Clostridium thermocellum JW20 (ATCC 31549), which was isolated from a Louisiana cotton bale, grew on cellulose, cellobiose, and xylooligomers and, after adaptation, on glucose, fructose, and xylose in the pH range of 7.5 to 6.1 with T(opt) of 60 degrees C, T(max) of 69 degrees C, and T(min) of above 28 degrees C. Doubling times during growth on cellulose and cellobiose were 6.5 and 2.5 h, respectively. The G+C content of the DNA was 40 mol% (chemical analysis). Growth on cellulose as substrate was totally inhibited in the presence of more than 125 mM sodium sulfate, 300 mM sodium chloride, 250 mM potassium chloride, 200 mM calcium chloride, 125 mM magnesium chloride, 40 mM lactate, or 250 mM acetate. The ratio of the fermentation products ethanol to acetate plus H(2) decreased when the culture was agitated. Agitation otherwise increased the rate of cellulose degradation in a growing culture but not under nongrowth conditions or with cell-free culture supernatant containing the extracellular cellulase. Shaking lowered the concentration of H(2) in the culture broth and thus minimized inhibition by the H(2) formed. Externally added H(2) caused an increased formation of ethanol during growth on cellulose or cellobiose. However, at an atmospheric pressure as high as 355 kPa (50 lb/in), H(2) did not cause significant growth inhibition beyond an increasing lag phase (up to 24 h). Several criteria to specifically prove the purity of C. thermocellum cultures were suggested.