Three new sulfur- or non-sulfur-dependent archaeal isolates, including a Pyrococcus strain, from Guaymas Basin hydrothermal vents (Gulf of California; depth, 2,010 m) were characterized and physiologically compared with four known hyperthermophiles, previously isolated from other vent sites, with an emphasis on growth and survival under the conditions particular to the natural habitat. Incubation under in situ pressure (200 atm [1 atm = 101.29 kPa]) did not increase the maximum growth temperature by more than 1 degrees C for any of the organisms but did result in increases in growth rates of up to 15% at optimum growth temperatures. At in situ pressure, temperatures considerably higher than those limiting growth (i.e., > 105 degrees C) were survived best by isolates with the highest maximum growth temperatures, but none of the organisms survived at temperatures of 150 degrees C or higher for 5 min. Free oxygen was toxic to all isolates at growth range temperatures, but at ambient deep-sea temperature (3 to 4 degrees C), the effect varied in different isolates, the non-sulfur-dependent isolate being the most oxygen tolerant. Hyperthermophiles could be isolated from refrigerated and oxygenated samples after 5 years of storage. Cu, Zn, and Pb ions were found to be toxic under nongrowth conditions (absence of organic substrate), with the non-sulfur-dependent isolate again being the most tolerant.