Open AccessDevelopment and Application of Small-Subunit rRNA Probes for Assessment of Selected Thiobacillus Species and Members of the Genus Acidiphilium
Author(s) -
Jordan Peccia,
Éric Marchand,
JoAnn Silverstein,
Mark Hernandez
Publication year - 2000
Publication title -
applied and environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.552
H-Index - 324
eISSN - 1070-6291
pISSN - 0099-2240
DOI - 10.1128/aem.66.7.3065-3072.2000
Subject(s) - thiobacillus , bacteria , biology , microbiology and biotechnology , oligonucleotide , ribosomal rna , 16s ribosomal rna , molecular probe , oligomer restriction , biochemistry , hybridization probe , chemistry , dna , gene , genetics
Culture-dependent studies have implicated sulfur-oxidizing bacteria as the causative agents of acid mine drainage and concrete corrosion in sewers.Thiobacillus species are considered the major representatives of the acid-producing bacteria in these environments. Small-subunit rRNA genes from all of theThiobacillus andAcidiphilium species catalogued by the Ribosomal Database Project were identified and used to design oligonucleotide DNA probes. Two oligonucleotide probes were synthesized to complement variable regions of 16S rRNA in the following acidophilic bacteria:Thiobacillus ferrooxidans andT. thiooxidans (probe Thio820) and members of the genusAcidiphilium (probe Acdp821). Using32 P radiolabels, probe specificity was characterized by hybridization dissociation temperature (Td ) with membrane-immobilized RNA extracted from a suite of 21 strains representing three groups of bacteria. Fluorochrome-conjugated probes were evaluated for use with fluorescent in situ hybridization (FISH) at the experimentally determinedTd s. FISH was used to identify and enumerate bacteria in laboratory reactors and environmental samples. Probing of laboratory reactors inoculated with a mixed culture of acidophilic bacteria validated the ability of the oligonucleotide probes to track specific cell numbers with time. Additionally, probing of sediments from an active acid mine drainage site in Colorado demonstrated the ability to identify numbers of active bacteria in natural environments that contain high concentrations of metals, associated precipitates, and other mineral debris.