In situ colonization experiments were performed to study the pioneer populations of bacteria at Loihi Seamount, Hawaii. Over a ten-year sampling period, 41 microbial growth chambers (MGCs) were deployed and recovered in Pele's Pit and the surrounding area after short-term (4–10 days) and long-term (∼1–6 years) incubations in the flow of hydrothermal effluent. Terminal-restriction fragment length polymorphism (T-RFLP) analysis of the small subunit rRNA gene (SSU rDNA) revealed that the short-term MGC communities exhibited a low number of represented populations when compared to the long-term MGC communities and naturally occurring microbial mats. Cluster analysis of T-RFLP fingerprints showed the short-term MGC communities all had similar richness but were separated into three distinct groups with different arrays of colonizing populations. Clone library analysis showed that cooler vents (T ave = 40°C) were primarily colonized by Mariprofundus ferrooxydans, a neutrophilic Fe-oxidizing ζ −Proteobacteria while warmer vents (T ave = 71°C) were colonized by Sulfurimonas spp. and other sulfur-cycling members of the ε −Proteobacteria. Vents with an intermediate temperature (T ave = 51°C) were colonized by representatives of both ζ -Proteobacteriaand ε -Proteobacteria. Long-term MGC communities did not cluster with any of the short-term communities and exhibited higher richness, indicating a greater number of bacterial populations were able to colonize and grow in the long-term growth chambers.