the impact of grazing by microzooplankton in northern hiroshima bay, the seto inland seam, japan

Marine Biology (1994) 119:77-88 �9 Springer-Verlag 1994

T. Kamiyama

The impact of grazing by microzooplankton in northern Hiroshima Bay, the Seto Inland Seam, Japan

Received: 30 September 1993 ! Accepted: 24 November 1993

Abst rac t The abundance of microzooplankton and their grazing impact on phytoplankton were studied using the dilution technique from May 1990 to November 1991 in northern Hiroshima Bay, a typical eutrophic area in the Seto Inland Sea. Microzooplankton, dominated in number by tintinnid ciliates, were abundant from June to Septem- ber when chlorophyll-a concentrations were high. Maxi- mum density of microzooplankton ranged from 3.8• to

3 1 25.4x10 ind 1- . During the period of investigation, mean microzooplankton density and mean chlorophyll-a concentration of the <20-gin fraction increased toward the inner region of the bay. The microzooplankton grazing on phytoplankton increased from summer to early autumn, and decreased from late autumn to winter. At an offshore station, the annual means of the daily grazing loss for total chlorophyll-a and the chlorophyll-a of the <20%tm fraction were 12 and 15% of the initial standing stock, respectively. At an estuarine station, the microzooplankton grazed 19 and 29% of the total and <20-~tm initial standing stock, respectively. The quantity of grazed chlorophyll- a correlated positively and linearly with the potential production of chlorophyll-a at both stations. The quantity of chlorophyll-a grazed by microzooplankton and the potential production of chlorophyll-a were nearly equivalent in the <20-gm fraction at the estuarine station. This suggests that the microzooplankton assemblage was able to con- sume almost all the nanoplankton newly produced in the eutrophic estuary.

Various field investigations and laboratory experiments have demonstrated that these ciliates can grow rapidly, oc- cur in abundance, and feed efficiently on nanoplankton (Beers et al. 1971; Taniguchi and Kawakami 1983; Gif- ford 1985; Verity 1985, 1987; Kamiyama and Aizawa 1987). Because of their abundance, they may play important roles not only as consumers of nanoplankton but also as promoters of nutrient cycling (Matsukawa 1990; Sasaki 1991). In coastal waters, microzooplankton have been observed to fluctuate considerably under the influence of the size and species composition of phytoplankton communities (Verity 1987). Evaluation of the seasonal impact of microzooplankton grazing on phytoplankton has been at- tempted using the dilution technique (Landry and Hassett 1982), but knowledge about this subject is quite limited (Gifford 1988; Paranjape 1990). Little is known about the seasonal variation of the microzooplankton community in eutrophic embayments in Japan.

In the present study the seasonal variation of the microzooplankton community was examined in relation to hy- drography and chlorophyll-a distribution in the northern part of Hiroshima Bay, a highly eutrophic embayment in the Seto Inland Sea. Simultaneously, monthly grazing impacts of microzooplankton on phytoplankton biomass were estimated using the dilution technique of Landry and Has- sett (1982) to evaluate the ecological role of the microzooplankton feeding in the estuarine and offshore areas of the bay.

Introduction

Microzooplankton have been observed to be the direct consumers of nanoplankton in most zooplankton communities.

Communicated by T. Ikeda, Nagasaki

Dr. Takashi Kamiyama Nansei National Fisheries Research Institute, Ohno-cho, Saeki-gun, Hiroshima 739-04, Japan

Materials and methods

Field investigation

Measurements and sample collections were made at five stations lo- cated in northern Hiroshima Bay (Fig. 1). Temperature and salinity were measured at four to seven discrete depths at all stations. Water samples were collected from 2-m depth with a 6-liter Van Dorn water sampler at about monthly intervals from May 1990 to November 1991. 1 liter of the seawater was immediately preserved with buf- fered formalin at a final concentration of 1 to 2% for counting microzooplankton, and other samples were brought to the laboratory in polyethylene bottles under dark conditions. These water samples

78

~ 4 ~ 2o'N

lo'

Fig. 1

132~ 30'

Location of the sampling stations in northern Hiroshima Bay

were divided into two subsamples to measure the chlorophyll-a of unfractionated seawater (total chlorophyll-a) and the chlorophyll-a of the seawater filtered through 20-gm mesh-size screen (<20-~tm chlorophyll-a). The seawater treated as above was filtered through Whatman GF/C glass-fiber filters. The filters and filtrates were frozen until chlorophyll and nutrients were analyzed. Chlorophyll-a was extracted by grinding the filters in 90% acetone and measured with a Turner Designs fluorometer (Yentsch and Menzel 1963; Holm-Han- sen et al. 1965). Nutrients (NO3, NO2, NH 4 and PO4) in the filtrates were analyzed on a TrAAcs 800 autoanalyzer (Bran Lubbe Co. Ltd.). The preserved liter of seawater was concentrated to 1 -3 ml by set- tling. In the present paper microzooplankton is defined a zooplank- ter with body width less than 200 ~tm. Counts were made under a phase contrast microscope at magnification of 100 or 200x using a Sedgwick-Rafter chamber. In the present study, heterotrophic dino- and nano-flagellates were not included in the defined "microzooplankton", since I could not distinguish the heterotrophic flagellates from autotrophic phytoplankton under the light microscope only. Tintinnids were identified to species based primarily on Kofoid and Campbell (1929), Marshall (1969), Hada (1937, 1938) and Bakker and Phaff (1976) for the genus Tintinnopsis.

Estimation of grazing rate

Grazing rates of microzooplankton on phytoplankton were estimated using the dilution technique at an estnarine station (Stn 2) and an offshore station (Stn 5) during the same period as the field investigation. 18 liters of seawater were collected with a Van Dorn water sampler and screened through a 200-gin mesh net to eliminate the net zooplankton. The treated samplewas immediately poured into a polyethylene tank protected from the light and brought to the laboratory. About half of the water sample in the tank was filtered through a Whatman GF/F filter. This filtered seawater was then combined with the remaining unfiltered seawater, and seven or eight dilution mix- tures were prepared. One treatment consisted of undiluted seawater. Dilution factors were determined by the ratio of the chlorophyll-a concentration in each dilution mixture to that in the undiluted seawater before the incubation. Nitrate (10 gM) and phosphorus (1 gM) were added to each dilution mixture to prevent potential overesti- mates of grazing due to nutrient limitation (cf. Landry and Hassett 1982; Burkill et al, 1987). l-liter, transparent plastic bottles (TPX | or polycarbonate) were filled with each dilution mixture. These bot-

ties were incubated in situ for 24 h at 2-m depth from a pier in Hi- roshima Bay. Before and after the incubation, 100-ml subsamples were taken from each dilution mixture to measure total chlorophyll- a and <20-gm chlorophyll-a. 100 ml of each subsample was filtered through a Whatman GF/F filter, which was frozen for subsequent chlorophyll analysis. At the same time, filtrate from the undiluted seawater was frozen for nutrient measurements. Chlorophyll-a and nutrients were anaiyzed by the same methods as in the field investigation. A change in phytoplankton concentration with time (t) is ex- pressed by the exponential equation following Landry and Hassett (1982):

(I/t) In (Pt/Po)=k-g, (1)

where Po and Pt are phytoplankton densities at the beginning and end of the experiment; k and g are the specific rates of phytoplankton growth and mortality due to grazing, respectively. In the experiments using the dilution technique, when a water sample is diluted, k is in- dependent of the dilution, but g is proportional to the dilution factor (predator density). Hence, the change under dilution conditions is

(l/t) In (Pt/Po)=k-x g, (2)

where x is the proportion of undiluted seawater. The values of k and g were determined from linear regression analysis between the apparent growth rate of chlorophyll-a [(l/t) In (Pt/Po)] in each bottle and the proportion of undiluted seawater (x). The significance of the regression line was tested by the Student's t-test at a level of p<0.05. Significant k and g were regarded as the growth rate of phytoplankton and the grazing rate by microzooplankton, respectively, and 95% confidence intervals were then calculated for k and g.

The dilution technique requires three assumptions: first, no nutrient limitation during the experiment; second, absence of a feeding threshold for the microzooplankton; third, adequacy of the Eq. (1). The first and second assumptions are important for the practice of dilution experiments. In this experiment, there was a possibility that nutrient limitation occurred since NO3-N concentration in the undiluted treatment after the 24-h incubation decreased below 1.0 gM in some experiments in the summer. Thus, the first assumption was evaluated by eliminating the results based on the undiluted treat- ments from the calculation of the linear regression, by recalculating k and g, and by testing for significant differences in these coefficients between the complete and altered data sets according to Gifford (1988). The second assumption was confirmed from examination of the relationship between the dilution factor and the apparent growth rate.

Results

Hydrographic condit ions

Fig. 2 shows the temperature and salinity at the 2-m depth, and the water -column stability at each station. Tempera- ture ranged from 10.5 to 26.5 ~ at all stations. The max- i m u m temperature recorded in 1990 was 0.8 to 1 .6~ higher than that in 1991. The salinity was low at Stns 1 through 4, presumably because of the inf luence of the river inflow. Sal ini ty decreased markedly in July and August , part iculary at Stn 2. The water co lumn stability, represented by the difference of water density (at) between surface and bottom, was high at Stn 1 in the estuarine area and low at Stn 5 in the offshore area. In the study area, vertical stratification began in May or April, and vertical mixing began in August or September.

Fig. 3 shows nutr ient concentrat ions at the 2-m depth. In general, dissolved inorganic ni trogen (DIN, NH 4 - N + N O 3 - N + N O 2 - N ) increased during the mix ing period

28

26

24

22

20

18

16

14

12

10

32

30

._c 28 r ~o

24

26

[] Stn 3

zx Stn 4

�9 Stn 5

18

16

14

12

4

2

0

*" 10 a= E ._m 8 o3 <] 6

79

0 0

r

a. E

i

M J J A S O N D J F M A M J J A S O N

1990 1991

Fig. 2 Seasonal variation of temperature and salinity at the 2-m depth, and of water column stability (A Sigma-t) in northern Hiroshima Bay. A Sigma-t represents the difference in density (at) between surface and bottom waters

and decreased during the stratification period. At Stn 1 it increased suddenly in May and June. DIN was higher at Stns 1 and 2 than at Stn 5 except in the summer. PO4-P also tended to be high during the mixing period and low during the stratification period at alFstations. PO4-P con-

centrations were higher at Stn 1 than at the other stations during the stratification period, while there was not much difference in PO4-P concentrations between Stns 2 and 5. The ratio of DIN to PO4-P was the highest in April and June,

80

Fig. 3 Seasonal variation of dissolved inorganic nitrogen (DIN = NHg-N+NO 3- N+NO2-N), phosphorus (PO4-P), and the ratio of DIN to PO4-P at the 2-m depth in northern Hiroshima Bay

20

16

i'i?

i O.!. ! I~. 0.4I

~ I 200

100 Z

o!

A

(

M J J A S O N D J F M A M J J A S O N

1990 1991

81

80

60

40

20

O,

6O

40

Sin

> 2 0 j J m

<20jUra

Stn 2

] ~--10 o~ < 20 p.m

"~, 6 20 pm

2 4

0 1 2 3 4 5

Station

Fig. 5 Mean chlorophyll-a of the <20-gin fraction and >20-gm fraction at the 2-m depth in northern Hiroshima Bay from May 1990 to November 1991

20 80[ N

4O V

L"~I 20 I

o 60[ , o

2O

Stn 3

Stn 4

88

60

40

20

0

Fig. 4

Stn 5

M ' J ' J A S O N D J F M A M d d ' A ' S ' O ' N ' 1 9 9 0 1991

Seasonal variation of chlorophyll-a at the 2-m depth in northern Hiroshima Bay. Filled area indicates the <20-gm fraction

Chlorophyll

Fig. 4 shows the seasonal variation of chlorophyll-a. At Stns 1 and 2, chlorophyll-a was present at high levels, above 10 gg 1-1, during much of the period. There were two peaks in chlorophyll-a values, in June and in August or September. During both peaks, total chlorophyll-a was dominated by that of the <20-gm fraction. In June 1990 and 1991 total chlorophyll-a at both stations consisted almost entirely of the <20-gm chlorophyll-a, due to blooms of Heterosigma akashiwo during these periods. At Stns 1 to 3 in July 1991, the >20-gm chlorophyll-a contributed

more than 50% to the total chlorophyll-a due to an abundance of the diatoms, Chaetoceros spp. During the winter through early spring, the total chlorophyll-a content was low, and the <20-gin fraction constituted less than 50% of the total chlorophyll-a. At Stn 5, chlorophyll-a never exceeded 10 gg 1-1. The <20-gm fraction exceeded 50% of total chlorophyll-a during June and July 1990 and during June to September 1991. Mean chlorophyll-a values during the period of investigation show that the contribution of the <20-gin fraction to total chlorophyll-a values increased toward the inner region of the bay, whereas the >20-gm fraction exceeded the <20-gm fraction at Stn 5, the outermost station iFig. 5).

Microzooplankton

Fig. 6 shows the seasonal abundance of microzooplankton. The maximum and minimum densities were observed in June through September and in January through March, respectively. The highest densities at Stns 1 to 4 ranged from 15.3• to 25.4• ind 1-1 in June or September 1990. At Stn 5 the maximum density was only 3.8• ind 1-1, which amounted to only 15% of the density at Stn 3 on the same day. The microzooplankton community was com- posed mostly of tintinnids, non-tintinnid ciliates, and nauplii. Tintinnids usually dominated during all seasons, although non-tintinnid ciliates dominated at Stns 1 and 2 in August 1990. Nauplii numerically accounted for 30% of the microzooplankton at Stn 2 in June 1990. The dominant species of tintinnids were HelicostomeIla fusiformis, H. Ionga, Amphorella quadrilineata, Eutintinnus tubulosus, Tintinnopsis beroidea, T. tubulosoides, and T. corniger. These species were abundant in June through September.

The mean densities during the investigation tended to increase toward the inner region of the bay, corresponding with the abundance of chlorophyll-a (Fig. 7). The microzooplankton density at Stn 5 was only about one-quarter to one-half those at the other, more inshore stations. Tin- tinnids constituted more than 50% of the total microzooplankton at all stations. The ratios of non-tintinnid ciliates to total microzooplankton at Stns 1 and 2 were higher than those at the other stations.

82

Fig. 6 Seasonal variation of microzooplankton at the 2-m depth in northern Hiroshima Bay

2 5

2 0

15

10

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0

Stn 1

20 Stn 2

X 0 / ~ ..... , - - , " , . . . . . . . . . . .

. ; 20 Stn 3 TinUnnld, 15 ~ Non- t in t lnn id

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0 M J J A S 0 N D J F M A M J J A S O N

1990 1991

83

Tab le 1 Growth rates (k), g raz ing rates 9) and effects on the initial s tanding s tock o f ch lorophyl l -a in nor thern H i rosh ima Bay. NS: Not s ignif icant (P_2 5 %); CI: conf idence intervals; r = correlat ion coeff icient

Stn Date Unf rac t iona ted seawater < 2 0 - g m size fract ion

Growth rate of Graz ing rate r 2 Initial s tanding Growth rate o f Graz ing rate r 2 Initial s tanding ch lorophyl l -a s tock grazed ch lorophyl l -a s tock grazed (d-1)_+95% CI (d-1)+95% CI d -1 (%) (d-1)_+95% CI (d-1)_+95% CI d -1 (%)

Stn 2

Stn 5

1990 17 May 1.53+_0.23 0.34_+0.32 a 0.60 29.1a 0.77+-0.19 0.67_+0.29 0.88 49.0 19 Jun 0 .37+0.22 NS - - 0.45-+0.07 0.39_+0.11 0.94 32.5 26 Jul 1.48+0.17 0.45+0.25 0.81 36.1 1.56+0.23 1.08+-0.33 0.93 65.9

6 Sep 0.91+-0.20 NS - - 1.20+_0.23 0.57+_0.30 0.82 43.3 26 Sep 1.13+_0.07 0.36+_0.09 0.95 30.5 0.90-+0.20 NS - - 22 Oct 0.67+_0.14 0.51+0.21 0.89 39.6 0.63_+0.14 0.48+_0.21 0.88 37.9

4 Dec 0.85+_0.12 NS - - 0.69_+0.18 NS - -

1991 16 Jan 0.76+_0.08 0.14_+0.12 0.64 13.0 0.71+_0.14 0.23_+0.22 0.58 20.6 15 Feb 0.50_+0.16 NS - - 0.12_+0.09 NS - - 13 Mar 0.50_+0.06 0.15_+0.11 0.72 13.6 0.50_+0.20 NS - - 17 Ap t 0 .59+0.10 NS - - 0.42_+0.12 0 .26+0.20 0.65 23.1

4 Jun 0.32+0.05 0.28+_0.10 0.88 24.4 0.26-+0.07 0.37+_0.13 0.89 30.8 25 Jun 0 .57+0.06 0.37+_0.14 0.88 30.8 0.58+0.05 0 .39+0.10 0.94 32.6 15 Jul 1.77-+0.21 0 .58+0.46 a 0.61 43.9 a 1.39-+0.34 1.16-+0.64 0.77 68.7

7 A u g 1.65+-0.12 1.23_+0.26 0.96 70.7 1.57+-0.09 1.39_+0.21 0.98 75.2 11 Sep 1.87_+0.07 0.52+_0.14 0.93 40.3 1.32_+0.15 0.61+_0.34 0.76 45.4 21 Oct 0.37_+0.09 NS - - 0.41_+0.14 NS - - 12 Nov 0.40_+0.10 NS - - 0.48_+0.15 0.73_+0.33 0.83 51.9 A n n u a l m e a n (4 Dec 1 9 9 0 - 1 2 Nov 1991) 19.1 b 29.0

1990 17 May 1.34+0.32 NS - - 0 .86+0.32 a 0 .55_0.45 a 0.67 42.2 a 19 Jun 0.75+0.25 0.56+0.37 0.75 42.7 0 .72+0.20 0 .79+0.30 0.92 54.7 26 Jul 1.53_+0.11 NS - - 1.50_+0.21 NS - -

6 Sep 1.15_+0.16 0.40_+0.25 0.78 33.0 0 .84+0.30 NS - - 26 Sep 0.97+_0.10 0 .20+0.16 0.68 18.0 0.72-+0.16 NS - - 22 Oct 1.05+_0.16 0.50_+0.24 0.85 39.2 0.77+_0.40 NS - -

4 Dec 0.30+_0.29 NS - - 0.34-+0.21 NS - -

1991 16 Jan 0.26+_0.09 NS - - NS NS - - 15 Feb NS NS - - NS NS - - 13 Mar 0.45+_0.12 NS - - NS NS - - 17 Apr 1.06-+0.30 0.59_+0.51 0.57 44.6 0 .70+0.26 0.55+-0.44 0.62 42.4

4 Jun 0.64+0.07 NS - - 0 .58+0.10 0.19+-0.16 0.59 17.5 25 Jun 1.04+-0.34 0.83+_0.73 0.56 56.3 0.85+_0.37 NS - - 15 Jul 1.88+_0.06 NS - - 1.38+_0.12 0.31+_0.23 0.65 26.5 7 A u g 1.67+0.12 0.31-+0.22 0.66 26.3 1.85-+0.20 0.60_+0.38 0.71 44.8

11 Sep 1.83+0.11 0.62_+0.21 0.90 46.4 1.34+_0.15 0.63+_0.30 0.81 46.6 21 Oct 0.33+_0.16 NS - - 0.31+_0.17 NS - - 12 Nov NS NS - - NS NS - -

Annua l m e a n (4 Dec 1 9 9 0 - 1 2 Nov 1991) 11.6 15.3

a Nutr ien t l imitat ion apparent; b Data for July 1990 was used for calculat ion in place o f the data for July 1991

Grazing impact of microzooplankton

F i g s . 8 a n d 9 s h o w t h e r e s u l t s o f t h e d i l u t i o n e x p e r i m e n t s .

T h e a s s u m p t i o n t h a t n u t r i e n t l i m i t a t i o n d i d n o t o c c u r , w a s

n o t c o n f i r m e d f o r t h e e x p e r i m e n t s o n t o t a l c h l o r o p h y l l - a

a t S t n 2 i n M a y 1 9 9 0 a n d i n J u l y 1 9 9 1 , n o r f o r t h e e x p e r i -

m e n t o n t h e < 2 0 - g m c h l o r o p h y l l - a a t S t n 5 i n M a y 1 9 9 0 ,

b e c a u s e t h e a l t e r e d c o e f f i c i e n t s , 9, o b t a i n e d b y e l i m i n a t -

i n g t h e u n d i l u t e d t r e a t m e n t s w e r e n o t s i g n i f i c a n t i n t h o s e

c a s e s . H e n c e , t h e s e c o e f f i c i e n t s w e r e r e g a r d e d a s t h e r e -

s u l t o f n u t r i e n t l i m i t a t i o n . T h e o t h e r a s s u m p t i o n t h a t a f e e d -

i n g t h r e s h o l d d i d n o t o c c u r w a s r e a l i z e d i n a l l c a s e s ( F i g s .

8 a n d 9) .

B a s e d o n t h e c o e f f i c i e n t s , k a n d 9, d e t e r m i n e d e x p e r i -

m e n t a l l y , t h e i m p a c t o f m i c r o z o o p l a n k t o n g r a z i n g o n t h e

5 A

4 _= n o

8 2 e - -

"o c = 1

0 1 2 3 4 5

Station �9 Tlntinnlds [ ] Non-tintlnnld clllates [ ] Nauplli [ ] Others

Fig. 7 M e a n abundance o f mic rozoop lank ton at the 2-m depth in nor thern H i rosh ima Bay. Values represent m e a n abundance o f mi- c rozooplankton f rom M a y 1990 to N o v e m b e r 1991

84

2

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21 O c t 1881

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0.2 0.4 1 0 0.6 o12 0.4

12 NoV 1881

0

1 0 012 014 0'.6 0~8

Fraction undiluted seawater

Fig. 8 Relationships between apparent growth rate (based on chlorophyll-a) in unfractionated seawater (filled circles and continuous line) and in the <20-1~m size fraction (open circles and dotted line) at Stn 2 from May 1990 to November 1991. Lines fitted by linear regression

2

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Frac t ion u n d i l u t e d s e a w a t e r

21 Oct 1991

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12 Nov lg91

( ~ 0 0 C O � 9 0 0 o

m 0 0 m 0.2 oA o'.a o'.8

F i g . 9 R e l a t i o n s h i p s b e t w e e n a p p a r e n t g r o w t h r a t e in u n f r a c t i o n a t e d s e a w a t e r ( f i l l ed c i r c l e s and c o n t i n u o u s l i ne ) and in the < 2 0 - g m s ize f r a c t i o n (open c i r c l e s and d o t t e d l ine ) at S tn 5 f r o m M a y 1990 to N o v e m b e r 1991. L i n e s f i t t ed b y l i n e a r r e g r e s s i o n

86

initial standing stock was calculated at both stations (Ta- ble 1). At Stn 2, growth coefficients for total chlorophyll- a and for the <20-gin chlorophyll-a ranged from 0.32 to 1.87, and from 0.12 to 1.57, respectively. Grazing coefficients for total chlorophyll-a and for the <20-gin fraction were ~_1.23 and ~1.39, respectively. These levels were low during the winter-early spring period, and high during the summer-early autumn period. The grazing coefficients for < the 20-gin fraction were usually larger than those for total chlorophyll-a. The daily loss in the initial standing stock of chlorophyll-a due to grazing reached 71% for total chlorophyll-a and 75% for the <20-gin fraction chlorophyll-a, respectively. The annual mean (December 1990 to Novem- ber 1991) daily consumption of total chlorophyll-a and <20-gm chlorophyll-a was estimated to be 19 and 29%, respectively, using the assumption that non-significant data are equal to 0.

At Stn 5, the maximum growth coefficients for total chlorophyll-a and for the <20-gm chlorophyll-a were 1.88 and 1.85, respectively. The grazing coefficients, 9, were ~_0.83 and ~_0.79, respectively. These values were low com- pared to those at Stn 2. The grazing coefficients, 9, for the <20-gm chlorophyll-a were insignificant during the autumn-winter period. The maximum daily loss in the initial standing stock of total chlorophyll-a and <20-gm chlorophyll-a was 56 and 55%, respectively. Furthermore, the annual mean daily consumption of total chlorophyll-a and <20-gm chlorophyll-a was 12 and 15%, respectively, which was less than at Stn 2.

Discussion

Since Hiroshima Bay receives the inflow of several rivers in the northern region, Stns 1 and 2 are influenced by fresh water, resulting in the relatively low salinity at these stations (Fig. 2). DIN and PO4-P concentrations were highest at Stn 1 throughout the year, but among the other stations the concentrations of these nutrients were not clearly different. This can be explained by the information that nutrients supplied by river water are rapidly taken up by phy- tgplankton in the vicinity of the river mouths in Hiroshima Bay in the summer (Yuasa et al. 1990). The high ratio of DIN to PO4-P in March through June at Stns 1 and 2 is due to the paucity of PO4-P despite the high levels of DIN during this period. High N/P ratios in the spring have been reported for an estuary of the Chesapeake Bay (Fisher et al. 1992).

During the period os the mean level of chlorophyll-a in the <20-gm fractions increased toward the inner region of the bay (Fig. 5). This also suggests the influence of river inflow on the inner stations of the bay. Ma- lone (1977) reported that nanoplankton growth rates increase more rapidly with increasing DIN than do net-phytoplankton growth rates. Yamochi (1989) showed that trace metals are necessary to increase the yield of Heterosigrna akashiwo, a representative nanoplankter in Hiroshima Bay. Accordingly, the DIN and/or trace metals in river water

may affect the size composition of phytoplankton at each station.

The fluctuation of microzooplankton was similar to that of chlorophyll-a. In particular, tintinnids, the dominant group of microzooplankton, were abundant at the inner stations where the <20-gm chlorophyll-a values were high. This suggests high growth ability of tintinnids in eutrophic embayments. In addition, the microzooplankton were generally abundant during the stratification period. Reverante and Gilmartin (1983) showed that microzooplankton stocks increased ten-fold in abundance, and two-fold in biomass, from the mixed to the stratified period. The abundance of microzooplankton during the stratification period may be due to an increase during the same period of nano- phytoplankton which are satisfactory food for the microzooplankton.

Non-tintinnid ciliates represented by oligotrichs are generally more abundant than tintinnids by a factor of 2 to 10 (Pierce and Turner 1992). However, the tintinnids occurred more abundantly than non-tintinnid ciliates in Hiroshima Bay. Similarly, a higher density of tintinnids than non-tintinnid ciliates is recognized in the eutrophic coastal area, Solent Estuary on the southern English coast and Tokyo Bay in Japan (Burkill 1982; Nomura et al. 1992). It was prob- ably essential for this phenomenon that nano-phytoplankton dominated phytoplankton assamblage in these eutrophic areas.

Recently, heterotrophic dinoflagellate have been fo- cused in microzooplankton assemblage (Hansen 1991;Ve- rity et al. 1993). In Hiroshima Bay, <20-gin chlorophyll-a was much abundant in the estuarine stations, suggesting the possibility that this group was much abundant and grazed on nano-phytoplankton in large quantities in the certain period of summer. Unfortunately, there are no data about abundance of heterotrophic dinoflagellates in this study. Further studies are needed to estimate biomass and the grazing impacts of heterotrophic dinoflagellates in this area.

In the present investigation, the maximum densities of microzooplankton were recorded at all stations in June 1990 when a bloom of Heterosigma akashiwo occurred in the inner region of the bay. However, tintinnids generally reject this species as food (e.g. Taniguchi and Takeda 1988). Furthermore, Olisthodiscus luteus, somewhat similar to 11. akashiwo, repressed the growth of tintinnids (Ver- ity and Stoecker 1982). In the present study, the abundance of microzooplankton decreased markedly at Stns 2 through 4 in June 1991 when a H. akashiwo bloom occurred (Fig. 6), an observation inconsistent with that in June 1990. Nielsen (1990) reported that ciliate densities were low in a layer where the noxious flagellate Chrysochromulina polylepis was abundant and then increased as the bloom dissipated. Furthermore, Hansen (1989) and Carlsson et al. (1990) observed that the growth phases or growth conditions of noxious flagellates influenced the degree of inhi- bition of microzooplankton. Similarly, in the present investigation the difference in abundance of microzooplankton between June 1990 and June 1991 may be ascribed to differences in growth phases and/or growth conditions of H. akashiwo.

The impact of microzooplankton grazing using dilution experiments have been studied in several localities. In coastal waters off Washington, USA, 6 to 24% of the phytoplankton standing stock was consumed daily by microzooplankton (Landry and Hassett 1982). In the Celtic Sea, the impact of microzooplankton grazing ranged from 13 to 42% of the standing stock of phytoplankton within the ther- mocline in the summer and from 30 to 65% in the autumn (Burkill et al. 1987). In Jones Sound and in central Baffin Bay, the microzooplankton community removed daily 8 to 15% and 9 to 15% of the initial standing stock in the summer, respectively (Paranjape 1987). In Halifax Harbor, Canada, the daily consumption of the phytoplankton crop by microzooplankton ranged from 0 to 50% in five experiments during the year. These values are in general agree- ment with the present estimates, which range from 0 to 71% at Stn 2 and from 0 to 56% at Stn 5. However, in the present study, the grazing impact at Stn 2 was estimated under high chlorophyll-a conditions, exceeding 10 gg 1-1 from June through October. There are only a few studies in which dilution experiments were conducted under such high chlorophyll conditions (Gallegos 1989; McManus and Ederington-Cantrell 1992).

In order to analyze the relationship between chlorophyll production and grazing impact, the potential production of chlorophyll (PPC) and the actual production of chlorophyll (APC) were calculated for these dilution experiments using the methods of Gifford (1988); grazed chlorophyll (GC) was obtained by subtracting APC from PPC. The re-

120

100 Stn 2 / ,~i

so ........ 9 " / / / / / Y=-1.73+O,93X / "

60 r~-o.s7 .... / / /

.//"//Y=..-5.OO+O.81X 40 / ~ r"=O.Se

o , , ,~= 0 20 40 60 80 100 120

aO 2 Stn 5 o 25 5

O 15 ............ �9 ............

I- Y=.O 5~ +o.ssx - o ....... [ ,"=o.ga ..~ '

l O [ ~. ................

5 �9 o " � 9 1 7 6 vo,

0 ~ - = ' ~ ' r I 0 5 10 15 20 25 30

1 1 Potential production chlorophyll (l~g Iday" ) Total chl~.orophyll < 20 pm.~h!orophyll

Fig. 10 Relationships between potential production of chlorophyll and grazed chlorophyll Lines fitted by linear regression

87

lationships between PPC and GC at both stations are shown in Fig. 10. At Stn 2, GC increased with increasing PPC, and GC for the <20-gm chlorophyll exceeded that for the total chlorophyll at the same PPC level. In particular, the GC value rose linearly with PPC, and the slope of the line fitted by linear regression was significant at the 5% level. The slope for the <20-gm chlorophyll was nearly 1. These results demonstrate that the chlorophyll production of the <20-gm fraction was mostly consumed by microzooplankton. A daily grazing loss equivalent to the potential production of chlorophyll has been reported in the other stud-. ies (Gifford 1988; Burkill et al. 1993). In the present experiments, this balance between GC and PPC for the <20:gm fraction was observed frequently. This notable feature may relate to the high chlorophyll levels and abundant microzooplankton in eutrophic embayments.

At Stn 5, the relationships between PPC and GC for total chlorophyll was also significant at the 5% level (Fig. 10), but the slope for total chlorophyll was lower than that for the <20-gm chlorophyll, and for total chlorophyll undetectable amounts of GC were observed at relatively high levels of PPC (>10 gg 1-1 d-l). The maximum values of PPC and GC at Stn 5 are less than one-quarter of those at Stn 2, for the respective fractions of chlorophyll. Fur- thermore, the slopes of linear regressions between PPC and GC for total and <20-gm chlorophyll are only 37 and 60% of those at Stn 2, respectively. These results indicated a re- duced efficiency of consumption of phytoplankton at Stn 5 relative to Stn 2.

In the inner region of the bay, influenced by river inflow, features of the hydrographic conditions and plankton community are the following: high concentration of nutrients, high level of chlorophyll dominated by the <20-gm size fraction, and high density of microzooplankton con- sisting mainly of tintinnids. The microzooplankton af- fected the nanoplankton communities during the period of phytoplankton abundance. Based on these results, primary production due largely to nanophytoplank- ton may be linked efficiently to secondary producers, which are predominantly microzooplankton. Although the food of microzooplankton was considered as phytoplankton in the present study, recent works showed that an important taxon of microzooplankton, oligotrich, can efficiently ingest bacteria. (Fenchel and Jonsson 1988; Ras- soulzadegan et al. 1988). Verity (1991) indicated that ciliates (oligotrichs and tintinnids) can ingest aplastidic (=nonpigmented) nanoplankton as well as phytoplankton. Further study is necessary to estimate the role of microzooplankton in planktonic and microbial food webs in eutrophic embayments.

Acknowledgements I thank Dr. A. Tsuda for giving me helpful sug- gestions. I am grateful to Drs. K. Kawaguchi, M. Terazaki, S. Nish- ida, and T. Honjo for a critical reading of this manuscript. Thanks are also due to Dr. K. Tamai, K. Tanaka, M. Itaoka and M. Gotoh for invaluable assistance with the field investigations. This work was partly supported by a grant from the Fisheries Agency, Ministry of Agriculture, Forestry and Fisheries, Japan.

88

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the impact of grazing by microzooplankton in northern hiroshima bay, the seto inland seam, japan

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