Activity of an Attached and Free-Living Vibrio sp. as Measured by Thymidine Incorporation, p -Iodonitrotetrazolium Reduction, and ATP/DNA Ratios

Abstract

Three independent techniques, [3H]thymidine incorporation, the reduction rate of p-iodonitrotetrazolium violet (INT) to INT formazan normalized to DNA, and the ratio of ATP to DNA, were adapted to measure the activity of attached and unattached estuarine bacteria. In experiments employing the estuarine isolate Vibrio proteolytica, nutrient concentrations were manipulated by varying the concentration of peptone-yeast extract. In the presence of exogenous nutrients, the activity of free-living cells was greater than that of attached cells as measured by [3H]thymidine incorporation and ATP/DNA ratios. In the absence of peptone-yeast extract, however, the activity of attached cells surpassed that of free-living cells as determined by [3H]thymidine incorporation and INT formazan normalized to DNA. Of the three techniques, [3H]thymidine incorporation was deemed most sensitive for detecting changes in activity resulting from slight differences in nutrient concentration. By this technique, attached cells were much less sensitive to changing nutrient concentrations than were free-living cells. Below a threshold concentration, attached cell activity remained constant, while the activity of unattached cells decreased as a function of decreasing nutrient concentration. The results suggest that loss of cell surface area available for substrate uptake due to attachment may be an important factor in determining the relative activities of attached and free-living cells.

Comparison of biodegradation kinetic parameters for naphthalene in batch and sand column systems by pseudomonas putida

“…Because the ko, km, and kc o 2 values re p resent a series of mass-transfer and reaction processes occurring within the cell membrane, the dependence on surf a c e a rea likely has a mechanistic basis. Activity decreases with attached bacteria have also been observed in batch assays by other workers [5,25]. Jeffery and Paul [5] found that the thymidine incorporation rate and the ATP/DNA ratio of free suspended cells were greater than those of attached cells.…”

“…Activity decreases with attached bacteria have also been observed in batch assays by other workers [5,25]. Jeffery and Paul [5] found that the thymidine incorporation rate and the ATP/DNA ratio of free suspended cells were greater than those of attached cells. They ascribed the reduced rates to physiological diff e rences between the two systems, primarily a re d u ction in the amount of cell surface available for nutrient uptake.…”

“…did not find conclusive evidence that the adhesion of cells to solid surfaces directly influences bacterial metabolism. They concluded that the differences were l a rgely indirect, resulting from decreases in the " a p p a rent" substrate affinity because of a reduction in the cell surface exposed to the bulk solution, and an i n c reased mass-transfer resistance to the non-exposed surface [2][3][4][5][6][7][8][9][10][11]. It should be noted that all of these studies were conducted in well-mixed systems, and may not be applicable to degradation in flow thro u g h porous media.…”

Responses of Bacterial Assemblages on Standing‐Decaying Blades of Smooth Cordgrass to Additions of Water and Nitrogen

“…Note that MUSCHENHEIM et al (1989) found evidence that bacteria attached to particles from smooth-cordgrass shoots moved to the water column in laboratory incubations. Cells having just left the surface of decomposing leaves, where both leaf lysates and algal exudates (FALLON et al, 1985) could be available, might reasonably be expected to be more productive (thymidine incorporation per cell) than most of those in the bacterioplankton (see, e.g., JEFFREY and PAUL, 1986;TURLEY, 1993;AZAM et af., 1994;HEISSENBERGER et al, 1996;SCHUT et al, 1997). Cell emigration might also be involved in the explanation of changes in bacterial morphotypes during tidal excursions onto the marsh (NEWELL and KRAMBECK, 1995).…”

DNA:ATP RATIOS IN MARINE MICROALGAE AND BACTERIA: IMPLICATIONS FOR GROWTH RATE ESTIMATES BASED ON RATES OF DNA SYNTHESIS1

“…For many years, for example, it was customary to measure ATP in the firefly bioluminescence assay by integrating light emission for 1 min or more (Holm-Hansen and Booth 1966), often beginning 5-15 s after the mixing of reagents (e.g. Jeffrey and Paul 1986) . Karl (1978), however, showed that this procedure can overestimate ATP concentrations by more than a factor of 2 due to the presence of non-adenine nucleotide triphosphates in microorganisms.…”