Determination of soil fungal biomass from soil ergosterol analyses

Determination of fungal biomass (FB) and FB-carbon (FB-C) from soil ergosterol concentration is difficult because of unknown ergosterol-to-fungal biomass (E-to-FB) conversion factors and inefficient ergosterol extraction methods. We applied a microwave-assisted extraction (MAE) and high performance liquid chromatographic (HPLC) procedure to measure ergosterol in soil samples. The E-to-FB conversion factors were determined in six species of fungi grown in vitro. The MAE method was fast and extracted up to nine times more soil ergosterol than a classical refluxing saponification method. Soil ergosterol was separated and quantified rapidly (< 10 min) by HPLC. Alternaria alternata, Chaetomium globosum, Fusarium oxysporum, Penicillum chrysogenum, Rhizopus stolonifer and Trichoderma harzianum isolated from soil and plant matrices were grown in batch. Ergosterol and biomass content were determined in mycelia harvested during the stationary and exponential phases of growth. Total mycelial ergosterol ranged from 180 to 2178 microgram, and total dry biomass ranged from 17 to 595 mg. Total ergosterol and fungal dry biomass were strongly associated (r2 = 0.95). The C content in mycelial mats averaged 43% (+/- 1.1; SD), and was similar among fungal species and growth phases. The analyses of variance showed that the E-to-FB ratio was similar among fungal species or growth phase. An average ergosterol concentration of 4 microgram mg-1 dry biomass was determined for the six species of fungi, which gave a conversion factor of 250 microgram dry biomass microgram-1 ergosterol. The MAE method recovered an average of 62% (+/- 11%, SD) of the ergosterol added in mycelial mats to soils prior to extraction, and its recovery was independent of soil properties. The E-to-FB ratio and percent recovery of mycelial ergosterol helped establish for the first time relationships determining soil FB and FB-C from soil ergosterol concentration. The amount of FB ranged from 155 to 4745 microgram g-1 and that for FB-C ranged from 67 to 2040 microgram g-1 for different soils, and was higher in samples taken from native undisturbed land than in samples taken from adjacent cultivated fields. Measurement of soil ergosterol concentration is a useful estimate content of the living soil FB.