Biomedical subjects

Philip Hugenholtz

Publications and source records attributed to Philip Hugenholtz.

At least 19 recordsLinked to original sources

PubMed2026

The planktonic microbiome of the Great Barrier Reef.

Large genome databases have markedly improved our understanding of marine microorganisms1-5. Although these resources have focused on prokaryotes, genomes from many dominant marine lineages, such as Pelagibacter and Prochlorococcus, are conspicuously underrepresented. Here we present the Great Barrier Reef Microbial Genomes Database (GBR-MGD), comprising 5,283 prokaryotic genomes obtained from Great Barrier Reef seawater samples using Nanopore and Illumina sequencing, including a collection of high-quality genomes of underrepresented groups. We show that standard short-read assemblies miss these populations owing to a combination of strain heterogeneity and low-GC-percentage sequencing bias. The GBR-MGD also comprises 20 chromosome-level picoeukaryote and 808,585 viral genomes, including a newly described clade of marine Crassvirales. We demonstrate the utility of the GBR-MGD to identify indicator taxa that can reliably predict the effects of reef management practices, such as the establishment of marine protected zones.

Bacteria↗

PubMed2007

Riding giants.

Explore the source record for details and available documents.

Computational Biology↗

PubMed2006

Lineages of acidophilic archaea revealed by community genomic analysis.

Novel, low-abundance microbial species can be easily overlooked in standard polymerase chain reaction (PCR)-based surveys. We used community genomic data obtained without PCR or cultivation to reconstruct DNA fragments bearing unusual 16S ribosomal RNA (rRNA) and protein-coding genes from organisms belonging to novel archaeal lineages. The organisms are minor components of all biofilms growing in pH 0.5 to 1.5 solutions within the Richmond Mine, California. Probes specific for 16S rRNA showed that the fraction less than 0.45 micrometers in diameter is dominated by these organisms. Transmission electron microscope images revealed that the cells are pleomorphic with unusual folded membrane protrusions and have apparent volumes of <0.006 cubic micrometer.

Amino Acid Sequence↗

PubMed2006

Accurate phylogenetic classification of variable-length DNA fragments.

Metagenome studies have retrieved vast amounts of sequence data from a variety of environments leading to new discoveries and insights into the uncultured microbial world. Except for very simple communities, the encountered diversity has made fragment assembly and the subsequent analysis a challenging problem. A taxonomic characterization of metagenomic fragments is required for a deeper understanding of shotgun-sequenced microbial communities, but success has mostly been limited to sequences containing phylogenetic marker genes. Here we present PhyloPythia, a composition-based classifier that combines higher-level generic clades from a set of 340 completed genomes with sample-derived population models. Extensive analyses on synthetic and real metagenome data sets showed that PhyloPythia allows the accurate classification of most sequence fragments across all considered taxonomic ranks, even for unknown organisms. The method requires no more than 100 kb of training sequence for the creation of accurate models of sample-specific populations and can assign fragments >or=1 kb with high specificity.

Animals↗

PubMed2006

Metagenomic analysis of two enhanced biological phosphorus removal (EBPR) sludge communities.

Enhanced biological phosphorus removal (EBPR) is one of the best-studied microbially mediated industrial processes because of its ecological and economic relevance. Despite this, it is not well understood at the metabolic level. Here we present a metagenomic analysis of two lab-scale EBPR sludges dominated by the uncultured bacterium, "Candidatus Accumulibacter phosphatis." The analysis sheds light on several controversies in EBPR metabolic models and provides hypotheses explaining the dominance of A. phosphatis in this habitat, its lifestyle outside EBPR and probable cultivation requirements. Comparison of the same species from different EBPR sludges highlights recent evolutionary dynamics in the A. phosphatis genome that could be linked to mechanisms for environmental adaptation. In spite of an apparent lack of phylogenetic overlap in the flanking communities of the two sludges studied, common functional themes were found, at least one of them complementary to the inferred metabolism of the dominant organism. The present study provides a much needed blueprint for a systems-level understanding of EBPR and illustrates that metagenomics enables detailed, often novel, insights into even well-studied biological systems.

Adaptation, Biological↗

PubMed2006

The integrated microbial genomes (IMG) system.

The integrated microbial genomes (IMG) system is a new data management and analysis platform for microbial genomes provided by the Joint Genome Institute (JGI). IMG contains both draft and complete JGI genomes integrated with other publicly available microbial genomes of all three domains of life. IMG provides tools and viewers for analyzing genomes, genes and functions, individually or in a comparative context. IMG allows users to focus their analysis on subsets of genes and genomes of interest and to save the results of their analysis. IMG is available at http://img.jgi.doe.gov.

Bacteriophages↗

PubMed2006

The Genomes On Line Database (GOLD) v.2: a monitor of genome projects worldwide.

The Genomes On Line Database (GOLD) is a web resource for comprehensive access to information regarding complete and ongoing genome sequencing projects worldwide. The database currently incorporates information on over 1500 sequencing projects, of which 294 have been completed and the data deposited in the public databases. GOLD v.2 has been expanded to provide information related to organism properties such as phenotype, ecotype and disease. Furthermore, project relevance and availability information is now included. GOLD is available at http://www.genomesonline.org. It is also mirrored at the Institute of Molecular Biology and Biotechnology, Crete, Greece at http://gold.imbb.forth.gr/

Databases, Nucleic Acid↗

PubMed2006

Numbers and locations of native bacteria on field-grown wheat roots quantified by fluorescence in situ hybridization (FISH).

Native bacteria, Pseudomonas and filamentous bacteria were quantified and localized on wheat roots grown in the field using fluorescence in situ hybridization (FISH). Seminal roots were sampled through the season from unploughed soil in a conservation farming system. Such soils are spatially heterogeneous, and many roots grow slowly through hard soil with cracks and pores containing dead roots remnant from previous crops. Root and rhizosphere morphology, and contact with soil particles were preserved, and autofluorescence was avoided by observing sections in the far-red with Cy5 and Cy5.5 fluorochromes. Spatial analyses showed that bacteria were embedded in a stable matrix (biofilm) within 11 microm of the root surface (range 2-30 microm) and were clustered on 40% of roots. Half the clusters co-located with axial grooves between epidermal cells, soil particles, cap cells or root hairs; the other half were not associated with visible features. Across all wheat roots, although variable, bacteria averaged 15.4 x 10(5) cells per mm(3) rhizosphere, and of these, Pseudomonas and filaments comprised 10% and 4%, respectively, with minor effects of sample time, and no effect of plant age. Root caps were most heavily colonized by bacteria along roots, and elongation zones least heavily colonized. Pseudomonas varied little with root development and were 17% of bacteria on the elongation zone. Filamentous bacteria were not found on the elongation zone. The most significant factor to rhizosphere populations along a wheat root, however, was contact with dead root remnants, where Pseudomonas were reduced but filaments increased to 57% of bacteria (P < 0.001). This corresponded with analyses of root remnants showing they were heavily colonized by bacteria, with 48% filaments (P < 0.001) and 1.4%Pseudomonas (P = 0.014). Efforts to manage rhizosphere bacteria for sustainable agricultural systems should continue to focus on root cap and mucilage chemistry, and remnant roots as sources of beneficial bacteria.

Agriculture↗

PubMed2006

Non-sulfate-reducing, syntrophic bacteria affiliated with desulfotomaculum cluster I are widely distributed in methanogenic environments.

The classical perception of members of the gram-positive Desulfotomaculum cluster I as sulfate-reducing bacteria was recently challenged by the isolation of new representatives lacking the ability for anaerobic sulfate respiration. For example, the two described syntrophic propionate-oxidizing species of the genus Pelotomaculum form the novel Desulfotomaculum subcluster Ih. In the present study, we applied a polyphasic approach by using cultivation-independent and culturing techniques in order to further characterize the occurrence, abundance, and physiological properties of subcluster Ih bacteria in low-sulfate, methanogenic environments. 16S rRNA (gene)-based cloning, quantitative fluorescence in situ hybridization, and real-time PCR analyses showed that the subcluster Ih population composed a considerable part of the Desulfotomaculum cluster I community in almost all samples examined. Additionally, five propionate-degrading syntrophic enrichments of subcluster Ih bacteria were successfully established, from one of which the new strain MGP was isolated in coculture with a hydrogenotrophic methanogen. None of the cultures analyzed, including previously described Pelotomaculum species and strain MGP, consumed sulfite, sulfate, or organosulfonates. In accordance with these phenotypic observations, a PCR-based screening for dsrAB (key genes of the sulfate respiration pathway encoding the alpha and beta subunits of the dissimilatory sulfite reductase) of all enrichments/(co)cultures was negative with one exception. Surprisingly, strain MGP contained dsrAB, which were transcribed in the presence and absence of sulfate. Based on these and previous findings, we hypothesize that members of Desulfotomaculum subcluster Ih have recently adopted a syntrophic lifestyle to thrive in low-sulfate, methanogenic environments and thus have lost their ancestral ability for dissimilatory sulfate/sulfite reduction.

Culture Media↗

PubMed2005

SNP-VISTA: an interactive SNP visualization tool.

BACKGROUND: Recent advances in sequencing technologies promise to provide a better understanding of the genetics of human disease as well as the evolution of microbial populations. Single Nucleotide Polymorphisms (SNPs) are established genetic markers that aid in the identification of loci affecting quantitative traits and/or disease in a wide variety of eukaryotic species. With today's technological capabilities, it has become possible to re-sequence a large set of appropriate candidate genes in individuals with a given disease in an attempt to identify causative mutations. In addition, SNPs have been used extensively in efforts to study the evolution of microbial populations, and the recent application of random shotgun sequencing to environmental samples enables more extensive SNP analysis of co-occurring and co-evolving microbial populations. The program is available at http://genome.lbl.gov/vista/snpvista1. RESULTS: We have developed and present two modifications of an interactive visualization tool, SNP-VISTA, to aid in the analyses of the following types of data: A. Large-scale re-sequence data of disease-related genes for discovery of associated and/or causative alleles (GeneSNP-VISTA). B. Massive amounts of ecogenomics data for studying homologous recombination in microbial populations (EcoSNP-VISTA). The main features and capabilities of SNP-VISTA are: 1) mapping of SNPs to gene structure; 2) classification of SNPs, based on their location in the gene, frequency of occurrence in samples and allele composition; 3) clustering, based on user-defined subsets of SNPs, highlighting haplotypes as well as recombinant sequences; 4) integration of protein evolutionary conservation visualization; and 5) display of automatically calculated recombination points that are user-editable. CONCLUSION: The main strength of SNP-VISTA is its graphical interface and use of visual representations, which support interactive exploration and hence better understanding of large-scale SNP data by the user.

Algorithms↗

PubMed2005

Comparative metagenomics of microbial communities.

The species complexity of microbial communities and challenges in culturing representative isolates make it difficult to obtain assembled genomes. Here we characterize and compare the metabolic capabilities of terrestrial and marine microbial communities using largely unassembled sequence data obtained by shotgun sequencing DNA isolated from the various environments. Quantitative gene content analysis reveals habitat-specific fingerprints that reflect known characteristics of the sampled environments. The identification of environment-specific genes through a gene-centric comparative analysis presents new opportunities for interpreting and diagnosing environments.

Animals↗

PubMed2005

Genome-directed isolation of the key nitrogen fixer Leptospirillum ferrodiazotrophum sp. nov. from an acidophilic microbial community.

Analysis of assembled random shotgun sequence data from a low-diversity, subsurface acid mine drainage (AMD) biofilm revealed a single nif operon. This was found on a genome fragment belonging to a member of Leptospirillum group III, a lineage in the Nitrospirae phylum with no cultivated representatives. Based on the prediction that this organism is solely responsible for nitrogen fixation in the community, we pursued a selective isolation strategy to obtain the organism in pure culture. An AMD biofilm sample naturally abundant in Leptospirillum group III cells was homogenized, filtered, and serially diluted into a nitrogen-free liquid medium. The resulting culture in the terminal dilution grew autotrophically to a maximum cell density of approximately 10(6) cells/ml, oxidizing ferrous iron as the sole energy source. 16S rRNA-internal transcribed spacer region clone library analysis confirmed that the isolate is a member of Leptospirillum group III and that the culture is axenic. We propose the name Leptospirillum ferrodiazotrophum sp. nov. for this iron-oxidizing, free-living diazotroph. This study highlights how environmental sequence data can provide insights for culturing previously uncultured microorganisms.

Bacteria↗

PubMed2004

Bellerophon: a program to detect chimeric sequences in multiple sequence alignments.

SUMMARY: Bellerophon is a program for detecting chimeric sequences in multiple sequence datasets by an adaption of partial treeing analysis. Bellerophon was specifically developed to detect 16S rRNA gene chimeras in PCR-clone libraries of environmental samples but can be applied to other nucleotide sequence alignments. AVAILABILITY: Bellerophon is available as an interactive web server at http://foo.maths.uq.edu.au/~huber/bellerophon.pl

Algorithms↗

PubMed2004

Community structure and metabolism through reconstruction of microbial genomes from the environment.

Microbial communities are vital in the functioning of all ecosystems; however, most microorganisms are uncultivated, and their roles in natural systems are unclear. Here, using random shotgun sequencing of DNA from a natural acidophilic biofilm, we report reconstruction of near-complete genomes of Leptospirillum group II and Ferroplasma type II, and partial recovery of three other genomes. This was possible because the biofilm was dominated by a small number of species populations and the frequency of genomic rearrangements and gene insertions or deletions was relatively low. Because each sequence read came from a different individual, we could determine that single-nucleotide polymorphisms are the predominant form of heterogeneity at the strain level. The Leptospirillum group II genome had remarkably few nucleotide polymorphisms, despite the existence of low-abundance variants. The Ferroplasma type II genome seems to be a composite from three ancestral strains that have undergone homologous recombination to form a large population of mosaic genomes. Analysis of the gene complement for each organism revealed the pathways for carbon and nitrogen fixation and energy generation, and provided insights into survival strategies in an extreme environment.

Archaea↗

PubMed2004

Reclassification of Sphaerobacter thermophilus from the subclass Sphaerobacteridae in the phylum Actinobacteria to the class Thermomicrobia (emended description) in the phylum Chloroflexi (emended description).

Sphaerobacter thermophilus was originally classified as the deepest branching member of the phylum Actinobacteria (high-G+C, Gram-positive bacteria) based on 16S rRNA gene comparative analysis. However, the analysis lacked suitable outgroups, and additional 16S rRNA gene sequences indicate that it is most closely related to Thermomicrobium roseum, which it also resembles phenotypically. Furthermore, both species are reproducibly affiliated with the phylum Chloroflexi (green non-sulfur bacteria), despite T. roseum currently being classified in its own phylum, the Thermomicrobia. Transfer of Sphaerobacter to the class Thermomicrobia, and transfer of the class Thermomicrobia to the phylum Chloroflexi, are proposed. Descriptions for the phylum Chloroflexi and the class Thermomicrobia are emended to reflect the proposed changes in classification.

Actinobacteria↗

PubMed2004

Use of stable-isotope probing, full-cycle rRNA analysis, and fluorescence in situ hybridization-microautoradiography to study a methanol-fed denitrifying microbial community.

A denitrifying microbial consortium was enriched in an anoxically operated, methanol-fed sequencing batch reactor (SBR) fed with a mineral salts medium containing methanol as the sole carbon source and nitrate as the electron acceptor. The SBR was inoculated with sludge from a biological nutrient removal activated sludge plant exhibiting good denitrification. The SBR denitrification rate improved from less than 0.02 mg of NO(3)(-)-N mg of mixed-liquor volatile suspended solids (MLVSS)(-1) h(-1) to a steady-state value of 0.06 mg of NO(3)(-)-N mg of MLVSS(-1) h(-1) over a 7-month operational period. At this time, the enriched microbial community was subjected to stable-isotope probing (SIP) with [(13)C]methanol to biomark the DNA of the denitrifiers. The extracted [(13)C]DNA and [(12)C]DNA from the SIP experiment were separately subjected to full-cycle rRNA analysis. The dominant 16S rRNA gene phylotype (group A clones) in the [(13)C]DNA clone library was closely related to those of the obligate methylotrophs Methylobacillus and Methylophilus in the order Methylophilales of the Betaproteobacteria (96 to 97% sequence identities), while the most abundant clone groups in the [(12)C]DNA clone library mostly belonged to the family Saprospiraceae in the Bacteroidetes phylum. Oligonucleotide probes for use in fluorescence in situ hybridization (FISH) were designed to specifically target the group A clones and Methylophilales (probes DEN67 and MET1216, respectively) and the Saprospiraceae clones (probe SAP553). Application of these probes to the SBR biomass over the enrichment period demonstrated a strong correlation between the level of SBR denitrification and relative abundance of DEN67-targeted bacteria in the SBR community. By contrast, there was no correlation between the denitrification rate and the relative abundances of the well-known denitrifying genera Hyphomicrobium and Paracoccus or the Saprospiraceae clones visualized by FISH in the SBR biomass. FISH combined with microautoradiography independently confirmed that the DEN67-targeted cells were the dominant bacterial group capable of anoxic [(14)C]methanol uptake in the enriched biomass. The well-known denitrification lag period in the methanol-fed SBR was shown to coincide with a lag phase in growth of the DEN67-targeted denitrifying population. We conclude that Methylophilales bacteria are the dominant denitrifiers in our SBR system and likely are important denitrifiers in full-scale methanol-fed denitrifying sludges.

Autoradiography↗

PubMed2004

Chthoniobacter flavus gen. nov., sp. nov., the first pure-culture representative of subdivision two, Spartobacteria classis nov., of the phylum Verrucomicrobia.

The phylum Verrucomicrobia is increasingly recognized as an environmentally significant group of bacteria, particularly in soil habitats. At least six subdivisions of the Verrucomicrobia are resolved by comparative analysis of 16S rRNA genes, mostly obtained directly from environmental samples. To date, only two of these subdivisions (1 and 4) have characterized pure-culture representatives. We have isolated and characterized the first known pure-culture representative of subdivision 2. Strain Ellin428 is an aerobic heterotrophic bacterium that is able to grow with many of the saccharide components of plant biomass but does not grow with amino acids or organic acids other than pyruvate. Cells are yellow, rod-shaped, nonmotile, and gram-stain negative, and they contain peptidoglycan with direct cross-linkages of the A1 gamma meso-Dpm type. The isolate grows well at 25 degrees C on a variety of standard biological media, including some used in the routine cultivation of bacteria from soil. The pH range for growth is 4.0 to 7.0. Low levels of menaquinones MK-10 and MK-11 were detected. The major cellular fatty acids are C(14:0), a-C(15:0), C(16:1 omega 7c), and/or 2OH i-C(15:0), and C(16:0). The G+C content of the genomic DNA is 61 mol%. We propose a new genus and species, Chthoniobacter flavus gen. nov., sp. nov., with isolate Ellin428 as the type strain, and a new class for the subdivision to which it belongs, Spartobacteria classis nov. Environmental sequences indicate that the class Spartobacteria is largely represented by globally distributed, abundant, and active soil bacteria.

Bacteria↗

PubMed2004

Liquid serial dilution is inferior to solid media for isolation of cultures representative of the phylum-level diversity of soil bacteria.

Representatives of only four well-characterized bacterial phyla were isolated from a pasture soil by using liquid serial dilution culture. In contrast, members of Acidobacteria, Verrucomicrobia, and Gemmatimonadetes and of other poorly represented bacterial lineages were isolated in earlier experiments with solidified versions of the same media. We conclude that, contrary to expectation, liquid serial dilution culture is inferior to culturing on solid media for isolating representatives of many bacterial phyla from soil.

Bacteria↗