The oxidation of hydrogen and NADH by membrane fractions of two autotrophically grown hydrogen bacteria, Pseudomonas facilis and strain 14 g, both lacking a hydrogen dehydrogenase, was studied by difference-spectrophotometric and manometric methods. The spectrophotometric data did not support the existence of two separate electron transport pathways for both the substrates. However, from the effect of rotenone, antimycin A, BAL, and HQNO on the oxygen uptake rate with H2 or NADH as substrates, separate pathways could be proposed: in strain 14 g at least to cytochrome b and in P. facilis at least to cytochrome c.