A comparison between biostimulation and bioaugmentation in a solid treatment of anaerobic sludge: Drug content and microbial evaluation.

Emerging pollutants can reach the environment through the sludge of Wastewater Treatment Plants. In this work, the use of Trametes versicolor in biopiles at lab-scale was studied, evaluating its capacity to remove the most hydrophobic Pharmaceuticals and assessing the evolution of the biopiles microbial communities. The total removal of drugs at real concentrations from sewage sludge was assessed for non-inoculated and fungal inoculated biopiles, testing if the re-inoculation of the biopiles after 22 days of treatment would improve the removal yields. It was found that 2 out of the 15 initially detected pharmaceuticals were totally degraded after 22 days, and re-inoculated fungal biopiles achieved higher removal rates than non-re-inoculated fungal biopiles for single compounds and for all the drugs simultaneously: 66.45% and 49.18% re-inoculated and non-re-inoculated biopiles, respectively. Finally, the study of the bacterial and fungal communities revealed that fungal inoculated and non-inoculated biopiles evolved to similar communities adapted to the presence of those drugs.

Pharmaceuticals removal and microbial community assessment in a continuous fungal treatment of non-sterile real hospital wastewater after a coagulation-flocculation pretreatment.

Hospital wastewaters are a main source of pharmaceutical active compounds, which are usually highly recalcitrant and can accumulate in surface and groundwater bodies. Fungal treatments can remove these contaminants prior to discharge, but real wastewater poses a problem to fungal survival due to bacterial competition. This study successfully treated real non-spiked, non-sterile wastewater in a continuous fungal fluidized bed bioreactor coupled to a coagulation-flocculation pretreatment for 56 days. A control bioreactor without the fungus was also operated and the results were compared. A denaturing gradient gel electrophoresis (DGGE) and sequencing approach was used to study the microbial community arisen in both reactors and as a result some bacterial degraders are proposed. The fungal operation successfully removed analgesics and anti-inflammatories, and even the most recalcitrant pharmaceutical families such as antibiotics and psychiatric drugs.

Stable Carbon Isotope Fractionation During 1,2-Dichloropropane-to-Propene Transformation by an Enrichment Culture Containing Dehalogenimonas Strains and a dcpA Gene.

A stable enrichment culture derived from Besòs river estuary sediments stoichiometrically dechlorinated 1,2-dichloropropane (1,2-DCP) to propene. Sequential transfers in defined anaerobic medium with the inhibitor bromoethanesulfonate produced a sediment-free culture dechlorinating 1,2-DCP in the absence of methanogenesis. Application of previously published genus-specific primers targeting 16S rRNA gene sequences revealed the presence of a Dehalogenimonas strain, and no amplification was obtained with Dehalococcoides-specific primers. The partial sequence of the 16S rRNA amplicon was 100% identical with Dehalogenimonas alkenigignens strain IP3-3. Also, dcpA, a gene described to encode a corrinoid-containing 1,2-DCP reductive dehalogenase was detected. Resistance of the dehalogenating activity to vancomycin, exclusive conversion of vicinally chlorinated alkanes, and tolerance to short-term oxygen exposure is consistent with the hypothesis that a Dehalogenimonas strain is responsible for 1,2-DCP conversion in the culture. Quantitative PCR showed a positive correlation between the number of Dehalogenimonas 16S rRNA genes copies in the culture and consumption of 1,2-DCP. Compound specific isotope analysis revealed that the Dehalogenimonas-catalyzed carbon isotopic fractionation (εC(bulk)) of the 1,2-DCP-to-propene reaction was -15.0 ± 0.7‰ under both methanogenic and nonmethanogenic conditions. This study demonstrates that carbon isotope fractionation is a valuable approach for monitoring in situ 1,2-DCP reductive dechlorination by Dehalogenimonas strains.

Morphological and ultrastructural characterization of an unusual purple sulfur bacterium from a marine microbial-mat community.

An unusual purple sulfur bacterium present in the pink layer of the Ebro Delta microbial mats has been identified through the study of its ultrastructural features. As pure cultures of this bacterium have not been obtained, due to its inability to grow in axenic conditions, only enrichment cultures where it reached at least 90% of total biomass have been considered. In enrichment cultures, the cells are rods that are 5.4+/-0.6 microm wide and 11.0+/-2.1 microm long. The color of the cell suspensions is pink to pinkish-red. Cells are motile by means of a polar tuft of flagella and multiply by binary fission. This bacterium possesses an extensive internal photosynthetic membrane system consisting of stacks of lamellae, contains bacteriochlorophyll a and carotenoids of normal spirilloxanthin series and deposits sulfur intracellularly. In natural samples, the most abundant phototrophic purple sulfur bacterium developing in this ecosystem has the same kind of intracytoplasmic membrane system, but the cells differ slightly in size and arrangement, in that they are ovoid, 4.2+/-0.5 microm wide and 6.0+/-1.4 microm long, and can be seen forming irregular non-motile clumps which are embedded in slime. Differences observed between field samples and enrichment cultures suggest that environmental conditions may bring about changes in the phenotypic characteristics of the cells. The morphological characteristics of the described bacterium resemble those of large Chromatia. However, this bacterium differs from known species in this group, particularly in terms of its photosynthetic membrane system and in its light absorption properties.

The biomass dynamics of cyanobacteria in an annual cycle determined by confocal laser scanning microscopy.

We recently published a method for estimating cyanobacteria biomass in delta microbial mats from the Ebro river using confocal laser scanning microscopy (CLSM). The present paper uses this method for identifying different groups of cyanobacteria and for determining their biomass dynamics. Microcoleus chthonoplastes and the Lyngbya-Oscillatoria were the most important contributors to the cyanobacterial biomass throughout the study. Biomass values ranged from 1.29 to 6.55 mgC/cm2 sediment for Microcoleus chthonoplastes, and from 128 microgC/cm2 to 3.16 mgC/cm2 sediment for Lyngbya-Oscillatoria. This technique is useful for determining biomass and for studying filamentous cyanobacteria as well as ramified eukaryotic cells. Confocal serial sections through the samples can be obtained. Two-dimensional images from the samples can be used to calculate the biomass of individual cells.

Molecular taxonomy of the genus Chlorobium.

This review summarizes molecular studies on the taxonomy of the genus Chlorobium. Furthermore, we try to introduce a comprehensive view of how joining traditional phenotypic-based taxonomy with the current molecular approaches leads to a more natural classification. Moreover, we introduce the current insights that molecular techniques have brought to the detection and identification of nonisolated Chlorobium strains in their environments.

Confocal laser scanning microscopy as a tool to determine cyanobacteria biomass in microbial mats.

We have developed a method based on confocal laser scanning microscopy for detection and quantification of cyanobacteria from the Ebro Delta microbial mats. Cyanobacteria play a major role as primary producers in microbial mats; it is difficult, however, to apply classical methods to estimate their biomass because they establish strong interactions with detritic particles. The protocol described here allows the localization of individual cells or filaments with micrometre precision without the need to either manipulate or stain the samples. This method is suitable for studying biomass 'in situ' in microbial mats.

Bacterial symbioses. Predation and mutually beneficial associations.

The endosymbiotic theory, which has proved to explain the origin of mitochondria and chloroplasts, also posits the origin of nucleus and other cellular organelles that could have derived from ancient relationships among bacteria. It seems that predation might have been a prerequisite to the establishment of symbiosis as a source of evolutionary novelty. This review describes current different examples of bacteria able not only to attack and degrade other bacteria, but also to establish stable symbiotic relationships with different eukaryotic organisms.

A mathematical model to determine the optimal number of fragments for comparison of bacterial chromosomic macrorestriction patterns.

To our knowledge, although comparison of chromosomic macrorestriction patterns has become one of the most feasible molecular tools of the current microbial taxonomy, a mathematical approach to optimize the choice of a restriction enzyme among the endonucleases tested for such comparison has not been previously described. The coincidence of restriction patterns for two tested bacterial strains with this chosen endonuclease will ensure a high genetic relatedness between them. We report a mathematical model to determine the probability of hazardously obtaining a particular chromosomic macrorestriction pattern by PFGE and to calculate the optimal number of fragments for its comparison. The model presented allows us to determine the optimal number of fragments in order to compare chromosomic restriction patterns. The model calculates this values as a function of the chromosome size and the restriction site length. The model is not useful for choosing a restriction enzyme previous to experimental steps, but as a tool for the choice of the restriction enzyme that yields the lowest probability of hazardously obtaining coincidences of chromosomic patterns. The applicability of this model has been exemplified by determining the optimal number of fragments for some well-characterized bacteria and by comparing these values with those that have been experimentally used.

Cellular content of storage inclusions in purple sulfur bacteria determined by ultrathin sections.

Phototrophic sulfur bacteria accumulate storage inclusions as a mechanism to adapt to several types of environmental stress. We compare the content of elemental sulfur and poly-beta-hydroxybutyrate (PHB) found in cultures growing under laboratory conditions with the content found in microorganisms in natural environments. Since natural communities are extremely heterogeneous in composition (they do not contain only phototrophic sulfur bacteria) and physiological state, it was not possible to apply conventional chemical analyses for the determination of the contents of sulfur and PHB. The study was performed by means of transmission electron microscopy, which turned out to be an excellent tool for this purpose. The results indicate that, in natural environments, cells have an extremely high content of storage inclusions, much higher than their laboratory grown counterparts, probably as a consequence of less favorable environmental conditions.

Genomic heterogeneity in Chlorobium limicola: chromosomic and plasmidic differences among strains.

Chromosome analysis by pulsed-field gel electrophoresis shows a high level of genetic heterogeneity between the two subspecies of the green sulfur bacteria Chlorobium limicola analysed: C. limicola and C. limicola f. s. thiosulfatophilum. Currently, they are differentiated only by the ability to utilize thiosulfate as photosynthetic electron donor, and by their %G + C content (51% and 58.1%, respectively). However, the capacity to utilize thiosulfate as photosynthetic electron donor does not appear to be a useful criterion to differentiate between some strains of this species, because this ability is encoded by plasmids that are different depending on the thiosulfatophilum strain analysed. In contrast, this study reveals that the comparison of chromosomal restriction patterns is very useful as an additional aid for the differentiation and identification of C. limicola strains.

Analysis of bacterial genomes by pulsed field gel electrophoresis.

The present article reviews the main achievements reached in bacterial genetics through the application of pulsed field gel electrophoresis. In particular, it summarizes the main subjects where the use of this technique has significant relevance for the analysis of bacterial genomes. With this purpose, it sums up some of the most relevant examples which illustrate the applicability of this technique in bacterial taxonomy, in epidemiological work and in the study of bacterial genome organization (including the assessment of the number of genomic elements and the size and topology of all of them). This technique, which makes it possible to make genomic comparisons in an easy, fast and repetitive way, has been demonstrated to be especially useful for those bacteria where the application of classical genetic techniques has not been possible.

Induced mutagenesis by bleomycin in the purple sulfur bacterium Thiocapsa roseopersicina.

Cell death and mutagenesis in bleomycin-treated cells of Thiocapsa roseopersicina (a purple sulfur bacterium) was studied by cultivation in a semisolid medium (agar-shake technique). This technique has also proven useful in assessing the frequency of antibiotic mutations by detecting and counting individual colonies of Thiocapsa roseopersicina. The frequencies of spontaneous mutants resistant to ampicillin, rifampicin, cloramphenicol, tetracycline, kanamycin, streptomycin, and neomycin were also studied: they ranged between 2 x 10(-9) and 9 x 10(-8). Bleomycin (4 micrograms/ml) sharply increased the frequency of ampicillin-resistant mutants, from 10(-8) (spontaneous) to 4 x 10(-4) (induced), in 17 h. An inducible, error-prone mechanism of DNA synthesis seems to be responsible for this enhancement of the mutagenic effect. this is the first report on the sensitivity to several antibiotics, and capacity of lethality and mutagenesis by bleomycin has been studied in a purple sulfur bacterium.

Transformation of Chlorobium limicola by a plasmid that confers the ability to utilize thiosulfate.

A modified transformation method for Chlorobium limicola confirms the role of an endogenous plasmid in thiosulfate metabolism. The plasmid was present in two forma specialis thiosulfatophilum (Tio+) strains and absent in one non-thiosulfate-utilizing (Tio-) strain. The plasmid (size, 14 kb) was transferred by transformation from Tio- to Tio+. The chromosomal restriction patterns, analyzed by pulsed-field gel electrophoresis, were used to distinguish between the different strains.

Distribution of predatory bacteria that attack chromatiaceae in a sulfurous lake.

Predatory bacteria that attack Chromatiaceae (purple sulfur bacteria) recovered from Lake Cisó (a mostly anaerobic holomictic lake) have been studied over two annual cycles. During the mixing period the lake was completely anaerobic; both predator and prey populations were found along the water column, and even at the surface. Throughout the stratification period maximum Chromatiaceae occurred between a depth of 1 and 3 m depth. The maximum numbers of predators and prey (Chromatiaceae) also occurred in this range.A collapse took place in the lake during the second annual cycle in 1986. It brought about changes in the physicochemical parameters of the lake, thus altering the population dynamics. Nevertheless, during both cycles the number of predatory bacteria was maximum immediately below the depth at which the maximum number of prey bacteria occurred.

Ciliates from a fresh water sulfuretum.

Ciliates were collected from a freshwater sulfuretum, Lake Cisó, which is part of a gypsum karstic area whose main feature is Lake Banyoles (Girona, Spain). Chromatium, Lamprocystis and Chlorobium are the major phototrophic sulfur bacteria in Lake Cisó. Blooms of a photosynthetic cryptomonad (up to 5 X 10(5) ind ml-1) were found at the metalimnion. The community of ciliates could be divided in three groups: aerobic, cosmopolitan, genera such as Stentor and Vorticella, in the epilimnion; a large population (up to 10(4) ind ml-1) of Coleps, adapted to low concentrations of both oxygen and sulfide, together with a few individuals of the equally sulfide-tolerant genus Paramecium, in the metalimnion, and anaerobic, true sulfide-loving genera such as Plagiopyla and Metopus, in the hypolimnion, where sulfide concentration was between 0.6 and 1.2 mM.