Effect of culture conditions on ergosterol as an indicator of biomass in the aquatic hyphomycetes

Abstract

Ergosterol is a membrane component specific to fungi that can be used to estimate fungal biomass using appropriate factors of conversion. Our objectives were to determine the limits of use of ergosterol content as a measure of biomass for aquatic hyphomycetes, and to evaluate a previously established ergosterol-to-biomass conversion factor. We varied inoculum quality, growth medium, and degree of shaking of four aquatic hyphomycete species. In cultures inoculated with homogenized mycelium, we found a significant effect of shaking condition and culture age on ergosterol content. In liquid cultures with defined medium, ergosterol content reached 10 to 11 microg/mg of mycelium (dry mass) and varied by factors of 2.2 during exponential growth and 1.3 during stationary phase. The increase in ergosterol content during exponential phase could be attributed, at least in part, to rapid depletion of glucose. Oxygen availability to internal hyphae within the mycelial mass is also responsible for the differences found between culture conditions. Ergosterol concentration ranged from 0.8 to 1.6 microg/mg in static cultures inoculated with agar plugs. Ergosterol content varied by a factor of 4 in two media of different richnesses. For different combinations of these parameters, strong (r(2) = 0.83 to 0.98) and highly significant (P << 0.001) linear relationships between ergosterol and mycelial dry mass (up to 110 mg) were observed. Overall, the ergosterol content varied by a factor of 14 (0.8 to 11 mg/g). These results suggest that care must be taken when the ergosterol content is used to compare data generated in different field environments.

FIG. 1

Mycelial dry mass (A), ergosterol (B), and ergosterol content (C) of T. elegans in liquid cultures as a function of time. Cultures were inoculated with homogenized mycelium in BP(●), SC (□), and UC (▴) series. Each point is the mean of three replicates, and bars denote 1 standard deviation.

FIG. 2

Glucose concentration (A) and oxygen saturation (B) in liquid cultures as a function of time. For details, see the legend to Fig. 1.

FIG. 3

Ergosterol as a function of dry mass in liquid cultures. Data are from Fig. 1A and B. The regression line was calculated from the combined data of the three series: BP (●), SC (□), and UC (▴) (y = 9.3x − 62; r² = 0.92). The equations for the individual series were y = 10x − 91 with r² = 0.92 (BP), y = 8.8x − 47 with r² = 0.93 (SC), and y = 8.2x − 29 with r² = 0.89 (UC).

FIG. 4

Relationship between ergosterol amount and mycelial dry mass for T. marchalianum grown in MEB (open symbols) or GMS-VAA (closed symbols) inoculated with agar plugs. The equations were y = 0.81x − 3 with r² = 0.95 (MEB) and y = 3.3x − 9.8 with r² = 0.98 (GMS-VAA).

FIG. 5

Relationship between ergosterol amount and mycelium dry mass for three strains of aquatic hyphomycetes, A. acuminata (A), C. aquatica CLAQ 28–185 (B), and C. aquatica CLAQ 102–299 (C), grown in MEB inoculated with agar plugs. The equations were y = 1.2x − 0.44 with r² = 0.96 (A), y = 1.4x − 0.34 with r² = 0.89 (B), and y = 1.6x − 0.25 with r² = 0.83 (C).