Magnetospirillum kuznetsovii sp. nov., a novel magnetotactic bacterium isolated from a lake in the Moscow region.

Strain LBB-42T was isolated from sediment sampled at Lake Beloe Bordukovskoe, located in the Moscow region (Russia). Phylogenetic analyses based on 16S rRNA gene sequencing results assigned the strain to the genus Magnetospirillum. Major fatty acids were C16 : 1ω7c, C16 : 0 and C18 : 1 ω9/C18 : 1 ω7. Genome sequencing revealed a genome size of 4.40 Mbp and a G+C content of 63.4 mol%. The average nucleotide identity and digital DNA-DNA hybridization values suggested that strain LBB-42T represents a new species, for which we propose the name Magnetospirillum kuznetsovii sp. nov., with the type strain LBB-42T (=VKM B-3270T=KCTC 15749T).

Magnetospirillum caucaseum sp. nov., Magnetospirillum marisnigri sp. nov. and Magnetospirillum moscoviense sp. nov., freshwater magnetotactic bacteria isolated from three distinct geographical locations in European Russia.

Three strains of helical, magnetotactic bacteria, SO-1T, SP-1T and BB-1T, were isolated from freshwater sediments collected from three distinct locations in European Russia. Phylogenetic analysis showed that the strains belong to the genus Magnetospirillum. Strains SO-1T and SP-1T showed the highest 16S rRNA gene sequence similarity to Magnetospirillum magnetotacticum MS-1T (99.3 and 98.1 %, respectively), and strain BB-1T with Magnetospirillum gryphiswaldense MSR-1T (97.3 %). The tree based on concatenated deduced amino acid sequences of the MamA, B, K, M, O, P, Q and T proteins, which are involved in magnetosome formation, was congruent with the tree based on 16S rRNA gene sequences. The genomic DNA G+C contents of strains SO-1T, SP-1T and BB-1T were 65.9, 63.0 and 65.2 mol%, respectively. As major fatty acids, C18 : 1ω9, C16 : 1ω7c, C16 : 0 and C18 : 0 were detected. DNA-DNA hybridization values between the novel strains and their closest relatives in the genus Magnetospirillum were less than 51.7 ± 2.3 %. In contrast to M. magnetotacticum MS-1T, the strains could utilize butyrate and propionate; strains SO-1T and BB-1T could also utilize glycerol. Strain SP-1T showed strictly microaerophilic growth, whereas strains SO-1T and BB-1T were more tolerant of oxygen. The results of DNA-DNA hybridization and physiological tests allowed genotypic and phenotypic differentiation of the strains from each other as well as from the two species of Magnetospirillum with validly published names. Therefore, the strains represent novel species, for which we propose the names Magnetospirillum caucaseum sp. nov. (type strain SO-1T = DSM 28995T = VKM B-2936T), Magnetospirillum marisnigri sp. nov. (type strain SP-1T = DSM 29006T = VKM B-2938T) and Magnetospirillum moscoviense sp. nov. (type strain BB-1T = DSM 29455T = VKM B-2939T).

Isolation and characterization of Magnetospirillum from saline lagoon.

Magnetotactic bacteria (MTB) are aquatic prokaryotes that orient themselves to earth's magnetic field with the help of intracellular organelle magnetosomes. Although many species of MTB have been identified, the isolation of MTB is a challenging task due to the lack of systematic isolation procedure and/or commercial media. In this study, we are reporting the isolation of magnetotactic spirillum from the Pulicat lagoon, India using a systematic and selective procedure. Sampling site was chosen on the basis of physicochemical properties of the ecosystem and the catalysed reporter deposition fluorescence in situ hybridization (CARD-FISH) analysis of sediment samples. In the current study, a combination of techniques including 'capillary racetrack' Purification and gradient cultivation resulted in the isolation of magnetotactic spirilla from aquatic sediments. Based on the 16S rRNA gene sequence analysis, the strain was identified as Magnetospirillum and was designated as Magnetospirillum sp. VITRJS1. The genes responsible for magnetosome formation (mamA, B, E, F, K, M, O, P, Q, T) were successfully detected using PCR amplification. The presence of cbbM gene confirmed that the isolate is chemolithoautotroph and utilises reduced sulphur as an electron source. Furthermore, magnetosomes extracted from VITRJS1 found to be cubo-octahedral in shape and 45 nm in size. Our results indicate that the systematic procedure using sediment analysis, CARD-FISH, and a combination of isolation methods enables the selective and rapid isolation of MTB from aquatic sediment sample.

Cytochrome cd1 nitrite reductase NirS is involved in anaerobic magnetite biomineralization in Magnetospirillum gryphiswaldense and requires NirN for proper d1 heme assembly.

The alphaproteobacterium Magnetospirillum gryphiswaldense synthesizes magnetosomes, which are membrane-enveloped crystals of magnetite. Here we show that nitrite reduction is involved in redox control during anaerobic biomineralization of the mixed-valence iron oxide magnetite. The cytochrome cd1-type nitrite reductase NirS shares conspicuous sequence similarity with NirN, which is also encoded within a larger nir cluster. Deletion of any one of these two nir genes resulted in impaired growth and smaller, fewer, and aberrantly shaped magnetite crystals during nitrate reduction. However, whereas nitrite reduction was completely abolished in the ΔnirS mutant, attenuated but significant nitrite reduction occurred in the ΔnirN mutant, indicating that only NirS is a nitrite reductase in M. gryphiswaldense. However, the ΔnirN mutant produced a different form of periplasmic d(1) heme that was not noncovalently bound to NirS, indicating that NirN is required for full reductase activity by maintaining a proper form of d1 heme for holo-cytochrome cd(1) assembly. In conclusion, we assign for the first time a physiological function to NirN and demonstrate that effective nitrite reduction is required for biomineralization of wild-type crystals, probably by contributing to oxidation of ferrous iron under oxygen-limited conditions.

Analysis of the CtrA pathway in Magnetospirillum reveals an ancestral role in motility in alphaproteobacteria.

Developmental events across the prokaryotic life cycle are highly regulated at the transcriptional and posttranslational levels. Key elements of a few regulatory networks are conserved among phylogenetic groups of bacteria, although the features controlled by these conserved systems are as diverse as the organisms encoding them. In this work, we probed the role of the CtrA regulatory network, conserved throughout the Alphaproteobacteria, in the magnetotactic bacterium Magnetospirillum magneticum strain AMB-1, which possesses unique intracellular organization and compartmentalization. While we have shown that CtrA in AMB-1 is not essential for viability, it is required for motility, and its putative phosphorylation state dictates the ability of CtrA to activate the flagellar biosynthesis gene cascade. Gene expression analysis of strains expressing active and inactive CtrA alleles points to the composition of the extended CtrA regulon, including both direct and indirect targets. These results, combined with a bioinformatic study of the AMB-1 genome, enabled the prediction of an AMB-1-specific CtrA binding site. Further, phylogenetic studies comparing CtrA sequences from alphaproteobacteria in which the role of CtrA has been experimentally examined reveal an ancestral role of CtrA in the regulation of motility and suggest that its essential functions in other alphaproteobacteria were acquired subsequently.

MamK, a bacterial actin, forms dynamic filaments in vivo that are regulated by the acidic proteins MamJ and LimJ.

Bacterial actins, in contrast to their eukaryotic counterparts, are highly divergent proteins whose wide-ranging functions are thought to correlate with their evolutionary diversity. One clade, represented by the MamK protein of magnetotactic bacteria, is required for the subcellular organization of magnetosomes, membrane-bound organelles that aid in navigation along the earth's magnetic field. Using a fluorescence recovery after photobleaching assay in Magnetospirillum magneticum AMB-1, we find that, like traditional actins, MamK forms dynamic filaments that require an intact NTPase motif for their turnover in vivo. We also uncover two proteins, MamJ and LimJ, which perform a redundant function to promote the dynamic behaviour of MamK filaments in wild-type cells. The absence of both MamJ and LimJ leads to static filaments, a disrupted magnetosome chain, and an anomalous build-up of cytoskeletal filaments between magnetosomes. Our results suggest that MamK filaments, like eukaryotic actins, are intrinsically stable and rely on regulators for their dynamic behaviour, a feature that stands in contrast to some classes of bacterial actins characterized to date.

Anaerobic degradation of 4-methylbenzoate via a specific 4-methylbenzoyl-CoA pathway.

The pathway for anaerobic degradation of 4-methylbenzoate was studied in the denitrifying alphaproteobacterium Magnetospirillum sp. strain pMbN1. Adaptation studies with whole cells indicated substrate-dependent induction of the capacity to degrade 4-methylbenzoate. Differential protein profiling (2D-DIGE) of 4-methylbenzoate- in comparison with benzoate- or succinate-adapted cells revealed the specific abundance increase of substrate-specific protein sets. Their coding genes form distinct clusters on the genome, two of which were assigned to 4-methylbenzoate and one to benzoate degradation. The predicted functions of the gene products agree with a specific 4-methylbenzoyl-CoA degradation pathway in addition to and analogous to the known anaerobic benzoyl-CoA degradation pathway. In vitro benzoyl-CoA and 4-methylbenzoyl-CoA reductase activities revealed the electron donor and ATP-dependent formation of the corresponding conjugated cyclic dienoyl-CoA/4-methyl-dienoyl-CoA products. The 4-methylbenzoyl-CoA reductase activity was induced in the presence of 4-methylbenzoate. In accordance, metabolite analysis of cultures grown with 4-methylbenzoate tentatively identified 4-methylcyclohex-1,5-diene-1-carboxylate. The 4-methylbenzoate induced genes were assigned to code for the putative 4-methylbenzoyl-CoA reductase; their products display pronounced sequence disparity from the conventional class I benzoyl-CoA reductase, which does not accept substituents at the para-position. Identification of 3-methylglutarate together with the formation of specific proteins for ring cleavage and ß-oxidation in 4-methylbenzoate-adapted cells suggest conservation of the methyl group along the specific 4-methylbenzoyl-CoA degradation pathway.

Magnetospirillum bellicus sp. nov., a novel dissimilatory perchlorate-reducing alphaproteobacterium isolated from a bioelectrical reactor.

Previously isolated dissimilatory perchlorate-reducing bacteria (DPRB) have been primarily affiliated with the Betaproteobacteria. Enrichments from the cathodic chamber of a bioelectrical reactor (BER) inoculated from creek water in Berkeley, CA, yielded a novel organism most closely related to a previously described strain, WD (99% 16S rRNA gene identity). Strain VDY(T) has 96% 16S rRNA gene identity to both Magnetospirillum gryphiswaldense and Magnetospirillum magnetotacticum, and along with strain WD, distinguishes a clade of perchlorate-reducing Magnetospirillum species in the Alphaproteobacteria. In spite of the phylogenetic location of VDY(T), attempted PCR for the key magnetosome formation genes mamI and mamL was negative. Strain VDY(T) was motile, non-spore forming, and, in addition to perchlorate, could use oxygen, chlorate, nitrate, nitrite, and nitrous oxide as alternative electron acceptors with acetate as the electron donor. Transient chlorate accumulation occurred during respiration of perchlorate. The organism made use of fermentation end products, such as acetate and ethanol, as carbon sources and electron donors for heterotrophic growth, and in addition, strain VDY(T) could grow chemolithotrophically with hydrogen serving as the electron donor. VDY(T) contains a copy of the RuBisCo cbbM gene, which was expressed under autotrophic but not heterotrophic conditions. DNA-DNA hybridization with strain WD confirmed VDY(T) as a separate species (46.2% identity), and the name Magnetospirillum bellicus sp. nov. (DSM 21662, ATCC BAA-1730) is proposed.

Characteristics and optimised fermentation of a novel magnetotactic bacterium, Magnetospirillum sp. ME-1.

Magnetotactic bacteria (MTB) can biosynthesise magnetosomes, which have great potential for commercial applications. A new MTB strain, Magnetospirillum sp. ME-1, was isolated and cultivated from freshwater sediments of East Lake (Wuhan, China) using the limiting dilution method. ME-1 had a chain of 17 ± 4 magnetosomes in the form of cubooctahedral crystals with a shape factor of 0.89. ME-1 was closest to Magnetospirillum sp. XM-1 according to 16S rRNA gene sequence similarity. Compared with XM-1, ME-1 possessed an additional copy of mamPA and a larger mamO in magnetosome-specific genes. ME-1 had an intact citric acid cycle, and complete pathway models of ammonium assimilation and dissimilatory nitrate reduction. Potential carbon and nitrogen sources in these pathways were confirmed to be used in ME-1. Adipate was determined to be used in the fermentation medium as a new kind of dicarboxylic acid. The optimised fermentation medium was determined by orthogonal tests. The large-scale production of magnetosomes was achieved and the magnetosome yield (wet weight) reached 120 mg L-1 by fed-batch cultivation of ME-1 at 49 h in a 10-L fermenter with the optimised fermentation medium. This study may provide insights into the isolation and cultivation of other new MTB strains and the production of magnetosomes.

Construction and preliminary analysis of a metagenomic library from a deep-sea sediment of east Pacific Nodule Province.

The Pacific Nodule Province is a unique ocean area containing an abundance of polymetallic nodules. To explore more genetic information and discover potentially industrial useful genes of the microbial community from this particular area, a cosmid library with an average insert of about 35 kb was constructed from the deep-sea sediment. The bacteria in the cosmid library were composed mainly of Proteobacteria including Alphaproteobacteria, Gammaproteobacteria and Deltaproteobacteria. The end sequences of some cosmid clones were determined and the complete insert sequences of two cosmid clones, 10D02 and 17H9, are presented. 10D02 has a length of 40.8 kb and contains 40 predicted encoding genes. It contains a partial 16S rRNA gene of Alphaproteobacteria. 17H9 is 36.8 kb and predicted to have 31 encoding genes and a 16S-23S-5S rRNA gene operon. Phylogenetic analysis of 16S and 23S rRNA gene sequence on the 17H9 both reveals that the inserted DNA from 17H9 came from a novel Alphaproteobacteria and is closely related to Magnetospirillum species. The predicted proteins of ORF 1-11 also have high identity to those of Magnetospirillum species, and the organization of these genes is highly conserved among known Magnetospirillum species. The data suggest that the retrieved DNA in 17H9 might be derived from a novel Magnetospirillum species.

In vitro assembly of the bacterial actin protein MamK from ' Candidatus Magnetobacterium casensis' in the phylum Nitrospirae.

Magnetotactic bacteria (MTB), a group of phylogenetically diverse organisms that use their unique intracellular magnetosome organelles to swim along the Earth's magnetic field, play important roles in the biogeochemical cycles of iron and sulfur. Previous studies have revealed that the bacterial actin protein MamK plays essential roles in the linear arrangement of magnetosomes in MTB cells belonging to the Proteobacteria phylum. However, the molecular mechanisms of multiple-magnetosome-chain arrangements in MTB remain largely unknown. Here, we report that the MamK filaments from the uncultivated 'Candidatus Magnetobacterium casensis' (Mcas) within the phylum Nitrospirae polymerized in the presence of ATP alone and were stable without obvious ATP hydrolysis-mediated disassembly. MamK in Mcas can convert NTP to NDP and NDP to NMP, showing the highest preference to ATP. Unlike its Magnetospirillum counterparts, which form a single magnetosome chain, or other bacterial actins such as MreB and ParM, the polymerized MamK from Mcas is independent of metal ions and nucleotides except for ATP, and is assembled into well-ordered filamentous bundles consisted of multiple filaments. Our results suggest a dynamically stable assembly of MamK from the uncultivated Nitrospirae MTB that synthesizes multiple magnetosome chains per cell. These findings further improve the current knowledge of biomineralization and organelle biogenesis in prokaryotic systems.

Exposure of magnetic bacteria to simulated mobile phone-type RF radiation has no impact on mortality.

The interaction of mobile phone RF emissions with biogenic magnetite in the human brain has been proposed as a potential mechanism for mobile phone bioeffects. This is of particular interest in light of the discovery of magnetite in human brain tissue. Previous experiments using magnetite-containing bacteria exposed directly to emissions from a mobile phone have indicated that these emissions might be causing greater levels of cell death in these bacterial populations when compared to sham exposures. A repeat of these experiments examining only the radio frequency (RF) global system for mobile communication (GSM) component of the mobile phone signal in a well-defined waveguide system (REFLEX), shows no significant change in cell mortality compared to sham exposures. A nonmagnetite containing bacterial cell strain (CC-26) with similar genotype and phenotype to the magnetotactic bacteria was used as a control. These also showed no significant change in cell mortality between RF and sham exposed samples. Results indicate that the RF components of mobile phone exposure do not appear to be responsible for previous findings indicating cell mortality as a result of direct mobile phone exposure. A further mobile phone emission component that should be investigated is the 2-Hz magnetic field pulse generated by battery currents during periods of discontinuous transmission.

[Geographical distribution of magnetotactic bacteria].

Magnetotactic bacteria (MTB) are widely distributed in aquatic environments. To assess the correlation between their evolutionary relatedness and geographic distribution, we analyzed 239 16S rDNA sequences available in the Genbank, and constructed phylogenetic trees based on the sequences. After elimination of redundant sequences by grouping those with identity > 97% into a single one, we analyzed in detail total 139 16S rDNA sequences, including 55 from marine MTB and 82 from freshwater sequences, and belonging to Proteobacteria and Nitrospirae. Phylogeny analysis based on those sequences suggests that the geographical distribution of MTB has certain regional distribution character: marine MTB is distinct from freshwater MTB, and off coast MTB are remotely related with ocean MTB. In contrast, the MTB from similar habitats, such as offshore in Brazil and the United States or freshwater lakes in Germany and China, are closely related. It is found that similar species have a large geographic distribution and tend to adopt the similar habitats, morphotypes of MTB and their living environment conditions have a significant relevance. This observation suggests that MTB may have multiple evolutionary origins. And also, it suggests the environmental conditions, as an important evolutionary pressure, play an important role in the long-term evolution of MTB.

Molecular mechanisms of magnetosome formation.

Magnetotactic bacteria are a diverse group of microorganisms with the ability to use geomagnetic fields for direction sensing. This unique feat is accomplished with the help of magnetosomes, nanometer-sized magnetic crystals surrounded by a lipid bilayer membrane and organized into chains via a dedicated cytoskeleton within the cell. Because of the special properties of these magnetic crystals, magnetotactic bacteria have been exploited for a variety of applications in diverse disciplines from geobiology to biotechnology. In addition, magnetosomes have served as a powerful model system for the study of biomineralization and cell biology in bacteria. This review focuses on recent advances in understanding the molecular mechanisms of magnetosome formation and magnetite biomineralization.