Discussion
The effect of leaf litter leachates on the growth, production, and cell stoichiometry of the green alga Scenedesmus varied significantly among the leaf litter leachates of different tree species. As nutrient concentrations were not the same among the culture media, growth rates in different types of leachates could not be directly compared with each other. Therefore, we plotted the C biomass accumulated during the 7-day incubation period against TDN and TDP in the leachate. The result showed that the accumulated C biomass increased with increasing TDN in the leachate regardless of the micronutrient amendment, although the increasing trend was attenuated at higher TDN concentrations. In addition, C biomass was highly limited in the leachates of Siebold’s beech, Japanese elm and Japanese hemlock regardless of high TDP concentrations (species ID 5, 6, 7 in Fig. 2g and h). These results suggest that the variation in C biomass among the different culture media from leaf litter leachate was mainly caused by TDN concentration. In a previous study, Ho et al. (2023) suggested that nitrogen is the most deficient element in the leaf litter leachates relative to phytoplankton demand, because the percentage of release relative to leaf litter elemental content was lowest for TDN compared to DOC and TDP in all 11 tree species studied. The present study confirmed the validity of this suggestion for phytoplankton production.
However, the algal growth rate, estimated from temporal changes in cell abundance, differed from that of C biomass. In the present study, it ranged from 0.2 to 0.3 d-1in for most of the leaf litter leachates, except for Japanese elm and Japanese hemlock, where the growth rate was <0.2 d-1. The result suggests that the nutrient conditions in the present study were moderate according to the growth rate of Scenedesmus under different culture conditions in previous studies . However, the growth rate was not closely related to either TDN or TDP in leaf litter leachate, especially when micronutrients were not amended. The result suggests that the factors regulating growth rate, i.e., cell division rate, and net C biomass accumulation rate, i.e., organic carbon production rate, are different in this algal species.
It should be noted that when micronutrients were added, the algal growth rate significantly increased in the leaf litter leachates of oak, Japanese elm and Japanese hemlock, although the opposite trend was found in the leachate of hinoki cypress. The results contrasted with the response of C biomass, which showed a significant difference between the culture medium with and without micronutrient amendment only in Japanese hemlock. These results suggest that the fitness of the present algal species in the terms of cell division rate is more sensitive to micronutrient supply than the net production rate. In fact, the algal growth rate was significantly correlated with TDP but not TDN when micronutrients were additionally supplied. This fact implies that the variation of algal growth rate among different types of leaf litter leachates was mainly regulated by P supply in concert with micronutrients, whereas that of net production rate was more regulated by N supply. The relative importance of P rather than N may be related to the fact that P is an essential element of DNA and RNA and can strongly limit cell division .
Micronutrients are known to control the enzyme activity that regulates the N and P use efficiency of phytoplankton . Indeed, in leaf litter leachates from Japanese red pine and hinoki cypress, a significant increase in N and P cell quotas was found when micronutrients were additionally supplied. In these types of leaf litter leachates, micronutrients may play a role in P and N use efficiency. In addition, micronutrients appear to play different roles in algal growth and C biomass accumulation. Several studies have shown that the factors affecting algal cell division rates are different from those affecting the photosynthetic production rates . For example, Li et al. showed that in the dinoflagellate Amphidinium carterae , the cell division cycle was halted but the photosynthesis continually accumulated C in cells under P deficiency. On the contrary, Negi et al. found that in the green alga Chlorella sorokiniana , N deficiency did not change the cell density, but decreased the photosynthetic rate.
As mentioned above, the addition of micronutrients tended to reduce the growth rate of Scenedesmus incubated in a culture medium from the leaf litter leachate of hinoki cypress. The result suggests that the leaf litter leachate of hinoki cypress may contain excessive amounts of some micronutrients that are harmful to Scenedesmus . Rueter et al. showed that Al and Cu tended to reduce the growth rate ofScenedesmus when supplied in excess. Because we added a mixture of micronutrients, we could not determine which micronutrients and trace elements were harmful when supplied in excess. However, the present study indicates that the water-soluble content of micronutrients and trace elements varies widely among the leaf litter of different tree species as suggested by Pourhassan et al. .
Studies have shown that decomposition and nutrient release rates somewhat differ between coniferous and broadleaf tree species . However, we could not determine whether leaf litter leachates from broadleaf or coniferous trees had a better nutrient composition for the growth rate of Scenedesmus due to the large variation of growth rate within each of these two groups. In addition, we could not find significant difference in the C biomass production rate of this green alga between the culture media from broadleaf and coniferous leaf litter leachates due to the large variations in the rate within these tree species. These results suggest that the effect of leaf litter on aquatic organisms cannot be generalized across broad taxonomic tree groups.
Similar to the growth and C biomass production rates, leaf litter leachates from different tree species had different effects on the C:N:P ratio of the algae. It is well known that the elemental contents of algae are important determinants of the quality of food for herbivorous zooplankton . In this study, the molar C:N and C:P ratios ofScenedesmus varied from 6.6 to 21 and 106 to 600, respectively, depending on the leaf litter leachate. In general, algal cells with C:N and C:P ratios close to Redfield ratios are good quality food for zooplankton growth . In the leaf litter leachates of Siebold’s beech, Japanese elm, Japanese red pine and hinoki cypress, the molar C:N and C:P ratios ranged from 6.6 to 12 and 106 to 160, respectively, which are close to the Redfield ratios. Thus, algae grown in lentic waters receiving nutrients from leaf litter of these species would be suitable food for herbivorous zooplankton. However, in the leaf litter leachates from Japanese maple and Japanese cedar, the C:P ratio ofScenedesmus cells was greater than the threshold elemental ratio (>300) above which the growth rate of zooplankton such asDaphnia is limited by phosphorus . Urabe and Watanabe also suggest that algal cells with C:N ratios greater than 15 are suboptimal food for zooplankton. In this study, the C:N ratio of Scenedesmuscells in the leaf litter leachates of Japanese maple and Japanese hemlock was greater than this value. In addition, depending on the leaf litter leachate of different tree species, algae may alter the cellular content of other chemical substances, such as fatty acids and amino acids, that affect food quality for herbivores. Together with such a possibility, the present results suggest that the mass transfer from algae to herbivorous zooplankton in a lake may be influenced by the vegetation composition in the watershed via subsidies of nutrient inputs from leaf litter.