Phylogenetic characterization of dwarf archaea and bacteria from a semiarid soil

Microscopic studies have shown that most microbes exist in soils as dwarf cells that are thought to be an adaptation to nutrient limitation. Most dwarfs are uncultured by current approaches and have not been identified phylogenetically. Only a few dwarf bacteria have been isolated in culture and dwarf archaea have received little study. We selected a semiarid creosotebush shrubland site for studying dwarf microorganisms because arid and semiarid soils are generally nutrient-poor. Soils were collected beneath creosotebush canopies and in open areas between shrubs. Cells were eluted in sodium pyrophosphate and filtered with a 0.45-μm pore-size filter. Filtrate DNA was extracted, PCR-amplified using universal bacterial and archaeal 16S rDNA primers, cloned, RFLP-screened, and sequenced. The eluted cell filtrates were also inoculated into R2B medium. After incubation, cultures were filtered to select against populations of dwarfs that formed large cells in the presence of nutrients (pleomorphic dwarfs) and to select for populations that retained dwarf size (intrinsic dwarfs). Dwarf archaea and bacteria were present in the initial filtrate and in the cultures. A single dwarf archaeon (SevArch-01) that is related to other soil Crenarchaeota sequences was found in the initial filtrates and in subsequent filtrate cultures, indicating an intrinsically dwarf archaeon. Dwarf bacteria fell into four bacterial phyla: Proteobacteria, Firmicutes, Actinobacteria, and the TM-7 group. Intrinsically dwarf bacteria in enrichment cultures were identified as α- and β-Proteobacteria. Dwarf bacteria related to Arthrobacter, Propionibacterium, and other actinobacteria were detected. Several sequences showed no close relationships to any microorganisms that have been grown in culture.