Peer Review #3 of "Evaluation of waterlogging tolerance and responses of protective enzymes to waterlogging stress in pumpkin (v0.2)"

Waterlogging caused by short and severe, or prolonged precipitation can be attributed to global warming.Pumpkin plants are drought-tolerant but not tolerate to waterlogging stress.Under frequent rain and waterlogging conditions, the production of pumpkins is of lower quality, sometimes rotten, and harvest failure occurs in severe cases.Therefore, it is of great significance to assess the waterlogging tolerance mechanism of pumpkin plants.In this study, 10 novel pumpkin varieties from Baimi series were used.The waterlogging tolerance level of pumpkin plants was evaluated by measuring waterlogging tolerance coefficient of biomass and physiological indices using waterlogging stress simulation method.The criteria to evaluate the waterlogging tolerance capacities of pumpkin plants were also explored.Using principal component and membership function analysis, waterlogging tolerance levels of the pumpkin varieties were ranked as follows: Baimi No. 10> Baimi No. 5> Baimi No. 1> Baimi No. 2> Baimi No. 3> Baimi No. 7> Baimi No. 9> Baimi No. 6> Baimi No. 4> Baimi No. 8. Based on the results, Baimi No. 10 was identified with strong waterlogging tolerance and Baimi No. 8 with weak waterlogging tolerance.The responses of malondialdehyde (MDA), proline, key enzymes responsible for anaerobic respiration, and antioxidant enzymes to waterlogging stress were studied in pumpkin plants.The relative expression levels of related genes were determined using real-time fluorescence quantitative PCR technique.The aim of our study was to assess the waterlogging tolerance mechanism of pumpkin plants, thus laying a theoretical foundation for breeding waterlogging-tolerant varieties in the future.After flooding stress treatment, the antioxidant enzyme activities, contents of proline and alcohol dehydrogenases of BaimiNo. 10 and Baimi No. 8 displayed an increase followed by a decrease.All indices of Baimi No. 10 were higher than Baimi No. 8. MDA contents gradually increased, with the content being higher in Baimi No. 8 than Baimi No. 10.The activities of pyruvate decarboxylases (PDCs) in Baimi No. 8 and Baimi No. 10 exhibited a decrease initially, followed by an increase, and then a decrease again.The PDC activity in Baimi No. 8 was generally higher