Effects of substrate and water depth on the physiological status of a submerged macrophyte, Vallisneria natans (Lour.) H. Hara, were determined by measuring biomarkers in leaves and roots, to understand factors limiting the re-establishment of V. natans in urban eutrophic ponds. Ramets of V. natans were grown in the laboratory using aquaria containing water and bottom mud from a eutrophic pond and maintained under sufficient light in an incubator. The growth and chlorophyll-a (Chl-a) content of leaves were greater in aquaria with mud than in those with sand, which was used as the reference substrate. The contents of a peroxidation product (malondialdehyde (MDA)) and three antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD)) in leaves and roots, used as stress biomarkers, changed during the experiment, although differences in these contents between mud and sand were not consistent across the experimental days. To control water depth in the field, ramets of V. natans were grown in cages with different substrates (mud and sand) installed at different depths (0.5, 1.2, and 2.0 m) in the pond. The mean light quantum during the experiment decreased with increasing depth, from 79.3 μmol/m2 s at 0.5 m to 7.9 μmol/m2 s at 2.0 m. The Chl-a content in leaves decreased, whereas the MDA content in both leaves and roots increased with increasing water depth. All enzyme activities increased at the beginning and then decreased to the end of the experiment at 2.0 m depth, suggesting deterioration of enzyme activities due to depth-related stress. The MDA content and CAT activity were higher for sand than for mud, whereas the difference in the growth and the leaf Chl-a content between substrates remained unclear in the pond. On comparing the laboratory and field experiments, the leaf Chl-a content was found to be lower and the MDA content and enzyme activities exhibited sharp increase for ramets grown in the pond, even at 0.5 m depth, when compared with those grown in the aquaria. Our results suggest that the bottom mud of the pond is not the major limiting factor in the re-establishment of V. natans. Because water depth and light attenuation exerted strong stress on V. natans, shallow areas or measures to improve water transparency are required to promote the introduction of V. natans in eutrophic ponds for successful restoration in urban areas.
Peer Review #2 of "Effects of substrate and water depth of a eutrophic pond on the physiological status of a submerged plant, Vallisneria natans (v0.2)"
