Abstract
Based on comparative phylogenetic analysis of 16S rRNA gene sequences deposited in an RDP database, we constructed a local database of thaumarchaeotal 16S rRNA gene sequences and developed a novel PCR primer specific for the archaeal phylum Thaumarchaeota. Among 9,727 quality-filtered (chimeral-checked, size >1.2 kb) archaeal sequences downloaded from the RDP database, 1,549 thaumarchaeotal sequences were identified and included in our local database. In our study, Thaumarchaeota included archaeal groups MG-I, SAGMCG-I, SCG, FSCG, RC, and HWCG-III, forming a monophyletic group in the phylogenetic tree. Cluster analysis revealed 114 phylotypes for Thaumarchaeota. The majority of the phylotypes (66.7%) belonged to the MG-I and SCG, which together contained most (93.9%) of the thaumarchaeotal sequences in our local database. A phylum-directed primer was designed from a consensus sequence of the phylotype sequences, and the primer’s specificity was evaluated for coverage and tolerance both in silico and empirically. The phylum-directed primer, designated THAUM-494, showed >90% coverage for Thaumarchaeota and <1% tolerance to non-target taxa, indicating high specificity. To validate this result experimentally, PCRs were performed with THAUM-494 in combination with a universal archaeal primer (ARC917R or 1017FAR) and DNAs from five environmental samples to construct clone libraries. THAUM-494 showed a satisfactory specificity in empirical studies, as expected from the in silico results. Phylogenetic analysis of 859 cloned sequences obtained from 10 clone libraries revealed that >95% of the amplified sequences belonged to Thaumarchaeota. The most frequently sampled thaumarchaeotal subgroups in our samples were SCG, MG-I, and SAGMCG-I. To our knowledge, THAUM-494 is the first phylum-level primer for Thaumarchaeota. Furthermore, the high coverage and low tolerance of THAUM-494 will make it a potentially valuable tool in understanding the phylogenetic diversity and ecological niche of Thaumarchaeota.
Highlights
The archaeal phylum Thaumarchaeota was proposed in 2008, distinguishing mesophilic ammonia-oxidizing archaeal (AOA) lineages from hyperthermophilic Crenarchaeota lineages [1]
This proposal was based on archaeal phylogeny inferred from rRNA and ribosomal protein sequences, which suggested that mesophilic Crenarchaeota constitute a distinct phylum that branches off near the root of Archaea
The sequence set for the backbone tree, B = {backbone sequences}, included 106 sequences from the group MG-I as key members of the phylum Thaumarchaeota, which were selected based on published literature [12,42–49], and 72 genus-level representative sequences from the phyla Euryarchaeota, Crenarchaeota, Nanoarchaeota, and Korarchaeota
Summary
The archaeal phylum Thaumarchaeota was proposed in 2008, distinguishing mesophilic ammonia-oxidizing archaeal (AOA) lineages from hyperthermophilic Crenarchaeota lineages [1]. A number of molecular ecological studies have revealed that the Archaea inhabit a wide variety of moderate environments [5], suggesting that they play a substantial role in global geochemical cycling. Based on 16S rRNA gene surveys, the Thaumarchaeota have been estimated to represent up to 20% and 5% of all prokaryotes in marine and terrestrial environments, respectively [6–8]. Another notable feature of the Thaumarchaeota is that all cultured or enriched members of this phylum are ammonia oxidizers [9– 16]. Recent studies have shown that the copy numbers of archaeal amoA are much higher than the copy numbers of bacterial amoA in many soil samples [24– 29], indicating the predominance of AOA over AOB
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
