Estimating Bacterial Production in Marine Waters from the Simultaneous Incorporation of Thymidine and Leucine

Abstract

We examined the simultaneous incorporation of [3H]thymidine and ['4C]leucine to obtain two independent indices of bacterial production (DNA and protein syntheses) in a single incubation. Incorporation rates of leucine estimated by the dual-label method were generally higher than those obtained by the single-label method, but the differences were small (dual/single = 1.1 + 0.2 [mean ± standard deviation]) and were probably due to the presence of labeled leucyl-tRNA in the cold trichloroacetic acid-insoluble fraction. There were no significant differences in thymidine incorporation between dual-and single-label incubations (dual/ single = 1.03 ± 0.13). Addition of the two substrates in relatively large amounts (25 nM) did not apparently increase bacterial activity during short incubations (<5 h). With the dual-label method we found that thymidine and leucine incorporation rates covaried over depth profiles of the Chesapeake Bay. Estimates of bacterial production based on thymidine and leucine differed by less than 25%. Although the need for appropriate conversion factors has not been eliminated, the dual-label approach can be used to examine the variation in bacterial production while ensuring that the observed variation in incorporation rates is due to real changes in bacterial production rather than changes in conversion factors or introduction of other artifacts.

Microbial degradation of organic carbon and nitrogen on diatom aggregates

“…Bacterial production-Bacterial production on aggregates was determined directly after the respiration measurements. Bacterial production was measured by incorporation of [ 14 C] leucine ( 14 C-Leu; Kirchman et al 1985;Simon and Azam 1989) and bacterial growth rate by incorporation of [ 3 H] thymidine ( 3 H-TdR; Fuhrman and Azam 1980) into the ice-cold trichloroacetic acid precipitate using the dual label approach (Chin-Leo and Kirchman 1988). Both radiotracers ( 3 H-TdR [75 Ci mmol Ϫ1 ] and 14 C-Leu [312 mCi mmol Ϫ1 ], Amersham) were added to three samples and a formalin-killed control and were incubated at in situ temperature for 1 h in the dark.…”

Bacterial growth and grazing on diatom aggregates: Respiratory carbon turnover as a function of aggregate size and sinking velocity

“…Bacterial production rates-Bacterial production rates were measured using the microcentrifugation method (Smith and Azam 1992). Incorporation of [ 3 H]-thymidine ( 3 H-TdR; Fuhrman and Azam 1980) and [ 14 C]-leucine ( 14 C-Leu; Kirchman et al 1985, Simon andAzam 1989) into the ice-cold trichloroacetic acid precipitate was measured with a dual label approach (Chin-Leo and Kirchman 1988). 3 H-TdR (75 Ci mmol Ϫ1 ) and 14 C-Leu (312 mCi mmol Ϫ1 , both from Amersham) were added to samples and a formalin-killed control and were incubated at in situ temperature for 1 h in the dark.…”

Bacterial production and growth efficiencies: Direct measurements on riverine aggregates

“…Bacterial production rates-Bacterial production rates were measured directly after respiration had been determined on the same aggregate using the microcentrifugation method (Smith and Azam 1992 Kirchman 1988). 3 H-TdR (75 Ci mmol Ϫ1 ) and 14 C-Leu (312 mCi mmol Ϫ1 , both from Amersham) were added to samples and to a formalinkilled control and incubated at in situ temperature for 1 h in the dark.…”