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Research in Microbiology xx (2010) 1e8
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Isolation and characterization of a marine magnetotactic spirillum
axenic culture QH-2 from an intertidal zone of the China Sea
Kailing Zhu a,1, Hongmiao Pan a,1, Jinhua Li b, Kui Yu-Zhang c, Sheng-Da Zhang a,
Wen-Yan Zhang a, Ke Zhou a, Haidong Yue a, Yongxin Pan b,e, Tian Xiao a,e,**, Long-Fei Wud,e,*
a
Key Laboratory of Marine Ecology & Environmental Sciences, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China
b
Key Laboratory of the Earth’s Deep Interior, Institute of Geology and Geophysics, Chinese Academy of Sciences, Beijing 100029, China
c
Laboratoire de Microscopies et d’Etude de Nanostructures, Université de Reims, 51687 Reims, France
d
Laboratoire de Chimie Bactérienne, Institut de Biologie Structurale et Microbiologie, UPR9043-CNRS, Marseille, France
e
Laboratoire International Associé de la Bio-Minéralization et Nano-Structures, CNRS-Marseille, 13009 Marseille, France
Received 4 December 2009; accepted 3 February 2010
Abstract
Magnetotactic bacteria (MTB) are ubiquitous in aquatic habitats. Because of their fastidious requirements for growth conditions, only very
few axenic MTB cultures have been obtained worldwide. In this study, we report a novel marine magnetotactic spirillum axenic culture,
designated as QH-2, isolated from the China Sea. It was able to grow in semi-solid or liquid chemically defined medium. The cells were
amphitrichously flagellated and contained one single magnetosome chain with an average number of 16 magnetosomes per cell. Phosphate and
lipid granules were also observed in the cells. Both rock magnetism and energy-dispersive X-ray spectroscopy characterizations indicated that
the magnetosomes in QH-2 were single-domain magnetites (Fe3O4). QH-2 cells swam mostly in a straight line at a velocity of 20e50 mm/s and
occasionally changed to a helical motion. Unlike other magnetotactic spirilla, QH-2 cells responded to light illumination. As a consequence of
illumination, the cells changed the direction in which they swam from parallel to the magnetic field to antiparallel. This response appears to be
similar to the effect of an increase in [O2]. Analysis of the QH-2 16S rRNA sequence showed that it had greater than 11% sequence divergence
from freshwater magnetotactic spirilla. Thus, the marine QH-2 strain seems to be both phylogenetically and magnetotactically distinct from the
freshwater Magnetospirillum spp. studied previously.
Ó 2010 Elsevier Masson SAS. All rights reserved.
Keywords: Magnetotactic bacteria; Response to light; Pure culture; Magnetosome; Phylogenetic analysis
1. Introduction
Magnetotactic bacteria (MTB) are a heterogeneous group
of aquatic microorganisms which share the ability to orient
themselves along magnetic field lines. The cell reaction to the
* Corresponding author. Laboratoire de Chimie Bactérienne, CNRS-Marseille,
13009 Marseille, France. Tel.: þ33 4 91164157; fax: þ33 4 91718914.
** Corresponding author. Key Laboratory of Marine Ecology & Environmental
Sciences, Institute of Oceanology, Chinese Academy of Sciences, Qingdao
266071, China.
E-mail addresses: [email protected] (T. Xiao), [email protected]
(L.-F. Wu).
1
These authors contributed equally to this work.
magnetic field is due to the presence of magnetosomes,
intracellular membrane-bound crystals of iron mineral which
consist of either magnetite (Fe3O4) or greigite (Fe3S4) within
the single domain (SD) size range (30e120 nm) (Bazylinski
and Frankel, 2004). Because of their high abundance and
their remarkable capacity for accumulating and precipitating
iron minerals, MTB are assumed to have great impact on the
biogeochemical cycling in natural sediments and are considered as an ideal model for understanding the mechanism of
biomineralization. MTB comprise a variety of morphological
types, such as coccoid, vibriod, rod-shaped, spiral-shaped, and
multicellular aggregates. They are distributed worldwide and
most of them are found at, or just below, the oxiceanoxic
0923-2508/$ - see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.resmic.2010.02.003
Please cite this article in press as: Zhu, K., et al., Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone
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transition zone (OATZ) or redoxocline in aquatic habitats
(Bazylinski and Frankel, 2004).
Considering their ubiquitous distribution and remarkable
diversity with respect to physiology and biomineralization,
axenic cultures of MTB are needed for a comprehensive
understanding of the mechanisms of magnetosome biogenesis,
magnetotaxis as well as of MTB ecological function. As a new
type of bioresource, magnetotactic bacteria have also attracted
much attention for their potential use in biotechnology, biomediation, and geobiology (Bazylinski and Frankel, 2004;
Lang and Schüler, 2006). However, only very few strains are
available in pure culture because of their fastidious growth
requirements and strong metabolic diversity (Flies et al., 2005).
Most cultivated MTB are affiliated with Alphaproteobacteria
(Flies et al., 2005) except for the magnetotactic sulfatereducing bacterium Desulfovibrio magneticus (Sakaguchi
et al., 2002). Axenic marine cultures include two magnetic
vibrios (MV-1, Bazylinski et al., 1988; MV-2, DeLong et al.,
1993), one magnetic spirillum (MMS-1, formerly known as
MV-4, Meldrum et al., 1993) and three magnetic cocci (MC-1,
Frankel et al., 1997; MC-2, Devouard et al., 1998; MO-1,
Lefèvre et al., 2009).
Here we report a novel marine magnetic spirillum axenic
culture, designated strain QH-2, isolated from an intertidal
zone of the China Sea. We will describe the growth features,
cell structure, magnetic properties and novel motility
characteristics.
2.4 ml of 0.8 M NaHCO3; 0.05 g sodium thioglycolate; and
900 ml filtered seawater collected from the pond through
a 0.45 mm filter membrane. Artificial seawater and 0.1 g/L
peptone were used instead of natural seawater and sediment
extract in order to make the chemically defined medium, called
QH-C medium. The pH value of the growth medium was
adjusted to 7.6e7.8. For semi-solid medium, 0.2e0.5 g agar
was added to 1 L of growth medium. After pH adjustment, the
medium was autoclaved at 120 C for 20 min. Cultures were
incubated at room temperature (between 22 and 26 C).
2.2. Optical and electron microscopy observations
The swimming behavior of magnetotactic bacteria was
analyzed by the ‘hanging drop’ method (Schüler, 2002) using
a microscope (OLYMPUS BX51) connected to a ChargeCoupled Device (CCD, OLYMPUS DP71). Morphological
examination of the cells was performed with fluorescence
microscopy after staining with 1% acridine orange (AO). Nile
red staining for lipid storage granules was performed according
to Greenspan et al. (1985).
Fresh cells were deposited on formvar carbon-coated copper
grids, either directly or after treatment with 0.1% uranyl
acetate. The grids were dried in air. TEM observations were
made using a Zeiss EM9 microscope at 80 kV. The size and
shape factors of magnetosomes were estimated using (length þ
width)/2 and width/length, respectively. The chemical composition of magnetosomes was studied by EDXSeTEM.
2. Materials and methods
2.3. Sequence analysis of the 16S rRNA gene
2.1. Isolation and cultivation of the QH-2 strain
The samples were collected from a seawater pond located
at Huiquan Bay in the city of Qingdao, China. The characteristics of the pond were previously described (Pan et al.,
2008). The sediments together with interface water, with
a ratio of 1:2, were collected and stored in 1-L glass bottles.
Magnetotactic bacteria were enriched by attaching the south
pole of permanent magnets (0.37 mT) outside the bottles
placed at the water/sediment interface. After 20e30 min, cells
accumulating as dark spots underneath the magnets were
removed with a Pasteur pipette and saved as magnetically
collected samples. These samples were further magnetically
purified in Pasteur pipettes according to the racetrack purification method (Wolfe et al., 1987), and inoculated into 5 ml
plastic tubes that were generally filled up to 4/5th of their
volume with various media and sealed with parafilm and
incubated at 22e26 C in dim light. Bacteria grew and formed
a sharp band or zone after several days of incubation.
The optimal growth medium for QH-2 (QH medium) was
modified from that used for the Magnetococcus sp. MC-1
(Frankel et al., 1997). It contained 100 ml extract solution of
seawater sediments (modified from the DSMZ, Medium 12:
Soil Extract Medium); 5 ml modified Wolfe’s mineral solution; 0.5 ml vitamin solution; 2.0 ml of 0.01 M ferric quinate;
1.0 g NH4Cl; 2.5 g Na2S2O3$5H2O; 1.5 ml of 0.5 M potassium
phosphate buffer, pH 7.6; 1.3 g sodium lactate (50e60%);
The 16S rRNA gene of the QH-2 was amplified between
positions 27 and 1492 (Escherichia coli 16S rRNA gene
sequence numbers), using primers 27F (50 -AGA GTY TGA
TCC TGG CTC AG-30 ) and 1492R (50 -GGT TAC CTI ‘GTI’
ACG ACT T-30 ) by polymerase chain reaction (PCR) carried
out with the following cycle: an initial denaturing step at 94 C
for 5 min, followed by 25 cycles of 1 min at 94 C, 45 s at
50 C and 1 min at 72 C, and a final extension step of 10 min
at 72 C. Then the PCR products were sequenced directly by
Sinogenomax Company in Beijing.
The sequences of the 16S rDNA gene were first analyzed
using Advanced BLAST search program on the NCBI
Website (http://www.ncbi.nlm.nih.gov/BLAST/). The related
sequences were preliminarily aligned with the default setting
of CLUSTALX (1.83). Phylogenetic analysis was performed
with the default setting of MEGA 4 using the neighbor-joining
method. Similarity was calculated using the BioEdit program.
The newly determined sequence is available from GenBank
under accession number EU675666.
2.4. Magnetic measurements
For magnetic measurements, about 1010 QH-2 cells were
collected by centrifugation from cultures. The centrifuged
cells were washed once with distilled water, and then placed in
a non-magnetic gelatin capsule. To avoid possible oxidization,
Please cite this article in press as: Zhu, K., et al., Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone
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the capsule was protected with N2 and stored at 80 C until
magnetic measurements were made.
Room-temperature magnetic experiments were performed
on a vibrating sample magnetometer Model 3900 (Princeton
Measurements Corporation, sensitivity is 5.0 1010 A m2).
A hysteresis loop was measured between þ500 and 500 mT
with an average time of 400 ms. Saturation magnetization
(Ms), saturation remanence (Mrs) and coercivity (Bc) were
determined after correction for paramagnetic phases. The
saturation isothermal remanent magnetization (SIRM) was
demagnetized in a backfield to obtain remanence coercivity
(Bcr). First-order reversal curves (FORCs) were measured
following the protocol as described by Roberts et al. (2000). A
FORC diagram was calculated using FORCine version 1.05
with a smoothing factor of 3 (Harrison and Feinberg, 2008).
This diagram showed a microcoercivity field along the horizontal axis (Hc) and a magnetostatic interaction field along the
vertical axis (Hb) (Chen et al., 2007).
Low-temperature magnetic experiments were performed on
a Quantum Design Magnetic Property Measurement System
(MPMS XP-5, sensitivity is 5.0 1010 A m2). Saturation
remanence acquired in a 2.5-T field at 5 K (hereafter termed
SIRM5K-2.5T) was demagnetized by warming from 5 to 300 K
after two different pre-treatments. The first was to cool the cell
sample from 300 down to 5 K in a zero field (ZFC), whereas
the second was to cool the cell sample from 300 to 5 K in
a 2.5-T field (FC). The Verwey transition temperature (Tv) was
defined as the temperature for the maximum of the first-order
derivative of dM/dT of the FC curve. The d ratio (dFC/dZFC)
was calculated according to Moskowitz et al. (1993), which
reflects the difference in remanence losses between the FC and
ZFC when warming through the Verwey transition.
3. Results and discussion
3.1. Isolation and cultivation of the QH-2 strain
Magnetotactic bacteria collected from the intertidal zone
sediments displayed various cell shapes including cocci, rods,
vibrios and spirilla, although the coccoid morphotype was
always the dominant one. After magnetic racetrack purification, MTB were inoculated in various semi-solid media in 5 ml
plastic culture tubes. More than two weeks, after the first
inoculation, a sharp bacterial band or zone was formed at the
oxiceanoxic interface in some of the tubes. Cells collected
afterwards from the band in QH-semi-solid media (see
‘‘Materials and methods’’) were examined under an optical
microscope. Unexpectedly, the dominant magnetotactic cocci
in the collected samples did not grow, but magnetotactic
spirilla together with non-magnetic rods and cocci were found
in the first culture. To obtain a pure culture of magnetotactic
spirilla, the mixed culture was repeatedly subjected to a new
cycle of racetrack purification and re-inoculated in fresh semisolid media. After more than 20 subsequent cycles of racetrack
purification and re-inoculation, we finally obtained a pure
culture of magnetotactic spirillum, designated strain QH-2, as
proven by 16S rRNA gene sequence analysis and genomic
Fig. 1. Cellular characteristics of QH-2 cells. QH-2 cells were stained with
acridine orange (A) or uracyl acetate (B and C) and inspected using fluorescence (A) or electron microscopy (B and C). The scale bars are 4 mm for A and
1 mm for B and C. Panel D shows the growth curve of QH-2 cultures incubated
in QH-C media (see Materials and methods) at 26 C. The values are means
plus standard deviation calculated from 3 sets of independent experiments.
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sequencing (see below). Both the fluorescence microscope
(Fig. 1A) and TEM observations (Fig. 1B) showed that the cells
are vibrioid-to-helicoid in morphology, with a mean width of
0.8 0.2 mm and variable lengths ranging from 1 to 3 mm, with
a mean length of 2.0 0.4 mm. They were amphitrichously
flagellated with a single flagellum at each pole (Fig. 1C).
Various marine culture media were analyzed for the growth
of the QH-2 culture, which seemed to be heterotrophic under
the conditions used, as growth of QH-2 required both the
sediment extract for the QH medium and low amounts of
peptone for QH-C medium. Nonetheless, we could not exclude
capacity for autotrophic growth if appropriate conditions were
provided. Up to now, QH and QH-C are the optimal media for
QH-2 cell growth. When grown statically within the QH-C
medium containing 0.05% agar at 26 C, the generation time
of QH-2 was about 16 h (Fig. 1D).
3.2. Phylogenetic lineage
Sequences of the 16S rRNA gene measured with about
20 independent QH-2 cultures showed 99.9% identity.
Chromosomal DNA was prepared from the QH-2 culture.
Genomic sequencing indicated that the QH-2 culture consists of
only one species of bacteria (data not shown), confirming the
pure culture of the QH-2 strain. Phylogenetic analysis showed
that QH-2 was affiliated with Alphaproteobacteria and its
16S rRNA sequence was more than 11.7% divergent from all
freshwater magnetotactic spirilla including Magnetospirillum
magnetotacticum MS-1, Magnetospirillum magneticum AMB-1,
Magnetospirillum gryphiswaldense MSR-1, and Magnetospirillum spp. MGT-1 and WM-1 (Blakemore et al., 1979; Li
et al., 2007; Matsunaga et al., 1991; Okamura et al., 2003;
Schüler and Koehler, 1992). Moreover, QH-2 was 88.9%,
82.4% and 84.7% identical to that of marine magnetotactic
vibrio MV-1, Magnetococcus sp. MC-1 and magnetotactic
ovoidal strain MO-1, respectively (Fig. 2). The closest relative was found to be the marine magnetic spirillum MMS-1
(97.2% identity) (Fig. 2), which was isolated from mud and
water from School Street Marsh, Woods Hole, MA, U.S.A.
(Meldrum et al., 1993). Therefore QH-2 together with MMS-1
may represent a novel phylogenetic lineage that is distinct from
other axenic magnetotactic bacterial cultures.
However, it is worth noting that although the 16S rRNA
gene sequence of QH-2 showed high identity with that of
MMS-1, they might not belong to the same species. In fact,
even with 97.7% identity in their 16S rRNA sequences, the
freshwater magnetospirillum strains AMB-1 (from Japan) and
WM-1 (from China) actually belong to different species
(Li et al., 2007).
3.3. Intracellular structures of QH-2
TEM observations and statistical analyses showed that each
QH-2 cell contained 7e28 magnetosomes, with an average
number of 16 5 per cell (Fig. 3A and B), which is similar to
the number of magnetosomes found in MMS-1 (Meldrum
et al., 1993). The magnetosome crystals of QH-2 cells were
identified as iron oxide (Fe3O4) by both EDXS and electron
diffraction (data not shown). The magnetosomes in a chain
displayed a relatively large size distribution compared to other
magnetospirilla (Fig. 3B and C). The growth conditions of
QH-2 cultures might not be optimal for the formation of
magnetosomes. The magnetite magnetosomes had an average
width of 58 20 nm, an average length of 81 23 nm and
a shape factor of 0.71 0.11. Similar to other magnetotactic
bacteria, the distribution of the QH-2 crystals was asymmetric,
with a cut-off toward larger size. Meanwhile, the length of
magnetosomes in QH-2 cells was longer than that of MMS-1
whereas their width was roughly the same. The shape factor
analysis of magnetosome crystals of QH-2 cells showed the
distribution bounded by one, with a maximum around 0.7
(Fig. 3C), which is similar to those of marine magnetotactic
bacteria MV-1 (shape factor was 0.65) (Devouard et al., 1998).
In addition to the formation of magnetosomes (Fig. 3A,
white arrow), one or two dark granules (Fig. 3A, labeled ‘P’)
and a number of white globules (Fig. 3A, labeled ‘L’) were
observed in QH-2 cells by TEM. EDXS analysis revealed that
the dark granules were rich in phosphorus and oxygen, but the
white globules could not be identified (data not shown).
Similar intracellular aggregates have been identified as lipid
Fig. 2. Neighbor-joining tree. The sequence determined in this study is written in bold. GenBank accession numbers of the sequences used are indicated in
parentheses. Bootstrap proportions are shown. Scale bar, 0.02 substitutions per nucleotide position.
Please cite this article in press as: Zhu, K., et al., Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone
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Fig. 3. Magnetosome and granule characteristics of QH-2 cells. Panel A shows transmission electron microscopic micrographs of QH-2 cells and panel B shows
a typical magnetosome chain inside cells. Magnetosome crystals (white arrow in panel A), phosphorouseoxygen-rich granules (‘P’ in panel A) and lipid storage
globules (‘L’ in panel A) are indicated. Size (panel C1) and shape factor (panel C2) distributions were obtained by analysis of 281 crystals of QH-2 cells at
stationary growth phase. Panels D1 and D2 are typical images of QH-2 cells inspected under an optical microscope as Nomarski contrast and fluorescence,
respectively. D2 shows the same cells as in D1 after Nile red staining. Lipid storage granules are shown as white spots. Scale bars ¼ 1 mm for panel A, 100 nm for
panel B and 4 mm for panel D.
storage granules in MTB (Lefèvre et al., 2009; Silva et al.,
2008). An efficient identification assay for the lipid storage
granules is Nile red staining as described by Greenspan et al.
(1985). Indeed, the white globules were specifically stained as
red spots in QH-2 cells (Fig. 3, Panel D2, represented by white
color), which confirmed their authenticity as lipid storage
granules. As previously reported for the MO-1 strain, some
magnetosome chains bent along the curvature of the lipid
storage globules in the QH-2 cells (Fig. 3A). The fact that they
contain lipid storage granules might be a common trait of
MTB (Lefèvre et al., 2009; Schultheiss et al., 2005; Silva
et al., 2008). The biosynthesis of lipid storage granules is
supposedly promoted in response to stress imposed on the
cells and during unbalanced growth. These white granules act
as storage compounds for energy and carbon needed for
maintenance of metabolism and synthesis of cellular
metabolites during starvation, in particular if growth causes
consumption to significantly increase (Waltermann and
Steinbuchel, 2005).
3.4. Magnetic property of the QH-2 strain
The thermal demagnetization curves of SIRM5K-2.5T are
shown in Fig. 4A. Both FC and ZFC curves show sharp drops in
remanence between 90 and 112 K. This confirms the magnetite
magnetosome composition as revealed by the TEM study. The
determined Tv was 108 K, and dFC and dZFC were 0.43 and
0.29, respectively, yielding a d ratio of 1.5. The higher dFC and
lower d ratios were possibly due to the chain arrangement of
magnetosomes in QH-2 cells, i.e. short axis alignment.
The hysteresis loop of the QH-2 cell samples was potbellied. The values of hysteresis parameters such as Bc and
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Fig. 4. Magnetic characteristics of QH-2 cells. (A) FC-SIRM5K-2.5T (solid squares) and ZFC-SIRM5K-2.5T (open squares) warming curves; (B) room temperature
hysteresis loop; (C) room temperature FORC diagram and profiles of coercivity field (Hc) and interaction field (Hb) distribution through the peak of FORC
distribution (r).
Bcr, and ratios Bcr/Bc and Mr/Ms were deduced as 29.0 mT and
36.1 mT, and 1.24 and 0.55, respectively. These indicated that
the QH-2 magnetosomes were all uniaxial SD particles
(Fig. 4B).
The FORC diagram of QH-2 cells had a closed concentric
contour around a central peak Hc,FORC ¼ 34.4 mT, indicating
a typical feature of SD magnetite magnetosomes. The characteristic interaction field, Hb1/2, which was defined as the
value of the Hb field where the peak of the FORC distribution
was reduced to half of its maximum value, was 2.4 mT. The
narrow vertical spread of QH-2 samples in the FORC diagram
indicated no or weak intercell or interchain magnetostatic
interactions (Fig. 4C).
3.5. Characterization of QH-2 motility
It was reported, by Spormann and Wolfe (1984) for spirilla
and by Frankel et al. (1997) for cocci, that migration in the
magnetic field is determined by an aerotactic sensory system.
There are two kinds of magneto-aerotactic behaviors: axial or
polar magnetotaxis. The axial magnetotactic Magnetospirillum
spp. cells swim in both directions along the magnetic field,
whereas the polar magnetotactic bacteria swim persistently
toward one direction in a magnetic field (this is the case for the
marine Magnetococcus sp. MC-1 strain) (Frankel et al., 1997).
QH-2 cells were amphitrichously flagellated (Fig. 1C).
Interestingly, QH-2 cells did not swim equally in both directions under oxic conditions in the hanging drop assay under
the microscope. Most QH-2 cells were north-seeking and
accumulated on the north side of the droplet (Fig. 5A). This is
consistent with the behavior of polar MTB in the northern
hemisphere (Qingdao: N36.1 300 , E120.3 100 ). However, the
proportion of QH-2 cells at the north side compared to those at
the south side was not as high as that measured with other
polar MTB from the northern hemisphere (such as the case of
MC-1) and the proportion varied with different culture
conditions and different growth phases. Moreover, once the
cells on the north side were removed and then analyzed in
a fresh droplet, the population was re-segregated to the north
and south sides. Polar magnetotaxis has also been observed for
other freshwater magnetospirilla that are normally axial
magneto-aerotactic (Frankel et al., 2006).
The motility track of the QH-2 cells was recorded by using
dark-field optical microscopy. They swam roughly along
a straight line with a velocity ranging from 20 to 50 mm/s
(Fig. 5B). Occasionally, a cell was observed to change from
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Fig. 5. Motility of QH-2 cells. Cells accumulated at the north and south edges of a droplet. (A) Motility tracks of QH-2 cells in geomagnetic field were recorded
using dark-field optical microscopy. (B) Trajectory sections ‘1’, ‘2’ and ‘3’, separated by arrows, show cell motion in straight line, helix and straight line,
respectively. Scale bar shows 30 mm, C1 shows that cells accumulated at the north end in a droplet before flashing with a blue light at 450e480 nm at the northern
edge of the droplet, whereas C2 shows swimming away of cells from the droplet edge 1 min after flashing. The geomagnetic field direction is indicated.
the straight line motion (Fig. 5B, trajectory section 1) to
a helix (section 2) and then back to the straight line (section 3).
The helical pattern in the trajectory section 2 was unlikely to
result from the sticking of the flagellum onto the glass cover
slip, since it corresponded to a 100 mm distance that the cell
covered without an obvious change in swimming velocity.
Such change of motion mode seemed to be in concordance
with the change in swimming direction of QH-2.
Bacteria can sense a wide range of environmental signals
that steer bacterial locomotion through the extensively studied
chemotaxis mechanism (Hazelbauer et al., 2008; Wadhams
and Armitage, 2004). The chemoreceptors MCPs (methylaccepting chemotaxis proteins) detect the stimuli and influence cellular locomotion via histidine protein kinase CheA,
the phosphorylation state of the response regulator CheY.
CheY-P interacts with flagellar motor and switches the rotation
direction. Rotation in one direction results in smooth swimming, whilst switching of the rotation direction may lead to
backward motion, tumbling or swimming pause (Hazelbauer
et al., 2008). Therefore, smooth swimming of a bacterium is
periodically interrupted by a change in direction.
The frequently used bacterial motility apparatus is the
rotatory flagella, as is the case for QH-2 cells. When propelled
by flagella, bacterial cells translate along and rotate around the
long cellular axis. The overview of the swimming trajectory
appears as a straight line when the translation axis overlaps
with the rotation axis. However, when the two axes deviate,
the trajectory displays itself as a helix. The helical trajectory
was occasionally observed for QH-2 cells, as indicated by the
trajectory section 2 in Fig. 5B. This point corresponds to
a change in swimming direction, from parallel to perpendicular to the magnetic field. It is possible that such a helical
trajectory was due to a poorly aligned magnetosome chain in
the cell, as observed for the cell in Fig. 1B. Moreover, the cell
might not have enough mature magnetosomes to enable
constant alignment of the cell along the magnetic field lines.
When the cell is well aligned along the magnetic field lines
(the sections 1 and 3), the three axes (translation, rotation and
magnetic dipolar moment) overlap and result in a straight
trajectory. In contrast, if the cell does not align with the
magnetic field lines, as observed for section 2 of which the
translation direction is perpendicular to the magnetic field
orientation, the three axes deviate. As a consequence, the
overview of the trajectory is helical.
It is well known that light is one of the major changing
stimuli for many bacterial species. Blue and UV lights cause
damage to almost all living systems. Blue light is particularly
damaging if oxygen is present. It was reported that cells of the
marine Magnetococcus sp. MC-1 strain in a capillary tube
exhibited a response to a short-wavelength light (500 nm),
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ARTICLE IN PRESS
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K. Zhu et al. / Research in Microbiology xx (2010) 1e8
which caused them to swim away from the meniscus (high
[O2]) and toward the petroleum jelly plug (low [O2]) persistently
parallel to the magnetic field during the illumination, resulting
in a reaction similar to an increase in [O2] (Frankel et al., 1997).
The QH-2 cells were studied for their response to shortwavelength light in a droplet. It was observed using a light
microscope that most QH-2 cells swam to the north side and
accumulated at the edge under normal conditions. When illuminating the cells (Fig. 5C1) with the wavelengths ranging from
330 to 550 nm, the emitted energy triggered the QH-2 cells
swimming away from the edge to the interior of the droplet
(from high to low [O2]) (Fig. 5C2). Such cell behavior was
similar to that observed for MC-1 cells. However, unlike the
case of MC-1 in which virtually all cells left the original position, nearly half of the QH-2 cells appeared to stop motility upon
illumination and become agglomerated instead of swimming
away (Fig. 5C2). In contrast, magnetotactic spirillum AMB-1,
one of the most extensively studied axial magneto-aerotaxis
models, apparently did not respond to the illumination process
in the same way (data not shown). The mechanism of QH-2 cell
reaction to light requires further investigation.
As the axenic culture of QH-2 was isolated from the
intertidal region, the bacteria in the sediments might be
regularly exposed to air and light or the changing chemical
gradients with the tides. Therefore, the pure culture of QH-2
could be used as a novel strain model for the study of the
mechanism of magneto-aerotaxis in combination with the
cellular reaction to light and oxygen.
Acknowledgments
This work was supported by the NSFC (Nos. 40776094 and
40906069), the Haiwaijiechuxuezhe Fund of the Chinese
Academy of Sciences (No. 2006-1-15), a CNRS scholar
fellowship (to K.L. Zhu), a special fund of Creative Projects
for the Postdoctors of Shandong Province (082318101N) and
K.C. Wong Education Foundation, Hong Kong.
We thank W. Jiang, T. Song, C. Chen, W.J. Zhang,
C.L. Santini and N. Philippe for discussions and advice, and
M. Jiang for assistance in electron microscopy observations.
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Please cite this article in press as: Zhu, K., et al., Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone
of the China Sea, Research in Microbiology (2010), doi:10.1016/j.resmic.2010.02.003