J. Mater. Environ. Sci., 2017 Volume 8, Issue 11, Page

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1 Journl of Mterils nd Environmentl Sciences ISSN : Copyright 2017, University of Mohmmed 1er Oujd Morocco J. Mter. Environ. Sci., 2017 Volume 8, Issue 11, Pge Propgtion of citrus rootstocks by stem cutting using uxin pretretments: the cse of citrumelo (Citrus prdisi Mcf. x Poncirus trifolit (L.) Rf.) A. Fdli 1,2, A. El Attoui 1,2, O. Chetto 1,2, D. Boudoudou 1,2, B. A. Tlh 1, R. Benkirne 2, H. Benyhi 1* 1. UR d Améliortion des Plntes et Conservtion des Ressource Phytogénétiques, Institut Ntionl de Recherche Agronomique, PB 256, Kenitr, Morocco. 2. Lbortoire de Botnique, Biotechnologie et Protection des Plntes, Fculté des Sciences, Université Ibn Tofil, PB 133, Kenitr, Morocco. Received 03 Sep 2016, Revised 16 Feb2017, Accepted 19 Feb 2017 Keywords Citrus; rootstock; propgtion; growth regultors; cutting. H. Benyhi hmidbenyhi2002@yhoo.fr Tel. : Abstrct In order to develop effective methods for rpid clonl multipliction of citrus rootstocks, we propose herein to investigte the effect of exogenously pplied uxins on rooting nd estblishment of spring cuttings of two citrumelo ccessions (Citrus prdisi Mcf. x Poncirus trifolit L. Rf), i.e. Swingle nd Scton. Cuttings were tken from young greenhouse seedlings nd exposed for 24 hours to different tretments including three uxin types (indole-3-cetic cid (IAA), indole-3-butyric cid (IBA) nd nphthlene cetic cid (NAA)) nd two concentrtions of ech one (100 nd 200 ppm). Treted cuttings were then rised long with control cuttings under greenhouse conditions for six weeks. The results tken t the end of this period hve shown significnt vritions in response to uxin tretments regrding survivl, rooting nd sprouting prmeters. These followed similr ptterns for the two ccessions nd were found to be dependnt on the nture of the uxin rther thn its concentrtion, the genotype or the interction of these fctors. Among the different combintions tested, IBA t 100 ppm level gve the highest survivl (100%), the highest rooting percentge (100%) nd ws selected s the most suitble for promoting sprouting of citrumelo cuttings. By contrst, the ppliction of NAA gve poor results nd exerted prticulr inhibitory effect on initition of Swingle citrumelo roots nd sprouts. 1. Introduction Cutting is often the preferred method of propgtion in horticulture since it is chep, fst, simple nd does not require the specil techniques necessry in grfting, budding or micro-propgtion. For citrus, - s mny fruit crops - this method is necessry not only to mintin desirble chrcteristics of the mother plnt [1], but lso to provide them with desirble quntities of true-to-type rootstocks. This fct my be of gret importnce since rootstocks re known to ffect mny scion chrcteristics such s vigor, yield, fruit qulity nd disese resistnce, conditioning therefore the success of citrus industries [ ]. As opposed to seedling, the products of cutting re uniform plnts tht do not exhibit the differences resulting from genetic vritions [7]. Furthermore, it is reported tht cutting hs the tendency to decrese the juvenile stge of plnts nd reduce the time of nursery development [8]. The study of Verm et l. [9], for exmple, hve shown higher sprout length in cuttings of kgzi lime s compred to budded plnts nd nucellr seedlings. A review of literture revels tht numerous studies were crried out to estimte the success of cutting in citrus species nd hve divided them into two groups: (i) Esy-to-root species such s lemons (Citrus limon), cid Fdli et l., J. Mter. Environ. Sci., 2017, 8 (11), pp

2 limes (Citrus urntifoli) nd citrons (Citrus medic), nd (ii) difficult-to-root species including sweet ornge (Citrus sinensis), mndrins (Citrus reticulte) nd trifolite ornge (Poncirus trifolit) [10]. This rnking ws confirmed lter by Vills Bos et l. [11] who studied the rooting of stem cuttings of 16 citrus cultivrs nd reported low rooting rtes in sweet ornge nd tngerines nd higher rtes in lemon Sicilino nd the citrons Etrog nd Dimnte. However, little is still known bout the rooting bility of hybrid cultivrs. Swingle Citrumelo, for exmple, which resulted from cross between grpefruit (Citrus prdisi) nd trifolite ornge in 1907 did not become widespred until the mid to lte 1980s when it strted to be used in Florid orchrds [12]. This dely ws minly due to shortge in supply of seeds nd nursery trees becuse of qurntines relted to citrus bcteril spot disese. It ppered lso tht some seeds of other citrumelos were used for propgtion nd mistkenly identified s Swingle during tht period which my explin the unexpectedly poor performnce of groves plnted on Swingle [12]. Nowdys, lthough citrumelo proved to be tolernt to slinity [13-14], lklinity [15], CTV (Citrus Tristez Virus), nemtodes nd Phytophthor diseses [16], its behvior is known to be ccession-dependnt [17]. Moreover, its use is still limited to some countries such s Brzil nd the U.S. It is therefore importnt to estblish sound nd ccurte methods for mss production of this rootstock before considering clonl selection or undertking breeding progrm. Besides genotypic influence, the rooting potentil of plnt cuttings depends on growth regultors, especilly uxins. Indeed, it hs been estblished tht endogenous nd exogenous levels of uxins plys key role in promoting root initition in plnts [18]. The indole-3-cetic cid (IAA) ws the first plnt hormone to be pplied exogenously for rooting purposes [19]. In the sme yer, Zimmermn nd Wilcoxon [20] discovered tht severl new synthetic uxins, mong them indole-3-butyric cid (IBA) lso promoted rooting. Lter, vrious workers demonstrted tht IBA is very effective in promoting rooting of mny difficult-to-root species [ ]. In citrus, Bhusl et l. [24] reported tht cuttings of different species treted with IBA displyed higher number nd length of roots reltive to non-treted. Similrly, Rossl nd Kersten [25] showed tht quick dipping of Vlenci sweet ornge cuttings before plnting incresed significntly the percentge of formed cllus in the bsl prt of the roots. However, mny inconsistencies cn be noticed mong these experiments due to differences in concentrtions, formuls, dditives or tretment durtions. As consequence, one cn still find vrieties nd cultivrs - in lmost every species - tht do not root even fter tretment with uxins. Thus, the objective of the present investigtion ws to evlute the performnce of spring cuttings of two citrumelo ccessions, i.e. Swingle nd Scton, in response to different uxin tretments in order to determinte n efficient method to produce uniform nd true-to-type cuttings of this rootstock with predictble chrcteristics. 2. Experimentl detils 2.1. Plnt mteril nd growth conditions In order to investigte the effect of uxin tretments on rooting nd estblishment of citrumelo cuttings (Citrus prdisi Mcf. x Poncirus trifolit L. Rf.), greenhouse experiment ws conducted t the Regionl Center for Agriculturl Reserch in Kenitr (Morocco) in lte April The choice of this period ws bsed on the fct tht cuttings collected in lte spring root most redily [26]. Uniform stem cuttings, with n verge length of 12 cm nd contining 3 to 4 buds were collected in the greenhouse from two-yer-old helthy seedlings belonging to two citrumelo ccessions: Swingle citrumelo nd Scton citrumelo. The use of smll, spindly nd ngulr wood ws voided when possible, s this type of wood wsreported to be difficult to root [27] Appliction of uxin tretments After tking ll the leves off, the bsl ends of the cuttings were soked for 24 hours in wter solutions contining Indole-3-cetic cid (IAA), Indole-3-butyric cid (IBA) nd Nphtlene cetic cid (NAA) t 100 nd 200 ppm concentrtions. Control lots were treted with tp wter for the sme period. All cuttings were then plnted in 60x40x12 cm sized plstic trys filled with sterilized pet moss nd kept in mini tunnels to mintin suitble environment for vegettive growth: C temperture nd 40-75% reltive humidity. The irrigtion ws pplied immeditely fter plnting nd every two dys therefter with tp wter. The experiment ws lid out in Split-Split-Plot Design with three replictions nd 10 cuttings per repliction. Min plots consisted of two genotypes (Swingle nd Scton), subplots consisted of the 3 tretments (IAA, IBA nd NAA) nd sub-subplots consisted of three uxin concentrtions (0, 100 nd 200 ppm). Fdli et l., JMES, 2017, 8 (11), pp

3 Survivl rte (%) 2.3. Dt collection nd Anlysis After six weeks of growth, the cuttings were uprooted nd relevnt dt were recorded. These included the number of roots per cutting, the number of sprouts per cutting, root length, sprout length nd sprout dimeter. Survivl rte nd rooting/sprouting percentges were lso clculted bsed on the observtions. Collected dt were nlyzed sttisticlly using SAS pckge (SAS 9.0). Dt were first subjected to nlysis of vrince t p=0.05 nd squre root techniques were pplied wherever necessry (percentges). The mens were compred using Duncn s multiple rnge test t 5% significnce level. 3. Results 3.1. Effect of uxin tretments on cutting survivl ANOVA results indicted tht survivl rte ws significntly ffected by uxin type nd concentrtion fter six weeks of growth (P<0.05). However, no significnt differences were found between tretments for Swingle citrumelo cuttings, lthough their survivl rtes rnged between 87% nd 100% (Figure 1). By contrst, the response of Scton citrumelo cuttings vried mong uxin tretments with mximum rtes t 100 ppm IAA nd 100 ppm IBA (100%) nd minimum rtes t 200 ppm IAA (84%) b b b b b Swingle Scton Control 100 ppm IAA 200 ppm IAA 100 ppm IBA 200 ppm IBA 100 ppm NAA 200 ppm NAA Figure 1: Effect of Auxin tretments on survivl of citrumelo cuttings 3.2. Effect of uxin tretments on rooting Six weeks fter plnting, cuttings of both citrumelo ccessions formed roots to vrying degrees. In instnces where roots were not induced, some clluses were often developed t the bsl end of cutting in response to uxin tretments. The bsl end ws swollen nd the color ws chnged from green to whitish green. The results recorded from the tested cuttings, including the percentge of rooting, the number of roots nd root length re presented in the tble 1. The nlysis of vrince showed highly significnt effect (P<0.001) of uxin type on ll these prmeters. By contrst, the effects of genotype nd uxin concentrtion were only significnt on the number of roots. In both ccessions, mximum rooting percentge of 100% ws chieved under the 100 ppm IBA tretment. However, cuttings treted with IAA t 200 ppm exhibited higher number of primry roots reching verges of nd roots per cutting, respectively for Swingle nd Scton ccessions. The lowest vlues were recorded under control nd NAA tretments with rooting percentge of 68 to 81% nd n verge number of roots rnging from 2.89 to 4.39 roots per cutting. Dt recorded in respect to root length showed lso vrition depending on the tretment pplied. In Scton citrumelo cuttings, the longest roots were obtined under 100 ppm IAA (7.01 cm), 100 ppm IBA (6.78 cm) nd 200 ppm IBA (6.47 cm), wheres it ws the tretment 200 ppm IAA which stimulted the best elongtion of Swingle citrumelo roots with n verge length of 8.07 cm. The shortest roots were recorded under NAA nd control tretments regrdless of the ccession studied. The corresponding vlues for these tretments vried from 4.12 nd 4.79 cm. Fdli et l., JMES, 2017, 8 (11), pp

4 Tble 1: Effect of uxin tretments on rooting of citrumelo cuttings Genotype Tretment Rooting 1 (%) Number of roots 2 Root lenght 2 (cm) C. Swingle Control bc c 4.70 b 100 ppm IAA b 8.72 b 6.65 b 200 ppm IAA b ppm IBA b 7.44 b 200 ppm IBA b 9.03 b 6.75 b 100 ppm NAA bc 2.33 c 4.79 b 200 ppm NAA c 2.62 c 4.48 b C. Scton Control b 3.12 d 4.12 b 100 ppm IAA b bc ppm IAA b b 100 ppm IBA c ppm IBA b b ppm NAA b 3.78 d 4.72 b 200 ppm NAA b 4.39 d 4.65 b Anlysis of vrince 4 Genotype (G) NS ** NS Type of uxin (T) *** *** *** Concentrtion (C) NS *** NS G x T NS NS NS G x C NS NS NS T x C NS *** NS G x T x C NS NS NS 1 Men of 3 replictions. 2 Men of 30 replictions. 3 For ech ccession, superscripts within columns indicte men seprtion between tretments by Duncn's multiple rnge test t 0.05 level. 4 Significnt effects re indicted by sterisks: * = P < 0.05, ** = P < 0.01 et *** = P < 0.001, while non significnt effects re indicted by NS Effect of uxin tretments on sprouting ANOVA results reveled significnt effect of uxin type on ll sprouting prmeters studied t 0.01 level. The effects of genotype, uxin concentrtion nd the interction Auxin type x Auxin concentrtion were lso significnt with respect to the number of sprouts, sprout length nd the number of leves per cutting. Among tretments, IBA t 100 ppm resulted in mximum number of sprouted cuttings fter 45 dys of growth with percentges of nd 100% respectively for Swingle nd Scton cultivrs, wheres control nd NAA tretments resulted in the lowest vlues, rnging from 73 to 78% (Tble 2). Similrly, dt pertining to the number of sprouts per cutting, the length of sprouts nd the number of leves per sprout were the highest under the concentrtion 100 ppm of IBA with mens of 2.06, 7.94 cm, respectively in Swingle nd 2.16, 7.34 cm, in Scton (Tble 2). This tretment ws closely followed by the 100 ppm IAA tretment nd contrsted to control nd NAA tretments which resulted in minimum vlues. We should prticulrly note tht NAA tretments resulted in lower vlues thn control in the cse of Swingle citrumelo cuttings suggesting specific toxic effect. In contrst to the forementioned prmeters, the results showed no or few differences mong tretments in regrds to sprout dimeter. Fdli et l., JMES, 2017, 8 (11), pp

5 Tble 2: Effect of uxin tretments on sprouting of citrumelo cuttings Genotype Tretment Sprouting 1 (%) Number of sprouts 2 Sprout lenght 2 (cm) Sprout dimeter 2 Number of leves 2 (mm) C. Swingle Control b bc 3.65 cd c 100 ppm IAA b 1.83 b 5.57 b b 200 ppm IAA b 1.52 bc 4.61 bcd bc 100 ppm IBA ppm IBA b 1.21 c 5.24 bc c 100 ppm NAA b 1.43 bc 2.81 d c 200 ppm NAA b 1.45 bc 3.05 d c C. Scton Control c 1.54 cd 3.25 c 0.98 b 6.37 c 100 ppm IAA b 2.04 b 6.89 b b 200 ppm IAA b 1.39 d 4.91 bc 1.09 b 8.61 bc 100 ppm IBA ppm IBA b 1.88 bc 6.86 b b 100 ppm NAA bc 1.50 cd 3.88 c 1.15 b 7.34 c 200 ppm NAA bc 1.68 bcd 3.89 c 1.14 b 7.97 c Anlysis of vrince 4 Genotype (G) NS * * NS ** Type of uxin (T) ** ** *** ** *** Concentrtion (C) NS *** ** NS *** G x T NS NS NS NS NS G x C NS NS NS NS NS T x C NS ** NS * ** G x T x C NS NS NS NS * 1 Men of 3 replictions. 2 Men of 30 replictions. 3 For ech ccession, superscripts within columns indicte men seprtion between tretments by Duncn's multiple rnge test t Significnt effects re indicted by sterisks: * = P < 0.05, ** = P < 0.01 et *** = P < 0.001, while non significnt effects re indicted by NS. 4. Discussion Most of tested cuttings survived fter six weeks of growth even in the non-treted control. This my be due to the suitble substrte nd/or environmentl conditions (temperture, humidity). Indeed, the effect of growth medium nd sesonl vritions on cutting estblishment is well documented nd pet medium ws prticulrly recommended for growth of citrus cutting [28-29]. Furthermore, both ccessions rooted successfully under wter tretment, which implies tht these cn be rnked s esy-to-root cultivrs. According to Hbermnn et l. [30] nd Alm et l. [31], rooting nd survivl of cuttings re correlted since the initition of roots cn promote rehydrtion of tissue nd prevent their deth from wter stress. All rooting prmeters tended to increse to similr extent in the two ccessions in response of exogenous uxin ppliction. We showed prticulrly in this study high effectiveness of the combintions 100 ppm x IBA nd 200 ppm x IAA. These findings re in line with previous works tht hve reported higher ctivity of IBA nd IAA in stimulting rooting of citrus species [24-32]. Some uthors lso stted tht uxin concentrtions my ffect rooting response of cuttings [32-29]. In experiments where 100, 200 nd 400 ppm concentrtions of IAA Fdli et l., JMES, 2017, 8 (11), pp

6 nd IBA were tested on cuttings of Eurek lemon, Homosss sweet ornge, sweet ornge seedlings, sour ornge, Clmindin, Cleoptr mndrin nd Thornton tngelo [27], it ws found (i) tht 100 ppm concentrtion ws no more effective thn tp wter (ii) 200 ppm concentrtion gretly incresed the number of roots induced on ll vrieties s compred with tht obtined with tp wter nd (iii) 400 ppm concentrtion cused some injury to the bse of cuttings of some vrieties nd cused no pprent injury to cuttings of other vrieties. Figure 2: Appernce of Swingle citrumelo cuttings fter six weeks of growth. A: Control ; B: 100 ppm IAA ; C: 200 ppm IAA ; D: 100 ppm IBA ; E: 200 ppm IBA ; F: 100 ppm NAA ; : 200 ppm NAA. In contrst to IAA nd IBA, cuttings treted with NAA hd more difficulty to root. This might be due to differences in bsorption nd trnsport of these hormones. Indeed, lthough NAA proved to stimulte rooting of mny plnt species [ ], it is thought tht low levels of this hormone cn be bsorbed by treted tissues [36]. On the other hnd, differences in uptke between IAA nd IBA re unlikely ccording to Ludwig-Müller et l. [37] who followed the uptke of these two uxins for 24h nd 48h nd reported no significnt differences during these time periods. The literture reports lso severl cellulr nd biochemicl explntions which my illustrte the differences observed in our study mong uxins. Some uthors ttributed the effectiveness of IAA nd IBA to their higher stbility [1-38]. Others [39-40] studied the metbolism of IAA nd IBA by mung cuttings nd ttributed the better ctivity of IBA to the rpid formtion of IBAsp, n IBA conjugte, which promoted rooting better thn IAA.. In the sme line, Pythoud nd Buchl [41] could not find ny oxidtion products of IBA in cuttings of Fdli et l., JMES, 2017, 8 (11), pp

7 Populus tremul nd ssumed tht this fct might be the cuse of the better ctivity of IBA in rooting. It ws lso extensively reported tht IAA nd IBA hve the prticulrity to increse the endogenous levels of free IAA nd free IBA, which seem to ply key role in root initition ( ). In citrus, experiments on Rngpur lime hve shown n increse in endogenous level of IAA in the brk of cuttings which ws more thn three times higher t the dy 19 fter exogenous tretment thn t dy 0 nd correlted well with ese of rooting [47]. Similrly, Epstein et l. [48] observed concomitnt increse in free IBA with the rooting of esy- nd difficult-to-root cultivrs of sweet cherry (Prunus vium L.). They postulted tht esy-to-root, s opposed to difficult-to-root cultivrs, hve the bility to hydrolyze the ester conjugte t the pproprite time to relese free IBA, which my promote root initition. This my be plusible explntion in our cse if we consider tht citrumelos re esy-to-root cultivrs. Figure 3: Appernce of Scton citrumelo cuttings fter six weeks of growth. A: Control ; B: 100 ppm IAA ; C: 200 ppm IAA ; D: 100 ppm IBA ; E: 200 ppm IBA ; F: 100 ppm NAA ; : 200 ppm NAA. Besides rooting, there were considerble vritions in sprouting of citrumelo cuttings depending on the nture nd the concentrtion of uxins pplied. In generl, NAA tretments did not differ from control, while IBA nd IAA gve stisfctory results, prticulrly t 100 ppm level. Previous workers hve lso noticed n incresing effect of IBA on bud sprouting of sweet lime [49] nd Stsum ornge [50] compred to untreted cuttings. Bhtt nd Tomr [22] obtined mximum sprouting percentge (68.50%), mximum sprout length (6.59 cm) nd mximum sprout dimeter (0.32 cm) under the concentrtion 500 ppm fter dipping Citrus urntifoli Fdli et l., JMES, 2017, 8 (11), pp

8 Swingle cuttings in concentrted solutions of IBA. According to the literture, the increse in sprouting my be due to more number of leves nd vigorous system, which my enhnce the bsorption of minerl nd wter nd promote the production nd ssimiltion of crbohydrtes tht re necessry for growth [31]. It is lso thought tht esy-to-root species cn bsorb more uxin nd trnsport more of it to the eril prts [18] where it cn increse liner growth of cutting tissues through stimultion of cell elongtion [51]. In this cse, the differences in effectiveness mong uxins my hve resulted from differences in trnsport velocity s suggested by Epstein nd Sgee [52] who found tht IBA ws trnsported in midribs of Citrus leves t somewht lower (pproximtely 60%) rte thn IAA. In conclusion, the results of the present investigtion clerly indicte tht cuttings of citrumelo root esily nd develop within short time fter tretment with low uxin concentrtions for 24 h before plnting. We prticulrly recommend the use of IBA nd IAA t the 100 ppm concentrtion for this purpose. We should lso note tht this technique gve better results thn those reported by erly workers on citrumelo [53] nd vrious other citrus species using the quick dip method [ ]. Thus, it should be generlized in order to be used in lrge scle for mss production of improved mteril nd effective estblishment of citrus nursery stocks. Acknowledgements-The uthors grtefully cknowledge the hed of the Regionl Center for Agriculturl Reserch in Kenitr, Dr. Hssn Benoud, for his encourgement nd support throughout the study periods. References 1. Hrtmnn H.T., Kester D.E. Plnt propgtion. Principles nd prctices, Prentice-Hll Inc., (1983). 2. Espinoz-Nunez E., Mourão Filho F. A. A., Stuchi E. S., Cnturis-Avilés T., dos Sntos Dis C. T. Sci. Hortic. 129 (2011) Grcıá-Sánchez F., Jifon J. L., Crvjl M., Syvertsen J. P. Plnt Sci. 162 (2002) Zmor O. P., Urruti V. M., Rodríguez S. B. Agrocienci 36 (2002) Zekri M. Proc. Fl. Stte Hort. Soc. 109 (1996) Zekri M., Al-Jleel A. J. Amer. Soc. Hort Sci. 121 (2002) Bose T.K., Mitr S. K., Sdhu M. K. Propgtion of tropicl nd subtropicl horticulturl crops. Ny Proksh, (1986). 8. Spiegel-Roy P., Goldschmidt E. E. The Biology of Citrus. Cmbridge University Press, (1996). 9. Verm S. K., Singh H., Bhrdwj P. N., Ary R. R. Progress. Hortic. 37 (2005) Pltt R. G., Opitz K. W. Propgtion of citrus. In: The Citrus Industry. Berkeley, USA, Univ. Clif. Press (1973). 11. Vills Bos R. M. F., Sntos R. F. A., Slibe A. A. In Congresso Brsileiro de Fruticultur. Cmpins- SP (1987) Cstle B., Stover E. Updte on use of Swingle citrumelo rootstock. Univ. Fl. Ext. Fct Sheet HS-801 (2001). 13. Ait El Aoud B., Fdli A., Aderdour T., Tlh A., Benkirne R., Benyhi H. Biolife 3 (2015) Fdli A., El Aymni I., Chetto O., Boudoudou D., Tlh A., Benkirne R., Benyhi H. Int. J. Recent Sci. Res. 6 (2015) Benyhi H., Beniken L., Omri F. E., Benzzouze A., Hndji N., Mstef Y., Olitrult P. Afr. J. Agric. Res. 6 (2011) Cstle W. S., Wutscher H. K., Youtsey C. O., Pelosi R. R. Proc. Fl. Stte Hort. Soc. 101 (1988) Fdli A., Chetto O., Tlh A., Benkirne R., Morillon R., Benyhi H. J. Life Sci. 8 (2014) Epstein E., Ludwig-Müller J. Physiol. Plnt. 88 (1993) Cooper W. C. Plnt Physiol. 10 (1935) Zimmermn P. W., Wilcoxon F. Contrib. Boyce Thompson Inst. 7 (1935) Alikhni L., Ansri K., Jmnezhd M., Tbtbie Z. Irn. J. Plnt Physiol. 1 (2011) Bhtt B. B., Tomr Y. K. J. Hill Agric. 2 (2011) Owis S. J. Pk. J. Biol. Sci. 13 (2010) Bhusl R. C., Mizutni F., Moon D. G., Rutto K. L. Pk. J. Biol. Sci. 4 (2001) Rossl P. A. L., Kersten E. Sci. Agric. 54 (1997) Ozcn M. Tr. J. Agric For. 14 (1990) Cooper W. C. Proc. Fl. Stte Hort. Soc. 53 (1940) 174. Fdli et l., JMES, 2017, 8 (11), pp

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