GENETIC IMPROVEMENT OF SUGARCANE BY CONVENTIONAL AND MOLECULAR APPROACHES
Location: Sugarcane Research Unit
Title: First report of ratoon stunt of sugarcane caused by Leifsonia xyl. subsp. xyli in Pakistan
| Hussnain, S - |
| Afghan, S - |
| Haq, M - |
| Mughal, S - |
| Shahazad, A - |
| Hussain, K - |
| Nawaz, K - |
| Jackson, P - |
| Batool, A - |
| Irfan, A - |
Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: August 1, 2011
Publication Date: August 10, 2011
Citation: Hussnain, S.Z., Afghan, S., Haq, M.I., Comstock, J.C., Mughal, S.M., Shahazad, A., Hussain, K., Nawaz, K., Pan, Y.-B., Jackson, P., Batool, A., Irfan, A. 2011. First report of ratoon stunt of sugarcane caused by Leifsonia xyl. subsp. xyli in Pakistan. Plant Dis. 95:1581.
Interpretive Summary: Sugarcane is grown over one million hectares in Pakistan with an average tonnage of about 50 tons per hectare. There are several reasons for this low yield and the most important is the lack of cultivars that are resistant to the red rot, ratoon stunting (RSD) and rust diseases. A field disease survey was conducted during 2007 to 2010 in Punjab province of Pakistan. Typical RSD symptoms were obvious on plants of cultivars CP77-400, SPF-241, CP72-2086 and NCo-310. Chemical tests indicated that up to 25% plants surveyed had RSD. The causal RSD bacteria were isolated and antibody was raised against the bacterium. Cultivars SPF-213, CPF-237, HSF-240, NSG-555, SPSG-26, SPSG-79, SPF-238 and CP77-400 were resistant, SPF-241 was moderate resistance, and CP72-2086 and NCo-310 were highly susceptible to RSD. The RSD-specific antibodies have permitted the immunoassay-based RSD disease detection and artificial inoculation during RSD disease screening experiments in Pakistan to improve sugarcane breeding for disease resistance to RSD.
During a survey of the sugarcane crop in the area of Faisalabad, Sargodha and the Dera Ghazi Khan Division of the Punjab province of Pakistan from 2007 to 2010, symptoms consistent with ratoon stunting, including stunted growth and reddening of the vascular bundles at the nodal regions were observed on sugarcane varieties CP77-400, SPF-241, CP72-2086 and NCo-310. The two varieties CP72-2086 and NCo-310 showed severely stunted growth. Chemical test was performed for detecting ratoon stunt from the field. Longitudinal sections of mature nodes were treated with a combination of hydrogen peroxide and hydrochloric acid. Healthy canes developed a blue-green color in the parenchymatous tissue around the fibrovascular bundles, whereas diseased cane did not. This field test illustrated that up to 25% plants were infected by ratoon stunt in the survey area. Aerobic bacteria were isolated from a stunted plant of variety NCo-310 on a modified culture medium and incubated for 3-4 week at 28°C. Light off-white, round and raised growth bacterial colonies were observed. The size of the bacteria was 1.5-4.5 µm by 0.2-0.35 µm. Isolates were positive for the Gram and catalase reactions and negative for oxidase, aesculin hydrolysis, urease production, and motility. The pathogen was identified as Leifsonia xyli subsp. xyli (Lxx) (formerly Clavibacter xyli subsp. xyli) based on its morphological characteristics. A DAC-ELISA was developed with antiserum raised against Lxx at the National Institute for Biotechnology and Genetic Engineering (NIBGE) Faisalabad, Pakistan. Infected/suspected to be infected plants of different varieties were used for ELISA test. This ELISA showed that sugarcane varieties NCo-310 (Log 1.342 CFU/ml) and CP72-2086 (Log 0.118 CFU/ml) had higher Lxx titres than the other varieties tested (SPF-213 (Log 0.071CFU/ml), CPF-237 (Log 0.077CFU/ml), HSF-240 (Log 0.069 CFU/ml), NSG-555 (Log 0.060 CFU/ml), SPSG-26 (Log 0.076 CFU/ml), SPSG-79 (Log 0.074 CFU/ml), SPF-238 (Log 0.057 CFU/ml) and CP77-400 (Log 0.063 CFU/ml). Variety SPF-241 (Log 0.107 CFU/ml) was weakly positive for ratoon stunt. The axillary buds of sugarcane were injected via a sterile hypodermic syringe with an 18-gauge needle to deliver a bacterial suspension of 109 cells/ml. Inoculated sugarcane plants were examined at intervals over a period of nine months for the development of symptoms and the presence of bacteria. Varieties were evaluated on the bases of average number of colonized vascular bundles. SPF-213, CPF-237, HSF-240, NSG-555, SPSG-26, SPSG-79, SPF-238 and CP77-400 were resistant; SPF-241 showed moderate resistance and CP72-2086 and NCo-310 were highly susceptible to ratoon stunt. The pathogen was re-isolated from the inoculated plants and identified as Lxx by bacteriological tests and its serological reaction. To our knowledge this is the first report of ratoon stunt of sugarcane in Punjab province of Pakistan.