Extraction of microorganisms from soil: evaluation of the efficiency by counting methods and activity measurements

Different procedures for the release of microorganisms from soils were tested. The extraction efficiency was evaluated by combining biochemical activities (fluorescein diacetate hydrolysis, dehydrogenase activity and dimethylsulfoxide reduction) and cell counting. In contrast to the counting methods these activities have the advantage that there is no interference from the soil matrix or from the clay and silt in the extracts. We determined the distribution of activities and cell counts among the successive extracts and the soil residue of a three-step and a five-step extraction procedure. Centrifugation at 600×g, used in order to separate soil particles, causes a significant proportion of the bacteria and nearly all fungi to enter the sediment. The influence of relative centrifugal forces <600×g on the recording of microorganisms was investigated in detail. g-Values≥200 lead to unacceptable losses of microorganisms. Because the satisfactory separation of microorganisms and clay or silt is not possible, a compromise between removing the soil particles and quantifying the microorganisms is necessary. We recommend the one-step extraction of the soil by shaking and ultrasonication with subsequent centrifugation at ≤100×g. This results in extracts containing about 50% of the biochemical activities and 45% of the cell counts of the soils. This method thus corresponds in its extraction efficiency to a conventional three-step extraction, but is much simpler to carry out.