Structural alterations in non-stoichiometric magnetite induced by microorganisms are studied by nuclear methods. Magnetite samples were exposed to fungal strain known for its immobilization capabilities. Neutron activation analysis identified iron as the dominant element. The presence of maghemite in non-stoichiometric magnetite complicates the determination of iron sites by Mössbauer spectrometry. Mössbauer spectra recorded at 4.2 K display overlapping lines corresponding to tetrahedral and octahedral iron sites. Application of external magnetic field of 6 T improved the spectral resolution and revealed a presence of distinct sextets. Interaction with Aspergillus niger did not notably affect magnetite´s magnetic properties, indicating its stability.