The origin of recently growing irregular cave pisoids in Perlova Cave (Velka Fatra Mts, Slovakia) seems to be due to the activity of hydrogen-oxidizing bacteria. Several samples of water and freshly deposited calcite from cave pisoids were analysed for stable oxygen isotope ratios. The obtained values were checked using the O’Neil equation. Almost all the calcite samples display values more positive than their calculated counterparts. This phenomenon is ascribed to a fractionation process mediated by bacteria. The light isotope of oxygen is preferentially taken up by the hydrogen-oxidizing bacteria. It causes the relative enrichment in the heavy isotope in the bacterias’ surroundings, which is recorded in the calcite precipitated around the bacterial cells.