38. Evaluation of durable resistance of transgenic hybrid maintainer line IR58025B for bacterial blight disease of rice

1) Plant Breeding, Genetics, and Biochemistry Division, International Rice Research Institute, DAPO Box 7777,Metro Manila, Philippines.
2) Directorate of Rice Research, Hyderabad-500 030, India
3) Central Rice Research Institute, Cuttack-753006, India

Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Ishiyama) Swings et al. (Xoo), is the most destructive disease of rice worldwide which causes significant yield losses annually (Mew, 1987). Yield loss from this disease can be as high as 50% (Adhikari et al. 1995). In Punjab and Haryana states of India, major epidemics occurred in 1979 and 1980; severe kresek was observed and total crop failure was reported (POS, 1979 &1980; Mew, 1987). Compared with conventional inbred varieties, rice hybrids are more responsive to nitrogenous fertilizer and exhibit vigorous growth. These attributes have made most of the rice hybrids highly sus-ceptible to bacterial blight disease (Mew et al. 1988; Reddy et al. 1998). Cai and Zhong (1980) reported that bacterial blight is a common disease on hybrid rice in Zhejiang Province in China.

In view of the limited success in chemical control of the disease, the importance of hostplant resistance has been well recognized. So far, 24 resistance genes have been identified in rice and its wild relatives (Kinoshita, 1995; Lin et al. 1998). The dominant gene Xa21 introgressed from Oryza longistaminata into O. sativa confers broad-spectrum resistance to most of the isolates of Xoo, making it a valuable gene for resistance to bacterial blight disease (Khush et al. 1990). However, the improvement of hybrid parental lines through conventional breeding is a time-consuming process and linkage with undesirable characters is a problem. Song et al. (1995) cloned the Xa21 gene through map-based strategy. This is the first wellcharacterized disease-resistance gene that has been characterized in rice. Tu et al. (1998) introduced the Xa21 gene into elite indica cultivar IR72 via the biolistic transformation method. The improved transgenic IR72 carrying Xa21 exhibited broad-spectrum resistance in field tests (Tu et al. 2000).

The maintainer line IR58025B (designated here as ML1) was transformed with bacterial blight resistance gene Xa21 using the particle bombardment method. We report here the reaction of homozygous T3 transgenic ML1 lines to two Philippine races of Xoo, race 4 (PXO 61)

and race 6 (PXO 99), and the stable integration of Xa21 in subsequent generations. A total of 140 T3 homozygous transgenic ML1 plants from seven different lines were grown in the IRRI CL4 containment green house. Non-transformed ML1 plants served as the control. Standard agronomic practices were followed for growing the plants, except that no chemical pesticides were applied to allow for an optimum evaluation of the resistance reaction. The plants were inoculated at the maximum tillering stage following the leaf-clipping technique (Kauffman et al. 1973). For this purpose, the bacteria were mass-multiplied on peptone sucrose agar or modified Wakimoto's medium. The bacteria were harvested from 72 h growth and concentration was adjusted to 108-109 c.f.u./ml of suspension. In each line, sets of 10 plants were inoculated with race 4 and another 10 plants were inoculated with race 6 of Xoo. Observations were recorded 14 days after inoculation on lesion length on transgenic and non-transgenic plants.

All the lines tested were highly resistant to race 4 and possessed a good level of resistance to race 6 (Figure 1). Philippine race 6 is a highly virulent race. The mean lesion length against race 4 varied from 4.04 to 5.74 cm in transgenic plants, whereas the average lesion length in non-transformed ML1 plants was 34.11 cm (Table 1). Against race 6, the average lesion length varied from 8.79 to 11.16 cm in transgenic plants compared with 24.97 cm in nontransformed plants. The variation in the bacterial blight reaction among the lines may be due to differences in the level of expression of the transgene. Among the different lines tested, ML1-7-9-1-8 and ML1-7-9-24-2 were the most promising. The bioassay results were further

supplemented by Southern blot analysis of the Xa21 gene. The presence of the 3.8-kb band indicated the presence of the Xa21 gene (Figure 2). The study revealed that transgenic ML1 plants carrying the Xa21 gene showed a high level of resistance to both races of the bacterial blight pathogen. The study also revealed that the increased level of resistance to the bacterial blight pathogen persists in transgenic plants over generations, indicating its stable inheritance. This homozygous transgenic maintainer line can be useful in a hybrid rice development program.


Thanks are due to Dr. Pamela Ronald of UC-Davis for providing the Xa21 gene, the Rockefeller Foundation for partial financial support, and Bill Hardy for editorial assistance.


Adhikari, T.B., C.M. Vera Cruz, Q. Zhang, R.J. Nelson, D.Z. Skinner, T.W. Mew and J.E. Leach, 1995. Genetic diversity of Xanthomonas oryzae pv. oryzae in Asia. Appl. Environ. Microbiol. 61: 966-971.

Cai, S. F. and M.Y. Zhong, 1980. Control experiment of diseases and pests of hybrid late rice [In Chinese]. Zhejiang Nongye Kexue 4: 166-169.

Kauffman, H.E., A.P.K. Reddy, S.P.Y. Hsieh and S.D. Merca, 1973. An improved technique for evaluating resistance of rice varieties to Xanthomonas oryzae. Plant Dis. Rep. 57: 537- 541.

Khush, G.S., E. Bacalangco and T. Ogawa, 1990. A new gene for resistance to bacterial blight from O. longistaminata. Rice Genet. Newsl. 67: 121-122.

Kinoshita, T., 1995. Report of committee on gene symbolization, nomenclature and linkage groups. Rice Genet. Newsl. 12: 9-115.

Lin, X., C., Wang, G. Wen, D. Zhang, Y. Xie and Q. Zhang, 1998. Progress in map-based cloning of Xa22(t), a new gene for bacterial blight resistance in rice. Paper presented at the Plant and Animal Genome Conference, 18-22 Jan 1998, San Diego, Calif., USA.

Mew, T.W., 1987. Current status and future prospects of research on bacterial blight of rice. Annu. Rev. Phytopathol. 25: 359-382.

Mew, T. W., F.M. Wang, J.T. Wu, K.R. Lin and G.S. Khush, 1988. Disease and insect resistance in hybrid rice. In: Hybrid rice. Manila (Philippines): International Rice Research Institute. p. 189-200.

POS (1979 & 1980). Production-oriented survey, Directorate of Rice Research, Hyderabad, India.

Reddy, A.P.K., K. Krishnaiah, Z.T. Zhang and Y. Shen, 1998. Managing vulnerability of hybrid rice to biotic stresses in China and India. In: Proceedings of the 3rd International Symposium on Hybrid Rice Technology: Advances in hybrid rice technology (Virmani, S.S., E.A. Siddiq, and K. Muralidharan, eds.). Hyderabad, India & International Rice Research Institute, Philippines. p 147-155.

Song, W.Y., G.L. Wang, L. Chen, H-S. Kim. Li-Ya Pi. J. Garner, B. Wang, T. Holsten, W.X. Zhai, L.H. Zhu, C. Fauquet and P. Ronald, 1995. A receptor kinase-like protein encoded by the rice disease resistance gene Xa21. Science 270: 1804-1806.

Tu, J., I. Ona, Q. Zhang, T.W. Mew, G.S. Khush and S.K. Datta, 1998. Transgenic rice variety IR72 with Xa21 is resistant to bacterial blight. Theor. Appl. Genet. 97: 31-36.

Tu, J., K. Datta, G.S. Khush, Q. Zhang and S.K. Datta, 2000. Field performance of Xa21 transgenic indica rice (Oryza sativa L.), IR72. Theor. Appl. Genet. 101: 15-20.