scholarly journals Comparative Diversity of Ammonia Oxidizer 16S rRNA Gene Sequences in Native, Tilled, and Successional Soils

1999 ◽  
Vol 65 (7) ◽  
pp. 2994-3000 ◽  
Author(s):  
Mary Ann Bruns ◽  
John R. Stephen ◽  
George A. Kowalchuk ◽  
James I. Prosser ◽  
Eldor A. Paul
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Ammonia Oxidizer ◽  
Most Probable Number ◽  
Rrna Gene ◽  
Probable Number ◽  
Rdna Sequences ◽  
16S Rdna Sequences ◽  
Native Soils

ABSTRACT 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 forNitrosospira 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 Nitrosospiracluster 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.

scholarly journals Phenotypic characterization of Astragalus glycyphyllos symbionts and their phylogeny based on the 16S rDNA sequences and RFLP of 16S rRNA gene

2014 ◽  
Vol 105 (6) ◽  
pp. 1033-1048 ◽  
Author(s):  
Sebastian Gnat ◽  
Magdalena Wójcik ◽  
Sylwia Wdowiak-Wróbel ◽  
Michał Kalita ◽  
Aneta Ptaszyńska ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Phenotypic Characterization ◽  
Rrna Gene ◽  
Rdna Sequences ◽  
16S Rdna Sequences ◽  
Astragalus Glycyphyllos Symbionts

scholarly journals Clover Proliferation Group (16SrVI) Subgroup A (16SrVI-A) Phytoplasma is a Probable Causal Agent of Dry Bean Phyllody Disease in Washington

Plant Disease ◽  
2004 ◽  
Vol 88 (4) ◽  
pp. 429-429
Author(s):  
I.-M. Lee ◽  
K. D. Bottner ◽  
P. N. Miklas ◽  
M. A. Pastor-Corrales
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Sequence Homology ◽  
Rrna Gene ◽  
16S Rrna Gene Sequences ◽  
Dry Bean ◽  
Rdna Sequences ◽  
Bean Plants ◽  
16S Rdna Sequences

During 2003, a new disease, dry bean phyllody (DBPh), was observed in the Columbia Basin of Washington in dry bean (Phaseolus vulgaris L.) cultivars of Andean origin grown in Mattawa and Paterson, WA that caused great reduction in dry bean production. Symptoms of DBPh became apparent during mid-to-late pod development and were characterized by leafy petals (phyllody) and aborted seed pods resembling thin, twisted, and corrugated leaf-like structures. Deformed sterile pods that were small, sickle-shaped, upright, and leathery were also observed. The infected plants generally exhibited chlorosis, stunting, or bud proliferation from leaf axils. Symptoms of DBPh were indicative of possible infection by phytoplasmas. Restriction fragment length polymorphism (RFLP) and phylogenetic analyses of amplified 16S rDNA sequences were used for phytoplasma identification. Four symptomatic bean plants were analyzed and tested positive for phytoplasma infection on the basis of results of initial polymerase chain reaction (PCR) and subsequent nested-PCR amplifications (2). RFLP analyses of 16S rDNA sequences with restriction enzymes, MseI, AluI, HhaI, RsaI, and HpaII indicated that the phytoplasma strains associated with DBPh belonged to the clover proliferation group (16SrVI) subgroup A (16SrVI-A) (2). This subgroup currently consists of three members, clover proliferation (CP; GenBank Accession No. AY500130), potato witches'-broom (PWB; GenBank Accession No. AY500818), and vinca virescence (VR; GenBank Accession No. AY500817), a strain of beet leafhopper-transmitted virescence agent (BLTVA) phytoplasmas (1,2). The taxonomic affiliations of the DBPh phytoplasma strains were confirmed by phylogenetic analysis of cloned 16S rRNA gene sequences (GenBank Accession Nos. DBPh2, AY496002; DBPh3, AY496003). Among the existing members of subgroup 16SrVI-A, the four DBPh strains were closely related to the VR strain with 99.7% 16S rDNA sequence homology and to the CP strain with 99.2% sequence homology. To gain further evidence on the role of 16SrVI-A phytoplasma strains in DBPh disease, a modified test of Koch's postulates was conducted. Infected tissue from one phytoplasma-positive dry bean sample was grafted onto three Pinto UI-114 bean seedlings in the greenhouse. Within 60 days, the bean seedlings exhibited corrugated leaf-like structures from aborted seedpods, a lack of flower formation, general chlorosis, and stunting similar to the original diseased plants. The lower leaves of the inoculated bean plants became epinastic and leathery. The transmitted phytoplasma was detected in each of the grafted symptomatic seedlings, and the RFLP patterns of its 16S rRNA gene sequences were identical to those of the phytoplasmas in the scions. A high correlation between the presence of disease symptoms and the presence of subgroup 16SrVI-A phytoplasmas in the bean plants suggests that these phytoplasmas play an etiological role in DBPh disease. To our knowledge, these findings provide the first confirmed case of phytoplasma-associated DBPh in the United States. References: (1) D. A. Golino et al. Plant Dis. 73:850, 1989. (2) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998.

Characterization and Identification of Numerically Abundant Culturable Bacteria from the Anoxic Bulk Soil of Rice Paddy Microcosms

1999 ◽  
Vol 65 (11) ◽  
pp. 5042-5049 ◽  
Author(s):  
Kuk-Jeong Chin ◽  
Dittmar Hahn ◽  
Ulf Hengstmann ◽  
Werner Liesack ◽  
Peter H. Janssen
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Dry Weight ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Most Probable Number ◽  
Rrna Gene ◽  
16S Rrna Gene Sequences ◽  
Probable Number ◽  
Clostridial Cluster

ABSTRACT Most-probable-number (liquid serial dilution culture) counts were obtained for polysaccharolytic and saccharolytic fermenting bacteria in the anoxic bulk soil of flooded microcosms containing rice plants. The highest viable counts (up to 2.5 × 108 cells per g [dry weight] of soil) were obtained by using xylan, pectin, or a mixture of seven mono- and disaccharides as the growth substrate. The total cell count for the soil, as determined by using 4′,6-diamidino-2-phenylindole staining, was 4.8 × 108cells per g (dry weight) of soil. The nine strains isolated from the terminal positive tubes in counting experiments which yielded culturable populations that were equivalent to about 5% or more of the total microscopic count population belonged to the divisionVerrucomicrobia, theCytophaga-Flavobacterium-Bacteroides division, clostridial cluster XIVa, clostridial cluster IX, Bacillus spp., and the class Actinobacteria. Isolates originating from the terminal positive tubes of liquid dilution series can be expected to be representatives of species whose populations in the soil are large. None of the isolates had 16S rRNA gene sequences identical to 16S rRNA gene sequences of previously described species for which data are available. Eight of the nine strains isolated fermented sugars to acetate and propionate (and some also fermented sugars to succinate). The closest relatives of these strains (except for the two strains of actinobacteria) were as-yet-uncultivated bacteria detected in the same soil sample by cloning PCR-amplified 16S rRNA genes (U. Hengstmann, K.-J. Chin, P. H. Janssen, and W. Liesack, Appl. Environ. Microbiol. 65:5050–5058, 1999). Twelve other isolates, which originated from most-probable-number counting series indicating that the culturable populations were smaller, were less closely related to cloned 16S rRNA genes.

scholarly journals Specific Detection, Isolation, and Characterization of Selected, Previously Uncultured Members of the Freshwater Bacterioplankton Community

2005 ◽  
Vol 71 (10) ◽  
pp. 5908-5919 ◽  
Author(s):  
Frederic Gich ◽  
Karin Schubert ◽  
Alke Bruns ◽  
Herbert Hoffelner ◽  
Jörg Overmann
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Most Probable Number ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Light Harvesting Complexes ◽  
Probable Number ◽  
Bacterioplankton Community ◽  
Isolation And Characterization

ABSTRACT High-throughput cultivation was combined with rapid and group-specific phylogenetic fingerprinting in order to recover representatives of three freshwater bacterioplankton communities. A total of 570 bacterial cultures were obtained by employing the most probable number and MicroDrop techniques. The majority of the cultured bacteria were closely related to previously uncultured bacteria and grouped with the α-Proteobacteria, β-Proteobacteria, Actinobacteria, Firmicutes, or Flavobacteria-Cytophaga lineage. Correspondingly, the natural bacterioplankton community was analyzed by high-resolution phylogenetic fingerprinting of these five bacterial lineages. 16S rRNA gene fragments were generated for each lineage and subsequently separated by denaturing gradient gel electrophoresis. By the combination of five group-specific PCR protocols, the total number of 16S rRNA gene fingerprints generated from the natural communities was increased sixfold compared to conventional (eubacterial) fingerprinting. Four of the environmental α-Proteobacteria 16S rRNA gene sequences obtained from the natural community were found to be identical to those of bacterial isolates. One of these phylotypes was detected in 14 different cultures and hence represented the most frequently cultured bacterium. Three of these 14 strains were characterized in detail. Their complete 16S rRNA gene sequences showed only 93% similarity to that of Sandaracinobacter sibiricus, the closest relative described so far. The novel phylotype of bacterium is a strict aerobe capable of using numerous organic carbon substrates and contains bacteriochlorophyll a bound to two different photosynthetic light-harvesting complexes. Dot blot hybridization revealed that the strains occur in lakes of different trophic status and constitute up to 2% of the microbial community.

scholarly journals Cryptic haplotypes of "Candidatus Liberibacter africanus"

2015 ◽  
Author(s):  
Warrick Nelson ◽  
Sandrine Eveillard ◽  
Marie-Pierre Dubrana ◽  
Joseph Bové
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Rdna Sequences ◽  
New Information ◽  
Candidatus Liberibacter ◽  
Citrus Leaves

“Candidatus Liberibacter africanus” (Laf) has long been recognised as a causal agent of the devastating citrus disease huanglongbing (HLB) or citrus greening. This species is currently restricted to Africa, the Arabian Peninsula and some Indian Ocean islands and vectored by the African citrus psyllid, Trioza erytreae. Blotchy mottle on citrus leaves is characteristic of the disease. Somewhat similar symptoms in the Rutaceous tree Calodendrum capensis (Cape Chestnut) resulted in the discovery of Laf outside commercial citrus crops in South Africa. This was classed as a subspecies of Laf (capensis, hence LafC). In subsequent surveys of both commercial citrus crops and Calodendrum, both natural and ornamental specimens, LafC was not found in the citrus crop, nor has Laf been found in C. capensis. HLB was reported from Madagascar in 1968 but no sequences from this source have so far been published. Until fairly recently, only the reference 16S rRNA gene sequences of Laf (L22533) and LafC (AF137368) had been deposited in GenBank. Both of these reference sequences contain a number of unresolved nucleotides. Resolving these nucleotide positions by aligning against more recently available sequences, it becomes evident that these unresolved positions represent one percentage point difference in similarity between Laf and LafC. The originally reported 97.4% similarity is therefore incorrect based on this new information. Recalculating the similarity on the full length 16S rDNA sequence results in 99.54% similarity, a value too high to justify a subspecies status. LafC should therefore be reduced to that of a haplotype of Laf. Further, the six 16S rRNA gene sequences currently available in GenBank identified as the species Laf separate into 2 haplotype groups. The 3 haplotypes of Laf are therefore LafA designated as the first accession sequenced (L22533), LafC for the former capensis subspecies and to recognise the prior use of this term, and LafB for the third haplotype not previously recognised. Thus the cryptic presence of 3 haplotypes is revealed by this review of the Laf 16S rDNA sequences.

The Diversity of Benthic Microorganisms in Acidic Mine Lake Sediments

2007 ◽  
Vol 20-21 ◽  
pp. 489-492
Author(s):  
Huynh A. Pham ◽  
Carolyn E. Oldham ◽  
Jason J. Plumb
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Microbial Communities ◽  
Lake Sediments ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Most Probable Number ◽  
Rrna Gene ◽  
Probable Number ◽  
Benthic Bacteria ◽  
Different Depths

The sediment microbial communities of a disused coal mine lake, Lake Kepwari (pH~4.5-5) were studied to understand how the natural microbial processes in an oligotrophic acidic mine lake system influence the iron and sulphur cycles. Most probable number (MPN) viable counts were used to enumerate the benthic bacteria at different depths. MPN results revealed an abundance of bacteria that were capable of growing in sulphate reducing medium with numbers in the range of 1 × 107 – 1 × 108 cells.g-1 of wet sediment. In contrast, MPN results showed much lower numbers of bacteria that were capable of growing in ferric reducing medium with 1 × 102 – 2 × 103 cells.g-1 of wet sediment detected. Serial decimal dilution cultures were used to isolate pure strains of benthic bacteria. Strains HP1, HP2 and HP3 were isolated from benthic lake sediments at 18 m, 0 m and 10 m water depths respectively. 16S rRNA gene sequence analysis of strain HP1 showed that the strain belonged to the genus Enterobacter, strain HP2 belonged to the Order Rhizobiales and strain HP3 belonged to the sub-order Micrococcineae. Denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene fragments was used to profile the diversity of the benthic microbial communities at different depths. DGGE profiling of benthic sediments revealed that sediments contained mostly members of the Proteobacteria, Actinobacteria and Firmicutes phyla.

scholarly journals Anaerobic Mineralization of Quaternary Carbon Atoms: Isolation of Denitrifying Bacteria on Dimethylmalonate

1999 ◽  
Vol 65 (8) ◽  
pp. 3319-3324 ◽  
Author(s):  
Olaf Kniemeyer ◽  
Christina Probian ◽  
Ramon Rosselló-Mora ◽  
Jens Harder
Keyword(s):  
Sewage Sludge ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Most Probable Number ◽  
Rrna Gene ◽  
Quaternary Carbon ◽  
Probable Number ◽  
Gene Similarity

ABSTRACT The microbial capacity to degrade simple organic compounds with quaternary carbon atoms was demonstrated by enrichment and isolation of five denitrifying strains on dimethylmalonate as the sole electron donor and carbon source. Quantitative growth experiments showed a complete mineralization of dimethylmalonate. According to phylogenetic analysis of the complete 16S rRNA genes, two strains isolated from activated sewage sludge were related to the genusParacoccus within the α-Proteobacteria (98.0 and 98.2% 16S rRNA gene similarity to Paracoccus denitrificans T), and three strains isolated from freshwater ditches were affiliated with the β-Proteobacteria (97.4 and 98.3% 16S rRNA gene similarity to Herbaspirillum seropedicae T andAcidovorax facilis T, respectively). Most-probable-number determinations for denitrifying populations in sewage sludge yielded 4.6 × 104dimethylmalonate-utilizing cells ml−1, representing up to 0.4% of the total culturable nitrate-reducing population.

scholarly journals Annual Variations in the Diversity, Viability, and Origin of Airborne Bacteria

2010 ◽  
Vol 76 (9) ◽  
pp. 3015-3025 ◽  
Author(s):  
Camilla Fahlgren ◽  
Åke Hagström ◽  
Douglas Nilsson ◽  
Ulla Li Zweifel
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Seasonal Pattern ◽  
Most Probable Number ◽  
Rrna Gene ◽  
Clone Libraries ◽  
Probable Number ◽  
Annual Variations ◽  
Annual Survey ◽  
Cultured Bacteria

ABSTRACT The presence of bacteria in aerosols has been known for centuries, but information on their identity and role in dispersing microbial traits is still limited. This study monitored the airborne bacterial community during an annual survey using samples collected from a 25-m tower near the Baltic Sea coast. The number of CFU was estimated using agar plates while the most probable number (MPN) of viable bacteria was estimated using dilution-to-extinction culturing assays (DCAs). The MPN and CFU values produced quantitatively similar results, displaying a pronounced seasonal pattern, with the highest numbers in winter. The most dominant bacteria growing in the DCAs all formed colonies on agar plates, were mostly pigmented (80%), and closely resembled (>97%) previously cultured bacteria based on their 16S rRNA gene sequences. 16S rRNA gene clone libraries were constructed on eight occasions during the survey; these revealed a highly diverse community with a few abundant operational taxonomic units (OTUs) and a long tail of rare OTUs. A majority of the cloned sequences (60%) were also most closely related to previously “cultured” bacteria. Thus, both culture-dependent and culture-independent techniques indicated that bacteria able to form colonies on agar plates predominate in the atmosphere. Both the DCAs and clone libraries indicated the dominance of bacteria belonging to the genera Sphingomonas sp. and Pseudomonas sp. on several sampling occasions. Potentially pathogenic strains as well as sequences closely resembling bacteria known to act as ice nuclei were found using both approaches. The origin of the sampled air mass was estimated using backward trajectories, indicating a predominant marine source.

scholarly journals Phylogenetic Analysis of Nonthermophilic Members of the Kingdom Crenarchaeota and Their Diversity and Abundance in Soils

1998 ◽  
Vol 64 (11) ◽  
pp. 4333-4339 ◽  
Author(s):  
Daniel H. Buckley ◽  
Joseph R. Graber ◽  
Thomas M. Schmidt
Keyword(s):  
16S Rrna ◽  
16S Rdna ◽  
Phylogenetic Analyses ◽  
Pcr Primers ◽  
Soil Samples ◽  
Molecular Techniques ◽  
Rrna Gene ◽  
Rdna Sequences ◽  
16S Rdna Sequences ◽  
Long Term Ecological Research

ABSTRACT Within the last several years, molecular techniques have uncovered numerous 16S rRNA gene (rDNA) sequences which represent a unique and globally distributed lineage of the kingdom Crenarchaeotathat is phylogenetically distinct from currently characterized crenarchaeotal species. rDNA sequences of members of this novel crenarchaeotal group have been recovered from low- to moderate-temperature environments (−1.5 to 32°C), in contrast to the high-temperature environments (temperature, >80°C) required for growth of the currently recognized crenarchaeotal species. We determined the diversity and abundance of the nonthermophilic members of the Crenarchaeota in soil samples taken from cultivated and uncultivated fields located at the Kellogg Biological Station’s Long-Term Ecological Research site (Hickory Corners, Mich.). Clones were generated from 16S rDNA that was amplified by using broad-specificity archaeal PCR primers. Twelve crenarchaeotal sequences were identified, and the phylogenetic relationships between these sequences and previously described crenarchaeotal 16S rDNA sequences were determined. Phylogenetic analyses included nonthermophilic crenarchaeotal sequences found in public databases and revealed that the nonthermophilic Crenarchaeota group is composed of at least four distinct phylogenetic clusters. A 16S rRNA-targeted oligonucleotide probe specific for all known nonthermophilic crenarchaeotal sequences was designed and used to determine their abundance in soil samples. The nonthermophilicCrenarchaeota accounted for as much as 1.42% ± 0.42% of the 16S rRNA in the soils analyzed.

scholarly journals Ecological Significance of Microdiversity: Identical 16S rRNA Gene Sequences Can Be Found in Bacteria with Highly Divergent Genomes and Ecophysiologies

2004 ◽  
Vol 70 (8) ◽  
pp. 4831-4839 ◽  
Author(s):  
Elke Jaspers ◽  
Jörg Overmann
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Niche Overlap ◽  
Most Probable Number ◽  
Ecological Niches ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Probable Number ◽  
Internally Transcribed Spacer

ABSTRACT A combination of cultivation-based methods with a molecular biological approach was used to investigate whether planktonic bacteria with identical 16S rRNA gene sequences can represent distinct eco- and genotypes. A set of 11 strains of Brevundimonas alba were isolated from a bacterial freshwater community by conventional plating or by using a liquid most-probable-number (MPN) dilution series. These strains had identical 16S rRNA gene sequences and represented the dominant phylotype in the plateable fraction, as well as in the highest positive dilutions of the MPN series. However, internally transcribed spacer and enterobacterial repetitive intergenic consensus PCR fingerprinting analyses, as well as DNA-DNA hybridization analyses, revealed great genetic diversity among the 11 strains. Each strain utilized a specific combination of 59 carbon substrates, and the niche overlap indices were low, suggesting that each strain occupied a different ecological niche. In dialysis cultures incubated in situ, each strain had a different growth rate and cell yield. We thus demonstrated that the B. alba strains represent distinct populations with genetically determined adaptations and probably occupy different ecological niches. Our results have implications for assessment of the diversity and biogeography of bacteria and increase the perception of natural diversity beyond the level of 16S rRNA gene sequences.

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