MARINE ECOLOGY PROGRESS SERIES Vol. 133: 263-273, 1996 1 Published March 28 Mar Ecol Prog Ser

Role of spp. in the productivity of the subtropical North Pacific

Ricardo M. Letelierl.*, David M. ~arl~

'college of Oceanic and Atmospheric Sciences, Oregon State University, Corvallis, Oregon 97331-5503, USA 'School of Ocean and Earth Science and Technology, University of Hawaii. Honolulu, Hawaii 96822, USA

ABSTRACT.The concentrations of filamentous Trichodesmium spp., present as free tri- chomes and In colonial assemblages, were measured at approximately monthly intervals at Stn ALOHA (22"45'N, 158°00'W) between October 1989 and December 1992. The average abundance of filaments in the upper 45 m of the water column was highly variable ranging from 1.1 to 7.4 X 10"tri- chomes m-3 and from 0.02 to 1.4 X 102 colonies m-? Colonies were composed, on average, of 182 fila- ments accounting for 12% of total (free filament plus colonies) Trichodesmium biomass. Low densities of single trlchomes were associated with, but not restricted to, deep mixing events and winter periods. During 1991 and 1992 the concentration of Trichodesmium spp. present in the water column increased relative to the pre 1991 observations. This increase coincided with increases in photosynthetic carbon assimilation and in the molar ratio of N:P of suspended particulate matter in the upper 45 m of the water column. However, the change in Trlchodesmium biomass alone does not account for the change ob- served in autotrophic carbon assimilation and elemental biomass composition Tricliodesniium spp. comprised, on average, 18":~of the chlorophyll a, 4'' of the photosynthetic carbon assimilation, 1OCtbof the particulate nitrogen and 5% of the part~culatephosphorus. We also estimate that Trichodesmium dlnitrogen fixation accounted for, on average, at least 27'7, of the new production at this study site. These observations, combined with primary production experiments conducted on isolated colonies, suggest that production is enhanced due to the release of NH,+ and dissolved organic nitrogen by Trichodesmium spp. during episodes of .

KEYWORDS: Trichodesmium . Nitrogen fixation . Nutrients . Productivity . Time-series

INTRODUCTION Trichodesmium in marine ecosystems and the marine nitrogen cycle remains a matter of debate (Carpenter Spatial and temporal distributions of the diazotrophic 1983, Codispoti 1989, Capone et al. 1992). (nitrogen-fixing)cyanobacterium Trichodesmium have Field observations have shown that Trichodesmium been extensively studied in the eastern section of the colonles become abundant in surface waters of tropical tropical North Atlantic Ocean and Caribbean Sea (Car- and subtropical marine ecosystems during periods of penter & Price 1976, Carpenter & Romans 1991), in the calm weather (Steven & Glombitza 1972, Carpenter Indian Ocean (Devassy et al. 1978), in the East China & Price 1976, Bryceson & Fay 1981, Karl et al. 1992). Sea (Marumo & Asaoka 1974a), and in the Pacific These colonies of Trichodesmium cells support an Ocean (Marumo & Asaoka 1974b, Marumo & Naga- entire localized ecosystem including epiphytic hetero- sawa 1976). The presence of Trichodesmium has also trophic bacterla (Carpenter & Price 1976, Borstad & been frequently reported in oligotrophic marine envi- Borstad 1977, Paerl et al. 1989), (Borstad & ronments in the context of phytoplankton bloom phe- Borstad 1977, Bryceson & Fay 1981), and hydrozoans nomena (Bowman & Lancaster 1965, Dupuoy et al. (Geiselman 1977, Borstad & Brinckmann-Voss 1979). 1988, Karl et al. 1992).Nevertheless, the importance of There is also evidence that the survival rate of larval stages of the Macrosetella gracilis (Dana) increases when Trichodesmium is present (Bjornberg 'E-mail: [email protected] edu 1965, Calef & Grice 1966, Roman 1978). Tnchodesmium

O Inter-Research 1996 Resale of fr~llarticle not per~nitled Mar Ecol Prog Ser 133: 263-273, 1996

also undergoes rapid lysis when senescent (Van Baalen bution of Trichodesmium in the euphotic zone of & Brown 1969, Borstad & Borstad 1984). Devassy et Stn ALOHA between August 1989 and December al. (1979) concluded from the succession following a 1992. We also analyze the variability of Trjchodesmium Trichodesmium bloom that rapid cell structure degra- biomass in the upper 45 m of the euphotic zone as it dation resulted in an increase of inorganic nutr~entsin relates to changes in the elemental ratio of particulate the water column, presumably the result of a rapid and matter and primary product~on over similar depth efficient microbial recycling of organic constituents ranges. following cell lysis. Furthermore, recent studies indi- cate that, even under healthy conditions, Trichodes- mium releases NH,' (Prufert-Bebout et al. 1993) and METHODS dissolved organic nitrogen (Capone et al. 1994, Glibert & Bronk 1994) during the process of nitrogen fixation. The Hawaiian Ocean Time-series (HOT) program These observations suggest that fluctuations in the investigators sample the water column of Stn ALOHA abundance of Trichodesmium in the pelagic marine at approximately monthly intervals as part of the environment may affect both the taxonomic and chem- United States-Joint Global Ocean Flux Study (US- ical composition of the ambient plankton community. JGOFS; Karl & Winn 1991, Karl & Lukas in press). As In the subtropical North Pacific gyre, where nltrogen is part of this program, water column suspended particu- generally considered to be the production rate lim~ting late carbon (PC),particulate nitrogen (PN),particulate nutr~entcontrolling productivity, nitrogen fixation and phosphorus (PP),and p~gmenlswere measured during its release In the form of ammonium, or other utilizable each cruise. Water samples (4 to 10 l), collected with a forms of nitrogen, may be an important source of 'new' 24-position rosette water sampler equipped with 12 1 nutrient for phytoplankton otherwise incapable of polyvinylchloride (PVC) bottles and SeaBird CTD, direct assimilation of gaseous nitrogen. wele pie-screeiie6 through ci 202 pm mesh and fi!tered Most field studies of Trichodesmium have concen- onto combusted 25 mm GF/F Whatman filters for PC trated on either the spherical or fusiform colony mor- and PN analyses, or onto combusted and acid washed phologies of this in spite of the fact that Marumo 25 mm GF/F filters for PP analyses. PC and PN were & Asaoka (1974a) and Marumo & Nagasawa (1976) measured simultaneously in a Perkin-Elmer model have reported that a major fraction of Trichodesmium 2400 CHN analyzer. PP was quantified spectrophoto- biomass occurs in the form of single trichomes. Histor- metrically. For this analysis samples were combusted ically, single trlchome morphology h.as been consid- (450 to 500°C), acidified (0.5 N HC1 at 90°C for 90 min) ered less important in the context the marine nitrogen and color proportional to the concentration of phos- cycle because it was thought that nitrogen fixatlon phorus in solution was developed by adding a mixed occurred only in sub-oxic microenvironments found in reagent freshly prepared (10 m1 ammonium molyb- the center of selected colonies (Carpenter & Price 1976, date, 25 m1 5 M sulfuric acid, 10 m1 5.4 % ascorbic Bryceson & Fay 1981, Paerl & Bland 1982, Paerl & acid and 5 m1 potassium antimony tartrate; Strickland Prufert 1987, Paerl & Bebout 1988, but see Saino & & Parsons 1972) Particulate samples for pigment Hattori 1982). This view has recently been challenged analyses were collected and quantified using the as a result of studies of nitrogen fixation performed in chromatographic method described by Letelier et a1 Trichodesmium cultures (Ohki & Fujita 1988, Prufert- (1993). Bebout et al. 1993) and by enzymatic analyses of In order to assess the temporal and vertical distribu- natural Trichodesmium assemblages (Capone et al. tions of Trichodesmium colonies, horizontal net tows of 1990, Carpenter et al. 1990, Bergman & Carpenter discrete subsurface layers were obtained using an 1991). This evolu.tion of thinking about the potential opening-closing plankton net (0m.ori 1965; 50 cm in role of free trichomes in nutrient cycles has prompted mouth diameter, 270 cm in filtering cloth length and renewed speculation about the quantitative Impor- 100 pm in mesh size, 9 cm bucket diameter with 60 pm tance of oceanic nitrogen fixation processes. mesh size) between March 1990 and October 1991. The Hawaiian Ocean Time-series (HOT) program is When not counted at sea, samples were preserved in designed to characterize oligotrophic habitat and 5 '!<, buffered formalin and analyzed at our shore-based ecosystem variability in the water column at Stn laboratories. A flow meter (General Oceanics) was ALOHA (A Long-term Oligotrophic Habitat Assess- mounted at the mouth of the net to calculate the total ment, 22" 45' N, 158O00' W). This program provides an volume of water filtered during each tow and convert optimum background data set to study the effects that colony numbers to densities. A typical horizontal tow changes in Trichodesmium abundances may have in sampled approximately 4.3 m3 min-' and lasted 10 min. the biochemical characteristics of the euphotic zone. In Vertical net tows (0 to 100 m; 16 m3 total) were also per- this paper we report results of the study of the distri- formed between. May 1990 and October 1991 to com- Letelier & Karl: Trjchodesmium role in productivity

pute an independent estimate of the depth-integrated scope to select only those that appeared to be intact concentration of colonies. and free of . Once the regular primary pro- To assess the elemental (PC, PN, PP) and pigment duction experiment had been deployed (see Letelier et compositions of Trichodesmium at Stn ALOHA, colo- al. in press), the remaining water was used to fill tripli- nies from net tow samples were isolated using a cate 500 m1 polycarbonate incubation bottles for each Pasteur pipette, rinsed and vortexed in 50 m1 filtered water depth. One Trichodesmium colony was added to seawater to produce a suspension of single trichomes. each bottle. This amounted to an increase in chloro- Subsamples were transferred onto combusted GF/F phyll a (chl a) of approximately 100%. Triplicate poly- filters, as above. Triplicate samples (>l0 colonies per carbonate bottles containing filtered 0.2 pm sterile fil- filter) were analyzed for each determination. tered seawater were also inoculated with 3 colonies To examine the natural distribution of single tri- each. NaHI4CO3 (final activity = 0.1 pCi ml-') was used chomes, seawater was collected at discrete depths as radiotracer to measure carbon assimilation. The using the rosette water sampler described above. samples were incubated on deck from approximately Water samples (4 to l0 l from each depth) were gently dawn to dusk under in situ simulated light and temper- filtered onto Nitex mesh (10 pm mesh size). The mesh ature conditions. was back rinsed with 50 m1 filtered seawater to resus- To correlate Trichodesnlium abundances measured pend the trichomes after manual removal of colonies, if during each cruise with antecedent weather condi- present. This suspension was then filtered onto 0.8 pm tions, the sea surface temperatures (SST) recorded at 25 mm Nuclepore filters prestained with irgalan black hourly intervals by the National Data Buoy Center and mounted on microscope slides. Trichomes were (NDBC) buoy no. 51001 (23" 24' N, 162" 18' W) were enumerated by epifluorescence microscopy based on analyzed. From these hourly averages, we calculated a autofluorescence. To avoid overestimat- relative stability index based on the amplitude of the ing the quantity of single trichomes in a sample as a diel SST cycle. This approach assumes that the diel result of filament breakage during processing, only SST warming cycle, due to the local heating and cool- apical cells were enumerated rather than total tri- ing of the sea surface, increases in amplitude under chomes (intact filaments have shaped apical cells at calm weather conditions as a direct result of a decrease either end). The abundance of single trichomes re- in turbulent mixing. Hence, a large daily amplitude in ported here corresponds to the number of apical cells the diel SST cycle indicates stratification of the upper divided by 2. water column, while a low amplitude indicates a well- The elemental (PC. PN, PP) composition of single tri- mixed water column (Karl et al. 1995). chomes was studied on 2 separate occasions. A total of 8 replicates of l0 l each collected at 45 m depth were screened onto 10 pm mesh as described above. All RESULTS replicates were combined into a single 500 m1 pooled sample from which five 100 m1 aliquots were drawn. The estimated densities of Trichodesmium colonies One of the samples was used for microscopic enumer- and single trichomes at Stn ALOHA are highly vari- ation, 1 for PP, 1 for pigment and 2 for PC/PN. able (Fig. 1). The concentration of single trichomes The abundance of trichomes found in colonial mor- averaged over the upper 45 m of the water column phology was compared to the abundance of single tri- ranges from 1.1 to 8.4 X 104 filaments m-3 (F = 4.6 X chomes by counting the number of filaments per 104 filaments m-3, s = 2.3 X 104 filaments m-", n = 22; colony. Triplicate 60 m1 samples of filtered seawater Fig. 2B). Colony densities averaged over the same containing 20 to 30 hand-picked colonies per sample depth intervals ranged between 2 and 140 colonies m-3 were vortexed until no visible colonies remained in the (7 = 31 colonies m-" S = 41 colonies m-3, n = 11; Fig. 2B). suspension (5 to 30 min). From each sample, triplicate Colony size also varied considerably in single samples 5 m1 subsamples were withdrawn and filaments were (250 cells tnchome-l), the average size for both desmium colonies to photoautotrophic production at morphologies (single trichomes and colonies) was Stn ALOHA. Colonies were collected at night from 5, similar (approximately 100 cells trichome-l). Tricho- 25, 45 and 75 m and transferred into filtered seawater desmium cells in colony morphology, therefore, com- corresponding to the respective sampling depth. All prised 12% (S = 11 %, n = 11) of the total Trichodes- colonies were observed under a stereoscopic micro- mium biomass. Mar Ecol Prog Ser 133: 263-273, 1.996

Colonies m'3 The slope of a linear regression (model I1 0 40 80 120 160 200 geometric mean) between integrated colony 0 concentration calculated from 0 to 100 m vertical net tows and from samples collected at discrete depths (horizontal net tows) was not different from 1.0 (95% confidence limits = 0.84 and 1.06, R2= 0.85, n = 9; Fig. 3) indi- cating that our sampling procedures were accurate. This result suggests that the recon- struction of the depth distribution of colonies i from discrete depth samples is a valid rep- resentation of the vertical structure of the colony distribution for the water column. The vertical distribution of both the fila- October through Marc October through March ment and colony morphologies displays a B subsurface maximum located between 20 and 50 m in all cruises where the mixed-laver 0 depth is c50 m (Fig. l). However, this sub- surface maximum disappears when the mixed-layer deepens and exceeds a depth of 40 50 m. A near surface maxlmum of single tri- chomes (2 to 5 m) is also observed in 40% of S 80 the cruises and, although an accumulation of S colonies at the sea surface was visually 6 a observed from the ship on 2 occasions 120 (August 1991 and June 1992), this feature was not visible at the time of sampling and was not resolved by the sampling procedures 160 160 employed. April through Septembe April through September The abundance of single trichomes in the first 45 m of the water column does not cor- 200 200 relate with the average die1 SST amplitude

Fig. 1. Trichodesmirlrn spp. Vertical distribution of single trichomes calculated from the NDBC buoy no. 51001. (A, C) and colonies (B, D) at Stn ALOHA between October 1989 and Several running averages, from l4 December l992 intervals, were performed to assess the cor-

Buoy no 51001

Fig. 2. Tnchodesrnium spp. Temporal variation of (A) diel amplitude of sea surface temperature (SST) measured at 23" 24' N, 163" 18' W (see 'Methods') and (B)average densities of single trichomes (A) and colonies (e) in the upper 45 m of OJAJ 0 JAJO JA JO J A JO the water column at Stn ALOHA. 0: October; J. January or July; A: April 1989 1990 1991 1992 Letelier & Karl: Trichodesmium role in productivity

Table 1. Trichodesmium spp. Elemental composition and chlorophyll a (chl a) content of single trichomes and colonies [mean (iSD)]. PC: particulate carbon; PN: particulate nitro- gen; PP. particulate phosphorus

Parameter Single Colonies" hday 90 Oct 91

Chl a (ng filament- PC (ng filament-') PN (ng filament-') PP (ng filament-') PC:chl a (w:wl 0-100 m mean colony abundancc esrimated from horizonralnet tows, coloniesm-> Fig. 3. Trichodesmiurn spp. Comparison between the mean "Assumes colonies composed of 182 filaments abundance colonies over the upper 100 m of the water column estimated from 0 to 100 m vertical net tows and from horizontal net tows performed at discrete depths in our samples [

Table 2. Elemental ratio of 0 to 45 m depth integrated particulate matter at Stn ALOHA and comparison of the median elemental ratio with the theoretical Redfield ratio of 106C:16N:lP

Elemental Period Mean Median Significance ratio (mol:mol) (mo1:mol) levela

Time

Fig. 4. Trichodesmium spp. Temporal variation of photosyn- thetic carbon assimilation (0) integrated over the upper 45 m depth and average density of single trichomes (m) over the "Based on Wilcoxon's ranks test same depth range Mar Ecol Prog Ser 133:263-273, 1996

Relativecontribution (%) bility of overestimating single tri.chome and underesti- 0 2 4 6 8 10 12 0 10 20 30 40 50 mating colony densities as a result of the mechanical disruption of colonies exists. However, the slope of the -* -f--H linear regression between integrated colony concen- tration calculated from vertical net tows and from sam- ples collected at discrete depths (horizontal net tows) was not different from 1.0 (95% confidence limits = 0.84 and 1.06, R* = 0.85, n = 9; Fig. 3). Because, on m average, each horizontal net tow filtered 43 m3 and I vertical net tows only filtered 16 m3 this result suggests that the percentage of colonies disrupted is not a func-

*Carbon asslmilalan tion of the volume filtered by the net. Given the aver- mChlorophyll a age abundance of colonies estimated for the upper 1 50 m of the water column (X = 31 colonies m-3), the March 1990 t August 1991 complete disruption of Trichodesmium bundles would account for less than 14% of the estimated average Fig. 5. Tnchodesmium spp. Depth distribution of the contribu- abundance of single trichomes. Furthermore, because tion of Trichodesmium spp., as percentage, to total photo- colonies are probably subjected to a higher stress synthetic carbon assimliatlon (ej and total chl a (m) during a net tow than during a rosette water sampling procedure, we bel.ieve that our calculations of single Wilcoxon's signed rank test analyses, the phosphorus trichome densities are not significantly affected by content in the suspended particulate matter for the colony disruption. per1.od 1991-1992 is on average significantly lower Carpenter & Romans (1991) calculated from histori- than the Redfield ratio (Redfield et al. 1963). cal data that Trichodesmium colonies contribute 60 % Even though chl a contributed by Tnchodesmlum of the total chl a measured in small sample volumes may be as high as 40% of the tota.1 chl a measured at a (200 to 300 ml) collected in the upper water column of discrete depth (Fig. 5),integrated values from 0 to 45 m the western North Atlantic Ocean. This high contribu- account for no more than 28% (X= 18%, s = 8%, n = tion indicates that most of the Trichodesmium biomass 22). Carbon assimilation contribution is always lower in the western North Atlantic Ocean must be i.n the than the chl a contribution (Fig. 5) suggesting that colonial morphology. Free trichomes were not enumer- the carbon assimilation efficiency of Trichodesmium ated in that study. A marked difference in morpho- is lower than the assimilation efficiency of the phyto- logical composition and in the difference of the esti- plankton community as a whole. mated contribution of Trichodesmium to organic matter (Table 3) suggests that there may be significant ecological differences between these 2 marine envi- DISCUSSION ronments. This conclusion precludes the possibility of freely extrapolating results from one ocean basin to The only report available on the spatial distribution another. of single filaments of Trichodesmium (trichomes) in the Although there is no significant correlation between water column derives from a study in the Central and calm weather periods, as distinguished by the die1 North Pacific Ocean. On a transect from 50" N to 10" S along 155" W, Marumo & Asaoka (1974b) found large Table 3. Trichodesmium spp. Temporal variation in the con- concentrations of filaments (>5 X 104 trichomes m-3) tribution of slngle trichomes to total suspended particulate restricted to the upper 50 m of the subtropical gyre carbon (PC), particulate nitrogen (PN), particulate phospho- (from 15"N to 29"N). These concentrations are higher, rus (PP), and chlorophyll a (Chl a] in the upper 45 m of the but not significantly different from the average abun- euphotic zone at Stn ALOHA dance of single trichomes found in the upper water column of Stn ALOHA (X= 4.6 X 104 trichomes m-3, Parameter 1989-1992 1989-1990 1991-1992 s = 2.3 X 104trichomes m-3, n = 22). By comparing these (X) (%l (Oh) Trichodesmiurn concentra.tions with the abundance of PC 9.2 * 3.4" 5.8 * 2.7 11.3* 3.3 cells in, the colony morphology found at Stn ALOHA, PN 10.8 k 4.5 7.2 k 2.9 13.3 t 3.7 we conclude tha.t colonies comprise a minor fraction PP 4.6 * 2.4 2.2 * 1.1 5.3 * 1.9 (12%) of the Trichodesmium population in our study Chl a 13.4* 8.1 8.1* 6.4 17.1* 7.0 Due to the methodologies used to collect single "Mean ~t 1 SD of the mean trichomes and colonies in the present study, the possi- Letelier & Karl: Trichodesn:rluln role in product~vlty 269

amplitude of the SST at 23"24'N, 162" 18'W, and the duction of particulate matter is close to C:N Redfield abundance of single trichomes in the upper 45 m of the ratio, the estimates of particulate exported production euphotic zone at Stn ALOHA, low Trichodesmium den- at 150 m [downward particulate flux X (total produc- sities appear consistently during deep mixing events tion)-'] range from 0.02 to 0.12 between 1989 and 1992 and winter months. Nevertheless, low concentrations (x= 0.06, s = 0.02, n = 39). observed during May to June 1990 and October 1991 Field incubations (Mague et al. 1977), as well as the do not appear to be the result of deep mixing events. nitrogen isotopic signature of Trichode~~ium(615N = These observations suggest that environmental factors 1.7to +0.5 ppm; Wada & Hattori 1976) indicate that other than wind driven events and sea surface tern- nitrogen fixation accounts for virtually all cellular peratures, perhaps cloud cover, solar irradiance and nitrogen required for this organism to grow. Because grazing pressure, may play an important role in con- the N:C ratio observed for Trichodesmium is not sig- trolling Trichodesn~ium distribution and abundance on nificantly different from the Redfield ratio, we can use short tlme scales (days). carbon assimilation as an estimate of nitrogen flxatlon. On larger time scales, the Interannual variability in If the values of carbon assimilated by Trichodesrnium Trichodesmium abundance observed in the upper 45 m measured during March 1990 and August 1991 are of the water column during 1991 to 1992 may be the representative of its contribution to production at Stn result of a decrease in the frequency of deep mixing ALOHA, then Trichodesn~ium accounts, on average, events (Karl et al. 1995). Extended periods with a for 27 O/oof new production (21 % during 1989 to 1990 shallow mixed-layer wlll benefit phytoplankton such and 35 '70 dunng 1991 to 1992). However, this estlmate as Trichodesn~ium that have the capacity to regulate should be viewed as a lower boundary value because it their buoyancy (Fogg 1982, Komopka 1984. Karl et does not take into account either bloom events (Karl et al. 1992). al. 1992) or the potential release of NH', and dissolved The elemental composition of Trichodesmium is sur- organic nitrogen (DON) by Trichodesmium during prisingly constant when comparing single trichomes nltrogen fixation (see below). and colonles (Table 1).The PC:PN molar ratio (6.3:l)is The highest carbon assimilation observed in the first close to the predicted Redfield ratio (6.6:l) but the 4 yr of the HOT program occurred during August 1989. PN:PP (45:l) is 3 times greater than Redfield stoichio- Although an increase in the abundance of Trichodes- metry (16:l).These results are consistent with observa- mium was observed at the time of this primary produc- tions reported in previous studies (Mague et al. 1977, tivity experiment, a large bloom did occur 3 d later at McCarthy & Carpenter 1979, Lewis et al. 1988, Karl et Stn ALOHA (Karl et al. 1992) This observation sug- al. 1992, 1995). A low phosphorus cell quota appears gests that trichomes were already abundant at the time as a characteristic of Trichodesmium cells whether in of the incubatlons. However, there is no evidence of an free trichome or colony morphology. Furthermore, the increase in phytoplankton biomass when comparing chemical analysis of actively photosynthetic colonies the concentration of chl a measured from the primary collected during a bloom event in the vlcinlty of Stn productivity cast with values obtained in either previ- ALOHA yielded a PN:PP ratio of 125:l (Karl et al. ous or subsequent months (Fig. 6A). The only strong 1992), suggesting a large flexibility in the cell quota evidence suggesting that a change occurred in the phosphorus in Trichodesmium. water column was fo.und in the carbon assimilation The contribution to the integrated photosynthetic efficiency of the phytoplankton (Pe) and the C:N ratio carbon assimilation by Trichodesmium during March of particulate matter. The carbon assimilation efficiency 1990 and May 1991 in the upper 45 m of the water increased by a factor of 2 in the upper 50 m the water column at Stn ALOHA was estimated to be 3.9 and column (Fig. 6B) while the C:N ratio decreased from

4.3 O/uof the total production, respectively (Fig. 5). Al- 7.3:l to 6.3:l (mo1:mol; Fig. 6C). Assuming that the though the primary production below 45 m accounted Redfield ratio for phytoplankton is 6.6:1,the shift in the for 65 and 43% of the 0 to 200 m integrated produc- particulate elemental ratio suggests that phytoplank- tion, the contribution by Trichodesmium was negligible ton were not nitrogen limited during August 1989. below 75 m. Based on these results, the estimated con- Trichodesmium carbon assimilation number (PE)re- tribution of Trjchodesmiu~n to total inorganic carbon ported in the literature and calculated for Stn ALOHA assimilation was approximately 1.7% durlng March over the period of this study are relatively low, ranging 1990 and 2.5% during August 1991 from 0.17 to 3.8 g C (g chl a)-' h-' (Carpenter & Price The fraction of autotrophic production attributed to 1977, Mague et al. 1977, Lewis et al. 1988, Karl et al. Trichodesmium does not appear to be important unless 1992; Fig. 7).Hence, the increase in PBobserved in Au- we conslder this contribution in terms of new produc- gust 1989 and during 1991-1992 cannot be explained tion. Based on th.e downward flux of PN measured at by an increase of the contribut~onof Trichodesmium to Stn ALOHA and assuming that photoautotrophic pro- the total p:roduction in the euphotic zone. 270 Mar Ecol Prog Ser 133: 263-273, 1996

mg Chl a m-3 g C (g Chl a)-' h-' PC:PN (mol:mol) 0.0 0.1 0.2 0.3 0.4 0 2 4 6 8 10 12 14 4 5 6 7 8 9 10 11

Fig. 6. Depth prof~lesof (A) chl a concentrations, (B)phytoplankton asslm- ilation effic~ency, and (C) molar ratlo of carbon to nitrogen In the sus- pended particulate matter measured in samples col- lected at Stn ALOHA be- tween July and November 1989. (m)July; (0) August; (m)September; (A) Novem- ber samples

However, 2 lines of evidence suggest the existence Nevertheless, we need to be careful with the inter- of a synerglstic eiiect between Tricnoaesmium and pretation of nlsroricai vaiues of "r given ior Tricho- other algae present in the water column. The first desmium, and with the results of the experiment dis- observation is derived from the primary productivity cussed above. Assimilation numbers in Tnchodesmium experiments performed during March 1990 and have been calculated only by using colonial morpho- August 1991 (Fig. 7). Jn both cases the incubations at logies where tnchomes are subject to self shading 25 and 45 m showed that the P' of samples containing effects. In the samples collected during this study, only 1 colony was higher than the calculated pB based on chl a concentration per unit biomass (PC) increases in results from the independent incubation of colonies colonies when compared to single trichomes (Table 1). and phytoplankton. This effect was not evident at 5 m Although the increase in this ratio is not significant depth and may have been inhibited by high light con- due to the variability associated with measurement in ditions (Li et al. 1980, Lewis et al. 1988). colonies, it suggests that some degree of photoadapta- If we consider the supply of nitrogen to be limiting the algal production in the upper euphotic zone it is C possible to speculate that a release of NH,' and dis- g C (g C~Ia)-' h-' g (g C~Ia)-' h-' solved organic matter rich in DON during nitrogen 0 123 4 5 6 7 8 01234567891011 fixation (Prufert-Bebout et al. 1993, Glibert & Bronk 1111111111111111111, 1994) will enhance the production of phytoplankton other than Trichodesmiurn. Under high irradiance (the 5 m depth sample) the production of oxygen due to could be inhibiting nitroge- nase activity thus suppressing nitrogen fixation. -E Mchodesmium colonies have high respiration rates (Kana 1992), and the light intensity of the compen- o5 sation point for oxygen evolution in these colonies is approximately 300 pE m S ' (or 13 E m-' d-l) cor- responding to the irradiance measured at 30 m 150 -'D depth at Stn ALOHA. One of the metabolic benefits of a high respiration rate IS the protection of nitro- March 1990 August 1991 genase activity by reducing the lntracellular oxygen 200 tension during daylight hours. From this perspec- Fig. 7. Trichodesmiumspp. Depth distribution of assirn~lat~oneffl- tivel located pE trichomes near the 300 m-2 S-' ciency measured s~multaneouslyIn culonles (A),ndtural phyto- isolume may have higher activity than plankton assemblagcs (m),and natural phytoplankton assemblages those located closer to the sea surface. amended wth colomes (m).Error bars represent standard devidtlons Lvteller & Karl. Ti.~chodesrniurnrole in productivity

tion may occur in colonies as a result of self shadlng. Table 4 Temporal variation In the elemental ratio of sus- This observation is also consistent with the decrease in pended particulate matter after removing the contribution of PC:chl a observed during bloom conditions (Karl et al. single tr~chomesIn the upper 45 m of the euphotic zone at Stn ALOHA 1992). However, the relative decrease in PC:chl a in colonies found under non-bloom conditions during this Elemental 1989-1992 1989-1990 1991-1992 study suggests that photoadaptatlon between different rat10 (mol mol) (mo1:mol) (mol.mol) morphologies is not strong and may account for no more than a 20 % increase of PB between colonies and PC:PN single tnchomes (Table 1). PN:PP 15 3 13.5 16 6 The possibility that handhng the colonies in the preparation of prlmary production experiments may have disrupted microenvironments (Carpenter & Price the remaining bulk particulate matter. This residual 1976, Martinez et al. 1983) and damaged trichomes PN:PP of the particulate organic matter in the upper cannot be excluded (but see Carpenter et al. 1987 and 45 m of the water column, excluding Trichodesmium, Scranton et al. 1987). Although colonies inspected varies from 13.5:l (mo1:mol) in 1989-1990 to 16.6:l under a microscope before the inoculat~ons did not (mo1:mol) in 1991-1992 (Table 4) suggesting that nitro- appear broken, 20% of the incubation bottles at the gen IS not a limiting nutrient during the latter period end of the incubation period had some single trl- of the present study. chomes. If damage occurred in the colonies the low P' Two different processes related to the presence of value calculated for colonies must be considered an Trichodesmium may be taking place in the euphotic artifact of the experimental manipulations. zone. The first, discussed above, is the release of NH,' A second line of evidence suggesting that Tricho- and DON during the process of nitrogen fixation. The desmiunl is a source of nitrogen for other phytoplank- second process IS the release of other essential nutri- ters comes from the obsel-ved shift in elemental ratio ents during the senescence of tnchomes. Evidence in of the particulate matter and its correlation with the the literature indicates that Trichodesmium degrades abundance of trichomes in the upper 45 in of the water very fast (hours to days) upon reaching senescence column (Fig. 8). Sixty percent of the variability in (Van Baalen & Brown 1969, Borstad & Borstad 1984). If PN:PP may be attributed to variations in the abun- this is the case at Stn ALOHA, a decrease in the con- dance of Trichodesmium. Nevertheless, an increase centration of trichomes between cruises could result in in the percentage contribution of Trichodesmium bio- the release of nutrients, such as phosphorous, to other mass to the total suspended particulate matter is not phytoplankton taxa. This interpretation is consistent sufficient to explain the change in the elemental ratio with the observed trend of photosynthetic carbon assi- of the suspended particulate matter (Table 2). milation and Trichodesmium densities at Stn ALOHA. By knowing the approximate contribution of carbon, The increase in abundance of trichomes in the upper nitrogen, and phosphorus in Trichodesmium (Table 3), water column during 1991 to 1992 coincides with an In- it is possible to calculate the elemental composition of crease of photosynthetic carbon assimilation although, on smaller temporal scales (weeks to month), primary production appears to be inversely correlated to Tricho- desmium abundances (Fig. 4). The increase in carbon assimilation rates, following a decrease in trichome abundance between cruises, may indicate that the release of nutrients as a result of Trichodesmium senescence is an important ecological process, even under non-bloom conditions. Karl et a1 (1995) have suggested that the biogeo- chemical shift observed between 1989 to 1990 and 1991 to 1992 may be a result of an increase of the

5 1 stratification in the water column dul-lng an El Nino- 0 20 40 60 80 100 Southern Oscillatlon (ENSO) event. If the coupling Tr~chomesI-' between E-ichodesmium abundance, water column Fig 8 Tnchodesmiun~spp Molar ratio of 0 to 45 m depth stratlfication, and ENSO periods proves to be correct, integrated nitrogen (PN)to phosphorus (PP) concentlatlon in we should expect to observe a relaxation of nitrogen suspended matter plotted against thc average concentrat~on deficiency of the pelagic ecosystem of the North Pacific of single tr~chomesover the same dt pth range A model I1 geometric mean hear regression was used to model the subtropical gyre durlng ENSO events. This relaxation relat~onbetween both parameters would result from the combined effect of the reduction 272 Mar Ecol Prog Ser 133: 263-273, 1996

of nutrient inputs due to deep mixing events and the (ed) Cooperative investigations of the Canbbean and increase of nitrogen fixation by Trichodesmium and adjacent regions. FAO, Rome, p 51-57 other . Borstad CA,Borstad LE (1984) Needle-like projectiles ob- served issuing from lysing Trichodesmium (Cyanophyta) In conclusion, the observed change in PN:PP in the cells. Limnol Oceanogr 29:1097-1099 upper 45 m of the water column at Stn ALOHA Borstad GA, Brinckmann-Voss A (1979) On Pelag~anatricho- appears to be the result of a decrease in the frequency desmjae n, gen , family Pandeidae, (Anthomedusae/Athe- of deep mixing events. Because of the capacity to reg- catae, Cnidana), a new hydrozoan associated with the planktonic cyanophyte . Can ulate its buoyancy to fix NZ, Trichodesmium may play a J Zool 57:1232-1237 significant role in the input of nutrients when the Bo~rmanTE, Lancaster LJ (1965) A bloom of the plankton~c euphotic zone water column is stratified for extended blue-green alga in the Tonga periods of time. Furthermore, an over production of Islands. Limnol Oceanogr 10:291-293 reduced nitrogen and the concomitant extracellular Bryceson I, Fay P (1981) Nitrogen fixation in Oscillatoria (Trichodesmium) erythraea in relation to bundle formation release of ammonium and DON by Trichodesmium and trichome differentiation. Mar Biol 16:159-166 may explain the relative enrichment in PN observed at Calef GW. Grice GD (1966) Relationship between the blue- Stn ALOHA during 1991 and 1992. green alga Trichodesmium thiebautii and the copepod Perhaps because the focus of research on oceanic Macrosetella gracil~sin the plankton off north-eastern South America Ecology 47:855-856 photoautotrophic production was historically influ- Capone DG, Ferrier MD, Carpenter 'EJ (1994) Amino acid enced by problems related to the management of cycling in colonies of the planktonic marine cyanobac- fisheries (Ryther 1969), the contribution of nitrogen tenum Tricnoaesmiu~~~liliebauti~. Appi ~i;viiofi Microbic! fixation to total production estimated for the North 60:3989-3995 Pacific subtropical gyre may have been considered of Capone DG, O'Nell JM, Zehr J, Carpenter EJ (1990) Basis for die1 variation in nitrogenase activity in the marine plank- little importance. However, the contemporaneous con- tonic cyanobacterium E-lchodesmium thiebautii. Appl cepts of new production and exported biogenic nidiiel environ iviic~ub~u;56.2532-3535 from the euphotlc zone have become central in the Capone DG, Reuter JR,Carpenter EJ (1992) Overvlew of the analysis of biogeochemical cycles in the ocean during ad~dncedresearch workshop on bloom-forming marine . In: Carpenter EJ, Capone DG, Reuter JG the past decade (Longhurst 1991). This is the result of (eds) Vanne pelagic cyanobacteria: Trichodesmium and our interest and necessity to estimate the role of the other diazotrophs. Kluwer Academic Publishers, Boston, In global carbon cycles. In this context, results p 1-8 obtained during the first 4 yr of the HOT-series pro- Carpenter EJ (1983) Physiology and ecology of the marine gram suggest that the contribution of nitrogen fixation plankton Oscillatoria (Trichodesrnium). Mar Biol Lett 4: 69-85 by Trichodesmium in the North Pacific subtropical Carpenter EJ, Chang J, Cottrell M, Schubauer J, Paerl HW, gyre may provide a significant fraction of the nitrogen Bebout BM, Capone DG (1990) Re-evaluation of nitro- supporting new production. genase oxygen-protective mechanisms in the planktonic manne cyanobacterium Trichodesmium. Mar Ecol Prog Ser 65.151-158 Acknowledgements. We thank the captains, crews and scien- Carpenter EJ. Price CC IV (1976) Marine Oscillatona (Tricho- tists participating in the HOT program for assistance in the desmium): explanation for aerobic nitrogen fixation with- collection and analysis of samples. We also thank Roger out . Science 191:1278-1280 Lukas and the HOT-WOCE team for the generation of hydro- Carpenter EJ, Price CC TV (1977) Xitrogen fix~~l~on,distribu- graphic data, the National Oceanic and Atmospheric Admin- tion and production of Osdllator~a(Tr~chodcsmiurn) thie- istration-National Data Buoy Center (NOAA-NDBC) for bautii in the eastern Sargasso Sea. Limnol Oceanogr 20: access to the ocean data recorded by buoy #51001, and E. J. 381-401 Carpenter and 2 anonymous reviewer for helpful comments Carpenter EJ, Romans K (1991) Major role of the cyanobac- and suggestions. The present study was supported by terium Trlchodesmium in ndtrient cycling in the North National Science Foundation grants OCE 88-00329. OCE Atlantic Ocean. Science 254.1356-1358 90-16090, and OCE 93-01368 (awarded to D.K.). and OCE Carpenter EJ, Scranton MI, Novelli PC, Michaels A (1987) 87-17195 and OCE 93-03094 (awarded to R. Lukas). 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This article was presented by B. and E. Sherr (Senior Manuscript first received: A4arch 8, 1995 Editorial Advisors), Corvallis, Oregon, L'S. \ Revised version accepted: September 27, 1995