A simple procedure for elimination of fungal contamination for enumeration of iron-oxidizing bacteria from bioleaching matrix of sewage sludge

Enumeration of iron-oxidizing bacteria from the bioleaching matrix of sewage sludge is always confronted by fungal contamination. The objective of the present study was to find a reliable and simple method to remove fungal growth and to shorten the incubation time for facilitating enumeration of the iron-oxidizers from complex sludge samples. The results demonstrated that filtering the sludge sample through sterile No. 5 Whatman filter paper before serial dilution was effective in eliminating the fungal growth on agarose media. Both the counts and the incubation time required for enumeration were highly dependent on the medium pH, with maximum counts at pH 2.5-2.75. Medium prepared at pH values outside of this range led to lower counts and a longer lag time for colony formation. However, the introduction of heterotrophic microorganisms into the solid medium did not show further improvement in enumeration efficiency and shortening of the incubation period. By incorporating the optimal conditions obtained, the incubation time could be reduced to 7 and 10 d for pure cultures and sludge samples, respectively.