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Tonle Sap Great Lake (TSL) is the largest freshwater lake ecosystem in Southeast Asia and receptacle of impressive biodiversity. However, there is surprisingly little knowledge of its ecosystem structure and functioning. The main objective of the current work was to quantify the food web structure and assess the ecosystem health status of the TSL system by constructing the first holistic food web model using Ecopath with Ecosim (EwE). The results indicate that the ecotrophic efficiency (EE) values were very high for most of functional groups (EE > 0.5) except molluscs (0.146) and macrophytes (0.102). The high EE values together with the MTI (Mixed Trophic Impact) analysis indicated the overexploitation and degradation of fishery resources in the TSL system. The discrete trophic levels varied from 1 (phytoplankton, macrophytes and detritus) to 3.17 (snakehead). The energy transfer in the TSL food web was based mostly on the detrital food chain (77.9%) rather than the grazing food chain (22.1%), with an average transfer efficiency of 8.27%. The ratios of total primary production to respiration (TPP/TR) and to biomass (TPP/TB) were 1.23 and 2.04, respectively, while the Ascendency and Finn cycling index (FCI) of the system were estimated at 27.4% and 23.62%. Nevertheless, the connectance index (CI: 0.253) and system omnivory index (SOI: 0.075) were in-between compared to other lake ecosystems, which indicated that the food web structure was characterized by linear, rather than web-like features. Systematic analysis and indicators suggested that the ecosystem was a relatively healthy ecosystem achieving a certain stage of maturity, albeit with a vulnerable food web structure. Accordingly, some ecosystem-based strategies are presented for the improvement of fishery management and ecosystem conservation in TSL. [ABSTRACT FROM AUTHOR]