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Acute ecological impacts of co-contamination of polycyclic aromatic hydrocarbons (PAHs) and heavy metals on diversity and composition of coastal benthic prokaryotes were unclear. We took pyrene (Pyr) and cadmium (Cd) as the representatives and mimicked an eight-week exposure of moderate and high levels of Pyr, Cd and their mixtures. 16S rRNA amplicon sequencing was used to investigate interaction of the contaminants in temporal succession of prokaryotes. Generally, concentrations of Pyr and HCl-extractable Cd in the sediments were stable over time. Effects and interaction of Pyr and Cd on prokaryotic α-diversity were temporally- and dose-dependent with a decreasing trend in richness and Shannon index under various contamination regimes, particularly in the single-Cd contaminated groups at the early stage. Temporal variability and Pyr-induced pattern in prokaryotic composition were observed. However, Pyr and Cd showed a persistent interaction in prokaryotic composition after 7 days, altering successional trajectories of communities. The communities under Pyr contamination regardless of Cd could be at a developing stage for an active PAH-degrading community with appearance of a pioneer Cycloclasticus phylotype, persistently showing a strong correlation with Pyr level. The associations of phylotypes and Cd level were short-lived and weak, corresponding to the overall resistance of prokaryotic composition to Cd. In the high-throughput sequencing era, using microcosm experiment, we renewed the knowledge about how prokaryotes vary in terms of α-diversity, composition and specific taxa in response to co-contamination of model contaminants at a temporal scale. [ABSTRACT FROM AUTHOR]