Appl Microbiol Biotechnol (1999) 51: 229±234
ORIGINAL PAPER
M. A. Kacena á G. A. Merrell á B. Manfredi
E. E. Smith á D. M. Klaus á P. Todd
Bacterial growth in space ¯ight: logistic growth curve parameters
for Escherichia coli and Bacillus subtilis
Received: 27 February 1998 / Received revision: 21 August 1998 / Accepted: 3 September 1998
Abstract Previous investigations have reported that
bacterial suspension cultures grow to higher stationary
concentrations in space ¯ight than on Earth; however,
none of these investigations included extensive ground
controls under varied inertial conditions. This study
includes extensive controls and cell-growth data taken at
several times during lag phase, log phase, and stationary
phase of Escherichia coli and Bacillus subtilis. The
Marquardt-Levenberg, least-squares ®tting algorithm
was used to calculate kinetic growth parameters from
the logistic bacterial growth equations for space-¯ight
and control growth curves. Space-¯ight cultures grew to
higher stationary-phase concentrations and had shorter
lag-phase durations. Also, evidence was found for in-
creased exponential growth rate in space.
Introduction
Microgravity research o€ers an opportunity to investi-
gate the relative roles of di€usion and metabolic rate in
biomass synthesis in the absence of any convective
forces, such as agitation which is normally provided to
keep cells suspended and sedimentation that moves cells
through fresh medium but ultimately crowds them at the
bottom of the vessel. Therefore, the objective of this
research was to develop a complete set of experimental
growth curves for Escherichia coli and Bacillus subtilis
under varying inertial conditions including microgravity.
Investigations of the growth of bacteria in space ¯ight
have generally reported higher ®nal cell concentrations
in space ¯ight than under normal unit-gravity (1 g)
M. A. Kacena á G. A. Merrell á B. Manfredi á E. E. Smith
D. M. Klaus á P. Todd (&)
University of Colorado, BioServe Space Technologies,
Engineering Center, ECAE 1B-01, Campus Box 429,
Boulder, CO 80309-0429, USA
e-mail: todd@spot.colorado.edu
Tel.: +1-303-492-5936
Fax: +1-303-492-4341
Ó Springer-Verlag 1999
laboratory conditions (Mattoni et al. 1971; Mesland
1987; Mennigmann and Lange 1988; Klaus et al. 1994,
1997). Mennigmann and Lange (1988) found that
B. subtilis suspension cultures grown in space had an
increase in growth rate and total biomass yield. Also,
Klaus et al. (1997) found that E. coli, cultivated in space,
had a cell population that was an average of 88% higher
than controls. Several of these studies were performed in
the same ¯ight hardware as used in the experiments
described below, and the E. coli strain was identical.
However, Bouloc and D'Ari (1991) reported no di€er-
ences in ¯ight and ground E. coli bacterial cell biomass
concentrations.
Most of these earlier investigations did not include
extensive ground-control cultures modeling various ¯uid
and inertial conditions nor were data collected several
times throughout the growth cycle of the bacterial cul-
tures. Therefore, the current study includes both exten-
sive control culture conditions and cell-growth data
taken several times throughout the growth cycle of B.
subtilis and E. coli. The space-¯ight and control data
were analyzed on the basis of the logistic bacterial
growth equation (Bailey and Ollis 1986). This analysis
technique, along with high-precision, baseline laborato-
ry data, aids the determination of the kinetic parameters
for both B. subtilis and E. coli when grown in space, even
with limited data. According to this analysis, cultures in
space ¯ight grew to higher ®nal cell concentrations (cs)
with apparently increased growth rates (l) during com-
parable exponential growth periods (te), and had ap-
parently shorter lag periods (tl).
Materials and methods
Microorganisms
E. coli (ATCC 4157) and B. subtilis (ATCC 6051) were the two
bacterial strains selected because of the large pre-existing database
and because E. coli is a gram-negative species with lower motility
(compared to the B. subtilis strain used) while B. subtilis is a gram-
positive, highly motile species (Turnbull et al. 1990; Holt et al.
230

1994). The latter could be less a€ected by inertial forces because
swimming produces more movement than sedimentation alone.
Culture media
All bacterial samples were cultured in Vogel-Bonner synthetic
medium E growth medium (Vogel and Bonner 1956). Prior to
initiation of the space-¯ight experiments, the bacterial inocula were
held in minimal medium E (carbon source omitted) to minimize
metabolic activity. Then, at initiation, the bacteria in the minimal
medium E were mixed into complete medium E that contained
5 g l)1 glucose so that the ®nal concentration of glucose was ap-
proximately 4.3 g l)1.
Baseline cultures and cell counting
In separate, baseline experiments, samples were cultured in 9-ml
test-tubes completely ®lled with log-phase B. subtilis or E. coli cell
cultures (i.e. no headspace). The baseline experiments were used as
high-precision controls for which the Marquardt-Levenberg algo-
rithm was tested to ®t the data precisely to the growth kinetic
equations discussed below.
Hemacytometer counts of high-density, log-phase stock cul-
tures were determined, and the appropriate quantity of cells was
diluted into complete medium E to give a starting concentration of
5 ´ 105 cells ml)1. In Table 1 columns 2 and 3 show the various
inertial conditions and temperatures under which E. coli and
B. subtilis were grown in these studies. The inertial conditions in-
cluded static 1 g cultures in the vertical and horizontal positions at
both 23 °C and 37 °C, and vertical cultures that were grown on a
shaker bath (model 50, Precision Scienti®c Company, Chicago)
operating at 100 oscillations min)1 and 23 °C.
At the appropriate intervals (approximately every 4±8 h during
lag and stationary phases and about every 2 h during log phase),
the absorbance was measured in triplicate at 660 nm. Each culture
was used for only one measurement because the tubes had to be
completely mixed (by manual shaking and vortexing) to obtain
accurate absorbance readings. Absorbance was transformed into
cell concentration using calibration curves. These calibration
curves were obtained from previous experiments in which the ab-
sorbance had been measured, and then a total cell count was per-
formed on the same culture.
Experimental cultures for space ¯ight
The experiments were performed in the ¯uid processing apparatus
(FPA), space-¯ight quali®ed hardware that allows test-tube ex-
periments to be initiated and terminated in space (Luttges 1992).
Figure 1 illustrates the operation of the FPA and indicates how the
samples were loaded. Chamber A of the FPA contained 3.5 ml
complete medium E. To be consistent throughout the experiments,
0.5 ml minimal medium E, initially containing 5 ´ 105 cells ml)1
(on the basis of hemacytometer counts as just described) of either
E. coli or B. subtilis, were loaded into chamber B of the FPA.
Finally, 1.5 ml absolute ethanol was placed into chamber C. All
chambers were completely ®lled with liquid without headspace and
were completely sealed from other oxygen sources. Therefore, all
oxygen received by the cells was that which was initially dissolved
within the medium. After about 24 h the dissolved oxygen was
depleted (data not shown). This depletion, in the ground experi-
ments (performed at 23 °C), coincided with the end portion of the
lag-phase period after which cells continued to multiply in log
phase and then reached saturation. Therefore, all subsequent
growth was anaerobic. This information aids the interpretation of
our growth data, since our growth rate is below that generally

Table 1 Growth parameters of Escherichia coli and Bacillus subtilis
baseline cultures grown at 23 °C or 37 °C in 9-ml test-tubes under
agitation and sedimentation (above line). Growth kinetic para-
meters of E. coli and B. subtilis ¯ight and ground control (static
sedimentation or 1 g) cultures grown at 23 °C or 37 °C in a ¯uid
processing apparatus (below line). cs saturation cell concentration,
l exponential growth rate, tl lag-phase duration, ts stationary-phase
entry, te exponential growth period. Cell concentrations were
measured experimentally. l, tl, ts and te were calculated by applying
the Marquardt-Levenberg algorithm to the experimental data. C
control, used to compare two di€erent types of cultures. Quantities
in parentheses indicate the percentage di€erence from the control

Bacteria Conditions T (°C) 10)6 ´ ci 10)8 ´ cs l (h)1) tl (h) ts (h) te (h)

(cells ml)1) (cells ml)1)

E. coli 1 g vertical (C) 23 0.6 ‹ 0.1 5.8 ‹ 0.2 0.148 ‹ 0.008 31 ‹ 3 76 ‹ 4 44 ‹ 5

1 g horizontal 23 0.6 ‹ 0.1 5.1 ‹ 0.2 0.15 ‹ 0.01 23 ‹ 3 69 ‹ 5 46 ‹ 6
(13%)* ()2%)* (25%)* (9%)* ()3%)*
Shaker vertical 23 0.6 ‹ 0.1 5.1 ‹ 0.3 0.15 ‹ 0.03 13 ‹ 4 62 ‹ 7 49 ‹ 8
(13%)* ()2%) (60%)* (19%)* ()11%)*
1 g vertical (C) 37 0.6 ‹ 0.1 8.3 ‹ 0.1 0.32 ‹0.03 10 ‹ 2 33 ‹ 2 23 ‹ 2
1 g horizontal 37 0.6 ‹ 0.1 7.6 ‹ 0.2 0.31 ‹ 0.04 6‹2 30 ‹ 2 24 ‹ 3
(9%)* (2%) (3%)* (8%)* ()7%)
B. subtilis 1 g vertical (C) 23 0.6 ‹ 0.2 4.7 ‹ 0.1 0.17 ‹ 0.01 29 ‹ 3 68 ‹ 4 39 ‹ 5
1 g horizontal 23 0.6 ‹ 0.2 4.2 ‹ 0.1 0.19 ‹ 0.01 32 ‹ 3 66 ‹ 4 34 ‹ 5
(11%)* ()15%)* ()8%)* (3%)* (12%)*
Shaker vertical 23 0.6 ‹ 0.2 4.7 ‹ 0.2 0.19 ‹ 0.02 30 ‹ 3 67 ‹ 5 37 ‹ 5
()3%) ()10%)* ()3%)* (1%)* (5%)
1 g vertical (C) 37 0.6 ‹ 0.2 5.9 ‹ 0.1 0.33 ‹ 0.02 10 ‹ 1 31 ‹ 2 20 ‹ 2
1 g horizontal 37 0.6 ‹ 0.2 6.4 ‹ 0.1 0.31 ‹ 0.02 9‹1 31 ‹ 2 21 ‹ 2
()9%)* (4%)* (7%) (0.04%) ()3%)

E. coli 1 g vertical (C) 37 11 ‹ 2 2.7 ‹ 0.1 0.15 ‹ 0.02 18 ‹ 4 36 ‹ 4 18 ‹ 6

Flight 37 11 ‹ 2 11.6 ‹ 0.7 0.49 ‹ 0.06 4‹1 18 ‹ 1 15 ‹ 1
()325%)* ()224%)* (79%)* (49%)* (21%)*
B. subtilis 1 g vertical (C) 23 70 ‹ 40 3‹2 0.06 ‹ 0.04 32 ‹ 11 75 ‹ 16 43 ‹19
Flight 23 70 ‹ 40 8‹1 0.07 ‹ 0.03 19 ‹ 7 55 ‹ 10 36 ‹ 13
()125%)* ()23%)* (42%)* (26%)* (14%)
1 g vertical (C) 37 13 ‹ 3 1.3 ‹ 0.3 0.27 ‹ 0.08 4‹1 24 ‹ 1 19 ‹1
Flight 37 13 ‹ 3 7.3 ‹ 0.6 0.48 ‹ 0.01 4‹1 19 ‹ 2 15 ‹ 2
()474%)* ()79%) (5%) (20%)* (23%)*
* Statistically signi®cant di€erence compared to control (Student's t-test, P < 0.05)

Fig. 1 The ¯uid processing apparatus (FPA), a space-¯ight quali®ed
device, was used for all experiments in space and their synchronous
ground controls. Its function was to separate three ¯uids in the ``as
loaded'' condition. To initiate the experiment the plunger was
depressed, mixing chamber B into chamber A via laminar mixing.
When the experiment was to be terminated the plunger was depressed
once again to combine the contents of chamber C (®xative) with the
previously combined chambers A and B. For these experiments
chamber A contained 3.5 ml medium E supplemented with glucose.
Chamber B contained 0.5 ml unsupplemented medium E with the
bacteria. Chamber C held 1.5 ml 100% ethyl alcohol
reported for E. coli and because of the strict aerobic requirements
of B. subtilis (Bailey and Ollis 1986).
On and during space shuttle mission STS-63, bacterial cultures
were subjected to various environmental conditions as described in
Table 1. Synchronous ground controls were loaded into FPA and
were activated and terminated at the same time (in duplicate) as the
corresponding ¯ight cultures. Synchronous controls were static 1 g
samples in the vertical position and shaker-bath (as described
above) controls (because of the large number of replicate samples
required, cultures could not be grown on an in-¯ight 1 g centri-
fuge). After predetermined periods of time, bacterial growth was
terminated by the addition of ethanol (1.5 ml added to 4.0 ml
medium and cells). When the orbiter landed and the samples were
recovered, the ®xed cultures were shipped back to the laboratory in
Colorado where cell counts were performed at 400´ in a hemacy-
tometer, and data were recorded as cell concentration (cells ml)1).
The cell concentration was the mean of at least eight independent
counts.
Cell growth curve analysis
Cellular growth rates were analyzed quantitatively using the lo-
gistic bacterial growth equations (Bailey and Ollis 1986) and the
Marquardt-Levenberg, least-squares curve-®tting algorithm (Mar-
quardt 1963) using the SigmaPlot scienti®c graph system (Jandel
231
Scienti®c, Calif.). Inputs to the algorithm were time (t), the cor-
responding average cell density (ct), and the standard deviation of
the corresponding cell density, the inverse square of which was
used as the weighting function for each data point. Equations 1±4,
as previously published (Kacena and Todd 1997), were ®tted to
obtain the following parameters: co, the extrapolated cell density at
t ˆ 0; cs, the saturated(stationary)-phase cell density; ci, the lag-
phase cell density; and l, the exponential growth rate. After these
best-®t values had been obtained, the following three time pa-
rameters were established: tl, the lag-phase duration; ts, the sta-
tionary-phase entry time; and te, the duration of the exponential
growth phase. Errors were calculated by the propagation of the
measured errors, and signi®cance was determined by a Student's
t-test with P < 0.05.
Results
Baseline experiments
Figure 2A illustrates the most relevant and typical E.
coli, baseline growth curves (vertical, 1 g static cultures)
at 23 °C and 37 °C and their respective ®tted curves. The
growth parameters (initial cell density, ®nal cell density,
rate of growth, lag phase duration, logarithmic growth
duration, and time of entrance into stationary phase) for
all baseline experiments were collected and evaluated.
No lysis was seen for E. coli or B. subtilis cells during
200 h. Baseline controls were performed for the fol-
lowing four reasons: (1) to model growth of both cell
types under conditions similar to space ¯ight (com-
pletely ®lled small containers); (2) to explore gravity
conditions relevant to space ¯ight (particle suspension
compared to falling particles due to inertial forces),
namely, orientation, agitation, and temperature (listed
above); (3) to demonstrate a reproducible method of
analysis that gives biologically relevant parameters, such
as ®nal cell density (cs), rate of growth (l), lag phase
duration (tl), logarithmic growth duration (te), and time
of entrance into stationary phase (ts) (Table 1); (4) to
establish high-precision, baseline growth curves for
comparison to space-¯ight experiments, which are typi-
cally constrained to lower experimental precision owing
to the limits imposed by ¯ight-quali®ed incubation
hardware. Such a systematic, quantitative investigation
of bacterial cell culture relevant to space-¯ight e€ects
has not been published previously.
E€ects of space ¯ight
Figure 2B±D shows the ¯ight and simultaneous
ground-control experimental growth data and ®tted
curves for E. coli at 37 °C (Fig. 2B), B. subtilis at 37 °C
(Fig. 2C), and B. subtilis at 23 °C (Fig. 2D). Because
samples prepared for ¯ight experimentation had to be
stored in minimal medium at their respective experi-
mental temperatures (23 °C or 37 °C) for approxi-
mately 36 h prior to launch, some cell multiplication
occurred. Simulating these storage conditions in ground
experiments with and without shaking showed the
232
Fig. 2 A Experimental data with ®tted growth curves for vertical,
Escherichia coli, baseline controls grown at 23 °C (right-hand
curve) and 37 °C (left-hand curve). These curves were chosen as
being representative of the data from all of the baseline
experiments, in which growth curves of the same species at the
same temperature were visually indistinguishable. It can be seen
that bacteria grown at 37 °C exhibit faster growth rates (l), shorter
lag-phase periods (tl), and higher ®nal cell concentrations (cs) than
at 23 °C. B, C, D Experimental data with ®tted growth curves for
space-¯ight and ground-control bacterial cultures grown at 23 °C
and 37 °C. B, C Growth of E. coli and Bacillus subtilis, respectively,
at 37 °C. D growth of B. subtilis cultures at 23 °C. In all cases, the
¯ight cultures have a faster growth rate (l), shorter lag-phase
periods (tl), and higher ®nal cell concentrations (cs). All error bars
on the experimental data are shown and were calculated as the
standard deviation of the mean (average cell concentration)
initial cell densities to be those indicated as ci in Ta-
ble 1. The growth parameters and percentage change
from control samples for the space-¯ight experiments
are given in Table 1.
These experiments and their analyses seem to con®rm
the earlier suspicion (Klaus et al. 1994; TheÂvenet et al.
1996) that a shortened lag phase occurs in low-gravity
cultures. In all cases, ®nal cell densities (cs) were higher
in low-gravity cultures.
Discussion
The data point to three interesting ®ndings. (1) All space
cultures grew to higher ®nal cell density without excep-
tion. (2) Earlier evidence for a shorter lag phase in E. coli
appears to have been corroborated (Klaus et al. 1994,
1997; TheÂvenet et al. 1996). (3) The duration of the
logarithmic growth phase was very similar in space-
¯own cultures and ground controls, suggesting that a
faster growth rate (l) occurred; this was dicult to
measure owing to the small numbers of data points be-
tween tl and ts for establishing l. A detailed discussion
of each of these three ®ndings follows.
Final cell concentration
B. subtilis cultures grown at 23 °C aboard the Space
Shuttle experienced an average 125% increase in cs
compared to controls. At 37 °C, the space-¯ight B. sub-
tilis cultures grew to a stationary-phase cell density that
averaged 474% higher than that of the corresponding
controls. Similarly, E. coli cultures grown in micro-

gravity at 37 °C had a ®nal cell population that averaged
325% above that of static 1 g controls. These data
suggest either that, in space, bacteria have more favor-
able access to nutrients and better dissemination of
waste products, owing to lack of sedimentation to the
bottom of the culture container (indirect gravity e€ect),
or that low gravity causes more ecient metabolism of
nutrients, and cells can multiply more often before the
nutrients are exhausted (direct gravity e€ect). This dis-
tinction was tested in other experiments.
Studies using the same bacterial strains, grown in the
same hardware modi®ed for agar cultures, and ¯own on
the same Space Shuttle missions showed that cultures on
solid agar substrates grew to statistically identical ®nal
cell concentrations in space and on Earth (Kacena et al.
1997). Therefore, it would appear that space-¯own
bacteria do not have more ecient metabolic capabili-
ties in view of a lack of enhanced growth on agar.
The closest analogy in biotechnology practice to
quiescent suspension, such as occurs in microgravity, is
fed-batch fermentation. At very high glucose levels, too
much acetate and/or lactate is produced, and biomass
synthesis is attenuated (Crabtree e€ect). When the glu-
cose concentration is held low by batch feeding, more
biomass is synthesized (Frude et al. 1994; Todd and
Klaus 1995). Similarly, in low gravity without agitation,
the cell depletes its immediate unstirred surroundings of
glucose thereby dropping the glucose concentration to
an eventual steady-state value based on the material
balance between the transport (di€usion) rate to the cell
and the rate of substrate consumption. Indeed, when
glucose is fed to bacterial suspension cultures at a lower
concentration than is used in single-batch fermentations,
higher cell densities are achieved. In the absence of
mixing, cells are expected to deplete glucose in their
immediate environment and should therefore also be
expected to grow to higher cell densities. In the presence
of gravity this does not occur, because cells sediment,
become overcrowded, and rapidly deplete precursor
pools. This appears to be con®rmed by the baseline
studies. Static 1 g E. coli cultures had a ®nal cell con-
centration that was signi®cantly lower (13%) than
simultaneous 1 g shaker-bath samples. It is possible that
the shaker samples have better access to nutrients while
the cells are falling to the bottom of the culture vessel
(zig-zag pattern) and, once sedimented, the shaking ac-
tion allows more cell layers than just the top layers to be
exposed to the nutrients. There was no signi®cant dif-
ference between baseline 1 g and shaker B. subtilis
studies, probably because of the motility of B. subtilis
cells.
Lag-phase duration
B. subtilis cultures at 23 °C had much shorter lag phases
in ¯ight than on the ground, as re¯ected by a succession
of data prior to growth (Fig. 3C). At 37 °C, E. coli
cultures also appeared to have shorter lag phases in
233
¯ight than on the ground. However, B. subtilis samples
at 37 °C experienced essentially identical lag phases ac-
cording to the logistic curve ®ts; the lag phase was
shorter in ¯ight, but by only about 5%, which is not
statistically signi®cant. The non-signi®cant di€erence in
lag-phase duration for B. subtilis cultured at 37 °C, may
possibly be explained by the motility of B. subtilis cells
(compared to less motile E. coli cells grown at 37 °C) in
combination with the higher temperature (compared to
the B. subtilis cells grown at 23 °C). The exact duration
of the lag phase for all cases, associated errors, and
percentage di€erence from the control (Table 1) may
demonstrate a shortened lag phase in E. coli (37 °C) and
B. subtilis (23 °C) where the di€erences are shown to be
signi®cant. Klaus et al. (1994, 1997) also noted a de-
crease in lag-phase duration for E. coli grown in space
with absorbance monitoring; however, this decrease was
not reported to be signi®cant. In a separate study by
TheÂvenet and associates (1996), the E. coli cultures
grown in space also appeared to have a reduced lag
period. In any case, there is no evidence supporting a
prolonged lag period in low gravity.
Growth rate
Growth rate determinations were very dicult in space-
¯ight experiments, in which data intervals exceeded 1/l.
Nevertheless, it can be inferred, not only from the values
of l in the logistic curve ®ts, but also from straight lines
connecting ci to cs on the 37 °C growth curves for E. coli
and B. subtilis, that both cell types grew more rapidly in
the space cultures. B. subtilis cultures at 23 °C exhibited
a signi®cantly higher growth rate, on the basis of a data
set with more log-phase measurements, but the di€er-
ence was not as large (23%). These data suggest that
space-¯own, bacterial cultures grow more rapidly than
simultaneous controls.
In another study, Gasset et al. (1994) examined the
growth of E. coli by taking absorbance measurements on
samples that had been terminated (by decreasing the
temperature) at 6 assay times. They calculated the
doubling time to be 46 min in ¯ight and 59 min on
Earth. Also, they found that the ¯ight cultures entered
stationary phase 9 h earlier than did the ground sam-
ples. They believed this was due to either a more rapid
growth rate or a shorter lag period; but, because the
di€erences were not considered signi®cant, they con-
cluded that space ¯ight had ``no e€ect on the growth rate
of exponentially growing E. coli cells''. Mennigmann
and Lange (1988) studied B. subtilis cultures in space
(via absorbance measurements) and found them to have
a higher growth rate and ®nal biomass yield than con-
trols. Overall, evidence has been found that is consistent
with a higher growth rate in E. coli and B. subtilis sus-
pension cultures in low gravity.
Acknowledgements This work was supported by NASA grant
NAGW-1197, Achievement Rewards for College Scientists, and the
234
Zonta International Foundation. We would like to thank Pat
Leonard, John Doyle, Dave Hanna, Doug Kacena, Sue Ellis, and
Emily Wilson for their help over a 2-year period and Mike Pecaut
and Elizabeth Stephenson for taking readings over the weekend
after Thanksgiving Day.
References
Bailey JE, Ollis DF (1986) Biochemical engineering fundamentals,
2nd edn. McGraw Hill, New York, NY
Bouloc P, D'Ari R (1991) Escherichia coli metabolism in space.
J Gen Microbiol 137: 2839±2843
Frude MJ, Read A, Kennedy LD (1994) Scale-up production of a
recombinant Mycobacterium leprae antigen. Ann NY Acad Sci
72: 100±104
Gasset G, Tixador R, Eche B, Lapchine L, Moatti N, Toorop P,
Woldringh C (1994) Growth and division of Escherichia coli
under microgravity conditions. Res Microbiol 145: 111±120
Holt JG, Krieg NR, Sneath PHA, Staley JT, Williams ST (1994)
Facultatively anaerobic gram negative rods. In: Hensyl WR
(ed) Bergey's manual of determinative bacteriology, 9th edn.
Williams & Wilkins, Baltimore, Md, p 179
Kacena M, Todd P (1997) Growth characteristics of E. coli and B.
subtilis cultured on an agar substrate in microgravity. Micro-
gravity Sci Technol 10: 58±62
Kacena MA, Leonard PE, Todd P, Luttges MW (1997) Low
gravity and inertial e€ects on the growth of E. coli and B.
subtilis in semi-solid media. Aviat Space Environ Med 68: 1104±
1108
Klaus DM, Luttges MW, Stodieck LS (1994) Investigation of space
¯ight e€ects on Escherichia coli growth. SAE Technical Paper
Series, SAE 941260
Klaus D, Simske S, Todd P, Stodieck L (1997) Investigation of
space ¯ight e€ects on E. coli and a proposed model of under-
lying physical mechanisms. Microbiology 143: 449±455
Luttges MW (1992) Recognizing and optimizing ¯ight opportuni-
ties with hardware and life sciences limitations. Trans Kansas
Acad Sci 95: 76±86
Marquardt DW (1963) An algorithm for least squares estimation of
non-linear parameters. J Soc Ind Appl Math 11: 431±441
Mattoni RH, Ebersold WT, Eiserling FA, Keller ED, Roming WR
(1971) Induction of lysogenic bacteria in the space environment.
In: Saunders JW (ed) The experiments of biosatellite II. NASA
SP-104. pp 304±324
Mennigmann HD, Lange M (1988) Growth and di€erentiation of
Bacillus subtilis under microgravity conditions. In: Longdon
N, David V (eds) Biorack on Spacelab D1: an overview of the
®rst ¯ight of Biorack, an ESA Facility for Life Sciences Re-
search in Microgravity, ESA SP-1091. Noordwijk, Nether-
lands, pp 37±44
Mesland DAM (1987) Biorack experiments in Spacelab D-1 and
IML-1: further developments in gravitational biology. In:
Proceedings of the 3rd European Symposium on Life Sciences
Research in Space. pp 307±312
TheÂvenet D, D'Ari R, Bouloc P (1996) The SIGNAL experiment in
Biorack: Escherichia coli in microgravity. J Biotechnol 47:
89±97
Todd P, Klaus DM (1995) Theories and models on the biology of
cells in space. Adv Space Res 17: 3±10
Turnbull P, Kramer J, Melling J (1990) Bacillus. In: Parker MT,
Duerden BI (eds) Topley & Wilson's principles of bacteriology,
virology and immunity, 8th edn. Decker, Philadelphia, Pa,
p 189
Vogel HJ, Bonner DM (1956) Acetylornithinase of Escherichia coli:
partial puri®cation and some properties. J Biol Chem 218: 97±
106