Seasonal growth, δ13C in leaves and stem, and phloem structure of birch (Betula pendula) under low ozone concentrations

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Trees (1992) 6:69-76

9 1992

Original articles

Seasonal growth, 513C in leaves and stem, and phloem structure of birch (Betula pendula) under low ozone concentrations

Rainer Matyssek l, Madeleine S. Giinthardt-Goerg l, Matthias Saurer 2, and Theodor Keller 1 1 Swiss Federal Institute of Forest, Snow and Landscape Research, Ztircherstrasse 111, CH-8903 Birmensdorf, Switzerland 2 Physics Institute, University of Berne, Sidlerstrasse 5, CH-3012 Berne, Switzerland

Received April 2, 1991/June 26, 1991

Summary. The growth of potted birch cuttings (one clone of

B e t u l a p e n d u l a )

was studied under low 03 concentra- tions (0, 0.050, 0.075, 0.100 ~tl 1-1) throughout an entire growing season. With increasing 03 dose, 2 0 - 5 0 % o f all leaves formed were prematurely shed, while 4 0 - 7 0 % of the remaining foliage displayed advanced discoloration by the end of the season. Ozonation affected the S, P and N concentration of leaves and increased ~13C in leaves and stem, while the CO2 assimilation rate declined with in- creasing CO2 concentration in mesophyll intercellulars.

While whole-plant production correlated negatively with the 03 dose, ozone increased the specific leaf weight (i. e.

leaf weight/leaf area, SLW) but decreased the ratios of stem weight/stem length and root/shoot biomass. Neither the latter ratio nor SLW changed in experimentally defoliated control plants, whereas in ozonated plants starch accumulated along leaf veins and phloem tissue was deformed in the leaf petioles and the stem. Only in early summer was the relative growth rate higher in the ozonated than in the control plants. The ratio of whole-plant biomass production versus total foliage area formed was lowered under 03 stress. However, when relating biomass to the actual foliage area present due to leaf loss, this ratio did not differ between treatments. Similarly the ratio of actual foliage area versus basal stem area in cross-section did not differ. Overall, whole-plant production was strongly deter- mined by O3-caused changes in crown structure and began to be limited at 03 doses (approximately 180 ~tl 1-1 h) similar to those o f rural sites in Central Europe.

Key words: Ozone -

B e t u l a p e n d u l a -

Growth analysis -

~13C - Phloem structure

Introduction

Environmental factors are known distinctly to change the carbon allocation in trees (Schulze and Chapin 1987), but

Offprint requests to:

R. Matyssek

are non-detrimental as long as the tree metabolism copes with the stress (Waring 1987). While the understanding of mechanisms predisposing trees to injury is growing in rela- tion to some natural stress factors, knowledge concerning the seasonal impact of low concentrations of air pollutants is limited (Winner et al. 1988). Trees with mesophytic leaves are thought to be prone to high pollutant uptake (Reich 1987). A recent study with mesophytic

B e t u l a p e n d u l a

showed that under low 03 concentrations the de- cline o f carbon uptake and water-use efficiency of leaves was coupled with the gradual collapse of tissue structure (Matyssek et al. 1991). The relevance o f such 03 effects for tree growth can only be established through relating them to the production and carbon allocation in the whole plant (Mooney and Winner 1988). Even though the 03 impact on the foliage seems to be coupled with favoured above- ground versus below-ground carbon allocation at declining whole-plant production, findings do not necessarily repre- sent the low 03 stress o f many rural regions (Reich 1987).

Findings on the 03 effect on carbon allocation conflict (cf.

Reich and Lassoie 1985), as the developmental state of plants at the harvest time may bias growth analysis.

Aiming to understand the seasonal 03 impact on the whole plant (as requested by Mooney and Winner 1988), the present study follows the biomass development throughout the entire growing season of the same birch individuals, which in parallel were investigated at the leaf level (Matyssek et al. 1991). Mechanisms o f plant response are addressed as to whether gas exchange and carbon allo- cation under low 03 concentrations are associated with changes in 513C of leaves and stem (Farquhar et al. 1989) and in phloem structure (cf. Spence et al. 1990), and how partial defoliation in O3-free air affects carbon allocation relative to O3-caused leaf loss.

Materials and methods

Plants and treatments.

From 14 April through 2 October 1989, cuttings

of one birch clone

(Betulapendula,

Roth) were grown in 10-1 pots filled

with sand and a basal layer of inert synthetic clay beads (1 plant/pot,

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Table 1. Nutrient concentrations of birch leaves from mid-stem positions in August 1989 (~tg g I; 5 trees per 03 treatment) 03 concentration of fumigation (~tl 1-1):

0 (control) 0.050 0.075 0.100

Nutrient:

Ca 10667+_1 914 9504+_1 100 10461 --+848

K 17950___ 606 16 149+_t 288 17891 +_396

Mg 3485___ 196 3432+- 315 3498_+292

Fe 64--+ 12 51-+ 4 69-+ 11

P 2655-+ 268 2846-+ 212 2977-+367

S 1671-+ 311 1863_+ 337 1847+_440

10(107_+ 628 17 251 _+ 1343 3190+_ 111 5 5 + 3 3237+_ 471 1918+_ 341 Values in italics differ significantly from control at 5% according to Wilcoxon u-test

fertilized, well-watered). When transferred into the field fumigation chambers on 16 May 1989, the plants (shoot 3 cm long) were separated into four 03 treatments (20 plants/treatment, 4 plants/chamber). The 03 concentrations were 0 (control), 0.050, 0.075, and 0.100 ktl 1-1, and were monitored by a Monitor Labs 8810 instrument. Ozone was generated from pure oxygen (Fischer, model 502) and continuously added to char- coal-filtered air. Plants, fumigation chambers and O3 regimes were the same as described in Matyssek et al. (1991; see also Landolt et al. 1989).

On 27 July 1989, when the control had reached about one-quarter of its final biomass (see Fig. 4A), five further control plants were partially defoliated by excising 66% of all leaves on stem and branches (beginning at the basal axis ends). This treatment is denoted as defoliated control (DC plants).

Biomass analysis. On t7 July, t5 August, and 3 October (before autumnal discoloration), 5 - 1 0 individual trees in each 03 treatment were harvested (the 5 DC plants on 3 October) for the biomass analysis of the whole plant. The actualfoliage area was determined from one-sided area measurements of all leaves attached at each harvest (Delta-T areameter MK2), while the potential foliage also includes the shed leaf area. The latter was calculated from the missing leaf number and the mean leaf area on both stem and branches. Basal stem area in cross-section was calculated from diameter measurements (with calipers) at 10 cm above the stem basis. The plant organs were dried at 65~ to constant weight (4 days). The ratio of root/shoot biomass comprises only plant organs developed from the originally planted cutting (about 13 cm long, 0.6 g in April); otherwise the cutting increment is part of the whole-plant dr), weight. The relative growth rate (RGR) is derived from the mean whole- plant biomass in a treatment at each harvest as AWI A t * (W) -I, where A W is the increment of mean biomass during the time interval A t between two harvests; W = (Wl+W2)/2 is calculated from the mean biomass at the beginning (W0 and at the end (W2) of the interval At. The efficiency of production in terms of foliage availability was estimated by basing whole-plant biomass either on the actual or the potential foliage area at each harvest (reciprocal of leaf area ratio). The specific leaf weight (SLW) of the foliage is given as dry weight/area of the actual (= attached) foliage.

The extent of O3-induced discoloration in the attached foliage was determined by counting and assigning leaves to three classes of visual injury: (1) no symptoms, (2) early symptoms (little, light-green yellowish or black dots), and (3) established yellowish-bronze discoloration including large necroses.

The concentration of cations, S and P in leaves from similar stem position was assessed by ICP-AES (inductive coupled plasma-atomic emission spectroscopy) ICP, that of N with a Carlo Erba NA1500 analyser.

Gas exchange experiments. These were conducted on attached complete leaves with a thermo-electrically climate-controlled cuvette system (Walz) as described by Matyssek et al. (1991). The steady-state rates of net CO2 uptake and transpiration were determined after 90 min of constant cuvette conditions (see legend of Fig. 2).

813C analysis. A 50 mg aliquot was prepared from the ground total stem axis of each tree. Cellulose was separated from wood (Brenninkmeijer 1983) by the following steps: Soxhlet extraction with toluene-enthanol azeotrope, delignification with acidified NaC102 solution at 70~

(3 times 8 h), incubation in NaOH (4%) and washing in distilled water (24 h, 70 ~ C). Heating of the cellulose samples occurred at 950 ~ C (2 h), and the isotopic composition of the released and purified CO2 was determined with the mass spectrometer MAT 250 (Finnigan MAT) as described by Becker et al. (1989). 813C values of the samples were calculated by comparing the relative abundance of 13C with that in the PDB standard (Craig 1957). The 813C of the leaves investigated by gas exchange experiments (see Matyssek et al. 1991) was determined without preceding cellulose extraction.

Light microscopy. At each harvest date, discs were excised in the afternoon from two fully developed leaves at the stem of each tree (disc diameter = 8 mm; 2 discs each from the central left and right half of one lamina). After fixation and bleaching in hot methanol the starch in these samples was stained with JJK solution. Phloem tissue was investigated in samples cut from the central part of the leaf petiole and from the mid- position of the stem internode underneath that leaf. Sections (2.5 gun thick) were prepared after sample fixation in glutaraldehyde solution (2.5%, at 5 ~ C) and embedding in Technovit 7000. Acid fuchsin and toluidine blue solution were used for staining.

Results

N u t r i e n t a n d c a r b o n relations o f leaves

T h e c a t i o n c o n c e n t r a t i o n s o f l e a v e s d i d n o t c l e a r l y r e s p o n d to o z o n e ( T a b l e 1), w h e r e a s P a n d S t e n d e d to i n c r e a s e w i t h t h e 0 3 c o n c e n t r a t i o n . C o m p a r e d w i t h t h e l e a v e s f o r m e d in s p r i n g ( F i g . 1 A , l o w e r s t e m ) , N o f l e a v e s f o r m e d in s u m - m e r i n c r e a s e d in p a r a l l e l w i t h t h e l e a f a r e a in all b u t t h e 0.1 ~tl 1-1 0 3 t r e a t m e n t (Fig. 1 A , u p p e r s t e m ) . H o w e v e r at 0.1 ~tl 1-1, S L W w a s r a i s e d i n the s u m m e r l e a v e s ( F i g . 1 B).

B y A u g u s t , m a i n l y t h e ( o l d e r ) l e a v e s at t h e l o w e r s t e m h a d d e v e l o p e d v i s u a l 0 3 i n j u r y a n d d e c l i n e d in p h o t o s y n t h e t i c c a p a c i t y , d e p e n d i n g o n t h e 0 3 d o s e ( F i g . 2; s e e M a t y s s e k et al. 1991). T h i s d e c l i n e w a s c o u p l e d w i t h a r a i s e d ci/ca r a t i o (i. e. C O 2 c o n c e n t r a t i o n o f t h e m e s o p h y l l i n t e r c e l l u l a r s p a c e s v e r s u s that o f a m b i e n t air; F i g . 2), w h i l e t h e 813C o f t h e O 3 - i n j u r e d l e a v e s w a s h i g h e r ( - 3 1 . 5 + 0 . 4 % 0 ) t h a n in t h e c o n t r o l (-34.4___ 0.47oo). V i s u a l 0 3 i n j u r y s p r e a d f r o m t h e l o w e r s t e m , a n d b y S e p t e m b e r a b o u t t w o - t h i r d s o f t h e f o l i a g e d i s p l a y e d a d v a n c e d d i s c o l o r a t i o n in t h e 0 . 0 7 5 a n d 0.1 g l 1-1 t r e a t m e n t s ( T a b l e 2).

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71

120-[

1 0 0 1 A

~ 8oj

~ 60

4 0

20 0 ._~ a'-

~ o ~

5

Betula pendula

- - T r 1

Aug. 1989 T - -

+

r + ' r ~ _ ~ ; r

._r ~ Ozone (i.tl I~): +

~ .

4 .

co o 0 (control) 9

0.075

co 9 0.050 9 0.100 9 Iowerstem

3. r 1 T r

0 1 2 3 4 5

Nitrogen concentration

(%)

Fig.

1. A Leaf area, and B specific leaf weight (i. e. leaf weight/leaf area) of single leaves in relation to the nitrogen concentration. Two leaves each from the lower stem (formed in spring) and the upper stem (formed in summer) were analysed per plant. Thus, data points are means -+ SD of 10 leaves (from 5 plants) per treatment and stem position (little dots beside symbols mark leaves from lower stem; symbols without dots represent upper stem)

B i o m a s s d e v e l o p m e n t . T h e m e a n w h o l e - p l a n t b i o m a s s p r o - d u c e d in e a c h 0 3 t r e a t m e n t b y O c t o b e r d e c l i n e d l i n e a r l y w i t h the 0 3 d o s e a c c u m u l a t i n g o v e r the g r o w i n g s e a s o n (Fig. 3). R e m a r k a b l y , the p r o d u c t i o n o f this b i r c h c l o n e was i n h i b i t e d b y a s i m i l a r 0 3 d o s e to that o f the a m b i e n t air o f the e x p e r i m e n t a l site. T h e a n n u a l c o u r s e o f g r o w t h r e s u l t i n g in the r e l a t i o n o f Fig. 3 d i f f e r e d s t r o n g l y b e t w e e n t r e a t m e n t s (Fig. 4 A ) . W h i l e the b i o m a s s o f the c o n t r o l i n c r e a s e d t h r o u g h o u t the e x p e r i m e n t , p r o d u c t i o n o f the o z o n a t e d p l a n t s m o s t l y s t a g n a t e d as the f o l i a g e a r e a d e - c l i n e d after m i d - s u m m e r (Fig. 4 A , B). T h e d e c l i n e resulted f r o m the p r e m a t u r e loss o f O3-injured l e a v e s (Fig. 4 C ) . I n O c t o b e r , the D C p l a n t s d i s p l a y e d a b i o m a s s and f o l i a g e a r e a s i m i l a r to the 0.05 g l 1-1 0 3 treatment, al- t h o u g h the p r o p o r t i o n a l l e a f loss e q u a l l e d that at 0.1 g l 1-1 (Fig. 4 A - C ) .

O z o n e n o t o n l y r e d u c e d p r o d u c t i o n b u t also c h a n g e d the c a r b o n a l l o c a t i o n : S L W o f the a t t a c h e d (= actual) f o l i a g e w a s e n h a n c e d b y O c t o b e r in all 0 3 t r e a t m e n t s (Fig. 5 A ; cf.

Fig. 1B); the w e i g h t / l e n g t h ratio o f the stem and the r o o t / s h o o t b i o m a s s ratio were, h o w e v e r , h i g h e r in the c o n - trol (Fig. 5 B, C). T h e s e d i f f e r e n c e s in a l l o c a t i o n w e r e es- t a b l i s h e d d u r i n g the s e c o n d h a l f o f the season. I n the D C plants, o n l y the w e i g h t / l e n g t h ratio o f the s t e m s h o w e d a s i m i l a r d e c r e a s e to that u n d e r the 0.05 g l 1-1 0 3 r e g i m e (Fig. 5). A t the g i v e n O 3 - i n d u c e d l e a f loss (cf. Fig. 4 B , C), the d e c r e a s e d r o o t b i o m a s s r e l a t e d to p r o p o r t i o n a l l y m o r e f o l i a g e a r e a in o z o n a t e d than in c o n t r o l p l a n t s ( i n c l u d i n g D C plants; Fig. 6 A ) . I n parallel, the r o o t b i o m a s s was m o r e s t r o n g l y l i m i t e d t h a n that o f s t e m a n d b r a n c h axes, as S L W o f the a t t a c h e d f o l i a g e i n c r e a s e d w i t h the 0 3 c o n c e n t r a t i o n (Fig. 6 B ) .

T h r o u g h o u t the e x p e r i m e n t , the ratio o f a c t u a l (= attached) f o l i a g e a r e a to b a s a l s t e m a r e a in c r o s s - s e c t i o n

Betula pendula

1.0

0 . 9 - 0 . 8 - 0.7

0

June-Aug. 1989

1 i i.ll I I

Ozone (Lal): c 0 (control) both

ages

o

)

E 2 9 == 0.050 11-

week-old

0.075

~ =d~=" 9 ~ 9 0.100. 5-week-old

r 9 ~ O O O

o~- . . . ;~176 ~2~

10 20 30 40

510 60

CO 2 assimilation rate. A,.b (mg g~ h -~)

Fig. 2. Ratio of the CO2 concentration in the mesophyll intercellular spaces of the leaf, ci (cf. Farquhar and Sharkey 1982), versus that in the ambient air, Ca, as related to the CO2 assimilation rate per unit of single- leaf dry weight Aa m b (at Ca = 340 gl 1-1). Steady-state response after 90 min of single leaves from the lower stem to constant light intensity (>1200Hmol photons m -2 s-l), leaf temperature (20~ and leaf/air difference in vapor mole fraction (10 mmol mol-1); each data point represents one plant. The 03 dose (84 gl 1-1 h) in 5-week-old leaves at 0.100gl 1-1 was similar to that in ll-week-old leaves at 0.050 gl 1-1 (92 gl 1 -] h) but 139 gl 1 -] h for ll-week-old leaves at 0.075 gl 1 -] (cf.

Matyssek et al. 1991)

Betula pendula

1989

150 / ~ ~ .

A I O~ doses

12o ~. I I

~ 1 ~ near Zurich

- " . 5 ~ 60 o ~ z

$'=-

9 0.050

~ 3

0.075

9

0.100

I I I

100

200 300 400

Ozone dose (i.tl Ph): May-Oct.

Fig. 3. Whole-plant dry weight in early October (before autumnal leaf loss) in relation to the 03 dose of the growing season (means + SD of 7 - 10 plants/O3 treatment). Horizontal bar marks the range of the 03 doses in the ambient air of the experimental site and in the nearby rural vicinity of Zurich (see Matyssek et al. 1991)

Table 2. Leaves with visual 03 injury in relation to the total number of leaves attached to a tree (%); values given as means _+ SD of 5 trees/O3 treatment in September

03 concentration of fumigations (H11-1)

0 (control) 0.050 0.075 O. 1 O0

Symptoms:

(l)None 100___0 9-+-4 5--+ 4 1-+ 2

(2) Early - 50-+8 33-+11 30-+15

(3) Established - 42 -+ 7 62 -+ 12 69 -+ 14

discoloration

The three classes of 03 injury are: (1) no visual symptoms, (2) early visual symptoms (little, light-green yellowish or black dots), (3) established yellowish-bronze discoloration including large necroses

(4)

72

Betula pendula 1989

150 , , j

Ozone (ixl I1):

m 120-

"~ A o 0 (control) /

.~m 90- D 0 (DC) / T

x~ 9 0.050 ~ ~

:O:Oo, 1

T- ao- A

0 i I i

| B

,9.o 0,300

0.100

0 i I

C

= ~v 6 0 T

"~ N 40

~ 2 0

0 9

J A S O

Fig. 4. Seasonal courses of A whole-plant dry weight, B foliage area attached, and C proportional leaf loss of the total leaf number formed.

Data points are means __ SD of 5 - 10 plants/treatment at each harvest. At

points without bars,

SD falls within symbol size; treatment symbols are grouped around each harvest date for graphical reasons.

Straight lines

connect data populations with either no or strongest response to ozone

6')

E o ~

.~|

~[u.. 2-

Betula pendula 8

A total crown

0

.~

~ 180-

~ 12o-

"10 - - O'J

6o-

0

I I I

Ozone (ixl I1): o 0 (control) 9 0.050 0 (DC) 9 0.075 9 0.100

i r I T

t

~ 1989

I

o.,. o

0.0 J ' A ' S ' ' 0

Fig. 5. Seasonal courses of A dry weight/area of the attached foliage (SLW), B stem dry weight/stem length, and C the biomass ratio of the root versus the shoot (data points, arrangement of symbols and straight lines as in Fig. 4)

(Fig. 7 A) did not differ among all treatments (including DC plants), nor did the ratio of whole-plant biomass versus actual foliage area (Fig. 7B; see Materials and methods).

This is remarkable, as growth (reflected in biomass and stem diameter) integrates over time, but actual foliage area is accidentally determined by the time of O3-caused leaf loss. However, when relating the biomass to the potential foliage area, this ratio was reduced in ozonated and DC plants (Fig. 7C). This latter ratio reflects carbon invest- ments into green tissue, which did not pay off due to the reduced life span of leaves. The chronic 03 exposure of the foliage was recorded in the cellulose of the stemwood by increased 813C (Fig. 7 D).

During summer, R G R was first enhanced in ozonated plants (Table 3) but was then strongly depressed relative to the control. For the total period observed ( J u l y - O c t o b e r ) , R G R of DC plants was about the same as for the 0.075 and 0.100 ktl 1-1 03 regimes but lower than in the control and 0.05 pl 1-1 treatments (Table 3).

P h l o e m structure

When leaves were sampled in the afternoon, starch was found accumulated along the veins of leaves with early visual 03 injury (Fig. 8A) but not in the control (Fig. 8B).

The phloem tissue of ozonated plants was deformed in the

petioles of leaves with established discoloration (Fig. 8 C) and in the stem axes (Fig. 8 E; see corresponding controls in Fig. 8 D, F).

D i s c u s s i o n

H o w does ozone determine whole-plant production? Total

foliage area and arrangement of leaves usually more

strongly define undisturbed plant growth than does the

CO2 assimilation rate, A, as based on leaf weight or area

(Gifford 1974; Lange et al. 1987; Matyssek and Schulze

1987). When compensating ozonated birches for prema-

ture leaf loss and reduced A (Fig. 9, step 1, see legend), a

similar whole-plant dry weight is calculated for the 0.05 ktl

1-1 03 treatment to that in the control. However, for the

other 03 treatments, increases in branch number (step 2)

and mean leaf size (step 3) are additionally required to

obtain the production of the control. Obviously, the

changed crown structure (reduced number of branches, leaf

size and premature leaf loss) due to ozone more drastically

limits growth than does the decline in A, as the latter is only

part of step 1 and strongly varies between leaves, depend-

ing on leaf age and thus 03 dose (Table 2, Fig. 2; Matyssek

et al. 1991). The above-mentioned principles determining

undisturbed plant growth apparently also hold for 03

stress.

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Betula pendula 500

E A o 4 0 0 -

•

300

~_ 200

0 0

Q)

0 3 "10 0

t - r l - i ~

I 0 9

Ozone (p.I I"): ^o0 (control)

o (DC)

"~'~: 5 9 0.050

9 9 0.075

9 . = 9 * 0.100

October,1989

o

A

i i / i

10 2O 3 0 40

Root dry weight (g)

I i

o

".,.o o

cl:] D O 9

" .

o 9 O o

d m 9 9

o

9

2 4

Foliage dry weight / foliage area (mg/cm 2)

100 75

50

25

0 0

50

Fig. 6. October harvest, A ratio of foliage area versus root dry weight in relation to root dry weight, and B ratio of root by the sum of stem and branch dry weight (without leaves) in relation to SLW (dry weight/area) of the attached foliage

Nevertheless, the impact of

0 3 o n

A was well reflected in disturbed leaf gas exchange and raised ~13C of leaves and stems. This change in 813C seems to be a bioindication of even low stress by air pollutants (Becker et al. 1989;

Greitner and Winner 1988) and to provide a long-term stress record in the wood formation (Martin et al. 1988).

However, the increasing

~13C was

not related in the birch leaves to decreasing ci, as might be expected from stomatal closure due to air pollutants (Farquhar et al. 1989). We found a more rapid decline in A than in stomatal conduc- tance (Matyssek et al. 1991), resulting in raised ci (Fig. 2).

Thus, the raised ci may indicate that 813C was less affected by stomatal limitation than perhaps by changes in carbon fixation (e. g. by raised PEP-carboxylase activity relative to

73 E E 150

~ 1 2 0

v

t~ ~ 9 0

:;oo.

g ' ~ 3o-

O ~

300.

f,, ~

250.

~= o 2 0 0

~-.~

=_ I~_ 15o

2:

*6 ~ 100.

5 0 E 300 250 J

2~176

~ 150 1

,-Z ,ool

"6 504

"~ -26

~- ~ - 2 8

Betula pendula

I

A

-30

1989 9 TT Ozone (txl I~): o 0 (control) 9 0.050

~,| u 0 (DC) 9 0.075

i11t ~

⁹

o.too

% j . : ~ "

C

D

I I t

I 1 i

J A S 0

Fig. 7. Seasonal courses of A attached foliage area by basal stem area in cross-section, B whole-plant dry weight in ratio to the attached (actual) foliage area, C whole-plant dry weight in ratio to the potential foliage area (see Materials and methods), and D 8a3C of the cellulose in the stemwood (data points, arrangement of symbols and straight lines as in Fig. 4)

that of rubisco as found in ozonated pine and poplar;

LiJthy-Krause et al. 1990; Landolt, personal communica- tion).

Although the calculations of Fig. 9 reconstruct the total production, they cannot reflect changes of carbon alloca- tion in the plant. Compensating the DC plants for leaf loss

Table 3. Relative growth rate, RGR = A W / A t * (W) -1 (mg g-1 day-l), as calculated from the mean whole-plant biomass per treatment at each harvest;

/x W = increment of mean biomass during time interval A t between two harvests; W = (W1+W2)/2 with W1 as the mean biomass at the beginning and W2 at the end of the time interval A t

Ozone concentration of fumigation (B11-1)

0 (control) 0.050 0.075 0.100 Defoliated control (DC)

July 17-August 15: 24 38 35 44

August 15 - October 2: 20 10 6 0

July 17 -October 2: 19 19 16 15 15

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74 Fig. 8, Lamina of 9:week-old birch leaves sampled in the afternoon (3 p.m.); A 0.05 111 1-1 03 treatment, 03 dose = 79 I-ti 1-1 h, starch

(ar- rows)

accumulating along leaf veins; B control. C and D show phloem in the leaf petioles of birch, C 0.075 ~tl 1-1 03 treatment, 03 dose = 135 p.1 1-1 h, leaf with established O3-induced discoloration; D control. E and D

show phloem in the stem axis of birch; E 03 concentration and dose as in C; F control. Petiole and stem section depicted in C and E, and in D and F from adjacent positions each;

white arrows

= deformed cells;

black arrows

= declining cell contents (C- F by Ig. K~ilin)

(Fig. 9, step 1) raised the production to that of the intact control, though the ratio stem weight/length was lowered (cf. Ericsson et al. 1980) as in the ozonated plants (Fig. 5).

This limitation in both DC and ozonated plants may result f r o m the reduced foliage density along the stem; otherwise, unlike ozonated plants, the allocation in DC plants did not differ f r o m the intact control. Thus, other causes than leaf loss per se reduced the root/shoot biomass ratio and en- hanced S L W of the foliage in ozonated plants.

Phloem transport m a y play a key role in changing the carbon allocation under 03 stress. Tracer experiments have shown reduced assimilate concentration and transport in the phloem and to the roots of ozonated pine (Spence et al.

1990). As ozone m a y have a direct impact on the phloem in the mesophyll, a disturbed phloem loading m a y result in the starch accumulation observed along the veins of the birch leaves (Fig. 8 A), even though phloem seems to be less prone to complete disintegration than mesophyll cells (Fink 1989). Ozone probably does not act directly inside the petioles and stems. Therefore, the deformed phloem seen here (Fig. 8C, E) m a y arise from limited tissue growth and maintenance due to the reduced amount of available assimilates (reduced photosynthesis, starch reten- tion in leaves). Thus, similar to findings on the effects of

802 (Michin and Gould 1986), inhibited phloem loading in the leaf by ozone m a y limit root growth and m a y affect leaf differentiation as suggested by increased S L W and changes in S, P and N.

In parallel with problems in phloem transport, root growth m a y be limited due to a favoured carbon allocation into the green biomass under 03 stress (Mooney and Win- ner 1988). In fact, R G R tended to be raised in ozonated plants but only during the early growing season, whereas the foliage area based plant production as well as A, size and total number of leaves were

n o t

enhanced as compared with the control. Perhaps 03 stress requires high carbon costs for leaf growth (Reich 1983) and for a potential delay of premature leaf loss in order to prevent an even stronger decline in whole-plant production than that observed (cf.

Mooney et al. 1988). The unchanged foliage area based

plant production (referring to the attached foliage) and area

ratio between stem sapwood and attached foliage found in

the ozonated plants m a y thus be part of such potential

acclimation to 03 stress.

(7)

Fig. 8 for legend please see p. 74

(8)

76

actual biomass

in October

Betula pendula

Total plant dry

weight in October 1989 (g)

0 40 80 20

I I I i I

, A I I

t - - i - - I

no leaf

^loss,

no decline in

CO=

assimilation

~ , ~

rate (step 1) - t

-t-

same

branch number

^as

control (step 2 ) s i z e

^same^as

leaf ~ i zon ii i i l l l ~ ~ J . I) I I control (step 3) 9 0.075

9 0.100

0 2'0

^4'0

60

^8'0 ¹⁰⁰

% of control

Fig. 9. Of each treated (i. e. ozonated or DC) tree harvested in October,

the whole-plant production is converted into that of the control by three calculations (results given as means _ SD); step 1: the potential foliage area multiplied by the mean foliage area based plant production of the control (i. e. whole-plant biomass/attached foliage area) provides the whole-plant production of treated trees as corrected for leaf loss and reduced CO2 assimilation rate, A; the obtained production is then enhanced by enlarging the potential foliage area as dependent on the branch number (step 2) and leaf sizes (step 3) of the corresponding means in the control:

03 (~tl 1-]): 0 0.05 0.075 0.100

Branch number: 5 _+ 2 3 _+ 2 2 _+ 2 1 ___ 1

Stem leaf (cm2): 65--+6 66-+7 56-+5 48-+6

Branch leaf (cm2): 37-+4 36-+5 32-+5 25-+1

Acknowledgements. We gratefully acknowledge technical assistance by Mr. U. Btihlmann, Mr. P. Bleuler, Mr. A. Burkart, and help in chemical analysis by Mr. D. Pezzotta and Prof. H. Sticher (FITZ). The support in microscopical work by Mr. lg. Kalin is much appreciated. We thank Dr.

R. H~isler and Dr. W. Landolt as well as both reviewers for helpful suggestions concerning the manuscript and Mrs. M. J. Sieber for editing the English text.

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