RESISTANCE OF RUBBER CLONES RECOMMENDED IN INDONESIA TO CORYNESPORA AND COLLETOTRICHUM LEAF FALL DISEASES

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1 RESISTANCE OF RUBBER CLONES RECOMMENDED IN INDONESIA TO CORYNESPORA AND COLLETOTRICHUM LEAF FALL DISEASES Alchemi Putri Juliantika Kusdiana, Afdholiatus Syafaah, and Tri Rappani Febbiyanti Sembawa Research Centre, Indonesian Rubber Research Institute Abstract Leaf fall disease caused by Corynespora cassiicola and Colletotrichum sp. can causes reduction of latex production in rubber plantations. Hence screening for disease development in each rubber clone needs to be carried out. Resistance level of 25 rubber clones recommended in Indonesia to leaf fall diseases were examined on rubber plants grown in polybags placed in a greenhouse using three isolates each of C. cassiicola and Colletotrichum sp. isolated from the rubber clones RRIM 600, GT 1, and PR 303. The results showed that 9 rubber clones i.e AVROS 2037, BPM 1, BPM 109, IRR 42, IRR 107, IRR 220, PB 260, PB 330, and RRIC 100 had a high resistance to both the leaf fall diseases while 4 clones i.e BPM 24, PR 261, PR 300, and PR 303 had a low resistance. The other clones had a high resistance against only one of the diseases. Keywords: Colletotrichum sp., Corynespora cassiicola, leaf fall disease, rubber clones. INTRODUCTION Rubber (Hevea brasiliensis Muell. Arg) is one of the important plantation crops in several Asian and African countries. attack can be a limiting factor in rubber production. Corynespora leaf fall caused by Corynespora cassiicola (Berk. & M.A. Curtis) C.T. Wei and Colletotrichum leaf fall caused by Colletotrichum sp. are important leaf diseases. Leaf diseases attack plants of all age levels, both in nurseries or in gardens (Suryaningtyas, 2012; Jayashinge, 2000). Leaf fall disease intensity is affected by the condition of plants, weather, and the clones (Pawirosoemardjo, 2004). Corynespora leaf fall disease occurs during the dry months after the refoliation (Manju et al, 2001) and Colletotrichum leaf fall is more prominent during the rainy and humid months (Manju et al., 2014). Epidemics and recurrent attack of this disease can cause economic losses at all ages of the plant. C. cassiicola attack young copper brown and late copper brown leaf stages (Fernando et al., 2010a). The early symptoms is black patches on the leaf and extend into the smaller leaf veins and the spots will appear 811

2 Proceedings of International Rubber Conference 2017 like fish bones (Jinji et al., 2007). Later, the spots become more widespread, round or irregular with dried center and may fall off leaving a hole. This symptom is due the toxins accumulation at the site of infection. Furthermore, the lesion vicinity becomes chlorotic due to the breakdown of chloroplasts (Jayasinghe & Fernando, 2011). The affected leaves become yellow or brown gradually and then fall. In addition to attacking the leaves, pathogens also attack the petiole, shoots, stems, or branches (Suryaningtyas, 2012). Colletotrichum sp. in rubber can attack the young or old leaves, but usually the young rubber leaf is the main target showing higher disease intensity (Febbiyanti & Kusdiana, 2013). Young leaves when attacked limp and turn black, wrinkled, and finally fall. Infected mature leaves have raised brown or black spot, wrinkle, and rarely fall down (Situmorang & Budiman, 2003). Although cultivation of resistant clones is suggested for disease management, resistance can be broken by the emergence of new more virulent races of pathogen. Monitoring of disease in each clone needs to be done systematically (Situmorang et al., 2004). This study is an attempt to screen the rubber clones recommended in Indonesia, for disease resistance. MATERIALS AND METHODS The study was carried out in the Greenhouse of Sembawa Research Centre, South Sumatra from December 2015 to September 2016, on rubber plants grown in polybags using three replications. Twenty five rubber clones recommended in Indonesia used in this research were AVROS 2037, BPM 1, BPM 24, BPM 107, BPM 109, GT 1, IRR 5, IRR 32, IRR 39, IRR 42, IRR 104, IRR 107, IRR 112, IRR 118, IRR 220, PB 260, PB 330, PB 340, PR 255, PR 261, PR 300, PR 303, RRIC 100, RRIM 600, and RRIM 712. The pathogen isolates consisted of three isolates of C. cassiicola (CC-RRIM 600, CC-GT 1, and CC-PR 303) and three isolates of Colletotrichum sp. (CS-RRIM 600, CS-GT 1, and CS-PR 303) from RRIM 600, GT 1, and PR 303 rubber clones. Those isolates were high of virulence levels (Oktavia et al. 2016b). 812

3 Resistance of rubber clones recommended in Indonesia to corynespora and Preparation of Pathogen Isolate Leaves infected by C. cassiicola and Colletotrichum sp. were collected from three rubber clones RRIM 600, GT 1, and PR 303 from the field. Pathogens were isolated on potato dextrose agar (PDA) media purified and identified by microscopic. Production of C. cassiicola Inoculum C. cassiicola pathogen inoculum was prepared by Situmorang (2002) method. A total of 8-10 discs (Ø 5 mm) of isolate cultures were placed in reverse position on the abaxial surface of steriled young leaves in petri dishes, so the mycelia were in direct contact to the leaf surface. The leaf position was reversed in 3 to 4 days after incubation. The conidia were produced on the adaxial surface after 4-5 days. The leaves were dried for one day at room temperature and the conidia formed were removed using a brush, mixed with sterile water and filtered using gauze filter to separate the clumped conidia. Conidia concentrations were calculated and adjusted using a haemocytometer. Production of Colletotrichum sp. Inoculum Colletotrichum sp. inoculum was obtained from two weeks old cultures. The conidia were removed from the potato dextrose agar media using a brush, mixed with sterile water and filtered using gauze filter to separate the clumped conidia. Conidia concentrations were calculated and adjusted using a haemocytometer. Inoculation of Rubber Plants Inoculation was carried out by spraying one ml of inoculum with a concentration of 4x10 4 conidia/ml using a sprayer on the abaxial surface of the young brown leaves. The leaves were covered with transparent plastic bags to retain moisture during incubation. The plastic bags were removed after four days of incubation. Observation and Data Analysis Observation of disease intensity was carried out 12 days after inoculation. was graded using the scale as shown in Table

4 Table 1. attack scale Proceedings of International Rubber Conference 2017 Scale 0 No symptoms 1 Few blackish brown spots % of the leaf area is brownish yellow % of the leaf area is brownish yellow or leaf fall The disease intensity (%) was calculated using the Towsendt & Hueberger formula, 1943 (Sinaga, 2006): n n. v DI i 1 Z. N x100% where: DI : disease intensity; n : number of plant scale v; v : scale to-i; Z : highest score; N : number of plants observed. Based on the percentage of disease intensity and virulence levels of pathogen isolates the clones were classified as follows (modified from Situmorang, 2002): Table 2. of virulence level Attacks scale 0-5% highly resistant 6-33% resistant 34-67% susceptible % highly susceptible RESULTS AND DISCUSSION The results of screening for resistance against C. cassiicola on the 25 rubber clones recommended in Indonesia showed the presence of different virulence levels of each isolate ranging from highly resistant to highly susceptible levels (Table 3). The results of previous studies suggest that each rubber clone has a different resistance level to C. cassiicola isolates (Situmorang et al., 2000; Hadi et al., 2004; Oktavia et al., 2016a). 814

5 Resistance of rubber clones recommended in Indonesia to corynespora and Table 3. Corynespora leaf fall disease intensity of recommended rubber clones to three of C. cassiicola isolates CC-RRIM 600, CC-GT 1, and CC-PR 303. CC-RRIM 600 CC-GT 1 CC-PR 303 Clone Intensity (%) AVROS susceptible resistant 0.00 highly resistant BPM resistant 2.78 highly resistant 0.00 highly resistant BPM highly susceptible highly susceptible highly susceptible BPM highly resistant 0.00 highly resistant 0.00 highly resistant BPM resistant 0.00 highly resistant 8.33 resistant GT susceptible highly susceptible highly susceptible IRR highly susceptible highly susceptible susceptible IRR highly susceptible highly susceptible susceptible IRR highly susceptible highly susceptible highly susceptible IRR resistant resistant resistant IRR highly susceptible 2.28 highly resistant resistant IRR resistant resistant 0.00 highly resistant IRR highly susceptible highly susceptible resistant IRR highly susceptible highly susceptible susceptible IRR susceptible 5.56 highly resistant susceptible PB highly resistant 5.56 highly resistant 0.00 highly resistant PB resistant 0.00 highly resistant 0.00 highly resistant PB highly susceptible highly susceptible 0.00 highly resistant PR resistant resistant resistant PR highly susceptible highly susceptible highly susceptible PR highly susceptible resistant resistant PR highly susceptible highly susceptible highly susceptible RRIC resistant resistant 0.00 highly resistant RRIM susceptible highly susceptible resistant RRIM highly resistant 5.56 highly resistant resistant 815

6 Proceedings of International Rubber Conference 2017 C. cassiicola spores of CC-RRIM 600 had the greater effect of 54.33% compared to other spores i.e CC-GT 1 of 43.54% and CC-PR 303 of 32.55%. This can be due to higher virulence level. Situmorang et al. (2007) also reported that RRIM 600 rubber clone is an important source of infection in Indonesian rubber plantations. The testing of Colletotrichum sp. on the rubber clones showed only two different virulence levels, resistant and susceptible (Table 4). The effect of spore treatment on the disease intensity showed that the spores of Colletotrichum sp. CS-PR 303 had the greater effect of 32.94% compared to other spores CS-RRIM 600 of 31.37% and CS-GT 1 of %. The testing of resistance of Corynespora against 25 recommended rubber clones showed different resistance responses. Two clones i.e BPM 107 and PB 260 with the disease intensity value range from 0.93% to 1.85% are highly resistant. 11 clones i.e AVROS 2037, BPM 1, BPM 109, IRR 42, IRR 104, IRR 107, IRR 220, PB 330, PR 255, RRIC 100 and RRIM 712 are found resistant with disease intensity values ranging from 6.48% to 30.56%. Seven clones i.e IRR 5, IRR 32, IRR 39, IRR 112, PB 340, PR 300, and RRIM 600 are found susceptible with disease intensity values ranging from 44.44% to 66.67%. Five clones i.e BPM 24, GT 1, IRR 118, PR 261, and PR 303 are found highly susceptible with disease intensity value ranging from 72.22% to % (Table 5). These results are similar to previous studies which stated that clones PB 260 and RRIC 100 are resistant to Corynespora leaf fall disease (Fernando et al., 2010b), GT 1 and RRIM 600 were included in highly susceptible group on the basic of laboratory and field evaluation (Narayanan & Mydin, 2012; Jacob, 2003; Jayasinghe, 2000). The testing of recommended rubber clone by Kusdiana & Syafaah (2015) on laboratory scale revealed nine resistant clones that have the same resistance level as observed now in the greenhouse test i.e AVROS 2037, BPM 1, BPM 107, BPM 109, IRR 42, IRR 104, IRR 107, PB 260, and PB 330. In addition, there were two clones that are reported as susceptible i.e PR 300 and RRIM

7 Resistance of rubber clones recommended in Indonesia to corynespora and Table 4. Colletotrichum leaf fall disease intensity of recommended rubber clones to three of Colletotrichum sp. isolates CS-RRIM 600, CS- GT 1, and CS-PR 303. CS-RRIM 600 CS-GT 1 CS-PR 303 Clone AVROS resistant resistant resistant BPM resistant resistant susceptible BPM susceptible susceptible susceptible BPM susceptible susceptible susceptible BPM resistant resistant resistant GT resistant susceptible susceptible IRR resistant resistant resistant IRR resistant resistant susceptible IRR susceptible resistant resistant IRR resistant resistant resistant IRR susceptible resistant susceptible IRR resistant resistant resistant IRR resistant resistant resistant IRR resistant resistant resistant IRR resistant resistant susceptible PB susceptible resistant resistant PB resistant resistant resistant PB susceptible resistant resistant PR susceptible susceptible susceptible PR susceptible resistant resistant PR susceptible resistant susceptible PR resistant resistant susceptible RRIC susceptible resistant resistant RRIM resistant susceptible resistant RRIM susceptible susceptible susceptible 817

8 Testing to Colletotrichum leaf fall disease in greenhouses showed that 17 clones are resistant i.e AVROS 2037, BPM 1, BPM 109, GT1, IRR 5, IRR 32, IRR 39, IRR 42, IRR 107, IRR 112, IRR 118, IRR 220, PB 260, PB 330, PB 340, RRIC 100, and RRIM 600 with disease intensity values ranging from 15.43% to 33.03%. Eight clones were found susceptible i.e BPM 24, BPM 107, IRR 104, PR 255, PR 261, PR 300, PR 303, and RRIM 712 with disease intensity value ranging from 30.25% to 48, 26% (Table 5). Narayanan & Mydin (2012) reported that PB 260 and RRIM 600 are resistant to Colletotrichum leaf fall disease. Table 5. Corynespora and Colletotrichum leaf fall disease intensity of recommended rubber clones in greenhouse C. cassiicola C. gloeosporioides Clones Virulence Level Virulence Level AVROS resistant resistant BPM resistant resistant BPM highly susceptible susceptible BPM highly resistant susceptible BPM resistant resistant GT highly susceptible resistant IRR susceptible resistant IRR susceptible resistant IRR susceptible resistant IRR resistant resistant IRR resistant susceptible IRR resistant resistant IRR susceptible resistant IRR highly susceptible resistant IRR resistant resistant PB highly resistant resistant PB resistant resistant PB susceptible resistant PR resistant susceptible PR highly susceptible susceptible PR susceptible susceptible PR highly susceptible susceptible RRIC resistant resistant RRIM susceptible resistant RRIM resistant susceptible CONCLUSION Testing of resistance of Corynespora and Colletotrichum leaf fall disease in greenhouse showed that nine clones are resistant to both the diseases i.e AVROS 2037, BPM 1, BPM 109, IRR 42, IRR 107, IRR 220, PB 260, PB 330, and RRIC 100. Four clones were susceptible i.e BPM 818

9 Physiological characters and production toward tapping panel dryness of PB , PR 261, PR 300, and PR 303. The other clones were only resistant to one of the diseases. REFERENCES Febbiyanti, T. R. & Kusdiana, A. P. J. (2013). Pengaruh infeksi jamur Colletotrichum gloeosporioides terhadap kerusakan daun tanaman karet. Prosiding Konferensi Nasional Karet (pp ). Bogor, Indonesia: Pusat Penelitian Karet. Fernando, T. H. P. S., Jayasinghe, C. K., Wijesundera, R. L. C, & Siriwardena, D. (2010a). Susceptibility of different leaf stages of Hevea to Corynespora cassiicola. Journal of the Rubber Research Institute of Sri Lanka, 90, Fernando, T. H. P. S., Jayasinghe, C.K., Wijesundera, R.L.C., Silva, W.P.K., & Nishantha, E.A.D.N. (2010b). Evaluation of screening methods against Corynespora leaf fall disease of rubber (Hevea brasiliensis). J. Plant Dis Protec, 117(1), Hadi, H., Hartana, A., & Sinaga, M. (2004). Analisis genetika pewarisan sifat ketahanan tanaman karet terhadap penyakit gugur daun corynespora. Hayati, 11(1), 1-5. Jacob, C. K. (2003). of potential threat to rubber in India. Planters Chronicle, 92(10), Jayashinge, C. K. (2000). Corynespora leaf fall: The most challenging rubber disease in Asian and African Continents. Bull Rub Res Inst Sri Lanka, 42, Jayasinghe, C. K., & Fernando, T. H. P. S. (2011). Corynespora Leaf Fall of Hevea Rubber The Most Threatening Leaf in Asia & African Continents. Common Fund for Commodities & International Rubber Research and Development Board. Jinji, P., Xin, Z., Yangxian, Q., Yixian, X., Huiqiang, Z., & He, Z. (2007). First record of Corynespora leaf fall disease of Hevea rubber tree in China. Australasian Plant Notes, 2, Kusdiana, A. P. J., & Syafaah, A. (2015). Pengujian resistensi klon rekomendasi tanaman karet terhadap penyakit gugur daun Corynespora. Prosiding Dies Natalis Fakultas Pertanian UNSRI ke- 52 tahun 2015 (pp ). Palembang, Indonesia: Fakultas Pertanian, Universitas Sriwijaya. 819

10 Proceedings of International Rubber Conference 2017 Manju, M. J., Benagi, V. I., Shankarappa, T. H., Jacob, C. K., & Vinod, K. K. (2014). Dynamics of Corynespora leaf fall and Colletotrichum leaf spot diseases of rubber plants (Hevea brasiliensis). J. Mycol Plant Pathol, 44(1), Manju, M. J., Idicula, S. P., Jacob, C. K., Vinod, K. K., Prem, E. E., Suryakumar, M., & Kothandaraman, R. (2001). Incidence and severity of Corynespora leaf fall (CLF) disease of rubber in coastal Karnataka and North Malabar Region of Kerala. Ind J Natural Rubber Res, 14(2), Narayanan, C. & Mydin, K. K. (2012). Breeding for disease resistance in Hevea spp. - status, potential threats, and possible strategies. Proceedings of the 4 th International Workshop on Genetics of Host - Parasite Interactions in Forestry (pp ). Albany, CA: Pacific Southwest Research Station, Forest Service, U.S. Department of Agriculture Oktavia, F., Kuswanhadi, Sudarsono, Dinarti, D., & Widodo. (2016a). Diversity analysis of Corynespora cassiicola isolated from Indonesian Rubber Research Plantations. Proc Int Rub Conf 2016 (pp ). Siem Reap, Cambodia: Int Rub Res and Dev Board & Cambodian Rubber Research Institute. Oktavia, F., Sudarsono, Kuswanhadi, Dinarty, D., & Widodo. (2016b). Identifikasi ketahanan plasma nutfah karet IRRDB 1981 terpilih terhadap penyakit gugur daun Corynespora berdasarkan aktivitas toksin cassiicolin. Jurnal Penelitian Karet, 34(1), Pawirosoemardjo, S. (2004). Manajemen pengendalian penyakit penting dalam upaya mengamankan target produksi karet nasional tahun Prosiding pertemuan teknis strategi pengelolaan penyakit tanaman karet untuk mempertahankan potensi produksi mendukung industri perkaretan Indonesia tahun 2020 (pp ). Palembang, Indonesia: Balai Penelitian Sembawa, Pusat Penelitian Karet. Sinaga, M.S. (2006). Dasar-Dasar Ilmu Penyakit Tumbuhan. Jakarta: Penebar Swadaya. Situmorang, A. & Budiman, A. (2003). Penyakit Tanaman Karet dan Pengendaliannya. Palembang: Balai Penelitian Sembawa, Pusat Penelitian Karet. Situmorang, A. (2002). Sebaran penyakit gugur daun, virulensi dan genetika Corynespora cassiicola asal sentra perkebunan karet Indonesia (Disertasi), Institut Pertanian Bogor, Indonesia. 820

11 Physiological characters and production toward tapping panel dryness of PB 260 Situmorang, A., Sinaga, M. S., Suseno, R., Hidayat, S. H., Siswanto, & Darussamin, A. (2007). Virulensi isolat Corynespora cassiicola asal sentra perkebunan karet Indonesia terhadap beberapa klon karet anjuran. Jurnal Penelitian Karet, 25 (2), Situmorang, A., Sinaga, M. S., Suseno, R., Hidayat, S. H., Siswanto, & Darussamin, A. (2004). Status dan manajemen pengendalian penyakit gugur daun Corynespora di perkebunan karet. Prosiding pertemuan teknis strategi pengelolaan penyakit tanaman karet untuk mempertahankan potensi produksi mendukung industri perkaretan Indonesia tahun 2020 (pp ). Palembang, Indonesia: Balai Penelitian Sembawa, Pusat Penelitian Karet. Situmorang, A., Suryaningtyas, H., Pawirosoemardjo, S., & Sinaga, M. S. (2000). The development and virulence of races of Corynespora cassiicola on the rubber plant. Indonesian Rubber conference and IRRDB Symposium (pp ). Bogor, Indonesia: Indonesian Rubber Research Institute. Suryaningtyas, H. (2012). Pengendalian Penyakit. In M. Lasminingsih, H. Suryaningtyas, C. Nancy, & A. Vachlepi (Ed.). Saptabina usaha tani karet rakyat (pp ). Palembang, Indonesia: Balai Penelitian Sembawa. 821