Juan Iriberri

Temporal variability of attached and free-living bacteria in coastal waters

The temporal variability of the abundance and the incorporation of 3H-thymidine and 14C-glucose by attached and free-living bacteria, as well as their relation with environmental factors, were analyzed in a coastal marine ecosystem during a year. Both communities were quantitatively very different. Attached bacteria represented only 6.8% of the total bacterial abundance, whereas free-living bacteria represented 93.2%. The environmental factors most closely linked to the abundance and activity of free-living bacteria were temperature and the concentration of dissolved nutrients. Moreover, the free-living community showed similar temporal variations in abundance and in activity, with lower values in the cold months (from October to May). The attached community did not present the same pattern of variation as the free-living one. The abundance of the attached bacteria was mainly correlated to the concentration of particulate material, whereas their activity was correlated to temperature. We did not find a significant correlation between the abundance and the activity of the attached community. On the other hand, the activity per cell of the two communities did not present a clear temporal variation. Attached bacteria were more active than free-living ones in the incorporation of radiolabeled substrates on a per cell basis (five times more in the case of glucose incorporation and twice as active in thymidine incorporation). However, both communities showed similar specific growth rates. The results suggest that the two aquatic bacterial communities must not be considered as being independent of each other. There appears to be a dynamic equilibrium between the two communities, regulated by the concentrations of particulate matter and nutrients and by other environmental factors.

Ectoenzymatic Activity and Uptake of Monomers in Marine Bacterioplankton Described by a Biphasic Kinetic Model.

A bstractThe kinetics of bacterial hydrolytic ectoenzymatic activity and the uptake of monomeric compounds were investigated in the Northwestern Mediterranean Sea. Aminopeptidase and α- and β-glucosidase activities were analyzed by using fluorogenic substrates at 15–22 concentrations ranging from 1 nM to 500 μM. Radiolabeled glucose and a mixture of amino acids were chosen as representatives of monomeric compounds, and the bacterial uptake rates (assimilation plus respiration) were determined over a wide range of substrate concentrations (from 0.2 nM to 3 μM). We found biphasic kinetics both for hydrolytic enzymes and uptake systems: high affinity enzymes at low concentrations of substrates (Km values ranged from 48 nM to 2.7 μM for ectoenzymes and from 1.4 nM to 42 nM for uptake systems), and low affinity enzymes at high concentrations of substrates (Km values ranged from 18 μM to 142 μM for ectoenzymes and from 0.1 μM to 1.3 μM for uptake systems). Transition between high and low affinity enzymes was observed at 10 μM for aminopeptidase and from 1 μM to 25 μM for glucosidases, and it was more variable and less pronounced for the uptake of glucose (40 nM–0.28 μM) and amino acids (10 nM–0.16 μM). Results showed that the potential rates of hydrolysis and uptake are tightly coupled only if the high affinity hydrolytic ectoenzymes and the low affinity uptake systems are operating simultaneously.

Bacterial Colonization and Ectoenzymatic Activity in Phytoplankton-Derived Model Particles: Cleavage of Peptides and Uptake of Amino Acids

A bstractPhytoplankton-derived model particles were created in laboratory from a mixture of autoclaved diatom cultures. These particles were colonized by a marine bacterial community and incubated in rolling tanks in order to examine the relationship between aminopeptidase activity and leucine uptake. Bacteria inhabiting particles and ambient water were characterized for abundance, biovolume, aminopeptidase activity, leucine uptake, and growth rate. Particles were a less favorable habitat than ambient water for bacterial growth since growth rates of particle-attached bacteria were similar or even lower than those of free-living bacteria. During the first ∼100 h of the particle decomposition process, there were not statistically significant differences in the aminopeptidase activity:leucine uptake ratio between attached and free-living bacteria. From ∼100 h to ∼200 h, this ratio was higher for attached bacteria than for free-living bacteria. This indicates an uncoupling of aminopeptidase activity and leucine uptake. During this period, attached and free-living bacteria showed similar hydrolytic activities on a cell-specific basis. In the free-living bacterial community, variations in aminopeptidase activity per cell were associated with variations in leucine uptake per cell and growth rates. However, in the attached bacterial community, when leucine uptake and growth rates decreased, aminopeptidase activity remained constant. Thus, after ∼100 h, particle-attached bacteria were not taking advantage of their high aminopeptidase activity; consequently the hydrolysed amino acids were released into the ambient water, supporting the growth of free-living bacteria. These results demonstrate that over the particle decomposition process, the relationship between hydrolysis and uptake of the protein fraction shows different patterns of variation for attached and free-living bacterial communities. However, in our experiments, this uncoupling was not based on a hyperproduction of enzymes by attached bacteria, but on lower uptake rates when compared to the free-living bacteria.

Direct viable count of Gram-positive and Gram-negative bacteria using ciprofloxacin as inhibitor of cellular division

Abstract Following the method described by Kogure et al. (Can. J. Microbiol. 25 (1979) 415–420), we used ciprofloxacin, a quinolone which inhibits cell division of Gram-positive and Gram-negative bacteria, to quantify the number of viable bacteria in a series of cultures. As test bacteria we used three Escherichia coli strains, two of them sensitive to nalidixic acid and a third resistant to this inhibitor, and three Gram-positive bacteria: one rod ( Lactobacillus plantarum ) and two cocci ( Enterococcus faecalis and Micrococcus varians ). Ciprofloxacin is an efficient inhibitor of cell division in all six strains, although the effect on the morphology of the rods and cocci was clearly different. The percentage of viable bacteria was determined in the case of the rods by the increase in either length or biovolume and, for the cocci, by the increase in cell biovolume. Image analysis was required to detect variations in cell biovolume in the case of cocci and rods. For nalidixic acid-sensitive strains, it was possible to use both, as similar percentages of viable bacteria were detected with both inhibitors. The differences between the colony forming units (CFU) and direct viable (DVC) counts were much higher in the strains showing typical arrangements than in those strains without them. Finally, we found no significant differences in the number of viable bacteria in the natural population of the Butron river when we used either ciprofloxacin or nalidixic acid as inhibitors of cell division.

Factors affecting the survival of E. coli in a river

The relationships between physical, chemical and microbial characteristics of an aquatic ecosystem and the survival of E. coli have been studied. Two conditions of the ecosystem (warm and cold) are considered. T90 (time necessary for 90% of a bacterial population to die) in the warm situation shows an inverse exponential relationship with water temperature. Besides the direct relationship temperature-T90, there is an indirect effect of temperature upon T90 through the natural microflora of the water. The relationships between temperature and the heterotrophic population, and between the heterotrophic population and the bacterial consumers (P.F.U.), are exponential and linear, respectively.

Influence of light and natural microbiota of the Butrón river on E. coli survival

The survival of an E. coli strain in water samples from the Butrón river has been studied. The input of E. coli cells in the aquatic system breaks down the established balance among the components of the natural microbiota: E. coli becomes the object of the active protozoal predation whereas the autochtonous heterotrophic community become alternative preys. As a result of this new situation, the natural microbiota increases but returns to the initial values once the E. coli cells have been removed from the system. The effect of the temperature of incubation on the survival is exerted through the effect of this parameter on the predatory activity of the protozoa. Light has a lethal and direct action on the E. coli cells, the effect of this parameter is even superior to that of predation.

Channeling of bacterioplanktonic production toward phagotrophic flagellates and ciliates under different seasonal conditions in a river

The objective of this study was to analyze the flux of biomass through the communities of bacteria and phagotrophic protists in the cold and warm conditions occurring seasonally in Butrón River. Bacterial and heterotrophic protistan (flagellate and ciliate) abundance was determined by epifluorescence direct counts; protistan grazing on planktonic bacteria was measured from fluorescently labeled bacteria uptake rates; and the estimate of bacterial secondary production was obtained from [3H]thymidine incorporation rates. The abundance of bacterial, flagellate, and ciliate communities was similar during cold and warm situations. However, we observed that estimates of dynamic parameters, i.e., secondary bacterial production and protistan grazing, in both situations were noticeably different. In the warm situation, grazing rates of flagellates and ciliates (bacteria per protist per hour) were, respectively, 7 times and 18 times higher than those determined in the cold situation, and the grazing rates of the protistan communities (bacteria per protists present in 1 ml of water per hour) increased up to 5 times in the case of flagellates and 42 times in the case of ciliates. Estimates of bacterial secondary production were also higher during the warm situation, showing a ninefold increase. The percentage of bacterial production preyed upon by flagellates or ciliates was not significantly different between the two conditions. These results showed that in the different conditions of a system, the flux of biomass between the trophic levels may be quite different although this process may not be reflected in the abundance of each community of bacteria, flagellates, and ciliates.

Enumeration, Isolation and Some Physiological Properties of Actinomycetes from Sea Water and Sediment

Summary Nine selective culture media were used for counting actinomycetes in marine water and sediments. The presence of glycerol and/or asparagine in the media favoured the growth of these microorganims. Thermic treatment of sediment samples resulted in a selective reduction of the non-actinomycetal heterotrophic microflora. However, the same treatment of water samples had a negative effect on the actinomycetal flora. Actinomycetes in water and sediment represented 3–4% and 5–6% of the total flora, respectively. 72.7% of the isolated strains were Streptomyces, 20.5% Micromonospora and 7% Nocardia. The isolated actynomycetes strains, especially Streptomyces, showed a high hydrolytic activity for macromolecules and non-viable Gram-negative cells. Antimicrobial activity increased when sea water was present in the culture medium.

Influence of a Survival Process in a Freshwater System upon Plasmid Transfer Between Escherichia coli Strains

A bstractSurvival and potential ability to act as recipient or donor during the survival process for one plasmid-free and four plasmid-bearing Escherichia coli strains under nonilluminated and illuminated conditions in freshwater systems were studied. The five E. coli strains showed the same behavior with respect to the microbial parameters used to characterize the survival process (culturability and viability). Under nonilluminated conditions, recipient cells did not show variation in the ability to receive and express plasmid material, while the culturability of the recipient strain remained stable. Under the same conditions, donor cells lost their ability for plasmid transfer during the survival process, in all cases more than a 90% decrease of the number of transconjugants was found after 4 days of experimentation, although viable and culturable cells of donor strains maintained the capacity to express some plasmidic genes. Under illuminated conditions, transconjugants were not detected after 2 days of experimentation. The number of transconjugants formed was dependent not only on the time donor strains remained in the water but also on the temperature (20 or 37°C) at which the mating assays were conducted.

Grazing rates of diverse morphotypes of bacterivorous ciliates feeding on four allochthonous bacteria.

Aims: The permanence in aquatic systems of allochthonous bacteria coming from sewage effluents is a risk for public health. This work aimed to analyse the elimination of the bacteria Staphylococcus epidermidis, Enterococcus faecalis, Escherichia coli and Klebsiella pneumoniae by a riverine ciliate community.