Screening Resistant Rotala Indica To Sulfonylurea And Its Resistant Molecuar Mechanism And Distribution In Paddy Field In Eastern China

Rotala indica (also known as R. rotundifolia, Lythraceae), an excellent annual aquarium plant adapts to most typical water conditions, was listed as the number one weed encountered in wet places and paddy fields in China. Rotala indica has become a widespread problem throughout rice field, especially late rice. In some field Rotala indica cannot be controlled by Sulfonylureas (such as bensulfuron-methyl and pyrazosulfuron-ethyl) at recommended rate. Rotala indica become more serious in south-west, south of Jiangsu province over the last several years.The survey and confirmed ability of Rotala indica resistance and its mechanism to bensulfuron-methyl was necessary for control Rotala indica effectively and use Sulfonylureas in a rational, scientific way. Rotala indica seeds were collected from paddy fields in Anhui, Jiangsu and Zhejiang provinces with various herbicide application histories. Rotala indica sensitivity to bensulfuron-methyl was determined by whole plant experiments in greenhouse and in vivo fluorescence technique, its resistant level, molecular mechanism and possible spread route were also invesgated. The results showed as follows:1. This study was conducted to determine the level and distribution of resistance of Rotala indica biotypes to acetolactate synthase (ALS) inhibitors and the mechanism of resistance from Jiangsu, Anhui, Zhejiang province in China (Experiment Ⅰ). A random survey was conducted across in Jiangsu province to establish the frequency and distribution of acetolactate synthase (ALS)-inhibiting herbicides in Rotala indica biotypes infesting rice fields in 2008 (Experiment I) and 2010 (Experiment Ⅱ). Some Rotala indica biotypes were resistant to ALS-inhibitor herbicide due to their possession of an ALS enzyme with decreased sensitivity to bensulfuron-methyl. To comparison of the required herbicide rate to reduce plant fresh weight 50%(GR50) and Syngenta ’RISQ’(Resistance In-Season Quick) test between the resistant biotypes and the susceptible biotypes was conducted to obtain resistance factors. Dose-response experiments using four-leaf stage (Experiment I) and 5 cm length (Experiment II) of Rotala indica plants in the greenhouse to bensulfuron-methyl was confirmed 21 Resistant(south of Jiangsu province),4 Developing resistant,3 susceptible biotypes in Rotala indica in Jiangsu province (Experiment Ⅱ). For bensulfuron-methyl, resistance factors ranged from 5.34 to 132.28 (Experiment Ⅰ) and from less than 5.71 to 808 (Experiment Ⅱ) for Rotala indica.2. To investigate the molecular basis of resistance in Rotala indica, the ALS gene was sequenced and compared between susceptible and resistant biotypes. cloning the ALS gene by PCR, got the 2000bp target bind. Also we were isolated and were found to contain introns and to encode an ALS protein of 670 amino acids. ALS nucleotide sequences of resistant and susceptible 27 Rotala indica biotypes were compared. The comparison revealed amino acid substitutions within and among the resistant R. indica biotypes (Lishui, Jurong, Jiaxing 2 and Kunshan) at position 197 were Pro to Leu. And the wild-type gene with mutant ALS gene revealed three(include position 197-Pro to Leu) differences in their amino acid sequences..3. Bensulfuron-methyl has not directly relationship with photosynthesis of weed. But difference of resistance revel between weed cause change chlorophyll a. The measurements of chlorophyll a fluorescence excitations with a variable number of saturating flashes in Rotala indica show characteristic differences between the resistant (Zhejiang2) and the wild-type (Danyangl) plants. Resistance to ALS inhibitor Rotala indica have more chlorophyll a than non resistance one.In general, the resistant levels and distributions of Rotala indica to bensulfuron-methyl were first confirmed in eastern China. The resistant mechanism being conferred to the point mutation in ALS gene and the possible spread route were first revealed. This knowledge could be useful in research of ALS inhibitor resistance mechanism and management of resistant weeds in rice field.