Geoarchaeota: a new candidate phylum in the Archaea from high-temperature acidic iron mats in Yellowstone National Park.

Geothermal systems in Yellowstone National Park (YNP) provide an outstanding opportunity to understand the origin and evolution of metabolic processes necessary for life in extreme environments including low pH, high temperature, low oxygen and elevated concentrations of reduced iron. Previous phylogenetic studies of acidic ferric iron mats from YNP have revealed considerable diversity of uncultivated and undescribed archaea. The goal of this study was to obtain replicate de novo genome assemblies for a dominant archaeal population inhabiting acidic iron-oxide mats in YNP. Detailed analysis of conserved ribosomal and informational processing genes indicates that the replicate assemblies represent a new candidate phylum within the domain Archaea referred to here as 'Geoarchaeota' or 'novel archaeal group 1 (NAG1)'. The NAG1 organisms contain pathways necessary for the catabolism of peptides and complex carbohydrates as well as a bacterial-like Form I carbon monoxide dehydrogenase complex likely used for energy conservation. Moreover, this novel population contains genes involved in the metabolism of oxygen including a Type A heme copper oxidase, a bd-type terminal oxidase and a putative oxygen-sensing protoglobin. NAG1 has a variety of unique bacterial-like cofactor biosynthesis and transport genes and a Type3-like CRISPR system. Discovery of NAG1 is critical to our understanding of microbial community structure and function in extant thermophilic iron-oxide mats of YNP, and will provide insight regarding the evolution of Archaea in early Earth environments that may have important analogs active in YNP today.

Effect of repeat copy number on variable-number tandem repeat mutations in Escherichia coli O157:H7.

Variable-number tandem repeat (VNTR) loci have shown a remarkable ability to discriminate among isolates of the recently emerged clonal pathogen Escherichia coli O157:H7, making them a very useful molecular epidemiological tool. However, little is known about the rates at which these sequences mutate, the factors that affect mutation rates, or the mechanisms by which mutations occur at these loci. Here, we measure mutation rates for 28 VNTR loci and investigate the effects of repeat copy number and mismatch repair on mutation rate using in vitro-generated populations for 10 E. coli O157:H7 strains. We find single-locus rates as high as 7.0 x 10(-4) mutations/generation and a combined 28-locus rate of 6.4 x 10(-4) mutations/generation. We observed single- and multirepeat mutations that were consistent with a slipped-strand mispairing mutation model, as well as a smaller number of large repeat copy number mutations that were consistent with recombination-mediated events. Repeat copy number within an array was strongly correlated with mutation rate both at the most mutable locus, O157-10 (r2= 0.565, P = 0.0196), and across all mutating loci. The combined locus model was significant whether locus O157-10 was included (r2= 0.833, P < 0.0001) or excluded (r2= 0.452, P < 0.0001) from the analysis. Deficient mismatch repair did not affect mutation rate at any of the 28 VNTRs with repeat unit sizes of >5 bp, although a poly(G) homomeric tract was destabilized in the mutS strain. Finally, we describe a general model for VNTR mutations that encompasses insertions and deletions, single- and multiple-repeat mutations, and their relative frequencies based upon our empirical mutation rate data.

Strain typing of Borrelia burgdorferi, Borrelia afzelii, and Borrelia garinii by using multiple-locus variable-number tandem repeat analysis.

Human Lyme borreliosis (LB) is the most prevalent arthropod-borne infection in temperate climate zones around the world and is caused by Borrelia spirochetes. We have identified 10 variable-number tandem repeat (VNTR) loci present within the genome of Borrelia burgdorferi and subsequently developed a multiple-locus VNTR analysis (MLVA) typing system for this disease agent. We report here the successful application of MLVA for strain discrimination among a group of 41 globally diverse Borrelia isolates including B. burgdorferi, B. afzelii, and B. garinii. PCR assays displayed diversity at these loci, with total allele numbers ranging from two to nine and Nei's diversity (D) values ranging from 0.10 to 0.87. The average D value was 0.53 across all VNTR loci. A clear correlation exists between the repeat copy number and the D value (r = 0.62) or the number of alleles (r = 0.93) observed across diverse strains. Cluster analysis by the unweighted pair-group method with arithmetic means resolved the 30 observed unique Borrelia genotypes into five distinct groups. B. burgdorferi, B. afzelii, and B. garinii clustered into distinct affiliations, consistent with current 16S rRNA phylogeny studies. Genetic similarity and diversity suggest that B. afzelii and B. garinii are close relatives and were perhaps recently derived from B. burgdorferi. MLVA provides both phylogenetic relationships and additional resolution to discriminate among strains of Borrelia species. This new level of strain identification and discrimination will allow more detailed epidemiological and phylogenetic analysis in future studies.