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  • in situ hybridization  (1)
  • microcosms  (1)
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  • 1
    ISSN: 1573-5117
    Keywords: microcosms ; bacterial populations ; 16S rRNA oligonucleotide probes ; lignin ; coastal seawater
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Most studies of marine bacterial communities focus on functional attributes of the community, rather than on population or community structure, at least in part, because of the difficulty in enumerating individual species within complex communities. Here, we describe a study in which populations of three bacterial species were followed over time in replicate marine enrichment cultures using 16S rRNA-based oligonucleotide probes. Three identical enrichment microcosms were established with lignin-rich pulp mill waste as a sole carbon source, inoculated with coastal seawater, and transferred at two-week intervals. Population levels were assessed throughout a six-week period using species-specific 16S rRNA-based oligonucleotide probes directed toward three bacterial species that were numerically important (and culturable) members of the enrichments. Substantial differences in the population levels of each bacterial species were found among the triplicate incubations, despite the fact that the enrichments were inoculated and treated identically. Stochastic differences in the composition of the inoculum and/or ecological interactions within the enrichment replicates may have been important in determining final population levels. Functional ability, assessed as rates of degradation of a synthetic lignin preparation, were fairly similar among the three replicate enrichments (within 70%), despite the substantial differences in population levels of the representative lignin-degrading species.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1573-5117
    Keywords: in situ PCR ; in situ hybridization ; todC1 gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The toluene dioxygenase (todC1) gene and its mRNA transcripts were amplified by in situ PCR and in situ RT-PCR, respectively, in intact cells of the bacterium Pseudomonas putida F1. In situ amplicons of DNA and mRNA were then detected by hybridizing to a fluorescently labeled oligonucleotide. In situ PCR protocols were developed to distinguish between cells of P. putida F1 (possessing the todC1 gene) and P. putida AC10R (lacking the todC1 gene); the method was sensitive enough to detect amplified products from a single copy of the todC1 gene. P. putida F1 cells were also introduced into seawater with toluene addition. Cells expressing todC1 and total cells were detectable by in situ RT-PCR and Yo-Pro 1 counterstaining, respectively. Nearly 90% of cells expressing the todC1 gene were detected in seawater amended with toluene at day 3, but no cells expressing todC1 were detected in seawater not exposed to toluene. Our results suggest that in situ PCR amplification can be a useful technique for studying presence or absence of a specific gene and gene expression of bioremediative bacteria at the individual cell level following release into natural environments.
    Type of Medium: Electronic Resource
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