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Measurement of biological activity in materially closed microbial ecosystems
205
BioSystems, 14 (1981) 205--209 © Elsevier/North-Holland Scientific Publishers Ltd.
MEASUREMENT OF BIOLOGICAL ACTIVITY IN MATERIALLY CLOSED MICROBIAL ECOSYSTEMS
E L I Z A B E T H A. K E A R N S
and C L A I R E. F O L S O M E
Laboratory for Primordial Biology, Department of Microbiology, University of Hawaii at Manoa, Honolulu, Hawaii 96822, U.S.A. (Received March 4th, 1981)
Gas phase oxygen concentrations of materially closed, energetically open miniature microbial ecosystems were measured periodically. Our results indicate: (i) closed systems remain biologically active for at least 9 years, (ii) Po 2 values might serve as an indicator of stability, (iii) each closed ecosystem seems to seek its own unique final Po 2 state, and (iv) ecosystem response to experimentally depleted Po 2 suggests the presence of positive feedback control.
Introduction The prospect of long-term manned space missions has increased interest in the properties of closed ecologies (Gitel'son, 1976; Taub, 1974; Brown, 1966). A reliable metric to assess biological activity and stability in closed systems has yet to be developed. While some theoretical studies have been performed (Botkin, 1978; Board, 1976; Margulis and Lovelock, 1974), there has been little experimental research. To gain qtmntitative insight on the nature of ecosystem stability and activity under closure, we are investigating miniature materially closed, energetically open complex microbial ecosystems. We propose that gaseous oxygen concentration changes can be used as a measure of biological activity. Oxygen concentration is an attractive metric as a potential descriptor of ecosystem states as it is almost entirely biologically produced. Geological mechanisms of production are negligible in comparison (Holland, 1962; Walker, 1974) and the presence of oxygen in the atmosphere is closely associated with the origin and evolution of life (Berkner and Marshall, 1965; Rutten, 1971; Com-
monet, 1965). Furthermore, the thermodynamic properties of this gas have been thoroughly studied and its concentrations are easily measured in small samples.
Materials and methods We devised small closed ecosystems constructed with carbonate sand, water and sediment from shallow coral reefs. Twoliter Pyrex flasks were filled with 100 ml sea water and 100 cc sand and sediment taken from oligotrophic coral reefs at 15 m depths. These flasks were sealed immediately under ambient air with ground glass high vacuum stoppers to prevent matter exchange with the external environment. Flasks were fitted with renewable multilayer teflonfaced sampling ports and were placed by a window receiving NE sun light. As a control, one flask was only loosely covered to allow gas exchange without significant water evaporation. An 8-year-old materially closed system was also employed in these studies. We are aware that these closed miniature ecosystems contain a complex assemblage of of microbial biotypes. Even small volume
206 ecosystems as these presumably contain a range of micro-niches for principal forms of microbial metabolism; photo- and chemoautotrophic and photo-and chemoorganotrophic modes. To date we have not attempted to assay these ecosystems for their specific biotype distribution profiles since experiments as this could perturb a s m a l l
closed ecosystem by removing a metabolic link or by physically disturbing the near isolation of that link. Thus our initial experiments reported here were designed to treat the entire ecosystem flask as a "white box" in which the input (sun light) was eitherunchanged or removed and the system output (P02) was periodically measured.
2~
2~
o~
22 ¸
2
20~21] I "'"" 20"
~B 0
~0
30
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0
I
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I
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~0
6O
8O
~00
I 80
I00
T, days
0
T, days
21
o~ ." . !
..
0
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0
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! 0
• i
!
!
2O
I z,o
I 60 T, days
I 80
I Loo
12
~ 20
I ~0
I 6o
I
T, d a y s
Fig. i . The kinetics o f oxygen concentration change for four microbial ecosystems. Data are presented as linear regression lines derived from 3-day moving average values. Mean and standard deviation were calculated for all horizontal portions. (A) a closed ecosystem, measurements performed from the day o f closure: section 1 comprises 20 data points; the coefficient of determination (r 2) is 0.86; section 2 comprises 43 data points, standard deviation (s), 0.20; section 3, 52 data points, r ~ -- 0.73• (B) a replicate d o s e d ecosystem, taken from the same site as A, measurements performed from the day o f closure: section 1, 25 data points, r ~ = 0.84; section 2, 56 data points, s = 0•16; section 3, 119 data points, r~ = 0.91. ( C ) a n open control system collected from the_~_me site as A and B; 166 data points; s = 0.20. (D) an 8 -year-old closed ecosystem; the initial P o 2 increase marked by 28 data points, r 2 = 0.79; the later horizontal portion comprises 177 data points, s = 0.21.
207 Overt signs of microbial activity are continually manifested in closed ecosystems by the appearance of green, blue~areen, brown and reddish colonies and mats upon the sand and inner walls of the flask. Gas bubbles of varying size enmeshed in these pigmented photosynthetic communities appear and dissolve. Deeper, presumably anaerobic, portions of the sediment show periodically varying black spots 1--5 mm across. Closed microbial ecosystems kept in our laboratory since 1967 still show all these facets of continued biological activity, although it generally is believed that materially closed systems cannot survive for any substantial length of time (Taub, 1974). The gas phases of each experimental system were analyzed by gas chromatography using 2 mm × 2 m molecular seive columns and a Carle micro thermistor bead detector. Small samples of 25 or 50 ~1 were taken daffy and compared to samples of dry air at ambient temperature and pressure. The ratio of the oxygen peak height to the total peak heights of nitrogen and oxygen was assigned a value of 20.95% for a standard dry air. The sample ratio was compared to this standard ratio to compute the relative oxygen content. Changes in total gas pressure within closed flasks were less than 5 mm (less than 0.7%) over the total span of sampling.
Results The kinetics of PO2 change for several ecosystems are shown in Fig. 1. The data were fitted using linear regression analyses. A and B are two flasks monitored daffy for 3 months from the moment of closure. Both show an overall increase in PO2: the PO2 of A increased from 19.5% to 22% by day 40, while the PO2 o f B increased from 19.7% to 24% by day 85. This corresponds to increments of 6% (A) and 16% (B) higher than the control value of 20.7% oxygen of Fig. 1C (note that 20.7% is the standard
oxygen content of moist air, while 20.95% is the value for dry air). While the rates of oxygen increase differed slightly between these two systems, several features are common: (i) both show a step pattern of PO~ increase, and (ii) the rate of PO2 increase before a step is distinctly greater than after a step. For A these values are 0.14%/day, followed by 0.07%/day: for B the rates are 0.10%/day, followed by 0.06%/day. This suggests that a lower PO2 in the gas phase is correlated with a greater rate of oxygen production. The open ecosystem, Fig. 1C, and the long-term closed ecosystem, Fig. 1D, show no overall change in the rate of oxygen production under constant light conditions. The brief rise observed for ecosystem D was due to a change in its location to an area receiving greater sun light (a response we have observed frequently). All the closed ecosystems of Fig. 1 show apparent oscillations of PO2 with periods greater than 24 h. Because this phenomenon could obscure a conventional regression line, the data were analyzed by fitting a regression line using a moving average of successive 3-days' span. The nature of these longer term PO2 periodic fluctuations awaits further analysis. To show that a net production of oxygen did occur, the atmosphere of ecosystem A was evacuated to 14 mm pressure and replaced with nitrogen at 750 mm pressure. The PO2 after evacuation and equilibration of dissolved gas was about 4%. The PO2 began to increase at a rate almost three times greater than that measured before removal (Fig. 2A). The rate before removal of oxygen was 0.07%/day, while after removal it increased to 0.21%/day. This result suggests that low PO2 stimulates a greater rate of oxygen production and implies the presence of a positive feedback mechanism. The restoring capacity of this mechanism appears to be very high since within 35 days the oxygen content had reached half of its amount prior to evacuation.
208 A
B 22 J 17
22
15-
2'
13-
20
..
o~
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o
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,,,
I 6o
I so
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T, days
,s
o
I ,o
I ~o
~ ...... 3'o 4'o T, days
I so
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Fig. 2. The kinetics o f o x y g e n c o n c e n t r a t i o n change in t w o closed microbial e c o s y s t e m s w h i c h have b e e n pert u r b e d . (A) a closed e c o s y s t e m f r o m w h i c h t h e a t m o s p h e r e was evacuated t o 14 m m pressure and replaced with 750 m m nitrogen. T h e first 37 data p o i n t s are a best fit t o a linear regression, r ~ = 0.99. T h e last 37 data p o i n t s are a best fit t o a p o w e r curve, r 2 = 0.97. (B) a closed e c o s y s t e m w h i c h h a d b e e n covered in a l u m i n u m foil and black c l o t h t o p r e v e n t t h e e n t r a n c e o f sun light. T h e initial P o 2 increase b e f o r e p e r t u r b a t i o n is a best fit to a linear regression (22 d a t a p o i n t s , r 2 = 0.77), w h e r e a s t h e later decrease is a b e s t fit t o an e x p o n e n t i a l curve (48 data p o i n t s , r 2 = 0.98). T h e insert, a m o r e detailed p r e s e n t a t i o n o f t h e data for B t l ~ o u g h d a y 12, gives an e x a m p l e o f a long p e r i o d P o 2 oscillation.
A long-term 8-year-old ecosystem was completely covered to ~ e v e n t the entrance of sun light. The Po2 rate of consumption is shown in Fig. 2B. The arrow marks the point at which the sygtem was covered. After a lapse of 2 days, a definite decrease in PO2 became apparent. The initial oxygen consumption rate appeared to be line,r: continued measurements show the emergence of a deviation from linearity. Both an initial linear ( P 0 2 = m t + b) and then an exponential
curve (Po2 = ae bt) fit the data best. The exponential curve is a slightly closer overall fit with a coefficient of determination of 0.98 (the linear coefficient of determination was 0.97). A curvilinear fit appears reasonable, since the rate of respiration should decrease as heterotrophic communities within the ecosystem become nutrient limited by the absence of continued primary photosynthetic productiori. Thus the Po2 should ultimately approach zero asymtotically.
209
Discussion There are some interesting general aspects to these experimental results. One is the lack of factors controlling final Po2. For example, both A and B ecosystems have oxygen contents significantly above 20.95%, the present day terrestrial value. Indeed, there is still no evidence that B has reached a steady state PO2 value. Since oxygen is produced biologically and apparently is under feedback control, the difference in steady state PO2 must be due to some systematic variation in the distribution of production or consumption mechanisms. Although it is evident that biological consumption factors are present in all ecosystems, many geological factors are missing. The combustion of fossil fuels and weathering are significant geological consumption factors and serve as a sink for about 3% of the total biologically produced oxygen each year {Walker, 1974). Other consumption processes, as volcanism and forest fires, are far less significant, accounting for the use of some 0.0039% of that oxygen produced each year (Walker, 1974; Bakuzis, 1969). It is conceivable that the PO2 of the Earth is not constant over geologic time spans, but is still rising (Silldn, 1964). There are insufficient data to appraise this possibility. Another feature of our results is that there appears to be more than one possible steady state PO2: each ecosystem appears to reach its own unique PO2 value. All our closed ecosystems are exposed to the same sun light intensity of 720 ~E m -2 day -1, but each seems to reach different steady state levels (for example, the oxygen content of the long term system of Fig. 1D is about 14%). We do n o t know at this time whether these various possible steady state PO2 levels are continuous or discrete or ecosystem dependent. Our results show: (i) the biological production and consumption of oxygen can continue in materially closed energetically open microbial ecosystems for at least 8
years, (ii) PO~ changes within closed systems might serve as a metric for ecosystem stability, (iii) a positive feedback control manifest by increased oxygen production under conditions of oxygen removal is suggested, and (iv) various closed ecosystems seem each to approach unique PO2 steady state concentrations ranging from 14% to 30%
Acknowledgement This work was supported by research grant NGR 12-001-109 from the National Aeronautics and Space Administration, and by the University of Hawaii at Manoa.
References Bakuzis, E.V., 1969, The Ecosystem Concept in Natural Resource Management, G.M. Van Dyne (ed.) (Academic Press, N.Y.) pp. 189--254. Berkner, L. and L. Marshall, 1965, Limitation on oxygen concentration in a primitive planetary atmosphere, J. Atmos. Sci. 23, 133--143. Board, P., 1976, Anaerobic regulation of atmospheric oxygen. Atmos. Environ. 10, 339--341. Botkin, D., 1978, Closed regenerative life support systems for space travel: Their development poses fundamental questions for ecological science. Life Sci. Space Res. 17, 3--12. Brown, A.H., 1966, Human ecology in space flight, D. Calloway (ed.) (N.Y. Acad. Sci., N.Y.)pp. 82--119. Commoner, B., 1965, Biochemical, biological, and atmospheric evolution. Proc. Natl. Aead. Sci. USA. 53, 1184--1194. Gitel'son, I.I., 1976, Problems of Space Biology, Vol. 28 (National Aeronautics and Space Administration Technical Translation from the Russian) No. TTF-16993. Holland, H., 1962, in Petrologic Studies: A Volume to Honor A.F. Buddington, A.E.J. Engel, H.L. James and B.F. Leonard (eds.) (Geological Soc. of Amer., Boston) pp. 447--477. Margulis, L. and Lovelock, J., 1974, Biological modulation of the Earth's atmosphere, Icarus 21, 471. Rutten, M.G., 1971, The Origin of Life by Natural Causes (Elsevier Publishing Co.). Sill~n, L.G., 1964, Processes regulating the oxidation state of the system (air + s e a + sediments) in past and present. Acta Chem. Scand. 18, 1016--1018. Taub, F., 1974, Closed ecological systems. Annu. Rev. Ecol. Syst. 5,139--160.
BioSystems, 14 (1981) 205--209 © Elsevier/North-Holland Scientific Publishers Ltd.
MEASUREMENT OF BIOLOGICAL ACTIVITY IN MATERIALLY CLOSED MICROBIAL ECOSYSTEMS
E L I Z A B E T H A. K E A R N S
and C L A I R E. F O L S O M E
Laboratory for Primordial Biology, Department of Microbiology, University of Hawaii at Manoa, Honolulu, Hawaii 96822, U.S.A. (Received March 4th, 1981)
Gas phase oxygen concentrations of materially closed, energetically open miniature microbial ecosystems were measured periodically. Our results indicate: (i) closed systems remain biologically active for at least 9 years, (ii) Po 2 values might serve as an indicator of stability, (iii) each closed ecosystem seems to seek its own unique final Po 2 state, and (iv) ecosystem response to experimentally depleted Po 2 suggests the presence of positive feedback control.
Introduction The prospect of long-term manned space missions has increased interest in the properties of closed ecologies (Gitel'son, 1976; Taub, 1974; Brown, 1966). A reliable metric to assess biological activity and stability in closed systems has yet to be developed. While some theoretical studies have been performed (Botkin, 1978; Board, 1976; Margulis and Lovelock, 1974), there has been little experimental research. To gain qtmntitative insight on the nature of ecosystem stability and activity under closure, we are investigating miniature materially closed, energetically open complex microbial ecosystems. We propose that gaseous oxygen concentration changes can be used as a measure of biological activity. Oxygen concentration is an attractive metric as a potential descriptor of ecosystem states as it is almost entirely biologically produced. Geological mechanisms of production are negligible in comparison (Holland, 1962; Walker, 1974) and the presence of oxygen in the atmosphere is closely associated with the origin and evolution of life (Berkner and Marshall, 1965; Rutten, 1971; Com-
monet, 1965). Furthermore, the thermodynamic properties of this gas have been thoroughly studied and its concentrations are easily measured in small samples.
Materials and methods We devised small closed ecosystems constructed with carbonate sand, water and sediment from shallow coral reefs. Twoliter Pyrex flasks were filled with 100 ml sea water and 100 cc sand and sediment taken from oligotrophic coral reefs at 15 m depths. These flasks were sealed immediately under ambient air with ground glass high vacuum stoppers to prevent matter exchange with the external environment. Flasks were fitted with renewable multilayer teflonfaced sampling ports and were placed by a window receiving NE sun light. As a control, one flask was only loosely covered to allow gas exchange without significant water evaporation. An 8-year-old materially closed system was also employed in these studies. We are aware that these closed miniature ecosystems contain a complex assemblage of of microbial biotypes. Even small volume
206 ecosystems as these presumably contain a range of micro-niches for principal forms of microbial metabolism; photo- and chemoautotrophic and photo-and chemoorganotrophic modes. To date we have not attempted to assay these ecosystems for their specific biotype distribution profiles since experiments as this could perturb a s m a l l
closed ecosystem by removing a metabolic link or by physically disturbing the near isolation of that link. Thus our initial experiments reported here were designed to treat the entire ecosystem flask as a "white box" in which the input (sun light) was eitherunchanged or removed and the system output (P02) was periodically measured.
2~
2~
o~
22 ¸
2
20~21] I "'"" 20"
~B 0
~0
30
2O
z,O
SO
0
I
I
t
I
I
2O
~0
6O
8O
~00
I 80
I00
T, days
0
T, days
21
o~ ." . !
..
0
20¸ 19
0
15-
! 0
• i
!
!
2O
I z,o
I 60 T, days
I 80
I Loo
12
~ 20
I ~0
I 6o
I
T, d a y s
Fig. i . The kinetics o f oxygen concentration change for four microbial ecosystems. Data are presented as linear regression lines derived from 3-day moving average values. Mean and standard deviation were calculated for all horizontal portions. (A) a closed ecosystem, measurements performed from the day o f closure: section 1 comprises 20 data points; the coefficient of determination (r 2) is 0.86; section 2 comprises 43 data points, standard deviation (s), 0.20; section 3, 52 data points, r ~ -- 0.73• (B) a replicate d o s e d ecosystem, taken from the same site as A, measurements performed from the day o f closure: section 1, 25 data points, r ~ = 0.84; section 2, 56 data points, s = 0•16; section 3, 119 data points, r~ = 0.91. ( C ) a n open control system collected from the_~_me site as A and B; 166 data points; s = 0.20. (D) an 8 -year-old closed ecosystem; the initial P o 2 increase marked by 28 data points, r 2 = 0.79; the later horizontal portion comprises 177 data points, s = 0.21.
207 Overt signs of microbial activity are continually manifested in closed ecosystems by the appearance of green, blue~areen, brown and reddish colonies and mats upon the sand and inner walls of the flask. Gas bubbles of varying size enmeshed in these pigmented photosynthetic communities appear and dissolve. Deeper, presumably anaerobic, portions of the sediment show periodically varying black spots 1--5 mm across. Closed microbial ecosystems kept in our laboratory since 1967 still show all these facets of continued biological activity, although it generally is believed that materially closed systems cannot survive for any substantial length of time (Taub, 1974). The gas phases of each experimental system were analyzed by gas chromatography using 2 mm × 2 m molecular seive columns and a Carle micro thermistor bead detector. Small samples of 25 or 50 ~1 were taken daffy and compared to samples of dry air at ambient temperature and pressure. The ratio of the oxygen peak height to the total peak heights of nitrogen and oxygen was assigned a value of 20.95% for a standard dry air. The sample ratio was compared to this standard ratio to compute the relative oxygen content. Changes in total gas pressure within closed flasks were less than 5 mm (less than 0.7%) over the total span of sampling.
Results The kinetics of PO2 change for several ecosystems are shown in Fig. 1. The data were fitted using linear regression analyses. A and B are two flasks monitored daffy for 3 months from the moment of closure. Both show an overall increase in PO2: the PO2 of A increased from 19.5% to 22% by day 40, while the PO2 o f B increased from 19.7% to 24% by day 85. This corresponds to increments of 6% (A) and 16% (B) higher than the control value of 20.7% oxygen of Fig. 1C (note that 20.7% is the standard
oxygen content of moist air, while 20.95% is the value for dry air). While the rates of oxygen increase differed slightly between these two systems, several features are common: (i) both show a step pattern of PO~ increase, and (ii) the rate of PO2 increase before a step is distinctly greater than after a step. For A these values are 0.14%/day, followed by 0.07%/day: for B the rates are 0.10%/day, followed by 0.06%/day. This suggests that a lower PO2 in the gas phase is correlated with a greater rate of oxygen production. The open ecosystem, Fig. 1C, and the long-term closed ecosystem, Fig. 1D, show no overall change in the rate of oxygen production under constant light conditions. The brief rise observed for ecosystem D was due to a change in its location to an area receiving greater sun light (a response we have observed frequently). All the closed ecosystems of Fig. 1 show apparent oscillations of PO2 with periods greater than 24 h. Because this phenomenon could obscure a conventional regression line, the data were analyzed by fitting a regression line using a moving average of successive 3-days' span. The nature of these longer term PO2 periodic fluctuations awaits further analysis. To show that a net production of oxygen did occur, the atmosphere of ecosystem A was evacuated to 14 mm pressure and replaced with nitrogen at 750 mm pressure. The PO2 after evacuation and equilibration of dissolved gas was about 4%. The PO2 began to increase at a rate almost three times greater than that measured before removal (Fig. 2A). The rate before removal of oxygen was 0.07%/day, while after removal it increased to 0.21%/day. This result suggests that low PO2 stimulates a greater rate of oxygen production and implies the presence of a positive feedback mechanism. The restoring capacity of this mechanism appears to be very high since within 35 days the oxygen content had reached half of its amount prior to evacuation.
208 A
B 22 J 17
22
15-
2'
13-
20
..
o~
f8
17
7"
f6
3
o
i ~o
I L,o
,,,
I 6o
I so
I ,oo
T, days
,s
o
I ,o
I ~o
~ ...... 3'o 4'o T, days
I so
I 6o
I
Fig. 2. The kinetics o f o x y g e n c o n c e n t r a t i o n change in t w o closed microbial e c o s y s t e m s w h i c h have b e e n pert u r b e d . (A) a closed e c o s y s t e m f r o m w h i c h t h e a t m o s p h e r e was evacuated t o 14 m m pressure and replaced with 750 m m nitrogen. T h e first 37 data p o i n t s are a best fit t o a linear regression, r ~ = 0.99. T h e last 37 data p o i n t s are a best fit t o a p o w e r curve, r 2 = 0.97. (B) a closed e c o s y s t e m w h i c h h a d b e e n covered in a l u m i n u m foil and black c l o t h t o p r e v e n t t h e e n t r a n c e o f sun light. T h e initial P o 2 increase b e f o r e p e r t u r b a t i o n is a best fit to a linear regression (22 d a t a p o i n t s , r 2 = 0.77), w h e r e a s t h e later decrease is a b e s t fit t o an e x p o n e n t i a l curve (48 data p o i n t s , r 2 = 0.98). T h e insert, a m o r e detailed p r e s e n t a t i o n o f t h e data for B t l ~ o u g h d a y 12, gives an e x a m p l e o f a long p e r i o d P o 2 oscillation.
A long-term 8-year-old ecosystem was completely covered to ~ e v e n t the entrance of sun light. The Po2 rate of consumption is shown in Fig. 2B. The arrow marks the point at which the sygtem was covered. After a lapse of 2 days, a definite decrease in PO2 became apparent. The initial oxygen consumption rate appeared to be line,r: continued measurements show the emergence of a deviation from linearity. Both an initial linear ( P 0 2 = m t + b) and then an exponential
curve (Po2 = ae bt) fit the data best. The exponential curve is a slightly closer overall fit with a coefficient of determination of 0.98 (the linear coefficient of determination was 0.97). A curvilinear fit appears reasonable, since the rate of respiration should decrease as heterotrophic communities within the ecosystem become nutrient limited by the absence of continued primary photosynthetic productiori. Thus the Po2 should ultimately approach zero asymtotically.
209
Discussion There are some interesting general aspects to these experimental results. One is the lack of factors controlling final Po2. For example, both A and B ecosystems have oxygen contents significantly above 20.95%, the present day terrestrial value. Indeed, there is still no evidence that B has reached a steady state PO2 value. Since oxygen is produced biologically and apparently is under feedback control, the difference in steady state PO2 must be due to some systematic variation in the distribution of production or consumption mechanisms. Although it is evident that biological consumption factors are present in all ecosystems, many geological factors are missing. The combustion of fossil fuels and weathering are significant geological consumption factors and serve as a sink for about 3% of the total biologically produced oxygen each year {Walker, 1974). Other consumption processes, as volcanism and forest fires, are far less significant, accounting for the use of some 0.0039% of that oxygen produced each year (Walker, 1974; Bakuzis, 1969). It is conceivable that the PO2 of the Earth is not constant over geologic time spans, but is still rising (Silldn, 1964). There are insufficient data to appraise this possibility. Another feature of our results is that there appears to be more than one possible steady state PO2: each ecosystem appears to reach its own unique PO2 value. All our closed ecosystems are exposed to the same sun light intensity of 720 ~E m -2 day -1, but each seems to reach different steady state levels (for example, the oxygen content of the long term system of Fig. 1D is about 14%). We do n o t know at this time whether these various possible steady state PO2 levels are continuous or discrete or ecosystem dependent. Our results show: (i) the biological production and consumption of oxygen can continue in materially closed energetically open microbial ecosystems for at least 8
years, (ii) PO~ changes within closed systems might serve as a metric for ecosystem stability, (iii) a positive feedback control manifest by increased oxygen production under conditions of oxygen removal is suggested, and (iv) various closed ecosystems seem each to approach unique PO2 steady state concentrations ranging from 14% to 30%
Acknowledgement This work was supported by research grant NGR 12-001-109 from the National Aeronautics and Space Administration, and by the University of Hawaii at Manoa.
References Bakuzis, E.V., 1969, The Ecosystem Concept in Natural Resource Management, G.M. Van Dyne (ed.) (Academic Press, N.Y.) pp. 189--254. Berkner, L. and L. Marshall, 1965, Limitation on oxygen concentration in a primitive planetary atmosphere, J. Atmos. Sci. 23, 133--143. Board, P., 1976, Anaerobic regulation of atmospheric oxygen. Atmos. Environ. 10, 339--341. Botkin, D., 1978, Closed regenerative life support systems for space travel: Their development poses fundamental questions for ecological science. Life Sci. Space Res. 17, 3--12. Brown, A.H., 1966, Human ecology in space flight, D. Calloway (ed.) (N.Y. Acad. Sci., N.Y.)pp. 82--119. Commoner, B., 1965, Biochemical, biological, and atmospheric evolution. Proc. Natl. Aead. Sci. USA. 53, 1184--1194. Gitel'son, I.I., 1976, Problems of Space Biology, Vol. 28 (National Aeronautics and Space Administration Technical Translation from the Russian) No. TTF-16993. Holland, H., 1962, in Petrologic Studies: A Volume to Honor A.F. Buddington, A.E.J. Engel, H.L. James and B.F. Leonard (eds.) (Geological Soc. of Amer., Boston) pp. 447--477. Margulis, L. and Lovelock, J., 1974, Biological modulation of the Earth's atmosphere, Icarus 21, 471. Rutten, M.G., 1971, The Origin of Life by Natural Causes (Elsevier Publishing Co.). Sill~n, L.G., 1964, Processes regulating the oxidation state of the system (air + s e a + sediments) in past and present. Acta Chem. Scand. 18, 1016--1018. Taub, F., 1974, Closed ecological systems. Annu. Rev. Ecol. Syst. 5,139--160.