John A. Fuerst

Missing lithotroph identified as new planctomycete

With the increased use of chemical fertilizers in agriculture, many densely populated countries face environmental problems associated with high ammonia emissions. The process of anaerobic ammonia oxidation (‘anammox’) is one of the most innovative technological advances in the removal of ammonia nitrogen from waste water,. This new process combines ammonia and nitrite directly into dinitrogen gas. Until now, bacteria capable of anaerobically oxidizing ammonia had never been found and were known as “lithotrophs missing from nature”. Here we report the discovery of this missing lithotroph and its identification as a new, autotrophic member of the order Planctomycetales, one of the major distinct divisions of the Bacteria. The new planctomycete grows extremely slowly, dividing only once every two weeks. At present, it cannot be cultivated by conventional microbiological techniques. The identification of this bacterium as the one responsible for anaerobic oxidation of ammonia makes an important contribution to the problem of unculturability.

New concepts of microbial treatment processes for the nitrogen removal in wastewater

Many countries strive to reduce the emissions of nitrogen compounds (ammonia, nitrate, NOx) to the surface waters and the atmosphere. Since mainstream domestic wastewater treatment systems are usually already overloaded with ammonia, a dedicated nitrogen removal from concentrated secondary or industrial wastewaters is often more cost-effective than the disposal of such wastes to domestic wastewater treatment. The cost-effectiveness of separate treatment has increased dramatically in the past few years, since several processes for the biological removal of ammonia from concentrated waste streams have become available. Here, we review those processes that make use of new concepts in microbiology: partial nitrification, nitrifier denitrification and anaerobic ammonia oxidation (the anammox process). These processes target the removal of ammonia from gases, and ammonium-bicarbonate from concentrated wastewaters (i.e. sludge liquor and landfill leachate). The review addresses the microbiology, its consequences for their application, the current status regarding application, and the future developments.

Endocytosis-like protein uptake in the bacterium Gemmata obscuriglobus

Endocytosis is a process by which extracellular material such as macromolecules can be incorporated into cells via a membrane-trafficking system. Although universal among eukaryotes, endocytosis has not been identified in Bacteria or Archaea. However, intracellular membranes are known to compartmentalize cells of bacteria in the phylum Planctomycetes, suggesting the potential for endocytosis and membrane trafficking in members of this phylum. Here we show that cells of the planctomycete Gemmata obscuriglobus have the ability to uptake proteins present in the external milieu in an energy-dependent process analogous to eukaryotic endocytosis, and that internalized proteins are associated with vesicle membranes. Occurrence of such ability in a bacterium is consistent with autogenous evolution of endocytosis and the endomembrane system in an ancestral noneukaryote cell.

Discovery of a New Source of Rifamycin Antibiotics in Marine Sponge Actinobacteria by Phylogenetic Prediction

ABSTRACT Phylogenetic analysis of the ketosynthase (KS) gene sequences of marine sponge-derived Salinispora strains of actinobacteria indicated that the polyketide synthase (PKS) gene sequence most closely related to that of Salinispora was the rifamycin B synthase of Amycolatopsis mediterranei. This result was not expected from taxonomic species tree phylogenetics using 16S rRNA sequences. From the PKS sequence data generated from our sponge-derived Salinispora strains, we predicted that such strains might synthesize rifamycin-like compounds. Liquid chromatography-tandem mass spectrometry (LC/MS/MS) analysis was applied to one sponge-derived Salinispora strain to test the hypothesis of rifamycin synthesis. The analysis reported here demonstrates that this Salinispora isolate does produce compounds of the rifamycin class, including rifamycin B and rifamycin SV. A rifamycin-specific KS primer set was designed, and that primer set increased the number of rifamycin-positive strains detected by PCR screening relative to the number detectable using a conserved KS-specific set. Thus, the Salinispora group of actinobacteria represents a potential new source of rifamycins outside the genus Amycolatopsis and the first recorded source of rifamycins from marine bacteria.

Linking Ultrastructure and Function in Four Genera of Anaerobic Ammonium-Oxidizing Bacteria: Cell Plan, Glycogen Storage, and Localization of Cytochrome c Proteins

Anaerobic ammonium oxidation (anammox) is an ecologically and industrially important process and is performed by a clade of deeply branching Planctomycetes. Anammox bacteria possess an intracytoplasmic membrane-bounded organelle, the anammoxosome. In the present study, the ultrastructures of four different genera of anammox bacteria were compared with transmission electron microscopy and electron tomography. The four anammox genera shared a common cell plan and contained glycogen granules. Differences between the four genera included cell size (from 800 to 1,100 nm in diameter), presence or absence of cytoplasmic particles, and presence or absence of pilus-like appendages. Furthermore, cytochrome c proteins were detected exclusively inside the anammoxosome. This detection provides further support for the hypothesis that this organelle is the locus of anammox catabolism.

Phylum Verrucomicrobia representatives share a compartmentalized cell plan with members of bacterial phylum Planctomycetes

BackgroundThe phylum Verrucomicrobia is a divergent phylum within domain Bacteria including members of the microbial communities of soil and fresh and marine waters; recently extremely acidophilic members from hot springs have been found to oxidize methane. At least one genus, Prosthecobacter, includes species with genes homologous to those encoding eukaryotic tubulins. A significant superphylum relationship of Verrucomicrobia with members of phylum Planctomycetes possessing a unique compartmentalized cell plan, and members of the phylum Chlamydiae including human pathogens with a complex intracellular life cycle, has been proposed. Based on the postulated superphylum relationship, we hypothesized that members of the two separate phyla Planctomycetes and Verrucomicrobia might share a similar ultrastructure plan differing from classical prokaryote organization.ResultsThe ultrastructure of cells of four members of phylum Verrucomicrobia – Verrucomicrobium spinosum, Prosthecobacter dejongeii, Chthoniobacter flavus, and strain Ellin514 – was examined using electron microscopy incorporating high-pressure freezing and cryosubstitution. These four members of phylum Verrucomicrobia, representing 3 class-level subdivisions within the phylum, were found to possess a compartmentalized cell plan analogous to that found in phylum Planctomycetes. Like all planctomycetes investigated, they possess a major pirellulosome compartment containing a condensed nucleoid and ribosomes surrounded by an intracytoplasmic membrane (ICM), as well as a ribosome-free paryphoplasm compartment between the ICM and cytoplasmic membrane.ConclusionA unique compartmentalized cell plan so far found among Domain Bacteria only within phylum Planctomycetes, and challenging our concept of prokaryote cell plans, has now been found in a second phylum of the Domain Bacteria, in members of phylum Verrucomicrobia. The planctomycete cell plan thus occurs in at least two distinct phyla of the Bacteria, phyla which have been suggested from other evidence to be related phylogenetically in the proposed PVC (Planctomycetes-Verrucomicrobia-Chlamydiae) superphylum. This planctomycete cell plan is present in at least 3 of 6 subdivisions of Verrucomicrobia, suggesting that the common ancestor of the verrucomicrobial phylum was also compartmentalized and possessed such a plan. The presence of this compartmentalized cell plan in both phylum Planctomycetes and phylum Verrucomicrobia suggest that the last common ancestor of these phyla was also compartmentalized.

Porphyrobacter neustonensis gen. nov., sp. nov., an aerobic bacteriochlorophyll-synthesizing budding bacterium from fresh water.

Four strains of orange- or red-pigmented bacteria isolated from freshwater surfaces were shown to synthesize bacteriochlorophyll under aerobic conditions. These strains shared unusual morphological features, such as acellular stalks, crateriformlike structures, and buds, with bacteria in the order Planctomycetales. However, comparisons of 16S rRNA sequences showed them to be members of the alpha-4 subdivision of the class Proteobacteria and most closely related to the marine aerobic bacteriochlorophyll-synthesizing bacterium Erythrobacter longus. They also differ from members of the Planctomycetales phenotypically in their synthesis of bacteriochlorophyll and possession of a peptidoglycan cell wall. They can be distinguished from E. longus on the basis of their 16S rRNA sequence, the G+C content of their DNA, cellular fatty acid composition, and carbon substrate spectrum. A new genus, Porphyrobacter, with a single species, P. neustonensis gen. nov., sp. nov., is proposed for these strains. The type strain is ACM 2844.

A comparison of five methods for assaying bacterial hydrophobicity

Five methods for assaying bacterial surface hydrophobicity, namely, bacterial adherence to hydrocarbons, salt aggregation, hydrophobic interaction chromatography, adhesion to polystyrene and latex particle agglutination were used to compare the hydrophobic surface properties of Escherichia coli, Acinetobacter calcoaceticus, Staphylococcus aureus and Streptococcus mitis. Two strains of A. calcoaceticus, including RAG-1, gave strong positive results by all five methods. S. mitis gave weak or negative results by all methods. The results for the other bacteria varied with the method. We conclude that reliance on one method for such tests is inadequate.

Isolation of Gemmata-like and Isosphaera-like planctomycete bacteria from soil and freshwater

ABSTRACT New cultured strains of the planctomycete division (order Planctomycetales) of the domain Bacteria related to species in the genera Gemmata and Isosphaera were isolated from soil, freshwater, and a laboratory ampicillin solution. Phylogenetic analysis of the 16S rRNA gene from eight representative isolates showed that all the isolates were members of the planctomycete division. Six isolates clustered with Gemmata obscuriglobus and related strains, while two isolates clustered with Isosphaera pallida. A double-membrane-bounded nucleoid was observed in Gemmata-related isolates but not in Isosphaera-related isolates, consistent with the ultrastructures of existing species of each genus. Two isolates from this study represent the first planctomycetes successfully cultivated from soil.

The anammox case-a new experimental manifesto for microbiological eco-physiology.

The anaerobic ammonium oxidation process is a new process for ammonia removal from wastewater. It is also a new microbial physiology that was previously believed to be impossible. The identification of Candidatus Brocadia anammoxidans and its relatives as the responsible bacteria was only possible with the development of a new experimental approach. That approach is the focus of this paper. The approach is a modernisation of the Winogradsky/Beyerinck strategy of selective enrichment and is based on the introduction of the molecular toolbox and modern bioreactor engineering to microbial ecology. It consists of five steps: (1) postulation of an ecological niche based on thermodynamic considerations and macro-ecological field data; (2) engineering of this niche into a laboratory bioreactor for enrichment culture; (3) black-box physiological characterisation of the enrichment culture as a whole; (4) phylogenetic characterisation of the enriched community using molecular tools; (5) physical separation of the dominant members of the enrichment culture using gradient centrifugation and the identification of the species of interest in accordance with Kochs postulates; (6) verification of the in situ importance of these species in the actual ecosystems. The power of this approach is illustrated with a case study: the identification of the planctomycetes responsible for anaerobic ammonium oxidation. We argue that this was impossible using molecular ecology or conventional ‘cultivation based techniques’ alone. We suggest that the approach might also be used for the microbiological study of many interesting microbes such as anaerobic methane oxidisers.