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Fruit set, young fruit and leaf growth of Citrus unshiu in relation to assimilate supply
Scientia Horticulturae, 53 ( 1993 ) 99-107
99
Elsevier Science Publishers B.V., Amsterdam
Fruit set, young fruit and leaf growth of Citrus unshiu in relation to assimilate supply Yong-Ling Ruan Department of Horticulture, Zhejiang Agricultural, University, Hangzhou 310029, People's Republic of China (Accepted 2 June 1992)
ABSTRACT Ruan, Y.-L., 1993. Fruit set, young fruit and leaf growth of Citrus unshiu in relation to assimilate supply. Scienttia Hortic., 53: 99-107. Fruit set, frc itlet and young leaf growth at an early stage of fruit ontogeny were promoted significantly by girdling at the base of autumn-shoots of the preceding year on Day 10 pre-full bloom. In contrast, defoi/iation or darkening of the old leaves on the autumn-shoots aggravated fruit drop and inhibited fruitlet and young leaf growth. During this period the net photosynthetic rate (Pn) of old leaves rose sharply. These observations clearly indicate that assimilate supply to fruitlets and young leaves of Citrus unshiu during this period is mainly affected by the current photosynthates from the old leaves instead of from the remobilization of stored reserves within the trunks. On Day 20 after full bloom the young leaves reached 48% of their final area and increased their Pn rapidly. Subsequently, the Pn of old leaves began to decrease dramatically. The impeding effect of darkening of olid leaves on fruit growth from Day 20 after full bloom was absent. The results show that the young leaves gradually substitute for the old leaves to supply photosynthates to the developing fruits from about Day 20 after full bloom. Keywords: assimilate supply; Citrus unshiu; defoliation; fruit set; girdling; leaf darkening; photosynthesis; young fruit and leaf growth. Abbreviations: ABA = abscisic acid; AFB = after full bloom; FB = full bloom; GA3 = gibberellic acid; PFB = pre-full bloom; Pn = net photosynthetic rate; ZT = zeatin.
INTRODUCTION
The initiation and subsequent development of floral organs, fruitlets and young leaves are most vulnerable to deficiencies in assimilate supply (Patrick, 1988). It has been proposed that carbohydrate levels exert a regulatory role in fruit set of citrus crops (Erner, 1989). The potential for carbohydrate Correspondence to: Y.-L. Ruan, Department of Biological Sciences, The University of Newcastle, N.S.W. 2308, Australia.
© 1993 Elsevier Science Publishers B.V. All rights reserved 0304-4238/93/$06.00
1O0
Y.-L. RUAN
consumption during fruit set may exceed current photosynthesis and hence the carbohydrate supply may become a limiting factor for fruit set of citrus crops (Garcia-Luis et al., 1988). Undoubtedly, better understanding of the relationship between the development of these meristematic sinks and assimilate supplies will be both of theoretical and horticultural significance. Assimilates, for the developing sinks, can come from stored reserves and from current photosynthates (Weaver and Johnson, 1985). In the east of China, shading satsuma mandarin trees in winter does not affect the bud break and flowering next spring (Wu et al., 1988). Furthermore, no positive net photosynthetic rate (Pn) value of the field-grown trees was observed from December to mid-April, at the time of new spring-flush emergence (Ruan et al., 1987). It is concluded that the assimilates for initiation and development of floral and leaf primordium come from stored reserves (Ruan et al., 1987). However, the old leaves on autumn-shoots of the preceding year (old leaves in the following text) resume their photosynthetic capacity in the flowering period (Akao et al., 1981; Ruan et al., 1987). In contrast, the y~ung springflush leaves (young leaves in the following text) do not export photosynthate at this stage (Liu et al., 1989). Thus, it appears that the old leaves might export photosynthates to flowers and fruitlets. Garcia-Luis et al. (1988) assumed that most of the dry matter consumed in the fruiting process of citrus crops is drawn from both current photosynthesis and from reserves stored in the 'old' parts of the tree. Nevertheless, experimental evidence on fruit set and subsequent development of Citrus unshiu in relation to photoassimilate source (s) is sparse. Assimilate partitioning to meristematic sinks could be regulated by components of the source-path system (Patrick, 1988). Girdling, leaf darkening or shading (Beruter and Droz, 1991 ) and defoliation (Ryugo, 1985) are often used as means to manipulate the assimilate supply to developing sinks. Studies reported in this paper attempt to determine the source(s) of assimilate supply for the fruitlets and young leaves of C. unshiu based on the effects of girdling, old leaf darkening and defoliation. Changes of Pn of old and young leaves of control shoots were monitored during the flowering and early fruit growth periods. MATERIALS AND METHODS
The field experiments were conducted in the agricultural experiment station of Zhejiang Agricultural University with 5-year-old bearing satsuma mandarin (C. unshiu, Marc. ) trees. The climate is subtropical with an average annual rainfall of 1616.0 ram, average daily minimum temperature in January of 2.7 ° C and average daily maximum temperature in July of 32.6 ° C. The site is located at 30 ° 0' N and 119 ° 5' E.
GROWTH OF C UNSHIU IN RELATION TO ASSIMILATES
101
Experiment I Treatments. - Ten autumn-shoots of the preceding year from each of six trees were tagged and divided into five groups belonging to five treatments. The selected shoots were about 0.5 cm thick and 18.0 cm in length with uniform flower and leaf loads. The criterion of full bloom (FB) was taken as :~heday when most flowers were open. The following treatments were applied on Day 10 pre-full bloom (PFB): ( 1 ) girdling phloem of the shoots at their bases with a grafting knife to prevent the influx or effiux of assimilates; (2) darkening the old leaves with black paper ~gags (9 cm X 6 cm), leaving the young leaves and floral organs exposed; (3) defoliation of all the old leaves on each shoot; (4) girdling and defoliation; ( 5 ) control. Each treatment contained 12 shoots. Leafless inflorescences, which are not responsible for the yield (Erner, 1889) and less than 10% of the total inflorescences in our experiments, were removed. Thus only leaf inflorescences were left for comparison. Morphological investigation andPn measurements. - On Days 10 PFB and 20 after full bloom (AFB), the numbers of old leaves and flowers or fruitlets on each treated shoot were counted. The fresh weights of 12 flowers or fruitlets at the nodal Position 5-7 on the shoots of each treatment were measured. Twelve young leaves of each treatment were labelled for leaf area measurement with a leaf area instrument (LI 3000, LI-COR, Lincoln, NE). The Pn of 12 young leaves and 6 old leaves from control shoots were monitored with an infra-red gas analyser (FQ-W, Fushan analytical plant, Fushan City, China), using an open system at 22-24 °C and 400 gE m -2 s-~ light intensity according to Ruan et al., ( 1988 ). Both the leaf area and Pn were determined from the Day 10 PFB to Day 50 AFB at 10 day intervals. Gibberellic acid (GAa), zeatin (ZTO and abscisic acid (ABA) determinations. Three fruits from each treatment were picked and weighed on Day 20 AFB. The fruit were immediately snap frozen in liquid nitrogen and then stored at - 2 0 ° C . The extraction of endogenous GA3, ZT and ABA from the fruit was carried out according to the procedure described by Ruan et al. ( 1991 ). An HPLC (Waters 201 model) method was used for determination of the hormone content ZT was separated and detected on a 250 m m × 5 mm column of Lichorosorb Rp-18 in the mobile phase of methanol:acelomitrile:water = 33:23:44 (pH 4.0 with acetic acid) with a 270 nm UV detector. GA3 and ABA were separated and detected on a 300 mm × 4 mm column of g-Bondapak C~8 in the mobile phase of methanol: acelomitrile:water = 1"1:3 (pH 4.0 with acetic acid) with a 254 nm UV detector. For more details see Ruan et al. ( 1991 ).
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Y.-L. RUAN
Experiment 2 On Day 20 AFB, six autumn-shoots from each of six trees were divided into three groups with two shoots in each for the following treatments: ( l ) girdling; (2) leaf darkening; (3) control. The methods of the treatments were the same as in Experiment 1. The numbers and fresh weights of fruits were counted and measured respectively as duscdbed in Experiment 1 on the Days 20, 35 and 50 AFB. RESULTS AND D I S C U S S I O N
Effects of treatments applied PFB (Experiment 1) A girdling treatment of 30 day from Day 10 PFB increased fruit set two fold and promoted fruitlet and young leaf growth significantly, whereas defoliation with or without girdling exerted opposite effects (Tables l and 2 ). Further, darkening treatment of old leaves accentuated fruit drop and inhibited frtqtlet and young leaf growth (Tables I and 2 ). The results indicate that fruit set and subsequent fruitlet and young leaf growth of C. unshiu at the early stage of fruit ontogeny are strongly influenced by the old leaves instead of remobilization of stored reserves within tree trunks. The old leaves rapidly resume their photosynthetic capacity in the flowering period (Akao et al., 1981; Ruan et al., 1987 and also see Fig. l ). Assimilate export rate is proportional to photosynthetic rate in both short- and longterm experiments (Ho et al., 1989). Therefore, it is considered that the contribution of old leaves to the fruitlets and young leaves is attributed to their TABLE ! Effect of girdling, old leaf darkening and defoliation applied at Day 10 PFB on fruit set (fruits per shoot) and fruit weight (grams FW per fruit) of C. unshiu trees Treatment
Days after treatment 0
Control Girdling Defoliation Leaf darkening Girdling+defoliation
30
Flower set '
Flower weight
Fruit set
Fruit weight
12.6 a 2 11.8 a 10.8 a 10.9 a 11.2 a
0.25 0.25 0.24 0.26 0.25
1.6 a 4.8 b 0.6 c 0.3 d 0.0 e
1.0 a 1.2 b 0.8 c 0.4 d -
a a a a a
IThe units of flower set and flower weight are the same as those of fruit set and fruit weight respectively. eMeans followed by different letters within each column differ significantly at 5% level according to Duncan's multiple range test.
103
GROWTH OF C. UNSHIU IN RELATION TO ASSIMILATES
TABLE 2 Effect of girdling, old leaf darkening and defoliation applied at Day 10 PFB on leaf area (cm 2) of young spring-flush leaves of C. unshiu trees Treatment
Days after treatment
Control Girdling Defoliation Leaf darkening Girdling + defoliation
0
30
0.67 a 0.68 a 0.68 a 0.66 a 0.68 a
5.20 a 7.12 b 4.71 c 3.90 d 3.50 d
'Means followed by different letters within each column differ significantly at 5% level according to Duncan's multiple range test
8
,-8
;-
young leaf
A
m
0 0
6
old leaf
4
0
E e,. O.
2
0
0 -20
0
20
40
60
days after full bloom Fig. 1. Comparison of Pn changes between young spring-flush leaves and old leaves on autumnshoots of the preceding year. Each point represents the mean + SE.
current photosynthate supply. Results from a '4CO2 labelling experiment indicate that more than one third of current photosynthates in old leaves are exported to young fruit and leaves during the fruiting process of C. unshiu (Akao et al., 1981 ). The inhibition of leaf photosynthesis results in similar negative effects on fruit set and growth in sweet orange (Moss, 1976), peach (Byers et al., 1985) and apple (Byers et al., 1985; Beruter and Droz, 1991 ). Our results also support the view that the supply of photoassimilate is critical
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Y.-L. RUAN
for fruit retention in early stage of fruit development (Patrick, 1988; Beruter and Droz, 1991 ). For fruit bearing satsuma mandarin trees, carbohydrate reserves are mainly stored in large roots and the crown (Akao et al., 1981 ). In this study the pos. sible contribution of stored reserves in these organs could not be directly determined. However, the obvious positive effect of girdling with maintenance ofold leaves on fruit set and growth (Tables 1 and 2), suggests that the amount of assimilates transported to fruitlet and young leaves from the stored reserves in the roots and crown, were not a significant assimilate source to support the development of these organs. The defoliation treatment with girdling led to no fruit retention and inhibited young leaf growth to a greater extent than defoliation without girdling (Tables 1 and 2 ), suggesting the stored assimilates within trunks might be transported to fruitlets and young leaves in case of old leaf abscission. Previous studies indicated that carbohydrate reserves in old leaves of C. unshiu will not be mobilized until senescence usually about 2-3 weeks before leaf drop (Ruan et al., 1987; Wu et al., 1988). In the east of China, the old leaf abscission occurs about 4 months after flowering. Thus, it is unlikely that carbohydrate reserves in the old leaves are exported to fruitlets and young leaves. It has been demonstrated that in fruit trees nitrogen is mainly stored in perennial woody tissues (roots and stems) over winter and is always used predominately for young fruit and leaf growth next spring regardless of the current nitrogen supply (Erner, 1989; Millard and Thomson, 1989). Thus, defoliation of old leaves appears not to alter the nitrogen supply to fruitlets and young leaves. As the major transfer process involved in providing nitrogen to young developing organs in the stem is via xylem (Dasilva and Shelp, 1990), girdling the phloem of the shoots would not affect the nitrogen transport to the developing sinks. In order to determine the effect of girdling, defoliation and leaf darkening on the hormonal balance of the young fruits, their endogenous GA3, ZT and ABA contents were analysed. No significant difference was found among these treatments, the average contents of GA3, ZT and ABA in the fruitlets at Day 20 AFB were 0.35, 2.60 and 0.21/~g g-~ FW, respectively. This result further verifies that the effects of girdling, defoliation and leaf darkening on fruitier growth is mainly a function of photoassimilate supply. The fact that the hormonal balance in the fruitlets was not affected by girdling, old leaf darkening and defoliation implies that hormones in the fruitlets may be synthesized locally as proposed by Erner (1989). It is interesting to note that fruit drop and growth inhibition caused by defoliation were less than that caused by leaf darkening (Tables I and 2 ). Darkening the old leaves may have changed them from a source to sink model owing to their considerable dark respiration (Akao et al., 1981; Ruan et al.,
GROWTH OF C. UNSHIU IN RELATION TO ASSIMILATES
105
TABLE 3 Effect of girdling and old leaf darkening applied at Day 20 AFB on fruit weight (grams FW per fruit ) ofC. unshiu trees Treatment
Control Girdling Leaf darkening
Days after treatment 0
15
30
1.2 a ~ 1.2 a 1.1 a
3.8 a 3.6 a 2.6 b
6.5 a 6.7 a 6.2 a
1Means followed by different letters within each column differ significantly at 5% level according to Duncan's multiple test.
1988). Thus the darkened old leaves might compete with the meristematic sinks for assimilate supply, accentuating fruit drop and growth inhibition.
Effects of treatments applied AFB (Experiment 2) Assimilate patterns are altered by the changes in demand by developing sinks (Patrick, 1988). Therefore the effects of assimilate supply manipulation AFB were studied. Sink strength is positively correlated with sink volume (Patrick, 1988). The assimilate supply for the rapidly growing citrus fruit would be enhanced after fruit set, which could lead to the decrease, even cessation, of photoassimilate export out of the girdling position. The fact that girdling applied at Day 20 AFB did not affect young fruit weight increase (Table 3 ) is consistent with this proposal. The Pn of old leaves began to decrease dramatically from Day 20 AFB (Fig. l ). It suggests that current photosynthate supply from old leaves to developing fruits decreases after Day 20 AFB. This is consistent with the fact that 30 days of darkening old leaves did not result in fruit growth inhibition (Table 3 ). The net photosynthate export of young leaves of C. unshiu commences at the stage of 40% of their final leaf area (Liu et al., 1989 ). In this experiment the young leaves reached 48% of their final leaf area at Day 20 AFB and thus can be regarded as a 'net source leaves'. This corresponds to the time that the Pn of the young leaves exceeds that of the old leaves which continues to decline (Fig. l ). These results indicate that young leaves gradually substitute for old leaves to supply current photosynthates to the developing fruits after Day 20 AFB. CONCLUSIONS
The assimilate supply for fruitlets and young leaves at the early stage of fruit ontogeny mainly originates from current photosynthesis of the old leaves.
106
Y.-L.RUAN
This conclusion is verified by the fact that in south and east China old leaf abscission of field-grown trees, owing to cold injury in winter and early spring, leads to poor fruit set (Ruan et al., 1987; Wu et al., 1988). Fruit drop was greatly accentuated by leaf darkening during flowering and fruiting processes. This suggests that reduced solar radiation by shading or by overcast weather in this period could be a major hindrance to the fruit sei. The response emphasises the importance of maintaining an optimum light environment within citrus groves. On Day 20 AFB, the young leaves, considered to be a net exporter of photosynthates, increased in Pn rapidly and substituted for the old leaves to supply current photosynthates to the developing fruits. ACKNOWLEDGEMENT
I would like to thank Professor John W. Patrick for his helpful comments and critical reading of the manuscript. REFERENCES Ak~,:, S., Tsukahara, S., Hisada, H. and Ona, S., 1981. Contribution of photosynthetic assimilates to development of flower and spring flush in Citrus unshiu Marc. J. Jpn. Soc. Hortic. Sci., 50: 1-9. Beruter, J. and Droz, Ph., 1991. Studies on locating the signal for fruit abscission in the apple tree. Scientia Hortic., 46:201-214. Byers, R.E., Lyons, Jr., J.C., Yoder, K.S., Barden, J.A. and Young, R.W., 1985. Peach and apple thinning by shading and photosynthetic inhibition. J. Hortic. Sci., 60: 465-475. Dasiva, M.C. and Shelp, B.J., 1990. Xylem-to-phloem transfer of organic nitrogen in young soybean plants. Plant Physiol., 92: 797-80~. Erner, Y., 1989. Citrus fruit set: carbohydrate, hormone and leaf mineral relationships. In: C.J. Wright (Editor), Manipulation of Fruiting. Academic Pre~s, Butterworths, London, pp. 233242. Garcia-Luis, A., Fornes, F., Sanz, A. and Guardiola, J.L., 1988. The regulation of flowering and fruit set in citrus: relationship with carbohydrate levels. Israel J. Bot., 37:189-201. Ho, L.C., Grange, R.I. and Shaw, A.F., 1989. Source/sink regulation. In: D.A. Baker and J.A. Milburn (Editors), Transport of Photoassimilates. Longman, Harlow, UK, pp. 319-320. Liu, D.-H., Zhang, S.-L., Wu, G.-L. and Zhang, L.-C., 1989. Studies on photosynthate translocation in Citrus unshiu. Acta Hortic.. 16:191 - 198 (in Chinese with English abstract ). Millard, P. and Thomson, C.M., 1989. The effect ofautumn senescence of leaves on the internal cycling of nitrogen for the spring growth of apple trees. J. Expl. Bot., 40:1285-1289. Moss, G.L., 1976. Thinning 'Washington' navel and 'late valencia' sweet orange fruits with photosynthetic inhibitor. HortScience, I 1: 48-50. Patrick, J.W., 1988. Assimilate partitioning in relation to crop prc,ductivity. HortScience, 23: 33-40. Ruan, Y.-L., Zhang, L.-C., Wu, G.-L. and Zhang, S.-L., 1987. Changes of net photosynthetic rate of wintering citrus leaves. Acta Agric., 13:392-395 (in Chinese with English abstract ). Ruan, Y.-L., Zhang, L.-C., Wu, G.-L. and Zhang, S.-L., 1988. Effect of water stress on photosynthesis of Citrus unshiu. Acta Hortic., 15:93-98 (in Chinese with English abstract ). Ruan, Y.-L., Zhang, S.-L., Zhu, K., Wu, G.-L. and Li, S.-J., 1991. The types of cytokinins and
GROWTH OF C. UNSHIU IN RELATION TO ASSIMILATES
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their changes and abscisic acid in the stem of Citrus unshiu during the period of flower bud initiation. Scientia Agric., 24:55-59 (in Chinese with English abstract ). Ryugo, K., 1985. Promotion and inhibition of flower initiation and fruit set by plant manipulation and hormones. Acta Hortic., 1:301-305. Weaver, R.J. and Johnson, J.O., 1985. Relation of hormones to nutrient mobilization and the internal environment of the plant: The supply of mineral nutrients and and photosynthate. In: R.P. Pharis, and D.M. Reid (Editors), Hormonal Regulation of Development. III Role of Environmental Factors. Encyclopedia of Plant Physiology, Vol. II. Springer, Berlin, pp. 3-36. Wu, G.-L., Zhang, L.-C., Zhang, F., Ruan, Y.-L. and Liu, J.-S., 1988. A study on the physiology of overwintering satsuma trees under straw sheds. China Citrus, 17:3-6 (in Chinese ).
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Elsevier Science Publishers B.V., Amsterdam
Fruit set, young fruit and leaf growth of Citrus unshiu in relation to assimilate supply Yong-Ling Ruan Department of Horticulture, Zhejiang Agricultural, University, Hangzhou 310029, People's Republic of China (Accepted 2 June 1992)
ABSTRACT Ruan, Y.-L., 1993. Fruit set, young fruit and leaf growth of Citrus unshiu in relation to assimilate supply. Scienttia Hortic., 53: 99-107. Fruit set, frc itlet and young leaf growth at an early stage of fruit ontogeny were promoted significantly by girdling at the base of autumn-shoots of the preceding year on Day 10 pre-full bloom. In contrast, defoi/iation or darkening of the old leaves on the autumn-shoots aggravated fruit drop and inhibited fruitlet and young leaf growth. During this period the net photosynthetic rate (Pn) of old leaves rose sharply. These observations clearly indicate that assimilate supply to fruitlets and young leaves of Citrus unshiu during this period is mainly affected by the current photosynthates from the old leaves instead of from the remobilization of stored reserves within the trunks. On Day 20 after full bloom the young leaves reached 48% of their final area and increased their Pn rapidly. Subsequently, the Pn of old leaves began to decrease dramatically. The impeding effect of darkening of olid leaves on fruit growth from Day 20 after full bloom was absent. The results show that the young leaves gradually substitute for the old leaves to supply photosynthates to the developing fruits from about Day 20 after full bloom. Keywords: assimilate supply; Citrus unshiu; defoliation; fruit set; girdling; leaf darkening; photosynthesis; young fruit and leaf growth. Abbreviations: ABA = abscisic acid; AFB = after full bloom; FB = full bloom; GA3 = gibberellic acid; PFB = pre-full bloom; Pn = net photosynthetic rate; ZT = zeatin.
INTRODUCTION
The initiation and subsequent development of floral organs, fruitlets and young leaves are most vulnerable to deficiencies in assimilate supply (Patrick, 1988). It has been proposed that carbohydrate levels exert a regulatory role in fruit set of citrus crops (Erner, 1989). The potential for carbohydrate Correspondence to: Y.-L. Ruan, Department of Biological Sciences, The University of Newcastle, N.S.W. 2308, Australia.
© 1993 Elsevier Science Publishers B.V. All rights reserved 0304-4238/93/$06.00
1O0
Y.-L. RUAN
consumption during fruit set may exceed current photosynthesis and hence the carbohydrate supply may become a limiting factor for fruit set of citrus crops (Garcia-Luis et al., 1988). Undoubtedly, better understanding of the relationship between the development of these meristematic sinks and assimilate supplies will be both of theoretical and horticultural significance. Assimilates, for the developing sinks, can come from stored reserves and from current photosynthates (Weaver and Johnson, 1985). In the east of China, shading satsuma mandarin trees in winter does not affect the bud break and flowering next spring (Wu et al., 1988). Furthermore, no positive net photosynthetic rate (Pn) value of the field-grown trees was observed from December to mid-April, at the time of new spring-flush emergence (Ruan et al., 1987). It is concluded that the assimilates for initiation and development of floral and leaf primordium come from stored reserves (Ruan et al., 1987). However, the old leaves on autumn-shoots of the preceding year (old leaves in the following text) resume their photosynthetic capacity in the flowering period (Akao et al., 1981; Ruan et al., 1987). In contrast, the y~ung springflush leaves (young leaves in the following text) do not export photosynthate at this stage (Liu et al., 1989). Thus, it appears that the old leaves might export photosynthates to flowers and fruitlets. Garcia-Luis et al. (1988) assumed that most of the dry matter consumed in the fruiting process of citrus crops is drawn from both current photosynthesis and from reserves stored in the 'old' parts of the tree. Nevertheless, experimental evidence on fruit set and subsequent development of Citrus unshiu in relation to photoassimilate source (s) is sparse. Assimilate partitioning to meristematic sinks could be regulated by components of the source-path system (Patrick, 1988). Girdling, leaf darkening or shading (Beruter and Droz, 1991 ) and defoliation (Ryugo, 1985) are often used as means to manipulate the assimilate supply to developing sinks. Studies reported in this paper attempt to determine the source(s) of assimilate supply for the fruitlets and young leaves of C. unshiu based on the effects of girdling, old leaf darkening and defoliation. Changes of Pn of old and young leaves of control shoots were monitored during the flowering and early fruit growth periods. MATERIALS AND METHODS
The field experiments were conducted in the agricultural experiment station of Zhejiang Agricultural University with 5-year-old bearing satsuma mandarin (C. unshiu, Marc. ) trees. The climate is subtropical with an average annual rainfall of 1616.0 ram, average daily minimum temperature in January of 2.7 ° C and average daily maximum temperature in July of 32.6 ° C. The site is located at 30 ° 0' N and 119 ° 5' E.
GROWTH OF C UNSHIU IN RELATION TO ASSIMILATES
101
Experiment I Treatments. - Ten autumn-shoots of the preceding year from each of six trees were tagged and divided into five groups belonging to five treatments. The selected shoots were about 0.5 cm thick and 18.0 cm in length with uniform flower and leaf loads. The criterion of full bloom (FB) was taken as :~heday when most flowers were open. The following treatments were applied on Day 10 pre-full bloom (PFB): ( 1 ) girdling phloem of the shoots at their bases with a grafting knife to prevent the influx or effiux of assimilates; (2) darkening the old leaves with black paper ~gags (9 cm X 6 cm), leaving the young leaves and floral organs exposed; (3) defoliation of all the old leaves on each shoot; (4) girdling and defoliation; ( 5 ) control. Each treatment contained 12 shoots. Leafless inflorescences, which are not responsible for the yield (Erner, 1889) and less than 10% of the total inflorescences in our experiments, were removed. Thus only leaf inflorescences were left for comparison. Morphological investigation andPn measurements. - On Days 10 PFB and 20 after full bloom (AFB), the numbers of old leaves and flowers or fruitlets on each treated shoot were counted. The fresh weights of 12 flowers or fruitlets at the nodal Position 5-7 on the shoots of each treatment were measured. Twelve young leaves of each treatment were labelled for leaf area measurement with a leaf area instrument (LI 3000, LI-COR, Lincoln, NE). The Pn of 12 young leaves and 6 old leaves from control shoots were monitored with an infra-red gas analyser (FQ-W, Fushan analytical plant, Fushan City, China), using an open system at 22-24 °C and 400 gE m -2 s-~ light intensity according to Ruan et al., ( 1988 ). Both the leaf area and Pn were determined from the Day 10 PFB to Day 50 AFB at 10 day intervals. Gibberellic acid (GAa), zeatin (ZTO and abscisic acid (ABA) determinations. Three fruits from each treatment were picked and weighed on Day 20 AFB. The fruit were immediately snap frozen in liquid nitrogen and then stored at - 2 0 ° C . The extraction of endogenous GA3, ZT and ABA from the fruit was carried out according to the procedure described by Ruan et al. ( 1991 ). An HPLC (Waters 201 model) method was used for determination of the hormone content ZT was separated and detected on a 250 m m × 5 mm column of Lichorosorb Rp-18 in the mobile phase of methanol:acelomitrile:water = 33:23:44 (pH 4.0 with acetic acid) with a 270 nm UV detector. GA3 and ABA were separated and detected on a 300 mm × 4 mm column of g-Bondapak C~8 in the mobile phase of methanol: acelomitrile:water = 1"1:3 (pH 4.0 with acetic acid) with a 254 nm UV detector. For more details see Ruan et al. ( 1991 ).
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Y.-L. RUAN
Experiment 2 On Day 20 AFB, six autumn-shoots from each of six trees were divided into three groups with two shoots in each for the following treatments: ( l ) girdling; (2) leaf darkening; (3) control. The methods of the treatments were the same as in Experiment 1. The numbers and fresh weights of fruits were counted and measured respectively as duscdbed in Experiment 1 on the Days 20, 35 and 50 AFB. RESULTS AND D I S C U S S I O N
Effects of treatments applied PFB (Experiment 1) A girdling treatment of 30 day from Day 10 PFB increased fruit set two fold and promoted fruitlet and young leaf growth significantly, whereas defoliation with or without girdling exerted opposite effects (Tables l and 2 ). Further, darkening treatment of old leaves accentuated fruit drop and inhibited frtqtlet and young leaf growth (Tables I and 2 ). The results indicate that fruit set and subsequent fruitlet and young leaf growth of C. unshiu at the early stage of fruit ontogeny are strongly influenced by the old leaves instead of remobilization of stored reserves within tree trunks. The old leaves rapidly resume their photosynthetic capacity in the flowering period (Akao et al., 1981; Ruan et al., 1987 and also see Fig. l ). Assimilate export rate is proportional to photosynthetic rate in both short- and longterm experiments (Ho et al., 1989). Therefore, it is considered that the contribution of old leaves to the fruitlets and young leaves is attributed to their TABLE ! Effect of girdling, old leaf darkening and defoliation applied at Day 10 PFB on fruit set (fruits per shoot) and fruit weight (grams FW per fruit) of C. unshiu trees Treatment
Days after treatment 0
Control Girdling Defoliation Leaf darkening Girdling+defoliation
30
Flower set '
Flower weight
Fruit set
Fruit weight
12.6 a 2 11.8 a 10.8 a 10.9 a 11.2 a
0.25 0.25 0.24 0.26 0.25
1.6 a 4.8 b 0.6 c 0.3 d 0.0 e
1.0 a 1.2 b 0.8 c 0.4 d -
a a a a a
IThe units of flower set and flower weight are the same as those of fruit set and fruit weight respectively. eMeans followed by different letters within each column differ significantly at 5% level according to Duncan's multiple range test.
103
GROWTH OF C. UNSHIU IN RELATION TO ASSIMILATES
TABLE 2 Effect of girdling, old leaf darkening and defoliation applied at Day 10 PFB on leaf area (cm 2) of young spring-flush leaves of C. unshiu trees Treatment
Days after treatment
Control Girdling Defoliation Leaf darkening Girdling + defoliation
0
30
0.67 a 0.68 a 0.68 a 0.66 a 0.68 a
5.20 a 7.12 b 4.71 c 3.90 d 3.50 d
'Means followed by different letters within each column differ significantly at 5% level according to Duncan's multiple range test
8
,-8
;-
young leaf
A
m
0 0
6
old leaf
4
0
E e,. O.
2
0
0 -20
0
20
40
60
days after full bloom Fig. 1. Comparison of Pn changes between young spring-flush leaves and old leaves on autumnshoots of the preceding year. Each point represents the mean + SE.
current photosynthate supply. Results from a '4CO2 labelling experiment indicate that more than one third of current photosynthates in old leaves are exported to young fruit and leaves during the fruiting process of C. unshiu (Akao et al., 1981 ). The inhibition of leaf photosynthesis results in similar negative effects on fruit set and growth in sweet orange (Moss, 1976), peach (Byers et al., 1985) and apple (Byers et al., 1985; Beruter and Droz, 1991 ). Our results also support the view that the supply of photoassimilate is critical
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for fruit retention in early stage of fruit development (Patrick, 1988; Beruter and Droz, 1991 ). For fruit bearing satsuma mandarin trees, carbohydrate reserves are mainly stored in large roots and the crown (Akao et al., 1981 ). In this study the pos. sible contribution of stored reserves in these organs could not be directly determined. However, the obvious positive effect of girdling with maintenance ofold leaves on fruit set and growth (Tables 1 and 2), suggests that the amount of assimilates transported to fruitlet and young leaves from the stored reserves in the roots and crown, were not a significant assimilate source to support the development of these organs. The defoliation treatment with girdling led to no fruit retention and inhibited young leaf growth to a greater extent than defoliation without girdling (Tables 1 and 2 ), suggesting the stored assimilates within trunks might be transported to fruitlets and young leaves in case of old leaf abscission. Previous studies indicated that carbohydrate reserves in old leaves of C. unshiu will not be mobilized until senescence usually about 2-3 weeks before leaf drop (Ruan et al., 1987; Wu et al., 1988). In the east of China, the old leaf abscission occurs about 4 months after flowering. Thus, it is unlikely that carbohydrate reserves in the old leaves are exported to fruitlets and young leaves. It has been demonstrated that in fruit trees nitrogen is mainly stored in perennial woody tissues (roots and stems) over winter and is always used predominately for young fruit and leaf growth next spring regardless of the current nitrogen supply (Erner, 1989; Millard and Thomson, 1989). Thus, defoliation of old leaves appears not to alter the nitrogen supply to fruitlets and young leaves. As the major transfer process involved in providing nitrogen to young developing organs in the stem is via xylem (Dasilva and Shelp, 1990), girdling the phloem of the shoots would not affect the nitrogen transport to the developing sinks. In order to determine the effect of girdling, defoliation and leaf darkening on the hormonal balance of the young fruits, their endogenous GA3, ZT and ABA contents were analysed. No significant difference was found among these treatments, the average contents of GA3, ZT and ABA in the fruitlets at Day 20 AFB were 0.35, 2.60 and 0.21/~g g-~ FW, respectively. This result further verifies that the effects of girdling, defoliation and leaf darkening on fruitier growth is mainly a function of photoassimilate supply. The fact that the hormonal balance in the fruitlets was not affected by girdling, old leaf darkening and defoliation implies that hormones in the fruitlets may be synthesized locally as proposed by Erner (1989). It is interesting to note that fruit drop and growth inhibition caused by defoliation were less than that caused by leaf darkening (Tables I and 2 ). Darkening the old leaves may have changed them from a source to sink model owing to their considerable dark respiration (Akao et al., 1981; Ruan et al.,
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TABLE 3 Effect of girdling and old leaf darkening applied at Day 20 AFB on fruit weight (grams FW per fruit ) ofC. unshiu trees Treatment
Control Girdling Leaf darkening
Days after treatment 0
15
30
1.2 a ~ 1.2 a 1.1 a
3.8 a 3.6 a 2.6 b
6.5 a 6.7 a 6.2 a
1Means followed by different letters within each column differ significantly at 5% level according to Duncan's multiple test.
1988). Thus the darkened old leaves might compete with the meristematic sinks for assimilate supply, accentuating fruit drop and growth inhibition.
Effects of treatments applied AFB (Experiment 2) Assimilate patterns are altered by the changes in demand by developing sinks (Patrick, 1988). Therefore the effects of assimilate supply manipulation AFB were studied. Sink strength is positively correlated with sink volume (Patrick, 1988). The assimilate supply for the rapidly growing citrus fruit would be enhanced after fruit set, which could lead to the decrease, even cessation, of photoassimilate export out of the girdling position. The fact that girdling applied at Day 20 AFB did not affect young fruit weight increase (Table 3 ) is consistent with this proposal. The Pn of old leaves began to decrease dramatically from Day 20 AFB (Fig. l ). It suggests that current photosynthate supply from old leaves to developing fruits decreases after Day 20 AFB. This is consistent with the fact that 30 days of darkening old leaves did not result in fruit growth inhibition (Table 3 ). The net photosynthate export of young leaves of C. unshiu commences at the stage of 40% of their final leaf area (Liu et al., 1989 ). In this experiment the young leaves reached 48% of their final leaf area at Day 20 AFB and thus can be regarded as a 'net source leaves'. This corresponds to the time that the Pn of the young leaves exceeds that of the old leaves which continues to decline (Fig. l ). These results indicate that young leaves gradually substitute for old leaves to supply current photosynthates to the developing fruits after Day 20 AFB. CONCLUSIONS
The assimilate supply for fruitlets and young leaves at the early stage of fruit ontogeny mainly originates from current photosynthesis of the old leaves.
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This conclusion is verified by the fact that in south and east China old leaf abscission of field-grown trees, owing to cold injury in winter and early spring, leads to poor fruit set (Ruan et al., 1987; Wu et al., 1988). Fruit drop was greatly accentuated by leaf darkening during flowering and fruiting processes. This suggests that reduced solar radiation by shading or by overcast weather in this period could be a major hindrance to the fruit sei. The response emphasises the importance of maintaining an optimum light environment within citrus groves. On Day 20 AFB, the young leaves, considered to be a net exporter of photosynthates, increased in Pn rapidly and substituted for the old leaves to supply current photosynthates to the developing fruits. ACKNOWLEDGEMENT
I would like to thank Professor John W. Patrick for his helpful comments and critical reading of the manuscript. REFERENCES Ak~,:, S., Tsukahara, S., Hisada, H. and Ona, S., 1981. Contribution of photosynthetic assimilates to development of flower and spring flush in Citrus unshiu Marc. J. Jpn. Soc. Hortic. Sci., 50: 1-9. Beruter, J. and Droz, Ph., 1991. Studies on locating the signal for fruit abscission in the apple tree. Scientia Hortic., 46:201-214. Byers, R.E., Lyons, Jr., J.C., Yoder, K.S., Barden, J.A. and Young, R.W., 1985. Peach and apple thinning by shading and photosynthetic inhibition. J. Hortic. Sci., 60: 465-475. Dasiva, M.C. and Shelp, B.J., 1990. Xylem-to-phloem transfer of organic nitrogen in young soybean plants. Plant Physiol., 92: 797-80~. Erner, Y., 1989. Citrus fruit set: carbohydrate, hormone and leaf mineral relationships. In: C.J. Wright (Editor), Manipulation of Fruiting. Academic Pre~s, Butterworths, London, pp. 233242. Garcia-Luis, A., Fornes, F., Sanz, A. and Guardiola, J.L., 1988. The regulation of flowering and fruit set in citrus: relationship with carbohydrate levels. Israel J. Bot., 37:189-201. Ho, L.C., Grange, R.I. and Shaw, A.F., 1989. Source/sink regulation. In: D.A. Baker and J.A. Milburn (Editors), Transport of Photoassimilates. Longman, Harlow, UK, pp. 319-320. Liu, D.-H., Zhang, S.-L., Wu, G.-L. and Zhang, L.-C., 1989. Studies on photosynthate translocation in Citrus unshiu. Acta Hortic.. 16:191 - 198 (in Chinese with English abstract ). Millard, P. and Thomson, C.M., 1989. The effect ofautumn senescence of leaves on the internal cycling of nitrogen for the spring growth of apple trees. J. Expl. Bot., 40:1285-1289. Moss, G.L., 1976. Thinning 'Washington' navel and 'late valencia' sweet orange fruits with photosynthetic inhibitor. HortScience, I 1: 48-50. Patrick, J.W., 1988. Assimilate partitioning in relation to crop prc,ductivity. HortScience, 23: 33-40. Ruan, Y.-L., Zhang, L.-C., Wu, G.-L. and Zhang, S.-L., 1987. Changes of net photosynthetic rate of wintering citrus leaves. Acta Agric., 13:392-395 (in Chinese with English abstract ). Ruan, Y.-L., Zhang, L.-C., Wu, G.-L. and Zhang, S.-L., 1988. Effect of water stress on photosynthesis of Citrus unshiu. Acta Hortic., 15:93-98 (in Chinese with English abstract ). Ruan, Y.-L., Zhang, S.-L., Zhu, K., Wu, G.-L. and Li, S.-J., 1991. The types of cytokinins and
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their changes and abscisic acid in the stem of Citrus unshiu during the period of flower bud initiation. Scientia Agric., 24:55-59 (in Chinese with English abstract ). Ryugo, K., 1985. Promotion and inhibition of flower initiation and fruit set by plant manipulation and hormones. Acta Hortic., 1:301-305. Weaver, R.J. and Johnson, J.O., 1985. Relation of hormones to nutrient mobilization and the internal environment of the plant: The supply of mineral nutrients and and photosynthate. In: R.P. Pharis, and D.M. Reid (Editors), Hormonal Regulation of Development. III Role of Environmental Factors. Encyclopedia of Plant Physiology, Vol. II. Springer, Berlin, pp. 3-36. Wu, G.-L., Zhang, L.-C., Zhang, F., Ruan, Y.-L. and Liu, J.-S., 1988. A study on the physiology of overwintering satsuma trees under straw sheds. China Citrus, 17:3-6 (in Chinese ).