Pergamon

I

Deep-sea Research I, Vol. 44, No. 7, pp. 1255-1275, 1997 0 1997 Elsevier Science Ltd

All rights reserved. Printed in Great Britain P l l S0967-0637(97)00014-9 0967-0637/97 %17.00+ 0.00

Biomass, growth rates and limitation of Equatorial Pacific diatoms

STÉPHANE BLAIN,* AUDE LEYNAERT," PAUL TRÉGUER," MARIE-JOSÈPHE CHRÉTIENNOT-DINET? and MARTINE RODIERt

(Received 24 July 1996; in revised form 13 January 1997; accepted 28 January 1997)

Abstract-Biomass, growth and species composition of siliceous phytoplankton were studied in the Equatorial Pacific during October 1994. Experiments were carried out in different nutrient conditions along the Equator. An oligotrophic area, with nitrate concentrations as low as 10 nM in the upper layer, was encountered in the western part of the transect (166"E-17OoW). The concentration of biogenic silica varied from 10 nmol 1-' in the surface layer up to 40 nmol i-' in the deep chlorophyll maximum located near the nutracline. Biogenic silica production, measured by the 32Si method, showed a similar vertical pattern in the nitrate-depleted water, and the mean assimilation rate for Si was 0.4k0.2 nmol 1-' h-' (integrated mean value: 63 p o l m-' h-I). In contrast, nitrate concentration ranged from 2 to 4pM in the surface layer in the high-nutrient low-chlorophyll (HNLC) area located from 170 to 150"W and biogenic silica increased to 200nmo11-1. The Si assimilation rate was 1.7+ 1.0 nmol 1-' h-' (integrated mean value: 162 pmol m-' h-I). In both areas, 80% of Si biomass was concentrated in larger cells (> 10 pm). Scanning electron microscopy was used to estimate diatom numbers and cell surface areas. This latter parameter correlates well with biogenic silica and warrants a discussion of the contribution of different species to the total biogenic silica. The measured values for specific uptake rate of Si never reached the optimum uptake or growth rate deduced from environmental parameters and kinetic constants reported in the literature. In addition the mean growth rate (0.9 + 0.3 doubling per day) for the nitrate-depleted water does not differ from the mean value (0.850.2 doubling per day) in the HNLC area. Therefore it can be concluded that diatom growth is severely limited in both regions. This agrees well with the nutrient balance study. In the oligotrophic area, N supply appears to be the limiting factor in the upper layer. In the HNLC region, the results of this study are consistent with the hypothesis that diatom growth might be limited by a micro nutrient such as iron. Carbon production by diatoms was estimated to be 31 +4 mmol C m-2 d-', which is one-third of the total carbon production of the nutrient-enriched area. 0 1997 Elsevier Science Ltd

INTRODUCTION

Because of the large oceanic source of carbon dioxide located in the Equatorial Pacific (Tans et al., 1990), this area has received a great'deal of attention during recent years. Murray et al. (1994) concluded that physical, rather than biological, processes control the CO2 flux in this region. However, several major biological features of the region are still poorly understood. Diatoms may play a major role in oceanic new production (Goldman, 1993) and biogenic fluxes (Takahashi, 1987). The Equatorial Pacific is one of the three large oceanic areas considered to be a representative high-nutrient low-chlorophyll (KNLC) regime. Among

* UMR CNRS 6539, Institut Universitaire Européen de la Mer, UBO 6, Avenue le Gorgeu, BP 809 29 285, Brest

-f Observatoire Ockanologique de Banyuls, F-66650, Banyuls sur mer, France. Cedex, France.

' f: Ce<Ee'O;rstorsrl-de-Nopka, BP A5, Nouméa Cedex, New Caledonia. \

1255 Fonds Documentaire ORSTOM Cote s $fi13838 EX :

1256 ‘

‘ it.

S. Blain et al.

the different hypotheses attempting to solve this paradox, grazing pressure and limitation by the micro nutrient iron are the most debated (Chisholm and Morel, 1991; Martin et al., 1991; Cullen, 1995). In this context the role of large phytoplankton, especially diatoms, is suspected to be a corner-stone of the problem (Price et al., 1994).

Diatoms also produce very particular events as described by Yoder et al. (1994). The physical accumulation of diatoms along a thermal front created a dramatic increase in biomass and could help to explain the vast laminated diatom mat deposits observed by Kemp and Baldauf (1993) in the eastern Equatorial Pacific Ocean. TrCguer et al. (1995) identified the Equatorial Pacific as one of the main sinks for silica in the world ocean, and Sancetta et al. (1991) asked “Massive fluxes of rhizosolenid diatoms: a common occurrence?”.

Despite the potential role of diatoms in all of these critical issues, few experiments have been devoted to the specific study of biomass, growth and limitation of siliceous plankton species in this area. The first (Pavillard, 1935) and subsequent (Hasle, 1959; Desrosière, 1969) studies about the abundance and composition of phytoplankton mentioned some taxonomic or quantitative observations of diatoms. Thomas and Dodson (1975) measured silicic acid requirements for three oceanic diatoms in the eastern tropical Pacific Ocean. More recently Chavez et al. (1990), using epifluorescence microscopy, described the abundance of the major taxonomic groups in the picoplankton to microplankton range and their relation to physical and chemical properties and primary production. They concluded that primary production along the Equator could be highly correlated to the abundance of pennate diatoms. These conclusions were confirmed by Iriarte and Fryxell (1993, who reported detailed observations on the micro-phytoplankton (species composition, cell number and biomass) in the central Equatorial Pacific. They suggested that most of the variability of the carbon fluxes in the Equatorial Pacific could be explained by variability in the biomass and species composition of diatoms, both driven by environmental variability in this area, particularly El Niño events versus “cool” conditions.

Few data are available on the contribution of diatoms to primary production in the Equatorial Pacific. Some indirect indications (Chavez et al., 1991) were deduced from the growth rate of the > 5 pm fraction of the natural population. Dugdale et al. (1995), comparing nutrient distributions and the results of a model, suggested that a “silica pump” may control and dominate new production in the HNLC area off Peru. At the same time, using 228Ra-derived nutrient budgets, Ku et al. (1995) proposed that productivity may be limited by the availability of new silicate.

In this paper the first direct measurements of biogenic silica (BSi) and of silicic acid uptake rate by phytoplankton are reported, together with the phytoplankton species composition in two contrasting areas of the Equatorial Pacific: the western Equatorial Pacific, where oligotrophic conditions prevailed, and the central Equatorial Pacific, representative of HNLC systems. The comparison indicates that neither in the oligotrophic area nor in the HNLC area are diatoms growing at rates close to their maximum growth rate.

MATERIALS AND METHODS

As part of the France-JGOFS Equatorial Pacific Process Study, sea water samples and hydrographic data were collected during the FLUPAC cruise (September 23-October 29, 1994), aboard the French R/V L’Atalante. This cruise took place during the 1994 warming event (Liu et al., 1995). The cruise plan consisted of two legs along 165”E and along the

- I.!{- : . . : i ~ ~ ! ! 4 > l x ; “ I t j \ ,>:<)4)j

.I ì '

Equatorial Pacific diatoms 1257

Equator, and two Equatorial time-series stations (7 days): at 166"E and 150"W. Details on data and data acquisition are available in the cruise reports (Le Borgne et al., 1995; Le Borgne and Gesbert, 1996). In this paper only the Equatorial transect data set of 10 stations from 167"E to 150"W are considered. The light intensity on the deck was monitored continuously with a solar radiation sensor (2770 Orca, France). Hydrographic data were obtained at each station of the Equatorial leg with a SeaBird SBE-9llplus CTD and sensors, equipped with Aquatracka III (Chelsea Instruments) fluorometer and SeaTech transmissometer (more details are given in Eldin et al., submitted). From these routine CTD casts, discrete water-column samples for nutrients, pigments, and particulate matter and microscopic determinations were collected at 12-14 depths with a SeaBird "Carrousel" mounted with 12 Niskin and 12 NOEX bottles.

The nutrients (nitrate, nitrite, ammonium, phosphate and silicic acid) analyses were done by standard automated colorimetric methods (Strickland and Parsons, 1972). A high sensitivity method (Oudot and Montel, 1988) for the determination of nitrate and nitrite at nanomolar concentrations was applied when concentrations of both ions were less than 1.5 pM. Details of our routine analytical techniques can be found in Bonnet (1995).

Biogenic silica (BSi)

One or two litres of sea water were filtered immediately after collection under gentle vacuum through 0.6 pm Nuclepore membrane filters. The filters were dried for 12 h at 60°C and stored in a plastic petri dish before analysis. Biogenic silica analyses were performed using the NaOH digestion method (Paasche, 1973) modified by Ragueneau and Tréguer (1994). The blank was 8 nmol 1-I, and the limit of detection, determined as three times the standard deviation of the blank, was 5 nmol 1-l.

Silicon uptake rate (psi) and specific uptake rate ( Vsi)

Two hundred and fifty millilitre polycarbonate bottles were carefully filled with sea water collected with Niskin bottles retro-fitted with silicone rubber. Each bottle was covered with a nickel light attenuator to simulate in situ light intensity at different depths (100,25, 10,3, 1 and 0% of the incident light). The recent production of the radionuclide 32Si with a very high specific activity (42165 Bq/pg Si) enabled use of it in waters where ambient silicic acid concentrations were very low (less than 2 pM). The stock solution was diluted in order to add 30 pl or 42 850 & 400 cpm per sample. The addition of 32Si increased the in situ silicic acid concentration by less than 0.1 yM. The radio tracer 32Si was injected just after sampling, and the bottles were placed in a deck incubator maintained at surface sea water temperature by sea water circulation. A11 the incubations started before sunrise and were of 24 hours duration. Incubations were terminated by filtration of the sample through 0.6 pm Nuclepore membrane filters, which were then rinsed with filtered sea water and placed in plastic counting vials. The final counting was carried out 6 months later by Cerenkov radiation, when secular equilibrium had been attained. The Si steady-state uptake rate (p nmol I-' h-') was deduced from the final activity on the filter, the silicic acid concentration and the initial injected activity. The specific uptake rate, Vsi(day- I), was calculated from the equation Vsi = psi/BSi. We also calculated the growth rate (doubling per day) for Si biomass as psi = Vsi/ln2.

1258 S. Blain et al.

Microscopy

For scanning electron microscopy (SEM), sea water sub-samples (500 ml) were collected from Niskin bottles and immediately filtered onto Nuclepore filters (Costar Corporation, 0.8 pm) using a manual vacuum pump. The filters were rinsed with distilled water to remove salt, air dried, placed on stubs and coated with gold before examination under an Hitachi S 250 scanning electron microscope.

Cell counts and surface estimations

Cell counts were made on stubs with SEM. The whole stub was examined for larger cells, and a magnification of x 1000 was used along a transect for the smaller ones. After calculations according to the scanned surface area and the filtration volume, diatom numbers were estimated. In order to get a corresponding amount of silica, cell surfaces were calculated from size measurements of the different species. For each sample examined, a total diatom surface was calculated as a silica equivalent.

RESULTS Nutrient distribution

General conditions observed along the Equatorial leg are described in detail in Eldin et al. (submitted). Their description of the nutrient distributions (Fig. 1) reveals that two different structures were successively sampled from 167"E to 150"W, with a transition zone between 172 and 168"W. West of 172"W, the waters were oligotrophic and characterized by a two- layer system. In the surface waters, nitrate concentrations were < O. 1 pM (most of the time under the 0.002 pM limit of detection) and silicic acid concentrations were approximately 1 pM. At greater depth, the thermocline separated the nutrient-rich deep water from the nutrient-impoverished surface euphotic zone. East of 172"W, nutrient distributions changed through the transition zone, from an oligotrophic situation to a high-nutrient low- chlorophyll (HNLC) regime. Between 172 and 168"W, the top of the nitracline (as defined by the O. 1 pM NO3- isoline) rose from 80 m to the surface. Further east, significant surface nitrate enrichment up to 3.55 pM was observed. Silicic acid presented similar features, with surface concentrations up to 2.6 pM.

The molar ratio Si(OH)4/(N03- +NO2- +NH4+) is also shown in Fig. 1. From 170"E to 167"W, this ratio was greater than 1 due to surface nitrate depletion in this region. This ratio was even higher (up to 40) between 177.5 and 178.5"E (Stations #66, #68 and #70) because of a low ammonium level. Near the thermocline and in the eastern part of the transect (Stn 76-80), the ratio was close to 1.

Biogenic silica distribution along the Equator

Figure 2 shows the distribution of biogenic silica (BSI) along the Equator. The concentrations in the oligotrophic region were very low in the upper layer, close to the limit of detection of our method (5 nmol 1-'). A deep maximum of BSi was found near the nitracline, as shown in Fig. 3a, but the concentration of BSi never exceeded 40 nmol 1- '. In contrast, the enriched water east of 170°W, presented BSi profiles (Fig. 4a) with homogenous concentrations (> 100 nmol 1-I) in the mixed layer. Then BSi values decreased with depth to 20 nmol 1-'.

Y I ' f

Equatorial Pacific diatoms 1259

# station 62 64 6 6 6 8 70 I2 14 76 78 80

O

. . . . 100

[NO;] ( pmol. 1.')

./. . . 2 1 ; . . . . . . ' . . f - . . . . . . . . . . .

zoo! , , , , . . . a I . . . . I . a . I I u . . . I * - I ' * ' r 180" 170"W 16O"W 155"W 170"E

[Si(OH4)] ( p o l . 1.')

170"E 180" 170"W 16O"W 155"W

Fig. 1. Contour plot of nitrate, silicic acid and the ratio silicic acid concentration versus total nitrogen concentration expressed as nitrate plus nitrite plus ammonium in the upper water column

(0-200 m) for the transect along the equator between 170"E and 155"W.

1260 S. Blain et al.

# stations

62 64 66 68 70 72 74 76 78 80

-30

-60

n - -90 B

-120

-30

-60

-90

-120

-150 1 ' 1-150

-180 -180 l ~ ~ ~ ~ " ~ " ~ " ~ " ~ ~ ~ ' l ~ ' ~ ' l ~ ~ ~ ' ~ ' 170"E 175"E 180"E 175"W 170"W 165"W 160"W 155"W

-1 Biogenic siiica nmol.1

Fig. 2. Contour plot of biogenic silica along the Equator.

Taxonomic composition, diatom counts and species contribution to biogenic silica

Results are shown in Table 1 for an oligotrophic site (Stn 64), and in Table 2 for an HNLC area (Stn 78). About 30 different taxa were taken into account, but only 20 were present in the water column at each station. Nitzschia bicapitata Cleve was found to be the most common species with two size types (10 and 30 pm in length). However, its relatively small size as compared to large centric or pennate diatoms excluded it from being a dominant element of the whole community. As biogenic silica correlated well with the diatom surface area, the percentage of biogenic silica referable to each species is included in Tables 1 and 2.

The two sites differ not only in their taxonomic composition but also in the distribution and abundance of diatoms. At station #64 the maximum diatom number was found at 90 m and was associated with the deep chlorophyll maximum. The community was highly diversified with 16 species present at this depth. Pennate diatoms were dominant in the water column, and centric forms were absent from subsurface water (30 m) as well as below the thermocline (140 m), where Pseudo-nitzschia accounted for 36%. At station #78, the partition between pennate and centric diatoms was more bal,anced, with slight dominance of centric diatoms in surface and sub-surface waters (5 and 40m). The maximum diatom biomass was also related to the chlorophyll maximum found at 40 m, with 14 species almost exclusively represented by centric diatoms. Chaetoceros was the main component (47%), in terms of biogenic silica, of centric diatoms and Pseudo-nitzschia accounted for 36% of the pennate diatoms. At 80 m, the centric diatom contribution was reduced to 6%, and the dominant element was again Pseudo-nitzschia.

I

Equatorial Pacific diatoms 1261

Biogenic silica (nmol. 1-')

O b I

O 20 40 60 80

20- (a) I / \\,t.

40- 60-

3 9 & 100- 80-

120- 140-

160- 180-

a

1 -1 -1 Biogenic silica production (nmol. 1 h )

o 0.1 Q.2 Q.3 Q.4 O 1

20- (b) O \ '

i 40-

n 60- 7

3 e 100- \ - E 80- .l.

i a 120- 140- 0 '

160-

180- Fig. 3. Biogenic silica concentration (a), and production rate (b), profiles at the Equator and 170"E,

station #62 (oligotrophic site).

Silicon uptake rate

Strengths and limitations of the 32Si method. The measurement of silicic acid uptake is generally performed using isotope tracer. The stable isotopes 30Si and 29Si have been used successfully (Nelson and Goering, 1977a,b) in various systems. However, the method is tedious in preparing samples for final mass spectrometric determination. Moreover, although recent improvements in the precision of the analysis by mass spectrometry have been made (Nelson and Brzezinski, in press), the limit of detection of the method requires the addition of significant amounts of tracer, which can modify the initial concentration of silicic acid in the sample (especially in oligotrophic systems).

In the last few years, a new method based on the radioisotope 32Si has been developed (Tréguer et al., 1991). Recently, the production of a new 32Si source (Phillips et al., 1995), with a much higher specific activity (42 165 Bq/pg Si), has enhanced the potential of that

" i 1 2

1262 S. Blain et al.

,

O

20-

40- - 60- 0

80- 8 100- a

120-

140-

160-

Biogenic silica (nmol. 1-l)

O 50 100 150 200 O I

20- 40- - 60- E - 80- s!

e 100- J' 'Lt

120-

160

I I I I I I 2 0) ./'

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method for measuring silica production rates, enabling us to use it in oligotrophic areas. The major advantage of this method lies in the high sensitivity of the detection, which allows reduction of the amount of the injected tracer. This method is very suitable for oligotrophic sea water, as demonstrated during the present cruise, because the injection of the 32Si contributed less than 10% of the initial concentration of silicic acid. Clearly the method measures the steady state uptake rate (see Morel, 1987 for definition) of 32Si by the plankton community, but it is necessary to discuss briefly the real significance of this parameter and its link with those measurements describing how the ecosystem is functioning. The regeneration of Si is still poorly understood and documented, but on this time scale, it is realistic to assume that the silica dissolution principally affects older parts of the frustule. This implies limited dissolution of newly absorbed 32Si. Therefore, the main event that would release 32Si could be the release of intracellular Si in living cells by grazers. But this pathway seems to be minor because first, Binder and Chisholm (1980) reported that the

Equatorial Pacific diatoms 1263 c

Table 1. Taxonomic composition, cell size, surface area andpercentage of biogenic silica of diatoms at station #64 (oligotrophic site)

Depth

Diatom species Size Surface 30m 30m 90m 90m 140m 140m w pmz Cell.l-' % BSi Cel1.1"' % BSi Cell.1-' % BSi

Nitzschia bicapitata Cleve Nitzscia closterium (Ehr.) W. Smith Pennate diatom Nitzschia bicapitata Cleve Psmmadictyon panduriformis (Gregory) Mann Pseudo-nitzschia sp. 3 Thalassionema nitzschioides (Grun.) Grunow ex Hustedt Chaetoceros dichaeta Ehrenberg Pseudo-nitzschia sp. 1 Leptocylindrus danicus Cleve Pseudo-nitzschia sp. 2 Chaetoceros sp. 2 Thalassiosira cf. lineata Jousé Thalassiosira sp. Actinocyclus Synedra sp. Mastogloia Thalassiosira cf. lentiginosa (Janisch) G. Fryxell Centrique diatom Planktonietta sot (Wallich) Schiitt Chaetoceros sp. 1 Bacteriastrum cf. furcatum Shadbolt

10 x 2.5 15x3

15 x 2.5 30x3 18x5

30x5 30 x 4.5

20x 10 50 x 3 5 10 x 40 85x5

10 20

20-25 25x5 150x5 50 x 30

35

40 70 30 10

Proboscia sp. 500 x 2.5-5 Chaetoceros peruvianus Brightwell 25 x 20 Hemidiscus sp. 200 Asteromphalus sp. 1 1 O0 Asteromphalus sp. 2 1350 Thalassiothrix gibberula Hasle 1000 x 8 Rhizosolenia sp. 400 x 50

1 O0 120 200 300 460

500 600

630 650

1200 1350 1800 2000 2300 2350 2350 2500 2500

2600 4200 4500 5600 7850

12 O00 15 O00 17 300 20 O00 25 O00 62 800

3500

1500 23 500 500 12

9500

3500 2500

5 500

35 25 5

5 - 500 5

5 10

500 10

5 64 75

2 2500 8 4000 22

1

32

12 5 - 9 2000 36 - 5 - 4

- 15 "1

7 500 32

- -

-

22 -

II

soluble silica pool represents only 1-2% of the intracellular Si, and second, large grazers are underrepresented in a 250 ml volume incubation. The redissolution of biogenic silica could also contribute bias to the silica uptake rate measurement by decreasing the 32Si specific activity during the incubation. This is assumed to be negligible in the current study, because the BSi concentration did not exceed 10% of the silicic acid concentration.

Finally, in the Equatorial Pacific, the uptake of Si by organisms other than diatoms, principally radiolarians, must be examined carefully. Scanning electron microscopic observation allowed us to correlate biogenic silica, not only with the number of diatoms [r = 0.7789 (n = 6)], but also with an estimation of the surface of the cell [r = 0.8673 (n = 6)] . This good correlation implies that diatoms represent a large part of the biogenic silica. This conclusion was confìrrned by the observed contribution of radiolarians and acantharians to

1 , '

1264 S. Blain et al.

Table 2. Taxonomic composition, cell size. surface area andpercentage of biogenic silica of diatoms at station #78 (HNLC site)

Depth

Size Surface 5m 5m 40m 40m 80m 80m Diatom species Pm p2 Celll-' % BSi Celll-' % BSi Ce11.1-I % BSI

Nitzschia bicapitata Cleve 10x2 Nitzschia closterium ('Ehr.) W. Smith". 15 x 3 Pennate diatom Nitzschia bicapitata Cleve Psmmadictyon panduriformis (Gregory) Mann Pseudo-nitzschia sp. 3 Thalassionema nitzschioides (Grun.) Grunow ex Hustedt Chaetoceros dichaeta Ehrenberg Pseudo-nitzschia sp. 1 Leptocylindrus danicus Cleve Pseudo-nitzschia sp. 2 Chaetoceros sp. 2 Thalassiosira cf. Iineata Jous6 Thalassiosira sp. Actinocyclus Synedra sp. Mastogloia Thalassiosira cf. lentiginosa (Janisch) G. Fryxell Centrique diatom Planktoniella sol (Wallich) Schütt Chaetoceros sp. 1 Bacteriastrum cf. furcatum Shadbolt Proboscia sp. Chaetoceros peruvianus Brightwell Hemidiscus sp. Asteromphalus sp. 1 Asteromphalus sp. 2 Thalassiothrix gibberula Hasle Rhizosolenia sp.

Total

15 x 2.5 30x3 18x5

30x5 30 x 4.5

20x 10 50 x 3-5 10 x 40 85x5

10 20

20-25 25x5 150 x 5 50 x 30

35

40 10 x 8

70 30 10

500 x 2.5-5 25 x 20

200 100 150

1000 x 8

400 x 50

100 1500 - 120 200 300 3000 4 2500 4 4500 17 460 500 1

500 600 1000 2

630 650

1200 1350 1800 2000 2300 2350 2350 2500 2500

2600 2700 4200 4500 5600 7850

12 O00 15 O00 17 300 20 O00 25 O00

5500 34

500 1 90 - 25 - 16000 42 11 O00 36

1500 25

35 - 45 - 10 -

20 - 5 -

5500 40 5500 50

20 -

15 -

25 - 15 - 20 - 40 2 30 1

5 -

5 - 5 - 5 -

10 - 10 1 50 15

62 800 30 8 20 6 5 4

the total mesozooplankton dry weight (Le Borgne and Rodier, submitted). To get an upper limit of the contribution of these organisms to biogenic silica it was assumed that their dry weight was entirely made of silica and it was compared to measured biogenic silica. Radiolarians and acantharians did not contribute more than 2% of the biogenic silica in the mixed layer above the thermocline.

If all of these preliminary remarks are considered, 32Si uptake rate determination is a good proxy for the determination of gross uptake rate of Si by phytoplankton and gives relevant information about the gross growth rate of siliceous phytoplankton in the studied areas. Finally, the specific uptake rate measured isotopically is a minimum, estimate because a part of the biogenic silica could be detrital.

. . Equatorial Pacific diatoms

I I I I I 1 1 1 1 1 1 1 1 1 1 ~ 1 I I I I I I I I I I I 1 I l I t I I l

h v E -20 -40 -60- - - ; :e;i: f? a

-80 - 0.6

1265

-

-

Measurements. Figure 5 shows the distribution of the Si uptake rate along the Equator. The lowest value was found in the surface water of station #62 ( < 0.2 nmol 1- ' h- I). The production rate increased with depth as illustrated in Fig. 3b and was maximum near the depth of the chlorophyll maximum. There was a tendency for the uptake rate to increase from west to east in the oligotrophic area, where the value was 0.6 nmol 1-' h-' at station #74. In the surface water of the HNLC area, the Si uptake rate was roughly 10 times higher than in nitrogen-depleted waters. A representative vertical profile (station #78) of this area is plotted in Fig. 4b. The uptake rate was maximal in the surface layer and decreased with depth. In both regions, the production rate tended to coincide vertically with the distribution of biogenic silica.

In both areas, siliceous assemblages were producing biogenic silica at a depth where little or no photosynthesis was taking place. For the dark bottles the mean uptake rates were, respectively, 0.7 f0.3 nmol I-' h-' in the oligotrophic water and 0.6 10.2 nmol 1-' h-' in the HNLC water.

Integrated from the surface to the 0.1 % light depth, the daily rates of biogenic silica production averaged 1.5 mmol m-2 day-' in the oligotrophic area and 3.8 mmol m-' day-' in the HNLC zone. In the oligotrophic zone, the deep sub-surface maximum in p contributed more than 50% to the integrated production rate.

The specific uptake rate V(day- ') or the growth ïate p (doubling per day) were calculated from the measured BSi and p. It should be noted that the precision of V (or p) is the sum of the relative uncertainty on BSi and p. Three depths at station #68, where the BSi concentrations were close to the limit of detection, were deleted from the results because they introduced a large uncertainty to the calculated growth rate. An isolated high value (4.6 doubling day-') in surface water at station #76 was also excluded, because the lack of other information, such as species composition, did not allow the ruling out of an experimental artefact. The different profiles for the oligotrophic stations are presented in Fig. 6. For the samples that received more than 10% of the light intensity arriving to the surface, the

1266 S. Blain et al.

growth rates increased from station #62 to station #66. This increase coincided with the increase in the daily integrated light intensity from 2.7 x lo5 J m-' day-' to 4.3 J m-' day- '. A light limitation at stations #62 and #64 is confirmed by comparison of the PAR measured during the incubation (450 and 1000 pE mP2 s-l respectively for the maximum value) and the Photosynthesis-Irradiance curve determined by André and Sadoudi (1995). For stations, #66 and #70 the daily integrated light intensity was constant, and the mean profile is shown in this paper (mean light intensity was 4.6 x 105$0.2 J m-2 day-' and the maximum PAR was 1800 pE m-' s-'). This mean profile is compared in Fig. 7 with the mean profile for specific Si uptake rate in the HNLC system (stations #76, #78 and #80). The integrated light intensity was 4.6 x 105k0.2 x lo5 J m-' day-'. The growth rate in surface water (light intensity 100 and 25% of the surface incident light) averaged 0.9k0.3 doubling day-' for the oligotrophic area, where no nitrate was detected, and 0.8 k0.2 doubling day- ' for the HNLC system, where the nitrate concentration was 2.6 pM. The two means are statistically indiscernible. Stations #72 and #74 have been excluded from this comparison because they were located in the transition zone between the two typical areas.

DISCUSSION

Evidence for limitation of diatom growth in oligotrophic und HNLC ureas

Kinetic approach. The uptake rate of nutrient by phytoplankton is generally described by a hyperbolic function of nutrient (Michaelis-Menten kinetics) linking the experimental

0.00 0.50 1.00 1.50

Growth rate (doubling per day)

Fig. 6 . Profiles of growth rate for the oligotrophic stations. Station #62 open diamond, station #64 open circle. The integrated incident light fluxes at the surface during the length of the incubation were 2.7 x lo5 and 3.2 x lo5 J m-' day-', repectively . The filled square represents the mean value for stations #66 and #70. The error bar represents the standard deviation. The mean integrated light flux

at the surface was 4.6 x 105_+0.2 x lo5 J m-' day-'.

Equatorial Pacific diatoms r"

4

1267

100

10

1 1 2 3

Gmwtb rate (doubling per &y) O 1 2 3

Growtb rate (donbliog per day)

Fig. 7. Comparison of the mean growth rate prome in the oligotrophic area (a), and in the HNLC area (b). The error bar represents one standard deviation. The integrated incident light fluxes during the incubations were 4.6 x lO'f0.2 x 10' J m-' day-' and 4.8k0.2 x 10' J m-' day-', repectively:

The nitrate (open square) and silicic acid (open circle) profiles are also shown.

parameters Vand [SI to theorical values Vmax and K, , by the relation V = Vmax A. It is possible to discuss the limitation of the uptake rate of a phytoplankton species by comparing the measured value with the opt i " value deduced from Vmax, Ksv and the ambient nutrient concentration. Vmax and Ksv were not measured during the cruise reported here, but some indications are available from the literature for other regions.

HNLC water. Goering et al. (1973) and Dugdale et al. (1981) report Ksv of 3 pM silicic acid and Vmax of 0.08 h- ' at 15"s in the Peru upwelling region. The ambient concentration of nitrate and silicic acid were, 12 and 2 pM, respectively. Therefore these constants were used to estimate the expected Si specific uptake rate for diatoms in the nutrient-enriched part of the transect, where [Si(OH)4] of 2 pM and [NO3-] of 4pM were measured. The calculated rate is 0.032 h-l. Nelson and Brzezinski (1990) also reported V& of 0.024 _+ 0.004 h- ' and 0.035 & 0.004 h- in a Gulf Stream warm-core ring, where [Si(OH)4] were 2.5 and 2.9 pM and the nitrate concentration 5 pM. The mean value of V measured in the HNLC area was 0.022&0.009 h-l and never reached the maximum value calculated above from Goering's data. This value also lies in the lower part of the range reported by Nelson and Brzezinski (1990).

oligotrophic waters. In a recent work Brzezinski and Nelson (1996) reported two sets of constants (V,,, and K ~ v ) deduced from kinetic experiments in the Sargasso Sea where nitrate was depleted. Using a [Si(OH)4] of 1.5 pM uptake rates within 0.017 h-' and 0.011 h-' for the first set and between 0.007 h-' and 0.003 h-' for the second set are

.

1268 S. Blain et al. ~

obtained. These values are lower than the measured values of 0.024 & 0.007 h- '. The kinetic constant measured in the Sargasso Sea appears to be significantly higher than those of the natural phytoplankton community of the western Equatorial Pacific.

Another way to describe phytoplankton growth is the Monod formulation A. The growth rate of the diatoms was not measured directly in the

incubation described in this study, but was estimated from the specific uptake rate. Furnas (1990) reviewed measured or estimated growth rates of phytoplankton assemblages. Most of the values reported for centric and pennate diatoms are above 2 doublings day-' in tropical area. Blasco et al. (1982) reported a maximum growth rate of diatoms around 2.5 doublings day- l.

In the upper layer of the oligotrophic area the diatoms were growing at less than 1 doubling day-'. This result is consistent with the limited Si uptake rate suggested above. Referring to Fig. 1 it is evident that the oligotrophic area is largely nitrogen, rather than silicic acid, deficient. So nitrogen would be expected to limit the growth rate of diatoms. A debate on the form of nitrogen (NO3- or NH4+) that may be preferentially used by large cells such as diatoms (i.e. Harrison and Wood, 1988; Taylor and Joint, 1990; Price et al., 1994) or on the nitrate uptake inhibition by nanomolar ammonium concentrations (Harrison et al., 1996) will not be entered into here. However, the N (nitrate, nitrite and ammonium) concentrations were low enough to limit the acquisistion of N by large cells by molecular diffusion (Chisholm, 1992). Brzezinski and Nelson (1996) demonstrate that Si uptake rate by diatoms in the Sargasso Sea may be limited by even at nanomolar mO3-1. They suggested that this paradoxical result may be not detrimental to diatoms, but an adaptive behaviour to survive with minor biochemical damage in severely depleted water. They also suggested this adaptive response may be common in most of the open- ocean- water with [Si(OH)4] < 3 pM. In the oligotrophic Pacific waters [Si(OH)4] is three times higher than in the oligotrophic Atlantic waters. So without kinetic experiments it is not possible to confirm this hypothesis, but it cannot be ruled out. However, the microscopic observations showed an increase in predominance of pennate diatoms over centric diatoms when the N-nutrient resources decreased. Chisholm (1992) suggested that this is a response of the organism to the diffusion-limited uptake rate. So this observation suggests N- rather than Si limitation.

Near the nitracline, the light intensity was less than 1 % of the incident light, but nutrients were more abundant. Goldman (1993) reported growth rate data of large diatom species from the Sargasso Sea cultured at a low light level (60 pE mF2 s-'): four of six species were growing at 1.OkO.1 doublings day-'. During the present cruise the PAR near the chlorophyll maximum was at a similar level. A mean growth rate of 0.9k0.3 doublings day-' was measured. The two sets of data are in good agreement and confirm the possible large contribution of diatoms to new primary production at the base of the euphotic zone, in response to episodic nutrient injections through the thermocline.

Thomas and Dodson (1 975) reported K,, and bax from three different diatom species in the eastern tropical Pacific. Mean K,, was 0.51 pM, and pmax was within the range 3.21-5.04 doublings day- ' . Although these high maximal rates were certainly associated with particular species composition and high biomass encountered during bloom conditions, they were used to estimate the growth rate during the incubations reported here. The growth rate of diatoms is expected to be between 2.6 and 4 doublings day-', which is much higher than the measured value 0.8 k 0.3 doublings day-'. Despite the uncertainty resulting from the estimation of a growth rate based on the measurement of uptake rate, it seems evident

lJ = Ksp+[sI

Equatorial Pacific diatoms 1269 ~

that diatoms growing at one-third or one-quarter of their optimal growth rate should be severely impaired.

The diatom limitation in the HNLC system is confìrmed by the comparison of the current growth rate measurements in both areas. So if a proven diatom limitation takes place in oligotrophic waters as discussed above, it should also be effective in HNCL water, because the growth rates were statistically indiscernible when light intensity was similar (Fig. 7).

Some authors have reported growth rates of diatoms in the Equatorial Pacific (150”W) from long bottle incubation experiments. These experiments were conducted to test possible iron limitation. Chavez et al. (1991) reported, for small Nitzschia sp., growth rates of 1 .O 0.3 doublings day- for control bottles and 1.35 & 0.06 doublings day- ’ for enriched cultures. Fryxell and Kaczmarska (1 994), using 6-day incubation experiments, measured growth rates of 1.9-3.4 divisions day-’, calculated on total diatom cell number. Chavez et al. (1991) also estimated a growth rate of 2 doublings day-’ for the > 5 pm fraction in the Equatorial Pacific. This fraction was assumed to be dominated by diatoms. They concluded that these organisms were growing faster than the overall population. Regarding measurements in the curent study it appears that diatom growth rates measured in bottle incubations conducted for several days are overestimated. The diatom population did not grow faster than other phytoplankton groups. For example Vaulot et al. (1995) reported growth rates of 0.73-0.86 doublings day-’ for Prochlorococcus measured in the central Equatorial Pacific (140”W).

Nutrient balance approach

The comparison of the environmental nutrient ratio with the elemental composition of the phytoplankton can give some information on the potential limitation of the phytoplankton by a major nutrient. The most complete data base on Si:C:N ratios for diatoms was compiled by Brzezinski (1985). The mean ratio Si:N determined for 18 species of diatoms was 0.95 & 0.23. If the Si:N ratio in seawater measured during the cruise (Fig. 1) is compared, it is evident that most of the samples from the upper layer of the oligotrophic area were above this range. This is consistent with the kinetic conclusion of the previous paragraph and the common vision of tropical ocean ecosystems limited by low nitrate flux (Lewis et al., 1986). In the HNLC area most Si:N environmental ratios are within the range previously reported, with Si(OH)4 concentrations slightly higher than NO3 - concentrations. Thus it is difficult to deduce which of these two elements would be exhausted first and would thus ultimately limit the production of diatoms. Another study (Wilkerson and Dugdale, 1996) reports the variability of surface NO3 and Si(OH)4 concentrations calculated from AVHRR derived SST data along the Equator. They calculated Si/N ratios < 1 east of 140”W in an “El Nino” situation and east of 170”W in non “El Nino” conditions, suggesting to be in minimal supply in these regions and most likely the limiting nutrient.

A more appropriate and relevant approach is based on the examination of the balance between growth and loss. The export of silica from the euphotic zone to deep water and the different pathways and rates for regeneration of nitrogen and silica contribute to the so- called “silica pump” (Dugdale et al., 1995). Based on modelling calculations these authors proposed that the main factor regulating the primary production offshore of the Peru upwelling may be silicic acid. Ku et al. (1995) reported the same idea in the Equatorial Pacific, where the availability of “new silicate” may limit the surface productivity. They

..

1270 S. Blain et al.

estimated upward silicic acid fluxes to the euphotic zone between 1.6 and 2.8 mmol Si m-' day-'. During the Flupac cruise drifting sediment traps were deployed. The detailed results will be reported elsewhere but the downward fluxes at 300 m at 150"W were between 0.35 and 0.85 mmol Si m-' day-'. During the cruise reported in this paper the upward Si flux was certainly not as high as reported by Ku, because the surface waters were partially isolated from the deep water because of a warm Kelvin wave propagating eastward. However during a &ed 7-day station at 150"W constant [BSi] and [Si(OH) .] were observed, suggesting that upward and downward Si fluxes were balanced. Although the measurements in October 1993 did not support the proposed Si-limitation by a "silica pump", such limitation could not be ruled out if particular events occurred: for example, a local diatom bloom initiated by shoaling of the thermocline as suspected by Bender and McPhaden (1990), or artificial iron-enrichment as performed in the mesoscale iron enrichment, experiments (Martin et al., 1994; Coale et al., 1996a).

THE CASE FOR IRON LIMITATION IN THE HNLC AREA

Both approaches discussed above indicate that diatom growth rates were considerably less than maximal in the HNLC region. The current data do not permit the limitation of the diatom growth rates by macro-nutrients to be ruled out. However, the evidence of diatom limitation is also consistent with the iron hypothesis (Martin, 1990). The diatoms from the central Pacific have the potential to grow faster if the lacking nutrient is added, as suggested from enclosed iron enrichment experiments (Takeda and Obata, 1995) or more clearly demonstrated in unenclosed experiments (Martin et al., 1994; Coale et al., 1996a). Studying the limitation of the growth rate of the Equatorial Pacific phytoplankton community by iron, Coale et al. (1996b) measured a saturation constant of 0.12 nM Fe and a pmax of 1 doubling day-'. This result is not in contradiction to the current finding that diatoms may grow faster if they are replete with iron, because their kinetic experiments were conducted with the whole phytoplankton community, which was dominated by nanoplankton. They calculated that the upwelled iron can support only 20% of the production required to draw down :the upwelled nitrate, and so could support a new primary production of 20 mmol C m-' day-'. Based on the Si-integrated primary production (4 f 1 mmol Si m-' day- ') reported here, the corresponding carbon production by diatoms was calculated. Using a Si:C ratio 0.13 (mole/mole), this comes to 31 f 8 mmol C m-' day-'. The two values are in good agreement.

CONTRIBUTION OF DIATOMS TO CARBON BIOMASS AND PRIMARY PRODUCTION

The contribution of diatoms to carbon biomass was estimated assuming Si:C of 0.13 f0.04 (Brzezinski, 1985). In order to compare with values previously reported in the literature for the HNLC area, the mean value of carbon biomass of diatoms in the upper 60m for stations #76, #78 and #SO was calculated. The results ranged between 7 and 22 pg C 1-' (10-35% of the total particulate organic carbon). The absolute estimation from this study is considerably higher than those deduced from microscopic biovolume determination and conversion to carbon biomass by other authors. Iriarte and Fryxell (1995) reported values of 0.83 and 2.18 pg C 1-'. The results from the current study are more compatible with the data of Buck and Chavez (1994), who reported a carbon biomass

Equatorial Pacific diatoms 1271

of 4.7 pg C I-' for diatoms associated with aggregates. The comparison of the different results is not easy, because it has now been recognized that the main factor responsible for the variability of phytoplankton biomass in the Equatorial Pacific is the variation in number and composition of the diatom population. However, the estimation-range reported here may be overestimated, because some detrital material or non-diatom species are included in the BSi measurements. In contrast the estimation of Iriarte and Fryxell (1995) reported above could be underestimated, as they did not take into account small diatoms ( < 20 pm). In the surface water of station #78, small Chaetoceros (10 pm) contributed 40% of the BSi.

In the same manner the contribution of diatoms to carbon primary production were calculated. The mean value (31 mm01 C m-' day-') is 34% of the whole carbon primary production in the HNLC area if one considers the mean carbon production value measured during the cruise, 92 & 3 mmol C m-' day- ' (Le Bouteiller, personal communication, 1996). This value is of the same order of magnitude as the mean integrated carbon production of 105 & 15 mmol C m-2 day-' reported by Murray et al. (1994).

It is also important to note that the estimated carbon production by diatoms is of the same order as the new primary production deduced from upwelled nitrate: 18 mmol C m-' day-' (Chavez and Barber, 1987) or 28 mmol C m-' day-' (Carr et al., 1995).

In oligotrophic water the contribution of diatoms to carbon biomass is low (< 6% of POC), except near the depth of the maximum of chlorophyll, where the mean value is 9% (range 5-1 8%) of the total particulate organic carbon. The mean integrated production of Si varied from 0.8 to 2.1 mmol Si m 3 d a y - l (equivalent to 6 to 16 mmol C m-' day-'), depending on the radiation availability. During the same cruise the integrated primary production of carbon in this area (166"E) was measured to be 25 and 75 mmol C m-' day- ' (A. Le Bouteiller personal communication, 1996). The carbon production associated with the diatom biomass near the chlorophyll maximum may represent up to 20% of the total primary production.

THE ANNUAL SI PRODUCTION IN THE EQUATORIAL PACIFIC: ESTIMATION AND COMPARISON

Table 3 summarises the integrated Si production rate in some oceanic regions. The mean daily silica production rate of 3.9 mm01 day-' in the HNLC area of the Equatorial Pacific allows a minimum estimate of the annual production of biogenic silica at this site: 1.4 mol Si m-' year-'. If this rate is representative of the HNLC area in the Equatorial Pacific (1.14 x lo6 km2) as defined by Wyrtki (1981), then the spatially integrated annual silica production in that region is 16 Tmol Si year-'. This estimate lies in the lower range reported for the Southern Ocean, another much larger, HNLC area (Nelson et al., 1995). This is a minimum estimate because:

(1) Vertically integrated production rates extended to at least the O. 1 % light depth. (2) The cruise took place during a warming event, and it is now well established that

primary production is greatly reduced during El Niño events (Murray et al., 1994; Murray, 1995)

(3) No particular bloom conditions were encountered during the cruise or were taken into account, although it was known that they could occur. Indeed, during Eqpac 1992 extremely high concentrations ( > 20 pg 1- of chlorophyll a) of buoyant diatoms

1272 - . , II

S. Blain et al. -

Table 3. Comparison of integratedproduction rates and biomass of biogenic silica in oligotrophic and HNLC arem

Zones Integratedproduetion BSi Surface Production m o l m-' mol m-' nmol I-' km2 Tmol year-' References

day- I year-'

Oligotrophic West Equatorial Pacific 0.8-2.1 0.5 10-40 4.8 x lo6 2.4 This study Sargasso Sea 0.2-1.6 0.2 <20 (a) 1.3 x lo8@) 26 Nelson and Brzezinski, in

press

HNLC: Pacific 3.9 1.4 1.1 x io7 16 This study Southern ocean 2.6-38 0.2-3.5 3.8 x io7 17-37 Leynaertkt al., 1993

World ocean 1.6-2.1 0.6-0.8 3.6 x lo8 200-280 Nelson et al., 1995 Quéguiner et al., in press

(a) Excepting blooming conditions ((BSi) = 580 nmol 1-I). (b) Surface of the mid-ocean gyres. (c) The expanse of

(Rhizosolenia sp.) were measured at 140"W, associated with an open-ocean front (Yoder et al., 1994). Assuming a C/Chla ratio of 60 and a Si/C ratio of O. 13 (Brzezinski, 1985) leads to a biogenic silica concentration > 10 pmoll-*, 60 times greater than the highest concentration encountered during the cruise reported here. As these diatoms cells were physiologically active and growing relatively fast (roughly one cell division per day), it is evident that such events could contribute a significant amount, or even most, of the annual silica production in that region.

As for the HNLC area, it is possible to estimate the annual production of biogenic silica in the oligotrophic part of the transect: 0.5 mol m-' year-'. To the authors' knowledge, the only other available data about biogenic production rate in oligotrophic areas are from the Sargasso Sea (Brzezinski and Nelson, 1996). The annual production reported here is about two times higher than the one they estimated. Several factors could explain that difference. The silicic acid concentration was higher in the western Equatorial Pacific (1.5 pM) than in the Sargasso Sea (0.6-0.9 pM), and the rate of silica production was found to be strongly limited by [Si(OH)4] in the Sargasso Sea. The temperature was also different, 30°C in the Pacific, 18-26°C in the Sargasso Sea.

CONCLUSION

In summary the comparison of diatom growth rates in the oligotrophic and HNLC Equatorial Pacific suggest that diatoms are limited in both regions. Nitrogen supply appears to be the limiting factor in the upper layer of the oligotrophic area. In the HNLC region the current results are consistent with the limitation by a micro nutrient such as iron. The contribution of diatoms to carbon production was estimated to be one-third of the total carbon production in the HNLC area and less than 20% near the deep maximum of chlorophyll in the oligotrophic waters. The estimates of annual production for HNLC and oligotrophic regions are respectively 1.4 mol m-' year-' and 0.5 mol m-' year-'.

Acknowledgements-The authors thank the officers and the crew of the research ship Atalante. The authors are indebted to Dave Nelson (Oregon State University on sabbatical stay in Brest) who offered his time, helpful advice

Equatorial Pacific diatoms 1273

and constructive criticisms to improve the manuscript. This work was supported by CNRS-INSU and ORSTOM. This is UMR CNRS 6539 contribution no. 97002.

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