Introduction

Magnetotactic bacteria comprise a diverse group of aquatic microorganisms with the

ability to orient along the lines of a magnetic field (Frankel, 2004). swimming property These
organisms were discovered by Salvatore Bellini in Italy in 1963 from the microscopic observation
of drops of freshwater samples. In 1975, Richard Blakemore observed the presence of
intracytoplasmic inclusions of iron, which were related to magnetic field orientation (Blakemore,
1975). These organisms are distributed in different classes of the phylum Proteobacteria and
also in other phyla and present different cellular morphologies. Most of described magnetotatic
bacteria are microaerophilic, anaerobic or facultative.

The common characteristic found in all magnetotactic bacteria is the presence of

intracytoplasmic magnetic particles and, consequently, the ability to respond to the magnetic
field (Blakemore & Maratea, 1979). These magnetic particles are specialized organelles called
magnetosomes, which are formed by the process of biomineralization (Komeili, 2006). The
magnetosomes are organelles composed of a magnetic crystal consisting of iron mineral
(magnetite - Fe3O4 - or greigite - Fe3S4), surrounded by a phospholipid membrane (Frankel &
Bazylinski, 2004). The membrane still contains associated proteins, which lead to the
biomineralization process (Arakaki, 2003; Gorby, 1988). Although variable among species, most
of the magnetosomes have a narrow range in size ranging from 35 to 120 nm when measured
along the major axis (Bazylinski & Schbbe, 2007). The morphology of crystals of magnetosomes
tends to be unique in a particular species of bacteria (Frankel & Bazylinski, 2004).

The precise control of the magnetosome formation process allows these structures to
have characteristics superior to those found in artificially synthesized iron particles (Xie et al,
2009), such as narrow size distribution, uniform morphology and unique magnetic domain of
the crystals. The presence of a biological membrane surrounding the magnetic crystal also
provides good dispersibility and easy functionalization (Alphandery, 2014). Such properties
favour the application of these magnetic nanoparticles in biomedicine and biotechnology.
However, due to the fastidious growth of magnetotactic bacteria and low yield of
magnetosomes during cultivation, obtaining these biologically friendly nanoparticles is still
challenging and their application is hindered (Yan et al, 2012).

To yield large quantities of the magnetosomes and make their application feasible,
large-volume cultures are needed when cultivating magnetotatic bacteria. A number studies
describe mass cultivation of magnetotactic bacteria in bioreactor using different strategies to
achieve maximum magnetite production and productivity (Heyen & Schuler, 2003; Liu et al,
2010; Sun et al, 2008a; Zhang, 2011). Those strategies included media modifications and culture
physical and chemical conditions (Araujo et al, 2015). To our knowledge, cultivation of
magnetotactic bacteria in bioreactors have been conducted in batch (Heyen & Schuler, 2003),
fed-batch (Liu et al, 2010; Sun et al, 2008a) and semicontinuous modes (Zhang, 2011). Most
studies report growth of Magnetospirillum genus and only one recent work made use of
Magnetovibrio blakemorei strain MV-1. The latter strain produced magnetosomes of a prismatic
shape and their volume and surface are larger than those of Magnetospirillum genus. These
characteristics might be advantageous for functionalization and application of these
magnetosomes (Silva et al,2013).
 As a hurdle in cultivation of magnetotactic bacteria, spontaneous nonmagnetic mutants
are often found in stationary growth of Magnetospirillum (Schubbe, 2003). The inability to
produce magnetosome in those cells is due to deletions in different sites of the magnetosome
island (Ullrich, 2005). In MV-1, spontaneous non-magnetic mutant cells are also found in culture
(Dubbels, 2004). These mutants do not produce several proteins present in wild type and they
lack an iron uptake system necessary for biomineralization. When growing in an fermentor it
might be a good strategy to avoid stationary phase in order to prevent loss of magnetosome
synthesis ability by cells. To keep cells growing in a steady state corresponding to exponential
grow, we need to make use of continuous growth. Continuous culture should be able to maintain
growth at a given rate indefinitely (e.g. maximum growth rate) as fresh sterile medium is
inserted in the reactor vessel and spent medium with metabolics and cell debris are being
removed (Hoskisson and Hobbs, 2005).

Here, we propose cultivation of magnetic vibrio Magnetovibrio blakemorei strain MV-1

in continuous culture in a chemostat strategy, where log phase conditions are kept constant,
including high nutrient concentration. Depletion of carbon, nitrogen and iron sources are thus
prevented. Viable cell density are kept by pumping fresh medium in as medium with cell
suspension is pumped out. The aim is to keep high productivity of magnetosome constant,
diminishing occurrence of late-growth-phase non-magnetic mutants.