Inoculation method of downy mildew of alfalfa and enzyme activity in infected plants

Downy mildew (Peronospora aestivalis) is a common disease found on alfalfa and has severely curbed the development of the alfalfa industry. To explore the effect of downy mildew infection on the protective enzyme activities in alfalfa and optimize the inoculation method of downy mildew of alfalfa, this study took alfalfa ‘Zhongcao No. 3’ as the research object, inoculated it with downy mildew, and investigated the enzyme activity determination of catalase (CAT), superoxide dismutase (SOD), peroxidase (POD), and phenylalanine ammonia-lyase (PAL) under different infection degrees. The results showed that, among the four inoculation methods, the diseased branch insertion method could not successfully inoculate diseased plants, and the diseased leaf friction, spray method in the incubator and outside the incubator could successfully inoculate diseased plants, with incidence rates of 11.1%, 88.9%, and 25.6%, respectively. The spray method in the incubator was observed to have the best inoculation effect. The inoculation had a significant impact on the enzyme activity in alfalfa, the mild downy mildew infection of alfalfa could cause CAT, and the activity was significantly enhanced (P < 0.05). Compared with healthy alfalfa, alfalfa infected with downy mildew could significantly enhance POD and SOD activity (P < 0.05), however, there was no significant difference in SOD activity between severe infection and healthy alfalfa. Moreover, the PAL activity showed a significant decrease as the degree of pathogen infection deepened (P < 0.05). This shows that the invasion of pathogenic bacteria can cause changes in the enzyme activity in alfalfa, and this study provides theoretical reference for the inoculation of downy mildew in alfalfa and the screening of resistant varieties by the changes in enzyme activities in alfalfa.