The cytoskeleton has a key function in the temporal and spatial organization of both prokaryotic and eukaryotic cells. Here, we report the identification of a new class of polymer-forming proteins, termed bactofilins, that are widely conserved among bacteria. In Caulobacter crescentus, two bactofilin paralogues cooperate to form a sheet-like structure lining the cytoplasmic membrane in proximity of the stalked cell pole. These assemblies mediate polar localization of a peptidoglycan synthase involved in stalk morphogenesis, thus complementing the function of the actin-like cytoskeleton and the cell division machinery in the regulation of cell wall biogenesis. In other bacteria, bactofilins can establish rod-shaped filaments or associate with the cell division apparatus, indicating considerable structural and functional flexibility. Bactofilins polymerize spontaneously in the absence of additional cofactors in vitro, forming stable ribbon-or rod-like filament bundles. Our results suggest that these structures have evolved as an alternative to intermediate filaments, serving as versatile molecular scaffolds in a variety of cellular pathways.

Insect intestinal tracts harbor several novel, deep-rooting clades of as-yet-uncultivated bacteria whose biology is typically completely unknown. Here, we report the isolation of the first representative of the termite group 1 (TG1) phylum from sterile-filtered gut homogenates of a humivorous scarab beetle larva. Strain Pei191T is a mesophilic, obligately anaerobic ultramicrobacterium with a gram-negative cell envelope. Cells are typically rod shaped, but cultures are pleomorphic in all growth phases (0.3 to 2.5 μm long and 0.17 to 0.3 μm wide). The isolate grows heterotrophically on sugars and ferments d-galactose, d-glucose, d-fructose, d-glucosamine, and N-acetyl-d-glucosamine to acetate, ethanol, hydrogen, and alanine as major products but only if amino acids are present in the medium. PCR-based screening and comparative 16S rRNA gene sequence analysis revealed that strain Pei191T belongs to the “intestinal cluster,” a lineage of hitherto uncultivated bacteria present in arthropod and mammalian gut systems. It is only distantly related to the previously described so-called “endomicrobia” lineage, which comprises mainly uncultivated endosymbionts of termite gut flagellates. We propose the name “Elusimicrobium minutum” gen. nov., sp. nov. (type strain, Pei191T = ATCC BAA-1559T = JCM 14958T) for the first isolate of this deep-branching lineage and the name “Elusimicrobia” phyl. nov. for the former TG1 phylum.

Phycobiliprotein aggregates were isolated from the prokaryote Acaryochloris marina, containing chlorophyll d as major pigment. In the electron microscope the biliprotein aggregates appear as rod-shaped structures of 26.0x11.3 nm, composed of four ring-shaped subunits 5.8 nm thick and 11.7 nm in diameter. Spectral data indicate that the aggregates contain two types of biliproteins: phycocyanin and an allophycocyanintype pigment, with very efficient energy transfer from the phycocyanin-to allophycocyanin-type constituent. The chromophore-binding polypeptides of the pigments have apparent molecular masses of 16.2 and 17.4 kDa. They crossreact with antibodies against phycocyanin and allophycocyanin from a red alga.

The use of dilution culture techniques to cultivate saccharolytic bacteria present in the anoxic soil of flooded rice microcosms allowed the isolation of three new strains of bacteria, typified by their small cell sizes, with culturable numbers estimated at between 1.2 ؋ 10 5 and 7.3 ؋ 10 5 cells per g of dry soil. The average cell volumes of all three strains were 0.03 to 0.04 m 3 , and therefore they can be termed ultramicrobacteria or "dwarf cells." The small cell size is a stable characteristic, even when the organisms grow at high substrate concentrations, and thus is not a starvation response. All three strains have genomic DNA with a mol% G؉C ratio of about 63, are gram negative, and are motile by means of a single flagellum. The three new isolates utilized only sugars and some sugar polymers as substrates for growth. The metabolism is strictly fermentative, but the new strains are oxygen tolerant. Sugars are metabolized to acetate, propionate, and succinate. Hydrogen production was not significant. In the presence of 0.2 atm of oxygen, the fermentation end products or ratios did not change. The phylogenetic analysis on the basis of 16S ribosomal DNA (rDNA) sequence comparisons indicates that the new isolates belong to a branch of the Verrucomicrobiales lineage and are closely related to a cloned 16S rDNA sequence (PAD7) recovered from rice paddy field soil from Japan. The isolation of these three strains belonging to the order Verrucomicrobiales from a model rice paddy system, in which rice was grown in soil from an Italian rice field, provides some information on the possible physiology and phenotype of the organism represented by the cloned 16S rDNA sequence PAD7. The new isolates also extend our knowledge on the phenotypic and phylogenetic depths of members of the order Verrucomicrobiales, to date acquired mainly from cloned 16S rDNA sequences from soils and other habitats.

Methyl-coenzyme M reductase (MCR) catalyses the methane-forming step in the energy metabolism of methanogenic Archaea. It brings about the reduction of methyl-coenzyme M (CH,-S-CoM) by 7-mercaptoheptanoylthreonine phosphate (H-S-HTP). Methanobacterium thermoautotrophicunz contains two isoenzymes of MCR, designated MCR I and MCR 11, which are expressed differentially under different conditions of growth. These two isoenzymes have been separated, purified and their catalytic and spectroscopic properties determined. Initial-velocity measurements of the twosubstrate reaction showed that the kinetic mechanism for both isoenzymes involved ternarycomplex formation. Double reciprocal plots of initial rates versus the concentration of either one of the two substrates at different constant concentrations of the other substrate were linear and intersected on AGO' = -45 kJImol. This methane-forming reaction is the terminal step in the energy metabolism of all methanogenic Archaea (Wolfe, 1991 ;Ferry, 1992;Weiss and Thauer, 1993). In addition, it is probably the rate-limiting step in methanogenesis. The specific activity of MCR, determined in cell extracts of methanogenic Archaea, is much lower than that of the other catabolic enzymes of the methane-forming pathway (Schworer and Thauer, 1991). The MCR concentration in methanogenic Archaea is relatively high. It can constitute more than 10% of the cellular protein (Rouvikre et al., 1988).It was discovered recently that Methanobacterium thermoautotrophicum, which grows on H, and C02, contains two isoenzymes of MCR, designated MCR I and MCR I1 (Rospert et al., 1990). The two isoenzymes, which can be separated by anion-exchange chromatography on a Mono-Q column, resemble each other in being composed of three different subunits in an (xzp2j~2 arrangement and in containing 2 mol of the nickel porphinoid F430 as tightly, but not covalently bound, prosthetic group. They differ significantly, however, in the size of the y subunit and in the amino acid sequences of all subunits, as revealed by a comparison of their N-terminal amino acid sequences and of their immunological properties. Southern-blot analyses indicate the presence of two related but clearly separate MCR operons in the genomic DNA (Reeve, 1992). Evidence has been presented that MCR I is the predominant form when growth of the methanogen is limited by the rate of the H, and CO, supply and that MCR I1 predominates when the concentration of H2 and of CO, is not growth-rate limiting (Bonacker et al., 1992). It was not clear why this Archaeon uses different iso-