Abstract: The macrozoobenthic community structures of the mainstream of Huai Rirer were investigated in March, June, September, and December 2011. A total of 94 macrozoobenthic species from 41 families were recorded. Results showed that there was a significant temporal variation in the macrozoobenthic density（p = 0.002). The maximum mean population density（332.5 ind./m2) occurred in June while the minimum population density（96.6 ind./m2) appeared in September. And yet, the temporal variation trend of biomass was not consistent with that of population densities. The maximum mean biomass（155.8 g/m2) occurred in September while the minimum biomass（40.0 g/m2) appeared in March, and there were no obvious differences among seasons. The redundancy analysis（RDA) results showed that the dissolved oxygen, salinity, total dissolved solids, temperature, and sediment types were the main environmental factors that affect the macrozoobenthic community structure. The temporal variation of Shannon-Wiener diversity index was not significant, but the Margalef richness index was showed significant seasonal variations（p = 0.000). Even so, the variation trends of the two indices were highly consistent from the upstream to the downstream. Overall, the water quality of the mainstream of Huai River was evaluated at the middle-level pollution.