corsica-Sardinia microplate disjunction to calibrate mitochondrial rDNA evolutionary rates in mountain newts (Euproctus)

Mitochondrial DNA (mtDNA) sequence variation was examined in the three species belonging to the newt genus Euproctus: E. asper, E. montanus, and E. platycephalus, and in three other species belonging to the same family: Triturus carnifex, T. vulgaris and Pleurodeles waltl. The Euproctus species inhabit mountain streams in the Pyrenean region, Corsica, and Sardinia, respectively. This vicariant distribution is believed to be a result of the disjunction and rotation of the Sardinia-Corsica microplate from the Pyrenean region and suggested dates for each cladogenetic event are available. A total of 915 bp from 12S and 16S ribosomal rRNA genes were compared for each taxon. These are the first mt-rDNA sequence data for salamanders. Sequences were used to reconstruct phylogenetic trees, investigate evolutionary rates for these genes, calibrate them with absolute time since divergence, and compare rates with published ones. Using P. waltl as the outgroup, all phylogenetic methods used (parsimony, maximum likelihood, and Neighbor Joining) produced trees with identical topologies and similar bootstrap values associated with each node. These sequence data cannot unambiguously resolve the splitting events leading to the main radiation of the genus Triturus and the origin of the genus Euproctus. These events may well have occurred very close in time, consistent with other sorts of data. Although it is unlikely strict linearity holds for all kinds of substitutions, relative rate tests of the molecular clock hypothesis could not reject clock-like behavior of sequence changes along Euproctus lineages. Estimates of absolute rates of base changes are 0.35% per Myr since divergence for all substitutions and 0.14% per Myr for transversions; these estimates are similar to other vertebrate estimates. A comparison with distance measures from allozyme studies agrees quite well with regard to relative divergences of the three Euproctus species.     

PHYLOGENETIC POSITION OF ATTHEYA LONGICORNIS AND ATTHEYA SEPTENTRIONALIS (BACILLARIOPHYTA)1

The phylogenetic position of diatoms belonging to the genus Attheya is presently under debate. Species belonging to this genus have been placed in the subclasses Chaetocerotophycidae and Biddulphiophycidae, but published phylogenetic trees based on 18S rDNA, morphology, and sexual reproduction indicate that this group of diatoms may be a sister group of the pennates. To clarify the position of Attheya, we studied the morphology, 18S rDNA, 16S rDNA of the chloroplasts, the rbcL large subunit (LSU) sequences of the chloroplasts, and the sterol composition of three different strains of Attheya septentrionalis (Østrup) R. M. Crawford and one strain of Attheya longicornis R. M. Crawford et C. Gardner. These data were compared with data from more than 100 other diatom species, covering the whole phylogenetic tree, with special emphasis on species belonging to the genera that have been suggested to be related to the genus Attheya. All data suggest that the investigated Attheya species form a separate group of diatoms, and there is no indication that they belong to either the Chaetocerotophycidae or the Biddulphiophycidae. Despite applying these various approaches, we were unable to determine the exact phylogenetic position of the investigated Attheya species within the diatoms.     

Cladistic tests of monophyly and relationships of biostratigraphically significant conodonts using multielement skeletal data –Lochriea homopunctatus and the genus Lochriea

Abstract: Since the 1960s, huge progress has been made in reconstructing the multielement skeletons of conodont species and developing a biologically defensible taxonomy. Nevertheless, a widespread prejudice remains that certain parts of the conodont skeleton, particularly the P₁ elements, are more informative than others with regard to taxonomy and evolutionary relationships. Here, we test these views. A new partial multielement reconstruction of the skeleton of the biostratigraphically significant conodont originally described as Gnathodus commutatus homopunctatus allows us to conduct a cladistic test of the alternative hypotheses of phylogenetic placement of this species. Our analysis also provides the first test of the hypothesis that Lochriea– species of which are markers for global correlation – is monophyletic and tests hypotheses concerning the origins of the genus. Our results demonstrate that homopunctatus is a species of Lochriea and that the genus is monophyletic. The widely held view that Lochriea arose from a species of Bispathodus is not supported. Our results show that it is difficult to predict a priori which parts of the conodont skeleton carry phylogenetic signal, and provide strong support for the hypothesis that similarity in the morphology of conodont P₁ elements alone is not a reliable guide to relationships and taxonomic groupings of conodont species. This is because P₁ elements with similar morphologies are convergently acquired in multiple conodont clades, because reliance on the characters of only one of the six or seven morphologically distinct elements of the conodont skeleton ignores phylogenetically significant data and because P₁ elements can lack characters that might seem to be diagnostic of a genus. Conodonts are no different to other organisms: ignoring data that have the potential to be phylogenetically informative is unlikely to produce the most reliable hypotheses of evolutionary relationships. We suggest that other biostratigraphically significant hypotheses of relationship between conodont taxa that are based on P₁ elements alone should be subject to cladistic testing.     

ULTRASTRUCTURE OF THE FEEDING APPARATUS AND MYONEMAL SYSTEM OF THE HETEROTROPHIC DINOFLAGELLATE PROTOPERIDINIUM SPINULOSUM1

The feeding veil or pallium of the thecate heterotrophic dinoflagellate Protoperidinium spinulosum Schiller is a highly vesiculate membranous sac containing several arched, sometimes bifurcated microtubular ribbons. It originates from an internal microtubular basket, passes through a sphincter-like osmiophilic ring located inside the posterior flagellar pore, and emerges from the cell at that pore. The osmiophilic ring is part of an interconnected myonemal system (composed of two striated collars and several striated connectives) that is anchored to the pore plate and to two inward protrusions composed of minute sulcal plates. A related species, Protoperidinium punctulatum (Paulsen) Balech, also possesses a microtubular basket/osmiophilic ring complex. Elongate electron-dense bodies within the basket resemble digestive secretory granules found in other protists. Granular, electron-lucent microbodies clustered at the anterior end of the basket may also have a role in prey digestion. Dense membranous whorls observed within a P. spinulosum cell presented as it was preparing to initiate feeding indicate a condensed storage site for pallium membranes. A narrow microtubule-strengthened pseudopodal appendage found in two non-feeding cells constitutes the tow filament that serves as the initial linkage between the dinoflagellate and its food. The structures that constitute the pallium and pallium precursors, described here for the first time, are unlike those of other known protists, although some similarities with the dinoflagellate peduncle are evident. The existence of this unique system of organelles may have important ramifications in the search for evolutionary relationships among protists.     

ORIGIN AND BIOGEOGRAPHY OF AESCULUS L. (HIPPOCASTANACEAE): A MOLECULAR PHYLOGENETIC PERSPECTIVE

Sequences of chloroplast gene matK and internal transcribed spacers of nuclear ribosomal RNA genes were used for phylogenetic analyses of Aesculus, a genus currently distributed in eastern Asia, eastern and western North America, and southeastern Europe. Phylogenetic relationships inferred from these molecular data are highly correlated with the geographic distributions of species. The identified lineages closely correspond to the five sections previously recognized on the basis of morphology. Ancestral character-state reconstruction, a molecular clock, and fossil evidence were used to infer the origin and biogeographic history of the genus within a phylogenetic framework. Based on the molecular phylogenetic reconstruction of the genus, sequence divergence, and paleontological evidence, we infer that the genus originated during the transition from the Cretaceous to the Tertiary (~65 M.Y.B.P.) at a high latitude in eastern Asia and spread into North America and Europe as an element of the “boreotropical flora”; the current disjunct distribution of the genus resulted from geological and climatic changes during the Tertiary.     

Phylogenetic Relationships of the Prodontria (Coleoptera; Scarabaeidae; Subfamily Melolonthinae), Derived from Sequence Variation in the Mitochondrial Cytochrome Oxidase II Gene

The beetle genus Prodontria is of importance to New Zealand conservation programs. All Prodontria species are brachypterous (having reduced wings), and the genus presents some interesting evolutionary and biogeographic questions that are testable using phylogenetic reconstruction. A phylogeny was produced for 14 flightless Prodontria species, 2 macropterous (fully winged) Odontria species, and single representatives of 2 outgroup genera using sequence data from the mitochondrial COII gene. The data support probable conspecificity of the morphologically similar P. modesta and P. bicolorata but do not support their hypothesized sister-species relationship with the geographically proximate P. lewisi. The alpine P. capito is found to be a paraphyletic group, with the most eastern population diverging after the western populations made their appearance. Many interesting biogeographic disjunctions are here proposed to be anomalous and the result of morphological convergence. The data do not support the idea of a common flightless ancester for Prodontria, but suggest that brachyptery has evolved numerous times. In some instances, this appears to have led to contemporaneous speciation resulting in little resolution of phylogenetic relationships in some parts of the tree. These data allow for a new interpretation of the origin and diversification of the southern New Zealand flightless melolonthine fauna. Multiple speciation events involving wing reduction are suggested to involve at least one widespread flighted ancestor that has given rise to brachypterous forms.     

Adaptive radiation in miniature: the minute salamanders of the Mexican highlands (Amphibia: Plethodontidae: Thorius)

The small size and apparent external morphological similarity of the minute salamanders of the genus Thorius have long hindered evolutionary studies of the group. We estimate gene and species trees within the genus using mitochondrial and nuclear DNA from nearly all named and many candidate species and find three main clades. We use this phylogenetic hypothesis to examine patterns of morphological evolution and species coexistence across central and southern Mexico and to test alternative hypotheses of lineage divergence with and without ecomorphological divergence. Sympatric species differ in body size more than expected after accounting for phylogenetic relationship, and morphological traits show no significant phylogenetic signal. Sympatric species tend to differ in a combination of body size, presence or absence of maxillary teeth, and relative limb or tail length, even when they are close relatives. Sister species of Thorius tend to occupy climatically similar environments, which suggests that divergence across climatic gradients does not drive species formation in the genus. Rather than being an example of cryptic species formation, Thorius more closely resembles an adaptive radiation, with ecomorphological divergence that is bounded by organism‐level constraints. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109 , 622–643.     

Oligonucleotide Probes for the Detection of Representatives of the Genus Thermoanaerobacter

Based on the analysis of 16S rRNA nucleotide sequences, oligonucleotide probes were designed for the detection of representatives of the genus Thermoanaerobacter. To increase the specificity of detection, the genus Thermoanaerobacter was divided into three groups. The probe Tab 827 (5"-GCTTCCGCDYCCCACACCTA-3") detected all known representatives of the genus Thermoanaerobacter; the probe Tab_1 844 (5"-TTAACTACGGCACGRAATGCTTC-3") was specific for the first group of species of the genus (T. wiegelii, T. siderophilus, T. sulfurophilus, T. brockii, T. kivui, T. ethanolicus, T. acetoethylicus, and T. thermohydrosulfuricus); the probe Tab_2 424 (5"-CACTAMYGGGGTTTACAACC-3") targeted the second group (T. thermocopriae, T. mathranii, and T. italicus); and the probe Tab_3 184 (5"-TCCTCCATCAGGATGCCCTA-3") was specific for the third group (T. tengcongensis, T. yonseiensis, T. subterraneus, and Carboxydibrachium pacificum, an organism related to the genus Thermoanaerobacter according to its 16S rRNA sequence). The oligonucleotide probes were labeled with Dig-11-dUTP. Hybridization with the probes showed the affiliation with Thermoanaerobacter of several pure cultures that were morphologically similar to representatives of this genus but possessed metabolic features unusual for it (capacity for agarose hydrolysis, anaerobic oxidation of CO, growth at low pH values) or were isolated from habitats previously unknown for Thermoanaerobacter (deep-sea hydrothermal vents).