Strain MSR-1 Research Articles

Study the effect of Enterobacter cloacae on the gene expression, productivity, and quality traits of Curcuma longa L. Plant.

Overuse of artificial chemical fertilizers could be detrimental to the environment. Utilizing beneficial microorganisms as biofertilizers is a sustainable technique that promotes soil health, crop yield, and ecosystem preservation. Curcuma longa L. is utilized as a medication since it has its antibacterial, anti-microbial, and anti-tumor characteristics, which reduce inflammation and hasten wound healing. The effect of E. cloacae strain MSR1, which is common in the roots of alfalfa grown in the Al-Ahsaa region, on C. longa plants is being investigated. C. longa rhizomes were planted under greenhouse conditions after being submerged in a solution of E. cloacae strain MSR1 (OD 500) or water treatment as a control for 12 hours. After 240 days of growing, ten randomly selected plants from each treatment were collected, and the vegetative growth and yield metrics were assessed. To investigate how E. cloacae influences C. longa production and chemical composition (photosynthetic pigment, nitrogen, phosphorus, potassium, and curcuminoid), measurements were conducted as well as genes diketide-CoA and curcumin synthases genes. Our research showed that C. longa's growth and yield were favorably impacted by E. cloacae. Significant increases in the related plants' chlorophyll a,b, carotenoid, nitrogen, phosphorus, and potassium levels were likewise a reflection of the enhanced effects shown in the growth and yield parameters. Treatment with E. cloacae raised the curcuminoid's three sub-components' compositions to varying degrees: bisdemethoxycurcumin, demethoxycurcumin, and curcumin. Comparing E. cloacae treated plants to the control, high expression levels of the genes diketide-CoA and curcumin synthase-1, -2, and 3 were also found. The treatment of E. cloacae is a good biostimulant candidate for boosting growth and yield as well as raising the medicinal qualities of C. longa, according to the overall results.

Electroactivity of the magnetotactic bacteria Magnetospirillum magneticum AMB-1 and Magnetospirillum gryphiswaldense MSR-1

Magnetotactic bacteria reside in sediments and stratified water columns. They are named after their ability to synthesize internal magnetic particles that allow them to align and swim along the Earth’s magnetic field lines. Here, we show that two magnetotactic species, Magnetospirillum magneticum strain AMB-1 and Magnetospirillum gryphiswaldense strain MSR-1, are electroactive. Both M. magneticum and M. gryphiswaldense were able to generate current in microbial fuel cells with maximum power densities of 27 and 11 µW/m2, respectively. In the presence of the electron shuttle resazurin both species were able to reduce the crystalline iron oxide hematite (Fe2O3). In addition, M. magneticum could reduce poorly crystalline iron oxide (FeOOH). Our study adds M. magneticum and M. gryphiswaldense to the growing list of known electroactive bacteria, and implies that electroactivity might be common for bacteria within the Magnetospirillum genus.

Effects of culture conditions on denitrification activity by Magnetospirillum gryphiswaldense strain MSR-1

The application of denitrification function of Magnetospirillum gryphiswaldense strain MSR-1 has not received attention. Here, we studied the effects of different culture conditions, including carbon (C) and nitrogen (N) sources, C/N ratio, iron source species and addition, initial pH, and temperature on the growth, magnetic production, and denitrification activity and application potential of MSR-1 under anoxic and nutrient environments similar to that of urban domestic sewage. Sodium lactate remains the best carbon source for denitrification by MSR-1, although acetic acid is also promising. Nitrate is the best nitrogen source, where addition and the ratio to nitrite should be >3 mM and 1, respectively. At C/N between 3 and 7, initial pH between 6.5 and 7.5, and culture temperature from 25 to 35 °C, nitrate content (84 mg N/L) decreased to the lowest value in 24 h, concomitant with the lowest residual nitrite. A maximum nitrate removal rate of 88.22 % was observed in 16 h by the optimization of response surface methodology with C/N = 5.13, initial pH = 6.83, and culture temperature = 30.3 °C. MSR-1 effectively removed nitrate, with the advantage of a low C/N demand; furthermore, optimal pH and temperature for denitrification are similar to those used in the sewage biological treatment system. Here, the consistency between optimal denitrification conditions of MSR-1 and actual wastewater conditions was considered for the first time, demonstrating the application potential of MSR-1 for wastewater denitrification and providing a theoretical basis for the application of the strain in wastewater treatment.

Progress in affinity ligand-functionalized bacterial magnetosome nanoparticles for bio-immunomagnetic separation of HBsAg protein

Efficient Bio-immunomagnetic separation (BIMS) of recombinant hepatitis B surface antigen (rHBsAg) with high binding capacity was studied using affinity ligand immobilized bacterial magnetosome nanoparticles (Magnetospirillum gryphiswaldense strain MSR-1 bacteria) as an immunomagnetic sorbent. Our results showed immunomagnetic adsorption, acted by affinity interactions with the immobilized monoclonal antibody, offered higher antigen adsorption and desorption capacities as compared with the commercially available immunoaffinity sorbents. Four different ligand densities of the Hep-1 monoclonal antibody were examined during covalent immobilization on Pyridyl Disulfide-functionalized magnetosome nanoparticles for HBsAg immunomagnetic separation. The average of adsorption capacity was measured as 3 mg/ml in optimized immunomagnetic sorbent (1.056 mg rHBsAg/ml immunomagneticsorbent/5.5 mg of total purified protein) and 5mg/ml in immunoaffinity sorbent (0.876 mg rHBsAg/ml immunosorbent/5.5 mg total purified protein during 8 runs. Immunomagnetic sorbent demonstrated ligand leakage levels below 3 ng Mab/Ag rHBsAg during 12 consecutive cycles of immunomagnetic separation (IMS). The results suggest that an immunomagnetic sorbent with a lower ligand density (LD = 3 mg Mab/ml matrix) could be the best substitute for the immunosorbent used in affinity purification of r-HBsAg there are significant differences in the ligand density (98.59% (p-value = 0.0182)), adsorption capacity (97.051% (p-value = 0.01834)), desorption capacity (96.06% (p-value = 0.036)) and recovery (98.97% (p-value = 0.0231)). This study indicates that the immunosorbent approach reduces the cost of purification of Hep-1 protein up to 50% as compared with 5 mg Mab/ml immunoaffinity sorbent, which is currently used in large-scale production. As well, these results demonstrate that bacterial magnetosome nanoparticles (BMs) represent a promising alternative product for the economical and efficient immobilization of proteins and the immunomagnetic separation of Biomolecules, promoting innovation in downstream processing.

Fusion expression of nanobodies specific for the insecticide fipronil on magnetosomes in Magnetospirillum gryphiswaldense MSR-1

BackgroundMagnetic nanoparticles such as magnetosomes modified with antibodies allow a high probability of their interaction with targets of interest. Magnetosomes biomineralized by magnetotactic bacteria are in homogeneous nanoscale size and have crystallographic structure, and high thermal and colloidal stability. Camelidae derived nanobodies (Nbs) are small in size, thermal stable, highly water soluble, easy to produce, and fusible with magnetosomes. We aimed to functionalize Nb-magnetosomes for the analysis of the insecticide fipronil.ResultsThree recombinant magnetotactic bacteria (CF, CF+ , and CFFF) biomineralizing magnetosomes with different abundance of Nbs displayed on the surface were constructed. Compared to magnetosomes from the wild type Magnetospirillum gryphiswaldense MSR-1, all of the Nb-magnetosomes biosynthesized by strains CF, CF+ , and CFFF showed a detectable level of binding capability to fipronil-horseradish peroxidase (H2-HRP), but none of them recognized free fipronil. The Nb-magnetosomes from CFFF were oxidized with H2O2 or a glutathione mixture consisting of reduced glutathione and oxidized glutathione in vitro and their binding affinity to H2-HRP was decreased, whereas that to free fipronil was enhanced. The magnetosomes treated with the glutathione mixture were employed to develop an enzyme-linked immunosorbent assay for the detection of fipronil in water samples, with average recoveries in a range of 78–101%.ConclusionsThe economical and environmental-friendly Nb-magnetosomes biomineralized by the bacterial strain MSR-1 can be potentially applied to nanobody-based immunoassays for the detection of fipronil or nanobody-based assays in general.

Novel Protein Mg2046 Regulates Magnetosome Synthesis in Magnetospirillum gryphiswaldense MSR-1 by Modulating a Proper Redox Status.

Magnetotactic bacteria (MTB) are a large, polyphyletic group of aquatic microorganisms capable of absorbing large amounts of iron and synthesizing intercellular nano-scaled nanoparticles termed magnetosomes. In our previous transcriptomic studies, we discovered that a novel gene (MGMSRv2_2046, termed as mg2046) in Magnetospirillum gryphiswaldense strain MSR-1 was significantly up-regulated during the period of magnetosome synthesis. In the present study, we constructed a MSR-1 mutant strain with deletion of mg2046 (termed Δmg2046) in order to evaluate the role of this gene in cell physiological status and magnetosome formation process. In comparison with wild-type MSR-1, Δmg2046 showed similar cell growth, but much lower cell magnetic response, smaller number and size of magnetosomes, and reduced iron absorption ability. mg2046 deletion evidently disrupted iron uptake, and redox equilibrium, and strongly inhibited transcription of dissimilatory denitrification pathway genes. Our experimental findings, taken together with results of gene homology analysis, indicate that Mg2046 acts as a positive regulator in MSR-1 under microaerobic conditions, responding to hypoxia signals and participating in regulation of oxygen metabolism, in part as a co-regulator of dissimilatory denitrification pathway with oxygen sensor MgFnr (MGMSRv2_2946, termed as Mg2946). Mg2046 is clearly involved in coupled regulation of cellular oxygen, iron and nitrogen metabolism under micro-aerobic or anaerobic conditions. Our findings help explain how MSR-1 cells initiate dissimilatory denitrification pathway and overcome energy deficiency under microaerobic conditions, and have broader implications regarding bacterial survival and energy metabolism strategies under hypoxia.

A Novel Highly Efficient Device for Growing Micro-Aerophilic Microorganisms.

This work describes a novel, simple and cost-effective culture system, named the Micro-Oxygenated Culture Device (MOCD), designed to grow microorganisms under particularly challenging oxygenation conditions. Two microaerophilic magnetotactic bacteria, a freshwater Magnetospirillum gryphiswaldense strain MSR-1 and a marine Magnetospira sp. strain QH-2, were used as biological models to prove the efficiency of the MOCD and to evaluate its specifications. Using the MOCD, growth rates of MSR-1 and QH-2 increased by four and twofold, respectively, when compared to traditional growing techniques using simple bottles. Oxystat-bioreactors have been typically used and specifically designed to control low dissolved oxygen concentrations, however, the MOCD, which is far less sophisticated was proven to be as efficient for both MSR-1 and QH-2 cultures with regard to growth rate, and even better for MSR-1 when looking at cell yield (70% increase). The MOCD enables a wide range of oxygenation conditions to be studied, including different O2-gradients. This makes it an innovative and ingenious culture device that opens up new parameters for growing microaerobic microorganisms.

Effects of Environmental Conditions on High-Yield Magnetosome Production by Magnetospirillum gryphiswaldense MSR-1

Background:Magnetotactic bacteria are a heterogeneous group of Gram-negative prokaryote cells that produce linear chains of magnetic particles called magnetosomes, intracellular organelles composed of magnetic iron particles. Many important applications have been defined for magnetic nanoparticles in biotechnology, such as cell separation applications, as well as acting as carriers of enzymes, antibodies, or anti-cancer drugs. Since the bacterial growth is difficult and the yield of magnetosome production is low, the application of magnetosome has not been developed on a commercial scale.Methods:Magnetospirillum gryphiswaldense strain MSR-1 was used in a modified current culture medium supplemented by different concentrations of oxygen, iron, carbon, and nitrogen, to increase the yield of magnetosomes.Results:Our improved MSR-1 culture medium increased magnetosome yield, magnetosome number per bacterial cell, magnetic response, and bacterial cell growth yield significantly. The yield of magnetosome increased approximately four times. The optimized culture medium containing 25 mM of Na-pyruvate, 40 mM of NaNO3, 200 µM of ferrous sulfate, and 5-10 ppm of dissolved oxygen (DO) resulted in 186.67 mg of magnetosome per liter of culture medium. The iron uptake increased significantly, and the magnetic response of the bacteria to the magnetic field was higher than threefold as compared to the previously reported procedures.Conclusion:This technique not only decreases the cultivation time but also reduces the production cost. In this modified method, the iron and DO are the major factors affecting the production of magnetosome by M. gryphiswaldense strain MSR-1. However, refining this technique will enable a further yield of magnetosome and cell density.

An Investigation of the Sensing Capabilities of Magnetotactic Bacteria.

We investigate the sensing capabilities of magnetotactic bacteria (Magnetospirillum gryphiswaldense strain MSR1) to MCF-7 breast cancer cells. Cancer cells are allowed to grow inside a capillary tube with depth of 200 $\mu \mathrm {m}$ and motion of magnetotactic bacteria is investigated under the influence of oxygen gradient and geomagnetic field. The influence of cancer cells is modeled to predict the oxygen gradient within the capillary tube in three-dimensional space. Our experimental motion analysis and count of motile magnetotactic bacteria indicate that they migrate towards less-oxygenated regions within the vicinity of cancer cells. Bands of magnetotactic bacteria with average concentration of 18.8±2.0% are observed in close proximity to MCF-7 cells $(h = 20~ \mu \mathrm {m})$, whereas the concentration at proximity of $190~ \mu \mathrm {m}$ is 5.0 ± 6.8%.

Reevaluation of the Complete Genome Sequence of Magnetospirillum gryphiswaldense MSR-1 with Single-Molecule Real-Time Sequencing Data.

ABSTRACTMagnetospirillum gryphiswaldense is a key organism for understanding magnetosome formation and magnetotaxis. As earlier studies suggested a high genomic plasticity, we (re)sequenced the type strain MSR-1 and the laboratory strain R3/S1. Both sequences differ by only 11 point mutations, but organization of the magnetosome island deviates from that of previous genome sequences.

Flow cytometry as a rapid analytical tool to determine physiological responses to changing O2 and iron concentration by Magnetospirillum gryphiswaldense strain MSR-1

Magnetotactic bacteria (MTB) are a diverse group of bacteria that synthesise magnetosomes, magnetic membrane-bound nanoparticles that have a variety of diagnostic, clinical and biotechnological applications. We present the development of rapid methods using flow cytometry to characterize several aspects of the physiology of the commonly-used MTB Magnetospirillum gryphiswaldense MSR-1. Flow cytometry is an optical technique that rapidly measures characteristics of individual bacteria within a culture, thereby allowing determination of population heterogeneity and also permitting direct analysis of bacteria. Scatter measurements were used to measure and compare bacterial size, shape and morphology. Membrane permeability and polarization were measured using the dyes propidium iodide and bis-(1,3-dibutylbarbituric acid) trimethine oxonol to determine the viability and ‘health’ of bacteria. Dyes were also used to determine changes in concentration of intracellular free iron and polyhydroxylakanoate (PHA), a bacterial energy storage polymer. These tools were then used to characterize the responses of MTB to different O2 concentrations and iron-sufficient or iron-limited growth. Rapid analysis of MTB physiology will allow development of bioprocesses for the production of magnetosomes, and will increase understanding of this fascinating and useful group of bacteria.

Magnetic nanoparticles are highly toxic to chloroquine-resistant Plasmodium falciparum, dengue virus (DEN-2), and their mosquito vectors.

A main challenge in parasitology is the development of reliable tools to prevent or treat mosquito-borne diseases. We investigated the toxicity of magnetic nanoparticles (MNP) produced by Magnetospirillum gryphiswaldense (strain MSR-1) on chloroquine-resistant (CQ-r) and sensitive (CQ-s) Plasmodium falciparum, dengue virus (DEN-2), and two of their main vectors, Anopheles stephensi and Aedes aegypti, respectively. MNP were studied by Fourier-transform infrared spectroscopy and transmission electron microscopy. They were toxic to larvae and pupae of An. stephensi, LC50 ranged from 2.563ppm (1st instar larva) to 6.430ppm (pupa), and Ae. aegypti, LC50 ranged from 3.231ppm (1st instar larva) to 7.545ppm (pupa). MNP IC50 on P. falciparum were 83.32μgml-1 (CQ-s) and 87.47μgml-1 (CQ-r). However, the in vivo efficacy of MNP on Plasmodium berghei was low if compared to CQ-based treatments. Moderate cytotoxicity was detected on Vero cells post-treatment with MNP doses lower than 4μgml-1. MNP evaluated at 2-8μgml-1 inhibited DEN-2 replication inhibiting the expression of the envelope (E) protein. In conclusion, our findings represent the first report about the use of MNP in medical and veterinary entomology, proposing them as suitable materials to develop reliable tools to combat mosquito-borne diseases.

Magnetic nanoparticles from Magnetospirillum gryphiswaldense increase the efficacy of thermotherapy in a model of colon carcinoma.

Magnetic nanoparticles (MNPs) are capable of generate heating power under the influence of alternating magnetic fields (AMF); this behaviour recently opened new scenarios for advanced biomedical applications, mainly as new promising tumor therapies. In this paper we have tested magnetic nanoparticles called magnetosomes (MNs): a class of MNPs naturally produced by magnetotactic bacteria. We extracted MNs from Magnetospirillum gryphiswaldense strain MSR-1 and tested the interaction with cellular elements and anti-neoplastic activity both in vitro and in vivo, with the aim of developing new therapeutic approaches for neoplastic diseases. In vitro experiments performed on Human Colon Carcinoma HT-29 cell cultures demonstrated a strong uptake of MNs with no evident signs of cytotoxicity and revealed three phases in the interaction: adherence, transport and accumulation in Golgi vesicles. In vivo studies were performed on subcutaneous tumors in mice; in this model MNs are administered by direct injection in the tumor volume, then a protocol consisting of three exposures to an AMF rated at 187 kHz and 23kA/m is carried out on alternate days, over a week. Tumors were monitored by Magnetic Resonance Imaging (MRI) to obtain information about MNs distribution and possible tissue modifications induced by hyperthermia. Histological analysis showed fibrous and necrotic areas close to MNs injection sites in mice subjected to a complete thermotherapy protocol. These results, although concerning a specific tumor model, could be useful to further investigate the feasibility and efficacy of protocols based on MFH. Magnetic nanoparticles naturally produced and extracted from bacteria seem to be promising candidates for theranostic applications in cancer therapy.

Manganese incorporation into the magnetosome magnetite: magnetic signature of doping

The biomineralization of magnetotactic bacterial magnetite nanoparticles is a topic of intense research due to the particles' narrow size distribution and magnetic properties. Incorporation of foreign metal ions into the crystal matrix of magnetotactic bacterial magnetite has been previously examined by a number of investigators. In this study, cells of a magnetotactic bacterium, Magnetospirillum gryphiswaldense strain MSR-1 were grown in the presence of manganese, ruthe- nium, zinc and vanadium, of which only manganese was incorporated within the magnetosome magnetite crystals. We demonstrate that the magnetic properties of magnetite crystals of magnetotactic bacteria can be significantly altered by the incorporation of metal ions, other than iron, in the crystal structure. The Verwey transition serves as a unique marker to probe the incorporation of the dopant within the magnetosome: manganese incorporation into the magnetite nanocrystals is signaled by a suppression oftheVerwey transition,aswell as by changes in the crystallinestructureand chemicalcomposition ofmagnetosome magnetite.

Surface expression of protein A on magnetosomes and capture of pathogenic bacteria by magnetosome/antibody complexes.

Magnetosomes are membrane-enclosed magnetite nanocrystals synthesized by magnetotactic bacteria (MTB). They display chemical purity, narrow size ranges, and species-specific crystal morphologies. Specific transmembrane proteins are sorted to the magnetosome membrane (MM). MamC is the most abundant MM protein of Magnetospirillum gryphiswaldense strain MSR-1. MamF is the second most abundant MM protein of MSR-1 and forms stable oligomers. We expressed staphylococcal protein A (SPA), an immunoglobulin-binding protein from the cell wall of Staphylococcus aureus, on MSR-1 magnetosomes by fusion with MamC or MamF. The resulting recombinant magnetosomes were capable of self-assembly with the Fc region of mammalian antibodies (Abs) and were therefore useful for functionalization of magnetosomes. Recombinant plasmids pBBR-mamC-spa and pBBR-mamF-spa were constructed by fusing spa (the gene that encodes SPA) with mamC and mamF, respectively. Recombinant magnetosomes with surface expression of SPA were generated by introduction of these fusion genes into wild-type MSR-1 or a mamF mutant strain. Studies with a Zeta Potential Analyzer showed that the recombinant magnetosomes had hydrated radii significantly smaller than those of WT magnetosomes and zeta potentials less than −30 mV, indicating that the magnetosome colloids were relatively stable. Observed conjugation efficiencies were as high as 71.24 μg Ab per mg recombinant magnetosomes, and the conjugated Abs retained most of their activity. Numbers of Vibrio parahaemolyticus (a common pathogenic bacterium in seafood) captured by recombinant magnetosome/Ab complexes were measured by real-time fluorescence-based quantitative PCR. One mg of complex was capable of capturing as many as 1.74 × 107 Vibrio cells. The surface expression system described here will be useful for design of functionalized magnetosomes from MSR-1 and other MTB.

A novel echinocandin MIG0310 with anticandida activity from newly isolated Fusarium sp. strain MS-R1

To isolate and identify an anticandida compound from newly isolated Fusarium strain MS-R1 and characterization of its activity against standard and clinical strains of Candida. The fungal strain, Fusarium strain MS-R1, was isolated from soil. Molecular identification according to the internal transcribed spacer region of the rRNA gene sequence showed the strain to be strongly related to Fusarium brachygibbosum complex. Successive extractions of the active compound showed activity against Candida albicans, including clinical strains. Inhibition of a clinical strain of Candida tropicalis, but not Candida krusei and Candida glabrata, was also shown as tested by the broth microdilution assay. The compound was purified by liquid chromatography coupled with thin-layer chromatography and bioautography and characterized by nuclear magnetic resonance spectroscopy. It was confirmed to have the molecular formula C(48)H(66)O(18) and was identified as an echinocandin with a novel structure. A novel echinocandin-type antifungal metabolite, MIG0310, was isolated and characterized. This is a second echinocandin reported from a Fusarium species, indicating this genus to have wider range of echinocandin compounds. The new compound may lead to new anticandidal drugs, broadening the treatment opportunities for Candida species, including those that are resistant to the current antifungal drugs.

Structure prediction of magnetosome-associated proteins

Magnetotactic bacteria (MTB) are Gram-negative bacteria that can navigate along geomagnetic fields. This ability is a result of a unique intracellular organelle, the magnetosome. These organelles are composed of membrane-enclosed magnetite (Fe3O4) or greigite (Fe3S4) crystals ordered into chains along the cell. Magnetosome formation, assembly, and magnetic nano-crystal biomineralization are controlled by magnetosome-associated proteins (MAPs). Most MAP-encoding genes are located in a conserved genomic region – the magnetosome island (MAI). The MAI appears to be conserved in all MTB that were analyzed so far, although the MAI size and organization differs between species. It was shown that MAI deletion leads to a non-magnetic phenotype, further highlighting its important role in magnetosome formation. Today, about 28 proteins are known to be involved in magnetosome formation, but the structures and functions of most MAPs are unknown. To reveal the structure–function relationship of MAPs we used bioinformatics tools in order to build homology models as a way to understand their possible role in magnetosome formation. Here we present a predicted 3D structural models’ overview for all known Magnetospirillum gryphiswaldense strain MSR-1 MAPs.

MamX encoded by the mamXY operon is involved in control of magnetosome maturation in Magnetospirillum gryphiswaldense MSR-1

BackgroundMagnetotactic bacteria produce membrane-enveloped magnetite crystals (magnetosomes) whose formation is controlled primarily by a gene island termed the magnetosome island (MAI). Characterization of single gene and operon function in MAI has elucidated in part the genetic basis of magnetosome formation. The mamX gene, located in the mamXY operon, is highly conserved in the MAI of all Magnetospirillum strains studied to date. Little is known regarding the function of mamX in the process of biomineralization.ResultsA mamX deletion mutant (∆mamX) and its complemented strain (CmamX) by conjugation in M. gryphiswaldense strain MSR-1 were constructed. There were no striking differences in cell growth among ∆mamX, CmamX, and wild-type strain (WT). ∆mamX displayed a much weaker magnetic response than WT. Transmission electron microscopy revealed the presence of irregular, superparamagnetic magnetite particles in ∆mamX, in contrast to regular, single-domain particles in WT and CmamX. The phenotype of ∆mamX was similar to that of an ftsZ-like deleted mutant and mamXY operon deleted mutant reported previously. Quantitative real-time RT-PCR (qPCR) results indicated that the deletion of mamX had differential effects on the transcription levels of the other three genes in the operon.ConclusionsThe MamX protein plays an important role in controlling magnetosome size, maturation, and crystal form. The four MamXY proteins appear to have redundant functions involved in magnetosome formation. Our findings provide new insights into the coordinated function of MAI genes and operons in magnetosome formation.

Magnetite Biomineralization in Magnetospirillum gryphiswaldense: Time-Resolved Magnetic and Structural Studies

Magnetotactic bacteria biosynthesize magnetite nanoparticles of high structural and chemical purity that allow them to orientate in the geomagnetic field. In this work we have followed the process of biomineralization of these magnetite nanoparticles. We have performed a time-resolved study on magnetotactic bacteria Magnetospirillum gryphiswaldense strain MSR-1. From the combination of magnetic and structural studies by means of Fe K-edge X-ray absorption near edge structure (XANES) and high-resolution transmission electron microscopy we have identified and quantified two phases of Fe (ferrihydrite and magnetite) involved in the biomineralization process, confirming the role of ferrihydrite as the source of Fe ions for magnetite biomineralization in M. gryphiswaldense. We have distinguished two steps in the biomineralization process: the first, in which Fe is accumulated in the form of ferrihydrite, and the second, in which the magnetite is rapidly biomineralized from ferrihydrite. Finally, the XANES analysis suggests that the origin of the ferrihydrite could be at bacterial ferritin cores, characterized by a poorly crystalline structure and high phosphorus content.

A key time point for cell growth and magnetosome synthesis of Magnetospirillum gryphiswaldense based on real-time analysis of physiological factors

Pure culture of magnetotactic bacteria with high magnetosome yield has been achieved for only a few strains. The major obstacles involve the nutritional requirements and culture conditions of the cells. To increase cell density and magnetosome production, it is necessary to elucidate the physiological characteristics of a particular strain during cell growth and develop an appropriate artificial control strategy. Large-scale culture of Magnetospirillum gryphiswaldense strain MSR-1 was successfully performed for 48 h in a 42-L autofermentor, and several key physiological parameters were measured in real time. Maximal values of cell density (OD565) (19.4) and cell yield (dry weight) (4.76 g/L) were attained at 40 h. The key time point for cell growth and magnetosome formation was found to be 18–20 h. At this point, cells entered the log phase of growth, the maximal values of Cmag (1.78), iron content (0.47%), and magnetosome number (26 ± 3 per cell) were observed, superoxide dismutase (SOD) activity began to decrease more rapidly, ATP content dropped to an extremely low level (0.17 fmol), and reducing power (NADH/NAD+ ratio) began to increase very rapidly. Excessive levels of dissolved oxygen (≥20 ppb) and lactic acid in the medium caused notable cytotoxic effects after 20 h. Artificial control measures for fermentation must be based on realistic cell physiological conditions. At the key time point (18–20 h), cell density is high and magnetosomes have matured. The process of magnetosome synthesis involves a high consumption of ATP and reducing power, and the cells require replenishment of nutrients prior to the 18–20 h time point. Culture conditions that effectively minimize dissolved oxygen accumulation, lactic acid content, and reducing power at this point will enhance magnetosome yield without obvious inhibition of cell growth.