Michael J. Ferris

Cyanobacterial ecotypes in the microbial mat community of Mushroom Spring (Yellowstone National Park, Wyoming) as species-like units linking microbial community composition, structure and function

We have investigated microbial mats of alkaline siliceous hot springs in Yellowstone National Park as natural model communities to learn how microbial populations group into species-like fundamental units. Here, we bring together empirical patterns of the distribution of molecular variation in predominant mat cyanobacterial populations, theory-based modelling of how to demarcate phylogenetic clusters that correspond to ecological species and the dynamic patterns of the physical and chemical microenvironments these populations inhabit and towards which they have evolved adaptations. We show that putative ecotypes predicted by the theory-based model correspond well with distribution patterns, suggesting populations with distinct ecologies, as expected of ecological species. Further, we show that increased molecular resolution enhances our ability to detect ecotypes in this way, though yet higher molecular resolution is probably needed to detect all ecotypes in this microbial community.

Biodiversity within hot spring microbial mat communities: molecular monitoring of enrichment cultures.

We have begun to examine the basis for incongruence between hot spring microbial mat populations detected by cultivation or by 16S rRNA methods. We used denaturing gradient gel electrophoresis (DGGE) to monitor enrichments and isolates plated therefrom. At near extincting inoculum dilutions we observed Chloroflexus-like and cyanobacterial populations whose 16S rRNA sequences have been detected in the ‘New Pit’ Spring Chloroflexus mat and the Octopus Spring cyanobacterial mat. Cyanobacterial populations enriched from 44 to 54°C and 56 to 63°C samples at near habitat temperatures were similar to those previously detected in mat samples of comparable temperatures. However, a lower temperature enrichment from the higher temperature sample selected for the populations found in the lower temperature sample. Three Thermus populations detected by both DGGE and isolation exemplify even more how enrichment may bias our view of community structure. The most abundant population was adap ted to the habitat temperature (50°C), while populations adapted to 65°C and 70°C were 102- and 104-fold less abundant, respectively. However, enrichment at 70°C favored the least abundant strain. Inoculum dilution and incubation at the habitat temperature favored the more numerically relevant populations. We enriched many other aerobic chemoorganotropic populations at various inoculum dilutions and substrate concentrations, most of whose 16S rRNA sequences have not been detected in mats. A common feature of numerically relevant cyanobacterial, Chloroflexus-like and aerobic chemorganotrophic populations, is that they grow poorly and resist cultivation on solidified medium, suggesting plating bias, and that the medium composition and incubation conditions may not reflect the natural microenvironments these populations inhabit.

PCR detection and analysis of the free-living amoeba Naegleria in hot springs in Yellowstone and Grand Teton National Parks.

ABSTRACT Free-living thermotolerant amoebae pose a significant health risk to people who soak and swim in habitats suitable for their growth, such as hot springs. In this survey of 23 different hot springs in Yellowstone and Grand Teton National Parks, we used PCR with primer sets specific for Naegleria to detect three sequence types that represent species not previously described, as well as a fourth sequence type identified as the pathogen Naegleria fowleri.

Legionella species diversity in an acidic biofilm community in Yellowstone National Park

ABSTRACT Legionella species are frequently detected in aquatic environments, but their occurrence in extreme, acidic, geothermal habitats has not been explored with cultivation-independent methods. We investigated a predominately eukaryotic algal mat community in a pH 2.7 geothermal stream in Yellowstone National Park for the presence of Legionella and potential host amoebae. Our analyses, using PCR amplification with Legionella-specific primers targeting 16S rRNA genes, detected four known Legionella species, as well as Legionella sequences from species that are not represented in sequence databases, in mat samples and cultivated isolates. The nonrandom occurrence of sequences detected at lower (30°C) and higher (35 to 38°C) temperatures suggests that natural thermal gradients in the stream influence Legionella species distributions in this mat community. We detected only one sequence, Legionella micdadei, from cultivated isolates. We cultured and sequenced partial 18S rRNA gene regions from two potential hosts, Acanthamoeba and Euglena species.

Species diversity in hot spring microbial mats as revealed by both molecular and enrichment culture approaches — relationship between biodiversity and community structure

We have conducted long-term studies of microbial mats of hot springs as model systems for investigating composition and structure of natural microbial communities and as modern analogs of stromatolites (Ward et al. 1984, 1987, 1989b). As recently as the last symposium on microbial mats our knowledge of the biodiversity within these communities was based solely on microbial species cultivated from such mat systems. The view has changed dramatically in the intervening years, because of the application of culture-independent techniques to recover and probe ribosomal RNAs (rRNAs) as biomarkers representing individual mat community members (Ward et al. 1992; Weller et al. 1992 and references cited therein), and more recently because of the renewed attempt to cultivate more relevant species. Here, we summarize what is currently known. The evidence suggests that the molecular methods we are using (Ward et al. 1992) may still not permit a complete understanding of the true complexity of the community. However, the approach does provide insight into understanding the basis behind this large biodiversity and into how more relevant species can be cultivated.

Prevalence and abundance of uncultivated Megasphaera-like bacteria in the human vaginal environment.

ABSTRACT Cultivation-independent analysis of 16S rRNA gene sequences in vaginal samples revealed two previously unrecognized, uncultivated Megasphaera-like phylotypes. Phylogenetic analysis and environmental distribution suggest that these Megasphaera types may be unique to the vaginal environment. Quantitative PCR suggests that both phylotypes are present in higher concentrations in women with bacterial vaginosis.

Detection of Naegleria sp. in a Thermal, Acidic Stream in Yellowstone National Park

Abstract An initial survey of sequences of PCR-amplified portions of the 18S rRNA genes from a community DNA clone library, prepared from an algal mat in a thermal, acidic stream in Yellowstone National Park, WY, USA, revealed among other sequences, several that matched Vahlkampfia. This finding prompted further investigation using primers specific for Naegleria. Sequences from a subsequent DNA clone library, prepared from the 5.8S rRNA gene and the adjacent internal transcribed spacer (ITS) regions of the rRNA, closely matched Naegleria and formed an independent lineage within a clade containing Naegleria sturti and Naegleria niuginiensis. The sequences may represent a new Naegleria species.