Spatial and temporal distributions of prochlorophyte picoplankton in the North Atlantic Ocean

Spatial and temporal distributions of prochlorophyte picoplankton in the North Atlantic Ocean

Deep-Sea Research, Vol. 37, No. 6, pp. 1033--1051. 1990. Printed in Great Britain. 0198-0149/90 $3.00 + 0.00 (~) 1990 Pergamon Press pie Spatial and...

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Deep-Sea Research, Vol. 37, No. 6, pp. 1033--1051. 1990. Printed in Great Britain.

0198-0149/90 $3.00 + 0.00 (~) 1990 Pergamon Press pie

Spatial and temporal distributions of prochlorophyte picoplankton in the North Atlantic Ocean ROBERT J. OLSON,* SALLIE W. CHISHOLM,t ERIK R. ZETrLER,* MARK g . ALTABET* and JEFFREY g . DUSENBERRY*t

(Received 12 October 1989; in revised form 21 February 1990; accepted I March 1990) Abstract---Over the past several years red-fluorescing picoplankton, believed to be prochlorophytes, have been shown to be extremely abundant in the North Atlantic and Pacific Oceans. The dim fluorescence of these tiny cells initially limited studies to the relatively highly pigmented cells near the bottom of the euphotic zone; however, improvements in sensitivity of flow cytometry now enable us to detect the prochlorophytes in surface waters as well. In the Sargasso Sea in May 1988 and May 1989prochlorophytes were present throughout the upper water column, and we observed the highest concentrations in surface waters within the Gulf Stream. Further south, the prochlorophytes formed subsurface maxima; the median depth of prochlorophyte (but not Synechococcus) populations followed the deepening of the nitracline. Prochlorophytes were not present in waters north of the Gulf Stream in May, although we had observed them there on a previous September cruise. They were present year round near Bermuda, with lowest concentrations in the winter, when Synechococcus was most numerous; the prochorophytes appear to "bloom" later than the Synechococcus, after the onset of seasonal stratification. The latitudinal variations in prochlorophyte and Synechococcus distributions during spring resembled the seasonal pattern near Bermuda.

INTRODUCTION EXTREMELY abundant (typically 105 ml - t ) and small ( < 0 . 8 / z m diameter) red-fluorescing cells have recently been observed by flow cytometry in the Atlantic and Pacific Oceans, and the Mediterranean Sea (CHISHOLM et al., 1988; LI and WOOD, 1988; NEVEUX et al., 1989; VAULOT et al., 1989). Although the identity of these cells is still not definitively established, the available evidence suggests that they are p r o k a r y o t e s without phycobiliproteins and with both chlorophylls a and b (CmsHOLM et al., 1988; GOEPaCKE, 1990). Thus they seem to fit LEWIN'S (1981) description of prochlorophytes, a group which was previously known to exist in the oceans only as a symbiont in colonial tunicates. However, H P L C analyses (CHISHOLM et al., 1988; GOEmCKE, 1990) suggest that both the chlorophyll a and the chlorophyll b in these cells are divinyl-like chlorophylls (BAZZAZ, 1981), and that an additional, chlorophyll c-like pigment is also present. These cells are thus different from other prochiorophytes, but for the present we will continue to use this name. Pigments with characteristics similar to those of the prochlorophytes we observed have been reported to occur in high concentrations in the oligotrophic tropical Atlantic and Caribbean Oceans (GmSKES and KgAAY, 1983), the B a n d a Sea (Indonesia) (GI~KES et al., 1988), the Sargasso Sea (GOERIeKE, 1990), the Pacific O c e a n (VERNET, 1983) and the *Woods Hole Oceanographic Institution, Woods Hole, MA 02543, U.S.A. tMassachusetts Institute of Technology, Cambridge, MA 02139. U.S.A. 1033

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Mediterranean Sea (VAULOT, personal communication). GOERICKE (1990) has reported that divinyl-like Chl a can account for nearly half of the total chlorophyll in waters off Bermuda over much of the year. These observations, in addition to the sheer numbers of these cells observed by flow cytometry, indicate that this previously unrecognized group of phytoplankton is an important component of oceanic ecosystems. Summer and autumn cruises in the North Atlantic and Pacific revealed prochlorophytes to be present at every oceanic station occupied except in shallow and well-mixed waters on Georges Bank off Cape Cod, and in most cases these were the most numerous phytoplankters present (CHISHOLMet al., 1988). The fluorescence detection limits of the instruments used by us (Coulter EPICS V) and by other workers [Becton-Dickinson FACS Analyzer (LI and WOOD, 1988); EPICS 741 (NEVEUX et al. 1989)] limited studies to cells in relatively low light conditions deep in the water column, in which photoadaptation had produced cells with high pigment (and hence fluorescence) levels. In typical ocean waters in summer we were not able to detect all the prochlorophytes in samples above the depth of the nitrite maximum (near the 1% isolume); shallower populations gradually disappeared with decreasing depth into the noise on the flow cytometric signatures, due to decreasing cell fluorescence (CHISHOLMet al., 1988; LI and Wooo, 1988; NEVEUXet al., 1989). Cell concentration was almost always declining with depth below the depth where we could detect all of the cells in a population, so our early results revealed relatively little about the preferred habitat of these cells. Based on this limited information of distributions, our initial impression of the prochlorophytes was one of ubiquitous populations primarily located at the bottom of the euphotic zone. The results reported herein were made with improved instrument sensitivity which allowed us to measure the distribution of prochlorophytes throughout the water column. In transects from Woods Hole to the Sargasso Sea and a series of profiles from a station off Bermuda taken over a seasonal cycle, we find that prochlorophyte distributions are similar to those Synechococcus populations during part of the year, but that the two kinds of picoplankton respond differently to summer stratification of the water column. The prochlorophytes apparently bloom later and can thrive deeper in the water column than Synechococcus. METHODS

Sampling Seasonal sampling was carried out at the 'OFP' site southeast of Bermuda (31°50'N, 64°10'W) (Fig. 1). Spatial coverage was provided by two cruises from Woods Hole to the Sargasso Sea: R.V. Endeavor cruise 177 (21 May-2 June 1988), and R.V. Oceanus cruise 206 (6-18 May 1989). Samples were taken with either Niskin or Go Flo bottles (General Oceanics) or with a plastic bucket when only surface samples were required. Temperature profiles were obtained with CTD or XBT. Nitrate, nitrite and Chl a were measured according to STRICKLANDand PARSONS(1972).

Flow cytometry Flow cytometry data from 0.2 ml samples were collected in list mode. The integrated

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Fig. I. Location of stations in the North Atlantic at which flow cytometric measurements of prochlorophytes were carried out. Filled symbols indicate high-sensitivity measurements were performed; open symbols indicate normal sensitivity. (Squares) September 1986; (circles) May 1988; (triangles) May 1989. Arrow indicates location of seasonal sampling site (OFP) off Bermuda (except for May 1988 sample, which was at 31°50'N, 66°10'W). Surface samples were obtained at 510 km intervals during the May 1989 cruise in the region of the five Gulf Stream stations.

forward scatter, right-angle scatter, orange fluorescence and red fluorescence signals from each cell in a water sample were measured as described in OLSO~ et al. (1990), transferred to a personal computer and analysed using software provided by D. Vaulot (CYTOPC). Analysis involved recognition of populations of various kinds of phytoplankton, and statistical analysis and normalization of these populations to standard fluorescent beads (0.57/zm diameter "Fluoresbrite", Polysciences, Inc., Warrington, PA). We divided the phytoplankton into three groups on the basis of their flow cytometric signatures: prochlorophytes, Synechococcus, and larger eukaryotic phytoplankton (Fig. 2). Synechococcus cells are easily recognized by the orange fluorescence of their phycoerythrin, while prochlorophyes have smaller scatter signals than Synechococcus and have only red fluorescence. The larger eukaryotic phytoplankton also have only red fluorescence but

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have larger scatter signals than Synechococcus, so that they are easily distinguished from prochlorophytes; there is very little overlap in these characteristics between the two cell types. Samples were kept in darkness at room temperature until analysed on board ship (generally within 2 h). Samples to be analysed in Woods Hole were kept in darkness on ice during transit, and then refrigerated at 4°C until analysed, generally 3-4 days after collection. Some samples were preserved by fixation in 1% glutaraldehyde and storage in liquid nitrogen (VAULOTet al., 1989)• Comparison of samples analysed immediately and again after storage demonstrated that, for prochlorophytes and Synechococcus, both these preservation methods were satisfactory for all the parameters of interest to us (Fig. 3). The difference in cell concentration of eukaryotic phytoplankton between samples analysed immediately and those refrigerated (data not shown) was also small (mean = 2%, S.D. = 18%, n = 12). An EPICS V (Coulter) flow cytometer was used for all measurements, except for the Oceanus 206 cruise, where a FACScan (Becton Dickinson) was also used. The FACScan was found to be adequate to detect prochlorophytes under most circumstances, except when the cells were especially dim (see Results). To examine the prochlorophytes in these waters, we used the EPICS to re-analyse replicate samples stored in liquid nitrogen. The configuration of the EPICS was modified for higher sensitivity in several ways, depending on the sensitivity required. In toto the modifications resulted in about 30-fold increased sensitivity over the standard configuration and thus allowed us to detect even dim prochiorophytes in oligotrophic surface waters (Fig. 4). The changes for high sensitivity were as follows: (1) The standard beam-shaping optics, which produce a beam spot of about 16 x 160pm, were replaced by a confocal lens system (Coulter EPICS) producing a beam

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spot of 16 × 40/~m. Subsequently, we learned that a single 38 mm spherical lens (Oriel, Inc.), producing a circular beam spot of about 20/~m diameter, gave even higher sensitivity. (2) The laser was run in "all lines" mode rather than at 488 nm and all the lines except 514.5 nm were selected by a 500 nm shortpass dichroic mirror (Omega Optical, NH). This both increases the overall power available for excitation and better matches the absorption

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characteristics of the Chl a and Chl b, the main photosynthetic pigments in the prochlorophytes. Laser power was 2.5 W (of which about half is <500 nm). (3) The standard jet-in-air flow cell was replaced by a "Profile" flow cell (Coulter EPICS), a 250 #m square quartz flow cell that incorporates a mirror and lens system to gather more light (N.A. approximately 1.4 compared to 0.6 for the jet-in-air system). This represents about a 4-fold gain in signal size. (4) The sheath pressure was reduced from 13 to 5 psi, and the outflow from the flow cell restricted. The velocity is thus reduced from the standard rate of 10 m s -t to as low as 1 m s - 1, so that integrated signal size was increased. A subset of these approaches, including beam shaping and sample velocity reduction, were previously used by ROBERTSON and BuTror~ (1989) to optimize an Ortho flow cytometer for examining marine bacteria. There are trade-offs when trying to increase sensitivity by these methods. Increasing excitation intensity does not always produce proportional increases in fluorescence (due to photobleaching), and slowing the flow and decreasing the beam spot size necessitate lower sample analysis rates. In addition, different optical configurations can influence signals from cells and beads in slightly different ways, so that fluorescence values from samples run in different ways may not be strictly comparable. However, we have determined that changes in the ceil:bead ratio are small (on the order of 10-20%) compared to the range of values we will be discussing. The dynamic range of the measurements must also be considered with these high sensitivity modifications. The large majority of eukaryotic phytoplankton cells are usually mixtures of small (2-5 #m) cells such as chlorophytes, which often form discrete populations (see Fig. 2). In samples run at high sensitivity, however, these cells often extended offscale in light scatter and fluorescence. We have therefore included everything larger than prochlorophytes in the "eukaryotic" category; thus we cannot speak strictly of "ultraplankton" (MURPHYand HAU6~N, 1985). Further, although we can count cells with offscale fluorescence, we cannot determine their fluorescence; thus we will not examine the fluorescence data of the eukaryotic phytoplankton here. RESULTS AND DISCUSSION

Seasonal studies off Bermuda The seasonal cycle of hydrography in the region of our sampling location off Bermuda ("OFP", 31°50'N, 64°10'W) has been well documented for many years (DEtJSER, 1986). It is characterized by wintertime deep mixing (to about 200 m) and strong thermal stratifiFig. 4. A depth profile of flow cytometric signatures of prochlorophytes run without (left) and with (right) high-sensitivity modifications to the flow cytometer. Uniform fluorescent beads (0.57/~m, Polysciences) were added to each sample as an internal standard. Synechococcuscells have been gated out. Samples from Sta, S off Bermuda were obtained on 15 September 1989 (by R. Sherriff-Dow) and were analysed in Woods Hole on 19 September 1989. (A) Laser excitation was 1 W at 488 nm, focused by a confocal lens assembly; the flow cell was the standard jet-in-air. Some cells were below the fluorescence detection limit in all samples above 75 m. (B) Laser excitation was approximately 1.2 W of all wavelengths shorter than 500 nm (principally 488 and 457 nm). A spherical lens and "Profile" flow cell was used and sheath pressure lowered from 13 to 5 psi to reduce sample velocity. Note that populations of prochlorophytes are present at all depths, though their numbers decline dramatically near the surface in this profile.

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cation in summer. The typical pattern of temperature structure, nitrate and chlorophyll distributions was observed in our samples between May 1988 and July 1989 (Fig. 5A). In July stratification was very strongly developed, with surface temperatures of nearly 27°C; nitrate was depleted to below detection limits (0.1/~M) down to about 100 m. We did not observe complete de-stratification in February 1989, but the surface temperature was below 20°C and the nitracline was only 20 m deep. The thermal structure suggests recent deep mixing to below 200 m. In spring and summer samples, deep chlorophyll maxima were present and surface waters were very low in chlorophyll; in autumn and winter samples, chlorophyll concentrations were relatively high up to the surface. High-sensitivity flow cytometry of the picophytoplankton in these samples revealed several important features (Fig. 5B, C). First, prochlorophytes were present year round at high concentrations. The integrated water column abundance of prochlorophytes was always equal to or greater than that of Synechococcus. However, the abundance of the two picoplankters exhibited different seasonal patterns (Fig. 6A): Synechococcus abundance was high in winter and low in summer, while prochlorophyte abundance variations were in the opposite direction. The lowest prochlorophyte abundance occurred when Synechococcus abundance was maximal; this was in the February 1989 sample, when the water column was least stratified and the nitracline was relatively shallow. Abundance of the larger eukaryotic phytoplankton followed the pattern of Synechococcus, not the prochlorophytes. The numerical preponderance of the prochlorophytes suggests that in all samples except February they account for the majority of the picophytoplankton biomass off Bermuda, even though we estimate from T E M and Nuclepore filter fractionation experiments that the volume of each prochlorophyte cell is about a third that of Synechococcus (i.e. mean diameters of 0.7 vs 1.0/~m) (CHISHOLMet al., 1988; WATERBURYet al., 1986). Although prochlorophytes were always present, their depth distributions varied dramatically with season (Fig. 5B). In late spring and summer profiles (May and July), prochlorophytes were present in pronounced subsurface maxima with very few cells at the surface, while in winter and early spring (November, February and March) cells were abundant to the surface. The profiles in February in particular resembled distributions from the wintertime Mediterranean (VAULOT,personal communication), where nitrate was present in surface waters and prochlorophytes and Synechococcus were distributed virtually identically. In contrast Synechococcus cells were always relatively uniformly distributed with depth in the upper layer, except in February 1989. These patterns are reflected in the median depths of each population: both Synechococcus and prochlorophyte populations were as shallow as 20 m in the winter, while in the summer the prochlorophytes were located much Fig. 5. Seasonal cycle of hydrographic and flow cytometricfeatures at a station off Bermuda ("OFP", 31°50'N, 64"10'W),except for the May 1988samples, whichwere from a station at about the same latitude but about 200 km to the west (31"50'N, 66"10'W). (A) Temperature, nitrate concentration and chlorophyllconcentration. (B) Cell concentrationsof prochlorophytes(filled squares), Synechococcus(open circles) and larger eukaryotic phytoplankton (multiplied by 10; dotted lines). The depth of the nitraclineis indicatedfor reference. Due to technicalproblemsthe samplesfromJuly 1988were not analysedwith the highestavailablesensitivity,and we wereunable to completelyenumerate the proehlorophytesfrom 50 m (open symbol)to the surface. However, we can see clearly a maximumin cell concentration. (C) Mean fluorescenceof prochlorophytes (CHL) and Synechococcus(PE) populations. Depth of the nitraclineis indicatedfor reference.

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deeper than Synechococcus (Fig. 6B); the prochlorophytes seem to track the depth of the nitracline. We speculate from these relationships that the prochlorophytes have a more stringent requirement for nitrate (or other nutrients that covary with nitrate) than do Synechococcus (also see below in latitudinal study). The median depth of the larger eukaryotic phytoplankton was more closely related to that of the prochlorophytes than to Synechococcus, in contrast to the total abundance patterns. As expected, mean fluorescence per cell increased with depth for both prochiorophytes and Synechococcus (CmsrtOLM et al., 1988; L~ and WOOD, 1988; NEVEUX et al., 1989; VAULOXetal., 1989; OLSON etal., 1985, 1990) and the increase with depth was of about the same magnitude for the two picoplankters (Fig. 5C). A similarity in patterns of cell fluorescence between prochlorophytes and Synechococcus also was noted by VAU LOTet al. (personal communication) for samples from the Mediterranean in winter. In addition to this general pattern, a marked seasonal pattern in fluorescence per cell also was observed (Fig. 6C). In winter the fluorescence intensity (and hence pigment content) of surface cells was at least five-fold higher than in summer. Fluorescence at depth showed the opposite pattern; the most fluorescent cells in winter profiles were less fluorescent than those in summer (Fig. 6D). These patterns could be determined purely by water column mixing dynamics: in stratified summer conditions deep cells photoadapt to low light intensities by making more pigment, and surface cells are isolated in high light and low nutrient conditions (far above the nitracline), leading to low pigment contents. In winter, more rapid mixing to greater depths would tend to average out fluorescence extremes: cells might not spend enough time at any given light level to completely photoadapt to it (LEwm et al., 1984; CULLEN and LEWm, 1988). However, lower insolation and greater availability of nutrients in winter would also contribute to higher pigment contents, and thus might play a role in changes in fluorescence per cell. The simplest explanation for the approach to the surface of all populations in winter is that light or nutrient conditions for growth near the surface were favorable then. The difference in insolation between summer and winter is less than two-fold at this latitude (KIRK, 1983). On the other hand, the shallow winter prochlorophytes (median depth 20 m) would experience nine-fold more light than the summer cells at 80 m (assuming day length in winter is half that in summer, and an attenuation coefficient of 0.05 m - t, as measured in May 1989). This suggests that increased nutrient availability in surface waters and not reduction in light alone is responsible for the growth of cells near the surface in winter.

"Spring bloom" of prochlorophytes near Bermuda Between 6 and 26 March 1989 we sampled four times at a station located midway between Bermuda and Sta. S, on the 2000 m isopleth ("S/2"; 32°18'N, 64°35'W). The Fig. 6. Summaryof seasonal cycle data from Fig. 4. Since the samples from July 1988were not analysed with the highest sensitivity,and we were unable to completelyenumerate the prochlorophytes from 50 m to the surface, these data are indicated by open squares. (A) Integrated water column abundance of prochlorophytes,Synechococcus,and eukaryotes, and surface temperature. (B) Median depth of prochlorophytes,Synechococcusand eukaryotes, and depth otthe nitracline. (C) Mean fluorescence of surface prochlorophytes (CHL) and Synechococcus (PE), relative to standard fluorescentbeads. The July 1988prochlorophytedata point (open square) is the detection limit of the instrument on this date. (D) Mean fluorescenceof the brightest prochlorophytesand Synechococcus in a given water column.

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timing of these samples coincided with the onset of water column stratification (Fig. 7A) and of nitrate depletion in the surface waters; measurable nitrate was present at the surface at the nearby Sta. S the week before this sampling period (data not shown). The chlorophyll in surface waters decreased dramatically over this period and a subsurface chlorophyll maximum developed. We observed striking changes in the picoplankton distributions during this interval (Fig. 7B), which are qualitatively similar to those in the Bermuda seasonal samples, but compressed in time. Prochlorophytes were present first at the surface and then in deep maxima, and became more abundant than Synechococcus as stratification/nutrient depletion progressed. Over the course of the four samplings, Synechococcus distributions remained nearly constant, with 2.5-4.0 x 105 cells ml- i in the surface waters and declining 6 Mar 89

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Picoplankton in the North Atlantic Ocean

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with depth. In contrast, prochiorophyte distributions showed great variability. In the first profile, prochlorophytes were low in number and uniformly distributed, and by the third they were far more abundant, with a large subsurface maximum. In the last profile this maximum, while still present, was shallower and of smaller magnitude. Mean fluorescence of prochlorophyte and Synechococcus populations followed similar patterns (Fig, 7C), as in the seasonal time series. Surface cells decreased in fluorescence after the first sampling, and the maximum fluorescence of deep cells was higher later in the month. The largest observed increase in prochlorophyte cell numbers in the above time series (about 10-fold between 16 and 22 March) can be accounted for by an average doubling time of a little less than 2 days. We have measured growth rates of this magnitude in laboratory cultures (unpublished data; B. PALENIK,personal communication), thus these patterns could have resulted from growth in situ. However, the changes we observed also could be the result of sampling different water masses with different histories. We have no data on either circulation or phytoplankton patchiness during this period, though on another occasion (two dates in May 1989) we observed picoplankton distributions to be essentially identical at Stas "S/2", S and OFP (Fig. 8). These observations are evidence of an extremely dynamic situation in either a spatial or a temporal regime, and suggest that the prochlorophytes respond at later stages than do Synechococcus during the onset of seasonal stratification and/or surface water nutrient depletion.

Horizontal distribution of prochlorophyte populations We have examined Synechococcus distributions and properties on several cruises in the Atlantic and Pacific Oceans (OLSON et al., 1988, 1990), and prochlorophyte populations during three cruises in the North Atlantic (Fig. 1). On the first, from Woods Hole to Dakar, Senegal in September 1986, we were not using high-sensitivity flow cytometry, and thus we could detect prochlorophytes only deep in the euphotic zone. However, it is 11-13 May 89

l-Z 2 0 0 '0

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Cells ml-i (thousands) Fig. 8. Depth profilesof prochlorophytc and Synechococcu~cellconccntrationsat thrcc stations off Bermuda, bctwccn II and 13 M a y 1989: Sta. O F P (31°50'N, 64°10'W); Sta. S (32010'N, 6 4 ° 3 0 ' W ) ; Sta. " S / 2 " ( 3 2 ° 1 8 ' N , 64°34'W).

1046

R . J . OLSON et al.

important to note that we observed these deep prochlorophytes at every station occupied, including two stations north of the Gulf Stream. Surface water temperatures were above 17°C at all stations. The second transect, in which we began using high sensitivity flow cytometry, was carried out in May 1988, and the third in May 1989. On neither of these spring cruises did we detect prochlorophytes north of the Gulf Stream (where surface temperatures were well below 17°C) as we had in the autumn, suggesting that growth limitation by low temperature can influence the distribution of prochlorophytes. Two major points emerged from these two cruises. First, the abundance and depth distributions of prochlorophytes in the Sargasso in May (Fig, 9) varied with latitude in a manner reminiscent of the seasonal variation off Bermuda, and in particular of the "spring bloom' there (Fig. 7). Part of the transect variations may thus reflect later onsets of stratification going north. In the more weakly stratified northern Sargasso (38°N) prochlorophytes were less abundant than Synechococcus, and were located in near-surface waters. Farther south (36°-37°N), prochlorophytes were more abundant than Synechococcus, but were still located in surface waters. South of about 35°N, in strongly stratified waters with a deep nitracline, the prochiorophytes formed subsurface maxima. These patterns are reflected in the differences between integrated and surface abundances (Fig. 10A, B). Synechococcus were relatively uniformly distributed in the surface waters, and decreased in abundance to the south (Figs 9B and 10A). These patterns are similar to those reported by ITU~I~IAGAand MA~RA(1988) for this region in April 1985. The abundance of the larger eukaryotic phytoplankton did not show any consistent pattern with latitude (data not shown), possibly reflecting changing community structure in this heterogeneous group of organisms (MtmPnv and HAUGEN, 1985; GLOVERet al., 1988). As we went south, the depth of the nitracline increased, as did the median depth of the prochlorophytes (but not Synechococcus) (Fig. 10C). There was a significant correlation between the nitracline and median cell depth for the prochlorophytes (Fig. 11) but not for Synechococcus or the eukaryotic phytoplankton (data not shown): if Z is median depth and N is nitracline depth (both in m), Zpro = 0.84N - 24

(r e = 0.66, P < 0.05, n = 17)

Zsy. = 0.26N + 14

(r 2 = 0.27, P > 0.05, n = 17)

ZEuk = 0.44N + 21

(r e = 0.18, P > 0.05, n = 17).

We note that if the time-series data from near Bermuda are added to this analysis, the correlation between nitracline and median prochlorophyte depth is still significant. Fluorescence intensity of surface prochlorophytes also decreased in the southern Sargasso Sea (Fig. 10D), as it did in the summer samples off Bermuda (Fig. 6C); at the five southernmost stations we were unable to detect all the prochlorophytes near the surface with the FACScan (see Methods). The second major finding of the May 1989 transect was that at all stations south of about 37°N, as off Bermuda, overall abundance of the prochiorophytes was several-fold higher than that of Synechococcus (Fig. 10A). However, the highest abundances were found in the Gulf Stream. In fact, the highest concentrations of prochlorophytes we have observed (2 x 105 m1-1) were in surface waters in the core of the Gulf Stream on this cruise. Within the core of the Stream, the abundance of surface prochlorophytes increased to the north

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Fig. 10. Summary of data from high-sensitivity measurements from Oceanus cruise 206 in May 1989 (open symbols) and Endeavor cruise 177 (closed symbols) as functions of latitude; prochlorophytes (squares); Synechococcus (circles). Surface temperature from May 1989 transect is indicated by solid line; note increased temperature in Gulf Stream. (A) Total water column abundance, and surface water temperature (solid line). (B) Surface cell concentrations and surface water temperature. (C) Median cell depths, and depth of nitracline (solid line). Dashed lines are "regression" lines for reference. (D) Mean fluorescence of prochlorophytes and Synechococcus, relative to standard beads.

Picoplankton in the North Atlantic Ocean

1049

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and Synechococcus remained constant or declined slightly (Fig. 10B). We do not know if the high concentrations of prochlorophytes we observed in the Gulf Stream were imported from other even more prolific regions, or if conditions were especially well suited for their growth there. Prochlorophyte concentrations fell to very low levels (<103 ml-i) within a few km north of the Gulf Stream, where the temperature dropped from 24 to 14°C. At the stations where the prochlorophytes were disappearing, Synechococcus were present at their highest concentrations, 2 x 105 ml -m. In fact, Synechococcus surface concentrations were lower in the Gulf Stream than to either side of it. We speculate that this pattern may reflect differing temperature optima between the two kinds of picoplankters, since the Gulf Stream was significantly warmer than the neighboring waters, or higher nutrient supply in the Gulf Stream, or both. CONCLUSIONS

A seasonal cycle of observations of prochlorophyte distributions using high-sensitivity flow cytometry has broadened our perception of the distribution of these cells. Our initial impression, that prochlorophytes were adapted for growth near the bottom of the euphotic zone, has been substantiated for summer samples. At other times of year the prochiorophytes were abundant higher in the water column, and we found that most of the cells in the population were usually shallower than the nitracline. The regulation of prochlorophyte abundance is clearly different from that of the other major component of the picophytoplankton, Synechococcus. Off Bermuda, the prochiorophytes bloom later in the spring than Synechococcus but reach higher abundance, and vary their depth distribution more, following the deepening of the nitracline in summer. If this pattern holds true, we would expect that in the southern Sargasso, where wintertime deep mixing is not a regular feature, prochlorophytes will not exhibit surface blooms but will always be found near the nitracline. The situation in more temperate waters north of the Gulf Stream is less clear since we have few observations of prochiorophyte distributions there; it appears that prochlorophytes may be restricted to summer growth in these waters and are probably less important than Synechococcus.

1050

R.J. OLSONet al.

Differences in photosynthetic pigment composition between the picoplankton groups may account for the typical s u m m e r t i m e features of the observed depth distributions. The prochlorophytes' major pigments, divinyl-like chlorophylls a (CmsHOL~ etal., 1988) and b (GoER1CKE, 1990), absorb efficiently at the wavelengths available in the blue lightdominated deep euphotic zone of the open ocean, as do the pigments of the eukaryotic ultraplankton (GLOvER et al., 1986). The phycoerythrin of Synechococcus, even of openocean high-urobilin strains, absorbs blue light much less efficiently (WATERBURYet al., 1986; CAMPBELLand ITURRIAGA,1988; OLSON et al., 1988, 1990). Thus the prochlorophytes are able to grow deep in the euphotic zone, where nutrients are m o r e available, but where Synechococcus would be severely light limited. The paucity of prochiorophytes in surface waters in s u m m e r suggests that these cells may actually be limited to regions of lower light intensity. Aiternatively, they may be less effective than Synechococcus at utilizing low concentrations of nutrients available in near-surface waters in summer. The changes in relative distributions of prochlorophytes and Synechococcus with latitude in the northern Sargasso in spring and with time off B e r m u d a during winter and early spring suggest that both kinds of cells are nutrient-replete and light-limited in the winter (so that growth is fastest near the surface), and that Synechococcus is able to grow faster (or to begin growth earlier in the year) than the prochlorophytes. In addition, differential grazing on the two kinds of picoplankters, which have not been investigated, could play a role in the patterns we observed.

Acknowledgements--We thank Rachel Sherrif-Dow for providing samples; Raffaella Cassotti, Sheila Frankel and Ellen Connors for technical support; John Waterbury for valuable discussions; and the captains and crews of the R.V.'s Gyre, Endeavor, Oceanus and Weatherbird. This work was supported in part by NSF grants OCE8416964, OCE8614332 (to R.J.O.), OCE8316616, OCE8421041, OCE8508032, OCE8614488 (to S.W.C.), OCE8717508 (to M.A.A.), ONR grants N00014-83-K0661, 84-C-0278 and 87-K-0007 (to R.J.O. and S.W.C.), and an NSF Graduate Fellowship (to J.A.D.). This is Woods Hole Oceanographic Institution contribution no. 7330 and Bermuda Biological Station contribution no. 1241. REFERENCES BAZZAZM. B. (1981) New chlorophyll chromophores isolated from a chlorophyll-deficient mutant of Maize. Photobiology and Photobiophysics, 2, 199-207. CAMPaELLL. and R. I~Um^~A(1988) Identification ofSynechococcus spp. in the Sargasso Sea by immunofluorescence and fluorescence excitation spectra performed on individual cells. Limnology and Oceanography, 33, 1196-1201. CHISHOLMS. W., R. J. OLSON, E. R. ZE'I['rLER,J. WATERRURV,R. GOER1CKEand N. WELSCHMEYER(1988) A novel frec-living prochlorophyte abundant in the oceanic euphotic zone. Nature, 334, 340-343. CULLENJ. J. and M. R. LEWlS(1988) The kinetics of algal photoadaptation in the context of vertical mixing. Journal of Plankton Research, 10, 1039--1963. DEOSEa W. G. (1986) Seasonal and intcrannual variations in deep-water particle fluxes in the Sargasso Sea and their relation to surface hydrography. Deep-Sea Research, 33, 225-246. Gl~zms W. W. C. and G. W. KRAAV(1983) Unknown chlorophyll a derivatives in the North Sea and tropical Atlantic Ocean revealed by HPLC analysis. Limnoiogy and Oceanography, 38, 757-766. GmsK~ W. W. C., G. W. KRAv,A. NONT#I,D. SImAPERMANAand SUTOMO(1988) Monsoonal alternation of a mixed and a layered structure in the phytoplankton of the euphotic zone of the Banda Sea (Indonesia): A mathematical analysis of algal pigment fingerprints. Netherlands Journal of Sea Research, 22, 123-137. GLOVERH. E., M. D. KELLERand R. R. L. GUILt.ARD(1986) Light quality and oceanic ultraphytoplankters. Nature, 319, 142-143.

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GLOVER H. E., B. R. PREZELIN,L. CAMPBELLand M. WYMAN(1988) Pico- and ultraplankton Sargasso Sea communities: variability and comparative distributions of Synechococcus spp. and algae. Marine Ecology Progress Series, 49,127-139. GOERtCKE R. (1990) Pigments as ecological tracers for the abundance and growth of marine phytoplankton. Ph.D. thesis, Harvard University, Cambridge, MA, 418 pp. ITURRIAGAR. and J. MARV.A(1988) Temporal and spatial variability of chroococcoid cyanobacteria Synechococcus spp. specific growth rates and their contribution to primary production in the Sargasso Sea. Marine Ecology Progress Series, 44, 175-181. KIRKJ. (1983) Light and photosynthesis in aquatic ecosystems. Cambridge University Press, 402 pp. LEWINR. A. (1981) The prochlorophytes. In: Theprokaryotes, Vol. 1, M. P. STAXR,H. STOLP.H. G. BALOWSand H. G. SCnLE6EL, editors, Springer, Berlin, pp. 257-266. LEwis M. R., J. J. CULL~Nand T. R. PLArr (1984) Relationships between vertical mixing and photoadaptation of phytoplankton: Similarity criteria. Marine Ecology Progress Series, 15, 141-149. LI W. K. W. and M. WOOD(1988) Vertical distribution of North Atlantic ultraphytoplankton: Analysis by flow cytometry and epifluorescenee miscroscopy. Deep-Sea Research, 35, 1615-1638. MuRPhy L. S. and E. M. HAUGEN(1985) The distribution and abundance of phototrophic ultraplankton in the North Atlantic. Limnology and Oceanography, 30, 47-58. NEveux J. D., D. VAULOX,C. C o u ~ and E. FtJv~I (1989) Green photosynthetic bacteria associated with the deep chlorophyll maximum of the Sargasso Sea. Comptes Rendus de l'Aeadamie des Sciences, 308, Serie III, %14. OLSON R. J., D. VAULOTand S. W. CmSHOLM (1985) Marine phytoplankton measured using shipboard flow cytometry. Deep-Sea Research, 10, 1273-1280. OLSON R. J., S. W. CmSnOLM, E. R. ZE't'rLERand E. V. ARMBRUST(1988) Analysis of Synechococcus pigment types in the sea using single and dual beam flow eytometry. Deep-Sea Research, 35,425--440. OLSON R. J., S. W. CmSnOLM, E. R. ZErrL~R and E. V. Ava~Bausv (1990) Pigments, size and distribution of Synechococcus in the North Atlantic and Pacific Oceans. Limnology and Oceanography, 35, 45-58. ROaERrSONB. R. and D. K. Btrrro~ (1989) Characterizing aquatic bacteria according to population, cell size and apparent DNA content by flow cytometry. Cytometry, 10, 70-76. S~ICKL^ND J. R. R. and T. R. PARSONS(1972) A practical handbook of seawater analysis. Fisheries Research Board of Canada, Ottawa, 310 pp. VAULOTD., C. CouRaa~s and F. P^RTENSXV(1989) A simple method to preserve oceanic phytoplankton for flow cytometric analysis. Cytometry, 10, 629--635. VERN~T M. (1983) Phytoplankton pigments: methods and ecology. Ph.D. dissertation, University of Washington, Seattle, 159 pp. WATEnBURYJ. B., F. W. VALOm and D. G. Fvo,NXS (1986) Biological and ecological characterization of the marine unicellular cyanobacterium Synechococcus. In: Photosynthetic picoplankton, T. PLArr and W. K. W. Lx, editors, Canadian Bulletin of Fisheries and Aquatic Science, 214, 71-120.