ORIGINAL PAPER

Spatial-temporal variability of ultraplankton vertical distributionin the Antarctic frontal zones within 60–66�E, 43–46�S

Received: 18 June 2003 / Accepted: 4 August 2003 / Published online: 9 September 2003� Springer-Verlag 2003

Abstract The vertical distribution of heterotrophicbacteria and four ultraphytoplanktonic (<10 lm)groups (Prochlorococcus, Synechococcus, pico- and na-noeukaryotes) was investigated by flow cytometry atthree process stations located in three different sub-sys-tems belonging to the Antarctic Circumpolar Currentfrontal zone and to the Southern Indian Ocean (60–66�E, 43–46�S; ANTARES 4 cruise, January-February1999): the Subtropical Zone (STZ), the ConvergenceZone and the Polar Frontal Zone (PFZ). In each sub-system, short-term variability of cell abundance and flowcytometric parameters (right-angle light scatter andchlorophyll autofluorescence) was assessed through atimes series of up to 24 h with a 2 h sampling frequency.The ultraphytoplankton vertical distribution exhibited ahigh spatial variability, with dominance of Prochloro-coccus in the STZ (mean: 762.85·1010 cells m)2),whereas picoeukaryotes (<3 lm) were dominant in thePFZ (55.46·1010 cells m)2), a typically high-nutrientlow-chlorophyll zone. Heterotrophic bacteria abun-dance was maximum (9.84·1013 cells m)2) in the frontalzone, between the Agulhas Front and the SubtropicalFront. Nanoeukaryotes showed the largest (up to 80%)variations between two consecutive sampling periods(2 h). Abundance variations could not be assigned to thesame water mass during the time series due to the highlyvariable hydrodynamics of the study area. Trends ofshort-term abundance variations were opposite betweenthe PFZ (lowest at night) and north of the Subtropical

Front (highest at night). The observed spatial and short-term variations illustrate the complexity of the watermasses in the Indian sector of the Southern Ocean, andhighlight the challenge of extrapolating discrete mea-surements over space and time for use in evaluatingcarbon budgets in such dynamic areas.

Introduction

The region delimited by the Crozet and KerguelenIslands on its western and eastern boundaries, respec-tively and between 42 and 47�S, encompasses a widerange of hydrographic regimes, including open-oceanwaters and dynamic fronts linked to the Antarctic Cir-cumpolar Current (Park 1989; Park et al. 1993, 2002;Park and Gamberoni 1997). The situation in this regionthat is hereafter referred to as the Crozet Basin, is par-ticularly complex due to the close juxtaposition of theAgulhas Front (AF), the Subtropical Front (STF) andthe Subantarctic Front (SAF) (Fig. 1). The Crozet Basinis one of the largest oceanic sinks for atmospheric CO2,due in part to biological uptake and export by phyto-plankton (Metzl et al. 1999). Like other oceanic frontalregions, it exhibits high chlorophyll concentrationsduring summer (Weeks and Shillington 1996), althoughthe Southern Ocean is considered to be an HNLC (high-nutrient low-chlorophyll) zone. This hydrographiccomplexity, coupled to extreme seasonality, results indramatic spatial and temporal variability in planktoncommunity composition and distribution.

For many years, large-size diatoms were consideredto form the basis of the Antarctic food-web structure inthe pelagic ecosystem (Guillard and Kilham 1977).However, this classical concept changed with subsequentinvestigations showing the importance of nano- andpicoplanktonic organisms in different Antarctic areas(Brockel 1981; Hewes et al. 1985; Weber and El-Sayed1987; Jacques and Panouse 1991; Detmer and Bathmann1997). In the past 30 years, it has been established that

Polar Biol (2003) 26: 734–745DOI 10.1007/s00300-003-0545-5

Celine Dubreuil Æ Michel Denis Æ Pascal ConanSylvie Roy

C. Dubreuil Æ M. Denis (&) Æ P. Conan Æ S. RoyLaboratoire d’Oceanographie et de Biogeochimie,UMR CNRS 6535, Centre d’Oceanologie de Marseille,Universite de la Mediterranee, 163 avenue de Luminy,Case 90113288 Marseille cedex 09 Marseille, FranceE-mail: denis@com.univ-mrs.frTel.: +33-4-91829114Fax: +33-4-91826548

Present address: P. ConanLaboratoire Arago, Universite Pierre et Marie Curie,UMR CNRS 7621, 66651 Banyuls-sur-Mer, France

algal taxa belonging to the ultraphytoplankton(<10 lm) may predominate in different areas of theSouthern Ocean, including small diatoms (5–10 lm),prymnesiophytes and other flagellates, and cyanobacte-ria (e.g. Prochlorococcus, Synechococcus sp.) (Jacquesand Panouse 1991). The size distribution of primaryproducers plays an important role in the trophic orga-nization of marine ecosystems and in the global flux oforganic matter. The size-structure distribution of phy-toplankton in the Indian sector of the Southern Oceanhas been essentially documented within the frame of theFrench ANTARES program (Fiala et al. 1998, 2002,2003; Kopczynska et al. 1998; Fouilland et al. 1999). Thebacterial distribution has received little attention (LokaBharathi et al. 2001; Delille 2003; Fiala et al. 2003). Inaddition, to our knowledge, their short-term variabilityremains unknown.

Determining the spatial-temporal variability inplankton distribution is critical for the interpretation ofdiscrete sampling and extrapolations over space andtime, and for the evaluation of carbon budgets. In thefrontal zone investigated here, spatial variations areessentially driven by the hydrodynamics. In addition, thedistribution of ultraplankton (<10 lm) can further beaffected by periodic and sporadic environmental changesthat may also induce variations in cell physiology. At thetime scale of the hour or day, the major periodic changesare the light-dark cycle and diel vertical migration, andthe most common sporadic perturbation is generated bywind stress. Nycthemeral periodicity was also reportedfor chlorophyll fluorescence (Dandonneau and Neveux1997; Vaulot and Marie 1999; Dusenberry et al. 2000;Binder and DuRand 2002) and phytoplanktonic celllight scatter (Olson et al. 1990b; DuRand and Olson1996; Vaulot and Marie 1999; Dusenberry et al. 2000;Binder and DuRand 2002).

To improve the knowledge of the spatial-temporalvariability of the ultraplankton distribution in the In-dian sector of the Southern Ocean, we investigated theultraphytoplankton and heterotrophic bacteria verticaldistributions at three process stations located in three

different sub-systems of the Crozet Basin: the Subtrop-ical Zone (STZ), the Convergence of the AF/STF andthe Polar Frontal Zone (PFZ) during the 1999 australsummer (January/February). In each sub-system, westudied the short-term variability of these vertical dis-tributions with a 2-h sampling frequency over a periodup to 24 h.

Materials and methods

Study area and sample collection

In January/February 1999, the ANTARES 4 cruise, a contributionto the SO-JGOFS program, focused on the frontal region of theCrozet Basin, between 60–66�E and 43–46�S, on board the Frenchvessel ‘‘Marion-Dufresne’’ (Fig. 1). Three process stations (Sta-3, 7and 8) were occupied in three different hydrological areas. Sta-8was located in the Subtropical Zone, north of the Agulhas Front, inwarm (>15�C) and saline (>35) subtropical waters (Fig. 1). Sur-face nitrate concentrations were low (<0.09 lM), characteristic ofan oligotrophic regime. Sta-7 was situated in the Convergencezone, tightly sandwiched between the AF and Subtropical Front,and was characterized by waters slightly cooler and less saline thansubtropical waters. Sta-3 was situated in the northernmost PolarFrontal Zone, south of the Subantarctic Front and corresponded toa typical high-nutrient low-chlorophyll (HNLC) zone, with nitratepresent in large amounts (>15 lM). Nitrate concentration in-creased southwards with sharp changes across the Subtropical andSubantarctic Fronts. The SAF marked the northern limit of cold(<10�C) and less saline (<34) polar waters.

A full description of physical and hydrological features andproperties of the investigated area can be found in the literature(Park 1989; Park et al. 1993, 2002; Park and Gamberoni 1997). Thehydrodynamic characteristics of the study area during theANTARES 4 cruise are reported in Park et al. (2002). Other fea-tures related to nutrient fluxes, and limitations are described byLeblanc et al. (2002) and Sedwick et al. (2002).

Sampling at the process stations was conducted in a Lagrangianmode by means of a drifting buoy. Time series of 24 h with afrequency of approximately 2 h were run at each process station,except at station 3, where they were interrupted by bad weatherconditions. Physical parameters were measured using a CTDprobe (Seabird 911+) mounted on a rosette of 12 Niskin bottles(12 l) for collection of seawater samples at 10 depths between 0 and200 m. The time series were made from 13, 10 and 7 vertical profilesat stations 8, 7 and 3, respectively. Hours are given in universaltime.

Fig. 1 Location of the time-series stations (++) in the Indiansector of the Southern Oceanduring the ANTARES 4 cruise.The oriented grey bands are aschematic representation of thesurface circulation (see Parket al. 2002) (AF Agulhas Front,STF Subtropical Front, SAFSubantarctic Front). Station 3(46�00¢S, 62�49¢E) was occupiedfrom 19/1/1999 to 20/1/1999.Station 7 (44�00¢S, 64�10¢E) wasoccupied from 8/2/1999 to 9/2/1999. Station 8 (43�00¢S,62�56¢E) was occupied from 13/2/1999 to 14/2/1999

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Flow cytometry analysis

Prefiltered (100 lm mesh size net) seawater samples were preservedwith 2% paraformaldehyde (Trousselier et al. 1995), frozen onboard and stored in liquid nitrogen. Back in the laboratory, sam-ples were thawed in a 30�C water bath and analysed with a flowcytometer (Cytoron Absolute, Ortho Diagnostic Systems) equippedwith an air-cooled 488 nm argon laser. The sample and sheath flowrates were 1 and 100 mm3 s)1, respectively. Fluorescent beads of 1or 10 lm in diameter were added to the sample in order to nor-malize the flow cytometer settings and provide a reference forconcentration calculations. Two different protocols were used, themaximum resolution capacity being set for discriminating Pro-chlorococcus, Synechococcus and picoeukaryotes. The mean anal-ysis volume was 350 mm3 for cyanobacteria analysis, and 700 mm3

for pico- and nanoeukaryote analysis. Heterotrophic bacteria werecounted after staining with Sybr Green II. Samples of 1 cm3 sea-water were supplemented with 10 mm3 Sybr Green II (from theMolecular Probes solution diluted 10 times) and incubated for10 min in the dark before analysis. The mean analysis volume was350 mm3. Counting reproducibility was better than 3.7% (SD,n=23) over a range of concentrations extending 1 order of mag-nitude. The spatial variability of cell abundance derived fromNiskin bottles closed at the same depth was ±10%.

Flow cytometry data were acquired and stored in list-mode withImmunocount software (Ortho Diagnostic Systems) and clusteranalysis was run with Winlist software (Verity Software House).The flow cytometric analysis was restricted to <10 lm phyto-planktonic cells. Four populations were discriminated on the basisof right angle light scatter (RALS) and fluorescence signals: Pro-chlorococcus characterized by low RALS values, low red fluores-cence, and no orange fluorescence; Synechococcus characterized byorange fluorescence due to the presence of phycoerythrin, and redfluorescence; picoeukaryotes (<3 lm) had larger RALS values andred fluorescence than cyanobacteria but no orange fluorescence,and nanoeukaryotes corresponded to the largest analysed cells (3–10 lm). Mean scatter and fluorescence signals were normalized byreference to the corresponding mean signal from the 1-lm fluoro-spheres.

We only focus here on the variability of cell parameters ofcyanobacteria (Prochlorococcus and Synechococcus) and notpico- and nanoeukaryotes. Flow cytometry provides a promptdetermination of RALS, used as a proxy of the cell size, and the cellautofluorescence (red and orange for Synechococcus), an indicatorof pigment content (chlorophyll and phycoerythrin, respectively forSynechococcus). It is worth noting that RALS can be affected bymany factors and, in particular, by sample preservation (Vaulotet al. 1989; Vaulot and Partensky 1991; Landry and Kirshtein 1993;Partensky et al. 1996), as in the present study. Indeed, preservationof cells induces shrinkage (Verity et al. 1992) and, consequently,affects cell scattering.

Results

Station 8 (Subtropical Zone)

Cell abundance

Sta-8, located in the oligotrophic subtropical zone(STZ), was characterized by warm and saline surfacewaters, as illustrated by the time course of temperatureand salinity vertical distributions in Fig. 2. At the timeof sampling, a thermocline was established at about75 m depth. Surface temperature was higher during thefirst night sampling than the following day.

Nanoeukaryotes (3–10 lm) reached a maximumabundance of 7·103 cells cm)3 (Fig. 3A); Synechococcusconcentrations ranged from 25 to 60·103 cells cm)3

(Fig. 3B), picoeukaryotes (<3 lm) from 2 to 8·103 cellscm)3 (Fig. 3C) and Prochlorococcus from 90 to 160·103cells cm)3 (Fig. 3D). Heterotrophic bacteria abundance(Fig. 3E) varied from 5 to 6.5·105 cells cm)3 above thethermocline. In the STZ, cyanobacteria were the mostabundant autotrophic organisms, 10–20 times more thanpico-and nanoeukaryotes, both present in similar con-centrations. Abundances of all cell groups exhibitedtheir highest values at night and their minimum in thelate morning (0900 to 1300 hours). There was a smalldrop at dusk for all populations except for heterotrophicbacteria. The time course of abundance changes variedwith depth. Ultraphytoplankton mainly occupied theupper 60 m during the first-night period, and its pres-ence extended to 100 m during the next night due to thedeepening of the mixed layer from about 70 to 90 m(derived from salinity and temperature profiles, Fig. 2).This change in the mixed-layer depth was mainly drivenby a salinity decrease (Fig. 2). At the surface (0–5 m),the abundance of ultraphytoplanktonic populationsexperienced a sharp decrease at dawn, and remained lowduring the daytime, whereas between 20 and 40 m,strong variations in cell concentrations were observed.During the night, the ultraphytoplankton abundancewas maximum and homogeneous within the wholemixed layer. Night cell concentrations were about 150,

Fig. 2 Time course of the vertical distribution of temperature (�C)and salinity at Sta-8 (STZ) during the time-series experiment

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Fig. 3A–E Time course of the vertical distribution of A nanoeuk-aryote, B Synechococcus, C picoeukaryote, D Prochlorococcusabundance (103 cells cm)3), and E heterotrophic bacteria (105 cellscm)3) at Sta-8 (STZ) during the time-series experiment

Table 1 Ranges and means ofphysical parameters(temperature and salinity) andintegrated (over the upper150 m) cell abundance(cyanobacteria, pico- andnanoeukaryotes, andheterotrophic bacteria) in theSTZ, the Convergence AF/STFand in the PFZ during cruiseAntares 4 (n1=number ofsamples, n2=number ofintegrated abundance profiles)

Station 8 Station 7 Station 3STZ Convergence AF/STF PFZ

Hydrologicalcharacteristics

Temperature (�C) Range 14.73–18.10 11.83–14.72 4.66–10.20Mean 16.59 13.53 7.91n1 117 89 63

Salinity Range 35.43–35.54 34.33–35.14 33.68–34.07Mean 35.5 34.71 33.78n1 117 89 63

Integrated cellabundance(1010cells m–2)

Prochlorococcus Range 597.99–931.07Mean 762.85n2 13

Synechococcus Range 218.63–287.74 66.18–140.39 26.99–55.49Mean 252.98 92.65 42.39n2 13 10 7

Picoeukaryotes(<3 lm)

Range 34.68–46.19 15.62–31.42 45.68–67.11

Mean 40.81 23.6 55.46n2 13 10 7

Nanoeukaryotes(3–10 lm)

Range 20.33–55.12 17.45–41.99 15.97–48.57

Mean 37.78 29.21 31.53n2 13 10 7

Heterotrophic bacteria Range 6853.5–9115.1 6248.8–17381.3 6333.8–12205.2Mean 8314.4 9843.6 8223.6n2 11 9 5

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50, 8 and 4·103 cells cm)3 for Prochlorococcus, Syn-echococcus, picoeukaryotes and nanoeukaryotes,respectively, and 6·105 cells cm)3 for heterotrophicbacteria. Below the thermocline (down to 150 m), lowpico- and nanoeukaryote concentrations (<1.5·103 cellscm)3) were observed, and the presence of cyanobacteriawas not detected. Heterotrophic bacteria were presentthroughout the whole water column, with lower abun-dance (1.5–3·105 cells cm)3) than in the mixed layer.

Plankton concentrations integrated over the upper150 m at station 8 are reported in Table 1. Nanoeuk-aryotes exhibited the largest abundance variations, witha 2.7-fold change from 20 to 55·1010 cells m)2. The fullrange of variation from minimum to maximum inte-grated cell concentration represents a 1.5- and 1.3-foldincrease for Prochlorococcus and the other groups,respectively (Table 1). The integrated abundance varia-tions during the 24-h cycle were similar for allautotrophic populations, with maximum cell concen-trations occurring during the night. The relative contri-bution of each population to the whole ultraplanktonabundance was stable during the time series, exceptfor pico- and nanoeukaryotes. Indeed, between 0641and 1248 hours, the mean integrated picoeukaryoteabundance (390·109 cells m)2, n=4, r=47) was almosttwice that of integrated nanoeukaryote abundance

Fig. 4A–D Time course of the vertical distribution of right-anglelight scatter (RALS; A and C) and red fluorescence (B and D) ofcyanobacteria at Sta-8 (A, B, C, D). RALS and fluorescence signalswere normalized with respect to corresponding signals of 1-lmbeads (see Materials and methods)

Fig. 5 Time course of the vertical distribution of temperature (�C)and salinity at Sta-7 (Convergence AF/STF) during the time-seriesexperiment

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(228·109 cells m)2, n=4, r=18), whereas their inte-grated abundances were not significantly different out-side this period.

Patterns of size and fluorescence parameters

At Sta-8, RALS vertical profiles of Prochlorococcus(Fig. 4A) and Synechococcus (Fig. 4C) were similarthroughout the water column. Minimum RALS signalswere observed at the end of the night and remained lowuntil the beginning of the afternoon. From 1300 hours,RALS increased, and reached a sub-surface (20 m)maximum at 1500 hours for both Prochlorococcus andSynechococcus. RALS then remained constant untilsunset and during the first part of the night. Below thethermocline, we observed a sharp increase of the Pro-chlorococcus RALS (below 60 m depth) and, to a lesserextent, of Synechococcus RALS (below 80 m depth).The range of RALS variations was much larger forProchlorococcus than for Synechococcus, and the largestvariations were observed at the beginning of the after-noon (1200–1600 hours).

Figure 4 (B, D) shows the temporal variations at Sta-8 of Prochlorococcus and Synechococcus red fluorescencenormalized to that of 1 lm beads. For Synechococcus,phycoerythrin and chlorophyll fluorescence profiles werevery well correlated. Consequently, only chlorophyll

fluorescence is reported in Fig. 4D. Fluorescence valuesin the upper 30 m were the lowest and remained rela-tively constant during the time series. Variations withtime were more pronounced at depth, with a morningdecrease of Synechococcus fluorescence, followed by anincrease from midday until early night (2300 hours).Over the entire time series, maximum fluorescence valueswere observed below the thermocline where fluorescenceincreased sharply with depth.

Station 7 (Convergence AF/STF)

Cell abundance

Sta-7, located in the frontal zone between the AF andthe STF (Fig. 1), was characterized by slightly coolerand less saline waters than the STZ (Fig. 5). Both thethermocline and halocline were deeper than at Sta-8.The depth of the thermocline fluctuated from 100 to60 m during the 24-h time series, whereas the haloclineremained stable at a depth of 60 m (Fig. 5). In the AF/STF convergence zone, picoeukaryotes exhibited lowermean integrated abundance (23.6·1010 cells m)2) than atSta-8 (40.8·1010 cells m)2, Table 1). Synechococcus wasabout half as abundant as in the subtropical zone(Fig. 6B) and Prochlorococcus was not detected south of44�S. Nanoeukaryote abundance was of the same order(�7·103 cells cm)3) (Fig. 6A) as in the STZ. In contrast,heterotrophic bacteria were more abundant (up to 9·105cells cm)3) than in the STZ (Fig. 6D). Both eukaryoticpopulations occupied a thinner layer at Sta-7 than at

Fig. 6A–D Time course of the vertical distribution of A nanoeuk-aryote, B Synechococcus, C picoeukaryote abundance (103 cellscm)3), and D heterotrophic bacteria (105 cells cm)3) at Sta-7(Convergence AF/STF) during the time-series experiment

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Sta-8. Synechococcus was the dominant autotrophicorganism (up to 24·103 cells cm)3), 3 times more abun-dant than pico- and nanoeukaryotes, whose cell con-centrations were of the same order (up to 7–8·103 cellscm)3). As in the STZ, abundance of all ultraplanktonicpopulations varied with time in the Convergence AF/STF. Autotrophic cell concentrations exhibited a sharpdecrease at dawn (Fig. 6), and a peak in late afternoon(11.8, 28 and 10·103 cells cm)3 for picoeukaryotes,Synechococcus and nanoeukaryotes, respectively). Het-erotrophic bacteria and picoeukaryotes exhibited a peakof abundance in the upper 50 m, between 0948 and1436 hours, which exactly corresponded to the lowestnanoeukaryote and Synechococcus surface abundances.

Integrated cell concentrations (Table 1) were highestduring the night and lowest at dawn, as in the STZ. Allultraplanktonic populations exhibited larger variations intheir integrated abundance than in the STZ (ratio max/min >2). The largest ratio (2.8) was observed for het-erotrophic bacteria. It was 2.4 for nanoeukaryotes, and2.0 and 2.1 for picoeukaryotes and Synechococcus,respectively (Table 1). The relative contribution of auto-trophic populations remained stable during the time ser-ies. Although Synechococcus was threefold less abundantin the Convergence AF/STF than in the warm waters ofthe STZ, it remained the dominant autotrophic organism,pico- and nanoeukaryote integrated abundance beingtwofold lower than in the STZ (Table 1). In contrast,heterotrophic bacteria were more abundant in the Con-vergence AF/STF than in the STZ (Table 1).

Patterns of size and fluorescence parameters

As for Sta-8, the thermocline in the frontal zone (Sta-7)marked the beginning of the Synechococcus RALS in-crease with depth, but RALS profiles were uniform atsunset in the mixed layer (Fig. 7A). RALS maximumvalues were threefold lower than in the STZ (Sta-8). Thelowest RALS values were observed at the surface duringthe whole of the afternoon and early night. The verticalfluorescence profile for Synechococcus exhibited thesame trend as in the subtropical zone (Fig. 7B), with asharp vertical gradient coinciding with the level of thethermocline. The lowest chlorophyll fluorescence valuescoincided with the peak of Synechococcus abundance inthe surface layer.

Station 3 (Polar Frontal Zone)

Cell abundance

Sta-3 was located in the northernmost area of the PFZ.The time series was stopped at about half way (the lastsampling occurred at 1345 hours) due to very rough seaconditions. Temperature and salinity profiles (Fig. 8)suggest that the same water mass was followed duringthe time series above the thermocline (around 70 m) butnot below it, where waters were more saline and warmer

during nighttime than daytime. The picophytoplank-tonic organisms reached their maximum abundance at60–80 m depth in cold polar waters (Fig. 9). The lowestconcentrations of Synechococcus (2–4·103 cells cm)3)were observed at the surface, and the maximum value atdepth was 3 times lower than in the STZ (13·103 cellscm)3) (Fig. 9B). Picoeukaryotes exhibited their maxi-mum abundance in the PFZ, up to 12·103 cells cm)3 (at60 m depth) (Fig. 9C) while nanoeukaryotes were evenlydistributed (4·103 cells cm)3) (Fig. 9A). Abundance ofheterotrophic bacteria was relatively homogeneousabove the thermocline at around 5·105 cells cm)3. Belowthe thermocline, abundance of eukaryotic cells (Fig. 9A,C) exhibited a sharp decline, cyanobacteria were absent(Fig. 9B) and heterotrophic bacteria abundance(Fig. 9D) was lower (2·105 cells cm)3). In contrast withstations 8 and 7, peak abundance was observed duringdaytime for all ultraphytoplanktonic populations. Pic-oeukaryotes and cyanobacteria were present down to80–100 m, and nanoeukaryotes down to 150 m, deeperthan at stations 8 and 7. Picoeukaryotes and Synecho-coccus patterns were similar, with maximum cell con-centrations following the thermocline oscillations(Fig. 8). As with other stations, nanoeukaryotes had thehighest range of abundance variations (Table 1).

In terms of integrated cell concentration (Table 1),picoeukaryotes were the numerically dominant auto-trophic organism in the PFZ. Nanoeukaryote abun-dance was of the same order at Sta-3 and 7, but theamplitude of variation was twofold higher at Sta-3.Mean integrated cell abundance of heterotrophic bac-teria was similar at Sta-3 and 7. Sta-3 was characterizedby the dominance of the smallest eukaryotic cells.

Patterns of size and fluorescence parameters

In the PFZ, at Sta-3, the Synechococcus RALS wasrelatively constant in the upper 60 m throughout thetime course (Fig. 10A). As for Sta-8, a sharp increase ofRALS with depth was observed below the thermocline.This vertical gradient was more pronounced during theday than during the night.

The spatial-temporal pattern of Synechococcus redfluorescence (Fig. 10B) was similar to that in the otherstations for the comparable portion of the time series,between 0500 and 1330 hours. Indeed, the lowest chlo-rophyll fluorescence values were observed in the upper50 m, and the fluorescence increased sharply with depth,following oscillations of the thermocline.

Discussion

Spatial distribution

A strong contrast in ultraphytoplankton compositionand vertical distribution was observed between the threeprocess stations. The oligotrophic waters of the STZ

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were characterized by the dominance of cyanobacteria,with Prochlorococcus exhibiting the highest concentra-tions (up to 150·103 cells cm)3, Fig. 3D) while Syn-echococcus had the maximum integrated abundance(218–287·1010 cells m)2, Fig. 3B). The dominance ofProchlorococcus in the subtropical waters was consistentwith all previous observations (see review in Partensky etal. 1999b), since its discovery by Chisholm et al. (1988).Prochlorococcus abundance was probably underesti-mated near the surface where, under high irradiance, thecell red fluorescence is very low (Partensky et al. 1999a,1999b).

The absence of Prochlorococcus south of the AF is inagreement with previous reports of its disappearance atlatitudes >43�S (Fouilland et al. 1999; Fiala et al. 2003).Temperature is considered as the main controlling factorof its abundance (Buck et al. 1996; Partensky et al.1999a; Cavender-Bares et al. 2001).

The PFZ was characterized by the dominance ofpicoeukaryotes (<3 lm; Fig. 9). Synechococcus inte-grated abundance was 20 times lower in the PFZ coldwaters (<10�C) than in the STZ where temperature was>15�C. This is consistent with observations by Murphyand Haugen (1985) who reported that the decrease in

cyanobacteria abundance correlates with decreasingtemperature in the North Atlantic Ocean. Between theAgulhas Front and the Subtropical Front, all integratedabundance values were lower than in subtropical (Sta-8)and Antarctic (Sta-3) waters. It is worth noting thatnanoeukaryotes exhibited the highest integrated abun-dance in the STZ. Another major difference between theSTZ and the PFZ was the vertical distribution of pic-oeukaryotes and Synechococcus, homogeneous withinthe upper surface layer (0–60 m) in the STZ (Fig. 3), andin the PFZ, concentrated at the thermocline (Fig. 9).This can be explained by very different meteorologicalconditions between these two locations during the studyperiod. In the STZ, the absence of wind stabilized thewater column, favouring the development of cyanobac-teria within the entire surface layer. In contrast, in thePFZ, there was a strong wind (up to 35 knots) gener-ating much turbulence within the upper surface layerand, probably, inducing higher concentrations of pic-oeukaryotes and Synechococcus at the basis of the mixedlayer, between 60 and 80 m depth, less affected by windmixing.

Detmer and Bathmann (1997) also reported that aparticular feature of HNLC systems was the dominanceof pico- and nanoplanktonic phyto-flagellates to thevirtual exclusion of diatoms. The observations at the

Fig. 7A, B Time course of Synechococcus vertical distribution ofright-angle light scatter (RALS) (A) and red fluorescence (B) at Sta-7 during the time-series experiment. RALS and fluorescence signalswere normalized with respect to corresponding signals of 1-lmbeads (see Materials and methods)

Fig. 8 Time course of the vertical distribution of temperature (�C)and salinity at Sta-3 (Polar Frontal Zone) during the time-seriesexperiment

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process stations are consistent with the parallel meso-scale study (Fiala et al. 2003) reporting that cyanobac-teria contributed 68% to total phytoplanktonabundance in the STZ, and picoeukaryotes 76% to totalphytoplankton in the PFZ. Heterotrophic bacteriaabundance was highest in the Convergence AF/STFzone, and of the same order in the STZ and the PFZ.Summer bacterial abundance (1.5–5.5·105 cells cm)3,Fig. 9D) observed in the PFZ was similar to thatobserved near the Kerguelen Islands during the‘‘Antares 3’’ cruise in October 1995 (M. Denis, unpub-lished data).

Short-term variations

In addition to the spatial variations of its distribution,ultraphytoplankton abundance also exhibited largeshort-term variations. Nanoeukaryotes showed thelargest (up to 80%) variations between two consecutivesamplings (2 h) in the STZ (station 8) between 1248 and1451 hours. The amplitude of the short-term variabilitywas different between the investigated hydrologicalzones. Picoeukaryotes showed the largest (up to 73%)abundance variation in the Convergence zone (Fig. 6C,between 1605 and 1749 hours). For Synechococcus, itslargest (41%) abundance variation was observed in the

PFZ (Fig. 8B, between 0619 and 0750 hours). Short-term abundance variations have been shown (Sherry andWood 2001; Binder and DuRand 2002) to result from acombination of biological processes such as cell divisionand cell mortality (e.g. grazing, viral lysis, UV-inducedlysis), and physical processes, such as cell export orimport (mixing episode, wind stress), which makesinterpretation of the abundance data very difficult. Inaddition, the highly dynamic nature of the study areamay generate advection at different levels in the watercolumn. For example, the sharp abundance decreaseobserved around 1000 hours in the STZ could have re-sulted from the advection of a different water mass witha lower planktonic content rather than grazing. Thehigher surface temperature at night than in daytime(Fig. 2) supports the change of water mass during thetime series despite the Lagrangian sampling mode. If weconsider the period of the time series at station 8 startingfrom 0700 hours, which seems to belong to the samewater mass (Fig. 2), the general trend of the short-termvariations corresponds to an increase in abundanceduring daytime leading to a peak in late afternoon. Theabundance decreased at dusk (2044 hours) and thenreturned to maximum values throughout the night. Thecyanobacteria concentration increase (Fig. 3C, D) wasprobably due to cell division, as observed in the equa-torial (Vaulot et al. 1995; Vaulot and Marie 1999; Binderand DuRand 2002) and subtropical Pacific Ocean (Liuet al. 1997). Cyanobacteria and picoeukaryotes exhibitedvery similar patterns of abundance variation, which wasalso observed in the equatorial Pacific (Partensky et al.1996; Blanchot et al. 1997). The short-term variability

Fig. 9A–D Time course of the vertical distribution of A nanoeuk-aryote, B Synechococcus, C picoeukaryote abundance (103 cellscm)3), and D heterotrophic bacteria (105 cells cm)3) at Sta-3 (PolarFrontal Zone) during the time-series experiment

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observed in the PFZ was different from that in the STZfor the comparable period. In contrast with the STZ, thelowest abundance values in the PFZ were observedduring the night. Mayzaud et al. (2002) reported thatmaximum mesozooplanktonic biomass occurred duringthe night in the PFZ, and during daytime in the STZ.These authors suggested that mainly salps, pteropodsand copepods to a lesser extent, contributed to thegrazing pressure. Pteropods can feed on particles assmall as 1-lm bacteria (Gilmer 1974, 1990) and, there-fore, could be responsible for ultraplankton decrease, atleast to some extent, when swarming in the surface layer.This is consistent with the large pigment content foundin the pteropod guts by Mayzaud et al. (2002) in thePFZ where picophytoplankton was dominant.

Ultraphytoplankton optical properties

A decrease in mean cell size over a time period can betaken as indicating the occurrence of cell division, andinversely, an increase of cell size can be associated withcell growth. However, the lack of observed cell size de-crease does not necessarily mean absence of cell division(Binder and DuRand 2002). In the STZ, Prochlorococ-

cus and Synechococcus exhibited similar variation pat-terns of RALS, used as a proxy of cell size (Fig. 5A, C).Such a similarity was also observed in the equatorialPacific (Binder et al. 1996; DuRand and Olson 1996;Blanchot et al. 1997; Vaulot and Marie 1999; Mann andChisholm 2000). Above the thermocline, cyanobacteriaRALS was the lowest before midday and then increasedsharply, probably due to cell growth. There was a goodcorrespondence between decrease in RALS and increasein abundance, which strongly suggests the occurrence ofcell division. RALS displayed increased values withdepth, more sharply below the thermocline, as previ-ously observed by several authors (Olson et al. 1990b;Partensky et al. 1996; Vaulot and Marie 1999;Dusenberry et al. 2000; Shalapyonok et al. 2001). ThisRALS depth dependence was less pronounced duringnighttime. In the Convergence zone and PFZ, Syn-echococcus RALS displayed the same pattern, suggestinga sharp increase of cell size below the thermocline. As inthe STZ, maximum abundance values correspondedwith minimum RALS values, and inversely.

The short-term variability of cell red fluorescence wasless pronounced in contrast with other investigationswhere the authors found strong short-term changes inred fluorescence (Partensky et al. 1996; Vaulot andMarie1999; Dusenberry et al. 2000). This is probably due to thefact that these studies were conducted on live samplesand not on fixed cells as in the present work, which wouldaffect the cell fluorescence, as shown by several authors(Vaulot et al. 1989; Vaulot and Partensky 1991; Landryand Kirshtein 1993; Partensky et al. 1996). We usedautofluorescence as an indicator of single cell pigmentcontent. In the STZ (Figs. 5B, D, 7B), normalized fluo-rescence patterns were quite similar for Prochlorococcusand Synechococcus. Red fluorescence did not vary greatlywith time above the thermocline, the lowest valuescorresponding to peaks of abundance. Like RALS, flu-orescence displayed a very strong vertical gradient, dueto the opposing effects of photoinhibition in the surfaceand photoacclimation at depth. The photoacclimation ofcyanobacteria to low light intensities has been reported inseveral areas: North Atlantic (Li and Wood 1988; Olsonet al. 1990a, 1990b; Moore et al. 1995; Partensky et al.1996; Dusenberry et al. 2000, 2001), Mediterranean Sea(Bautista and Jimenez-Gomez 1995), Arabian Sea(Sherry and Wood 2001) and in the equatorial Pacific(Olson et al. 1990b; Campbell and Vaulot 1993; Durandand Olson 1996; Partensky et al. 1999a, 1999b). Fluo-rescence increase with depth implies that the rate ofvertical mixing is slower than the kinetics of photoacc-limation. Blanchot and Rodier (1996) reported thatphotoacclimation of both Synechococcus and Prochlo-rococcus also depends on nitrate concentration (thresh-old of 0.1 lM), which would explain the absence ofphotoacclimation for Synechococcus and Prochlorococ-cus in the upper 30 m at Sta-8 in the STZ (Fig. 4B, D), atypical oligotrophic zone, where nitrate concentrationswere<0.09 lM above 60 m depth. This nitrate limitationdid not apply to Sta-7 and Sta-3 (PFZ) where surface

Fig. 10A, B Time course of the vertical distribution of normalizedSynechococcus right-angle light scatter (RALS) (A) and redfluorescence (B) at Sta-3 during the time-series experiment. RALSand fluorescence signals were normalized with respect to corre-sponding signals of 1-lm beads (see Materials and methods)

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nitrate concentrations were about 8 and 15 lM, respec-tively. The presence of two different ecotypes ofProchlorococcus in the STZ (Olson et al. 1988), couldalso account for its deeper red fluorescence increase withdepth (Fig. 4B). Indeed, several studies concluded thatthe depth-dependence of fluorescence and RALS ofProchlorococcus would be linked to a transition betweenphylogenetically distinct groups of organisms charac-terized by different red fluorescence levels (Campbell andVaulot 1993; Moore et al. 1995, 1998). The dimmestfluorescence ecotype, adapted to high light intensities,would be dominant in the upper layer and the brightest,adapted to low light intensities, in the subsurface layer(Campbell and Vaulot 1993; Partensky et al. 1996,1999b).

The surface mesoscale study of Fiala et al. (2003)revealed the low contribution of large cells (>20 lm), aswell as the low contribution of siliceous organisms to thetotal phytoplankton in the study area during australsummer. The present investigation, conducted in australsummer, contributed to documentation of the verticaldistribution of ultraplankton in the Crozet Basin, andprovided additional evidence of the dominance of thesmallest cells (<3 lm) in the upper 150 m of the dif-ferent hydrological domains sampled. The objective toseparately interpret spatial and short-term variations didnot fully succeed because of the intense hydrodynamicprocesses occurring in this area. Consequently, relyingon a Lagrangian sampling mode in such an area wouldnot be realistic. Nevertheless, the observed variationsdemonstrate that abundance (and consequently bio-mass) may vary by a factor of 3 within a period of 24 h.This variability should be taken into account whenderiving carbon budgets from discrete measurements,and extrapolating over time and space.

Acknowledgements This work is a French contribution to theSO-JGOFS program. It was supported by the French Polar Institute(IPEV) and the INSU-CNRS. We thank the captain, officers andcrew of RV ‘‘Marion Dufresne’’ and B. Ollivier for outstanding on-board support. Special thanks go to Y.-H. Park for his advice andstimulating discussions. We are grateful to the anonymous reviewersfor their constructive remarks and manuscript improvements.

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