PHYLOGENETIC ANALYSIS OF NONTHERMOPHILIC MEMBERS OF THE KINGDOM CRENARCHAEOTA AND THEIR DIVERSITY AND ABUNDANCE IN SOILS

Within the last several years, molecular techniques have uncovered num erous 16S rRNA gene (rDNA) sequences which represent a unique and glob ally distributed lineage of the kingdom Crenarchaeota that is phylogen etically distinct from currently characterized crenarchaeotal species. rDNA sequences of members of this novel crenarchaeotal group have bee n recovered from low- to moderate-temperature environments (-1.5 to 32 degrees C), in contrast to the high-temperature environments (tempera ture, >80 degrees C) required for growth of the currently recognized c renarchaeotal species. We determined the diversity and abundance of th e nonthermophilic members of the Crenarchaeota in soil samples taken f rom cultivated and uncultivated fields located at the Kellogg Biologic al Station's Long-Term Ecological Research site (Hickory Corners, Mich ,). Clones were generated from 16S rDNA that was amplified by using br oad-specificity archaeal PCR primers. Twelve crenarchaeotal sequences were identified, and the phylogenetic relationships between these sequ ences and previously described crenarchaeotal 16S rDNA sequences were determined. Phylogenetic analyses included nonthermophilic crenarchaeo tal sequences found in public databases and revealed that the nontherm ophilic Crenarchaeota group is composed of at least four distinct phyl ogenetic clusters. A 16S rRNA-targeted oligonucleotide probe specific for all known nonthermophilic crenarchaeotal sequences was designed an d used to determine their abundance in soil samples. The nonthermophil ic Crenarchaeota accounted for as much as 1.42% +/- 0.42% of the 16S r RNA in the soils analyzed.