Tracing Nitrogen Distribution and Biotic Responses in Spring-Fed Karst Rivers: A Pilot Study

Understanding nitrogen distribution in spring-fed karst rivers is important for interpreting ecosystem responses in populated Mediterranean landscapes. Nitrogen, in its various forms, is a key physicochemical quality element influencing biological communities and ecological quality of freshwater ecosystems. Elevated nitrogen availability may trigger eutrophication and other processes associated with biodiversity loss, posing risks to both aquatic ecosystem integrity and drinking water quality. However, translating nitrogen measurements into effective monitoring and management strategies remains challenging. Monitoring programs are often resource-intensive and require site-specific adaptation, particularly in heterogeneous systems such as karst catchments. General guideline values may not fully capture local hydrological variability, groundwater–surface water interactions, or combined stressors, including nutrient mixtures and salinity intrusion. These factors introduce uncertainty and complicate the interpretation of nitrogen dynamics. This pilot-scale exploratory study assessed total nitrogen (TN) across four environmental matrices—water and sediments, as well as tissue TN in aquatic bryophytes, and in benthic macroinvertebrates—at four spring-fed sites within the Koiliaris River Basin (Crete, Greece). The Koiliaris Critical Zone Observatory (CZO) is a representative karst watershed with highly permeable carbonate geology and long-term human pressures. TN concentrations were low in water (0.9–1.4 mg/L) and sediments (0.2–1.1 g/kg) but substantially higher in biotic compartments, particularly in macroinvertebrates (29.8–47.1 g/kg), while moss tissue TN ranged between 16.9 and 20.4 g/kg. Spatial variability among sites was observed, with consistently higher TN values at the coastal spring influenced by seawater intrusion. Although the limited sample size precluded formal statistical inference, exploratory analyses indicated positive associations between water TN and tissue TN in mosses and macroinvertebrates. These preliminary findings suggest that dissolved nitrogen may represent an important pathway of nitrogen availability to aquatic biota in this karst system. The study provides an exploratory framework for integrating abiotic and biotic nitrogen measurements and may inform the design of future, larger-scale investigations in Mediterranean spring-fed rivers.