Kevin J. Purdy

Evolutionary divergence and biogeography of sympatric niche-differentiated bacterial populations

Using multiple lines of evidence from denaturing gradient gel electrophoresis, environmental sequences and TaqMan quantitative PCR assays targeting a functional gene for sulfate respiration (dsr) affiliated with the geochemically important genus Desulfobulbus, we revealed strongly restricted distributions of specific genotypes and populations correlated with sampling position along an estuarine gradient free of dispersal barriers. Evidence of evolutionary divergence of populations was provided by three complementary analyses. First, analysis of molecular variance rejected the null hypothesis that genetic diversity within each sampling site was not significantly different than that of all sites pooled together (P<0.0001). Second, UniFrac and Parsimony tests showed phylogenetic clustering of sampling sites was highly significant (P<0.001). Third, pairwise FST statistics showed significant evolutionary divergence of populations based on the location in the estuary. To test the hypothesis that environmental niche-driven evolutionary divergence can create and maintain microbial biogeography, we used both statistical inference and an experimental manipulation to assess the independent effects of environment and geography. Significant effects of each on genotype distributions and population divergence supported the hypothesis. Our data are consistent with both sympatric and parapatric models of speciation, and suggest niche partitioning can contribute to evolutionary divergence and observable biogeographic patterns in microbial communities even among closely related taxa at limited spatial scales without significant barriers to dispersal.

Systems Biology for Ecology: From Molecules to Ecosystems

Summary Ecology stands on the edge of a true paradigm shift, fuelled by a recent technological revolution in our ability to measure both taxonomic and functional biodiversity via the application of metagenomics and transcriptomics. The advent of ‘next generation sequencing’ (NGS) in molecular biology is rapidly opening the black box of microbial ecology, providing us with some of the first glimpses of a previously hidden world. This is now enabling microbial ecology to become firmly embedded as a core subdiscipline within ecology, and to test general theories about biodiversity, biogeography and ecosystem functioning using a combination of molecular and more traditional techniques. In addition, NGS offers a means of not only measuring the abundance and diversity of the main drivers of many of the planets key biogeochemical processes, but also of linking the microscopic and macroscopic worlds that have, until now, been largely studied in isolation. We provide a detailed review of the rise of NGS, as well as highlighting areas that offer special promise for addressing general ecological questions across a range of levels of organisation, from individuals to ecosystems: essentially, how a ‘systems biology for ecology’ might be developed. We consider the current limitations and future prospects for NGS, and also how it offers potential economic benefits, for instance via bioprospecting the environment for commercially valuable genes and their products within the metagenome of natural ecosystems.

Chapter 3 - Systems Biology for Ecology: From Molecules to Ecosystems

Summary Ecology stands on the edge of a true paradigm shift, fuelled by a recent technological revolution in our ability to measure both taxonomic and functional biodiversity via the application of metagenomics and transcriptomics. The advent of ‘next generation sequencing’ (NGS) in molecular biology is rapidly opening the black box of microbial ecology, providing us with some of the first glimpses of a previously hidden world. This is now enabling microbial ecology to become firmly embedded as a core subdiscipline within ecology, and to test general theories about biodiversity, biogeography and ecosystem functioning using a combination of molecular and more traditional techniques. In addition, NGS offers a means of not only measuring the abundance and diversity of the main drivers of many of the planets key biogeochemical processes, but also of linking the microscopic and macroscopic worlds that have, until now, been largely studied in isolation. We provide a detailed review of the rise of NGS, as well as highlighting areas that offer special promise for addressing general ecological questions across a range of levels of organisation, from individuals to ecosystems: essentially, how a ‘systems biology for ecology’ might be developed. We consider the current limitations and future prospects for NGS, and also how it offers potential economic benefits, for instance via bioprospecting the environment for commercially valuable genes and their products within the metagenome of natural ecosystems.

Manuka honey inhibits adhesion and invasion of medically important wound bacteria in vitro

AIM To characterize the effect of manuka honey on medically important wound bacteria in vitro, focusing on its antiadhesive properties. MATERIALS & METHODS Crystal violet biofilm assays, fluorescent microscopy, protein adhesion assay and gentamicin protection assay were used to determine the impact of manuka honey on biofilm formation, human protein binding and adherence to/invasion into human keratinocytes. RESULTS Manuka honey effectively disrupted and caused extensive cell death in biofilms of Staphylococcus aureus, Pseudomonas aeruginosa and Streptococcus pyogenes. Sublethal doses of manuka honey inhibited bacterial adhesion to the fibronectin, fibrinogen and collagen. Manuka honey impaired adhesion of laboratory and clinical isolates of S. aureus, P. aeruginosa and S. pyogenes to human keratinocytes in vitro, and inhibited invasion by S. pyogenes and homogeneous vancomycin intermediate S. aureus. CONCLUSION Manuka honey can directly affect bacterial cells embedded in a biofilm and exhibits antiadhesive properties against three common wound pathogens.

Contrasting patterns of niche partitioning between two anaerobic terminal oxidizers of organic matter.

Understanding the ecological principles underlying the structure and function of microbial communities remains an important goal for microbial ecology. We examined two biogeochemically important taxa, the sulfate-reducing bacterial genus, Desulfobulbus, and the methanogenic archaeal genus, Methanosaeta, to compare and contrast niche partitioning by these two taxa that are ecologically linked as anaerobic terminal oxidizers of organic material. An observational approach utilizing functional gene pyrosequencing was combined with a community-based reciprocal incubation experiment and characterization of a novel Desulfobulbus isolate. To analyze the pyrosequencing data, we constructed a data analysis pipeline, which we validated with several control data sets. For both taxa, particular genotypes were clearly associated with certain portions of an estuarine gradient, consistent with habitat or niche partitioning. Methanosaeta genotypes were generally divided between those found almost exclusively in the marine habitat (∼30% of operational taxonomic units (OTUs)), and those which were ubiquitously distributed across all or most of the estuary (∼70% of OTUs). In contrast to this relatively monotonic distribution, for Desulfobulbus, there were many more genotypes, and their distributions represented a wide range of inferred niche widths from specialist genotypes found only at a single site, to ubiquitous or generalist genotypes found in all 10 sites examined along the full estuarine gradient. Incubation experiments clearly showed that, for both taxa, communities from opposite ends of the estuary did not come to resemble one another, regardless of the chemical environment. Growth of a Desulfobulbus isolated into pure culture indicated that the potential niche of this organism is significantly larger than the realized niche. We concluded that niche partitioning can be an important force structuring microbial populations, with biotic and abiotic components having very different effects depending on the physiology and ecology of each taxon.

The distribution and diversity of Euryarchaeota in termite guts

Publisher Summary The Euryarchaeota are a critical component of all termite guts, acting as hydrogen sinks in both lower and higher termites. In lower termites and in the wood- and fungus-feeding higher termites, the role of the methanogens appears to be to mop up trace hydrogen. In the soil-feeding higher termites, methanogenesis lies at the heart of termite nutrition and represents an essential gut process. New analyses of termite evolution suggest that the evolution of the higher termites probably occurred via an externalization of the gut in the fungus-feeding Macrotermitinae followed by the evolution of soil-feeding. It is plausible that a first step in this process would have been the acquisition of a Methanomicrobiales strain that had the potential to reduce hydrogen partial pressures to levels that allowed the effective exploitation of soil organic matter.

Phenotypic and genotypic characteristics of small colony variants and their role in chronic infection

Small colony variant (SCV) bacteria arise spontaneously within apparently homogeneous microbial populations, largely in response to environmental stresses, such as antimicrobial treatment. They display unique phenotypic characteristics conferred in part by heritable genetic changes. Characteristically slow growing, SCVs comprise a minor proportion of the population from which they arise but persist by virtue of their inherent resilience and host adaptability. Consequently, SCVs are problematic in chronic infection, where antimicrobial treatment is administered during the acute phase of infection but fails to eradicate SCVs, which remain within the host causing recurrent or chronic infection. This review discusses some of the phenotypic and genotypic changes that enable SCVs to successfully proliferate within the host environment as potential pathogens and strategies that could ameliorate the resolution of infection where SCVs are present.

Metabolic flexibility as a major predictor of spatial distribution in microbial communities.

A better understand the ecology of microbes and their role in the global ecosystem could be achieved if traditional ecological theories can be applied to microbes. In ecology organisms are defined as specialists or generalists according to the breadth of their niche. Spatial distribution is often used as a proxy measure of niche breadth; generalists have broad niches and a wide spatial distribution and specialists a narrow niche and spatial distribution. Previous studies suggest that microbial distribution patterns are contrary to this idea; a microbial generalist genus (Desulfobulbus) has a limited spatial distribution while a specialist genus (Methanosaeta) has a cosmopolitan distribution. Therefore, we hypothesise that this counter-intuitive distribution within generalist and specialist microbial genera is a common microbial characteristic. Using molecular fingerprinting the distribution of four microbial genera, two generalists, Desulfobulbus and the methanogenic archaea Methanosarcina, and two specialists, Methanosaeta and the sulfate-reducing bacteria Desulfobacter were analysed in sediment samples from along a UK estuary. Detected genotypes of both generalist genera showed a distinct spatial distribution, significantly correlated with geographic distance between sites. Genotypes of both specialist genera showed no significant differential spatial distribution. These data support the hypothesis that the spatial distribution of specialist and generalist microbes does not match that seen with specialist and generalist large organisms. It may be that generalist microbes, while having a wider potential niche, are constrained, possibly by intrageneric competition, to exploit only a small part of that potential niche while specialists, with far fewer constraints to their niche, are more capable of filling their potential niche more effectively, perhaps by avoiding intrageneric competition. We suggest that these counter-intuitive distribution patterns may be a common feature of microbes in general and represent a distinct microbial principle in ecology, which is a real challenge if we are to develop a truly inclusive ecology.

Improving the isolation of anaerobes on solid media: the example of the fastidious Methanosaeta

Methanosaeta spp. are globally important biogenic methane producers with only three strains described due to isolation difficulties. Here, clonal axenic isolates from estuarine sediments were obtained using alternative gelling and reducing agents. It is suggested that more systematic approaches with various combinations of media components will help to cultivate difficult-to-isolate anaerobes.

Nitrous oxide as a function of oxygen and archaeal gene abundance in the North Pacific.

Oceanic oxygen minimum zones are strong sources of the potent greenhouse gas N2O but its microbial source is unclear. We characterized an exponential response in N2O production to decreasing oxygen between 1 and 30 μmol O2 l−1 within and below the oxycline using 15NO2−, a relationship that held along a 550 km offshore transect in the North Pacific. Differences in the overall magnitude of N2O production were accounted for by archaeal functional gene abundance. A one-dimensional (1D) model, parameterized with our experimentally derived exponential terms, accurately reproduces N2O profiles in the top 350 m of water column and, together with a strong 45N2O signature indicated neither canonical nor nitrifier–denitrification production while statistical modelling supported production by archaea, possibly via hybrid N2O formation. Further, with just archaeal N2O production, we could balance high-resolution estimates of sea-to-air N2O exchange. Hence, a significant source of N2O, previously described as leakage from bacterial ammonium oxidation, is better described by low-oxygen archaeal production at the oxygen minimum zones margins.