CHEMISTRY AND THE AQUARIUM by RANDY HOLMES-FARLEY

Iron: A Look at Organisms Other than Macroalgae   

In my article two months ago, I discussed iron in reef tanks.  Much of that article was directed toward the effect that iron has on macroalgae, and in that article I concluded that it was generally positive.  Of course, macroalgae are not the only things in our tanks, and if iron is having an effect on other organisms, it is important to understand it.  Some hobbyists, for example, have suggested that iron may “supercharge” cyanobacteria, while others have suggested just the opposite.  It has also been suggested that iron supplements may actually be “toxic” to corals when dosed to a reef tank.   In looking at the literature, it would appear that diatoms and other microorganisms can also be impacted by iron additions.

Speciation of Iron in Seawater

Before proceeding to discuss the effect of iron on various marine organisms, there are some important issues that relate to the bioavailability of iron that should be understood.  Bioavailability is a concept that has applications that range from medicine to ecology.  In short, its premise is that not all of some particular agent is available to organisms.  For example, if a pharmaceutical that is intended to treat brain cancer is given orally and is not absorbed from the gastrointestinal tract, it will not likely be effective.  In this case, the drug would be described as having poor bioavailability.  That is, it is not available where it is needed.

In discussing metals such as iron in seawater, bioavailability can be critically important.  Iron can take many forms in seawater, some of which are readily available to organisms and some of which are not.  For many organisms, iron that is complexed to certain organic ligands is not bioavailable.  In a sense, the organisms cannot use the iron, and are not impacted by it.  In other cases, the organic/iron complex can be absorbed and used, and in some of the most interesting cases, these ligands are specifically designed by organisms to “go out and collect iron”.   Consequently, it behooves us to understand as much as possible about the speciation of iron in seawater (and tank water) in order to understand the effects that it might be having on organisms.

While these general concepts apply to all metals in seawater, it has recently become clear that iron in particular is extensively bound to organic chelators.  For example, one research group recently claimed in the journal Nature that “Dissolved Fe(III) in the upper oceans occurs almost entirely in the form of complexes with strong org. ligands presumed to be of biol. origin.” ¹

Others researchers claim “Previous coastal marine studies concluded that total dissolved Fe may affect initiation of algal blooms, including brown tides of Aureococcus anophagefferens.  However, the existence of unavailable colloidal and organically-complexed Fe make the dissolved pool a poor indicator of what is bioavailable for phytoplankton assimilation.”⁴

Consequently, if one were assessing the status of a tank with respect to iron, one would have to understand the specific nature of the iron present.  In other words, one cannot use measurements of total iron, such as those provided by most test kits or by the analytical technique ICP, to know whether a given tank “needs” iron or not.  This unfortunate circumstance prohibits reefkeepers dosing iron from having the type of simple relationship between concentration and effect that is enjoyed by many other ions of interest, such as calcium.  One might be able to use these techniques to follow bioavailability as a function of time in a single reef tank with increases or decreases in iron dosing since the level of available chelators may remain mostly unchanged (but I have not done this experiment).

Whole System Studies in the Ocean

The iron enrichment literature as a whole falls into 3 categories.  In the first, individual species are studied in isolation.  In the case of microorganisms, these studies would be the least relevant to aquaria where there is competition between many organisms for the same nutrients, space, etc, as well as predation.  The second category examines the effect of iron in the open ocean, but does not compare changes in growth of one organism to changes in growth of other organisms.  In the third category are those studies that have added iron to systems (such as the ocean) and looked at the absolute AND the relative populations of various organisms, including the effects of predation. 

It is this last category of study that I will describe first, since it most accurately reflects what one might expect to happen when one adds iron to a whole ecosystem: a reef tank.  Unfortunately, there are no studies of this kind that describe the effects of iron on corals.  Consequently, the effects of iron on corals will have to be inferred from less grand studies.

In one study, the researchers added iron to a variety of ocean environments and tracked the population change in various organisms.⁵  What they found is very interesting.  The primary organisms that increased relative to the others were diatoms.  The also found that cyanobacteria and dinoflagellates declined, and bacteria remained largely unchanged.  They state “…at least eight diatom genera and an undetermined number of different autotrophic nanoplankton taxa were present in higher numbers in the Fe+ carboys, whereas cyanobacteria, one diatom group, and dinoflagellates were more numerous in the controls”.  Also, “In contrast [to studies adding things besides iron], the HNLC bacterial communities in our experiments showed only a small response, despite large Fe-induced changes in biological and biochemical parameters.”

Similar results were seen in the well-publicized IRONEX experiments.  These studies have shownthat when there is adequate nitrate and phosphate, iron may become the limiting factor for phytoplankton growth. ^6-10    In IRONEX II, for example, diatoms accounted for 17% of the biomass growth prior to iron enrichment, and 79% after iron addition.¹¹  In this test, in fact, diatoms experienced a 70x increase in population.¹²  It was also found in this study that cyanobacteria did not increase growth as much as diatoms.  One common cyanobacterium, Prochlorococcus, actually decreased.¹³  Heterotrophic (nonphotosynthetic) dinoflagellate and ciliate populations increased substantially (>3x), presumably as they consumed the diatom bloom. ¹²  Heterotrophic bacteria experienced a smaller population increase (1.7 x).¹⁴

On the basis of these types of studies, the literature points to diatoms as one of the most likely things to become relatively enhanced by iron additions.

Cyanobacteria and Iron

It has been suggested by some hobbyists that iron additions to reef tanks may drive the growth of cyanobacteria, and that one should not dose it for that reason.  I’ve not seen that effect in my tank, nor have I heard it reported in others. It was also not reported to be significant in the open ocean studies described above.  Nevertheless, since cyanobacteria are a problem for many reefkeepers, this issue is an especially important one to address.

Fortunately, there is a great deal of literature on the relationship between iron and cyanobacteria.  Most of the literature indicates that cyanobacteria are especially well suited to low iron environments because they are able to release siderophores that bind to iron and give them a competitive advantage over other organisms.  This is, in fact, one of the reasons why they have been studied so extensively with respect to iron.  If they do generally have a competitive edge at low iron levels, then adding iron supplements and swamping out this competitive advantage may make strategic sense if reducing cyanobacteria is a goal. 

In one paper, for example, the researchers conclude “that cyanobacteria are efficiently adapted to grow in low-Fe environments (providing sufficient light for photosynthesis is available)…” ¹⁵  In a different paper, the researchers state: “This review focuses on how cyanobacteria respond to growth-limiting levels of available iron and on how siderophores potentially alter the biological availability of iron in the system thereby allowing the cyanobacteria to exist at low iron availabilities.” ¹⁶  In another paper  the researchers show that “The growth rates and intracellular and total cellular
iron levels for Synechococcus PCC 7002 demonstrate that iron availability does not directly dictate the maximum growth rate of these cyanobacteria…”¹⁷ 

Finally, in this paper, it appears that the three species of cyanobacteria tested are not iron limited for growth in the ocean (2 are phosphate limited, the other may be phosphate limited) while one of the species of macroalgae may be iron limited (Dictyota bartayresiana). ¹⁸ 

To be fair, some researchers do make seemingly contrary claims, though the fact that there are many species of cyanobacteria makes that result not overly surprising. One research group states that “Our results suggest that in 75% of the global ocean, iron availability limits nitrogen fixation by this organism.”¹⁹  Previously, other researchers had shown that iron does indeed stimulate growth of this particular cyanobacterium.²⁰

These results for cyanobacteria are essentially what one would expect for an organism that can grow well even in low iron conditions.  This fact does not demonstrate that the cyanobacteria won’t become iron limited under the higher phosphate and nitrate conditions present in a reef tank, but it also does not indicate that there is, at present, any cause for alarm about cyanobacteria and iron additions.

Effects of Iron on the coral Stylophora pistillata

Unlike the research areas described above, the work on corals has been very limited, with three papers meriting attention.  The first and most important of these papers, Response of a scleractinian coral, Stylophora pistillata, to iron and nitrate enrichment, was published in the Journal of Experimental Marine Biology and Ecology.²¹  The experiments in this paper essentially consisted of putting coral fragments (7 cm long) in a series of tanks.  The fragments were monitored for 4 weeks, the water was altered chemically, and the fragments were monitored for another 3 weeks.  This monitoring consisted of several measurements: coral growth rate, zooxanthellae density, photosynthesis rate, and chlorophyll content.

The starting water itself was pumped from the Mediterranean. It is low in nutrients, such as nitrate (< 0.06 ppm; < 1 mM); phosphate (< 0.02 ppm< 0.2 mM), and iron (< 0.2 ppb; < 4 nM). For test purposes, iron was raised to 0.3 ppb, and nitrate was raised to 0.15 ppm (yes, even the enriched nitrate level was far below that in most reef tanks).  The water volume of each tank was replaced continually so that a 100% change took place each hour.

Let’s begin, as they did, by discussing the zooxanthellae density.  This density is a measure of the number of zooxanthellae cells present.  It can be calculated as the number of zooxanthellae per animal cell, or by the total density of zooxanthellae.  By both of these measures, the zooxanthellae increased substantially in all of the test aquaria compared to the control.  The largest increase in zooxanthellae was found in that with just iron additions.  Essentially the same results were found for the chlorophyll content.  On the face of it, these experiments suggest that the growth rate of the zooxanthellae is both iron and nitrate limited. 

I’ll discuss later whether it is advantageous or not to boost zooxanthellae growth rates, but from this experiment is not clear is whether the iron and nitrate additions cause ongoing growth of the zooxanthellae, with excess zooxanthellae continually being expelled or otherwise lost, or whether the zooxanthellae have simply multiplied and reached a new plateau in density where they continue to grow at rates similar to before the iron and nitrate enrichment.  This issue is important because it bears on coral growth rates (below) and whether the changes seen in this 3 week test will continue for additional time periods.

The authors also measured various aspects of photosynthesis for all of these corals.  They found that photosynthesis was increased in all of the test tanks, with iron alone being the largest increase.  On a per zooxanthellae basis, however, the photosynthesis was unchanged.  This suggests that more zooxanthellae photosynthesize more as a whole (just as a large field of corn photosynthesizes more than a small field), but that the individual cells are photosynthesizing at about the same rate.  Apparently the added nutrients have increased the numbers of zooxanthellae, but did not otherwise impact photosynthetic activity.

Finally, the researchers measured coral growth by weight.  The growth rate was found to decrease substantially in each of the test tanks compared to control.  That is, iron and nitrate, individually and in combination, had a big effect on coral growth rates, with all of them reducing growth by about 30%.  The authors state that “Iron seems, therefore, toxic to the coral host, even if it increases the total number of algae.” 

The nature of this “toxicity”, however, may be somewhat less worrisome to reefkeepers than the word implies.  They authors propose a number of mechanisms for this effect, of which they seem to prefer the simple suggestion that if the zooxanthellae are growing more rapidly, they won’t be delivering as much in the way of useful photosynthetic byproducts to the host animal, decreasing its growth rate.  The do state that the relationship between calcification and iron additions remains unclear. 

This explanation begs the question, suggested above, of whether this was a one time change caused by a one time increase in zooxanthellae, or whether there will be an ongoing lower level of delivery of photosynthetic byproducts.  Only longer-term experiments would easily answer this question.

I have no reason to doubt the possibility of reduced delivery of photosynthetic byproducts, and it makes perfect sense, but I’d like to propose another possible explanation that they did not address: phosphate limitation.  For phytoplankton, phosphorus limitation in the ocean is usually secondary to nitrogen limitation.  That is, nitrogen is more limiting.  Still, according to Millero²²:

 “Above a phosphate concentration of 0.3 mM the rate of growth of many species of phytoplankton is independent of the concentration of P.  Below 0.3 mM cell division becomes inhibited and P-deficient cells are produced.  This probably does not occur in the oceans since NO₃^- is usually exhausted before PO₄^--- falls to this critical level”

The same is largely true of most macroalgae, with nitrate and phosphate being the limiting factors under different conditions and for different species.^23-25  Of course, those studies relate to phytoplankton and macroalgae, and not a coral.  However, it is not an unreasonable hypothesis that corals may be similarly limited at the low levels of phosphate encountered in this test (< 0.2 mM) when given extra nitrate.

Additionally, the IRONEX studies have shown that when there is adequate nitrate and phosphate, iron may become the limiting factor for phytoplankton growth.^6-10  In IRONEX I, for example, the phosphate level was1 mM and nitrate was 12 mM, both well above the values in this test, even after nitrate enrichment.  In that experiment, iron appeared to limit phytoplankton growth.

Putting these two ideas together, and retaining the caveat that these various studies involved phytoplankton and macroalgae, and not a coral, it is a reasonable possibility that the corals in this study with enriched iron and nitrate may be phosphate limited.  In such a scenario, the rapidly growth zooxanthellae may use up the absorbed phosphate, making the host unable to obtain enough phosphate, and thereby grow more slowly.

If phosphate limitation is a factor here, then it may not be so in reef tanks, where phosphate levels are typically far higher than in this test (and often equivalent to that in the IRONEX I study).  Additionally, since nitrate alone at 0.15 ppm had almost as big of an effect as nitrate plus iron, then perhaps all, or nearly all reef tanks (those with nitrate above 0.15 ppm) are already experiencing this reduced coral growth rate due to increased zooxanthellae growth, and the iron may not make the problem substantially worse.  Moreover, there may already be more than enough iron in reef tanks for this effect to have happened even in the absence of any iron supplements.

Whether one is concerned with coral growth rates, or even zooxanthellae density (which may impact the colors of corals), there is obviously substantial uncertainty here with any extrapolation of these results to real reef tanks.   Nevertheless, the experiment is interesting and helps us to understand issues involving iron in our reef tanks better.  In particular, it helps guide us in what types of problems to be on the lookout for.  These potential effects are detailed at the end of this article.

A third paper on corals is titled Indications from photosynthetic components that iron is a limiting nutrient in primary producers on coral reefs. ²⁷   This paper discusses biochemical evidence of iron limitation in a variety of organisms on a coral reef, but does not actually test elevated iron levels to see if growth limitation is real.  In it, these authors suggest that zooxanthellae in Sinularia sp. may be iron deficient.  They conclude “The degree and extent of Fe-stress in primary producers on a coral reef thus may influence growth rates, biomass, and distribution of species”.

In addition to these papers on coral growth, there are many papers on the presence of iron in coral tissue²⁸ and skeletons.^{29, 30}  These papers frequently show that iron levels in corals are increased when the level of iron in the water is increased.  Whether that is generally “good or bad” is not typically addressed.

References

1. Response of a scleractinian coral, Stylophora pistillata, to iron and nitrate enrichment.     Ferrier-Pages, Christine; Schoelzke, Vanessa; Jaubert, Jean; Muscatine, Len; Hoegh-Guldberg, Ove.    Observatoire Oceanologique Europeen, Centre Scientifique de Monaco,  Monaco,  Monaco.    Journal of Experimental Marine Biology and Ecology  (2001),  259(2),  249-261. For those without a subscription or a handy library that has it, it is available online for $30 for most individuals (some people seem to get a different message at that site and I don’t understand why; my two computers even get different messages; this may relate to whether the computer is “thought” by the web site to be part of an institution that does or does not already have a relationship with the publisher). 

2. Chemical Oceanography, Second Edition.     Millero, Frank J.; Editor.    USA.    (1996),  496 pp.Publisher: (CRC,  Boca Raton, Fla.) .

3.  Effects of nitrate, phosphate and iron on the growth of macroalgae and benthic cyanobacteria from Cocos Lagoon, Guam.     Kuffner, Ilsa B.; Paul, Valerie J.    UOG Station,  University of Guam Marine Laboratory,  Guam,  USA.    Marine Ecology: Progress Series  (2001),  222  63-72. 

4. Nutrient-limited growth of the coral reef macroalga Sargassum baccularia and experimental growth enhancement by nutrient addition in continuous flow culture.     Schaffelke, Britta; Klumpp, David W.    PMB 3,  Australian Institute Marine Science,  Townsville,  Australia.    Marine Ecology: Progress Series  (1998),  164  199-211. 

5. Nutrient limitation of the macroalga, Penicillus capitatus, associated with subtropical seagrass meadows in Bermuda.     McGlathery, Karen J.; Howarth, Robert W.; Marino, Roxanne.    Div. Biol. Sci.,  Cornell Univ.,  Ithaca,  NY,  USA.    Estuaries  (1992),  15(1),  18-25.

6 IronEx-I, an in situ iron-enrichment experiment: experimental design, implementation and results.     Coale, Kenneth H.; Johnson, Kenneth S.; Fitzwater, Steve E.; Blain, Stephane P. G.; Stanton, Tim P.; Coley, Teresa L.    Moss Landing Marine Laboratories,  Moss Landing,  CA,  USA.    Deep-Sea Research, Part II: Topical Studies in Oceanography  (1998),  45(6),  919-945. 

7. The behavior of iron and other trace elements during the IronEx-I and PlumEx experiments in the Equatorial Pacific.    Gordon, R. M.; Johnson, K. S.; Coale, K. H.  Deep-Sea Research, Part II: Topical Studies in Oceanography  (1998),  45(6),  995-1041. 
8. Mesozooplankton grazing manipulations during in vitro iron enrichment studies in the NE subarctic Pacific.Boyd, P. W.; Goldblatt, R. H.; Harrison, P. J.  Deep-Sea Research, Part II: Topical Studies in Oceanography  (1999),  46(11-12),  2645-2668.

9. Biological response to iron fertilization in the eastern equatorial Pacific (IronEx II). I. Microplankton community abundances and biomass.    Landry, M. R.; Ondrusek, M. E.; Tanner, S. J.; Brown, S. L.; Constantinou, J.; Bidigare, R. R.; Coale, K. H.; Fitzwater, S.  Marine Ecology: Progress Series  (2000),  201  27-42.

10. Confirmation of iron limitation of phytoplankton photosynthesis in the equatorial Pacific Ocean.    Behrenfeld, Michael J.; Bale, Anthony J.; Kolber, Zbigniew S.; Aiken, James; Falkowski, Paul G.  Nature (London)  (1996),  383(6600),  508-511.

11.  Metal tolerance in the scleractinian coral Porites lutea.     Harland, A. D.; Brown, B. E.    Cent. Trop. Coastal Manage.,  Univ. Newcastle upon Tyne,  Newcastle upon Tyne,  UK.    Mar. Pollut. Bull.  (1989),  20(7),  353-7. 

12.  Indications from photosynthetic components that iron is a limiting nutrient in primary producers on coral reefs.     Entsch, B.; Sim, R. G.; Hatcher, B. G.    Aust. Inst. Mar. Sci.,  Townsville,  Australia.    Mar. Biol. (Berlin)  (1983),  73(1),  17-30.

13. Metal concentration in the tissue and skeleton of the coral Montastrea annularis at a Venezuelan reef.     Bastidas, C.; Garcia, E.    Departamento de Biologia de Organismos, Universidad Simon,  Caracas,  Venez.  Editor(s): Lessios, Harilaos A.; Macintyre, Ian G.    Proceedings of the International Coral Reef Symposium, 8th, Panama, June 24-29, 1996  (1997),  2  1847-1850. 

14. Metal content on the reef coral Porites astreoides: an evaluation of river influence and 35 years of chronology.     Bastidas, C.; Garcia, E.    Depto. de Biologia de Organismos,  Universidad Simon Bolivar,  Caracas,  Venez.    Marine Pollution Bulletin  (1999),  38(10),  899-907.

15. Trace elements found to be variable in two coral reef species, Heliofungia actiniformis and Galaxea fascicularis, collected from the Ryukyu Islands.     Yamada, Gen; Fujimori, Ken; Yamada, Masa-Oki; Minami, Takeshi; Tohno, Setsuko; Tohno, Yoshiyuki.    Department of Cellular and Developmental Biology, Research Center of Innovative Cancer Therapy,  Kurume University,  Fukuoka,  Japan.    Biological Trace Element Research  (1998),  65(2),  167-180. 

16. Physiological changes in the coastal marine cyanobacterium Synechococcus sp. PCC 7002 exposed to low ferric ion levels.     Trick, Charles G.; Wilhelm, Steven W..    Department of Plant Sciences, University of Western Ontario,  London,  ON,  Can.    Marine Chemistry  (1995),  50(1-4),  207-17.

  17. Ecology of iron-limited cyanobacteria: a review of physiological responses and implications for aquatic systems Wilhelm SW, Aquatic Microbial Ecology (1995), 9:295-303.

18. Growth, iron requirements, and siderophore production in iron-limited Synechococcus PCC 7002.    Wilhelm, Steven W.; Maxwell, Denis P.; Trick, Charles G.  Limnology and Oceanography  (1996),  41(1),  89-97. 

19.  Effects of nitrate, phosphate and iron on the growth of macroalgae and benthic cyanobacteria from Cocos Lagoon, Guam.     Kuffner, Ilsa B.; Paul, Valerie J.    UOG Station,  University of Guam Marine Laboratory,  Guam,  USA.    Marine Ecology: Progress Series  (2001),  222  63-72.

20. Iron availability, cellular iron quotas, and nitrogen fixation in Trichodesmium.     Berman-Frank, Ilana; Cullen, Jay T.; Shaked, Yeala; Sherrell, Robert M.; Falkowski, Paul G.    Institute of Marine and Coastal Sciences,  Rutgers University,  New Brunswick,  NJ,  USA.    Limnology and Oceanography  (2001),  46(6),  1249-1260. 

21. Iron-stimulated N2 fixation and growth in natural and cultured populations of the planktonic marine cyanobacteria Trichodesmium spp.     Paerl, Hans W.; Prufert-Bebout, Leslie E.; Guo, Chunzhi.    Inst. Mar. Sci.,  Univ. North Carolina,  Chapel Hill, Morehead City,  NC,  USA.    Appl. Environ. Microbiol.  (1994),  60(3),  1044-7.

22.  Response of marine bacterial community composition to iron additions in three iron-limited regimes.     Hutchins, David A.; Campbell, Barbara J.; Cottrell, Matthew T.; Takeda, Shigenobu.    College of Marine Studies,  University of Delaware,  Lewes,  DE,  USA.    Limnology and Oceanography  (2001),  46(6),  1535-1545.

23. 11.  Photochemical cycling of iron in the surface ocean mediated by microbial iron(III)-binding ligands.     Barbeau K; Rue E L; Bruland K W; Butler A    Department of Chemistry and Biochemistry, University of California, Santa Barbara, California 93106, USA      NATURE  (2001 Sep 27),  413(6854),  409-13. 

24. Algae and cyanobacteria release organic chelators in the presence of inorganic Fe(III) thus keeping iron dissolved.     Benderliev, Konstantin.    Institute of Plant Physiology,  Acad. M. Popov,  Sofia,  Bulg.    Bulgarian Journal of Plant Physiology  (1999),  25(1-2),  65-75.

25. Study of the Organic Iron Complexation in Two Contrasted Environments: The Southern Ocean and the North-East Atlantic Ocean. Boye, M. vanden Berg, C. M. G.; Timmermans, K. R. Nolting, R. F.; de Jong, J. T. M.; de Baar, H. J. W.;  University of Liverpool;  online abstracts of the European Geophysical Society: http://www.copernicus.org/EGS/egsga/nice00/programme/abstracts/aac3596.pdf

26. Physicochemical speciation of iron during coastal algal blooms.     Gobler, Christopher J.; Donat, John R.; Consolvo, John A.; Sanudo-Wilhelmy, Sergio A.    Marine Sciences Research Center,  State University of New York at Stony Brook,  Stony Brook,  NY,  USA.    Marine Chemistry  (2002),  77(1),  71-89.

27. Biological response to iron fertilization in the eastern equatorial Pacific (IronEx II). III. Dynamics of phytoplankton growth and microzooplankton grazing.     Landry, M. R.; Constantinou, J.; Latasa, M.; Brown, S. L.; Bidigare, R. R.; Ondrusek, M. E.    Department of Oceanography,  University of Hawai'i at Manoa,  Honolulu,  HI,  USA.    Marine Ecology: Progress Series  (2000),  201  57-72.

28. Biological response to iron fertilization in the eastern equatorial Pacific (IronEx II). I. Microplankton community abundances and biomass.     Landry, M. R.; Ondrusek, M. E.; Tanner, S. J.; Brown, S. L.; Constantinou, J.; Bidigare, R. R.; Coale, K. H.; Fitzwater, S.    Department of Oceanography,  University of Hawai'i at Manoa,  Honolulu,  HI,  USA.    Marine Ecology: Progress Series  (2000),  201  27-42.

29. Differential response of equatorial Pacific phytoplankton to iron fertilization.     Cavender-Bares, Kent K.; Mann, Elizabeth L.; Chisholm, Sallie W.; Ondrusek, Michael E.; Bidigare, Robert R.    Department of Civil and Environmental Engineering,  Massachusetts Institute of Technology,  Cambridge,  MA,  USA.    Limnology and Oceanography  (1999),  44(2),  237-246.

30. The heterotrophic bacterial response during a mesoscale iron enrichment experiment (IronEx II) in the eastern equatorial Pacific Ocean.     Cochlan, William P.    Romberg Tiburon Center for Environmental Studies,  San Francisco State University,  Tiburon,  CA,  USA.    Limnology and Oceanography  (2001),  46(2),  428-435.