Microbial growth rates in a cold-core gulf stream eddy of the northwestern Sargasso sea

Deep-Sea Research. Vol. 33, No. 4, pp. 427-446, 1986. Printed in Great Britain.

0198~1149/86$3.00 + 0.(~) © 1986PergamonPressLtd.

Microbial growth rates in a cold-core Gulf Stream eddy of the northwestern Sargasso Sea ROGER B . HANSON,* LAWRENCE R . POMEROYt a n d ROBERT E . MURRAY:~ (Received 5 August 1985; in revised form 4 November 1985; accepted 5 November 1985)

Abstract--Horizontal spatial variations of microbial and bacterial abundance and biomass, and rates of nucleic acid synthesis in surface waters across cold-core Gulf Stream eddy P were investigated in April 1982. The age of eddy P was estimated at 8 to 10 months. Chlorophyll a reached (I.38 p.g 1~ in an area of high chlorophyll a fluorescence located in 18 to 19°C waters. Bacteria were mostly (98%) free-floating, and microbial-ATP biomass correlated with chlorophyll a fluorescence across eddy P. Rates of microbial metabolism and growth, and bacterial production were not proportional across areas of high chlorophyll a fluorescence. Estimates of microbial growth rates (8), were 0.3 to (I.6 h ~in the mixed surface waters of eddy P and 0.06 to 0.08 h ~in adjacent waters of the northwestern Sargasso Sea. Bacterial production ranged from 0.12 to 1.41 ~gC I ~ d ~, and production:biomass ratios (turnover times) were 0.024 to 0.27 h ~ in the eddy and ca. 11.002 h 1 in the Sargasso Sea. Because microbial metabolism (RNA synthesis) and growth (DNA synthesis) across eddy P were not closely coupled, although intertwined with diurnal variations, we conclude that the microbial communities, based on the experimental time scale, were in a state of unbalanced growth.

INTRODUCTION

COLD-WATERanomalies in the Sargasso Sea were first recognized over four decades ago (IsELIN, 1936, 1940). Since then, these anomalies have been described and investigated. PARKER (1971) identified several low temperature fields that exhibited dome-shaped structures and FUGLISTER(1972) applied the term 'rings' to these special features, generally referred to as "Cold-Core Gulf Stream Eddies". Cold eddies are generally generated east of 70°W from unstable meanders of the Gulf Stream (FUGLISTER, 1977; DOBLARand CHENEY, 1977; RICtIARDSON, 1980); the circulation (150 cm s-I) is cyclonic with convergence towards the interior of the eddy (VASTANO and HAGAN, 1977) and a barotropic component that extends to 4500 to 4900 m (KELLY and WEATHERLY, 1985). Cyclonic eddies in the Sargasso Sea are highly variable and those formed in the north may interact with the Gulf Stream many times before coalescing with the Stream between Florida and the New England sea mount chain (LAI and RICHARDSON, 1977; RICHARDSON, 1980). Other cold-core eddies may move southeastward at 5 cm s-I , decay at a rate of 0.3 to 0.6 m d-~ and have a life span of 2 to 4 years in the northwestern Sargasso Sea (PARKER, 1971; CHENEYand RICttARDSON, 1976; RICHARDSON, 1980; THE RING GROUP, 1981; WlEI3E, 1982). * Skidaway Institute of Oceanography, P.O. Box 13687, Savannah, GA 31416, U.S.A. -t Institute of Ecology, University of Georgia, Athens, GA 30602, U.S.A. ~: Department of Microbiology, University of Georgia, Athens, GA 30602, U.S.A. 427

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Microbial growth rates in a cold-core Gulf Stream eddy

429

The planktonic assemblages in various cold-core eddies may be quite different due to seasonal variations in assemblages in slope waters when eddies form (WmBE et al., 1976; WIE~E, 1982). In addition to seasonal variations, planktonic assemblages within Gulf Stream eddies undergo several physiological and structural changes due to nutrient enrichment and geotrophic forces as the eddies interact and decay in the northwestern Sargasso Sea (W1EBEet al., 1976; THE RIN~ GROUP, 1981; YENTSCHand PHINNEV, 1985). The regional importance of phytoplankton and zooplankton assemblages in these Gulf Stream mesoscale eddies to the ecology of the northwestern Sargasso Sea has been reported (WIEBE et al., 1976; ORTNERet al., 1978, 1980; WIEBE, 1982). However, we know of no reports which describe the potential rates of microbial growth and metabolism in cold-core eddies of the northwestern Sargasso Sea. Our study fbcuses on the spatial variations of microbial metabolism and growth relative to measured physical, chemical and biological properties of cyclonic eddy P. This eddy was studied during R.V. Cape Hatteras cruise 09-82, 29 March to 19 April 1982. Our results are examined in reference to the physical, chemical and biological interactions of continental slope water (CSW), Gulf Stream (GS) and Sargasso Sea (SAR) waters (Table 2). M A T E R I A L S AND METHODS

Cold-core eddy P was located approximately 300 km east of North Carolina, U.S.A. (Fig. 1; transedt A). Eddy P was first sighted on 15 September 1981 and had interacted with the Gulf Stream on occasions until it was last seen on 15 May 1982 (Table--13-7. Sampling sites included stations within the core of colder slope water, mixed GS-SAR waters, and in the warmer and more saline SAR water (Table 2). Seawater samples were collected from a depth of 5 m in 5-1 Niskin bottles at several stations across eddy P on 11 April 1982 (Fig. 2). Additional samples were also collected in North American Slope Waters (NASW) off Virginia and an area south of eddy P to provide a reference with which to compare our observations in the eddy. All samples were processed within 1 h of collection. Hydrographic measurements

Surface water temperatures and chlorophyll fluorescence (see below) were continuously monitored. Salinity was determined with a precision salinometer on samples taken from Nisken bottles. Vertical temperature fields were examined with 250 m expendable bathythermographs. Chlorophyll and A TP measurements

Measurements of relative in vivo fluorescence were made by continuously pumping surface seawater through a Turner Designs fluorometer. Phytoplankton for pigment analysis was collected on Reeve Angel 984HA glass fiber filters, homogenized in 100% acetone and measured by the HPLC method of JACOBSEN(1978). Samples were not sparged with nitrogen before storing at -50°C in a light-tight box. Total microbial biomass was calculated from the concentration of particulate adenosine triphosphate (ATP). After seawater samples were filtered through 0.22 lam Millipore filters, ATP was extracted with 5 ml of boiling sodium bicarbonate buffer (0.1 M, pH 8.5) for 2 min. Samples were then cooled, placed in scintillation vials with 5 ml of

430

R . B . HANSON et al.

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Fig. 1. Study area across eddy P (33°N; 73°W) and transect B in the northwestern Sargasso Sea. Stations along transects A and B were made at major discontinuities in temperature and in vivo chlorophyll a fluorescence (11 to 13 April 1982).

cold Tris (tris[hydroxymethyl]aminomethane) buffer (0.1 M, pH 7.8) and stored frozen for later analysis. ATP concentrations were determined by the luciferin-luciferase method of HOLM-HANSENand BOOTH(1966). The fraction of microbial ATP associated with unattached bacteria and small bacterialsize organisms was determined by filtering seawater samples through 0.2 and 0.6 tam pore-size filters. Filters were extracted and samples analyzed as described above. The ATP associated with <0.6 gm fraction picoplankton was calculated from the difference between the ATP on the 0.2 p.m filter and the 0.6 p.m filter. Table 2.

Location of hydrographic station across eddy P (transect A) and along transect B in the northwestern Sargasso Sea on 11 April and 12 to 13 April 1982

Eddy P

(transect A) Sta. No. 75 76 77 78 79 80 Transect B 81 82 83 84 85

(Time)

Water types

(11211) (1335) (16311) (1922) (2128) ({)tl25)

SAR GS/SAR GS/SAR CSW GS/SAR GS/SAR

(1928) (2215) (23511) ({)252) ({)357)

Position

Surface sldinity (1{1 ~)

Bucket temperature (°C)

33°46.2'N 33°52.0'N 33°5(I.11'N 33°50.0'N 33°51.0'N 33051.0'N

71°53.0'W 72°11.0'W 72°35.5'W 73°1~1.0'W 73°17.7'W 73°44.{1'W

36.454 36.511 36.497 36.458 36.546 36.497

21.8 211.3 18.1) 17.5 18.7 19.8

35051 . { I ' N 31°13.5'N 31°26.6'N 31°51.0'N 32°{~1.0'N

74°115.3'W 74°114.4'W 74°111.9'W 73°53.4'W 73°53.lJ'W

36.4911 36.575 36.479 36.588 36.468

21. i 211.3 20.2 19.9 211.9

SAR, Sargasso Sea water; GS, Gulf Stream; CSW, continental slope water.

431

Microbial growth rates in a cold-core Gulf Stream eddy

15"

I0o

.~5o 80 °

70 °

80 °

70 °

Fig. 2. Oceanographic Analysis Charts of cold eddies in the northwestern Sargasso Sea on 13 and 15 April 1982. Eddy P is approximately located at 33°N and 73°W. To the southwest along 30°N, a cold eddy appears to be coalescing with the Gulf Stream as well. These drawings are an interpretation of satellite infrared imagery from NOAA, National Weather Service.

Bacterial abundance Bacterial samples were collected in 5-1 Niskin samplers. Samples were drawn at once and fixed in cold, cacodylate-buffered glutaraldehyde. After staining the cells with acridine orange (HOBmE et al., 1977), replicates of 1 or 2 ml of water were filtered gently through a 0.2 ~tm black Sartorius filter. We have shown previously that there is no difference in counts on 0.2 tam Sartorius cellulose filters and 0.2 ~m Nuclepore filters (PoMEROYet al., 1983). The filter was mounted in low-fluorescence immersion oil and the coverglass sealed with wax. The slides were then stored at 5°C in the dark and counted within 2 weeks after the cruise. A Zeiss standard epifluorescence microscope using a magnification of ×1250 was used for counting 40 fields. Counts of each group were interpreted by analysis of variance followed by F-tests.

Incorporation of [3H]thymidine Rates of [3H]thymidine (Thymidine deoxyribose: TdR) incorporation were determined by the method of FUHRMANand AZAM(1980, 1982). Triplicate 10-ml samples were incubated in the dark at approximate in situ temperature for 4 to 6 h with 3.5 nM [methyl-3H]thymidine (77.2 Ci mmol -~, New England Nuclear Corp.). We have determined that 3.5 nM of labeled TdR underestimates bacterial production by 14% in some mesotrophic waters; however, in oligotrophic systems, we suspect the difference is less. Time course uptake of [3H]TdR was linear for up to 8 h. After incubation, samples were chilled in ice water, extracted in an equal volume of ice-cold 10% trichloroacetic acid (TCA) and filtered through 0.22 lam Millipore filters. Filters were rinsed twice with 3 ml of cold 5% TCA and then transferred to vials containing 10 ml of Scintiverse (Fisher Scientific Co.). Radioactivity was assayed by liquid scintillation spectrometry. Formalinkilled controls were used to measure abiotic adsorption of [3H]thymidine. The rate of [3H]TdR incorporation by free-living bacteria was determined separately. Triplicate 10-ml subsamples were incubated for 4 to 6 h with [3H]TdR, and then gently

432

R.B. HANSONet al.

filtered through 0.6 gm Nuclepore filters. The TCA-insoluble material in the <0.6 lam fraction was collected on 0.22 lam Millipore filters and processed as described above. Bacterial production rates were estimated from [3H]TdR incorporation rates using conversion factor of 2 to 6 x 10 ~8 cells produced per mole of exogenous TdR incorporated (FUHRMANand AZAM, 1982; DUCKLOWand HILL, 1985b). Given the present state of knowledge the use of the TdR method is an acceptable and valid technique to study bacterial growth in the open ocean (DuCKLOWand HILL, 1985b).

Incorporation of [2-3H]adenine Microbial uptake of [3H]adenine was measured from time-course experiments, substrate uptake kinetics and rates of incorporation into microbial RNA (KARL, 1979, 1981; KARLet al., 1981). Water samples (1.5 i) were prescreened through 100 lam mesh Nitex to remove large zooplankton and particles. A 250 ml subsample was incubated in situ temperature in the dark with 0.1 p.Ci m1-1 of [3H]adenine (New England Nuclear Corp.). Subsamples (25 ml) were removed over 0 to 5 h and filtered through 0.2 p.m Nuclepore filters. After 3 and 5 h of incubation, an additional 25 ml was filtered through 0.6 lam Nuclepore filters. Filters were rinsed 3 times with 0.2 gm filtered seawater, reacted with 5 ml of cold 5% TCA for 5 min, and rinsed 3 times with cold 5% TCA. The filters were then placed in scintillation vials with 0.2 ml Soluene-350 for 1 h before adding 10 ml of scintillation fluor. Radioactivity was determined on board ship with a Beckman LS-100 liquid scintillation spectrometer. The percentage of [3H]adenine incorporated by 0.6 p.m filterable organisms was calculated as [(dpm on 0.2 t~m filter-dpm on 0.6 p.m filter)/dpm on 0.2 p.m filter] x 100. Microbial community growth rate (6) can be estimated from the time-course kinetics of labeled substrate uptake (CUHELe t al., 1983; LI, 1984). T h e growth rate (6) of cells incorporating [3H]adenine can, however, overestimate the specific growth rate (g) of the population when fast growers constitute a small proportion of the total microbial assemblage and when unbalanced growth occurs; that is, when incorporation and cellular growth are not coupled. Therefore, 8 and ta cannot be interchanged unless la determined from cell division or biomass measurements equals 6. To estimate ~ for microbial populations, time-course data were transformed to natural logarithms and then fitted to the equation: In ([3H]adenine = b + at), where a is the slope of the linear regression line. Substrate kinetics of [3H]adenine incorporation into TCA-insoluble material were examined by incubating 5 subsamples, ranging in volume from 200 to 50 ml, with 0.005 to 0.2 laCi ml-l resulting in a concentration range of added adenine of 1.67 to 66.8 nmoles 1-l. After an incubation period of 3 to 4 h, which insured adequate uptake of labeled adenine at the lowest concentration, 50 ml subsamples were extracted with 5% cold TCA, held on ice for 15 min and 25 ml aliquots were filtered on 0.45 ~tm HA Millipore filters. Filters were rinsed with cold 5% TCA and dissolved in 10 ml of scintillation fluor (PCS, Amersham Corporation) before assaying for radioactivity. Microbial affinity for [3H]adenine was assessed after data transformation, i.e. t/f vs substrate concentration, where f is the fraction of radioactivity assimilated into TCAinsoluble material relative to that added to the seawater and t is the incubation time in hours. Linear regression analysis was used to determine the equation that best describes the relation between.t/f and [3H]adenine concentration. This equation defines Vm,,x (1/ slope), turnover time (y-intercept) and K + Sn (x-intercept). A Hewlett Packard 9825

Microbial growth rates in a cold-core Gulf Stream eddy

433

statistical program provided correlation coefficients (r2) and ANOVA (F-tests) for each linear regression analysis. Rates of RNA synthesis were estimated from the incorporation of [3H]adenine into RNA (KARL, 1979, 1981). For these measurements, 0.1 gCi m l -I of [3H]adenine was added to three 250 ml subsamples; one subsample was treated immediately, and the other two were incubated in the dark at in situ temperature for 4 h. Macromolecules were precipitated with cold 5% TCA, RNA fraction hydrolyzed with NaOH and macromolecules reprecipitated with cold TCA. DNA fraction was hydrolyzed in 5% TCA at 100°C, and the protein fraction precipitated at 0°C and filtered. Rates of [3H]RNA and [3H]DNA synthesis were determined by difference (HANSONand LOWERY, 1983). RESULTS

Hydrography

Several cold eddies were identified in the northwestern Sargasso Sea by satellite infrared images (Oceanographic Analysis Charts from N O A A , National Weather Service) as possible cyclonic eddies (Fig. 2). There was no chart available for the day of the study ( l l April 1982), but from charts from 13 and 15 April 1982 (Fig. 2), it can be seen that eddy P had not moved substantially over the 3 days. The cold core of this eddy was located at approximately 33°50'N 73°00'W. The area (transect A) was first surveyed to determine the horizontal scales of surface temperature and relative in vivo fluorescence. Cyclonic circulation was noted by deflections in the ship's course at both western and eastern boundaries of eddy P. After the horizontal scales of the surface cold water and relative in vivo fluorescence anomalies were determined (Fig. 3), hydrographic (salinities and XBT) and microbiological properties were measured at selected locations along transect A. Initial recordings showed the presence of surface waters of different in vivo fluorescence and temperature entrained in the warmer Sargasso Sea (Fig. 3). Bucket temperatures (Table 2) and horizontal-vertical temperature (Figs 3 and 4) indicated that the core of lower temperature water was at Sta. 78. In vivo fluorescence and horizontal temperature fields indicated an entrainment and mixture of different waters across eddy 23-

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Fig. 3. Continuoustemperature and in vivo fluorescencemeasurementsin surface waters across eddy P in the northwestern Sargasso Sea on 11 April 1982.

434

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Station Numbers

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Station Numbers

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Vertical temperature fields determined from X B T across eddy P (transect A) on 11 April 1982 and along transect B on 12 to 13 April.

P (Fig. 3). The surface salinities and temperatures of the water near the center of the eddy were 36.458%0 and 17.5°C, or 1.472%o and 5.5°C higher than that measured in the slope waters (37°51'N 73°32.8'W) off Virginia on 1 April 1982. Cold eddies are generally defined by depressed salinities and upward displacement of isotherms from the mean temperature field (LAI and RICHARDSON, 1977). Vertical temperature fields down to 250 m across the cold water anomaly indicated dome-shape isotherms typical of cold Gulf Stream eddies (Fig. 4). In waters south of eddy P on 12 to 13 April 1982, surface temperature structure and in vivo fluorescence varied erratically across transect B (Fig. 5) as did the vertical temperature fields in the upper 250 m (Fig.

4).

Plankton abundance and biomass

High phytoplankton biomass, usually expected at the center of cold eddies, was not observed in eddy P. Rather, two major chlorophyll a fluorescence bands, with consider-

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Continuous temperature and chlorophyll a fluorescence m e a s u r e m e n t s in the surface waters along transect B in the northwestern Sargasso Sea on 12 to 13 April 1982.

435

Microbial growth rates in a cold-core Gulf Stream eddy

Table 3. Standing stocks o f free bacteria, attached bacteria, and microbial A TP in surface waters (5 m) of eddy P (transect A) and along transect B in the northwestern Sargasso Sea on 11 and 12 to 13 April 1982 Eddy P (transect A) Sta. No. 75 76 77 78 79 80 Transect B 81 82 83 84 85

Microbial Free bacteria (10 -~ ml 1)

Attached bacteria

6.33 + 0.65

22.7 _+ 44.6

nd

nd

6.10 + 0.55

18.6 _+ 34.6

(10 2 ml ~)

nd

nd

4.35 + 0.47 3.66 + 0.39

7.9 + 13.6 6.1 + 13.0

5.15 4.03 4.72 5.02 7.08

+ 0.44 + 0.34 + 0.57 _+ 0.51 + 0.56

23.0 19.4 28.3 12.4 8.3

+ 47.7 + 33.1 + 32.5 _+ 20.5 _+ 113.3

ATP (lag I ~) 0.167 0.309 0.215 0.141 0.167 0.139 0.097 0.068 0.102 0.094 0.209

nd = no data.

able variation within each band, were located at Stas 77 and 79 adjacent to the cooler waters at the center of eddy P (Fig. 3). Within each major band, the spatial scales of phytoplankton patches were about 5 to 10 km. Chlorophyll a and phaeophytin concentrations were highest at Sta. 79 (0.377 and 0.221 p.g 1-~, respectively) and at Sta. 77 (0.297 and 0.505 p.g 1-1, respectively). The chlorophyll a values were, however, approximately 50% lower than concentrations in slope waters (Table 4). Free-living and attached bacteria occurred at densities of 3 to 6 x 105 ml-~ and 0.62 to 2.27 x 10 3 ml-t, respectively, in the waters of eddy P (Table 3). Nearly 98% of the bacteria were free-floating in surface water of the eddy, not unlike those densities and percentages in slope waters (Table 4). The spatial distribution of microbial ATP (Table 3) followed that of the in vivo fluorescence (Fig. 3). ATP concentrations in the eddy were 4 to 10 times lower than those in slope waters (Table 4). Bacterial densities in the waters south of eddy P (transect B) were similar to those in the eddy while microbial ATP was 1 to 3 times lower than in the eddy (Table 3). Only the surface water at Sta. 85 had elevated levels of chlorophyll a (0.43 gg 1-~), microbial ATP and bacteria (Table 3). These elevated values might be associated with eddy P. Table 4. Physical, chemical and biological properties of NA S W at 37°51 'N : at 73°33'W on 1 April 1982. Water samples collected from 10 m water depth at 0945 local standard time

Temperature (bucket, °C) Salinity (%0) Chlorophyll a (lag 1 l) Phaeophytin (lag I ~) Microbial A T P (lag 1 i) % A T P in < 0 . 6 m fraction Free bacteria (10 5 ml -I)

Attached bacteria % Free-floating Thymidine incorporation (pmol 1 i d i) % Incorporation in < 0 . 6 lam fraction Specific incorporation rate (moles cell ~ d 1) Adenine incorporation (pmol 1-l d -I) % Incorporation in < 0 . 6 lain fraction

12 34.986 (I.59 I).16 1.10 26 4.16 + 0.45 (I.22 + 0.24 95 238 _+ 4 88.7 5.7 x 10 I,, 2090 + 161 88.0

436

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Fig. 6. Time course (0 to 300 min) of [3H]adenine accumulation and [3H]adenine saturation kinetics (1.67 to 66.8 nmoles 1 1) for microbial communities at 6 stations across eddy P in the northwestern Sargasso Sea on 11 April 1982. Time-course experiments received 0.1 I.tCi ml-~ of [3H]adenine (33.4 nmoles l-I). Saturation kinetics (0.005 to 0.2 laCi ml 1) were incubated for 3 to 4 h. All curves were hand drawn.

Kinetics of [3H]adenine uptake Time-dependent microbial uptake of [3H]adenine across eddy P indicated two sites of high potential growth rates in areas of high relative in vivo fluorescence (Stas 77 and 79; Fig. 6). Uptake of [3H]adenine continued for 5 h. In waters near the western margin of eddy P (Sta. 80), initial uptake rates were as rapid as those at Sta. 80, but uptake continued for only 1 h. At the eastern margin of the eddy (Stas 75 and 76) uptake rates 40~ oSta No 8 3

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437

Microbial growth rates in a cold-core Gulf Stream eddy

Table 5.

Adenine saturation kinetics for microbial population in eddy P (transect A) and along transect B in the northwestern Sargasso Sea on 11 and 12 to 13 April 1982

Eddy P (transect A) 75 76 77 78 79 80

Vm,x

K + Sn

(nmoles 1-l h l)

(nmoles 1'l)

r2

F-test

501 620 51 112 152 86

0.021 0.019 0.135 0.028 0.035 0.044

23.7 5.3 26.1 12.9 15.1 11.1

0.88 0.85 0.94 0.99 0.96 0.98

10.3" 7.8* 24.7* 116.0"* 39.1"* 81.3"*

368 + 94 719 + 150 155 + 18

0.031 0.019 0.066

11.4 14.0 10.4

0.99 0.99 1.00

139.0"* 136.0"* 950.0**

Tt + S.E. (h)

Sta. No.

1113 272 193 456 420 252

+ + + + + +

Transect B 83 84 85 * = 0.05, ** = 0.005.

were low and ceased after 2 h. [3H]Adenine uptake in the center of the eddy was similar to those on the eastern margin but rates were sustained for 4 h. Transformation of [3H]adenine time-course data to In vs t and statistical analysis of the transformed data showed that ~ (h -t) for microbial populations across eddy P were 0.24 + 0.6 (S.E.), 0.29 + 0.08, 0.47 + 0.01, 0.34 + 0.06, 0.42 + 0.07 and 0.47 + 0.17

at Stas 75 to 80, respectively. Correlation coefficients (r 2) for the transformed data ranged from 0.88 to 0.95 (P < 0.05, F-test). Outside the influence of eddy P (transect B), 6 rates of microbial populations were 0.066 + 0.036, 0.084 + and 0.552 + 0.240 h-l at Stas 83 to 85, respectively. [3H]Adenine saturation kinetics for microbial populations across eddy P indicate that most populations were saturated with 17 nM of added adenine (Fig. 6). Saturation curves, when linearized by t/f transformations, indicated that the population with the highest maximum uptake velocity and fastest turnover time (8d) for adenine was at Sta. 77 (Table 5). The slowest turnover time of 46 days was at the eastern margin of the eddy. South of eddy P the fastest uptake kinetics were measured at Sta. 85 in a high in vivo 50-, I= 40'

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Rates of [3H]adenine incorporation into microbial R N A (o) and [3H]thymidine incorporation into bacterial D N A ( e ) across eddy P in the northwestern Sargasso Sea on 1 i April 1982. For [3H]adenine experiments, bars represent range of duplicate samples. Coefficient of variation (S.D./x) for the [3H]thymidine experiment ranged from 0.30 to 1.00. Fig. 8.

438

R . B . HANSON et al.

flourescence area. Microbial populations were again saturated with 17 nM [3H]adenine. Values for the adenine saturation kinetics are given in Table 5. Rates of [3H]adenine incorporation into RNA and [3H]TdR incorporation into DNA showed two different patterns of incorporation across eddy P (Fig. 8). Highest rates of adenine incorporation were in the eastern band of the high in vivo fluorescence (Sta. 77), similar to saturation kinetics (Table 5). However, the highest rates of TdR incorporation were found in the western band of high in vivo fluorescence (Sta. 79). Microbial and bacterial incorporation rates of [3H}adenine and [3H]TdR in eddy P were 10 to 100 times lower than those measured in slope waters (Table 4) and similar to those measured in the waters south of eddy P (Table 6). Bacteria at Sta. 79 also had the highest specific incorporation rate, 10.8 × 10 -20 moles cell-~ d -~, whereas the lowest specific rate of 2.8 × 10-2o was at Sta. 75 near the eastern margin of the ring. These specific rates in the eddy were 5 to 20 times lower than that measured at the reference station in the NASW (Table 4). Rates of [3H]adenine incorporation into DNA across eddy P showed elevated rates of DNA synthesis in both high in vivo fluorescence areas (Fig. 9), similar to the spatial patterns of [3H]TdR incorporation into DNA (Fig. 8). The rates of [3H]adenine incorporation into DNA were 6 to 10% of its incorporation into RNA, except at Sta. 77 where the percent incorporated into DNA shifted to 33% (Fig. 9). The percentage of [3H]adenine incorporation into protein was only 3 to 6% of the amount incorporated into RNA. Size fractionation

Microbial biomass and percent activity associated with smaller than 0.6 jam microbes and larger than 0.6 jam microbes were measured in the surface waters of eddy P and south of eddy P (Table 7). Most (80 to 85%) microbial uptake of labeled adenine in eddy P was by large (>0.6 jam) microorganisms (phytoplankton, some bacteria, and maybe protozoans). At the eastern margin of the eddy at Sta. 75, almost 50% of the radioactivity was in the <0.6 jam size class while at the western margin at Sta. 80, 95% of the radioactivity was in the >0.6 jam fraction. We also observed a time-dependent decrease in the percentage of labeled adenine in the <0.6 jam fraction, especially east of Sta. 78 (Table 7). Unlike in most oceanic waters, the uptake of labeled TdR by bacteria in the <0.6 jam fraction was not high, e.g. 30 to 60% in the mixed waters of eddy P and in the Sargasso Sea south of eddy P (Table 7). Time-dependent uptake of labeled TdR in each size fraction was not measured. Table 6.

Rates oJ" [3H]adenine incorporation into microbial R N A and [~H]thymidine incorporation into bacterial D N A across transect B in the northwestern Sargasso Sea on 12 to 13 April 1982 Rates of adenine incorporation (pmoles 1 i d 1)

Sta. No. 81 82 83 84 85

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Microbial growth rates in a cold-core Gulf Stream eddy

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Table 7. Size fractionation of [3H]adenine uptake, [~H]thymidine incorporation and microbial A TP ill surface waters from eddy P (transect A) and along transect B in the northwestern Sargasso Sea on 1I and 12 to 13 April 1982

Eddy P (transect A) Sta. No.

% Adenine uptake by 0.6 p.m filterable organisms

% Thymidine incorporation by 10.6 ~tm filterable organisms

% of total ATP in <0.6 lam fraction

75 76 77 78 79 80 Transect B 81 82 83 84 85

46"(28)t 18 (11) 16 ( 4 ) 16 ( 0 ) 21 (29) 5 (8)

nd 60 nd 31 35 37

69 58 41 43 41 29

nd nd 28 (28) 52 (15) 17 (15)

nd nd 46 36 69

nd nd 46 58 56

* % Adenine uptake by 0.6 lam filterable organisms after 3h. i % Adenine uptake aftcr 5 h. nd = no data.

M o s t ( 6 0 % ) m i c r o b i a l b i o m a s s in the surface w a t e r s o f e d d y P was also a s s o c i a t e d with m i c r o o r g a n i s m s > 0 . 6 lam, p r e s u m a b l y p h y t o p l a n k t o n , large b a c t e r i o p l a n k t o n , a n d p r o t o - a n d m i c r o z o o p l a n k t o n ( T a b l e 7). In w a t e r s east of Sta. 76 in e d d y P a n d in the S a r g a s s o Sea, m o s t (60 to 7 0 % ) m i c r o b i a l b i o m a s s was a t t r i b u t e d to <(/.6 lam o r g a n i s m s , p r e s u m a b l y minicell b a c t e r i o p l a n k t o n , ultra small p h y t o p l a n k t o n a n d e u c a r y o t i c g r a z e r s (FUHRMAN a n d MCMANUS, 1984). DISCUSSION Spatial heterogeneity across eddy P

P l a n k t o n i c c o m m u n i t i e s in cyclonic G u l f S t r e a m e d d i e s are a f f e c t e d by the age o f the e d d y , e n t r a i n m e n t o f G S a n d S A R w a t e r s with s l o p e w a t e r , biological a n d s e a s o n a l fluctuations in s l o p e w a t e r d u r i n g e d d y f o r m a t i o n , t e m p e r a t u r e , salinity a n d n u t r i e n t

440

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al.

gradients along isopycnals, and gravitational, vorticity and Coriolis forces. Our results show that considerable spatial variation of microbial communities and processes exists across the surface waters of eddy P. Although cyclonic Gulf Stream eddies appropriate slope water on the scale of several hundred kilometers, phytoplankton and bacterioplankton were differentially distributed on scales of 5 to 10 km in eddy P. Two areas of high in vivo fluorescence (phytoplankton) were found adjacent to the core of cold water in eddy P (Fig. 3). The location of these areas relative to the center of the eddy suggests a concentric ring of high phytoplankton biomass around the ring core. However, this is unlikely because of the spatial distribution of microbial and bacterial growth rates (Figs 8 and 9, Table 5), surface temperature fields, and spatial heterogeneity of phytoplankton across each area (Fig. 3). There are several possible explanations for the spatial variation of biological components in eddy P. One explanation is that during eddy P formation and subsequent interaction with the Gulf Stream (Table 2) the cyclonic rotation of the eddy pulls streamers of mixed SAR and GS waters around itself. Satellite infrared images of Gulf Stream eddies show asymmetrical streamers surrounding the core of water appropriated from either side of the Gulf Stream (WIEBE, 1982). Another explanation for the spatial heterogeneity of plankton across eddy P is the resultant geostrophic and rotary currents that deliver deep nutrient-rich waters into the high velocity region (YENTSCHand PHINNY, 1985). Finally, frictional forces (e.g. winds) over the surface layer can distribute surface waters ('washover') disproportionately across the eddy.

Natural history of eddy P Many cyclonic cold-core eddies form east of 70°W in the North Atlantic. Based on monthly oceanographic charts eddy P had probably generated off the Carolinas near 33°N 74°W before 15 September 1981 and had interacted with the Gulf Stream on occasions (Table 1). The exact location and time of eddy P formation is difficult to fix due to a lack of suitable oceanographic (satellite images) charts for the month prior to September 1981 over this particular area off the Carolinas. Based on vertical temperature fields (Figs 3 and 4), the age was set at approximately 10 months, assuming the 17°C isotherm was at the depth of 50 m at formation time and sank at a rate of 0.6 m d-~ (PARKER, 1971). Consequently, from this estimation and that of oceanographic charts eddy P was at least 8 to 10 months old when our study was done. Eddie P was last seen 15 May 1982. The horizontal scale of eddy P was estimated at 200 to 250 km. We made our estimates on the separation of the 22°C isotherm. Eddy P was possibly wider than 200 to 250 km as transect A was not across the center of the eddy (Figs 1 and 2).

Microbial and bacterial growth dynamics Microbial growth and bacterioplankton dynamics in the waters of cyclonic eddies have not been previously reported, to our knowledge. Microbial growth rates (~) in the surface waters of eddy P ranged from 0.3 to 0.6 h-~ compared to 0.06 to 0.08 h-~ in other waters of the northwestern Sargasso Sea. The generation times (i.e. 1/~5) of the microbial community ranged from 2.1 to 4.2 h in the cyclonic eddy and 11.9 to 15.2 h in the Sargasso Sea. These estimates of microbial growth rates and generation times based on [3H]adenine incorporation time courses are similar to values reported for other oceanic waters (Table 8). However, we caution the extrapolation of 6 values to specific growth rates (ta). These values often do not equate as have recently been shown by DUCKLOWand

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HILL (1985a,b). Therefore, any attempt to estimate microbial production from our measurements of time-dependent [3H]adenine incorporation (6) and microbial biomass (ATP) would be misleading and incorrect. First, because internal adenine pool size is unknown, we are unable to estimate isotopic dilution in the community. D e n o v o adenine synthesis was assumed blocked at the level of added [3H]adenine. Second, we cannot assume that most of the active microbial population constitutes a significant fraction of the microbial ATP. Protozooplankton and microzooplankton can represent at times a substantial fraction of the microbial ATP biomass in pelagic ecosystems (KARL, 1980; BANSE, 1980). Third, even though nM concentrations of labeled adenine may be utilized by both bacteria and phytoplankton (KARL, 1981), protozooplankton and microzooplankton may have assimilated labeled adenine at 33 nM concentration. Finally, even if metazoans could assimilate labeled adenine, metazoans ATP:C ratios are 2 to 5 times lower than those for bacteria and phytoplankton (KARLet al., 1978; KARL, 1980) and metazoan growth rates can range over several orders of magnitude (BANSE, 1982a,b). So, without knowing how [3H]adenine incorporation is partitioned among various biological components, what is the biomass of the active microbial population, and how [3H]adenine is diluted in the adenine pool, we cannot reliably convert [3H]adenine incorporation growth rates (~5) to carbon on the assumption that ~ = ta and [ATP] = active microbial cells. Only a few estimates of bacterioplankton production have been reported for central oceanic regions. [3H]TdR incorporation into DNA (FUHRMANand AZAM, 1980, 1982), frequency of dividing bacterial cells (HAGSTROMet al., 1979), and experimental variations (e.g. seawater cultures, size fractionation, dilution, and time-dependent assays), can be used to evaluate bacterioplankton growth dynamics. Yet, available techniques are still limited in application because of various procedural, technical and biological problems inherent in each technique (CHRISTIANet al., 1982; FUHRMANand AZAM, 1982; DUCKLOW and HILL, 1985b). One particularly difficult problem with the thymidine technique is determining the correct conversion factor. A wide range of valid conversion factors of <1 to 70 × 1018 cells produced per mole of thymidine incorporated has been derived (FUHRMANand AZAM, 1982; KIRCHMANet al., 1982; MORIARTY, 1984; DUCKLOWand HILL, 1985b) indicating considerable variability in conversion factors. DUCKLOWand HILL (1985b) reported a conversion factor for oceanic waters (SAR and NASW) of 2 to 6 × 1018 cells per mole. Using this factor and 5 x 10-15 g carbon per cell for oceanic minibacteria (WATSONet al., 1979), we estimated that bacterioplankton production ranged from 0.12 to 1.41 p.gC 1-1 d-l in surface waters of cold eddy P. In warm-core Gulf Stream eddies, bacterial production measured by [3H]TdR incorporation ranged from 0.14 to 8.4 tagC 1-j d -j (PEELEet al., 1985; DUCKLOW, 1984; DUCKLOWand HILL, 1985b). Specific incorporation rates of 3 to 11 × l0 -2° moles cell-1 d -1 were within the range of 0.5 to 50 × 10-2o for warm-core eddies (DucKLOWand HILL, 1985b; PEELEet al., 1985). From seawater culture techniques, DUCKLOWand HILL (1985a) estimated bacterial production in warm-core Gulf Stream eddies at 0.24 to 16.3 lagC !-~ d -~ and specific growth rates (~t) of 0.72 to 2.8 d -l. Bacterioplankton production:biomass ratios (turnover times) in eddy P, calculated from production ([3H]TdR data) and biomass (AODC data), ranged from 0.056 to 0.65 d-l, whereas in warm-core Gulf Stream eddies, P:B ratios ranged from 0.48 to 3.6 d -I (PEELEet al., 1985; DUCKLOWand HILL, 1985a).

Microbialgrowthrates in a cold-coreGulfStreameddy

443

Microbial biosynthetic response Cold-core Gulf Stream eddies are not isolated bodies of slope water. The cyclonic rotation of cold eddies mixes slope water with the Sargasso Sea to considerable depths in the Sargasso Sea (ScHMITZand VASTANO, 1975, 1977; KELLYand WEATHERLY, 1985) resulting in new and different physical and biological conditions unlike either original water mass. Accompanying these physical and biological changes, geotrophic forces along the eddy isopycnals may supplement nutrients in the mesotrophic waters of the eddy from the deep Sargasso Sea (YENTSCHand PHINNEY,1985). Consequently, plankton in cold-core eddies experience many environmental conditions over the life span of the eddy. For example, biological properties are highly variable due to seasonal variations in slope water at the time of eddy formation and diel changes in the northwestern Sargasso Sea (WlEBEet al., 1976; ORTNERet al., 1978, 1980; THE RING GROUP, 1981; WIEBE, 1982). Intertwined with these variables are salt and heat exchanges with the Sargasso Sea. For example, as waters in the eddy warm, slope plankton that are trapped in cyclonic eddies must migrate down with the sinking isotherms. Eventually slope plankton are progressively infiltrated by smaller, less abundant forms of the Sargasso Sea (WIEBEet al., 1976; THE RlNG GROUP, 1981; WIEBE, 1982). Microbial communities exhibited differential growth responses across cold-core eddy P. For example, although most communities in the eddy saturated at <20 nM of added [3H]adenine, the community in the eastern phytoplankton patch of the eddy showed no tendency to saturate at 67 nM of added [3H]adenine (Fig. 6). This community also sustained metabolism (uptake of 33nM [3H]adenine) for more than 5 h and most (90%) of the [3H]adenine was incorporated into RNA and not DNA suggesting a high metabolic potential (RNA synthesis) of the microbial community. Microbial populations in the western phytoplankton patch, on the other hand, saturated at <20 nM of added [3H]adenine and metabolism continued for more than 5 h. However, a large fraction (33%) of [3H]adenine uptake was diverted from RNA synthesis to DNA synthesis, suggesting a high growth potential (DNA synthesis) of the microbial community. These differential growth dynamics of the microbial communities are probably intertwined with diel differences, change in community structure, and proportion of growing/metabolizing microbial cells across eddy P. Microbial populations enclosed in cyclonic Gulf Stream eddies are also experiencing daily changes in heat/salt flux and nutrient enrichment/deprivation over the life span of the eddy. The rapid growth rates (1 to 2 d-l) of many of these microbial populations provide these populations with the potential to respond and adjust their metabolism with a change in environmental conditions. Our results, although intertwined with diel and community differences, indicate that microbial populations are possibly in a state of unbalanced growth (JANNASCH,1974; GOLDMANet al., 1979; HANSONand LOWERY, 1983). For example, rates of RNA and DNA synthesis of microbial populations across eddy P and the preferential utilization of [3H]adenine in specific biosynthetic processes suggest that metabolism (RNA synthesis) and growth (DNA synthesis) of these communities were not tightly coupled. Therefore, we conclude that microbial communities are in a state of unbalanced growth as a result of the asynchrony of biosynthetic processes across eddy P due to diel changes, heat flux, Gulf Stream interactions, and nutrients upwelling in the high velocity region of cold-core eddies.

444

R . B . HANSONet al.

Acknowledgements--This research was supported in part by NSF grant OCE81-10707. We thank Philip McGillivary for scientific assistance and the officers and crew of the R.V. Cape Hatteras for their logistic support on the cruise. We also thank S. Nishino and S. Earhart for technical assistance. Anna Boyette and Suzanne Mclntosh drafted the figures and Linda Land typed the manuscript. We also thank Drs Robert Christian and David Menzel for reading and commenting on the manuscript. Oceanographic analysis charts were redrawn from satellite infrared image charts courtesy of Jennifer Clark of NOAA, National Weather Service, Washington, D.C.

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WIEBE P. H., E. M. HULBURT, E. J. CARPENTER, A. E. JAHN, G. P. KNAPP, Ili, S. H. BOYD, P. B. ORTNER and J. L. Cox (1976) Gulf Stream cold core rings: large-scale interaction sites for open ocean plankton communities. Deep-Sea Research, 23, 695-710. WINN C. D. and D. M. KARL (1984) Microbial productivity and community growth rate estimates in the tropical North Pacific Ocean. Biological Oceanography, 3, 123-145. YENTSCH C. A. and D. A. PHINNEY (1985) Rotary motion and connection as a means of regulating primary production in warm core rings. Journal of Geophysical Research, 90, 3237-3248.