The response of mesozooplankton is critical in assessing the health of an estuarine ecosystem. Reports on the spatial and temporal dynamics of mesozooplankton community in estuarine and backwaters of the Southern parts of India are scanty. In this scenario, we appraised the community structure of mesozooplankton and their spatio-temporal dynamics based on various multivariate statistical assessments. A total of 31 taxa were identified and the abundance was principally dominated by Copepoda followed by Luciferidae during three sampling seasons. The most abundant species were: Paracalanus parvus parvus, Pseudodiaptomus aurivillii, Temora stylifera, and Pseudodiaptomus serricaudatus. Canonical correspondence analysis and Spearman's correlation coefficients underlined that salinity, turbidity, conductivity, temperature, dissolved oxygen, chlorophyll a (Chl-a), and nutrients were the principal environmental variables strongly linked with mesozooplankton dynamics in Munroe Island. The highest abundance of mesozooplankton was recorded in MoN (monsoon), followed by PrM (pre-monsoon), and PoM (post-monsoon). Hierarchical clustering confirmed that the grouping of sampling stations is based on the estuarine and freshwater influences on mesozooplankton abundance. During the entire investigation, various ecological indices were observed in good condition. Moreover, the optimum environmental conditions during the PoM season are marked with the highest indices values. Overall, multivariate investigations undoubtedly proved the suitability of mesozooplankton communities as potential bioindicators for spatial and seasonal ecological assessments. Our investigation emphasizes the high assemblages of mesozooplankton and their responses to various environmental variables and highlights the significance of long-term ecological monitoring in a threatened ecosystem like Munroe Island.