Phenotypic Variability in Azospirillum brasilense Strains Sp7 and Sp245: Association with Dormancy and Characteristics of the Variants

The state of metabolic dormancy in diazotrophic bacteria Azospirillum brasilense Sp7 (non-endophytic strain) and Sp245 (endophytic strain) was found to be associated with phenotypic variability. The latter manifested itself in the extension of the spectrum of A. brasilense phenotypic variants resulting from plating of cyst-like resting cells (CRC) on solid media and was more pronounced in strain Sp7. The major colony's morphological variants of strain Sp7 were (1) the dominant S type; (2) the highly pigmented Pg type; (3) the R type; (4) the Sin type, forming small colonies; and (5) the Sg type, forming segmented colonies. In addition to their colony morphology, the variants differed in the phenotype stability during transfers on the standard solid medium and in their motility in semisolid agar. The occurrence frequency of the phenotypic variants depended on the conditions and duration of incubation (storage) of the CRC of strain Sp7, as well as on heat treatment (at 55 and 60 degrees C for 10 min) of the cells prior to inoculation. The maximum frequency of S -> Pg transitions (up to 74%) was observed during the germination of CRC stored in a spent culture medium at -20 degrees C for 4 months; the maximum frequency (up to 100%) of S -> Sin transitions was observed after inoculation of the CRC subjected to heat treatment. The Pg variants were the most stable, whereas other types reverted rapidly to the S or Pg variant. The S variant grown in semisolid agar exhibited the mixed type of motility (Swa(+)Gri(+), swarming and migration in the form of microcolonies); the Pg and Sg variants showed the Swa(+)Gri(-)(swarming) phenotype and the Sm variant was nonmotile (Swa(-)Gri(-) phenotype). The spectrum of phenotypic variants of the endophytic strain Sp245 was narrower than that of strain Sp7 and was represented by S, Sin, and M (mucoid) variants that differed in the patterns of cell motility: the dominant S type displayed the swarming pattern (Swa(+)Gri(-)), the mucoid M type showed the mixed type (Swa(+)Gri(+)) of motility, and the Sm variant was nonmotile. The differences between the nonendophytic strain Sp7 and the endophytic strain Sp245 in their capacity for phenotypic dissociation and cell motility in semisolid media may reflect their ability to adapt to changing ambient conditions and specificity of plant-microbial interactions.