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Decline of persimmon (Diospyros kaki L.) trees on Diospyros virginiana rootstocks
Scientia Horticulturae, 48 ( 1991 ) 6 1 - 7 0 Elsevier Science Publishers B.V., A m s t e r d a m
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Decline o f p e r s i m m o n (Diospyros kaki L. ) trees on Diospyros virginiana rootstocks Y. Cohen a, A. Gur ~, Z. Barkai b and A. Blumenfeld c aDepartment o f Horticulture, The Hebrew University o f Jerusalem, Rehovot, Israel bMinistry o f Agriculture, Extension Service, Tel-Aviv, Israel Department of Fruit Trees, Agricultural Research Organization, The Volcani Centre, Bet-Dagan. Israel (Accepted 25 January 199 i )
ABSTRACT Cohen, Y., Gur, A., Barkai, Z. and Blumenfeld, A., 1991. Decline o f persimmon (Diospyros kaki L. ) frees on Diospyros virginiana rootstocks. Scientia Horlic., 48:61-70. Persimmon trees (Diospyros kaki L. ) grafted on Diospyros virginiana L. rootstock seedlings frequently grow very poorly and show several decline symptoms. Rootstock and scion girths o f declining trees were smaller than those o f healthy trees. Trees grafted on D. kaki rootstocks did not decline. In declining trees, the wood colour close to the graft union changed from yellow to brown, and gum accumulated in the phloem rays and xylem vessels, partly blocking them. The water transport in this area was impeded and the water potential o f shoots o f declining trees was less than that o f healthy trees. Scions from both healthy and declining trees were grafted on the same healthy D. virginiana rootstocks. The healthy scions were influenced by their unhealthy neighbours and grew poorly. Plant material obtained from these originally healthy scions also grew poorly when grafted on healthy D. virginiana rootstocks, indicating that the healthy scions were somehow infected by their unhealthy neighbours. Removing scions o f young declining trees and replacing them with healthy scions also resulted in reduced growth, compared with healthy ~ootstocks grafted with healthy scions. These results suggest the presence o f a transmittable biotic factor, both in D. virginiana rootstocks and D. kaki scions, causing decline only in D. kaki scion tissues. Keywords: decline; graft incompatibility; persimmon. Abbreviation: C = the ratio o f the velocity o f the transpiration stream at the zone o f the graft union and its average velocity in the rootstock and the scion.
INTRODUCTION
In recent years, cases o f decline o f p e r s i m m o n trees (Diospyros kaki L.) grafted on Diospyros virginiana L. ('the American p e r s i m m o n ' ) rootstock have been observed in Israel by A. B l u m e n f e l d and S. A n t m a n ( u n p u b l i s h e d data, 1990). This p h e n o m e n o n o f decline is widespread in nurseries and or0 3 0 4 - 4 2 3 8 / 9 ! / $ 0 3 . 5 0 © 1991 Elsevier Science Publishers B.V. All rights reserved.
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Y. COHEN ET AL.
chards, and includes all cultivated varieties. A similar phenomenon has been reported in the USA (Chandler, 1951; Childers, 1978 ). The symptoms of this decline are feeble growth, small upwards-folded leaves, swollen buds, protruding lenticels, dense internodes and sometimes shallow bark cracks in the zone of the graft union. Branching of the trees is much reduced and mineral levels in the leaves are low (Tsur, 1984). It has been shown that D. kaki trees grafted on D. virginiana seedlings obtained from different trees vary markedly in their respective rates of decline (A. Blumenfeld and S. Antman, unpublished data, 1990). Persimmon trees subjected to stress conditions, such as drought, trunk ringing, bending of branches and insect attack, developed similar morphological symptoms to trees suffering from decline (Tsur, 1984). The causes of the decline of the persimmon trees on D. virginiana rootstocks are unknown. Trials to isolate microorganisms responsible for the decline have been unsuccessful so far (M. Bar-Yoseph and A. Steinberg personal communication, 1990). The decline of interspecific graft combinations due to graft incompatibility may be caused by biochemical or pathological factors. In both cases, anatomical abnormalities occur at the graft union or close to the union. Pathological factors causing such abnormalities include viruses and mycop~asmas active only in certain rootstock-scion combinations, e.g. tristeza in citrus (Wallace, 1978), certain virus diseases of apples (Stouffer et al., 1977) and cherries (Schneider, 1945). In certain cases of graft incompatibility, the transpiration stream was found to be impeded at the graft union (Gur and Bium, 1975). We investigated the possibility of a similar situation in the case of declining persimmon trees grafted on D. virginiana rootstocks by comparing the veloc|~y of the transpiration stream in the rootstock and scion to its velocity in the zone of the graft union (Gur and Blum, 1975 ). In the present paper, the possible involvement of a transmittable biotic factor in the incompatibility of D. kaki trees on D. virginiana rootstocks is also investigated. MATERIALS AND METHODS
Plant material. - Tests and observations were conducted in commercial persimmon orchards (cultivar "Triumph') grafted on D. virginiana seedling rootstocks planted in 1974 (Plot a), 1976 (Plot b) and 1979 (Plot c) in the coastal plain of Israel. Many of the trees showed severe or mild symptoms of decline, others were healthy. Another plot used was planted in 1980 in the Hula valley in the north of Israel. The trees were 'Triumph' grafted on seedling rootstocks of D. virginiana and D. kaki rootstocks (Plot d). While many of the trees on the D.
D E C L I N E OF PERSIMMON TREES ON ROOTSTOCKS
63
virginiana rootstocks were declining, all the trees on D kaki rootstocks were healthy. E v a l u a t i o n o f the graft union. - The zone of the :graft union of 10- a n d 12year-old trees ( f r o m Plots a and b ) was cut longitudinally, carefully s m o o t h e d a n d observed macroscopically. A n a t o m i c a l methods. - Small blocks o f wood and b a r k (2 × 1 × 1 cm ) were r e m o v e d from the graft union o f 6-year-old healthy a n d declining trees (Plot c ) . The blocks were softened with boiling water and 50% hydrofluoric acid (Berlyn and Miksche, 1976 ) and then kept in a mixture o f glycerol and ethanol
(1:1). W o o d sections 30/~m thick and b a r k sectio~,s 4 0 / ( m thick were p r e p a r e d with a rotary m i c r o t o m e . Staining was by the m e t h o d o f Pianese ( C o n n , 1977). T r u n k girth. - The t r u n k girth in trees o f Plot c was measured annually in winter during 1986-1989, 12 cm a b o v e the graft union a n d 8 em below the graft union. T h e relative velocity o f the transpiration stream. - Short heat pulses were applied to the young xylem and the t i m e the heat pulse reached a thermistor, placed 2 cm above the site of application of the pulse, was measured ( G u t a n d Blum, 1975 ). Tests were p e r f o r m e d during July a n d September between 06:00 and 09,00 h, a n d repeated on seven trees of each group. The m e a s u r e m e n t s were p e r f o r m e d on trees grafted on two rootstocks - 6year-old healthy a n d declining trees ( ' T r i u m p h ' ) grafted on D. virginiana rootstocks (Plots c a n d d ) , and healthy trees of the same age and cultivar grafted on D. k a k i rootstocks ( Plot d ). The velocity o f the transpiration stream was established: ( 1 ) in the rootstock, 10 cm below the graft union ( = a ); ( 2 ) in the scion, 10 cm above the graft union ( = b), ( 3 ) at the graft union ( = c). A coefficient t e r m e d the water t r a n s p o r t coefficient ( C ) was calculated according to the f o r m u l a C = 2 c / ( a + b ). T h e water p o t e n t i a l o f leafy shoots. - Apical shoots 6 - 8 cm long and bearing 2 - 3 leaves were severed from p e r s i m m o n tree branches void of fruits. The wa',er potential was established with the aid of a 250 ml pressure c h a m b e r ( S h o l a n d e r et al., 1965 ). Shoots were sampled from the healthy and declining trees o f Plot c, in S e p t e m b e r between 06:00 a n d 07:(~0 h, when the leaf w a t e r content is maximal. Two shoots were sampled from each of seven trees per tree group. I n v o l v e m e n t o f a transmittable biotic f a c t o r iii ihe decline p h e n o m e n o n . - Scion
64
V. COHEN ET AL.
wood from healthy and declining persimmon trees ( ' T r i u m p h ' ) was cleft grafted on l-year-old D. virginiana rootstocks varying in origin. "Healthy rootstocks' were D. virginiana seedlings originating from mother trees characterized by a 1o,;,, rate of decline of trees grafted on (6.4% in an experimental plot in Israel, a.-, against 23-43% of trees grafted on seedlings from other sources; A. Tzuk, personal communication, 1989). 'Non-healthy rootstocks' were obtained from declining 2-year-old trees which were cut back beneath the graft union. Two series of grafts were used in the experiments. The 1987 series included five graft combinations with five plants per combination. Two scions were grafted on each l-year-old rootstock and on each scion only one shoot was allowed to grow out. Plants were grafted in April 1987 and grown for two subsequent summer seasons. The graft combinations for 1987 and 1988 are presented in Fig. 2 (see below). The 1988 series included four combinations, with 20 plants per combination. One scion was side grafted on each rootstock and allowed to develop two shoots. Plants were grafted in April 1988 and grown for one season. At the end of the experiments (August-September, 1988), shoot elongation and the fresh weight of the scion were established. Statistics. - Data were analysed according to analysis of variance. In experi-
ments with F values equal or exceeding the 5% level of ~ignificance, separation of the means was by the S t u d e n t - N e w m a n - K e u l s multiple-range test. RESULTS AND DISCUSSION
Trees in Plot c were classified as healthy, mildly declining and severely declining according to the symptoms of decline specified in the Introduction. These groups differed significantly in their scion and rootstock girth at the age of 7 years, when annual measurements were started, and up to the age of 10 years when they were terminated (Table 1 ). Persimmon trees grafted on D. k a k i rootstocks (Plot d) were uniform in size and did not show symptoms &decline. TABLE i The scion and rootstock girth o f 10-year-old healthy and decl:.ning persimmon trees ( ' T r i u m p h ' ) grafted on D. virginiana rootstocks (Plot c, seven replicates per tree group) Condition of trees
Healthy Mildly declining Severely declining
Girth ( r a m ) Rootstock
Scion
493 a 452 b 396 c
349 a 313b 262 c
DECLINE OF PERSIMMON TREES ON ROOTSTOCKS
65
Anatomicalobservations. - A marked wood discoloration from yellow to brown occurred at the graft union of declining trees, both in the rootstock and the scion. In other cases, a dark line appeared along the graft union in the wood, marking the borderline between the scion and the rootstock. In a few trees, shallow cracks appeared in the bark along the graft union, apparently without causing any vascular discontinuity. The scion bark close to the graft union in declining trees was thinner than that in healthy ones and numerous necrotic brown spots appeared in the secondary phloem. In the rootstock bark, similar brown spots appeared, but less frequently. In the cells of the phloem and the xylem rays, and on the perforation plates of the xylem vessels close to the graft union of declining trees, a dark gum mass appeared (Fig. 1 (a) and 1 ( b ) ) . The gum blocked the perforation plates to a large extent (Fig. 1 ( c ) ) . No indication of vascular discontinuity between the scion and the rootstock of declining trees was found. Anatomical abnormalities, like those mentioned above, which were very pronounced in declining trees, appeared only very rarely in healthy trees. An accumulation of gum in phloem and xylem vessels has been shown to develop in a virus disease of cherries which only occurs in certain rootstockscion combinations (the buckskin disease) (Schneider, 1945 ). Cracks in tb~ bark close to the graft union have also been described in cases of graft incompatibility; however, contrary to persimmon on D. virginiana rootstock such cracks are generally deeper and interfere with the continuity of phloem elements (Gur et al., 1968 ). The wood discoloration close to the graft union of persimmon also occurred in other cases of graft incompatibility (Mosse, 1958; Feucht et al., 1986 ) and is, apparently, caused by the oxidation of phenols. Velocity o f ;he transpiration stream and water potential - The ratio betwee~a the velocity of the transpiration stream at the zone of the graft union and its average velocity in the scion and rootstock (water transport coefficient, C) was much higher in declining trees grafted on D. virginiana, compared with healthy ones in two orchards (Table 2 ). Healthy trees on D. kaki or D. virginiana rootstocks had C values close to one, which points towards a compatible graft union (Gur and Blum, 1975). Declining tr~,es had much higher C values and mildly declining trees had intermediate values. An increase in the velocity of the transpiration stream in the partly obstructed vessels, close to the graft union, is in accordance with the laws of hydrodynamics. The gum accumulation in the xylem vessels, and particularly on the perforation plates, of declining persimmon trees seems to be the reason for this obstruction. A negative correlation (R = --0.631") was found between the scion trunk girth ofD. kaki trees on D. virginiana rootstocks and their C values (in Plot c). As a result of the obstruction to the water flow in declining trees, the shoot water potential decreased from - 0 . 4 8 MPa in healthy to - 0 . 5 7 MPa in declining trees, the difference being statistically significant (at the 5% level).
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Y. C O H E N ET AL
Fig. I. Tangential sections at the zone of the graft union o f 6-year-old declining persimmon trees ('Triumph") grafted on D. vh'giniana rootstocks (Plot c). (a) Phloem ( magnification × 63 ): (b) xylem (magnification × 25 ): (c) tracheae (magnification × 250 ).
DECLINE OF PERSIMMON TREES ON ROOTSTOCKS
67
Fig. 1. C o n t i n u e d . TABLE 2 T h e w a t e r t r a n s p o r t c o e f f i c i e n t ( C ) o f 6 - y e a r - o l d h e a l t h y a n d d e c l i n i n g p e r s i m m o n trees ( ' T r i u m p h " ) g r a f t e d on two species o f p e r s i m m o n r o o t s t o c k s ( s e v e n replicates p e r tree g r o u p ) C o n d i t i o n o f trees
Healthy Mildly declining Severely d e c l i n i n g
Plot c
Plot d
D. virginiana ~
D. v i r g i n i a n a ~
D. k a k i
1.01 b i.20 a b 1.37 a
! .02 b 1.32 a
1.00 -
iRootstock.
The similarity of the symptoms of water stress and the decline phenomenon (Tzur, 1984) also points toward the close relationship between the two. I n v o h , e m e n t o f a t r a n s m i t t a b l e f a c t o r . - All the graft combinations of the 1987
68
Y. C O H E N ET A L
series, which contained at least one component derived from declining trees (scion or rootstock), grew poorly and their shoot elongation was lower than that of plants with scions and rootstocks obtained from normal trees (Fig. 2 ( A ) ) . Shoot elongation was most severely depressed when both the scion and the rootstock originated from declining trees. Healthy scions grafted together with scions from declining trees on the same rootstock also grew weakly. Scions from normal trees, when grafted on rootstocks from which declining scions had been removed previously, grew weaker than healthy controls. In the 1988 series of grafts (Fig. 2 ( B ) ) bud wood obtained from healthy scions, which were growing in 1987 together with scions from declining trees on the same rootstocks, was grafted on healthy rootstocks and showed poor shoot growth compared with healthy control plants. The results of the two series of grafts point toward the transmission of a biotic factor from declining scions to healthy ones and, in the following year, to the transmission of this factor by scions from the 'infected' shoots of 1987 to a second generation of grafts. Furthermore, transmittance of this biotic factor from 'infected' D. virginiana 100 -
o
7s-
I ®
I
50
Hl
o 2s
I
I
I
1
b bc
D 0 \
I
° I
°
Fig. 2. Shoot elongation of D. kaki scions obtained from healthy (H) and declining (D) trees, and grafted on healthy D. virginiana (H) rootstocks and on rootstocks of the same species from which declining scions had been removed prior to regrafting them with healthy and declining scions (D). (A) Two scions per plant were grafted in April 1987. Records are o f August 1988. There were five replicates per graft combination. (B) One scion per plant was grafted in April 1988 and records are of September 1988. In two combinations, the scions were obtained from grafts of the previous year, as indicated by arrows. There were 20 replicates per graft combination. Data marked with the same letter are not significantly different at the 5% level.
DECLINE OF PERSIMMON TREES ON ROOTSTOCKS
69
rootstocks, which had previously carried declining D. kaki scions, to healthy D. kaki scions was shown to occur as well, causing a reduction in the growth of such scions. However, the severity of the decline caused by 'infected' ~cions was more pronounced than that caused by infected rootstocks (Fig. 2 ). Diospyros virginiana seedlings obtained from different source trees var!ed in the amount of decline they induced in D. kaki scions grafted upon (see Materials and Methods). In non-grafted seedlings, the source of the seeds did not influence vigour considerably. Hence, a transmittable biotic factor (virus, viroid or mycoplasma) exists in latent form in certain D. virginiana trees without showing any symptoms. This factor may, apparently, be transmitted by seeds and passes from infected rootstocks into D. kaki scions grafted on them, causing the symptoms of decline there. The transmission of this factor by seeds may explain the differences in susceptibility to the disease of seedling rootstocks obtained from different tree sources. The factor is also easily transmitted by infected scion wood. In horticultural practice, the transmission of the disease by infected scions is unlikely as the disease is easily recognized in the orchard and bud wood from affected trees will certainly be avoided. However, the involvement of biological vectors in the transmission of the disease in the orchard is not impossible and should be investigated. The transmission of viruses and mycoplasmas by seeds is known to occur in several tree species such as cherries, apples, walnuts and avocados (Wallace and Drake, 1962; Nemeth, 1986). In the case of the avocado 'sun-blotch' viroid, the agent exists latently in the seedlings and develops symptoms in scions grafted on them, as is also the case in the persimmon decline dealt with here. ACKNOWLEDGEMENTS
We thank M. Simchi and Y. Moyal for their assistance in various phases of the research, and Z. Mor for her assistance in the statistical evaluation of the results. REFERENCES Berlyn, G.P. and Miksche, P., 1976. Botanical Microtechnique and Cytochemistry. Iowa State University Press, Ames, IA, 73 pp. Chandler, W.H., 195 I. Deciduous Orchards. 2nd Edn. Lea & Febiger, Philadelphia, PA, 386 PP. Childers, N.F., 1978. Modern Fruit Science. 8th Edn. Rutgers University Horticultural Publications, New Brunswick, N J, 500 pp. Conn, H.J., 1977. In: Biological Stains. R.D. Lillie (Editor), 9th Edn. Williams & Wilkins, Baltimore, MD, 504 pp. Feucht, W., Schmid, P.P.S. and Chrisl, E., 1986. Qualitative changes in the phloem of cherry trees: Standard and transmitted-light fluorescence microscopy. Zeiss Info, 28: 47-48.
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Gur, A. and Bium, A., 1975. The water conductivity o f defective graft unions. J. Am. Soc. Hortic. Sci., 100: 325-328. Gur, A., Samish, R.M. and Lifshitz, Z., 1968. The role of the cyanogenic glucoside of the quince in the incompatibility between pear varieties and quince rootstocks. Hortic. Res., 8: | 13134. Mosse, B., ! 958. Further observations on growth and union structure of double-grafted pear on quince. J. Hortic. Sci., 33: 186-193. Nemeth, M., 1986. Virus, Mycoplasma and Rickettsia Diseases o f Fruit Trees. Martinus Nijhoff Publishers, Budapest, 840 pp. Schneider, H., 1945. Anatomy o f buckskin-diseased peach and cherry. Phytopathology, 35:6 ! 0 635. Sholander, P.E., Hammel, H.T., Bradstreet, E.D. and Hemmengsen, E.A., 1965. Sap pressure in vascular plants. Science, 148: 339-346. Stouffcr, R.F., Hickey, K.D. and Welsh, M.F., 1977. Apple union necrosis and decline. Plant Dis. Rep., 61: 20-24. "Fsur, A., 1984. Leaf folding in persimmon. Alon Hanotea, 38:803 (in Hebrew). Wallace, J.M., 1978. Virus and virus like diseases. In: W. Reuther, E.C. Calavan and G.E. Carman (Editors), The Citrus Industry. Vol. IV. University of California Division of Agricultural Science, Riverside, CA, pp. 67-184. Wallace, J.M. and Drake, R.J., 1962. A high rate of seed transmission of avocado sun blotch virus from symptomless trees and the origin of such trees. Phytopathology, 52:237-241.
61
Decline o f p e r s i m m o n (Diospyros kaki L. ) trees on Diospyros virginiana rootstocks Y. Cohen a, A. Gur ~, Z. Barkai b and A. Blumenfeld c aDepartment o f Horticulture, The Hebrew University o f Jerusalem, Rehovot, Israel bMinistry o f Agriculture, Extension Service, Tel-Aviv, Israel Department of Fruit Trees, Agricultural Research Organization, The Volcani Centre, Bet-Dagan. Israel (Accepted 25 January 199 i )
ABSTRACT Cohen, Y., Gur, A., Barkai, Z. and Blumenfeld, A., 1991. Decline o f persimmon (Diospyros kaki L. ) frees on Diospyros virginiana rootstocks. Scientia Horlic., 48:61-70. Persimmon trees (Diospyros kaki L. ) grafted on Diospyros virginiana L. rootstock seedlings frequently grow very poorly and show several decline symptoms. Rootstock and scion girths o f declining trees were smaller than those o f healthy trees. Trees grafted on D. kaki rootstocks did not decline. In declining trees, the wood colour close to the graft union changed from yellow to brown, and gum accumulated in the phloem rays and xylem vessels, partly blocking them. The water transport in this area was impeded and the water potential o f shoots o f declining trees was less than that o f healthy trees. Scions from both healthy and declining trees were grafted on the same healthy D. virginiana rootstocks. The healthy scions were influenced by their unhealthy neighbours and grew poorly. Plant material obtained from these originally healthy scions also grew poorly when grafted on healthy D. virginiana rootstocks, indicating that the healthy scions were somehow infected by their unhealthy neighbours. Removing scions o f young declining trees and replacing them with healthy scions also resulted in reduced growth, compared with healthy ~ootstocks grafted with healthy scions. These results suggest the presence o f a transmittable biotic factor, both in D. virginiana rootstocks and D. kaki scions, causing decline only in D. kaki scion tissues. Keywords: decline; graft incompatibility; persimmon. Abbreviation: C = the ratio o f the velocity o f the transpiration stream at the zone o f the graft union and its average velocity in the rootstock and the scion.
INTRODUCTION
In recent years, cases o f decline o f p e r s i m m o n trees (Diospyros kaki L.) grafted on Diospyros virginiana L. ('the American p e r s i m m o n ' ) rootstock have been observed in Israel by A. B l u m e n f e l d and S. A n t m a n ( u n p u b l i s h e d data, 1990). This p h e n o m e n o n o f decline is widespread in nurseries and or0 3 0 4 - 4 2 3 8 / 9 ! / $ 0 3 . 5 0 © 1991 Elsevier Science Publishers B.V. All rights reserved.
62
Y. COHEN ET AL.
chards, and includes all cultivated varieties. A similar phenomenon has been reported in the USA (Chandler, 1951; Childers, 1978 ). The symptoms of this decline are feeble growth, small upwards-folded leaves, swollen buds, protruding lenticels, dense internodes and sometimes shallow bark cracks in the zone of the graft union. Branching of the trees is much reduced and mineral levels in the leaves are low (Tsur, 1984). It has been shown that D. kaki trees grafted on D. virginiana seedlings obtained from different trees vary markedly in their respective rates of decline (A. Blumenfeld and S. Antman, unpublished data, 1990). Persimmon trees subjected to stress conditions, such as drought, trunk ringing, bending of branches and insect attack, developed similar morphological symptoms to trees suffering from decline (Tsur, 1984). The causes of the decline of the persimmon trees on D. virginiana rootstocks are unknown. Trials to isolate microorganisms responsible for the decline have been unsuccessful so far (M. Bar-Yoseph and A. Steinberg personal communication, 1990). The decline of interspecific graft combinations due to graft incompatibility may be caused by biochemical or pathological factors. In both cases, anatomical abnormalities occur at the graft union or close to the union. Pathological factors causing such abnormalities include viruses and mycop~asmas active only in certain rootstock-scion combinations, e.g. tristeza in citrus (Wallace, 1978), certain virus diseases of apples (Stouffer et al., 1977) and cherries (Schneider, 1945). In certain cases of graft incompatibility, the transpiration stream was found to be impeded at the graft union (Gur and Bium, 1975). We investigated the possibility of a similar situation in the case of declining persimmon trees grafted on D. virginiana rootstocks by comparing the veloc|~y of the transpiration stream in the rootstock and scion to its velocity in the zone of the graft union (Gur and Blum, 1975 ). In the present paper, the possible involvement of a transmittable biotic factor in the incompatibility of D. kaki trees on D. virginiana rootstocks is also investigated. MATERIALS AND METHODS
Plant material. - Tests and observations were conducted in commercial persimmon orchards (cultivar "Triumph') grafted on D. virginiana seedling rootstocks planted in 1974 (Plot a), 1976 (Plot b) and 1979 (Plot c) in the coastal plain of Israel. Many of the trees showed severe or mild symptoms of decline, others were healthy. Another plot used was planted in 1980 in the Hula valley in the north of Israel. The trees were 'Triumph' grafted on seedling rootstocks of D. virginiana and D. kaki rootstocks (Plot d). While many of the trees on the D.
D E C L I N E OF PERSIMMON TREES ON ROOTSTOCKS
63
virginiana rootstocks were declining, all the trees on D kaki rootstocks were healthy. E v a l u a t i o n o f the graft union. - The zone of the :graft union of 10- a n d 12year-old trees ( f r o m Plots a and b ) was cut longitudinally, carefully s m o o t h e d a n d observed macroscopically. A n a t o m i c a l methods. - Small blocks o f wood and b a r k (2 × 1 × 1 cm ) were r e m o v e d from the graft union o f 6-year-old healthy a n d declining trees (Plot c ) . The blocks were softened with boiling water and 50% hydrofluoric acid (Berlyn and Miksche, 1976 ) and then kept in a mixture o f glycerol and ethanol
(1:1). W o o d sections 30/~m thick and b a r k sectio~,s 4 0 / ( m thick were p r e p a r e d with a rotary m i c r o t o m e . Staining was by the m e t h o d o f Pianese ( C o n n , 1977). T r u n k girth. - The t r u n k girth in trees o f Plot c was measured annually in winter during 1986-1989, 12 cm a b o v e the graft union a n d 8 em below the graft union. T h e relative velocity o f the transpiration stream. - Short heat pulses were applied to the young xylem and the t i m e the heat pulse reached a thermistor, placed 2 cm above the site of application of the pulse, was measured ( G u t a n d Blum, 1975 ). Tests were p e r f o r m e d during July a n d September between 06:00 and 09,00 h, a n d repeated on seven trees of each group. The m e a s u r e m e n t s were p e r f o r m e d on trees grafted on two rootstocks - 6year-old healthy a n d declining trees ( ' T r i u m p h ' ) grafted on D. virginiana rootstocks (Plots c a n d d ) , and healthy trees of the same age and cultivar grafted on D. k a k i rootstocks ( Plot d ). The velocity o f the transpiration stream was established: ( 1 ) in the rootstock, 10 cm below the graft union ( = a ); ( 2 ) in the scion, 10 cm above the graft union ( = b), ( 3 ) at the graft union ( = c). A coefficient t e r m e d the water t r a n s p o r t coefficient ( C ) was calculated according to the f o r m u l a C = 2 c / ( a + b ). T h e water p o t e n t i a l o f leafy shoots. - Apical shoots 6 - 8 cm long and bearing 2 - 3 leaves were severed from p e r s i m m o n tree branches void of fruits. The wa',er potential was established with the aid of a 250 ml pressure c h a m b e r ( S h o l a n d e r et al., 1965 ). Shoots were sampled from the healthy and declining trees o f Plot c, in S e p t e m b e r between 06:00 a n d 07:(~0 h, when the leaf w a t e r content is maximal. Two shoots were sampled from each of seven trees per tree group. I n v o l v e m e n t o f a transmittable biotic f a c t o r iii ihe decline p h e n o m e n o n . - Scion
64
V. COHEN ET AL.
wood from healthy and declining persimmon trees ( ' T r i u m p h ' ) was cleft grafted on l-year-old D. virginiana rootstocks varying in origin. "Healthy rootstocks' were D. virginiana seedlings originating from mother trees characterized by a 1o,;,, rate of decline of trees grafted on (6.4% in an experimental plot in Israel, a.-, against 23-43% of trees grafted on seedlings from other sources; A. Tzuk, personal communication, 1989). 'Non-healthy rootstocks' were obtained from declining 2-year-old trees which were cut back beneath the graft union. Two series of grafts were used in the experiments. The 1987 series included five graft combinations with five plants per combination. Two scions were grafted on each l-year-old rootstock and on each scion only one shoot was allowed to grow out. Plants were grafted in April 1987 and grown for two subsequent summer seasons. The graft combinations for 1987 and 1988 are presented in Fig. 2 (see below). The 1988 series included four combinations, with 20 plants per combination. One scion was side grafted on each rootstock and allowed to develop two shoots. Plants were grafted in April 1988 and grown for one season. At the end of the experiments (August-September, 1988), shoot elongation and the fresh weight of the scion were established. Statistics. - Data were analysed according to analysis of variance. In experi-
ments with F values equal or exceeding the 5% level of ~ignificance, separation of the means was by the S t u d e n t - N e w m a n - K e u l s multiple-range test. RESULTS AND DISCUSSION
Trees in Plot c were classified as healthy, mildly declining and severely declining according to the symptoms of decline specified in the Introduction. These groups differed significantly in their scion and rootstock girth at the age of 7 years, when annual measurements were started, and up to the age of 10 years when they were terminated (Table 1 ). Persimmon trees grafted on D. k a k i rootstocks (Plot d) were uniform in size and did not show symptoms &decline. TABLE i The scion and rootstock girth o f 10-year-old healthy and decl:.ning persimmon trees ( ' T r i u m p h ' ) grafted on D. virginiana rootstocks (Plot c, seven replicates per tree group) Condition of trees
Healthy Mildly declining Severely declining
Girth ( r a m ) Rootstock
Scion
493 a 452 b 396 c
349 a 313b 262 c
DECLINE OF PERSIMMON TREES ON ROOTSTOCKS
65
Anatomicalobservations. - A marked wood discoloration from yellow to brown occurred at the graft union of declining trees, both in the rootstock and the scion. In other cases, a dark line appeared along the graft union in the wood, marking the borderline between the scion and the rootstock. In a few trees, shallow cracks appeared in the bark along the graft union, apparently without causing any vascular discontinuity. The scion bark close to the graft union in declining trees was thinner than that in healthy ones and numerous necrotic brown spots appeared in the secondary phloem. In the rootstock bark, similar brown spots appeared, but less frequently. In the cells of the phloem and the xylem rays, and on the perforation plates of the xylem vessels close to the graft union of declining trees, a dark gum mass appeared (Fig. 1 (a) and 1 ( b ) ) . The gum blocked the perforation plates to a large extent (Fig. 1 ( c ) ) . No indication of vascular discontinuity between the scion and the rootstock of declining trees was found. Anatomical abnormalities, like those mentioned above, which were very pronounced in declining trees, appeared only very rarely in healthy trees. An accumulation of gum in phloem and xylem vessels has been shown to develop in a virus disease of cherries which only occurs in certain rootstockscion combinations (the buckskin disease) (Schneider, 1945 ). Cracks in tb~ bark close to the graft union have also been described in cases of graft incompatibility; however, contrary to persimmon on D. virginiana rootstock such cracks are generally deeper and interfere with the continuity of phloem elements (Gur et al., 1968 ). The wood discoloration close to the graft union of persimmon also occurred in other cases of graft incompatibility (Mosse, 1958; Feucht et al., 1986 ) and is, apparently, caused by the oxidation of phenols. Velocity o f ;he transpiration stream and water potential - The ratio betwee~a the velocity of the transpiration stream at the zone of the graft union and its average velocity in the scion and rootstock (water transport coefficient, C) was much higher in declining trees grafted on D. virginiana, compared with healthy ones in two orchards (Table 2 ). Healthy trees on D. kaki or D. virginiana rootstocks had C values close to one, which points towards a compatible graft union (Gur and Blum, 1975). Declining tr~,es had much higher C values and mildly declining trees had intermediate values. An increase in the velocity of the transpiration stream in the partly obstructed vessels, close to the graft union, is in accordance with the laws of hydrodynamics. The gum accumulation in the xylem vessels, and particularly on the perforation plates, of declining persimmon trees seems to be the reason for this obstruction. A negative correlation (R = --0.631") was found between the scion trunk girth ofD. kaki trees on D. virginiana rootstocks and their C values (in Plot c). As a result of the obstruction to the water flow in declining trees, the shoot water potential decreased from - 0 . 4 8 MPa in healthy to - 0 . 5 7 MPa in declining trees, the difference being statistically significant (at the 5% level).
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Y. C O H E N ET AL
Fig. I. Tangential sections at the zone of the graft union o f 6-year-old declining persimmon trees ('Triumph") grafted on D. vh'giniana rootstocks (Plot c). (a) Phloem ( magnification × 63 ): (b) xylem (magnification × 25 ): (c) tracheae (magnification × 250 ).
DECLINE OF PERSIMMON TREES ON ROOTSTOCKS
67
Fig. 1. C o n t i n u e d . TABLE 2 T h e w a t e r t r a n s p o r t c o e f f i c i e n t ( C ) o f 6 - y e a r - o l d h e a l t h y a n d d e c l i n i n g p e r s i m m o n trees ( ' T r i u m p h " ) g r a f t e d on two species o f p e r s i m m o n r o o t s t o c k s ( s e v e n replicates p e r tree g r o u p ) C o n d i t i o n o f trees
Healthy Mildly declining Severely d e c l i n i n g
Plot c
Plot d
D. virginiana ~
D. v i r g i n i a n a ~
D. k a k i
1.01 b i.20 a b 1.37 a
! .02 b 1.32 a
1.00 -
iRootstock.
The similarity of the symptoms of water stress and the decline phenomenon (Tzur, 1984) also points toward the close relationship between the two. I n v o h , e m e n t o f a t r a n s m i t t a b l e f a c t o r . - All the graft combinations of the 1987
68
Y. C O H E N ET A L
series, which contained at least one component derived from declining trees (scion or rootstock), grew poorly and their shoot elongation was lower than that of plants with scions and rootstocks obtained from normal trees (Fig. 2 ( A ) ) . Shoot elongation was most severely depressed when both the scion and the rootstock originated from declining trees. Healthy scions grafted together with scions from declining trees on the same rootstock also grew weakly. Scions from normal trees, when grafted on rootstocks from which declining scions had been removed previously, grew weaker than healthy controls. In the 1988 series of grafts (Fig. 2 ( B ) ) bud wood obtained from healthy scions, which were growing in 1987 together with scions from declining trees on the same rootstocks, was grafted on healthy rootstocks and showed poor shoot growth compared with healthy control plants. The results of the two series of grafts point toward the transmission of a biotic factor from declining scions to healthy ones and, in the following year, to the transmission of this factor by scions from the 'infected' shoots of 1987 to a second generation of grafts. Furthermore, transmittance of this biotic factor from 'infected' D. virginiana 100 -
o
7s-
I ®
I
50
Hl
o 2s
I
I
I
1
b bc
D 0 \
I
° I
°
Fig. 2. Shoot elongation of D. kaki scions obtained from healthy (H) and declining (D) trees, and grafted on healthy D. virginiana (H) rootstocks and on rootstocks of the same species from which declining scions had been removed prior to regrafting them with healthy and declining scions (D). (A) Two scions per plant were grafted in April 1987. Records are o f August 1988. There were five replicates per graft combination. (B) One scion per plant was grafted in April 1988 and records are of September 1988. In two combinations, the scions were obtained from grafts of the previous year, as indicated by arrows. There were 20 replicates per graft combination. Data marked with the same letter are not significantly different at the 5% level.
DECLINE OF PERSIMMON TREES ON ROOTSTOCKS
69
rootstocks, which had previously carried declining D. kaki scions, to healthy D. kaki scions was shown to occur as well, causing a reduction in the growth of such scions. However, the severity of the decline caused by 'infected' ~cions was more pronounced than that caused by infected rootstocks (Fig. 2 ). Diospyros virginiana seedlings obtained from different source trees var!ed in the amount of decline they induced in D. kaki scions grafted upon (see Materials and Methods). In non-grafted seedlings, the source of the seeds did not influence vigour considerably. Hence, a transmittable biotic factor (virus, viroid or mycoplasma) exists in latent form in certain D. virginiana trees without showing any symptoms. This factor may, apparently, be transmitted by seeds and passes from infected rootstocks into D. kaki scions grafted on them, causing the symptoms of decline there. The transmission of this factor by seeds may explain the differences in susceptibility to the disease of seedling rootstocks obtained from different tree sources. The factor is also easily transmitted by infected scion wood. In horticultural practice, the transmission of the disease by infected scions is unlikely as the disease is easily recognized in the orchard and bud wood from affected trees will certainly be avoided. However, the involvement of biological vectors in the transmission of the disease in the orchard is not impossible and should be investigated. The transmission of viruses and mycoplasmas by seeds is known to occur in several tree species such as cherries, apples, walnuts and avocados (Wallace and Drake, 1962; Nemeth, 1986). In the case of the avocado 'sun-blotch' viroid, the agent exists latently in the seedlings and develops symptoms in scions grafted on them, as is also the case in the persimmon decline dealt with here. ACKNOWLEDGEMENTS
We thank M. Simchi and Y. Moyal for their assistance in various phases of the research, and Z. Mor for her assistance in the statistical evaluation of the results. REFERENCES Berlyn, G.P. and Miksche, P., 1976. Botanical Microtechnique and Cytochemistry. Iowa State University Press, Ames, IA, 73 pp. Chandler, W.H., 195 I. Deciduous Orchards. 2nd Edn. Lea & Febiger, Philadelphia, PA, 386 PP. Childers, N.F., 1978. Modern Fruit Science. 8th Edn. Rutgers University Horticultural Publications, New Brunswick, N J, 500 pp. Conn, H.J., 1977. In: Biological Stains. R.D. Lillie (Editor), 9th Edn. Williams & Wilkins, Baltimore, MD, 504 pp. Feucht, W., Schmid, P.P.S. and Chrisl, E., 1986. Qualitative changes in the phloem of cherry trees: Standard and transmitted-light fluorescence microscopy. Zeiss Info, 28: 47-48.
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Gur, A. and Bium, A., 1975. The water conductivity o f defective graft unions. J. Am. Soc. Hortic. Sci., 100: 325-328. Gur, A., Samish, R.M. and Lifshitz, Z., 1968. The role of the cyanogenic glucoside of the quince in the incompatibility between pear varieties and quince rootstocks. Hortic. Res., 8: | 13134. Mosse, B., ! 958. Further observations on growth and union structure of double-grafted pear on quince. J. Hortic. Sci., 33: 186-193. Nemeth, M., 1986. Virus, Mycoplasma and Rickettsia Diseases o f Fruit Trees. Martinus Nijhoff Publishers, Budapest, 840 pp. Schneider, H., 1945. Anatomy o f buckskin-diseased peach and cherry. Phytopathology, 35:6 ! 0 635. Sholander, P.E., Hammel, H.T., Bradstreet, E.D. and Hemmengsen, E.A., 1965. Sap pressure in vascular plants. Science, 148: 339-346. Stouffcr, R.F., Hickey, K.D. and Welsh, M.F., 1977. Apple union necrosis and decline. Plant Dis. Rep., 61: 20-24. "Fsur, A., 1984. Leaf folding in persimmon. Alon Hanotea, 38:803 (in Hebrew). Wallace, J.M., 1978. Virus and virus like diseases. In: W. Reuther, E.C. Calavan and G.E. Carman (Editors), The Citrus Industry. Vol. IV. University of California Division of Agricultural Science, Riverside, CA, pp. 67-184. Wallace, J.M. and Drake, R.J., 1962. A high rate of seed transmission of avocado sun blotch virus from symptomless trees and the origin of such trees. Phytopathology, 52:237-241.