Suppressed Biofilm Formation Efficiency and Decreased Biofilm Resistance to Oxidative Stress and Drying in an Azospirillum brasilenseahpC Mutant

Azospirillum brasilense forms biofilms on various surfaces. It was previously shown that inactivation of the ahpC gene of alkyl hydroperoxide reductase in the SK586 mutant of A. brasilense Sp245 led to an increased sensitivity to peroxides and to changes in flagellation, motility, and cell surface properties. This work revealed that under nitrogen limitation, Sp245 switches predominantly to the biofilm mode of life, whereas this tendency is not characteristic of the mutant SK586. Inactivation of the ahpC gene negatively affects the viability of SK586 in biofilms, which is most noticeable under nitrogen limitation. While no significant morphological differences were found between cyst-like forms of Sp245 and SK586, the dormant mutant forms grown in a nitrogen-free medium were more sensitive to hydrogen peroxide than the dormant forms of Sp245. In Sp245 biofilms, viable forms resistant to drying were found even after 9 months of storage, while in SK586 biofilms, such bacteria remained viable for less than a month. In the course of adaptation to the plant root system, the number of Azospirillum cells resistant to drying increased, as compared to the number of such cells from mature biofilms on abiotic surfaces. Unlike Sp245, SK586 did not promote the development of the root system and the growth of the aerial parts of 10-day-old plants, possibly owing to its lower ability to colonize the roots of wheat seedlings incubated in a liquid medium. Alkyl hydroperoxide reductase is probably essential for better resistance of azospirilla to various stresses and for manifestation of their plant-growth-promoting activity.