The community structure and potential activities of nitrifying and denitrifying bacteria were studied in the rhizosphere of Typha latifolia and Phragmites australis present in a free water system constructed wetland (CW). Potential nitrate reduction and nitrification activities were shown to be significantly higher in the rhizosphere when compared with the nonvegetated sediment. Higher rates were generally obtained for P. australis. The community structure of denitrifying bacteria in the rhizosphere differed from that found at the bulk sediment, as revealed by PCR-denaturing gradient gel electrophoresis (DGGE) of the nitrous oxide reductase encoding gene nosZ. Results also show a greater nosZ genotype diversification and suggest a plant species effect in rhizosphere samples obtained during events of low hydraulic retention times. Ammonia-oxidizing communities were less complex on the basis of PCR-DGGE analysis of the 16S rRNA gene. Retrieved sequences were all related to Nitrosomonas marina and Nitrosomonas ureae, being both present in rhizosphere and bulk sediment regardless of environmental changes. The results demonstrate the effect of vegetation on the functioning and structure of bacterial communities involved in the removal of nitrogen in the treatment cells of a CW and point to the use of vegetation coverage to promote nitrification or denitrification in particular areas.

d Ammonia-oxidizing bacteria (AOB) and archaea (AOA) were quantified in the sediments and roots of dominant macrophytes in eight neutral to alkaline coastal wetlands. The AOA dominated in most samples, but the bacterial-to-archaeal amoA gene ratios increased with increasing ammonium levels and pH in the sediments. For all plant species, the ratios increased on the root surface relative to the adjacent bulk sediment. This suggests that root surfaces in these environments provide conditions favoring enrichment of AOB. N itrification is a critical process for nitrogen retention in coastal wetlands, which are important transition zones between marine and terrestrial environments. Nitrification rates are commonly higher in the rhizosphere of macrophytes than in the bulk sediment (16,24), suggesting that nitrifiers are stimulated by rhizosphere conditions of wetland plants. Ammonia oxidation, the first and rate-limiting step in nitrification, is performed by ammonia-oxidizing bacteria (AOB) and archaea (AOA). For freshwater macrophytes, plant-specific differences in the composition and abundance of AOB and AOA have been described both in the rhizosphere (11, 12) and on the epiphyton of submerged shoots (4, 9). The AOA generally outcompete AOB at low ammonia concentrations (10,13,14,26,29,30). Also, root exudates may play a role in the relative distribution of AOB and AOA, even though ammonia oxidation is mainly considered an autotrophic process, as mixotrophy at the expense of pyruvate has been proven for a soil AOA isolate (15,22,30). Aquatic macrophytes such as Phragmites australis can release up to 70 mg of dissolved organic carbon g Ϫ1 (root wet weight) day Ϫ1 (31) and may include organic compounds that are needed for AOA dependent on mixotrophy.Despite the importance of coastal wetlands as nutrient filters, little is known about ammonia oxidizers in these environments. However, based on the above-referenced findings and the fact that AOA dominate marine systems (6, 7), our hypothesis is that AOA are important in coastal wetlands and in particular in the rhizosphere of macrophytes. Our aim was to quantify the relative abundance of AOA and AOB and measure the potential nitrification rates in the sediment and at the root surface of the dominant macrophytes in coastal lagoons to determine if the two groups are differently favored due to their potential interactions with plants. All environments were slightly to moderately alkaline (pH 7.2 to 9.9) and were environments poorly studied in terms of AOB and AOA abundance in plant-associated microbial communities. Samples were obtained from eight coastal lagoons located in two protected areas in Spain with different climatic conditions, covering broad variations in salinity and eutrophication levels.At the Empordà and Baix Ter wetlands (42°14=N, 3°06=E) (2, 19, 23), two oligohaline lagoons, Ter Vell (TV) and Basses d'en Coll (BC), and two euhaline lagoons, Fra Ramon (FR) and Túries (TU), were sampled. The Doñana National Park (37°01=N, 6°25=W) is located in an arid area wi...

The microbial community structure of cork with marked musty-earthy aromas was analyzed using denaturing gradient gel electrophoresis of amplified ribosomal DNA. Cork stoppers and discs were used for DNA extraction and were analyzed by using selective primers for bacteria and fungi. Stoppers clearly differed from discs harboring a different fungal community. Moreover, musty-earthy samples of both types were shown to have a specific microbiota. The fungi Penicillium glabrum and Neurospora spp. were present in all samples and were assumed to make only a small contribution to off-odor development. In contrast, Penicillium islandicum and Penicillium variabile were found almost exclusively in 2,4,6-trichloroanisole (TCA) tainted discs. Conversely, Rhodotorula minuta and Rhodotorula sloofiae were most common in cork stoppers, where only small amounts of TCA were detected. Alpha-and gammaproteobacteria were the most commonly found bacteria in either control or tainted cork stoppers. Specific Pseudomonas and Actinobacteria were detected in stoppers with low amounts of TCA and 2-methoxy-3,5-dimethylpyrazine. These results are discussed in terms of biological degradation of taint compounds by specific microorganisms. Reliable and straightforward microbial identification methods based on a molecular approach provided useful data to determine and evaluate the risk of taint formation in cork.

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In this study, we have developed a rapid method for the simultaneous detection of Listeria monocytogenes and Salmonella spp. in foods, combining culture enrichment and a multiplex real-time polymerase chain reaction (PCR). The assay used two pre-existing primer-probe sets, labelled with different reporter dyes to enable the direct distinction of the original contaminating agent. Amplification efficiency and inclusivity/exclusivity of the combined assay was successfully assessed. The overall process included the culture enrichment based on the ISO standard, consisting of 24 h incubation in appropriate media (Half Fraser Broth for Listeria and buffered peptone water (BPW) for Salmonella), followed by a single DNA extraction of mixed enrichment aliquots, and real-time PCR detection of the hly and bipA genes of L. monocytogenes and Salmonella spp., respectively. An internal amplification control, co-amplified during the PCR run, was included in the assay to verify the results. The tool was evaluated with a variety of artificially inoculated samples of fresh products and ready to eat and cooked dishes, allowing the identification of the target pathogens down to 5 CFU/25 g of food sample. Moreover, the analysis saved a considerable amount of time compared to the ISO standard, being performed in less than 2 working days. Specificity, sensitivity and accuracy were satisfactorily tested by comparison to the standard methods ISO 11290-2:1998 and ISO 6579:2002, suggesting that the tool has a great potential as a reliable alternative for food safety assurance providing rapid detection of both pathogens in food samples.