Utility of 16S–23S rRNA spacer region methodology: how similar are interspace regions within a genome and between strains for closely related organisms?

Nagpal, Madan L.; Fox, Kevin M.; Fox, Alvin

doi:10.1016/s0167-7012(98)00054-2

Cited by 67 publications

(45 citation statements)

References 21 publications

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“…Cloning of the 16S rRNA gene and ITS region has allowed phylogenetic information to be attached to ARISA profiles, enabling tracking of specific species throughout samples. Interoperonic differences in spacer length occur within the genomes of microorganisms (Nagpal et al, 1998), such that a single species may contribute more than one peak to an ARISA profile. Previous studies (for example, Gugger et al, 2002;SA Wood, A Rueckert, SC Cary, unpublished) indicate that species of the order Nostocales commonly have two types of intergenic spacer regions (that is, two AFLs), whereas Chroococcales and Oscillatoriales have only one.…”

Section: Discussionmentioning

confidence: 99%

Sources of edaphic cyanobacterial diversity in the Dry Valleys of Eastern Antarctica

et al. 2008

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Cyanobacteria are major components of Antarctic Dry Valley ecosystems. Their occurrence in lakes and ponds is well documented, however, less is known about their distribution in edaphic environments. There has been considerable debate about the contribution of aquatic organic matter derived largely from cyanobacteria to terrestrial ecosystems. In this study, automated rRNA intergenic spacer analysis (ARISA) and 16S rRNA gene clone libraries were used to investigate cyanobacterial diversity in a range of soil environments within the Miers and Beacon Valleys. These data were used to elucidate the input of aquatic cyanobacteria to soil communities. Thirty-eight samples were collected from a variety of soil environments including dry and moist soils, hypoliths and lake and hydroterrestrial microbial mats. The results from the ARISA and 16S rRNA clone library analysis demonstrated that diverse cyanobacterial communities exist within the mineral soils of the Miers Valley. The soil samples from Beacon Valley were depauparate in cyanobacterial signals. Within Miers Valley, significant portions (29%-58%) of ARISA fragment lengths found in aquatic cyanobacterial mats were also present in soil and hypolith samples, indicating that lacustrine and hydroterrestrial cyanobacteria play a significant role in structuring soil communities. The influence of abiotic variables on the community structure of soil samples was assessed using BEST analysis. The results of BEST analysis of samples from within Miers Valley showed that total percentage of carbon content was the most important variable in explaining differences in cyanobacterial community structure. The BEST analyses indicated that four elements contributed significantly to species compositional differences between valleys. We suggest that the complete absence of lakes or ponds from Beacon Valley is a contributing factor to the low cyanobacterial component of these soils.

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Section: Discussionmentioning

confidence: 99%

Sources of edaphic cyanobacterial diversity in the Dry Valleys of Eastern Antarctica

et al. 2008

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“…Taton One caveat when interpreting ARISA data is that interoperonic differences in spacer length occur within the genomes of microorganisms (30), such that a single species may contribute more than one peak to an ARISA profile. Previous studies (13,53) indicate that members of the order Nostocales commonly have two types of intergenic spacer regions (i.e., two AFL), whereas members of the orders Chroococcales and Oscillatoriales have only one.…”

Section: Physical and Chemical Characteristics Of Pondsmentioning

confidence: 99%

Widespread Distribution and Identification of Eight Novel Microcystins in Antarctic Cyanobacterial Mats

Wood

Mountfort

Selwood

et al. 2008

Appl Environ Microbiol

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“…An overview of the size of the IGS within the main bacterial groups was obtained by comparing published data (5,8,15,28,45) and data from GenBank (NCBI) available as of September 2000. These data were from 99 bacterial genera and 332 species of bacteria (phylum ␣-, ␤-, ␥-, and ε-subdivisions of the genera Proteobacteria, low GϩC firmicutes, high GϩC firmicutes, Cyanobacteria, Chlamydiae, Cytophagales, and Spirochetes).…”

Section: Soil Origin and Microcosm Setupmentioning

confidence: 99%

Sequencing Bands of Ribosomal Intergenic Spacer Analysis Fingerprints for Characterization and Microscale Distribution of Soil Bacterium Populations Responding to Mercury Spiking

Ranjard

Brothier

Nazaret

2000

Appl Environ Microbiol

123

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Two major emerging bands (a 350-bp band and a 650-bp band) within the RISA (ribosomal intergenic spacer analysis) profile of a soil bacterial community spiked with Hg(II) were selected for further identification of the populations involved in the response of the community to the added metal. The bands were cut out from polyacrylamide gels, cloned, characterized by restriction analysis, and sequenced for phylogenetic affiliation of dominant clones. The sequences were the intergenic spacer between the rrs and rrl genes and the first 130 nucleotides of the rrl gene. Comparison of sequences derived from the 350-bp band to The GenBank database permitted us to identify the bacteria as being mostly close relatives to low G؉C firmicutes (Clostridium-like genera), while the 650-bp band permitted us to identify the bacteria as being mostly close relatives to ␤-proteobacteria (Ralstonia-like genera). Oligonucleotide probes specific for the identified dominant bacteria were designed and hybridized with the RISA profiles derived from the control and spiked communities. These studies confirmed the contribution of these populations to the community response to the metal. Hybridization of the RISA profiles from subcommunities (bacterial pools associated with different soil microenvironments) also permitted to characterize the distribution and the dynamics of these populations at a microscale level following mercury spiking.Recent major advances in the field of molecular biology have made it possible to develop community DNA fingerprinting methods with which to monitor complex bacterial communities in their natural environment without the need for isolation and cultivation steps. These methods involve in situ extraction of DNA from the microbial community, the PCR amplification of sequences of interest, and analysis of part of the genetic information. The sequences most commonly used as markers of communities are the genes of the ribosomal operon. The diversity of the amplified sequences is then resolved by differential electrophoretic migration on agarose or polyacrylamide gels, based on their size (terminal-restriction fragment length polymorphism, ribosomal intergenic spacer analysis [RISA]) or sequence (denaturing gel gradient electrophoresis, thermal gel gradient electrophoresis). These socalled fingerprinting methods provide band profiles that are representative of the genetic structure of the community as a whole or of a section of it, as defined by selected primers (for a review, see reference 26). These methods are valuable tools for characterizing complex bacterial communities and detecting shifts following environmental perturbations and are less time-consuming and labor-intensive than strategies such as small-subunit rRNA gene clone library construction (for a review, see reference 38). Individual bands can also be excised from a fingerprint profile, cloned and sequenced, or challenged with a range of probes providing precise information on the phylogenetic groups constituting a community (25,17).We previously looked...

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Utility of 16S–23S rRNA spacer region methodology: how similar are interspace regions within a genome and between strains for closely related organisms?

Cited by 67 publications

References 21 publications

Sources of edaphic cyanobacterial diversity in the Dry Valleys of Eastern Antarctica

Sources of edaphic cyanobacterial diversity in the Dry Valleys of Eastern Antarctica

Widespread Distribution and Identification of Eight Novel Microcystins in Antarctic Cyanobacterial Mats

Sequencing Bands of Ribosomal Intergenic Spacer Analysis Fingerprints for Characterization and Microscale Distribution of Soil Bacterium Populations Responding to Mercury Spiking

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