Colonization and disinfection of biofilms hosting coliform-colonized carbon fines.

The documented release of carbon fines from granular activated carbon filters is a concern for drinking water utilities, since these particles may carry coliform and even pathogenic bacteria through the disinfection barrier. Such a breakthrough could have an impact on distribution system biofilms. Using total cell counts, specific monoclonal antibody staining, and computerized image analysis, we monitored the colonization of introduced Klebsiella pneumoniae associated with carbon fines in mixed-population biofilms. The particles transported the coliforms to the biofilms and allowed successful colonization. Chlorine (0.5 mg/liter) was then applied as a disinfectant. Most K. pneumoniae along with the carbon fines left the biofilm under these conditions. The impact of chlorine was greater on the coliform bacteria and carbon fines than on the general fixed bacterial population. However, 10% of the introduced coliforms and 20% of the fines remained in the biofilm. The possibility that this represents a mechanism for bacteria of public health concern to be involved in regrowth events is discussed.

Effect of temperature on growth, respiration, and nutrient regeneration by an omnivorous microflagellate.

The effect of temperature on the rates and extent of carbon and nitrogen cycling by the heterotrophic microflagellate Paraphysomonas imperforata (diameter, 7 to 12 mum) fed with the diatom Phaeodactylum tricornutum was investigated over an ecologically pertinent temperature range (14 to 26 degrees C). All physiological rates investigated increased with increasing temperature. Q(10) values were similar for all rate changes and were comparable to those which have been reported for other protozoa. In contrast to all rates, microflagellate gross growth efficiency and cell volume were unaffected by temperature. Decreases in the concentrations of particulate carbon and particulate nitrogen from grazed diatom cultures also were similar when summed over the entire growth phase of the microflagellate population. Therefore, the proportions of ingested carbon and nitrogen which were incorporated or remineralized by the microflagellate were independent of temperature between 14 and 26 degrees C. At temperatures above 18 degrees C, growth rates of P. imperforata were greater than the maximum growth rates reported for most phytoplankton. We conclude that the impact of P. imperforata on natural phytoplankton communities is not controlled by temperature above 18 degrees C but may be affected by the rate at which zooplankton or microzooplankton prey on the microflagellate, as well as the inability of the microflagellate to graze efficiently when phytoplankton are present at low cell densities.

Carbon dioxide exchange between air and seawater: no evidence for rate catalysis.

It has been suggested that enzymatic catalysis plays a major role in regulating the mass transport of carbon dioxide from the atmosphere into the oceans. Evidence for this mechanism was not found in a series of gas exchange experiments in which the gas transfer rate coefficients for samples obtained from various natural seawaters, with and without the addition of carbonic anhydrase, were compared with those from artificial seawater. Wind-induced turbulence appears to be the major factor controlling the ocean's response to anthropogenic increases in atmospheric carbon dioxide.

Free ammonia inhibition of algal photosynthesis in intensive cultures.

The effect of free NH(3) inhibition on short-term photosynthesis was investigated in three microalgal species: the freshwater chlorophyte Scenedesmus obliquus, the marine diatom Phaeodactylum tricornutum and the marine chlorophyte Dunaliella tertiolecta. By performing a series of assays at various concentrations of added NH(4)Cl and culture pH, we demonstrated that the inhibitory compound was free NH(3) and that pH played no role in determining the magnitude of inhibition, other than in establishing the degree of dissociation of nontoxic NH(4) to toxic NH(3). When corrections were made for pH, all three species displayed the same sigmoidal response curve to free NH(3) concentration; 1.2 mM NH(3) led to 50% reduction in photoassimilation of C. Based on literature values, some marine phytoplankton appear to be significantly more sensitive to free NH(3) than were the test species, which are noted for their excellent growth characteristics. However, the combination of low algal biomass and strong pH buffering commonly found in most marine and many freshwater environments probably limits the possibilities for NH(3) toxicity to low alkalinity freshwaters and intensive algal cultures in which NH(4) is the main source of N. Such conditions occur commonly in algal wastewater treatment systems.

Effect of nitrogen-mediated changes in alkalinity on pH control and CO(2) supply in intensive microalgal cultures.

The freshwater alga Scenedesmus obliquus was grown in continuous culture at a fixed dilution rate of 0.5/day, but at varying pH in the range 4.17-10.67. The pH was regulated in the range 4.17-7.67 by continuously bubbling 1% CO(2)-enriched air into the cultures and by varying the source of nitrogen (NO(3) (-), NH(4) (+), or urea) in the growth medium, which, in turn, led to changes in culture alkalinity. Culture alkalinity and P(CO(2) ) were the sole determinants of pH. A pH-stat system, together with NO(3) (-) in the medium, was used to regulate the pH in the range 7.92-10.67. Maximum productivity, which occurred at pH 6.6, was dependent on N source only to the extent that culture alkalinity was a function of nitrogen uptake. The results demonstrate that the choice of N is a critical factor in controlling the pH of large-scale algal cultures. NH(4) (+) is a poor source of N because it leads to destruction of culture alkalinity and concomitant growth-inhibiting reductions in pH, whereas NO(3) (-) has an opposite effect, although pH is not so severely affected in this case. Urea is, by far, the most suitable N source for maximizing algal yield when it is supplied in combination with the proper amounts of HCO(3) (-) alkalinity in the growth medium and percent CO(2) in the bubbled gas that will lead to an equilibrium pH near the optimum pH.

Inorganic carbon limitation and chemical composition of two freshwater green microalgae.

Two freshwater chlorophytes, Chlorella vulgaris and Scenedesmus obliquus, were grown in inorganic carbon-limited continuous cultures in which HCO(3) was the sole source of inorganic carbon. The response of the steady-state growth rate to the external total inorganic carbon concentration was reasonably well described by the Monod equation; however, the response to the internal nutrient concentration was only moderately well represented by the Droop equation when the internal carbon concentration was defined on a cellular basis. The Droop equation was totally inapplicable when total biomass (dry weight) was used to define internal carbon because the ratio of carbon to dry weight did not vary over the entire growth rate spectrum. In batch cultures, maximum growth rates were achieved at the CO(2) levels present in atmospheric air and at HCO(3) concentrations of 3 mM. No growth was observed at 100% CO(2). Both nitrogen uptake and chlorophyll synthesis were tightly coupled to carbon assimilation, as indicated by the constant C/N and C/chlorophyll ratios found at all growth rates. The main influence of inorganic carbon limitation appears to be not on the chemical structure of the biomass, but rather on cell size; higher steady-state growth rates lead to bigger cells.

Problems in estimating growth rates of marine phytoplankton from short-term(14)C assays.

Growth rate estimates (µ') of phytoplankton populations that were sampled from nitrogen-limited continuous cultures and then incubated for short durations in batch culture with added(14)C-HCO3 (-) were significantly different than steady-state growth rates (µ) for 3 of 5 marine phytoplankton species. Two diatoms,Thalassiosira weissflogii andChaetoceros simplex, displayed virtually identical growth rates (µ=µ') over a wide range ofµ, whereasµ' for a third diatom,Phaeodactylum tricornutum, was overestimated by an average of 40% compared toµ. In contrast,µ' was underestimated by the(14)C technique for the two remaining species: up to 40% at a steady-stateµ of 1.0 day(-1) for the chlorophyteDunaliella tertiolecta and up to 100% atµ of 1.4 day(-1) for the haptophytePavlova lutheri. For the latter two species the divergence betweenµ andµ' appeared to increase with increasing steady-stateµ. A simple model of labeled and total carbon flow between the aqueous phase and cellular biomass was constructed to demonstrate that respiration was negligible whenµ=µ', but was significant whenµ'>µ. In the cases in whichµ'<µ, a rapid physiological alteration presumably took place once the steady state was disturbed and cells were placed in the incubation chambers, which perhaps was related to the nutritional state of the cultures at the time of sampling. Questions thus are raised regarding our ability to measure accurately primary productivity from shipboard experiments with confined samples of phytoplankton from nutrient-impoverished waters that probably are less hardy than the laboratory cultures used in these studies.

Steady-State Growth and Chemical Composition of the Marine Chlorophyte Dunaliella tertiolecta in Nitrogen-Limited Continuous Cultures.

The marine chlorophyte Dunaliella tertiolecta was grown in continuous cultures under NH(4)-N, NO(2)-N, NO(3)-N, and urea-N limitations. The effect of the nitrogen cell quota (Q(n)) on the steady-state growth rate (mu) was the same regardless of the N source. The relationship between mu and Q(n) was well described by the Droop equation, but only up to the true maximum growth rate ;mu (= cell washout rate). The ratio between the minimum cell quota (k(Q)) and the maximum cell quota (Q(m)) was 0.19. Hence, there is no substitute for determining ;mu experimentally. That there was no difference in growth response to different N sources suggests that no internal pooling of inorganic nitrogen occurred. Both the carbon (Q(c)) and phosphorus (Q(p)) cell quotas under N limitation increased with increasing mu in a threshold fashion: virtually no change in either cell quota up to approximately 0.8 ;mu, followed by a rapid and large increase up to ;mu. In addition, in the region of low mu, there was an increase in Q(p) with a decreasing medium N/P ratio of between 15 and 5 (by atoms). The results generally indicate the physiological limits in cellular constituency under N limitation. The usefulness of this information, however, in describing the response of natural populations of marine phytoplankton to transient nutrient exposures on the temporal and spatial microscales that most likely exist is of limited value.

Nitrogenous nutrition of marine phytoplankton in nutrient-depleted waters.

Variability in the small-scale temporal and spatial patterns in nitrogenous nutrient supply, coupled with an enhanced uptake capability for nitrogenous nutrients induced by nitrogen limitation, make it possible for phytoplankton to maintain nearly maximum rates of growth at media nutrient concentrations that cannot be quantified with existing analytical techniques.

Temperature effects on steady-state growth, phosphorus uptake, and the chemical composition of a marine phytoplankter.

The marine chrysophyteMonochrysis lutheri was grown in phosphorus-limited continuous cultures at temperatures of 15°, 18.8° and 23°C. The effect of temperature on the maximum growth rate was well-defined by the Arrhenius equation, but the Q10 for this alga (1.7) was somewhat lower than has been determined previously for many other phytoplankton species (2.0-2.2). The minimum phosphorus cell quota was relatively unaffected by temperature at 18.8°C and 23°C, but doubled in magnitude at 15°C. As a result, the internal nutrient equation of Droop described the relationship between specific growth rate and phosphorus cell quota well at 18.8° and 23°C, but was less successful at 15°C. The major limitation in using the Droop equation is that the ratio between the minimum and maximum cell quotas must be known, thus necessitating the need to establish the true maximum growth rate by the cell washout technique. In addition, the phosphorus uptake rate on a cell basis at a given steady state growth rate (=specific uptake rate) increased dramatically at 15°C, whereas the turnover rate of total available phosphorus was unaffected by temperature. Both the nitrogen and carbon cell quotas were relatively unaffected by growth rate at a given temperature, but the average values increased slightly with decreasing temperature. The overall conclusion is that phytoplankton growth and limiting-nutrient uptake rates are only synchronous at or near the optimum temperature. Because these types of responses are species specific, much additional data on temperature effects will be required before the importance of including such effects in phytoplankton-nutrient models can be determined.

Inorganic nitrogen removal from wastewater: effect on phytoplankton growth in coastal marine waters.

Algal bioassays were used to demonstrate the high efficiency of a comnbined tertiary wastewater treatment and marine aquaculture system in removing inorganic nitrogen, and to show that the coastal waters off Woods Hole, Massachusetts, are limited in nitrogen for marine phytoplankton growth. When nutrients were removed from secondarily treated domestic wastewater through assimilation by phytoplankton in an outdoor growth pond, the pond effluent, in varying dilutions with seawater, could not support more phytoplankton growth than the seawater alone. However, when nitrogen was added back to the mixtures of pond effluent and seawater, the phytoplankton growth response was similar to that with a mixture of wastewater and seawater. This is similar to the findings of other researchers, and suggests that nitrogen may be the key growth-limiting nutrient in many coastal marine waters. The combined tertiary treatment-marine aquaculture system appears to be an effective means of removing nitrogen from secondarily treated wastewater and controlling eutrophication of coastal marine waters.