CADMIUM TOXICITY IN CLONES OF POPULUS TREMULA

D.L. GODBOLD, M. LITZINGER and C. GRIESE* Forstbotanisehes Institut, *Institut for Waldbau, Universit~t O6ttingen, D-3400 G6ttingen, FRG.

ABSTRACT. Four clones of Populus tremula were exposed to 1 l lM Cd at pH 3.5 or 5.0. A differing sensitivity between the clones to both pH and Cd was found. The Cd content of the roots was approximately two times that of the leaves in all 4 clones. Cadmium accumulation was affected by the pH of the nutrient solution, but the magnitude of the effect was small. The gas exchange of the clones was found to be affected by pH and Cd supply. At pH 5.0 exposure to 1 pM Cd reduced CO 2 uptake by 20 to 60 % depending upon the clone investigated. The effect of Cd on gas exchange was greater at pH 5.0 than pH 3.5. At pH 5.0 exposure to 1 laM Cd reduced the chlorophyll level in the leaves by 17 to 34 %. The water content of the leaves was reduced by exposure to Cd at both pH levels.

I. Introduction

Cadmium is one of the most important metal contaminates in arable softs. Contamination results mainly from application of Cd containing mineral fertilizers and the use of sewage sludge (Andersson and Nilsson, 1973). Exposure to Cd reduces growth and developement in a number of broad-leaf and conifer tree species (Kelly et a/., 1979; White and Rolfe, 1980; Smith and Brennan, 1984). Exposure of Picea abies, Pinus strobus or Acer rubrum to Cd reduced root initiation and growth of laterals (Mitchell and Fretz, 1977). Pinus strobus accumulated more Cd than Acer rubrum and Picea abies but based on visual symptoms of toxicity appeared to be more tolerant. White and Rolfe (1980) demonstrated a differing sensitivity to Cd between two varieties of Populus deltoides. In these varieties lower survival of the Cd-sensitive seedlings was related to a greater Cd accumulation in the whole plant. In nutrient solutions Cd accumulation is affected by a number of factors. While Tyler and Mc Bride (1982) found that Cd uptake in Phaseolus ~Igaris was not influenced by pH and Ca supply, in other species Cd uptake was strongly reduced by low pH and high Ca supply (John, 1976; Hatch et at., 1988).

In addition to reducing tree yield, Cd also influences mineral nutrition and gas exchange. In seedlings of Acersaecharinum exposure to Cd decreased the levels of Fe in the leaves and increased leaf diffusive resistance to water (Smith and Brennan, 1984). Treatment with Cd decreased CO z uptake and transpiration in Picea abies (Schlegel et a/., 1987) and transpiration in Picea rubrum (Klein, 1985). In excised leaves of Acer

Water, Air, and Soil Pollution 57-58:209-215, 1991. © 1991 Kluwer Academic Publishers. Printed in the Netherlands.

210 D.L. GODBOLD ETAL.

saccharinum Cd decreased photosynthesis and transpiration, and increased dark respiration (Lamoreaux and Chancy, 1978). These authors attributed the decrease in transpiration to the direct effects of Cd on the stomata.

In the work presented here in clones of Populus tremula the effect of Cd and pH on growth, water relations and gas exchange has been investigated.

2. Materials and methods

2.1. CULTURE OF PLANTS

Clones of Populus tremula cv. Ahle were obtained as micro-propagated cuttings from Kleinwanzlebener Saatzueht, Einbeck, F.R.G.. The cuttings were transferred to constantly aerated nutrient solutions containing (laM): KNO a 400, Ca(NOa)2 400, MgSO4 150, NaH2PO4 150, MnSO4 2, Fe -EDTA 5, HaBOa 5, (NH4)6MovO24 0.5, NaC1 10, CoSO4 0.02, ZnSO4 0.1, CuSO4 0.1, at pH 5.0. Plants were grown in a controlled environment chamber with 23/21°C day/night temperatures, 35 +5% relative humidity, 100 pmol m -2 s -1 photon flux density (Osram L18w/25 lamps), and a 16 hr photoperiod.

2.2. CADMIUM-TREATMENT

After 3 weeks preeulture, the plants were transferred to nutrient solutions adjusted to pH 3.5 or 5.0, for 1 week. The plants were then transferred to nutrient solutions adjusted to pH 3.5 or 5.0 containing 1 laM Cd for 2 weeks. Cadmium was supplied as CdCh. Plants were grown singly in 500 cm z culture vessels. Each treatment and clone was replicated 6 times using a total of 96 culture vessels. The nutrient solutions were constantly aerated and renewed every 2 to 3 days.

2.3. CADMIUM-CONTENTS

Plants were separated into leaves, roots and f'me roots (third-order or higher), dried to a constant weight and wet ashed in 3:1 (v/v) HNO a:HClO4. Cadmium was determined by atomic absorption spectrophotometry (Perkin Elmer 3030) with 4 to 5 replicates per treatment.

2.4. GAS EXCHANGE MEASUREMENTS

Gas exchange rates were determined using a Walz mini cuvette photosynthesis measuring system. The measurements were made at a photon flux density of 1200 laE m -2 s -t, a CO2 partial pressure of 334 labar, chamber/leaf temperatures of 20.1/22.1 and relative humidity of 76%. Gas exchange was determined in the youngest fully expanded leaf on 5 to 6 plants per treatment.

2.5. CHLOROPHYLL CONTENTS

The youngest fully expanded leaves were frozen in liquid nitrogen and ground (Majcherczyk et 81., 1986). Chlorophyll was extracted in 80 % (v/v) acetone and determined at 645 and 663 nm (Amon, 1949), with 3 replicates per treatment.

CADMIUM TOXICITY IN CLONES OF POPULUS TREMULA

2.6, WATER CONTENTS

Water contents of the leaves were calculated using the difference between fresh and dry weight. Four to five replicates per treatment were determined.

211

3. Results

After a 2 week exposure to 1 laM Cd at pH 3.5 or 5.0, independent of clone and pH, higher Cd contents were found in fine roots than roots than leaves (Figure 1). At pH 3.5 in clone 9 and 14, higher Cd contents were found in all plants parts than in clones 4 and 6. At pH 5.0 this difference in Cd contents between the clones was less. At pH 5.0 the Cd contents of clones 4 and 6 were higher than at pH 3.5, whereas the Cd contents of clone 14 were lower at pH 5.0 than 3.5.

4 0

20 a

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o E =, 0

E

g 40 0

"o

2 0 ¸

[ ] f ine roots [ ] root [ ] leaf

pH 3-5

pH 5-0

4 6

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9 clone

14

Fig. 1. Cd contents in tissues of Populus tremula clones after exposure to 1 pM Cd at pH 3.5 of 5.0, for two weeks. I= S.E.

212 D.L. GODBOLD ET AL.

Rates of CO2 uptake (Table I) and transpiration (Table II) were influenced by both pH and Cd supply. Clones 4 and 14 had lower rates of CO 2 uptake and transpiration at pH 3.5 than at pH 5.0. In clones 6 and 9, CO 2 uptake was not affected by pH and rates of transpiration were only slightly lower at pH 3.5 than pH 5.0. After exposure to 1 t im Cd, a greater inhibition of CO ~ uptake and transpiration was found at pH 5.0 than at pH 3.5. Only in clone 9 were CO z uptake and transpiration reduced by Cd at both pH values. In clone 14 at pH 3.5, CO 2 uptake and transpiration were increased by exposure to Cd at pH 3.5, and were least inhibited by Cd at pH 5.0.

TABLE I. Influence of Cd on CO 2 uptake (pmol CO 2 m -2 s -1) of Populus tremula clones grown at two pH values. _+ SE. Percent of control in brackets.

clone control 1 pM Cd

pH 3.5 pH 5.0 pH 3.5 pH 5.0

4 3.2 *°'s 4.4 *0.3 3.1 *°'5 (96) 2.0 *°'3 (45) 6 4.3 *0.4 4.6 *°'2 4.5 *°.* (105) 1.9 *°'4 (40)

9 4.6 *°'4 4.6 .°"I 4.1 *o'4 (88) 2.4 *°'s (52) 14 2.7 *0.4 3.8 ~°'I 4.1 *°.5 (151) 3.1 *°'3 (81)

TABLE II. Influence of Cd on transpiration (lamol H 20 m =2 s-1) of Populus tremula clones grown at two pH values. + SE. Percent of control in brackets.

clone control 1 ~M Cd

pH 3.5 pH 5.0 pH 3.5 pH 5.0

4 443 *78 1294 .126 388 *64 (87) 538 .I.2 (41)

6 707 *a° 1186 *7o 664 *49 (93) 664 **o4 (56) 9 1063 .141 1253 *3o 575 *98 (54) 393 *47 (31)

14 407 *s8 1080 .5~ 747 *z~7 (184) 668 *3s (62)

CADMIUM TOXICITY IN CLONES OF POPULUS TREMULA 213

In all clones after exposure to Cd, the chlorophyll content (Table III) of the leaves was higher at pH 3.5 compared to the control, whereas at pH 5.0 exposure to Cd decreased the chlorophyll eontent of the leaves.

In all clones the water content (Table IV) of the leaves was greater at pH 5.0 than pH 3.5. Exposure to Cd decreased the water content of the leaves at both pH values. Between clones, at both pH values the greatest decrease in water content after exposure to Cd was shown in clone 9.

TABLE III. Influence of Cd on chlorophyll contents (mg g -1 Fwt) of Populus tremula clones grown at two pH values. +_ SE. Percent of control in brackets.

clone pH 3.5 pH 5.0

control 1 ~M Cd control 1 zM Cd

4 2.0 ~°'I 2.2 *°'I (ii0) 2.6 ~°'2 2.0 *°'2 (77) 6 2.2 *0.2 2.9 *°'s (131) 3.1 I°'2 2.1 *°'* (68) 9 2.0. o.2 2.3*o. I (113) 2.1to. I 1 . 7 ~ o . s (83)

14 2.5 ~°'3 2.5 *°'3 (i00) 2.5 ~°'I 1.7 I°'I (66)

TABLE IV. Influence of Cd on water contents (%) of Populus tremula clones grown at two pH values. _+ SE. Percent of control in brackets.

clone pH 3.5 pH 5.0

control 1 pM Cd control 1 pM Cd

4 8 3 . 3 ± 0 . 4 7 8 . 3 ~1"1 (94) 8 7 . 0 ~ ° ' 7 8 2 . 8 i ° ' 9 (95) 6 8 4 . 2 ~ ° ' 5 7 9 . 0 * ° ' 5 (94) 8 7 . 0 * ° ' z 8 5 . 6 ~ ° ' 7 (98 ) 9 8 3 . 6 ~ ° ' ~ 7 6 . 8 * ° ' 8 (92 ) 8 5 . 6 . 1 " ° 7 3 . 8 ~ ° ' e (86 )

14 8 0 . 0 ~1"1 7 8 . 8 ~ ° ' s (99 ) 8 6 . 7 ~ ° ' 3 7 7 . 8 i ° ' s (90 )

4. Discussion

In the 4 clones of Populus tremula used in this study differences in sensitivity to Cd were shown. White and Rolfe (1980) showed a difference in Cd tolerance between two varieties of cottonwood (Populus doltiodes). Steiner et al. (1984) found differences in tolerance to aluminium in a number of Populus hybrids.

Accumulation of Cd in plant parts varies among species (Haghiri, 1973). For example in seedlings of Pieea shies exposed to the same levels of Cd as used here, 95% of the Cd taken up was accumulated in the roots (Sehlegel, 1985). In this report only

214 D.L. GODBOLD ET AL.

30 to 38% of the Cd accumulated was retained in the roots of the Populus tremula clones. In rooted stem cuttings of Populus x Androseoggin, higher Cd contents were found in leaves than roots (Gingas et al., 1988). In seedlings of Populus deltoides higher levels of Cd were found in roots than shoots (White and Rolfe, 1980). Although a difference in accumulation of Cd in plant parts between plants grown from cuttings or seeds cannot be excluded, some Populus species or cultivars appear to readily transport Cd to the leaves.

Clones of Populus tremula differed not only in sensitivity to Cd, but also to the pH of the nutrient solution. In all clones, a greater inhibition of gas exchange by Cd was found at pH 5.0 than at pH 3.5. This was not related to higher Cd contents in the plants. With the exception of clone 9 which had the highest leaf Cd content and the greatest depression of gas exchange, no strong relationship between the Cd content of the leaves and the degree of inhibition of gas exchange could be found. This suggest that either the leaves of the clones have a different tolerance to Cd, or that the levels of Cd in the leaves were insufficient to directly effect gas exchange. In excised leaves of Aeersaccharinum treated with Cd inhibition of net photosynthesis and transpiration was correlated with the Cd contents of the leaves (Lamoreaux and Chancy, 1978). In Pieea abies ($ehlegel et al., 1987) and Lyeopersieon eseulentum (Baszynski et al., 1980) decreased rates of CO z uptake were accounted for by lower levels of chlorophyll and stomatal closure. In this study no clear relationship between the relative decrease in ehiorophyU content and the relative decrease in CO z uptake was evident.

In all clones, the water contents of the leaves were affected by both the pH of the nutrient solution and Cd supply. With some exceptions (Cd addition at pH 5.0) the relative changes in water content of the leaves had a similar pattern to the relative decrease in CO2 uptake due to both the pH of the nutrient solution and Cd supply. Lamoreaux and Chancy (1977) found decreased water conductivity of excised stem sections of Aeer saeeharinum after exposure to Cd. Robb eta/ . (1980) described blockage of xylem elements in stems of Phaseolus vulgaris treated with high concentrations of Zn. In Pieea abies seedlings treated with Hg decreased water contents in needle was attributed to root damage (Godbold and Htittermann 1988). Photosynthesis is reported to be sensitive to the water status of leaves (Kaiser, 1987). Lower water flow could account for the changes in water contents in the leaves of the clones and subsequently CO z uptake.

This work shows that clones of Populus tremula have differences in sensitivity to both Cd and pH. The effects of Cd were found to be greater at pH 5.0 than 3.5

5. Acknowledgements

This work was supported by the Bundesministerium ftir Forschung und Technologic in the renewable resources research programme.

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Physiol. P1antarum, 48, 365. Gingas, V.M., Sydnor, T.D. and Weidensaul, T.C.: 1988, Z Amer. Soe. Hort. Sei., 113,

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CADMIUM TOXICITY IN CLONES OF POPULUS TREMULA 215

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