Diversity and evolution of Bdellovibrio-and-like organisms (BALOs), reclassification of Bacteriovorax starrii as Peredibacter starrii gen. nov., comb. nov., and description of the Bacteriovorax-Peredibacter clade as Bacteriovoracaceae fam. nov. A phylogenetic analysis of Bdellovibrio-and-like organisms (BALOs) was performed. It was based on the characterization of 71 strains and on all consequent 16S rRNA gene sequences available in databases, including clones identified by data-mining, totalling 120 strains from very varied biotopes. Amplified rDNA restriction analysis (ARDRA) accurately reflected the diversity and phylogenetic affiliation of BALOs, thereby providing an efficient screening tool. Extensive phylogenetic analysis of the 16S rRNA gene sequences revealed great diversity within the Bdellovibrio (>14 % divergence) and Bacteriovorax (>16 %) clades, which comprised nine and eight clusters, respectively, exhibiting more than 3 % intra-cluster divergence. The clades diverged by more than 20 %. The analysis of conserved 16S rRNA secondary structures showed that Bdellovibrio contained motifs atypical of the d-Proteobacteria, suggesting that it is ancestral to Bacteriovorax. While none of the Bdellovibrio strains were of marine origin, Bacteriovorax included separate soil/freshwater and marine-specific groups. On the basis of their extensive diversity and the large distance separating the groups, it is proposed that Bacteriovorax starrii be placed into a new genus, Peredibacter gen. nov., with Peredibacter starrii A3.12 T (=ATCC 15145 T =NCCB 72004 T ) as its type strain. Also proposed is a redefinition of the Bdellovibrio and the Bacteriovorax-Peredibacter lineages as two different families, i.e. 'Bdellovibrionaceae' and a new family, Bacteriovoracaceae. Also, a re-evaluation of oligonucleotides targeting BALOs is presented, and the implications of the large diversity of these organisms and of their distribution in very different environments are discussed. Yaacov Davidov and Edouard Jurkevitch INTRODUCTIONThe obligate Gram-negative predatory bacteria belonging to the Bdellovibrio-and-like organisms (BALOs) are ubiquitously found in soil, in association with plant roots, in sea water, fresh water, biofilms and sewage Varon & Shilo, 1980;Williams et al., 1995). They were recently detected in the faeces of animals and humans (Schwudke et al., 2001). BALOs exhibit a unique biphasic life cycle in which a free-swimming cell infects the periplasmic space of a Gram-negative prey. Thereafter, the intracellular predator grows into a filament from which, at a later stage, progeny cells differentiate, lyse the host cell and start a new cycle (Jurkevitch, 2000). The molecular basis of this biphasic cycle and the determinants of the prey range (which varies greatly between BALO isolates) are unknown.BALOs consist of two poorly defined genera, Bdellovibrio and Bacteriovorax (Baer et al., 2000), which are affiliated with the d-Proteobacteria (Woese, 1987). Recently, the classification of these two genera in the family '...

Bdellovibrio-and-like organisms (BALOs) are widespread obligatory predators of other Gram-negative bacteria. Their detection by culture-dependent methods is complicated as their replication is totally dependent upon the availability of an appropriate prey. Because BALOs do not form numerically dominant groups within microbial communities, non-specific culture-independent tools also generally fail to detect them. We designed sets of 16S rRNA primers that specifically target BALOs. Polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) were combined, yielding partial 16S rDNA sequences. This simple method that allows specific in situ culture-independent detection of BALOs was applied to the soil environment. Bdellovibrio-and-like organisms were also isolated from the same soil and the phylogeny and prey range of the isolates analysed. Seventeen isolates retrieved using five different potential preys exhibited eight different spectra of prey utilization and formed nine operational taxonomic units (OTUs). These OTUs were affiliated with the Bdellovibrionaceae, Bacteriovorax, Peredibacter or Micavibrio, i.e. the known BALO groups. In comparison, 15 OTUs including 10 that were not detected by the culture-dependent approach were obtained using the specific primers in a PCR-DGGE approach. The occurrence of a complex BALO community suggests that predation occurs on a much wider range of prey than can be detected by the classical culture-dependent technique.

The life cycle, prey range and taxonomic status of a Bdellovibrio -like organism, strain JSST, were studied. Strain JSST was isolated from sewage in London, Ontario, Canada, in enrichment culture with Caulobacter crescentus prey cells. During predation, this strain remained attached to the outside of a stalked C. crescentus cell. No periplasmic growth stage was observed and no bdelloplast was formed. The stalked cells of C. crescentus retained their shape and, after predation, were devoid of cytoplasmic content, as shown by transmission electron microscopy. A periplasmic growth stage has been a definitive character in the description of members of the genera Bdellovibrio , Bacteriovorax , Bacteriolyticum and Peredibacter . This is the first description of an epibiotic predator in this group of prokaryotic predators. The G+C content of the genomic DNA of strain JSST was 46.1 mol%. 16S rRNA gene sequence analysis showed that this strain was related to Bdellovibrio bacteriovorus strains HD100T, 109J, 114 and 127 (90–93 % similarity). Phylogenetic analysis based on 16S rRNA gene sequences grouped strain JSST with the Bdellovibrio cluster, but at a distance from other Bdellovibrio isolates. On the basis of features of the life cycle and phylogenetic data, it was concluded that strain JSST merits classification as the type strain of a novel species, for which the name Bdellovibrio exovorus sp. nov. is proposed (type strain JSST  = ATCC BAA-2330T  = DSM 25223T).

Bdellovibrio-and-like organisms (BALOs) are peculiar, ubiquitous, small-sized, highly motile Gram-negative bacteria that are obligatory predators of other bacteria. Typically, these predators invade the periplasm of their prey where they grow and replicate. To date, BALOs constitute two highly diverse families affiliated with the delta-proteobacteria class. In this study, Micavibrio spp., a BALO lineage of epibiotic predators, were isolated from soil. These bacteria attach to digest and grow at the expense of other prokaryotes, much like other BALOs. Multiple phylogenetic analyses based on six genes revealed that they formed a deep branch within the alpha-proteobacteria, not affiliated with any of the alpha-proteobacterial orders. The presence of BALOs deep among the alpha-proteobacteria suggests that their peculiar mode of parasitism maybe an ancestral character in this proteobacterial class. The origin of the mitochondrion from an alpha-proteobacterium endosymbiont is strongly supported by molecular phylogenies. Accumulating data suggest that the endosymbiont's host was also a prokaryote. As prokaryotes are unable to phagocytose, the means by which the endosymbiont gained access into its host remains mysterious. We here propose a scenario based on the BALO feeding-mode to hypothesize a mechanism at play at the origin of the mitochondrial endosymbiosis.

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Accumulating data suggest that the eukaryotic cell originated from a merger of two prokaryotes, an archaeal host and a bacterial endosymbiont. However, since prokaryotes are unable to perform phagocytosis, the means by which the endosymbiont entered its host is an enigma. We suggest that a predatory or parasitic interaction between prokaryotes provides a reasonable explanation for this conundrum. According to the model presented here, the host in this interaction was an anaerobic archaeon with a periplasm-like space. The predator was a small (facultative) aerobic alpha-proteobacterium, which penetrated and replicated within the host periplasm, and later became the mitochondria. Plausible conditions under which this interaction took place and circumstances that may have led to the contemporary complex eukaryotic cell are discussed.

Inoculation with Azospirillum brasilense exerts beneficial effects on plant growth and crop yields. In this study, a comparative analysis of maize (Zea mays) root inoculated or not inoculated with A. brasilense strains was performed in two soils. Colonization dynamics of the rhizobacteria were tracked in various root compartments using 16S rRNA-targeted probes and 4',6'diamidino-2-phenylindole staining, and the structure of bacterial populations in the same samples was analyzed by denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction products of the 16S rRNA gene. Based on whole cell hybridization, a large fraction of the bacterial community was found to be active in both the rhizoplane-endorhizosphere and rhizosphere soil compartments, in both soil types. A DGGE fingerprint analysis revealed that plant inoculation with A. brasilense had no effect on the structural composition of the bacterial communities, which were also found to be very similar at the root tip and at zones of root branching. However, rhizobacterial populations were strongly influenced by plant age, and their complexity decreased in the rhizoplane-endorhizosphere in comparison to rhizosphere soil. A clone library generated from rhizosphere DNA revealed a highly diverse community of soil and rhizosphere bacteria, including an indigenous Azospirillum-like organism. A large proportion of these clones was only distantly related to known species.

The mechanism of function of the bacterial flagellar switch, which determines the direction of flagellar rotation and is essential for chemotaxis, has remained an enigma for many years. Here we show that the switch complex associates with the membrane-bound respiratory protein fumarate reductase (FRD). We provide evidence that FRD binds to preparations of isolated switch complexes, forms a 1:1 complex with the switch protein FliG, and that this interaction is required for both flagellar assembly and switching the direction of flagellar rotation. We further show that fumarate, known to be a clockwise/switch factor, affects the direction of flagellar rotation through FRD. These results not only uncover a new component important for switching and flagellar assembly, but they also reveal that FRD, an enzyme known to be primarily expressed and functional under anaerobic conditions in Escherichia coli, nonetheless, has important, unexpected functions under aerobic conditions.