Molecular phylogeny of some genera of Pamphagidae (Acridoidea, Orthoptera) from China based on mitochondrial 16S rDNA sequences

Based on the mitochondrial 16S ribosomal DNA partial sequences (473 bp) of 9 species of Pamphagidae (Acridoidea, Orthoptera) from China and of 4 species of Pamphagidae and 2 species of Pyrgomorphidae and Acrididae (as outgroups) retrieved from GenBank, we constructed the molecular phylogeny using the Neighbor Joining (NJ) and Minimum Evolution ( ME) methods based on the nucleotide Kimura 2-parameter model. The results of our study shown that: 1) the ranges of the 16S rDNA nucleotide divergence between two species of a genus were 0.21%, among genera of a subfamily were 0.42-3.38%, and among subfamilies of Pamphagidae were 1.90-8.88%, respectively. The phylogenetic tree shows that: 1) all Pamphagidae taxa form a monophyletic clade, and are well separated from the outgroup; 2) the African taxa Porthetinae (Lobosceliana brevicornis) and Akicerinae (Batrachotetrix sp.) are distinctly separated from the Chinese taxa Prionotropisinae; 3) Haplotropis bruneriana and Glauia terrea of Pamphaginae are nested in the middle of the tree, but their phylogenetic status is uncertain in this study; 4) 8 genera of Asiotmethis, Beybienkia, Mongolotmethis, Sinotmethis, Rhinotmethis, Filchnerella, Eotmethis and Pseudotmethis from China are all grouped into the subfamily Prionotropisinae, but their phylogenetic relationships are not clearly resolved.

Analysis of 16S rDNA sequences from pathogenic Leptospira serovars and use of single nucleotide polymorphisms for rapid speciation by D-HPLC

Leptospira have a worldwide distribution and include important zoonotic pathogens yet diagnosis and differentiation still tend to rely on traditional bacteriological and serological approaches. In this study a 1.3 kb fragment of the rrs gene (16S rDNA) was sequenced from a panel of 22 control strains, representing serovars within the pathogenic species Leptospira interrogans, Leptospira borgpetersenii, and Leptospira kirschneri, to identify single nucleotide polymorphisms (SNPs). These were identified in the 5' variable region of the 16S sequence and a 181 bp PCR fragment encompassing this region was used for speciation by Denaturing High Performance Liquid Chromatography (D-HPLC). This method was applied to eleven additional species, representing pathogenic, non-pathogenic and intermediate species and was demonstrated to rapidly differentiate all but 2 of the non-pathogenic Leptospira species. The method was applied successfully to infected tissues from field samples proving its value for diagnosing leptospiral infections found in animals in the UK. Crown Copyright (C) 2010 Published by Elsevier Ltd. All rights reserved.

Phylogenetic analysis of flatfish (order pleuronectiformes) based on mitochondrial 16s rDNA sequences

The phylogenetic relationships of the order Pleuronectiformes are controversial and at some crucial points remain unresolved. To date most phylogenetic studies on this order have been based on morpho-anatomical criteria, whereas only a few sequence comparisons based studies have been reported. In the present study, the phylogenetic relationships of 30 flatfish species pertaining to seven different families were examined by sequence analysis of the first half of the 16S mitochondrial DNA gene. The results obtained did not support percoids as the sister group of pleuronectiforms. The monophyletic origin of most families analyzed, Soleidae, Scophthalmidae, Achiridae, Pleuronectidae and Bothidae, was strongly supported, except for Paralichthyidae which was clearly subdivided into two groups, one of them associated with high confidence to Pleuronectidae. The analysis of the 16S rRNA gene also suggested the monophyly of Pleuronectiforms as the most probable hypothesis and consistently supported some major interfamily groupings.

Table 1 Microorganisms, identified with DOGE analysis. 16S rDNA sequences, obtained from sequencing of DOGE bands, were assigned to the corresponding microorganisms

Microorganisms play an important role in the transformation of material within the earth's crust. The storage of CO2 could affect the composition of inorganic and organic components in the reservoir, consequently influencing microbial activities. To study the microbial induced processes together with geochemical, petrophysical and mineralogical changes, occurring during CO2 storage, long-term laboratory experiments under simulated reservoir P-T conditions were carried out. Clean inner core sections, obtained from the reservoir region at the CO2 storage site in Ketzin (Germany) from a depth of about 650 m, were incubated in high pressure vessels together with sterile synthetic formation brine under in situ P-T conditions of 5.5 MPa and 40°C. A 16S rDNA based fingerprinting method was used to identify the dominant species in DNA extracts of pristine sandstone samples. Members of the alpha- and beta-subdivisions of Proteobacteria and the Actinobacteria were identified. So far sequences belonging to facultative anaerobic, chemoheterotrophic bacteria (Burkholderia fungorum, Agrobacterium tumefaciens) gaining their energy from the oxidation of organic molecules and a genus also capable of chemolithoautotrophic growth (Hydrogenophaga) was identified. During CO2 incubation minor changes in the microbial community composition were observed. The majority of microbes were able to adapt to the changed conditions. During CO2 exposure increased concentrations of Ca**2+, K**+, Mg**2+ and SO4**2- were observed. Partially, concentration rises are (i) due to equilibration between rock pore water and synthetic brine, and (ii) between rock and brine, and are thus independent on CO2 exposure. However, observed concentrations of Ca**2+, K**+, Mg**2+ are even higher than in the original reservoir fluid and therefore indicate mineral dissolution due to CO2 exposure.

Molecular characterization of the microbial species that colonize human ileal and colonic mucosa by using 16S rDNA sequence analysis

Aim: The aim of this study was to characterize the bacterial community adhering to the mucosa of the terminal ileum, and proximal and distal colon of the human digestive tract. Methods and Results: Pinch samples of the terminal ileum, proximal and distal colon were taken from a healthy 35-year-old, and a 68-year-old subject with mild diverticulosis. The 16S rDNA genes were amplified using a low number of PCR cycles, cloned, and sequenced. In total, 361 sequences were obtained comprising 70 operational taxonomic units (OTU), with a calculated coverage of 82.6%. Twenty-three per cent of OTU were common to the terminal ileum, proximal colon and distal colon, but 14% OTU were only found in the terminal ileum, and 43% were only associated with the proximal or distal colon. The most frequently represented clones were from the Clostridium group XIVa (24.7%), and the Bacteroidetes (Cytophaga-Flavobacteria-Bacteroides ) cluster (27.7%). Conclusion: Comparison of 16S rDNA clone libraries of the hindgut across mammalian species confirms that the distribution of phylogenetic groups is similar irrespective of the host species. Lesser site-related differences within groups or clusters of organisms, are probable. Significance and Impact: This study provides further evidence of the distribution of the bacteria on the mucosal surfaces of the human hindgut. Data contribute to the benchmarking of the microbial composition of the human digestive tract.

Phylogenetic analyses of some genera in Oedipodidae (Orthoptera : Acridoidea) based on 16S mitochondrial partial gene sequences

Based on the 16S mitochondrial partial gene sequences of 29 genera, containing 26 from Oedipodidae and one each from Tanaoceridae, Pyrgomorphidae and Tetrigidae (as outgroups), the homologus sequences were compared and phylogenetic analyses were performed. A phylogenetic tree was inferred by neighbor-joining (NJ). The results of sequences compared show that: (i) in a total of 574 bp of Oedipodidae, the number of substituted nucleotides was 265 bp and the average percentages of T, C, A and G were 38.3%, 11.4%, 31.8% and 18.5%, respectively, and the content of A+T (70.1%) was distinctly richer than that of C+G (29.9%); and (ii) the average nucleotide divergence of 16S rDNA sequences among genera of Oedipodidae were 9.0%, among families of Acridoidea were 17.0%, and between superfamilies (Tetrigoidea and Acridoidea) were 23.9%, respectively. The phylogenetic tree indicated: (i) the Oedipodidae was a monophyletic group, which suggested that the taxonomic status of this family was confirmed; (ii) the genus Heteropternis separated from the other Oedipodids first and had another unique sound-producing structure in morphology, which is the type-genus of subfamily Heteropterninae; and (iii) the relative intergeneric relationship within the same continent was closer than that of different continents, and between the Eurasian genera and the African genera, was closer than that between Eurasians and Americans.

Bacterial diversity of soil under eucalyptus assessed by 16S rDNA sequencing analysis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Identification of bacteria in endodontic infections by sequence analysis of 16S rDNA clone libraries

A significant proportion of oral bacteria are unable to undergo cultivation by existing techniques. In this regard, the microbiota from root canals still requires complementary characterization. The present study aimed at the identification of bacteria by sequence analysis of 16S rDNA clone libraries from seven endodontically infected teeth. Samples were collected from the root canals, subjected to the PCR with universal 16S rDNA primers, cloned and partially sequenced. Clones were clustered into groups of closely related sequences (phylotypes) and identification to the species level was performed by comparative analysis with the GenBank, EMBL and DDBJ databases, according to a 98 % minimum identity. All samples were positive for bacteria and the number of phylotypes detected per subject varied from two to 14. The majority of taxa (65(.)2 %) belonged to the phylum Firmicutes of the Gram-positive bacteria, followed by Proteobacteria (10(.)9 %), Spirochaetes (4(.)3 %), Bacteroidetes (6(.)5 %), Actinobacteria (2(.)2 %) and Deferribacteres (2(.)2 %). A total of 46 distinct taxonomic units was identified. Four clones with low similarity to sequences previously deposited in the databases were sequenced to nearly full extent and were classified taxonomically as novel representatives of the order Clostridiales, including a putative novel species of Mogibacterium. The identification of novel phylotypes associated with endodontic infections suggests that the endodontium may still harbour a relevant proportion of uncharacterized taxa.

Identificação de estirpes de rizóbios por seqüenciamento parcial dos genes 16S rDNA e nifH

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Assessment of prey overlap between a native (Polistes humilis) and an introduced (Vespula germanica) social wasp using morphology and phylogenetic analyses of 16S rDNA

In newly invaded communities, interspecific competition is thought to play an important role in determining the success of the invader and its impact on the native community. In southern Australia, the native Polistes humilis was the predominant social wasp prior to the arrival of the exotic Vespula germanica (Hymenoptera: Vespidae). Both species forage for similar resources (water, pulp, carbohydrate and protein prey), and concerns have arisen about potential competition between them. The aim of this study was to identify the protein foods that these wasps feed on. As many prey items are masticated by these wasps to the degree that they cannot be identified using conventional means, morphological identification was complemented by sequencing fragments of the mitochondrial 16S rRNA gene. GenBank searches using blast and phylogenetic analyses were used to identify prey items to at least order level. The results were used to construct complete prey inventories for the two species. These indicate that while P. humilis is restricted to feeding on lepidopteran larvae, V. germanica collects a variety of prey of invertebrate and vertebrate origin. Calculated values of prey overlap between the two species are used to discuss the implications of V. germanica impacting on P. humilis. Results obtained are compared to those gained by solely 'conventional' methods, and the advantages of using DNA-based taxonomy in ecological studies are emphasized.

Molecular characteristics of Camallanus spp. (Spirurida : Camallanidae) in fishes from China based on its rDNA sequences

In the,paper, we explored the intra- and interspecific evolutionary variation among species of Camallanus collected from different fish species in various regions of China. We determined the internal transcribed spacers of ribosomal DNA (ITS rDNA) sequences of these nematodes. The divergence (uncorrected p-distance) of ITS 1, ITS2, and ITS rDNA data sets confirmed 2 valid species of Camallanus in China, i.e., C. cotti and C. hypophthalmichthys. The 2 species were distinguished not only by their different morphologies and host ranges but also by a letranucleotide microsatellite (TTGC)n present in the ITS I region of C cotti. Phylogenetic analyses of the nematodes disclosed 2 main clades, corresponding to different individuals of C cotti and C. hypophthalmichthys from different fish species in various geographical locations, although the interior nodes of each clade received poor support.

Phylogeny of freshwater parasitic copepods in the Ergasilidae (Copepoda : Poecilostomatoida) based on 18S and 28S rDNA sequences

The phylogenetic relationships among the Ergasilidae genera are poorly understood. In this study, 14 species from four genera in the Ergasilidae including Sinergasilus, Ergasilus, Pseudergasilus, and Paraergasilus were collected in China, and their phylogenetic relationships were examined using neighbor-joining, maximum parsimony, maximum likelihood, and Bayesian inference methods based on partial sequences of 18S and 28S ribosomal deoxyribonucleic acid, respectively. All the analyses suggest that the Sinergasilus and Paraergasilus are both monophyletic, but the Ergasilus is polyphyletic rather than monophyletic. Considering the relationships among the four genera, the phylogenetic analyses and subsequent hypothesis tests all suggest that Pseudergasilus clustered with some Ergasilus species may have a closer relationship with Sinergasilus rather than with Paraergasilus. It is proposed that the Sinergasilus and the Pseudergasilus species might have evolved from Ergasilus species.

Phylogeny of Oedogoniales (Chlorophyceae, Chlorophyta) inferred from 18S rDNA sequences with emphasis on the relationships in the genus Oedogonium based on ITS-2 sequences

The phylogeny of Oedogoniales was investigated by using nuclear 18S rDNA sequences. Results showed that the genus Oedocladium, as a separated clade, was clustered within the clade of Oedogonium; whereas the genus Bulbochaete was in a comparatively divergent position to the other two genera. The relationship among the species of Oedogonium was discussed, focusing on ITS-2 phylogeny analyzed combining with some morphological characteristics. Our results showed that all the dioecious nannandrous taxa involved in this study were resolved into one clade, while all the monocious taxa were clustered into another clade as a sister group to the former. The report also suggests that the dioecious macrandrous taxa form a paraphyly and could be more basally situated than the dioecious nannandrous and the monoecious taxa by means of molecular phylogeny and morphotype investigations.

Phylogeny of diplozoids in five genera of the subfamily diplozoinae palombi, 1949 as inferred from ITS-2 rDNA sequences

The phylogenetic relationship of 5 genera, i.e. Diplozoon Nordmann, 1832, Paradiplozoon Achmerov, 1974, Inustiatus Khotenovsky, 1978, Sindiplozoon Khotenovsky, 1981, and Eudiplozoon Khotenovsky, 1985 in the subfamily Diplozoinae Palombi, 1949 (Monogenea, Polyopisthocotylea) was inferred from rDNA ITS-2 region using neighbour-joining (NJ), maximum likelihood (ML) and Bayesian methods. The phylogenetic trees produced by using NJ, ML and Bayesian methods exhibit essentially the same topology. Surprisingly, freshwater species of Paradiplozoon from Europe clustered together with species of Diplozoon, but separated from Chinese Paradiplozoon species. The results of molecular phylogeny and lower level of divergence (4(.)1-15(.)7%) in ITS-2 rDNA among Paradiplozoon from Europe and Diplozoon and, on the other hand, high level of divergence (45(.)3-53(.)7%) among Paradiplozoon species from Europe and China might indicate the non-monophyletic origin of the genus Paradiplozoon. Also, the generic status of European Paradiplozoon needs to be revised. The species of Paradiplozoon in China is a basal group in Diplozoinae as revealed by NJ and Bayesian methods, and Sindiplozoon appears to be closely related to European Paradiplozoon and Diplozoon. with their relationship to Eudiplozoon and Inustiatus being unresolved.

Is the genus Digramma synonymous to the genus Ligula (Cestoda : Pseudophyllidea)? Evidence from ITS and 5 ' end 28S rDNA sequences

The genus Digramma (Cestoda: Pseudophyllidea) described by Cholodkovsky in 1915 differs from the genus Ligula only by the number of the reproductive organs per proglottis. However, the occurrence of transitional forms in Digramma raises much confusion concerning its generic validity. In the present study, cestodes previously designated as Digramma and Ligula were collected from lakes in the lower and middle reaches of the Yangtze River, and also from Qinghai Lake on Qingzang plateau, China. The entire internal transcribed spacer of the ribosomal DNA (ITS rDNA) and 5' end of 28S rDNA were compared between the Digramma and Ligula specimens. The low level of nucleotide variation between the two genera may imply that cestodes in the genus Digramma are paraphyletic to the Ligula genus, and Digramma is a synonym of Ligula. However, whether previously identified Digramma cestodes represent different species in the genus Ligula requires further investigation.