# Magnetotactic bacteria

Magnetotactic bacteria (or MTB) are a class of bacteria discovered in the 1960s, that exhibit the peculiar ability to orient themselves along the magnetic field lines of Earth's magnetic field. The term magnetotaxis has been coined to describe the biological phenomenon upon which these microorganisms tend to move in response to the magnetic characteristics of the environment.

## Introduction

The first description of magnetotactic bacteria appeared in 1963 in a publication of the Istituto di Microbiologia (it: Microbiology Institute) of the University of Pavia written by Salvatore Bellini [1]. While observing bog sediments under his microscope, he noticed a group of bacteria that evidently oriented themselves in a unique direction. He realised that these microorganisms moved according to the direction of the North Pole and hence denominated them "magnetosensitive bacteria".

The first peer-reviewed report of magnetotactic bacteria appeared in a 1975 article in Science by Richard Blakemore, a microbiologist at the Woods Hole Oceanographic Institution, who had similarly observed bacteria capable of orienting themselves in a certain direction: Blakemore realised that these microorganisms were following the direction of Earth's magnetic field, from south to north, and thus coined the adjectival descriptor "magnetotactic" [2].

These bacteria have been the subject of many experiments: they have even been aboard the Space Shuttle to examine their magnetotactic properties in the absence of gravity, but a definitive conclusion was not reached [3].

The sensitivity of magnetotactic bacteria to the Earth's magnetic field arises from the fact that the bacteria precipitate within their cells chains of crystals of magnetic minerals; all magnetotactic bacteria reported to date precipitate either magnetite or greigite. These crystals, and sometimes the chains of crystals, can be preserved in the geological record as magnetofossils. The oldest reported magnetofossils come from the 1.9 billion year old Gunflint Chert [4]. There have also been claims of their existence on Mars based on the shape of magnetite particles within the Martian meteorite ALH84001, but these claims are highly contested [5].

### Biology

Several different morphologies (shapes) of MTB exist which further differ for number, layout and pattern of the "bacterial magnetic particles" (BMP) the bacteria contain [6]. The MTB can be subdivided into two categories, according to whether they produce particles of magnetite (${\displaystyle \mathrm {Fe} _{3}\mathrm {O} _{4}}$) or of greigite (${\displaystyle \mathrm {Fe} _{3}\mathrm {S} _{4}}$), although some species are capable of producing both. Magnetite possesses a magnetic moment three times that of greigite [5].

Magnetite-producing magnetotactic bacteria are usually found in oxic-anoxic transition zones (OATZ), the transition zone between oxygen-rich and oxygen-starved water or sediment. Many MTB are only able to survive in environments with very limited oxygen, and some can only exist in completely anaerobic environments. Quite possibly the evolutionary advantage of possessing a system of magnetosomes is linked to the ability of efficiently navigating within this zone of sharp chemical gradients by simplifying a potential three dimensional search for more favorable conditions to a single dimension (see the "Magnetism" subsection below for a description of this mechanism). Some types of magnetotactic bacteria can produce magnetite even in anaerobic conditions, using nitric oxide, nitrate, or sulfate as a final acceptor for electrons. The greigite mineralising MTB are usually strictly anaerobic. [7].

It has been suggested that MTB evolved in the early Proterozoic Era, as the increase in atmospheric oxygen reduced the quantity of dissolved iron in the oceans. Organisms began to store iron in some form, and this intracellular iron was later adapted to form magnetosomes for magnetotaxis. These early MTB may have participated in the formation of the first eukaryotic cells. [4] Biogenic magnetite not too different from that found in magnetotactic bacteria has been also found in higher organisms, from Euglenoid algae, to salmons, pigeons, and humans.

Magnetotactic bacteria produce their magnetic particles in chains, as is clearly visible in this microscopic image of Magnetospirillum magneticum. The magnetic dipole of the cell is therefore the sum of the dipoles of each BMP which is then sufficient to passively orientate the cell and overcome the casual thermal forces found in a water environment [7]. In the presence of more than one chain, the inter-chain repulsive forces will push these structures to the edge of the cell, inducing turgor [5].

The diversity of MTB is reflected by the high number of different morphotypes found in environmental samples of water or sediment. Commonly observed morphotypes include spherical or ovoidal cells (cocci), rod-shaped (bacilli), curved bacteria (vibrio) and helical (spirillum) of various dimensions. One of the more unique morphotypes is an apparently multicellular bacterium referred to as the Many-celled Magnetotactic Prokaryote (MMP). Regardless of their morphology, all MTB studied so far are motile by means of flagella and have a cell wall structure characteristic of Gram-negative bacteria. The arrangement of flagella differs and can be polar, bipolar, or in tufts.

Another trait which shows considerable diversity is the arrangement of magnetosomes inside the bacterial cell. In the majority of MTB, the magnetosomes are aligned in chains of various lengths and numbers along the cell's long axis, which is magnetically the most efficient orientation. However, dispersed aggregates or clusters of magnetosomes occur in some MTB usually at one side of the cell, which often corresponds to the site of flagellar insertion. Besides magnetosomes, large inclusion bodies containing elemental sulfur, polyphosphate, or poly-β-hydroxybutyrate are common in MTB.

The most abundant type of MTB occurring in environmental samples, especially sediments, are coccoid cells possessing two flagellar bundles on a somewhat flattened side. This "bilophotrichous" type of flagellation gave rise to the tentative genus "Bilophococcus" for these bacteria. In contrast, two of the morphologically more conspicuous MTB, regularly observed in natural samples, but never isolated in pure culture, are the MMP and a large rod containing copious amounts of hook-shaped magnetosomes (Magnetobacterium bavaricum).

### Magnetism

Physically, the development of a magnetic crystal is governed by two factors: one, moving to align the magnetic force of the molecules in conjunction with the developing crystal, while the other reduces the magnetic force of the crystal, allowing an attachment of the molecule while experiencing an opposite magnetic force. In nature this causes the existence of a magnetic domain, surrounding the perimeter of the domain, with a thickness of approximately 150 nm of magnetite, within which the molecules gradually change orientation. For this reason macroscopically the iron is not magnetic in the absence of an applied field. Similarly, extremely small magnetic particles do not exhibit signs of magnetisation at room temperature, their magnetic force is continually altered by the thermal motions inherent in their composition [5]. Instead, individual magnetite crystals in MTB are of a size between 35 and 120 nm, that is, big enough to have a magnetic field and at the same time small enough to remain a single magnetic domain [7].

The inclination of the Earth's magnetic field in the two respective hemispheres selects one of the two possible polarities MTB polarity model of the magnetotactic cells (with respect to the flaggelated pole of the cell), orienting the biomineralisation of the magnetosomes. Various experiments have clearly shown that magnetotaxis and aerotaxis work in conjunction in the magnetotactic bacteria. Aerotaxis is the response by which bacteria migrate to an optimal oxygen concentration in an oxygen gradient. It has been shown that, in water droplets, one-way swimming magnetotactic bacteria can reverse their swimming direction and swim backwards under reducing conditions (less than optimal oxygen concentration), as opposed to oxic conditions (greater than optimal oxygen concentration). The behaviour that has been observed in these bacterial strains has been referred to as magneto-aerotaxis.

Two different magneto-aerotactic mechanisms — known as polar and axial — are found in different MTB strains [8]. Some strains that swim persistently in one direction along the magnetic field (NS or SS[vague]) — mainly the magnetotactic cocci — are polar magneto-aerotactic. These magnetotactic bacteria will travel along the lines of the earth’s magnetic field according to their orientation, but will swerve as a group and reverse direction if exposed to a local, more powerful and oppositely oriented magnetic field. In this way they continue to travel in the same magnetic direction , but relative instead to the local field. Those MTB that swim in either direction along magnetic field lines with frequent, spontaneous reversals of swimming direction without turning around — for example, freshwater spirilla — are axial magneto-aerotactic and the distinction between NS and SS does not apply to these bacteria. The magnetic field provides both an axis and a direction of motility for polar magneto-aerotactic bacteria, whereas it only provides an axis of motility for axial types of bacteria. In both cases, magnetotaxis increases the efficiency of aerotaxis in vertical concentration gradients by reducing a three-dimensional search to a single dimension.

Scientists have also proposed an extension of the described model of magneto-aerotaxis to a more complex redoxtaxis. In this case, the unidirectional movement of MTB in a drop of water would be only one aspect of a sophisticated redox-controlled response. One hint for the possible function of polar magnetotaxis could be that most of the representative microorganisms are characterized by possessing either large sulfur inclusions or magnetosomes consisting of iron-sulfides. Therefore, it may be speculated that the metabolism of these bacteria, being either chemolithoautotrophic or mixotrophic, is strongly dependent on the uptake of reduced sulfur compounds which occurs in many habitats only in deeper regions at or below the OATZ due to the rapid chemical oxidation of these reduced chemical species by oxygen or other oxidants in the upper layers.

Microorganisms belonging to the genus Thioploca, for example, use nitrate, which is stored intracellularly to oxidize sulfide and have developed vertical sheaths in which bundles of motile filaments are located. It is assumed that Thioploca uses these sheaths to efficiently move in a vertical direction in the sediment, thereby accumulating sulfide in deeper layers and nitrate in upper layers [9]. For some MTB, it might also be necessary to perform excursions to anoxic zones of their habitat in order to accumulate reduced sulfur compounds.

## Magnetosomes

The biomineralisation of the magnetite requires regulating mechanisms to control the concentration of iron, the crystal nucleation, the redox potential and the pH. This is achieved by means of compartmentalisation in structures known as magnetosomes that allow the biochemical control of the above mentioned processes. After the genome of several MTB species had been sequenced, a comparative analysis of the proteins involved in the formation of the BMP became possible. Sequence homology with proteins belonging to the ubiquitous CDF (Cation Diffusion Facilitator) family and the "Htr-like" serine proteases has been found: while the first group is exclusively dedicated to the transport of heavy metals, the second group consists of heat shock proteins (HSPs) involved in the degradation of badly folded proteins. Other than the serine protease domain, some proteins found in the magnetosomial membrane (MM) also contain PDZ domains, while several other MM proteins contain TPR domains (Tetratric Peptide Repeat) [6].

#### TPR domain

The TPR domains are characterized by a folding consisting of two α-helices and include a highly conserved consensus sequence of 8 amino acids (of the 34 possible) which is the most common in nature. Apart from these amino acids, the remainder of the structure is found to be specialized in relation to its functional significance. The more notable compounds that comprise TPR domains include:

1. membrane bound transport complexes conveying proteins within mitochondria and/or peroxisomes
2. complexes that recognise DNA binding proteins and repress DNA transcription
3. the anaphase-promoting complex (APC}.

Examples of both the TPR-TPR interactions as well as TPR-nonTPR interactions have been reported [10].

#### PDZ domain

The PDZ domains are modular structures which consist of 6 β-filaments and 2 α-helices which recognise the C terminal aminoacids of proteins in a sequence specific manner. Usually the third residue from the C terminal is phosphorylated, thus preventing interaction with the PDZ domain. It's not surprising the only conserved residues in these structures are those involved in the recognition of the carboxy terminal (RKXXXGLGF). PDZ domains are quite widespread in nature since they constitute the basic structure upon which multi-proteic complexes are assembled: this is especially true for those associated with membrane proteins, such as the inward rectifier K+ channels or the β2-adrenergic receptors [11].

### Membrane and proteins

The formation of the MM requires at least three steps. During the first formation of an invagination in the citoplasmatic membrane is triggered by a GTPase. It is supposed this process can take place amongst eukaryote as well.

The second step requires the entrance of ferric ions into the newly formed vesicles from the external environment. Even when cultured in a Fe3+ deficient media, MTBs succeed at accumulating high intracellular concentrations of this ion by secreting upon need a siderophore, a low molecular weight ligand displaying an elevated affinity for Fe3+ ions. The "Fe3+-siderophore" complex is subsequently moved in the cytoplasm, where it is cleaved. The ferric ions must then be converted into the ferrous form (Fe2+), in order to be accumulated within the BMP; this is achieved by means of a trans-membrane transporter which exhibits sequence homology with a Na+/H+ antiporter. Actually, the complex is a H+/Fe2+ antiporter, which transports ions thanks to the proton gradient. These trans-membrane transporters are localised both in the cytoplasmic membrane and in the MM, but with in an inverted orientation: this configuration allows to generate an efflux of Fe2+ ions at the cytoplasmatic membrane, and an influx of this same ion at the MM. This step is strictly controlled by a cytochrome dependent redox system which is not yet fully explained and appears to be species specific.

During the final stage of the process the magnetite crystal nucleation by action of trans-membrane proteins with acidic and basic domains. One of these proteins, called Mms6, has also been employed for the artificial synthesis of magnetite, where its presence allows the production of crystals homogeneous in shape and size.

Likely, many other proteins associated with the MM could be involved in other roles, such as generation of supersaturated concentrations of iron, maintenance of reducing conditions, oxidisation of iron, and partial reduction and dehydration of hydrated iron compounds [12].

### Biomineralisation

Several clues led to the hypothesis that different genetic sets exist for the biomineralisation of magnetite and greigite. In cultures of Magnetospirillum magnetotacticum iron can't be replaced with other transition metals (Ti, Cr, Co, Cu, Ni, Hg, Pb) commonly found in the soil. In a similar manner oxygen and sulfur are not interchangeable as non-metallic substances of the magnetosome within the same species [7].

From a thermodynamical point of view, in presence of a neutral pH and a low redox potential, the inorganic synthesis of magnetite is favoured when compared to those of other iron oxides. It would thus appear that microaerophilic or anaerobic conditions create a suitable potential for the formation of the BMP. Moreover all iron absorbed by the bacteria is rapidly converted into magnetite, indicating the formation of crystals is not preceded by the accumulation of intermediate iron compounds; this also suggests the structures and the enzymes necessary for biomineralisatio are already present within the bacteria. This conclusions are also supported by the fact that MTBs cultured in aerobic conditions (and thus non magnetic) contain amounts of iron comparable to any other species of bacteria [13].

## Biotechnology Applications

In certain types of applications, bacterial magnetite offers several advantages compared to chemically synthesized magnetite [14]. Bacterial magnetosome particles, unlike those produced chemically, have a consistent shape, a narrow size distribution within the single magnetic domain range, and a membrane coating consisting of lipids and proteins. The magnetosome envelope allows for easy couplings of bioactive substances to its surface, a characteristic important for many applications.

Magnetotactic bacterial cells have been used to determine south magnetic poles in meteorites and rocks containing fine-grained magnetic minerals and for the separation of cells after the introduction of magnetotactic bacterial cells into granulocytes and monocytes by phagocytosis. Magnetotactic bacterial magnetite crystals have been used in studies of magnetic domain analysis and in many commercial applications including: the immobilization of enzymes; the formation of magnetic antibodies, and the quantification of IgG; the detection and removal of Escherichia coli cells with a fluorescein isothiocyanate conjugated monoclonal antibody, immobilized on magnetotactic bacterial magnetite particles; and the introduction of genes into cells, a technology in which magnetosomes are coated with DNA and "shot" using a particle gun into cells that are difficult to transform using more standard methods.

Unfortunately, the prerequisite for any large scale commercial application is mass cultivation of magnetotactic bacteria or the introduction and expression of the genes responsible for magnetosome synthesis into a bacterium, e.g., E. coli, that can be grown relatively cheaply to extremely large yields. Although some progress has been made, the former has not been achieved with the available pure cultures.

## Bibliography

1. Bellini, Salvatore, "Su di un particolare comportamento di batteri d’acqua dolce", 1963. Instituto di Microbiologia dell'Universita di Pavia.
2. Blakemore, Richard. "Magnetotactic Bacteria", 1975. Science 190: 377-379.
3. Urban, "Adverse effects of microgravity on the magnetotactic bacterium Magnetospirillum magnetotacticum", 2000 Acta Astronautica Elsevier
4. Chang, S. R. and J. L. Kirschvink. "Magnetofossils, the magnetization of sediments, and the evolution of magnetite biomineralization", 1989. Annual Review of Earth and Planetary Sciences 17: 169-195.
5. Cat Faber, Living Lodestones: Magnetotactic bacteria, Strange Horizons, 2001
6. Schüler, Dirk, "The biomineralization of magnetosomes in Magnetospirillum gryphiswaldense", 2002 Int. Microbiology
7. Bazylinski, Dennis, "Controlled biomineralization of magnetic minerals by magnetotactic bacteria", 1995 Chemical Geology Elsevier
8. Frankel, R. B., D. A. Bazylinski, and D. Schüler "Biomineralization of magnetic iron minerals in magnetotactic bacteria" J. Supramolecular Science, 5, 1998, pp. 383-390.
9. Huettel, M., S. Forster, S. Kloser, and H. Fossing. "Vertical migration in the sediment-dwelling sulfur bacteria Thioploca spp. in overcoming diffusion limitations" Appl. Environ. Microbiol., 62, 1996, pp. 1863-1872.
10. Lamb, Tugendreich, Hieter, Tetratrico peptide repeat interactions: to TPR or not to TPR?, 1995 TIBS Elsevier
11. Sheng, Sala, "PDZ domains and the organization of supramolecular complexes", 2001 Ann. Rev. Neuroscience
12. Matsunaga, Okamura, "Genes and proteins involved in bacterial magnetic particle formation", 2003 Trends in Microbiology Elsevier
13. Schüler D., Baeuerlein E. (1998). "Dynamics of iron uptake and Fe3O4 biomineralization during aerobic and microaerobic growth of Magnetospirillum gryphiswaldense". Journal of Bacteriology. 180 (1): 159–162. PMID 9422606.
14. Saiyed, Telang, Ramchand, "Application of magnetic techniques in the field of drug discovery and biomedicine", 2003 Biomag. Res. Tech. Biomed. Central