Field Evaluation of Citrus Cultivars for Greening Disease Resistance in Thailand

M. Koizumi, M. Prommintara, G. Linwattana, and T. Kaisuwan

ABSTRACT. Forty-three cultivars of citrus planted in the field at Nan Horticultural Research Station were evaluated for their resistance to greening disease. All the trees were propagated at the Station using Cleopatra mandarin rootstock, and were established in the field in 1986. Most of the scions were free from greening disease when propagated. Field observations in July 1992 revealed that a large number of trees had declined, showing typical symptoms. The cultivars Wilking, Ellendale, Pet-yala, Beauty, Onesco, Pong-chieng-ga (Tankan), Nian-ju, and sweet orange were most susceptible. Many mandarins including Fairchild, Murcott, Kinnow, Clementine, Fremont, Ponkan, King, and Som-keo-wan were next. Queen mandarin, Avon Ever Bearing (calamondin) and rough lemon grew well with mild symptoms. Ladu mandarin and Som-pan mandarin showed the most resistance, with good growth, few symptoms and yielding healthy fruit. Citrus greening is the most serious Horticultural Research Station. All disease in the southern part of Asia, the trees were propagated at the Sta- including Thailand. Many attempts tion by grafting on Cleopatra mandarin have been made to control the disease, rootstock. Alarge number of scions had namely production of disease-free bud- been introduced from foreign countries wood by heat therapy, antibiotic treat- and others were propagated from ments, shoot-tip grafting, propagation nucellar seedlings originating from in- of healthy plants under restricted con- troduced seeds. The trees were as- ditions, elimination of infected trees sumed to be greening-free when pro- from fields, control of vector by insec- pagated. However, one of the two bud- ticide sprays or release of natural lines of Clementine, Fremont and Som- enemies. However, citrus production keaw and all Som-keowan lines origi- is still limited by the disease. nated from infected mother trees Sweet orange and Bearrs lime were planted at the Station. reported to be somewhat tolerant to Ten trees of each cultivar or variety greening in the Philippines (1,2,3). In were planted in the field with a 6 x 8 India, Nariani (4) analyzed thereaction m spacing in 1986. Local mandarin of different species and cultivars of cit- Som-keowan which were propagated rus to greening under glasshouse con- by layering and showed typical green- ditions. He reported that sweet lime ing symptoms surrounded the field at seedlings and trees of Mosambi orange a distance of 50-100 m. Insecticides on sweet lime, Eureka lemon or Italian were occasionally sprayed to control lemon rootstock developed few the vector. symptoms. However, the use of toler- Observation. All the trees were in- ant rootstock and scion cultivars has spected on July 28-29,1992. Each tree not been practiced. was rated as -, + or + + based on the The objectives of this paper were tree vigor; also "few", + , + + or to analyze the field performance of + + + referred to the intensity of various citrus varieties in Thailand greening disease symptoms such as where greening disease is prevalent. yellow shoots, zinc deficiency, leaf Attempts were made to evaluate their mottling, and blotching. In addition, resistance to the disease. die-back, dwarfing, small leaves and feeding of psylla were noted. Other MATERIALS AND METHODS serious diseases were also noted. Data on the horticultural characteristics of Citrus varieties and cultivars. the trees such as sizes of canopy, yields, Forty-three citrus cultivars or bud- fruit quality and number of seeds in lines were planted in the field at Nan fruit were collected at the Station. Citms Greening Disease

Detection of pathogen. Scion ma- budline trees showed diversity in the terial of Hiroshima sweet orange and symptoms. Som-keowan from the field were Satsuma mandarin and Necked grafted onto virus-free seedlings of orange (Natsudaidai) showed few or rough lemon in a glasshouse. Small mild symptoms, but the trees lost their samples from young stems of these vigor and the attached leaves were varieties were fixed and embedded in small and scattered. Among satsuma resin for electron microscopy. U1- cultivars the early maturing ones such trathin sections were cut, stained and as Miyakawa and Sugiyama were more observed with a Hitachi H-300 electron susceptible than the late maturing ones microscope. such as Hayashi, Ishikawa, Owari, and Silver Hill. RESULTS Although Avon Ever Bearing (cala- mondin) and rough lemon trees grew By December 1989, local mandarin well, their foliage was yellow with mild Som-keowan trees had already de- symptoms such as mottling. Most of clined, showing yellowing foliar the Sripri, Queen mandarin and Som- symptoms. Some sweet orange trees keaw trees were vigorous with few or developed yellow shoots with typical mild symptoms, but one or two trees leaf symptoms such as mottling and declined with typical symptoms. blotching. However, most of the other All the trees of Ladu mandarin and trees were still healthy and were grow- Som-pan grew very well with few or ing well. very mild symptoms. Their foliage was The second observation in July 1992 still dark-green. All the trees set nor- revealed that most of the trees of the mal fruit without any abortive seeds. following cultivars had declined and However, the canopy of Ladu manda- had developed severe symptoms: Wilk- rin was flat. ing mandarin (MD), Ellendale MD, Ju- In another field, 23 cultivarsinclud- long-ka MD, Som-keowan MD, Pet- ing sweet orange and grapefruit were yala MD, Beauty MD, Onesco, Pon- planted. Most ofthe sweet orange trees chieng-ga (Tankan), unknown culti- were infected with greening disease vars of sweet orange and Nian-ju and declined. These trees developed (Table 1). Half or more of the trees of yellow shoots with typical symptoms. each cultivar were severely infected. In contrast, all the grapefruit trees The trees of the cultivars Wilking, El- were still vigorous and grew well. lendale, Ju-long-ga, Beauty, Tankan, However, some trees developed one or and Nian-ju showed severe dieback two yellow shoots on the outer canopy. while Pet-yala, Beauty, Onesco, and The psylla vector of greening dis- sweet orange seedlings were dwarfed. ease, Diaphorina citri was frequently The trees of the following cultivars observed in the fields. Foot rot oc- lost their vigor and developed typical curred on a few trees. Pink disease symptoms on the whole canopy, or par- caused by Corticiurn salrnonicolorwas tially in about half of the tree: Or- observed, but it did not cause serious tanique, Fairchild, Murcott, Kinnow, damages to the trees. Tristeza virus Bower, Clementine, Fremont, Pon- and brown aphids also occurred in the kan, King MD, Valencia orange, fields. However, severe stem-pitting Hiroshia sweet orange, Hybrid Fair- was not observed on the twigs of either child, Hybrid Som-keowan, and Ba- sweet orange or grapefruit, suggesting yue-ju. Leaf sizes of these trees were that severe strains of tristeza were not reduced. Fremont mandarin origina- common at the Station. ted from two different scion stocks, one An electron microscopy showed from Chieng Mai and the other from a many prokaryotic organisms in the de- tree planted at the Station. The trees generated cytoplasm of sieve paren- originated from the Station were se- chyma cells of Hiroshima and Som- verely infected, while the Chieng Mai keowan (Fig. 1). TABLE 1 PERFORMANCE OF CITRUS CULTIVARS GROWS IN A FIELD WHERE GREENING DISEASE WAS PREVALENT IN INLAND No. of trees in each pup

Total Tree vigd Internit). of symptomsx no. of trees + + + - Few + c + + + c Remarkk

Iiinnov; ID Bou-er MD Ellendale XD Ladu 5ID Cl~mpntinp Fremont ?rIDU Fremant lmv Ju-long-ka Pnnkan %&an mgXI3 Som-keoaan MD' Pet-yala MD Beauty MD Satsuma MD' Satsuma ND" Earlr satsgma MD srip;lU Queen MD one-w Keek orange (Natsudaidai) Valersda SW Himhima SW Pwhieng-ga (Tankan) Sweet oranger Avon Ever Heraing (Calamondin) 10 0 10 0 0 I0 0 0 Som-keaw 16 0 15 1 0 14 2 0 I% Hybrid Fairchild 10 0 6 5 0 8 2 0 DB,SL cs Hybr. Som-keowan 10 0 8 2 0 9 1 0 2 u h'ian-ju 9 0 1 8 0 0 9 0 DR 7.. $a-yue-ju 10 0 8 2 0 8 2 0 SL 9- Rough lemonq 10 0 10 0 0 10 0 0 ; "MD:mandarin, SW: Sweet orange. All the trees were propagated by grafting on Cleopatra mandarin rootstock and planted in the field in 1986. 9 ' + -I- = vigorous, + = intermediate, - = decline. "Based on legfsymptoms such as yellow shoots, zinc deficiency, mottling and blotch in^. "DB = dieback, SL = small leavea, DW = dwarf. "Propagated from a tree planted in Nan Station. "Propagated from xejons introduced from Chang Ma;. 'Late maturing varieties inclutling Ishikawa, Hayashi, Silverhill and Owari. 'Mid-maturing Sugigama. Wucellar seedlings of unhown eultivar. %elf rooted. Twelfth ZOCV Conference

intensity of tree decline or symptoms is a more suitable index for the evalu- ation of disease resistance. Satsuma mandarin trees uniformly declined in spite of showing few or mild mottle or zinc-deficiency. However, the attached leaves were small and scattered, presumably due to greening disease. The citrus cultivars were classified into four groups according to the dis- ease resistance. The most susceptible cultivars were Ortanique, Wilking, El- lendale, Pet-yala, Beauty, Onesco, Pon-chieng-ga (Tankan), Nian-ju, and sweet orange. The second group in- cluded the cultivars Fairchild, Mur- cott, Kinnow, Clementine, Fremont, Ponkan, King, Som-keowan, satsuma mandarin, Som-keaw and Ba-yue-ju. The third group of the following cul- tivars was more resistant than the sec- Fig. 1. Greening organisms (arrows) in the degenerated sieve tube of Som-keowan young ond group: Queen, Avon Ever Bearing shoot. Som-keowanwas collected at Nan Hor- (calamondin) and rough lemon. Grape- ticultural Research Station. The bar repre- fruit also belonged to the third group. sents 500 nm. Ladu mandarin and Som-pan mandarin that were the most resistant belonged to the fourth group. DISCUSSION Field observations in the Philip- pines showed that Szinkom-Ladu man- On the basis of our observations, darin hybrid, calamondin, lime and greening disease and its vector were lemon trees grew well in areas where prevalent in the field, andgreeningdis- the greening disease was prevalent (1). ease was the major cause of decline. However, subsequently it was re- The tree decline was correlated mainly ported that many mandarin trees in- with the intensity of the foliar greening cluding Ladu selection, Ladu hybrid symptoms. Most of the citrus cultivars selections, Szinkom hybrid selection, grew uniformly and developed similar Szinkom and Ponkan were removed symptoms in the same budline. How- from the field due to the severity of the ever, some cultivars or budlines mar- disorder (3). We observed that the kedly differed in tree vigor and symp- canopy of Ladu mandarin was flat and toms, possibly due to different infection the branches hanged. These would re- times. For example, local mandarin sult into over fruit-bearing and decline Som-keowan or Fremont trees propa- of trees in future. However, the differ- gated from budlines infected with the ence in Ladu mandarin between Philip- greening disease declined uniformly pines and Thailand should be further while in the others that were propa- studied, considering pathogenic strain gated from disease-free scions the or other destructive causal factors. We symptoms varied; some of the trees are carrying out to detect resistance to were healthy while others showed se- greening disease by analysis of the host vere symptoms. This observation reaction after artificial inoculation suggests that alower percentage of dis- under controlled conditions and popu- eased tree is not correlated with the lation trends of the pathogen in host degree of resistance. In contrast the tissues. Citms Greening Disease 279

ACKNOWLEDGMENT Fisheries, Japan and the Department The study was carried out under a of Agriculture, Thailand. We are cooperation project between the Trop- grateful to the National Research ical Agriculture Research Center, Council, Thailand for authorizing the Ministry of Agriculture, Forestry and cooperation. LITERATURE CITED 1. Altamirano, D. M., C. I. Gonzales, and R. C. Vinas 1976. Analysis of the devastation of leaf-mottling (greening) disease of citrus and its control program in the Philippines, p. 22-26. In: Proc. 7th Conf. IOCV. IOCV, Riverside. - 2. Gonzales, C. I., R. C. Vinas, and L. A. Vergara 1972. Observations on 110 citrus cultivars planted in anareaseverely infested by leafmottling, p. 3840. In: Proc. 5th Conf. IOCV. Univ. Florida, Gainseville. 3. Gonzales, C. I. and R. C. Vinas 1981. Field performance of citrus varieties and cultivars grown under control measures adopted against leaf mottling (greening) disease in the Philippines. Proc. Int. Soc. Citricult. 1: 463-464. 4. Nariani, T. K. 1981. Integrated approach to control citrus greeningdisease in India. Proc. Int. Soc. Citricult. 1: 471-472.