Ocean acidification (OA), a consequence of increased global carbon dioxide (CO2) emissions, is considered a major threat to marine ecosystems. Its effects on bacterioplankton activity, diversity, and community composition have received considerable attention. However, the direct impact of OA on heterotrophic bacterioplankton is often masked by the significant response of phytoplankton due to the close coupling of heterotrophic bacterioplankton and autotrophs. Here we investigated the responses of a heterotrophic bacterioplankton assemblage to high pCO2 (790-ppm) treatment in warm tropical western Pacific waters by conducting a microcosm experiment in dark for 12 days. Heterotrophic bacterioplankton abundance and production were enhanced by OA over the first 6 days of incubation, while the diversity and species richness were negatively affected. Bacterioplankton community composition in the high pCO2 treatment changed faster than that in the control. The molecular ecological network analysis showed that the elevated CO2 changed the overall connections among the bacterial community and resulted in a simple network under high CO2 condition. Species-specific responses to OA were observed and could be attributed to the different life strategies and to the ability of a given species to adapt to environmental conditions. In addition, high-throughput functional gene array analysis revealed that genes related to carbon and nitrogen cycling were positively affected by acidification. Together, our findings suggest that OA has direct effects on heterotrophic bacterioplankton in a low-latitude warm ocean and may therefore affect global biogeochemical cycles.