Based on the transposon-mutagenized library of Pantoea agglomerans BH18, mutant screens were conducted to obtain the strain with the highest Fe (III) reduction and hydrogen production. Of these transposon-mutagenized mutants, the mutant strain TB230 was screened for high Fe (III)-reducing efficiency and hydrogen production. The PCR amplification and kanamycin resistance selection results indicated that the transposon insertion of the mutant strain TB230 was stable. Hydrogen production of the mutant strain TB230 was (2.21 +/- 0.34) mol H-2/mol glucose, which increased hydrogen production by over 40% compared with that of the wild type strain. The accumulation concentration of Fe (II) in the medium of the mutant strain TB230 with Fe (OH)(3) as the sole electron acceptor was (7.39 +/- 0.49) mmol/l, which was approximately 3-fold greater than that of the wild type strain. The mutant strain TB230 showed high Fe (III)-reducing activity and hydrogen production by adopting glucose and pyruvate as the carbon source. In addition, the mutant strain TB230 was capable of Fe (III) reduction and hydrogen production under-fresh or marine conditions. This result indicates that the mutant strain with high microbial Fe (III) reduction and hydrogen production is beneficial for the improvement of anaerobic performance. (C) 2015 Elsevier Ltd. All rights reserved.