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PubMed1998

Evolutionary relationships among the Braconidae (Hymenoptera: Ichneumonoidea) inferred from partial 16S rDNA gene sequences.

Phylogenetic relationships among the Braconidae were examined using homologous 16S rDNA gene sequence data. Analyses recovered the few well-supported relationships evident in this family from morphological analyses, viz the monophyly of the microgastroid complex of subfamilies, the monophyly of the cyclostome complex of subfamilies (= braconoids), a sister-group relationship between the Alysiinae and Opiinae, and a close relationship between the Helconinae and Blacinae. With respect to the braconoid complex of subfamilies, a sister-group relationship was recovered between Aphidiinae and Mesostoinae, and a clade composed of Gnamptodontinae + Histeromerinae + Rhyssalinae + Aphidiinae + Mesostoinae was also recovered. The Doryctinae and Rogadinae sensu lato (s.l.) were generally not resolved as monophyletic. With respect to the helconoid complex of subfamilies, a sister-group relationship was recovered between Sigalphinae and Agathidinae, whereas Neoneurinae fell out among other helconoid subfamilies. Other relationships among the helconoid subfamilies were unclear from these analyses. With respect to the microgastroid complex of subfamilies, our data conform to morphological estimates, recovering ((Microgastrinae + Miracinae) + Cardiochilinae) + Cheloninae. The topology of our trees suggests that the cyclostome subfamilies are a natural derived group, inferring that endoparasitism (not ectoparasitism) is the ancestral state for the Braconidae, unless all of the ectoparasitic ancestors of the helconoid + microgastroid subfamilies are now extinct.

Animals↗

PubMed1990

Isolate B12, which harbours a virus-like element, represents a new species of the archaebacterial genus Sulfolobus, Sulfolobus shibatae, sp. nov.

The Sulfolobus isolate B12 and its endogenous virus-like element SSV1 have provided a fruitful system for detailed analysis of certain aspects of archaebacterial molecular biology, especially those concerning gene expression. In the course of clarifying this isolate's taxonomic position, we determined DNA base composition, ability to grow autotrophically, nucleotide sequence of 16S ribosomal RNA, and level of total genomic homology to other Sulfolobus strains. Although the results generally demonstrate a similarity to S. solfataricus, DNA-DNA hybridisation and 16S rRNA sequence data indicate that isolate B12 in fact represents a distinct species.

Archaea↗

PubMed2006

Phylogeny of selected Sepiidae (Mollusca, Cephalopoda) based on 12S, 16S, and COI sequences, with comments on the taxonomic reliability of several morphological characters.

Phylogenetic relationships among 11 species of sepiids from Japanese waters and Sepia officinalis from Mediterranean were studied using partial sequences of the mitochondrial 12S rRNA, 16S rRNA, and cytochrome c oxidase subunit I genes. These three genes had been analyzed in an Atlantic species S. elagans and was obtained from database. In the two-gene set analysis (16S+COI), sequence data of another 4 species were added from database. We also studied morphological characters of radulae, tentacular clubs, and cuttlebones. The molecular phylogeny was not congruent with relationships detected by the number of rows in radulae and the arrangement of suckers on the tentacular club. As to the cuttlebone shape, the molecular phylogeny suggests the separation of two groups, Doratosepion species with a lanceolate cuttlebone and the others with a broad cuttlebone. Our molecular phylogenetic study revealed these sepiids are separated into four clades. The first clade includes Sepia officinalis, S. hierrendda, S. bertheloti, S. pharaonis and Sepiella japonica. The second clade consists of S. latimanus and Metasepia tullbergi from sub-tropical waters. The third clade includes Sepia esculenta, S. madokai, S. aculeata and S. lycidas, which have a cuttlebone with a prominent spine. The fourth clade consists of Doratosepion species complex, S. kobiensis, S. lorigera, S. pardex, S. peterseni, and S. sp., which are characterized by a narrow cuttlebone with a distinct outer cone at the posterior end. The lack of membranous structures in the cuttlebone is a synapomorphy for this clade. S. elegans did not clearly belong to any of these clades and might represent the fifth clade.

Animals↗

PubMed2002

Simultaneous detection of Anaplasma marginale and a new Ehrlichia species closely related to Ehrlichia chaffeensis by sequence analyses of 16S ribosomal DNA in Boophilus microplus ticks from Tibet.

To identify ehrlichial agents in Boophilus microplus ticks, DNA samples of B. microplus collected from the Tibet Autonomous Region and Sichuan Province of China were screened by a nested PCR. Sixteen of 43 (37%) DNA samples of B. microplus from Tibet were positive in nested PCR analysis. All 27 samples from Sichuan were negative. The screen identified two ehrlichial agents based on different 16S rRNA genes that were found after amplifying and sequencing the 5'-end fragments of the 16S rRNA genes. One sequence was identical to that of the gene of Anaplasma marginale, an etiological agent of animal anaplasmosis. The other sequence was most similar to that of the gene of Ehrlichia chaffeensis, an etiological agent of human monocytic ehrlichiosis. The sequence of 1,501 bases from the novel ehrlichial agent was obtained and showed the greatest levels of sequence similarity (97 to 98%) to 16S rRNA gene sequences of the members of the E. canis group of the genus EHRLICHIA: Sequence comparison of the 16S rRNA gene with the members of the genus Ehrlichia reveals that the novel ehrlichial agent detected in B. microplus ticks is a new species of the genus Ehrlichia and is most closely related to E. chaffeensis.

Anaplasma↗

PubMed2004

Characterization and molecular methods for detection of a novel spiroplasma pathogenic to Penaeus vannamei.

Traditionally, Spiroplasma spp. have only been isolated from the surfaces of flowers and other plant parts, from the guts and hemolymph of various insects, and from vascular plant fluids (phloem sap) and insects that feed on these fluids. In this article, we report the first pathogenic spiroplasma to be discovered in shrimp and the results of its characterization through histological evaluation, in situ hybridization assays, transmission electron microscopy, 16S rRNA sequence homology, and injection infectivity studies. In addition, molecular methods are described that were developed for the detection of this microorganism, which was determined to be the causative disease agent in Colombian farm-raised Penaeus vannamei suffering from high mortalities. Using standard histological methods and in situ hybridization assays, it was confirmed that P. vannamei was infected with this pathogenic spiroplasma. Histological analysis revealed systemic inflammatory reactions in affected organs/tissues. In an attempt to identify the bacteria, frozen infected P. vannamei samples, from the initial epizootic, were used to sequence the 16S rRNA gene and develop molecular detection methods. The 16S rRNA gene was amplified by PCR and then sequenced. The sequence data were analyzed using the GenBank BLAST search and the results revealed a 98% homology with Spiroplasma citri, a pathogen of citrus trees. The 16S rRNA sequence data were evaluated for development of unique PCR primers to the putative spiroplasma. Using PCR primers developed for the spiralin gene of Spiroplasma spp., a digoxigenin-labeled probe was developed and tested. This probe was species-specific, with no positive reactions or cross-reactivity occurring with other bacterial samples tested in this format.

Animals↗

PubMed2003

Finding the common structure shared by two homologous RNAs.

MOTIVATION: CARNAC is a new method for pairwise folding of RNA sequences. The program takes into account local similarity, stem energy, and covariations to produce the common folding. It can handle all RNA types, and has also been adapted to align a new homologous sequence along a reference structured sequence. RESULTS: Using different data sets, we show that CARNAC provides a good partial prediction for a wide range of sequences (16S ssu rRNA, RNase P RNA, viruses) with only two sequences. In presence of a whole family of sequences, we also show that CARNAC can be used to detect whether the sequences actually share the same structure. AVAILABILITY: CARNAC is available at the URLhttp://www.lifl.fr/~perrique/rna/.

Algorithms↗

PubMed2001

Assessment of Changes in Biodiversity when a Community of Ultramicrobacteria Isolated from Groundwater Is Stimulated to Form a Biofilm.

The stimulation of groundwater bacteria to form biofilms, for the remediation of polluted aquifers, is subjected to environmental regulations that include measurement of effects on microbial biodiversity. Groundwater microorganisms contain a proportion of unidentified and uncharacterized ultramicrobacteria (UMB) that might play a major role in the bioclogging of geological materials. This study aimed to assess the changes in genetic and metabolic biodiversity when a community of UMB, isolated from groundwater, is stimulated to form biofilms on a ceramic surface. UMB were stimulated with aerobic conditions and injection of molasses, in reactors reproducing groundwater composition and temperature. Concentration of planktonic viable UMB, secretion of extracellular polymeric substances (EPS), and biofilm thickness were monitored. The assessment of changes in biodiversity was achieved by comparing the initial UMB community to the biofilm community, using the single strand conformational polymorphism (SSCP) method, the cloning and sequencing of 16S rRNA gene (16S rDNA) sequences, and the Biolog microplate system. The hypothesis stating that indigenous UMB would play a significant role of in the biofilm development was corroborated. Within 13 days of stimulation, the UMB produced 700 mg L?1 of planktonic EPS and formed a biofilm up to a thickness of 1100 mm. This stimulation led to a decrease in genetic diversity and an increase in metabolic diversity. The decrease in genetic diversity was shown by a reduced number of single strand DNA fragments in the SSCP profiles. As such, 16S rDNA sequences from the biofilm revealed the predominance of four bacterial groups: Zoogloea, Bacillus/Paenibacillus, Enterobacteriaceae, and Pseudomonads. A significant increase in metabolic diversity was shown by a highest substrate richness profile and a lower substrate evenness profile of the biofilm bacterial population (p = 0.0 and p = 0.09, respectively). This higher metabolic diversity might be a consequence of the stimulation that seemed to favor the growth of bacteria having a high nutritional versatility. Stimulation of UMB, isolated from groundwater, was effective to form a biofilm having a high metabolic biodiversity. This combination of molecular-based and metabolic-based methods expanded the insight into monitoring the changes in bacterial biodiversity.

Journal Article↗

PubMed1997

Phylogenetic analysis of the genus Chlamydia based on 16S rRNA gene sequences.

The phylogenetic relationships among Chlamydia spp. were investigated by comparing 16S rRNA gene sequences. In this analysis we used 14 strains of Chlamydia psittaci, including seven feline isolates, two avian isolates, two human isolates, one bovine isolates, one ovine isolate, and one koala isolate; and nine strains of Chlamydia trachomatis, including six human isolates, two swine isolates, and one mouse isolate. A phylogenetic analysis of the 16S rRNA gene sequences of these organisms and seven previously published sequences revealed eight genetic groups which formed two clusters. The first cluster was composed of C. pecorum, Chlamydia pneumoniae, and C. psittaci and included three genetic groups (one group containing avian, human, and ovine strains, one group containing feline strains, and one group containing guinea pig strains). The second cluster was composed of C. trachomatis and also included three genetic groups (one group containing human strains, one group containing swine isolates, and one group containing rodent strains). The strains in each genetic group exhibited similar genetic distances. The results of the phylogenetic analysis agreed with the results of previous genomic DNA, ompA gene allele, and biotyping studies. Therefore, the genetic groups based on genetic distances may be considered a criterion for species identification.

Animals↗

PubMed1998

A qualitative evaluation of the published oligonucleotides specific for the 16S rRNA gene sequences of the ammonia-oxidizing bacteria.

Over the past few years, there has been an increasing interest in making oligonucleotides specific for ammonia-oxidizing bacteria (AOB), in order to detect and monitor these slow growing bacteria in environmental samples, in enrichment cultures and in wastewater treatment plants. Based on 16S rDNA sequences, a broad selection of oligonucleotides have been designed, either encompassing all known AOB in the beta-subgroup of the Proteobacteria (beta AOB), or subclasses within beta AOB. Thirty different oligonucleotides have so far been published, with varying specificity. The first AOB-specific oligonucleotides published were obtained as a result of an alignment of only eleven 16S rDNA sequences from AOB. Including the present study, there are now forty nearly full length 16S rDNA sequences available from these bacteria, in addition to a number of partial sequences, so that an improved evaluation of the published oligonucleotides can be done. Two new 16S rRNA gene sequences from Nitrosospira are presented here, in a phylogenetic analysis containing every 16S rRNA gene sequences (> 1 kb) available from AOB. On the basis of an alignment of all these sequences, combined with searches in the nucleotide sequence databases, an evaluation of the thirty published oligonucleotides is presented. The analysis expose the strength and weakness of each oligonucleotide and discuss the use of oligonucleotides specific for 16S rRNA genes in future studies of AOB. The present work also identifies one new, broad range primer, specific for the AOB in the beta-subgroup of the Proteobacteria.

Ammonia↗

PubMed2004

Genetic and morphological data supporting the hypothesis of adaptive radiation in the endemic fish of Lake Matano.

Adaptive radiation resulting from differential selection acting on functional features is believed to be an important source of biodiversity. In this study, morphometric measures and mitochondrial DNA are used to test for adaptive radiation within four fish genera (Glossogobius, Oryzias, Dermogenys and Telmatherina) endemic to an ancient island lake (Lake Matano, Sulawesi, Indonesia), using the framework proposed by Schluter (The Ecology of Adaptive Radiation, Oxford University Press, 2000). We demonstrate common ancestry and rapid divergence in one genus (Telmatherina) based on 560 bp of 16S sequence data. We found higher levels of variation in feeding-related traits (N = 8) for Telmatherina relative to the other genera, while no differences were found for sexual display traits (N = 8) or neutral morphological traits (N = 8). Telmatherina also had the highest number of distinct colouration patterns among the four genera. These data, combined with the very low productivity of the lake, are indicative of selection driving adaptive radiation. The morphometric divergence in the Telmatherina likely results from selection acting on feeding traits in this low productivity lake, leading to trophic specialization among closely related morphotypes. These results provide indirect but compelling data supporting the adaptive radiation of Telmatherina in this system.

Adaptation, Biological↗

PubMed2003

Evaluation of broiler litter with reference to the microbial composition as assessed by using 16S rRNA and functional gene markers.

Very little is known about the microbial composition of animal bedding wastes, including poultry litter, and what is known has been deduced from standard culture methods, by which some fastidious organisms that exist in the environment may not be detected. We evaluated the bacterial composition of poultry litter by using a combination of culture and molecular detection. Total aerobic bacteria in poultry litter were detected by culture at 10(9) CFU/g of material. Enteric bacteria such as Enterococcus spp. and coliforms composed 0.1 and 0.01%, respectively, of the total aerobic cultivatable bacteria in poultry litter; no Salmonella strains were detected by culture. In order to characterize the most abundant bacterial groups, we sequenced 16S ribosomal DNA (rDNA) genes amplified by PCR with microbial community DNA isolated from poultry litter as the template. From the 16S rDNA library, 31 genera were identified. Twelve families or groups were identified with lactobacilli and Salinococcus spp. forming the most abundant groups. In fact, 82% of the total sequences were identified as gram-positive bacteria with 62% of total belonging to low G+C gram-positive groups. In addition to detection of 16S rDNA sequences associated with the expected fecal bacteria present in manure, we detected many bacterial sequences for organisms, such as Globicatella sulfidofaciens, Corynebacterium ammoniagenes, Corynebacterium urealyticum, Clostridium aminovalericum, Arthrobacter sp., and Denitrobacter permanens, that may be involved in the degradation of wood and cycling of nitrogen and sulfur. Several sequences were identified in the library for bacteria associated with disease in humans and poultry such as clostridia, staphylococci, and Bordetella spp. However, specific PCR targeting other human and veterinary pathogens did not detect the presence of Salmonella, pathogenic Escherichia coli, Campylobacter spp., Yersinia spp., Listeria spp., or toxigenic staphylococci. PCR and DNA hybridization revealed the presence of class 1 integrons with gene cassettes that specify resistance to aminoglycosides and chloramphenicol. Only from understanding the microbial community of animal wastes such as poultry litter can we manage animal disease and limit the impact of animal waste on the environment and human and animal health.

Animals↗

PubMed2003

Short report: variation in mitochondrial 12S and 16S ribosomal DNA sequences in natural populations of Triatoma infestans (Hemiptera: Reduviidae).

Mitochondrial DNA sequences of the 12S and 16S ribosomal RNA genes were analyzed in five natural populations of the Chagas' disease vector Triatoma infestans from Argentina. DNA sequence comparisons of 878 basepairs (12S plus 16S) revealed 13 haplotypes. A total of 10 private haplotypes were found in four of the populations analyzed, suggesting low current levels of genetic exchange. The levels of genetic differentiation between the population of Chancaní (Córdoba) and other two of the populations analyzed indicated significant deviation from a pattern of unrestricted gene flow. The haplotypic diversity and the private haplotypes found in the geographically closest localities of Chancaní and El Jardín (La Rioja) suggest that the reduction in the population size by insecticide treatment did not avoid the recovery of the populations apparently from survivors of the same area.

Animals↗

PubMed1999

Comparative diversity of ammonia oxidizer 16S rRNA gene sequences in native, tilled, and successional soils.

Autotrophic ammonia oxidizer (AAO) populations in soils from native, tilled, and successional treatments at the Kellogg Biological Station Long-Term Ecological Research site in southwestern Michigan were compared to assess effects of disturbance on these bacteria. N fertilization effects on AAO populations were also evaluated with soils from fertilized microplots within the successional treatments. Population structures were characterized by PCR amplification of microbial community DNA with group-specific 16S rRNA gene (rDNA) primers, cloning of PCR products and clone hybridizations with group-specific probes, phylogenetic analysis of partial 16S rDNA sequences, and denaturing gradient gel electrophoresis (DGGE) analysis. Population sizes were estimated by using most-probable-number (MPN) media containing varied concentrations of ammonium sulfate. Tilled soils contained higher numbers than did native soils of culturable AAOs that were less sensitive to different ammonium concentrations in MPN media. Compared to sequences from native soils, partial 16S rDNA sequences from tilled soils were less diverse and grouped exclusively within Nitrosospira cluster 3. Native soils yielded sequences representing three different AAO clusters. Probes for Nitrosospira cluster 3 hybridized with DGGE blots from tilled and fertilized successional soils but not with blots from native or unfertilized successional soils. Hybridization results thus suggested a positive association between the Nitrosospira cluster 3 subgroup and soils amended with inorganic N. DGGE patterns for soils sampled from replicated plots of each treatment were nearly identical for tilled and native soils in both sampling years, indicating spatial and temporal reproducibility based on treatment.

Ammonia↗

PubMed1991

Intrageneric structure of Streptococcus based on comparative analysis of small-subunit rRNA sequences.

The partial 16S rRNA sequences of 24 Streptococcus species were determined by reverse transcription. A comparative analysis of these sequences and the sequences of seven previously studied streptococcal species revealed the presence of several clusters within the genus. The clusters obtained from the sequence analysis agreed in general with the groups outlined on the basis of the results of nucleic acid hybridization studies, but there were some exceptions. The pyogenic group was extended to include Streptococcus agalactiae, S. parauberis, S. porcinus, and S. uberis. Four oral groups were discerned; these four groups centered on S. mutans, S. salivarius, S. anginosus, and S. oralis. Some species (e.g., S. suis and S. acidominimus) did not cluster with any particular group. Our findings are discussed in the context of data from other genetic and chemotaxonomic studies.

Base Sequence↗

PubMed1994

16S ribosomal DNA sequence identities of beta-proteobacterial endosymbionts in three Crithidia species.

The 16S ribosomal DNA sequences of endosymbionts from the trypanosomatid protozoa (Crithidia spp.) are most homologous to that of Bordetella spp. This finding extends the polyphyletic origin of endosymbionts for the first time to the beta Proteobacteria. Biased base transitions and compensatory mutations of the symbionts' sequences that may contribute to their identity in the three Crithidia spp. are noted.

Animals↗

PubMed2001

Microvariation artifacts introduced by PCR and cloning of closely related 16S rRNA gene sequences.

A defined template mixture of seven closely related 16S-rDNA clones was used in a PCR-cloning experiment to assess and track sources of artifactual sequence variation in 16S rDNA clone libraries. At least 14% of the recovered clones contained aberrations. Artifact sources were polymerase errors, a mutational hot spot, and cloning of heteroduplexes and chimeras. These data may partially explain the high degree of microheterogeneity typical of sequence clusters detected in environmental clone libraries.

Artifacts↗

PubMed1993

Restriction fragment length polymorphism analysis of Fusobacterium necrophorum using a novel repeat DNA sequence and a 16S rRNA gene probe.

A repeated DNA sequence was isolated from Fusobacterium necrophorum biotype AB, strain FnS1. The repeated sequence shared considerable homology with the transposase gene from the Pseudomonas syringiae insertion sequence IS801. The repeat sequence was used together with a 16S ribosomal RNA gene probe to type F. necrophorum isolates using restriction fragment length polymorphisms. The probes revealed differences between several clinical isolates and will be useful tools to study the epidemiology of ovine foot abscess and other diseases caused by F. necrophorum.

Amino Acid Sequence↗

PubMed1999

Detection of seven major evolutionary lineages in cyanobacteria based on the 16S rRNA gene sequence analysis with new sequences of five marine Synechococcus strains.

Although molecular phylogenetic studies of cyanobacteria on the basis of the 16S rRNA gene sequence have been reported, the topologies were unstable, especially in the inner branchings. Our analysis of 16S rRNA gene phylogeny by the maximum-likelihood and neighbor-joining methods combined with rate homogeneous and heterogeneous models revealed seven major evolutionary lineages of the cyanobacteria, including prochlorophycean organisms. These seven lineages are always stable on any combination of these methods and models, fundamentally corresponding to phylogenetic relationships based on other genes, e.g., psbA, rbcL, rnpB, rpoC, and tufA. Moreover, although known genotypic and phenotypic characters sometimes appear paralleled in independent lineages, many characters are not contradictory within each group. Therefore we propose seven evolutionary groups as a working hypothesis for successive taxonomic reconstruction. New 16S rRNA sequences of five unicellular cyanobacterial strains, PCC 7001, PCC 7003, PCC 73109, PCC 7117, and PCC 7335 of Synechococcus sp., were determined in this study. Although all these strains have been assigned to "marine clusters B and C," they were separated into three lineages. This suggests that the organisms classified in the genus Synechococcus evolved diversely and should be reclassified in several independent taxonomic units. Moreover, Synechococcus strains and filamentous cyanobacteria make a monophyletic group supported by a comparatively high statistical confidence value (80 to 100%) in each of the two independent lineages; therefore, these monophylies probably reflect the convergent evolution of a multicellular organization.

Biological Evolution↗