DNA Synthesis as a Measure of Bacterial Productivity in Seagrass Sediments

Abstract

B a c t e r~a l growth rates In sedlments have been calculated fiom measurements of the rate of lncolporatlon of trltlated thymldlne Into d e o x y r~b o n u c l e~c acld (DNA) T h e dllutlon of Isotope In DNA w a s used to estlmate the s u m of the pools of thymldlne In the sedlment and of other cellular precursors of t h y m~d~n eIn DNA Growth rates of bacterla In the surface zone of seagrass bed sedlrnents v a r~e d from 3 7 X 10' cell d l v~s~o n s h -' g -' dry weight of sediment on a hot autumn day to 3 3 X 10" cell d l v l s~o n s h -' g ' In w~n t e r By c o m b~n~n g growth rate measurements uslng the ~s o t o p e d~l u t~o n procedure with biomass measurements ~t IS now p o s s~b l e to obtaln redsondbly r e l~a b l e estimates of b a c t e r~a l product~vity In sedlments Seagrass beds are highly productive plant communltles, in which much of the primary production 1s not u t~l i z e d directly by animals, but enters h~g h e r trophlc levels through microorganisms, especially bacteria ( P h i l l~p s and McRoy, 1980) To quantify this process we need to know not only the blomass of the bacterial population, but also its growth rate Methods are available for measuring biomass in these sedim e n t~, w h~c h show that bacteria in seagrass beds of Moreton Bay Queensland constitute dbout 20 O/o of the sedlment organic matter excluding that which seagrass roots contribute ( M o r l a~t y ,1980) The measurement of growth rates of the whole population has not been possible by classical microb~ological techniques By measuring the rate of synthesis of d e o x y r~b o n u c l e~c acid (DNA), we hoped to be able to estimate the growth rate of bactena In the sedlment Bacteria In sedlments take up [ m e t h~l -~H ] thymidine (thymine-2-deoxyribose Tdr) and use lt for DNA synthesis (Tobln a n d Anthony, 1978) Fuhrman a n d Azam (1980) have used the rate of incorporation of Tdr into DNA in seawater to e s t~m a t e growth rate of planktonic bacterm They assumed that by adding a large excess of ~s o t o p e , the contribution of Tdr from other pools or pathways would be negligible In fact this may not be the case as Rosenbaum-Oliver and Zamenhof (1972) found that exogenous Tdr contributed only a portlon of the Tdr in DNA, varylng from 35 % to 63 % In a normal strain of Escherichia coli, depending on the culture conditions. The highest value of 63 % of exogenous Tdr was obtalned with l mg Tdr m l -' of culture medium. If natural populations of bacteria behave similarly, then to estlmate the rate of DNA synthesis, a n d thus obtain the rate of bacterial division, it is necessary to measure the dilution of a d d e d Tdr by pools in the cells a n d in the sediment. Thymidine-5'-triphosphate (dTTP), the final precursor in DNA synthesis, is synthesised only partly from exogenous Tdr and partly via other pathways within the cell (Rosenbaum-Oliver and Zamenhof, 1972). An isotope dilution experiment can be used to d e t e r m~n e the effect of added 'cold' precursor on the amount of labelled precursor incorporated into ...

Extracellular Proteolytic Activity in the Surface Sediment of a Eutrophic Inlet.

“…We emphasize, however, that at present there is no standard method to estimate the activities in sediment. Researchers for instance have measured the fluxes of oxygen and carbon dioxide 14) , the rates of nucleic acid synthesis 35,47) , and the extracellular hydrolytic activities 28,40) , to evaluate the heterotrophic processes in the sediment. Therefore, any comparisons with results from studies using other methods or model substrates for instance should be made with caution.…”

Mesure de la production bactérienne par incorporation de thymidine tritiée

“…FUHRMAN et AZAM (1982) en comparant les résultats de leurs mesures d'incorporation de thymidine à celles obtenues par incorporation de H 32 PO^ dans le DNA estiment la dilution isotopique à un facteur 3-7 pour des bactéries marines. MORIARTY et POLLARD (1981) proposent une approche cinétique (effet de l'addition de précurseur froid sur le taux d'incorporation dans le DNA) pour la détermination des pools internes de thymidine. MORIARTY et POLLARD (1982), POLLARD et MORIARTY (1984), BELL (1986) ont souligné la variabilité des résultats obtenus par cette méthode et la nécessité de déterminer ces pools lors de chaque mesure.…”

“…Ainsi cette méthode, moyennant quelques modifications expérimentales, est également utilisée pour la mesure de l'activité des bactéries dans les sédiments (MORIARTY et POLLARD, 1981, 1982 fixées à des supports solides (SERVAIS et al, soumis) ou formant des biofilms (MURREY et al, 1986) . Le marquage des bactéries par la H-thymidine a également été utilisé pour l'étude du broutage de celles-ci par le microzooplancton (HOLLIBAUGH et al, 1980) et l'étude des processus de mortalité bacté-rienne (SERVAIS et al 3 1985).…”

A critical examination of substoichiometric isotope dilution analysis using thymidine and leucine

“…In the method presented by Moriarty and Pollard (1981), the potential radioactivity incorporated (C max ) and thus the total thymidine incorporation during the incubation period (TTI) is then calculated:…”

“…In this form, the equation is analysed by linear regression (Moriarty and Pollard, 1981;Findlay et al, 1984;Bell, 1986). The X intercept, calculated as the ratio of (Y intercept / slope), equals -(L + P).…”