The cell wall of the filamentous anoxygenic phototrophic bacterium Oscillochloris trichoides.

The filamentous anoxygenic phototrophic bacterium Oscillochloris trichoides DG-6 has been studied, and it has been shown that there are no lipopolysaccharides on the cell surface. Fatty acids hydroxylated at the C3 position, amino sugars and phosphate-containing compounds characteristic of lipid A have also not been found. The genes encoding for proteins responsible for the synthesis of lipopolysaccharides and the genes for the transport system, usually localized in the outer membrane of Gram-negative bacteria, have not been detected in the genome. The rigid layer of the cell wall contains a peptidoglycan consisting of alanine, glutamine, ornithine and glycine, in the respective ratio 1.8 : 1.5 : 1.0 : 0.6. Thus, the investigated bacterium, Osc. trichoides, is a monoderm. The cell wall also contains a branched α-1,4-d-glucan with a repeating unit consisting of glucose residues linked by α-1→4 bonds (α-1→6 at the branching sites). Such polymers have not previously been reported in phototrophic bacteria.

Nitrolancea hollandica gen. nov., sp. nov., a chemolithoautotrophic nitrite-oxidizing bacterium isolated from a bioreactor belonging to the phylum Chloroflexi.

A novel nitrite-oxidizing bacterium (NOB), strain Lb(T), was isolated from a nitrifying bioreactor with a high loading of ammonium bicarbonate in a mineral medium with nitrite as the energy source. The cells were oval (lancet-shaped) rods with pointed edges, non-motile, Gram-positive (by staining and from the cell wall structure) and non-spore-forming. Strain Lb(T) was an obligately aerobic, chemolitoautotrophic NOB, utilizing nitrite or formate as the energy source and CO2 as the carbon source. Ammonium served as the only source of assimilated nitrogen. Growth with nitrite was optimal at pH 6.8-7.5 and at 40 °C (maximum 46 °C). The membrane lipids consisted of C20 alkyl 1,2-diols with the dominant fatty acids being 10MeC18 and C(18 : 1)ω9. The peptidoglycan lacked meso-DAP but contained ornithine and lysine. The dominant lipoquinone was MK-8. Phylogenetic analyses of the 16s rRNA gene sequence placed strain Lb(T) into the class Thermomicrobia of the phylum Chloroflexi with Sphaerobacter thermophilus as the closest relative. On the basis of physiological and phylogenetic data, it is proposed that strain Lb(T) represents a novel species of a new genus, with the suggested name Nitrolancea hollandica gen. nov., sp. nov. The type strain of the type species is Lb(T) ( = DSM 23161(T) = UNIQEM U798(T)).

Cell envelope architecture in the Chloroflexi: a shifting frontline in a phylogenetic turf war.

It is important that attempts to understand bacterial phylogeny take into account fundamental bacterial characteristics such as cell envelope composition and organization. Several prominent phylogenetic studies have assumed that the cell envelopes of members of the phylum Chloroflexi are 'gram-negative' (diderm, i.e. defined by both an inner plasma membrane and an outer membrane) and some of these studies have placed the branch leading to the extant Chloroflexi near the root of the bacterial phylogenetic tree. This Correspondence summarizes the compelling evidence that the Chloroflexi are in fact monoderm, i.e. have only a single cellular membrane. The phylogenetic implications of this conclusion are discussed. The data reviewed also shed interesting light on the distribution of protein secretion systems in diderm bacteria.

A life cycle of branched aerial mycelium- and multiple budding spore-forming bacterium Thermosporothrix hazakensis belonging to the phylum Chloroflexi.

As far as known, sporulation modes in prokaryotes include formation of endospores exemplified by Firmicutes bacteria, myxospores by myxobacteria and arthrospores by actinomycetes. Here we describe Thermosporothrix hazakensis strain SK20-1(T) belonging to the phylum Chloroflexi with a life cycle including a novel prokaryotic sporulation mode. Microscopic observations showed that strain SK20-1(T) formed multiple exospores per mother cell by budding in branched aerial mycelia. Although branched aerial mycelia are characteristic of actinomycetes, multiple budding sporulation has not been previously described in prokaryotes. The strain SK20-1(T) could be a model microbe for cellular differentiation with multiple budding spore formation.

Early colonization of thermal niches in a silica-depositing hot spring in central Tibet.

Thermophilic microbial mats dominated by the anoxygenic phototroph Roseiflexus castenholzii commonly develop around sinter-depositing geysers in the Daggyai Tso geothermal field of central Tibet. In this study we used morphological and molecular genetic techniques to reveal a diverse pioneer biofilm community including both archaea and bacteria involved in early colonization of such thermal niches at temperatures ranging from 46 to 77 degrees C. Sinter precipitation and biomineralization were evident at all locations, but the latter was selective between taxa and most evident on filamentous cells. Evidence for possible indirect biosignatures from biofilms overwhelmed by sinter deposition was found. Succession to a mature community appeared to relate to the growth rate for key taxa outpacing that of silicification within an optimum temperature range of 54-61 degrees C. The thin surface layer of silicification-resistant cyanobacteria that developed on the surface of mature mats may play a role in preventing biomineralization of the susceptible R. castenholzii beneath within these communities.

Initial characterization of the photosynthetic apparatus of "Candidatus Chlorothrix halophila," a filamentous, anoxygenic photoautotroph.

"Candidatus Chlorothrix halophila" is a recently described halophilic, filamentous, anoxygenic photoautotroph (J. A. Klappenbach and B. K. Pierson, Arch. Microbiol. 181:17-25, 2004) that was enriched from the hypersaline microbial mats at Guerrero Negro, Mexico. Analysis of the photosynthetic apparatus by negative staining, spectroscopy, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the photosynthetic apparatus in this organism has similarities to the photosynthetic apparatus in both the Chloroflexi and Chlorobi phyla of green photosynthetic bacteria. The chlorosomes were found to be ellipsoidal and of various sizes, characteristics that are comparable to characteristics of chlorosomes in other species of green photosynthetic bacteria. The absorption spectrum of whole cells was dominated by the chlorosome bacteriochlorophyll c (BChl c) peak at 759 nm, with fluorescence emission at 760 nm. A second fluorescence emission band was observed at 870 nm and was tentatively attributed to a membrane-bound antenna complex. Fluorescence emission spectra obtained at 77 K revealed another complex that fluoresced at 820 nm, which probably resulted from the chlorosome baseplate complex. All of these results suggest that BChl c is present in the chlorosomes of "Ca. Chlorothrix halophila," that BChl a is present in the baseplate, and that there is a membrane-bound antenna complex. Analysis of the proteins in the chlorosomes revealed an approximately 6-kDa band, which was found to be related to the BChl c binding protein CsmA found in other green bacteria. Overall, the absorbance and fluorescence spectra of "Ca. Chlorothrix halophila" revealed an interesting mixture of photosynthetic characteristics that seemed to have properties similar to properties of both phyla of green bacteria when they were compared to the photosynthetic characteristics of Chlorobium tepidum and Chloroflexus aurantiacus.

Pigment analysis of "Candidatus Chlorothrix halophila," a green filamentous anoxygenic phototrophic bacterium.

The pigment composition of "Candidatus Chlorothrix halophila," a filamentous anoxygenic phototrophic bacterium found in Baja California Sur, Mexico, was determined. Previous work showed that bacteriochlorophyll c (BChl c) was the major pigment in "Ca. Chlorothrix halophila," but it was not clear if this bacterium also contains BChl a (J. A. Klappenbach and B. K. Pierson, Arch. Microbiol. 181:17-25, 2004). Here we show that in addition to BChl c, a small amount of a pigment that is spectrally indistinguishable from BChl a is present in cell extracts of "Ca. Chlorothrix halophila." Nevertheless, the BChl a-like pigment from "Ca. Chlorothrix halophila" has a different molecular weight and a different high-performance liquid chromatography elution time than BChl a from other photosynthetic bacteria. Based on mass spectrometry and other spectroscopic analysis, we determined that the BChl a-like pigment in "Ca. Chlorothrix halophila" contains a tetrahydrogeranylgeraniol tail rather than the phytol tail that is present in BChl a. The carotenoids and major BChl c homologs in "Ca. Chlorothrix halophila" were also identified. BChls c were found to be farnesol esterified and geranylgeraniol esterified.

[A comparative study of the fluorescence properties of the chlorosomal antenna of the green bacterium from the family Oscillochloridaceae and the members from two other families of green bacteria].

The fluorescence properties of bacteriochlorophylls (BChl) of the chlorosomal light-harvesting antenna of Oscillochloris trichoides (strain DG-6) from a new family of green filamentous bacteria Oscillochloridaceae were investigated in comparison with green bacteria from two other families. A strong dependence of the fluorescence intensity of chlorosomal bacteriochlorophyll c of Osc. trichoides on the redox potential of medium was found, which previously was observed only in green sulfur bacteria. The presence of BChl a in chlorosomes did not appear in their absorption spectra but was visualized by fluorescence spectroscopy at 77 K. From the comparative analysis of fluorescence spectral data for the chlorosomal light-harvesting antenna of Osc. trichoides and similar spectral data for green bacteria from two other families, it was concluded that, in some fluorescence spectral features (spectral position of bacteriochlorophyll c/a fluorescence bands; shape and full width at half maximum fluorescence band of chlorosomal bacteriochlorophyll c; the Stokes shift value of bacteriochlorophyll c band; a high molar ratio of bacteriochlorophyll c : bacteriochlorophyll a in chlorosomes that makes the bacteriochlorophyll a fluorescence band unresolved at room temperature; and highly redox-dependent fluorescence intensity of chlorosomal bacteriochlorophyll c), Osc. trichoides chlorosomes are close to the chlorosomal antenna of Chlorobiaceae species.

Phylogenetic and physiological characterization of a filamentous anoxygenic photoautotrophic bacterium ' Candidatus Chlorothrix halophila' gen. nov., sp. nov., recovered from hypersaline microbial mats.

We report the phylogenetic and physiological characterization of a mesophilic and halophilic member of the filamentous anoxygenic phototrophic (FAP) bacteria, provisionally named ' Candidatus Chorothrix halophila' gen. nov. sp. nov., that has been maintained in a highly enriched culture in our laboratory for over a decade. Phylogenetic analysis of small-subunit RNA-encoding sequences places ' Candidatus Chlorothrix halophila' in a clade that includes cultivated members of the genera Chloroflexus and Oscillochloris. Physiological studies demonstrated sulfide-dependent photosynthetic uptake of (14)C-labeled bicarbonate. Enzymatic assays for the activity of propionyl-coenzyme A synthase indicated that ' Candidatus Chlorothrix halophila' does not use the 3-hydroxypropionate cycle of Chloroflexus aurantiacus OK-70-fl for autotrophic carbon assimilation. New concepts regarding the taxonomy and phylogeny of FAP bacteria have emerged from this work.

Anaerolinea thermophila gen. nov., sp. nov. and Caldilinea aerophila gen. nov., sp. nov., novel filamentous thermophiles that represent a previously uncultured lineage of the domain Bacteria at the subphylum level.

Two thermophilic, Gram-negative, non-spore-forming, multicellular filamentous micro-organisms were isolated from thermophilic granular sludge in an upflow anaerobic sludge blanket reactor treating fried soybean-curd manufacturing waste water (strain UNI-1(T)) and from a hot spring sulfur-turf in Japan (strain STL-6-O1(T)). The filaments were longer than 100 microm and of 0.2-0.3 microm (strain UNI-1(T)) or 0.7-0.8 microm (strain STL-6-O1(T)) in width. Strain UNI-1(T) was a strictly anaerobic organism. The optimum temperature for growth was around 55 degrees C; growth occurred in the range 50-60 degrees C. The optimum pH for growth was around 7.0; growth occurred in the range pH 6.0-8.0. Strain STL-6-O1(T) was a facultatively aerobic bacterium. The optimum temperature for growth was around 55 degrees C; growth occurred in the range 37-65 degrees C. The optimum pH for growth was around 7.5-8.0; growth occurred in the range pH 7.0-9.0. The two organisms grew chemo-organotrophically on a number of carbohydrates and amino acids in the presence of yeast extract. The G+C content of the DNA of strains UNI-1(T) and STL-6-O1(T) was 54.5 and 59.0 mol%, respectively. Major cellular fatty acids for strain UNI-1(T) were C(16 : 0), C(15 : 0), C(14 : 0) and C(18 : 0), whereas those for strain STL-6-O1(T) were C(18 : 0), C(16 : 0), C(17 : 0) and iso-C(17 : 0). MK-10 was the major quinone from aerobically grown STL-6-O1(T) cells. Phylogenetic analyses based on 16S rDNA sequences revealed that both strains belong to an uncultured, previously recognized clone lineage of the phylum Chloroflexi (formerly known as green non-sulfur bacteria). These phenotypic and genetic properties suggested that each strain should be classified into a new independent genus; hence, the names Anaerolinea thermophila and Caldilinea aerophila are proposed for strains UNI-1(T) (=JCM 11387(T)=DSM 14523(T)) and STL-6-O1(T)(=JCM 11388(T)=DSM 14525(T)), respectively. These strains represent the type and sole species of the genera Anaerolinea and Caldilinea, respectively.

Characterization of the chlorosome antenna of the filamentous anoxygenic phototrophic bacterium Chloronema sp. strain UdG9001.

The absorption and fluorescence properties of chlorosomes of the filamentous anoxygenic phototrophic bacterium Chloronema sp. strain UdG9001 were analyzed. The chlorosome antenna of Chloronema consists of bacteriochlorophyll (BChl) d and BChl c together with gamma-carotene as the main carotenoid. HPLC analysis combined with APCI LC-MS/MS showed that the chlorosomal BChls comprise a highly diverse array of homologues that differ in both the degree of alkylation of the macrocycle at C-8 and/or C-12 and the alcohol moiety esterified to the propionic acid group at C-17. BChl c and BChl d from Chloronema were mainly esterified with geranylgeraniol (33% of the total), heptadecanol (24%), octadecenol (19%), octadecanol (14%), and hexadecenol (9%). Despite this pigment heterogeneity, fluorescence emission of the chlorosomes showed a single peak centered at 765 nm upon excitation at wavelengths ranging from 710 to 740 nm. This single emission, assigned to BChl c, indicates an energy transfer from BChl d to BChl c within the same chlorosome. Likewise, incubation of chlorosomes under reducing conditions caused a weak increase in fluorescence emission, which indicates a small redox-dependent fluorescence. Finally, protein analysis of Chloronema chlorosomes using SDS-PAGE and MALDI-TOF-MS revealed the presence of a chlorosomal polypeptide with a molecular mass of 5.7 kDa, resembling the CsmA protein found in Chloroflexus aurantiacus and Chlorobium tepidum chlorosomes. Several minor polypeptides were also detected but not identified. These results indicate that, compared with other members of filamentous anoxygenic phototrophic bacteria and green sulfur bacteria, Chloronema possesses an antenna system with novel features that may be of interest for further investigations.