During screening for biosurfactant-producing, n-alkane-degrading marine bacteria, six heterotrophic bacterial strains were isolated from enriched mixed cultures, obtained from sea waterkediment samples collected near the Isle of Borkum (North Sea), using Mihagol-S (C,,,,-n-al kanes)as principal carbon source. These Gram-negative, aerobic, rod-shaped bacteria use a limited number of organic compounds, including aliphatic hydrocarbons, volatile fatty acids, and pyruvate and its methyl ether. During cultivation on n-alkanes as sole source of carbon and energy, all strains produced both extracellular and cell-bound surface-active glucose lipids which reduced the surface tension of water from 72 to 29 mN m-' (16). This novel class of glycolipids was found to be produced only by these strains. The 165 rRNA gene sequence analysis showed that these strains are all members of the y-subclass of the Proteobacteria. Their phospholipid ester-linked fatty acid composition was shown to be similar to that of members of the genus Halomonas, although they did not demonstrate a close phylogenetic relationship to any previously described species. On the basis of the information summarized above, a new genus and species, Akaniworax borkumensis, is described to include these bacteria. Strain SKZT is the type strain of A. borkumensis.

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We describe the first freshwater members of the class Actinobacteria that have been isolated. Nine ultramicro-size (<0.1 m 3 ) strains were isolated from five freshwater habitats in Europe and Asia. These habitats represent a broad spectrum of ecosystems, ranging from deep oligotrophic lakes to shallow hypertrophic lakes. Even when the isolated strains were grown in very rich media, the cell size was <0.1 m 3 and was indistinguishable from the cell sizes of bacteria belonging to the smaller size classes of natural lake bacterioplankton. Hybridization of the isolates with oligonucleotide probes and phylogenetic analysis of the 16S rRNA gene sequences of the isolated strains revealed that they are affiliated with the class Actinobacteria and the family Microbacteriaceae. The previously described species with the highest levels of sequence similarity are Clavibacter michiganensis and Rathayibacter tritici, two phytopathogens of terrestrial plants. The 16S rRNA gene sequences of the nine isolates examined are more closely related to cloned sequences from uncultured freshwater bacteria than to the sequences of any previously isolated bacteria. The nine ultramicrobacteria isolated form, together with several uncultured bacteria, a diverse phylogenetic cluster (Luna cluster) consisting exclusively of freshwater bacteria. Isolates obtained from lakes that are ecologically different and geographically separated by great distances possess identical 16S rRNA gene sequences but have clearly different ecophysiological and phenotypic traits. Predator-prey experiments demonstrated that at least one of the ultramicro-size isolates is protected against predation by the bacterivorous nanoflagellate Ochromonas sp. strain DS.The majority of the bacterial cells present in the bacterioplankton of marine and freshwater habitats are small (cell length, Ͻ1.5 m; volume, Ͻ0.3 m 3 ). Even very small bacteria, which are termed ultramicrobacteria (Ͻ0.1 m 3 ), are always present, and they frequently represent the numerically predominant fraction of typical marine and freshwater bacterioplankton. In contrast, in culture the cells of almost all bacteria isolated from bacterioplankton are much larger (cell length, Ͼ1.5 m; volume, Ͼ0.3 m 3 ) than most planktonic bacteria. In the case of several normally large-cell-size bacterial species, it has been shown that under strong starvation conditions the cell size decreases to dimensions typical of the majority of planktonic bacteria (23,26,27,31). There are only a few bacterial strains that have been isolated from marine and soil habitats whose cells are in the ultramicro size range (volume, Ͻ0.1 m 3 ) during growth under lab conditions (4,6,19,20,29,32,33,38). The most intensively investigated ultramicrobacterium is Sphingopyxis alaskensis (formerly Sphingomonas alsakensis [10,37]), a marine bacterium belonging to the class Alphaproteobacteria. Strains of this species have been isolated from Resurrection Bay, Alaska, from the North Sea, and from coastal waters near Japan (6,32,33). Recently, Rappé et...

Many heterotrophic bacteria have the ability to make polyhedral structures containing metabolic enzymes that are bounded by a unilamellar protein shell (metabolosomes or enterosomes). These bacterial organelles contain enzymes associated with a specific metabolic process (e.g. 1,2-propanediol or ethanolamine utilization). We show that the 21 gene regulon specifying the pdu organelle and propanediol utilization enzymes from Citrobacter freundii is fully functional when cloned in Escherichia coli, both producing metabolosomes and allowing propanediol utilization. Genetic manipulation of the level of specific shell proteins resulted in the formation of aberrantly shaped metabolosomes, providing evidence for their involvement as delimiting entities in the organelle. This is the first demonstration of complete recombinant metabolosome activity transferred in a single step and supports phylogenetic evidence that the pdu genes are readily horizontally transmissible. One of the predicted shell proteins (PduT) was found to have a novel Fe-S center formed between four protein subunits. The recombinant model will facilitate future experiments establishing the structure and assembly of these multiprotein assemblages and their fate when the specific metabolic function is no longer required.It has been recognized for more than 30 years that all cyanobacteria (1) and some other chemoautotrophic bacteria (2) contain carboxysomes. These polyhedral cellular inclusions consist of a proteinaceous shell enclosing an active enzyme, ribulose bisphosphate carboxylase/oxygenase (RuBisCO). 4Their function is to enhance the fixation of carbon dioxide (3), a reaction of planetary significance in that marine cyanobacteria are responsible for the majority of global carbon fixation (4, 5). More recently, sequence similarity was noticed between carboxysome shell genes and metabolic operon genes associated with propanediol utilization (pdu) and ethanolamine utilization (eut) in a variety of heterotrophic bacteria found in the mammalian gut (3) and the environment. In growth conditions that induce these metabolic operons, polyhedral organelles resembling carboxysomes were observed on electron microscopy of Salmonella enterica serovar Typhimurium (6), Klebsiella oxytoca, Citrobacter freundii, and Escherichia coli (7). Bioinformatics analysis also locates genes resembling carboxysome shell genes in metabolic operons in Clostridium perfringens (8), Clostridium tetani (9), Listeria monocytogenes and Listeria innocua (10), Enterococcus faecalis (11), Lactobacillus collinoides (12), Citrobacter rodentium, 5 and Yersinia enterocolitica (13) among other organisms. The non-carboxysome polyhedral structures have been referred to as enterosomes (3) or metabolosomes (14), emphasizing their role in cellular metabolism.There is some considerable interest in how these proteinaceous organelles form and the arrangement of protein subunits that give rise to these remarkable macromolecular assemblies. In carboxysomes, there are thought to be a number of shell prot...