An ensemble data assimilation scheme, Error Subspace Statistical Estimation (ESSE), is utilized to investigate the seasonal ecosystem dynamics of the Lagoon of Venice and provide guidance on the monitoring and management of the Lagoon, combining a rich data set with a physical-biogeochemical numerical estuary-coastal model. Novel stochastic ecosystem modeling components are developed to represent prior uncertainties in the Lagoon dynamics model, measurement model, and boundary forcing by rivers, open-sea inlets, and industrial discharges. The formulation and parameters of these additive and multiplicative stochastic error models are optimized based on data-model forecast misfits. The sensitivity to initial and boundary conditions is quantified and analyzed. Half-decay characteristic times are estimated for key ecosystem variables, and their spatial and temporal variability are studied. General results of our uncertainty analyses are that boundary forcing and internal mixing have a significant control on the Lagoon dynamics and that data assimilation is needed to reduce prior uncertainties. The error models are used in the ESSE scheme for ensemble uncertainty predictions and data assimilation, and an optimal ensemble dimension is estimated. Overall, higher prior uncertainties are predicted in the central and northern regions of the Lagoon. On the basis of the dominant singular vectors of the ESSE ensemble, the two major northern rivers are the biggest sources of dissolved inorganic nitrogen (DIN) uncertainty in the Lagoon. Other boundary sources such as the southern rivers and industrial discharges can dominate uncertainty modes on certain months. For dissolved inorganic phosphorus (DIP) and phytoplankton, dominant modes are also linked to external boundaries, but internal dynamics effects are more significant than those for DIN. Our posterior estimates of the seasonal biogeochemical fields and of their uncertainties in 2001 cover the whole Lagoon. They provide the means to describe the ecosystem and guide local environmental policies. Specifically, our findings and results based on these fields include the temporal and spatial variability of nutrient and plankton gradients in the Lagoon; dynamical connections among ecosystem fields and their variability; strengths, gradients and mechanisms of the plankton blooms in late spring, summer, and fall; reductions of uncertainties by data assimilation and thus a quantification of data impacts and data needs; and, finally, an assessment of the water quality in the Lagoon in light of the local environmental legislation.