全 文 ：Responses of Picoplankton to Nutrient Perturbation in the South China Sea ,
with Special Reference to the Coast_wards Distribution of Prochlorococcus
JIAO Nian_Zhi1＊ , YANG Yan_Hui1 , Hiroshi KOSHIKAWA2 , Shigeki HARADA2 , Masataka WATANABE2
(1.Key Laboratory for Marine Environmental Sciences , Ministry of Education , Xiamen University , Fujian 361005 , China;
2.National Institute for Environmental Studies , Tsukuba , Ibaraki 305_0053, Japan)
Abstract: Responses of Prochlorococcus (Pro), Synechococcus (Syn), pico_eukaryotes (Euk)and het-
erotrophic bacteria(Bact)in pelagic marine ecosystems to external nutrient perturbations were examined using
nitrogen_(N), phosphorus_(P), iron_(Fe), and cobalt_(Co)enriched incubations in the South China Sea
in November 1997.Variations in abundance of the 4 groups of microorganism and cellular pigment content of
the autotrophs during incubation were followed by flow_cytometric measurements for seven days.During the in-
cubation , Syn and Euk showed a relatively higher demand on Fe andN , while Pro required higher levels of Co
and P.The Fe was inadequate for all the organisms in the deep euphotic zone (75 m)of the study area.The
experimental results also implied that biological interaction among the organisms played a role in the community
structure shift during the incubation.It seemed that besides the effects of temperature , there are some other
physical and chemical limitations as well as impacts from biological interactions on Pro distribution in coast wa-
ters.
Key words: Prochlorococcus;picoplankton;nutrients;iron;cobalt;South China Sea
Since the discovery of Prochlorococcus (Pro), an
extremely small(mean cell size 0.6μm), divinyl chloro-
phyll containing prokaryotic oxy_phototrophic au-
totroph
[ 1] , new perspectives are required to understand
the structure of micro_communities and the relevant eco-
logical processes in the marine ecosystems.Over the past
decade , many field ecological investigations of Pro have
been conducted
[ 2-7] .Distribution of Pro has been ex-
tended from tropical and subtropical to near sub_arctic ar-
eas[ 8] and from oceanic waters to certain coastal wa-
ters[ 7 ,9 , 10] .The significance of Pro in total phytoplankton
biomass , production and energy_flow pathways , especially
its distribution and relationships with other picoplankton
such as Synechococcus (Syn), pico_eukaryotes (Euk)
and heterotrophic bacteria (Bact), has become a major
concern of biological oceanographers.Temperature is the
crucial environmental factor to the distribution of Pro
[ 5 , 6] ,
which was also verified by laboratory experiments[ 11] .
Physical conditions such as mixing and stratification have
also been proven to be important in describing depth pro-
files and seasonal fluctuations of Pro populations[ 5 , 12] .
Chemical conditions such as nutrients are also considered
to be affecting factors in the distribution of Pro[ 13 , 14] , yet
nutrient impacts are not as clear as the other factors.Ma-
jor nutrients such as nitrogen and phosphorus have been
the basic concerns in traditional concepts of limiting fac-
tors for phytoplankton in the marine environment.Recent
studies have revealed that micronutrients , such as
iron[ 15 ,16] and Cobalt[ 17 , 18] , are crucial to phytoplankton
in vast areas of the world oceans.Furthermore , in our
previous studies on the field ecology of picoplankton in the
East China Sea , we observed significant correlations be-
tween Pro and other picoplankton organisms such as Syn ,
Euk and Bact
[ 19] .Similar relationships were also found
across the Gulf Stream in the North Atlantic in summer
1996(Jiao , unpublished data).These evidence suggest-
ed that biological interaction as a possible factor influenc-
ing the distribution of Pro cannot be ignored.
Based on the above considerations , especially the
difference in nutrient composition and concentration be-
tween oceanic waters and coastal waters , this study was to
explore the response of Pro as well as the other co_existing
picoplankton components Syn , Euk and Bact , to enhance
nutrient concentrations from oligotrophic oceanic levels to
eutrophic coastal levels , and the biological interactions
among these organisms during the nutrient condition
shifts.We hoped to obtain some information for interpret-
ing the distribution of Pro in the transition areas of
marginal seas.We chose a typical marginal sea in the
western Pacific , the South China Sea , as the in situ ex-
perimental field , and employed long_time (seven days)
nutrient enriched incubation as the experimental strategy.
1 Materials and Methods
1.1 Experimental site
The South China Sea , 5°-20°N and 109°-120°
E , is a marginal sea in the western Pacific covering tropi-
cal and subtropical regions.It is characterized by high
temperature (surface water temperature was around 29 ℃
during the study period , November 1997)and meso_olig-
otrophic conditions.The nutrient enrichment experimental
Received:2001-09-11 Accepted:2002-03-28
Supported by the National Natural Science Foundation of China (39625008 , 49876033) and the State Key Basic Research and Development Plan of China
(G2000078500).
＊Author for correspondence.E_mail:
植 物 学 报
Acta Botanica Sinica 2002 , 44(6):731-739
site was chosen from 32 field investigation stations , the
station No.43 , located at latitude 6°N , longitude 110°
E , with water depth of 1 100 m and thermocline and nitr-
acline of 60-70 m.Water samples were taken for incu-
bation from the surface layer (0-5 m)and 75 m , the
maximum chlorophyll layer around the nitracline.Water
temperatures were 28.62 ℃ and 20.85 ℃ respectively.
Nutrient concentrations (μmol/L) at 5 m depth were
0.0 , 0.017 , 3.558 and 0.218 for NO3_N , NO2_N ,
NH4_N and PO4_P , respectively ;those at 75 m were
14.93 , 0.017 , 2.738 and 0.495.
1.2 Experimental design
In order to see the influence of coastal water on Pro ,
all experiments were designed with respect to shifting the
ecological conditions in the incubation bottles from ocean-
ic to typical coastal status.Four elements , nitrogen(N),
Phosphorus(P), iron(Fe)and cobalt(Co), were cho-
sen for the nutrient enrichment experiments based on the
following consideration.N and P are naturally more abun-
dant in coastal than in oceanic waters and are frequently
reported as limiting factors for phytoplankton;Fe is an el-
ement that phytoplankton require for synthesis of chloro-
phyll and nitrate reductase , and has been proven to be
deficient in most oceanic waters
[ 15 ,16 ,20] , although it is
much more abundant in coastal areas.Co has been report-
ed to be important to phytoplankton metabolism[ 17 ,18] .All
the nutrient enrichment levels were designed according to
high concentration levels recorded in the natural coastal
areas of the China Seas
[ 18 ,21 ,22] .The concentrations
added to the incubation bottles were:50 μmol/L N(NH4Cl), 10 μmol/L P (NaH2PO4), 0.117 μmol/L Fe(FeCl3), and 0.005 μmol/L Co (CoCl2), respectviely.
One litre glass bottles were used as incubators.The
bottles were cleaned according to the procedures described
by Fitzwater et al[ 23] .Water samples were taken from the
Go_flo bottles with almost no exposure to air.The incuba-
tion bottles were treated individually in a small area iso-
lated by clean plastic sheets[ 23] .The time lags of incuba-
tion after sampling was minimized to the best attempts.
Incubations were conducted in flowing surface water_
cooled water baths on the deck of the research vessel.
Blue plastic sheets were utilized as covers to provide light
at ambient levels of the sampling depths.However , we
could not keep the temperature of incubation bottles of the
75 m water the same as that at the sampling depth.Du-
plicate incubations were conducted for each treatment.
In order to observe the interactions of picoplankton
under conditions shifting away from their origin , incuba-
tions were lengthened up to seven days.Bottles were
shaken several times a day.1 mL subsamples were taken
from the incubation bottles every day during incubation for
cell abundance and cellular pigment content examina-
tions.Samples were placed in 1.2 mL cryogenic vials(Nalgene)and were fixed with glutaraldehyde (final con-
centration , 1%)in dark for 10 min , then stored in deep
freezers or liquid nitrogen until analysis.
1.3 Flow_cytometric(FCM)analysis
Samples were run on an FACSCalibur flow cytometer(Becton_Dickinson)equipped with an external quantita-
tive sample injector(Harvard Apparatus PHD 2000);the
injection flow rate was set at 10-20 μL/min for normal
enumeration.FCM data were acquired and analyzed by
CellQuest 2.2(Becton Dickinson).We used 0.474 μm
fluorescent beads(Fluorescence Scientific)as the internal
reference.The three autotrophs were distinguished ac-
cording to their positions in plots of chlorophyll (FL3)
vs.90°_angle light scatter (SSC), and phycoerythrin
(FL2)vs.SSC[ 5] .Cellular pigments were normalized to
bead units.SYBR Green_I (Molecular Probes)was ap-
plied as the DNA stain for heterotrophic bacteria enumera-
tion
[ 24] .Samples for FCM enumeration of autotrophs were
run separately from those for heterotrophs.
2 Results
2.1 Variations in cell abundance of the picoplank-
ters and in cellular pigment content of the autotrophs
in the surface water incubations
During the seven days incubation of control bottles ,
the Pro population continued to decline.Bact population
was relatively stable in the first three days , and then kept
increasing to the end.There was a significant inverse cor-
relation between the two(P<0.01).The Syn population
followed a trend similar to that of Bact except for a signifi-
cant drop on the second day.The population of Euk de-
creased at first and then increased to a relatively stable
level at about 1/3 of the initial population(Fig.1a).The
cellular pigment content of Syn in the first half of the in-
cubation was about twice as the initial level , then de-
creased from the fifth day to a level similar to the initial(Fig.2a).Pigment levels of Pro increased a little in the
first two days and then dropped down to levels similar to
or lower than the initial level.Euk s cellular pigment
contents dropped abruptly from the beginning of the incu-
bation and kept a low level till the end (Fig.2 , b , c).
In the bottles supplemented with Co (Fig.1b), Pro
responded rapidly in the first 24 h and a considerable in-
crease in cell abundance was observed.The cell abun-
dance then decreased to the initial level where it remained
for the next three days.On the fifth day another smaller
drop occurred and then it maintained a level near 3×104
cells/mL until the end of the experiment.In contrast , the
Syn population decreased in the first two days , increased
rapidly after the Pro population declined , and then held a
concentration of 8.6×103 cells/mL until the end of the
experiment.The Euk population , although fluctuating ,
finally reached a level almost twice its initial value.Bact
showed a pattern similar to that in the control bottles.In
this treatment cellular pigment content of autotrophs
reached high levels early or late days of the incubation.
That of Pro in particular was markedly different from that
in the controls(Fig.2).This was one of the two most fa-
vorite conditions for Pro (the other being P , below)in
terms of both cell abundance and cellular pigment con-
tent.
732 植物学报 Acta Botanica Sinica Vol.44 No.6 2002
Fig.1. Time course of cell abundance of picoautotrophs and heterotrophic bacteria in the nutrient enrichment experiments.
5 m(surface)water incubations:a.Control.b , c , d and e.Enriched with cobalt , iron , ammonia and phosphate respectively.Y axis units:
Syn×5 , Pro×1 , Euk×100 , Bact/ 10 cells/mL.75 m water incubations:f.Control.g , h , i and j.Enriched with cobalt , iron ammonia and
phosphate respectively.Y axis units:Syn×15 , Pro×1 , Euk×50 , Bact/10 cells/mL.
For the Fe_enriched bottles (Fig.1c), Syn had a
significant response in the first three days and the last two
days , reaching the highest abundance of 2×104 cells/mL
at the end of the incubation.Euk responded rapidly to Fe
addition in the first 24 h and then decreased.Then on the
fifth day , there was another increase in Euk , to a concen-
tration higher than the initial value , and which remained
to the end of the incubation.The Pro population remained
high for the first three days , but almost disappeared from
the community on the fifth day.Unlike in the other treat-
ments , the Bact population did not grow much , maintain-
ing relatively stable throughout the incubation.Interest-
ingly , the fast reproduction of Syn and Euk resulted in
relatively lower cellular pigment contents in comparison
with their pigment contents in other treatments(Fig.2 , a ,
c).The trend line of Pro cellular pigment was relatively
low and straight , indicating that Pro cells were not in
good condition(Fig.2b).
When enriched with a high level of NH+4 _N (Fig.
1d), Pro responded rapidly at first and then declined as
Syn and Bact began to grow.Pro was actually removed
from the community by the end of the experiment.Both
Syn and Bact reached their highest abundance in the latter
half of the incubation.The Euk population remained rela-
tively stable during the first two days , then increased and
maintained a higher level until the sixth day , and then
decreased to the initial level at the end of the incubation.
High cellular pigment content of Euk during the first four
JIAO Nian_Zhi et al:Responses of Picoplankton to Nutrient Perturbation in the South China Sea 733
days showed that these cells were very active initially be-
fore their numbers increased(Fig.2c).
In P_enriched bottles (Fig.1e), Pro responded
rapidly to the enrichment;its population almost doubled
in 24 h , reaching a concentration of 8.5×104 cells/mL ,
which was never equaled in any other treatments.From
the third day , cell abundance of Pro decreased to a level
lower than the initial and kept decreasing to the end of the
incubation.However , its cellular pigment content kept
increasing (Fig.2b), distinguishing Pro from the other
groups in terms of both cell numbers and cellular pigment
content.Populations of the other two autotrophs , Syn and
Euk , remained almost constant during the incubation , al-
though their cellular pigment contents increased after the
Pro population decreased(Fig.2 , a , c).Unlike in the Fe
treatment where Syn and Euk were the dominant au-
totrophs , in this case , Pro was the one that remained
dominant almost throughout the incubation.
2.2 Variations in cell abundance of the picoplank-
ters and in cellular pigment content of the autotrophs
in incubations of the 75 m water
The patterns of abundance variation of the four
groups of organism in the incubation of the 75 m water(Fig.1 , f-j)were less complex than that in the surface_
water experiments.Bact showed uniform patterns in both
the nutrient_enrichment and the control bottles.Owing to
the increase of temperature from 21 ℃ at the sampling
depth to 29 ℃of the cooling water from the surface layer ,
the Bact population increased dramatically to 6 ×105
cells/mL in the first two days , and after a little decline ,
increased slightly again.Syn , the most successful species
in the surface_water incubations , was less successful in
the deep_water incubations , basically due to its low initial
abundance(about 1 000 cells/mL on average)and per-
haps , to the limited remaining environmental carrying ca-
pacity after the Bact population thrived.Although cellular
pigment content of Syn increased in all the bottles as a re-
sult of the enhanced temperature(Fig.2d), the Syn pop-
ulation developed only in Fe_enriched incubations where
Syn population reached 6 000 cells/mL by the end of the
experiment.Euk populations grew only in Fe and N treat-
ments.Pro population , in all but one treatment , gradual-
ly decreased to around 6 000 cells/mL , equivalent to one_
fifth of their initial abundance.The exception was Fe en-
richment , in which the Pro population decreased for the
first five days and then increased to a level close to the
initial abundance at the end of the incubation.
2.3 Relative responses of picoplankton to nutrient
enrichments
Responses of Syn , Pro , Euk and Bact in cell
Fig.2. Time course of autotrophic picoplankton cellular pigment content(in bead units)in the nutrient enrichment experiments.
5 m(surface)water incubation:a.Syn.b.Pro.c.Euk.75 m water incubation:d.Syn.e.Pro.f.Euk.
734 植物学报 Acta Botanica Sinica Vol.44 No.6 2002
abundance to the different treatments can be arranged in
order.As seen from Fig.1 , in most cases , populations
responded in the first 24 h of incubation and then under-
went dramatic changes in the following two to three days ,
after that , relative stable trends can be seen from the later
days.We thus took the ratio of the cell abundance after
24 h incubation(D1)to the initial abundance(D0)as an
indicator of biological response to the treatments(Fig.3).
While took the cell abundance averaged over the last three
days of the incubation as an indicator of the adaptability of
each group of organism to the changed environments(Fig.
4).
Fig.3. Ratios of picoplankton cell abundance of D1 (after 24 h
incubation)to D0 (the initial cell abundance)in different nutrient
treatments of the surface water incubations.
Dashed line indicates D1/ D0 =1.C , control;Fe , enriched with
iron;Co , enriched with cobalt;N , enriched with ammonia;P , en-
riched with phosphate.
Fig.4. Responses of picoplankton cell abundance to different nu-
trient in the surface water (5 m)incubations.The different treat-
ments were arranged in order in light of cell abundance averaged
over the 5th , 6th and 7th incubation days.
C , control;Fe , enriched with iron;Co , enriched with cobalt;N ,
enriched with ammonia;P , enriched with phosphate.
In the surface_water experiments , after 24 h incuba-
tion , Syn had a dramatic response to the Fe enrichment ,
its D1/D0 ratio was 1.67.Ratios in the other treatments
and the control were around 1 , indicating there was no
significant stimulation of the addition of these nutrients to
Syn or the responses of Syn to the treatments were slow(Fig.3 , Syn).While , in the case of Pro , except for the
control(C), all the ratios were greater than 1 , suggesting
that Pro responded very quickly to all the nutrient addition
with best to P treatment(Fig.3 , Pro).With the D1/D0
ratio in Fe treatment reaching 2 , Euk s response to Fe
JIAO Nian_Zhi et al:Responses of Picoplankton to Nutrient Perturbation in the South China Sea 735
addition ranked No.1 among all the organisms to all the
treatments.While ratios of Euk in all the rest treatments
were rather low.The response order of Fe>N>P>Co> C along with the sharp decreasing gradients also
showed big differences in Euk s response to different
treatments(Fig.3 , Euk).Unlike the autotrophs , D1/D0
ratios of Bact in all the treatments and the control incuba-
tion were greater than 1 , indicating that the conditions in
the incubation bottles somehow better suited for Bact to
thrive (Fig.3 , Bact).
Cell abundance averaged on the last three days incu-
bation showed great similarity in nutrient enrichment con-
sequence of Syn , Euk and Bact(Fe>N>Co >P >
C)and distinct difference from Pro (Co>P>C >N >
Fe), indicating that Syn , Euk and even Bact were most
favored by the addition of Fe , followed by N , while Pro
was essentially pushed aside by them under such condi-
tions.On the other hand , Co and P favored Pro better
than the other picoplankters.Although Co and P were al-
so effective to Syn , Bact and Euk (by comparison with
Control treatments).Since abundance of Bact in the con-
trol bottles were the lowest either in the first day or the
later days of incubation , all the nutrients added were
likely to be favorable to Bact in competition(Fig.4).
For the incubations of 75 m water , although an obvi-
ous response of Bact to enhanced temperature made the
effects of nutrient addition vague , some trends were yet
distinct.First , Fe was responded by all the picoplankters
including Pro.Ammonia addition , again responded by
Syn , Euk and Bact though ambient nitrate was abundant.
Unlike the situation of the surface water incubation , in
the 75 m water , Co was responded not only by Pro in the
beginning but also by Syn and Euk at the end , and P was
responded by Pro and Bact.
3 Discussion
3.1 Incubation conditions
Although long_time incubation entails the risk of fail-
ure to maintain the biota in healthy condition , five or
more days are often employed in microplankton nutrient
enrichment experiments[ 25] .In our experiment , except for
the general decease in Pro population in the later days of
the incubations , which is usually encountered during in-
cubation due to Pro s somehow non_culturability , no se-
vere inhibition occurred to the whole communities in the
bottles within the seven days incubation period.Similar
results were also recorded by previous authors[ 25] .Owing
to “wall effects” , small bottles may produce an artificial
bias , especially in long_time incubation[ 26] .On the other
hand , we did observe Pro population increases in the sec-
ond day in most of the incubations which consistent with
results of some short period (24-28 h)incubations in
the Atlantic ocean[ 27] .Thus , the results from the latter
days of our experiments should not be extended to natural
ecosystems , but rather be used to explore the biological
tolerance to environmental changes and the biological in-
teractions of the organisms under conditions changing from
their original environments.
3.2 Picoplankton nutrient requirements
Although there is no doubt that Fe limits diatoms
growth in high_nutrient , low_chlorophyll(HNLC)ecosys-
tems , there are different explanations about the effects of
iron on picoplankton , especially prokaryotes.Experiments
with deferriferrioxamine B demonstrated that growth of
Synechococcus is not strongly limited by Fe in the HNLC
equatorial Pacific Ocean[ 28] .Other studies on photosyn-
thesis showed that growth of the dominant small phyto-
plankton is held below the physiological potential by iron
deficiency[ 29] .From our incubation results , we speculate
that Fe was a limiting factor for picoplankton in the study
area;In the 75 m water , in particular , the naturally low
Fe and light availability induced higher requirements for
Fe for physiological activities[ 30] and created higher de-
mand for Fe by the dominant picoplankton there.Rela-
tively high cellular pigment content in most of the Fe_sup-
plemented incubations(Fig.2)is obviously the result of
higher chlorophyll synthesis as a result of higher Fe avail-
ability[ 31-33] .Because nitrate reductase activity can be
enhanced by Fe
[ 33 ,34] , the strong response to the addition
of Fe by Syn and Euk in the surface water , and of all the
autotrophs in the deeper water , can be easily understood.
Even for Bact , Fe was shown to be essential in the 75 m
water.This is consistent with results from Antarctic wa-
ters
[ 35]
and from the subarctic Pacific where bacteria are
responsible for 20% to 45%of biological iron uptake[ 36] .
Major nutrients such as N and P have drawn the in-
terest of oceanographers for many years.N has been iden-
tified as the primary limiting nutrient for phytoplankton on
a region_wide and year_round basis.P , although fre-
quently reported to be more important than N in limiting
phytoplankton productivity , is apparently less consistent ,
temporally and spatially , in potential regulatory impor-
tance than N
[ 37-39] .These conclusions are basically for
whole phytoplankton assemblages and applicable to gener-
al understanding.With respect to cell size , small cells
prefer ammonia to nitrate , and the majority of nitrate up-
take is accounted for by large cells
[ 40 ,41] .Most recent
work has revealed that only low_light ecotype Pro strain
can take up nitrite and all Pro strains in culture do not
utilize nitrate due to lack of nitrate reduction genes[ 42] .
Thus high nitrate concentration favors others rather Pro.
In the present study , although there was plenty of NO-3
(15 μmol/L)in the 75 m water , addition of NH+4 in-
duced in Euk a great increase in cell abundance with high
cellular pigment content , indicating either that Euk pre-
ferred ammonium to nitrate or that there was inefficient ni-
trate uptake under conditions of iron deficiency as dis-
cussed above.Moreover , Syn and Euk had higher re-
sponses to N than to P , while Pro responded more to P
than N.
Cobalt has been reported to be essential to phyto-
plankton metabolism , especially where zinc is depleted.
Some species , e.g.Synechococcus bacillaris , needs Co
but not Zn for growth[ 17] .However , Segatto et al[ 18] re-
ported some neutral and negative effects of Co on growth
736 植物学报 Acta Botanica Sinica Vol.44 No.6 2002
rate and biomass accumulation:the diatom Ditylum
brightwellii Bailey was not affected by cobalt addition ,
and the dinoflagellate Prorocentrum minimum (Pavillard)
Schiller was inhibited by cobalt addition.Our results
showed significant response of Pro and weaker but still
positive responses of other picoplankton to Co in terms of
both cell abundance and cellular chlorophyll content.
Even in the incubations of 75 m water , although Bact
grew very well due to the temperature enhancement , all
the autotrophs had higher cellular pigment content in Co
than in the other treatments.
3.3 Implication of biological interactions
By plotting cell abundance over time in the different
treatments for each group of the organisms , inverse trends
between Pro and Bact , and Pro and Syn can be observed.
To find the relationship among these organisms under rela-
tively stable conditions , we removed the data from the
first two days to avoid the influence of the instantaneous
response to the nutrient pulses and averaged the data for
the remaining five days for correlation analysis.The cor-
relation coefficients were -0.74 between Pro and Bact ,
and -0.75 between Pro and Syn.Therefore , Bact and
Syn would be the major competitors of Pro when the envi-
ronmental conditions shifted away from typical oceanic
conditions.
Large cells such as big diatoms , which we did not
count in our incubations , might have had significant im-
pact on Pro in some cases , especially in the Fe_enriched
bottles.Although the natural abundance of large cells is
lower compared with that of picoplankton in the study area(chlorophyll a in the less than 2μm size fraction account-
ed for 84.3%of the total), the populations of large cells
might be able to grow very well in nutrient_enriched incu-
bation bottles , especially in those enriched with Fe[ 43] .
Thus , biological competition from large cells should be
taken into consideration in interpreting picoplankton popu-
lation dynamics under incubation conditions.
In HNLC regions and subtropical regions , grazing by
microzooplankton is the dominant cause of phytoplankton
mortality
[ 26] .In both systems , the major small phyto-
plankton groups grow rapidly and are cropped to low stable
levels by microzooplankton.Sustained high growth rates of
the phytoplankton depend on remineralization of the by_
products of grazing[ 44] .Assuming that the grazing mortali-
ty of Pro was balanced by its growth , as found in similar
ecological conditions
[ 4 ,45] , Pro responded rapidly when
pulsed with additional nutrients in the incubation bottles ,
but the grazing pressure remained unchanged or changed
less proportionally in a relative short time.Thus , this
balance was broken , resulting in a sudden increase in
phytoplankton cell abundance.Pro actually increase to
the maximum concentration at the first one or two days ,
and then decreased rapidly consequent upon the grazing
pressure.However grazing loss might not be responsible
for the major decrease in Pro population in the later days
of the incubation course , as also can be seen from that the
other groups , Syn , Euk and Bact , all reached their high-
est abundance in the latter days of incubation.Because
Syn , Euk and Bact could also be cropped by graziers their
populations would have dropped as Pro encountered in the
later days of the incubation.Therefore we come to the
point that grazing pressure was not the determining factor
for the total loss of Pro cells during incubation.
We would therefore attribute the major loss of Pro ,
besides grazing mortality , to cells dying when stressed by
biological competition and other environmental conditions.
Typical cases in point are N and Fe enrichments in the
surface water incubations , where nutrient enrichment in-
duced a bloom of Syn and Euk and in turn inhibited
growth of Pro.Indeed , we did notice a sub_population of
Pro sinking down into noise in the FCM plots.Those cells
actually had less and less cellular pigment content as in-
cubation proceeded.On the other hand , Pro might also
impacted upon the other organisms.An example is the
P_enriched incubation in which Syn , Euk and even Bact
were distinctly influenced.The later days of 75 m water
incubation were typical cases of biological interaction in
which temperature enhancement induced sharply increases
in the populations of bacteria[ 46] and exerted severe stress
on the autotrophs.
3.4 Factors of regulating Pro coast_wards distribu-
tion
Although temperature is crucial to Pro[ 5 ,6 , 11] , it is
not the sole factor controlling Pro distribution since the
recorded temperature lower limits vary from place to
place
[ 5 , 7 ,8] .The difference between winter and summer
in the East China Sea can be as large as 10 ℃①.In con-
sistent with other investigations that physical factors such
as mixing and stratification influence Pro s distribution
pattern[ 5 ,12] , we also recognized , in another study , the
great effects of ocean currents on distribution of Pro in the
continental shelf of the East China Sea①.Beyond that ,
one basic concern is about nutrient which is easy to be
thought of as one of the major differences between oceanic
and coastal environments but not easy to test out due to
their subtle effects and complicated consequences.Al-
though Pro is most abundant in oligotrophic oceanic wa-
ters , it dose not necessarily mean Pro dislike high nutri-
ents[ 13 ,43] .The quick response of Pro to nutrient addition
at the first 24 h in the current study is also a piece of evi-
dence for this point.However , since high nutrient levels
may favor other picoplankters better than Pro , as demon-
strated in the experiments , Pro would be finally pushed
aside in the system.Situation of Pro s distribution in the
coastal waters are most likely such cases.Beyond the di-
rect effects of nutrients , the consequent biological
influences are also not negligible.The relationship be-
tween Pro and other picoplankters during the incubation
course are quite similar to what observed in the field along
trophic gradients in the East China Sea[ 19] .Therefore , we
speculate that Pro would encounter severe physical and
①Jiao Nian_Zhi , Yang Yan_Hui , Hiroshi Koshikawa , Masataka Watanabe ,
2001.Coupling of hydrographic conditions and picoplankton distribution in
the East China Sea , a marginal sea of the Northwest Pacifi c.To be appear on
the journal of Aquatic Microbial Ecology.
JIAO Nian_Zhi et al:Responses of Picoplankton to Nutrient Perturbation in the South China Sea 737
chemical limitations as well as impacts from biological in-
teractions in its coastwards distribution.
4 Summary
In our experiments on nutrient enrichment , Syn and
Euk demonstrated high similarities in terms of nutrient re-
quirements , especially their high demand for Fe and N.
In contrast , Pro responded more to Co and P than to Fe
and N.
Fe was inadequate for the autotrophs in the deep eu-
photic zone (75 m)in the study area.Although ambient
nitrate concentration was high there , it could not be uti-
lized efficiently by phytoplankton due to the deficiency of
Fe.Co seemed to be particularly essential to Pro and
Bact.
Biological interactions should be taken into consider-
ation for analysis of shifts in microbial communities under
changing environmental conditions.In the present study ,
addition of N seemed to be eventually unfavorable to Pro
because high N favored Syn and Euk , which in turn in-
hibited the growth of Pro.The vice versus also appeared
to apply in the P and Co treatments where Pro had im-
pacts on the other groups of organisms.Bact had the least
difference in response to different treatments.Bact growth
in the enhanced temperature incubations of 75m water in-
hibited all the autotrophs.Pro had different nutrient re-
sponse orders from the other picoplankters and inversely
correlated with Bact in the incubation course.
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从营养扰动实验看原绿球藻在近海分布的制约因素
焦念志1＊ 杨燕辉1 Hiroshi KOSHIKAWA2 Shigeki HARADA2
Masataka WATANABE
2
(厦门大学海洋环境科学教育部重点实验室 , 厦门 361005;
2.National Institute for Environmental Studies , Tsukuba , Ibaraki 305_0053, Japan)
摘要: 地球上细胞最小 、丰度最大的放氧光合自养原核生物原绿球藻(Prochlorococcus)发现于热带大洋 , 并被证实
可在某些近海甚至近岸水域大量分布。但除温度之外 ,原绿球藻自然分布的控制因子尚不明了。从近海和大洋生
态条件的主要差别考虑 ,在南海进行了主要营养盐———氮 、磷和微量元素———铁 、钴扰动的现场培养实验 , 并应用
流式细胞技术监测原绿球藻及聚球藻(Synechococcus)、超微型真核浮游植物(pico_eukaryotes)的细胞丰度和单细胞色
素含量的响应以及细菌的影响。结果表明 ,磷和钴的添加有利于原绿球藻 , 而氮和铁的添加更有利于聚球藻和超
微型真核浮游植物。同时 ,由环境条件引起的生物响应又间接地导致超微型生物之间的相互作用。因而 , 原绿球
藻在近海的分布 ,可能受到营养盐组成等环境因子以及生物之间的相互作用等多方面的限制和影响。
关键词: 原绿球藻;超微型浮游生物;营养盐;铁;钴;南海
中图分类号:Q945.97 文献标识码:A 文章编号:0577-7496(2002)06-0731-09
收稿日期:2001-09-11 接收日期:2002-03-28
基金项目:国家自然科学基金(39625008 ,49876033);国家重大基础研究和发展规划项目(G2000078500)。
＊通讯作者。E_mai l:
(责任编辑:韩亚琴(实习))
JIAO Nian_Zhi et al:Responses of Picoplankton to Nutrient Perturbation in the South China Sea 739