Phylogeography, geographic structure, genetic variation, and potential species boundaries in Philippine slender toads

We investigated phylogeography of Philippine slender toads (genus Ansonia) and used a temporal framework for diversification, statistical tests of alternate topologies, and Bayesian approaches to test previous hypotheses concerning dispersal to, and colonization routes within, the southern Philippine island of Mindanao. Two species of Ansonia previously have been documented, with ranges separated by an east–west split corresponding to the approximate boundaries of Mindanao’s paleoisland precursors. We present new mtDNA sequence data (1946 bp from genes encoding ND1, 16S rRNA and tRNA^Leu) for 105 Ansonia specimens sampled from 20 localities on Mindanao Island. Our data suggest that Philippine Ansonia is composed of at least eight, well-supported population lineages, structured into a minimum of four highly divergent mtDNA clades. One clade corresponds to Ansonia mcgregori, a range-restricted species apparently limited to the distal portion of the Zamboanga Peninsula of western Mindanao. Two morphologically indistinguishable, but genetically divergent, lineages possibly are undescribed cryptic species from western Mindanao. We recognize the five remaining lineages as Ansonia muelleri pending data from morphology or bioacoustics that might diagnose separate species among these lineages. Regardless of their species status, the five allopatric lineages of A. muelleri should be viewed as important genetic units for future genetic conservation planning.     

Molecular phylogeny and systematics of the pseudoxyrhophiine snake genus Liopholidophis (Reptilia, Colubridae): evolution of its exceptional sexual dimorphism and descriptions of new taxa

The pseudoxyrhophiine snake genus Liopholidophis Mocquard, 1904 is endemic to Madagascar and consists of two distinct species groups. We performed molecular phylogenetic analyses using nucleotide sequences of c . 2200 bp from two mitochondrial genes (16S rRNA, cyt b ) and one nuclear gene (c-mos) to test the monophyly of the genus Liopholidophis and to investigate the relationships of the known species and an unidentified Liopholidophis population. Our phylogeny strongly supports the polyphyly of the genus Liopholidophis , confirms the monophyly of both species groups, and reveals that the unidentified population belongs to a new species. We therefore transfer the species of the former L. stumpffi group ( L. epistibes , L. infrasignatus , L. lateralis , L. martae , and L. stumpffi ) to a new genus Bibilava gen. n. and describe Liopholidophis dimorphus sp. n. from the rainforests of Montagne d'Ambre National Park in northern Madagascar. Finally, we compare our phylogeny with previous hypotheses and discuss the evolution of the extreme sexual dimorphism in tail length of Liopholidophis s. str.     

The complete mitochondrial genome of the spotted steed, Hemibarbus maculatus (Teleostei, Cypriniformes)

The complete mitogenome sequence of the spotted steed (Hemibarbus maculatus) was determined using long PCR reactions. The genome is 16,611 bp long and consists of 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and a control region. The gene order and composition of H. maculatus was similar to that of most other vertebrates. The base composition of H-strand in descending order is A (29.16%), C (27.41%), T (25.93%), and G (17.50%), with an AT content of 55.09%. The Kimura two-parameter distances between mitogenome sequences of H. maculatus and Hemibarbus labeo, Hemibarbus barbus, Hemibarbus longirostris, Hemibarbus mylodon were 2.09%, 2.35%, 12.77%, and 13.92%, respectively. The phylogenetic relationships among the five Hemibarbus species based on the whole mitogenome sequence data were (((H. barbus, H. labeo), H. maculatus), (H. longirostris, H. mylodon)).     

A new Ansonia from the Isthmus of Kra, Thailand (Amphibia, Anura, Bufonidae)

A new species of torrent-dwelling bufonid frog of the genus Ansonia is described from the Isthmus of Kra, Thailand. Ansonia kraensis is morphologically similar to Malaysian A. malayana, but differs from it in ventral coloration and larval morphology. Occurrence of A. kraensis in this region suggests a heterogeneous nature of the anuran fauna between northern and southern regions of the Malay Peninsula.     

Phylogenetic relationships within Aglaopheniidae (Cnidaria,Hydrozoa) reveal unexpected generic diversity

Morphology can be misleading in the representation of phylogenetic relationships, especially in simple organisms like cnidarians and particularly in hydrozoans. These suspension feeders are widely distributed in many marine ecosystems, and the family Aglaopheniidae Marktanner‐Turneretscher, 1890 is among the most diverse and visible, especially on tropical coral reefs. The taxonomy of this family is based on morphological characters with emphasis on reproductive structures for the identification of genera. This study is the most comprehensive molecular phylogeny of the Aglaopheniidae to date, including six genera and 38 species, of which 13 were investigated for the first time and sampled on tropical coral reefs throughout the Indo‐Pacific region. For newly sampled individuals, we sequenced the 16S rRNA, the nuclear locus comprising the complete ITS1‐5.8S rRNA gene‐ITS2 and the first intron of the calmodulin nuclear gene. Phylogenetic analyses of the data revealed and confirmed a general polyphyly, or doubtful monophyly, of all sampled genera in tropical regions based on both the mitochondrial and nuclear markers. Our results revealed that several morphological characters used today are unsuited to resolve phylogenetic relationships between species and genera, as well as the high phyletic diversity within this family. Future revision of the classification of this family will require extensive geographic sampling and the use of an integrative approach.     

Inbreeding depression in two seed-feeding beetles, Callosobruchus maculatus and Stator limbatus (Coleoptera: Chrysomelidae)

Inbreeding depression is well documented in insects but the degree to which inbreeding depression varies among populations within species, and among traits within populations, is poorly studied in insects other than Drosophila. Inbreeding depression was examined in two long-term laboratory colonies of the seed beetle, Callosobruchus maculatus (Fabricius), which are used frequently as models for experiments in ecology, evolution and behaviour. Inbreeding depression in these laboratory colonies are compared with one recently field-collected population of a different seed beetle, Stator limbatus Horn. Inbreeding reduced embryogenesis, egg hatch and larval survival in both species, such that eggs produced by sib matings were >17% less likely to produce an adult offspring. Inbred larvae also took 4-6% longer to develop to emergence in both species. Inbreeding depression varied among the measured traits but did not differ between the two populations of C. maculatus for any trait, despite the large geographic distance between source populations (western Africa vs. southern India). Inbreeding depression was similar in magnitude between C. maculatus and S. limbatus. This study demonstrates that these laboratory populations of C. maculatus harbour substantial genetic loads, similar to the genetic load of populations of S. limbatus recently collected from the field.     

Comparative sequence analyses on the 16S rRNA (rDNA) of Bacillus acidocaldarius, Bacillus acidoterrestris, and Bacillus cycloheptanicus and proposal for creation of a new genus, Alicyclobacillus gen. nov.

Comparative 16S rRNA (rDNA) sequence analyses performed on the thermophilic Bacillus species Bacillus acidocaldarius, Bacillus acidoterrestris, and Bacillus cycloheptanicus revealed that these organisms are sufficiently different from the traditional Bacillus species to warrant reclassification in a new genus, Alicyclobacillus gen. nov. An analysis of 16S rRNA sequences established that these three thermoacidophiles cluster in a group that differs markedly from both the obligately thermophilic organisms Bacillus stearothermophilus and the facultatively thermophilic organism Bacillus coagulans, as well as many other common mesophilic and thermophilic Bacillus species. The thermoacidophilic Bacillus species B. acidocaldarius, B. acidoterrestris, and B. cycloheptanicus also are unique in that they possess omega-alicylic fatty acid as the major natural membranous lipid component, which is a rare phenotype that has not been found in any other Bacillus species characterized to date. This phenotype, along with the 16S rRNA sequence data, suggests that these thermoacidophiles are biochemically and genetically unique and supports the proposal that they should be reclassified in the new genus Alicyclobacillus.     

Systematics of limbless scincid lizards from northern Madagascar: morphology,phylogenetic relationships and implications for classification (Squamata: Scincidae)

We report on the rediscovery of two limbless scincid species, Paracontias rothschildi Mocquard, 1905 and Paracontias minimus (Mocquard, 1906), after more than a century. The two species were found in syntopy in sandy soils of Forêt d’Orangea, Antsiranana Province, northern Madagascar, which probably constitutes the respective type locality and confirms the species’ Malagasy origin. Both taxa are redescribed based on newly collected material, and compared to other Malagasy species. In addition, Paracontias fasika n. sp. is described from the same locality and habitat. We discuss the taxonomy and origin of all three species and provide preliminary data on their natural history. Molecular relationships among seven Paracontias species are compared to external morphological characters formerly used in skink systematics. Our results indicate that morphology in fossorial skinks is well suited to distinguish species, but is of rather limited value to elucidate phylogenetic relationships. Similarities between these skinks in external characters apparently are the result of convergent evolution due to parallel selective pressures.     

The nucleotide sequences of 5S rRNAs from a rotifer, Brachionus plicatilis, and two nematodes, Rhabditis tokai and Caenorhabditis elegans.

The nucleotide sequences of 5S rRNAs from a rotifer, Brachionus plicatilis, and two nematodes, Rhabditis tokai and Caenorhabditis elegans have been determined. The rotifer has two 5S rRNA species that are composed of 120 and 121 nucleotides, respectively. The sequences of these two 5S rRNAs are the same except that the latter has an additional base at its 3'-terminus. The 5S rRNAs from the two nematode species are both 119 nucleotides long. The sequence similarity percents are 79% (Brachionus/Rhabditis), 80% (Brachionus/Caenorhabditis), and 95% (Rhabditis/Caenorhabditis) among these three species. Brachionus revealed the highest similarity to Lingula (89%), but not to the nematodes (79%).     

Extreme intraspecific mitochondrial DNA sequence divergence in Galaxias maculatus (Osteichthys: Galaxiidae), one of the world's most widespread freshwater fish

Biogeographic controversies surrounding the widespread freshwater fish, Galaxias maculatus, were addressed with DNA sequence data. Mitochondrial cytochrome b and 16S rRNA sequences were obtained from representatives of six populations of this species. Substantial levels of cytochrome b (maximum 14.6%) and 16S rRNA sequence divergence (maximum 6.0%) were detected between western Pacific (Tasmania-New Zealand) and South American (Chile-Falkland Islands) haplotypes. A considerable level of divergence was also detected between Tasmanian and New Zealand haplotypes (maximum 5.1%) and within and among Chilean and Falkland Island G. maculatus (maximum 3. 8%). The phylogenetic structure of haplotypes conflicts with the accepted pattern of continental fragmentation. Molecular clock calibrations suggest that haplotype divergences postdate the fragmentation of Gondwana. These findings point to marine dispersal rather than ancient vicariance as an explanation for the wide distribution. The phylogenetic structure of South American haplotypes was not consistent with their geographic distribution. We consider factors such as population divergence, population size, dispersal, secondary contact, and philopatry as potential causes of the high level of mtDNA nucleotide diversity in this species.     

Molecular systematics of African 20-chromosome toads (Anura: Bufonidae)

Intrageneric lineages and the historical biogeography of toads, genus Bufo, are poorly resolved due to their conservative morphology, their highly conserved karyotypes (typically 2N = 22), and erratic patterns of interspecific hybridisation. Here, we use mitochondrial and nuclear DNA sequence data to reconstruct relationships of the 20-chromosome toads, a major component of the African bufonid fauna. Mitochondrial 12S and 16S sequences from 29 species revealed two independent transitions between phenotypically distinct savannah and forest adapted forms. Analyses of mitochondrial 12S, 16S, and ND2, along with nuclear ACTC and Rhodopsin sequences from 12 species greatly increased bootstrap, and likelihood support for internal branches including a basal split into two pan-African 20-chromosome clades. Hybridisation is a weak indicator of phylogenetic relationship as it occurs across these deeply divergent clades, between Bufo rangeri and Bufo gutturalis. These analyses suggest a secondary reversion to 22-chromosomes in Bufo pardalis, within the 20-chromosome group, although we could not reject an alternative hypothesis that this lineage forms a sister to all 2N = 20 toads. Other informally recognised 22-chromosome groups form independent phylogenetic lineages outside the 20-chromosome group, such as the Angusticeps and Vertebralis divisions. Bufo lindneri, from the Taitanus division, is closely related to Stephopaedes anotis, and these species should be considered congeners.     

A broad-host-range, generalized transducing phage (SN-T) acquires 16S rRNA genes from different genera of bacteria

Genomic analysis has revealed heterogeneity among bacterial 16S rRNA gene sequences within a single species; yet the cause(s) remains uncertain. Generalized transducing bacteriophages have recently gained recognition for their abundance as well as their ability to affect lateral gene transfer and to harbor bacterial 16S rRNA gene sequences. Here, we demonstrate the ability of broad-host-range, generalized transducing phages to acquire 16S rRNA genes and gene sequences. Using PCR and primers specific to conserved regions of the 16S rRNA gene, we have found that generalized transducing phages (D3112, UT1, and SN-T), but not specialized transducing phages (D3), acquired entire bacterial 16S rRNA genes. Furthermore, we show that the broad-host-range, generalized transducing phage SN-T is capable of acquiring the 16S rRNA gene from two different genera: Sphaerotilus natans, the host from which SN-T was originally isolated, and Pseudomonas aeruginosa. In sequential infections, SN-T harbored only 16S rRNA gene sequences of the final host as determined by restriction fragment length polymorphism analysis. The frequency of 16S rRNA gene sequences in SN-T populations was determined to be 1 x 10(-9) transductants/PFU. Our findings further implicate transduction in the horizontal transfer of 16S rRNA genes between different species or genera of bacteria.     

Remotely-sensed land use patterns and the presence of Anopheles larvae (Diptera: Culicidae) in Sukabumi, West Java, Indonesia

Land use patterns and the occurrence of Anopheles species larvae were studied in Sukabumi District, West Java, Indonesia, from October 2004 to September 2005. Two land use maps derived using remote sensing were used. One map derived from Quickbird satellite images of 150 km2 of the Simpenan and Ciemas subdistricts (106 degrees 27' 53"-106 degrees 38' 38" E and 6 degrees 59' 59"-7 degrees 8' 46" S) in Sukabumi and one using ASTER images covering 4,000 km2 of Sukabumi District from 106 degrees 22' 15"-107 degrees 4' 1" E and 6 degrees 42' 50" - 7 degrees 26' 13" S. There was a total of 11 Anopheles spp. collected from 209 sampling locations in the area covered by the Quickbird image and a total of 15 Anopheles spp. collected from 1,600 sampling locations in the area covered by the ASTER map. For the area covered by the land use maps, ten species were found to have statistically positive relationships between land use class and species presence: Anopheles aconitus, An. annularis, An. barbirostris. An. flavirostris, An. insulaeflorum, An. kochi, An. maculatus, An. subpictus, An. sundaicus, and An. vagus. Quickbird and ASTER satellite images both produced land maps that were adequate for predicting species presence in an area. The land use classes associated with malaria vector breeding were rice paddy (An. aconitus, An. subpictus), plantation located near or adjacent to human settlements (An. maculatus), bush/shrub (An. aconitus, An. maculatus, An. sundaicus), bare land, and water body land use on the coast located < or = 250 m of the beach (An. sundaicus). Understanding the associations of habitat and species in one area, predictions of species presence or absence can be made prior to a ground survey allowing for accurate vector survey and control planning.     

Identical 5S rRNA nucleotide sequence of Vibrio cholerae strains representing temporal, geographical, and ecological diversity.

Enzymatic determination of the nucleotide sequence of 5S rRNA prepared from a Vibrio cholerae non-O1 serovar isolated from a canal in Louisiana in 1978 revealed it to be identical to that of V. cholerae O1 isolated in 1905 from a pilgrim at the E1 Tor quarantine station. The two strains are associated with sharply contrasting habitats. Ranges of variation of 5 to 15% in procaryotic 5S rRNA sequences between species are typical; variation within species may be 2% or greater. Because the 5S rRNA sequences were identical, it is concluded that they have a common phylogenic origin.     

A Broad-Host-Range, Generalized Transducing Phage (SN-T) Acquires 16S rRNA Genes from Different Genera of Bacteria

Genomic analysis has revealed heterogeneity among bacterial 16S rRNA gene sequences within a single species; yet the cause(s) remains uncertain. Generalized transducing bacteriophages have recently gained recognition for their abundance as well as their ability to affect lateral gene transfer and to harbor bacterial 16S rRNA gene sequences. Here, we demonstrate the ability of broad-host-range, generalized transducing phages to acquire 16S rRNA genes and gene sequences. Using PCR and primers specific to conserved regions of the 16S rRNA gene, we have found that generalized transducing phages (D3112, UT1, and SN-T), but not specialized transducing phages (D3), acquired entire bacterial 16S rRNA genes. Furthermore, we show that the broad-host-range, generalized transducing phage SN-T is capable of acquiring the 16S rRNA gene from two different genera: Sphaerotilus natans, the host from which SN-T was originally isolated, and Pseudomonas aeruginosa. In sequential infections, SN-T harbored only 16S rRNA gene sequences of the final host as determined by restriction fragment length polymorphism analysis. The frequency of 16S rRNA gene sequences in SN-T populations was determined to be 1 × 10⁻⁹ transductants/PFU. Our findings further implicate transduction in the horizontal transfer of 16S rRNA genes between different species or genera of bacteria.     

Identical 5S rRNA nucleotide sequence of Vibrio cholerae strains representing temporal, geographical, and ecological diversity

Enzymatic determination of the nucleotide sequence of 5S rRNA prepared from a Vibrio cholerae non-O1 serovar isolated from a canal in Louisiana in 1978 revealed it to be identical to that of V. cholerae O1 isolated in 1905 from a pilgrim at the E1 Tor quarantine station. The two strains are associated with sharply contrasting habitats. Ranges of variation of 5 to 15% in procaryotic 5S rRNA sequences between species are typical; variation within species may be 2% or greater. Because the 5S rRNA sequences were identical, it is concluded that they have a common phylogenic origin.     

Detection and identification of Theileria infection in sika deer ( Cervus nippon ) in China

The sika deer ( Cervus nippon ) is a first-grade state-protected animal in China and designated a threatened species by the World Conservation Union. To detect hemoparasite infection of sika deer, blood samples were collected from 24 animals in the Hubei Province Deer Center. Genomic DNA was extracted, and the V4 hypervariable region encoding 18S rRNA was analyzed by reverse line blot hybridization assay. PCR products hybridized with Babesia / Theileria genus-specific probes but failed to hybridize with any of the Babesia or Theileria species-specific probes, suggesting the presence of a novel, or variant, species. Here 18S rRNA and internal transcribed spacer (ITS) genes were amplified, cloned, and sequenced from 7 isolates. Alignment and BlastN of the cloned sequences revealed high similarities to the homologous 18S rRNA genes and ITS genes of Theileria cervi (AY735122), Theileria sp. CNY1A (AB012194), and Theileria sp. ex Yamaguchi (AF529272). Phylogenetic analysis based on the 18S rRNA gene and ITS sequences showed that all cloned sequences were grouped within the Theileria clade. Phylogeny based on the 18S rRNA gene divided the organisms into 2 groups. Group 1 was closest to Theileria sp. ex Yamaguchi (AF529272), and group 2 was distinct from all other identified Theileria and Babesia species. These results suggest the existence of Theileria sp. infection in sika deer in China. To our knowledge, this is the first report of cervine Theileria sp. in China.     

Burmoniscus kitadaitoensis Nunomura, 2009 (Crustacea,Isopoda, Oniscidea) from southern Japan,a junior synonym of B. meeusei (Holthuis, 1947)

Re-examination of the holotype of Burmoniscus kitadaitoensis Nunomura, 2009 from Kitadaitojima Island, southern Japan reveals that this species is a junior synonym of B. meeusei (Holthuis, 1947). Partial regions of mitochondrial COI, 12S and 16S rRNA genes, and nuclear 18S and 28S rRNA genes were detected for species identification in the future.     

Phylogenetic characterization of Centipeda periodontii, Selenomonas sputigena and Selenomonas species by 16S rRNA gene sequence analysis.

The nearly complete 16S rRNA gene sequences for oral Gram-negative anaerobic motile bacteria, Centipeda periodontii, Selenomonas sputigena and Selenomonas species (formerly S. sputigena type strain), were determined in order to unveil their relationship to other oral motile bacteria. To determine the phylogenetic characterization of these bacteria, their 16S rRNA gene sequences were obtained and compared with those from the ribosomal sequence databases previously reported. The 16S rRNA gene sequences of these bacteria were similar to those of Selenomonas ruminantium and Schwartzia succinivorans isolated from rumens, and to Pectinatus cerevisiiphilus isolated from spoiled beer. Among oral bacteria, the nucleotide sequence analysis of these bacteria revealed high nucleotide similarity to Veillonella species, whereas low similarity to oral motile bacteria such as Campylobacter species. Phylogenetic analysis clearly confirmed that C. periodontii and two Selenomonas species were classified as relatives of a group besides Selenomonas, Schwartzia, and Pectinatus species, and not as close relatives to oral motile bacteria, such as Campylobacter species. These results suggest that such oral Gram-negative anaerobic motile bacteria are close relatives of oral bacteria.