Diffuse reflectance infrared Fourier transform (DRIFT) and Mössbauer spectroscopic study of Azospirillum brasilense Sp7: Evidence for intracellular iron(II) oxidation in bacterial biomass upon lyophilisation.

ABSTRACT

Microbial cells are well known to be capable of remaining viable when desiccated, and a variety of beneficial microorganisms can thus be preserved for storage. For the ubiquitous widely studied soil bacterium Azospirillum brasilense (wild-type strain Sp7), which has a significant agrobiotechnological potential owing to its plant-growth-promoting capabilities perspective for its use in biofertilisers, Fourier transform infrared (FTIR) spectroscopy (in the diffuse reflectance mode, DRIFT) was used to control the state of biomass, together with 57Fe transmission Mössbauer spectroscopy to monitor intracellular iron speciation in live rapidly frozen cell suspension and in the lyophilised biomass (both measured at T = 80 K). It has been shown for the first time that a relatively large part of ferrous iron in live cells (22% of the whole cellular iron pool, represented by two high-spin Fe(II) forms, in the 18-h culture grown on 57Fe(III) complex with nitrilotriacetic acid as the sole source of iron) gets largely oxidised upon lyophilisation. The remaining part of iron(II) in the resulting dry biomass was found to be ca. 3% only. The major part of ferric iron in the dry biomass was shown to be comprised of ferritin-like ferric species (giving a typical magnetically split sextet at T = 5 K), while the iron(III) formed from cellular iron(II) by oxidation in air in the course of drying remained in a paramagnetic state even at T = 5 K. The possibility of intracellular iron(II) oxidation to iron(III) upon desiccation may be a specific natural strategy to avoid cell damage caused by Fenton-type reactions in dormant (frozen, dried) cells. The results obtained may have important implications related to iron speciation and redox transformations in dried bacterial preparations intended for long-term storage.