Microbial communities from different types of natural wastewater treatment systems: vertical and horizontal flow constructed wetlands and biofilters.

The prokaryotic microbial communities (Bacteria and Archaea) of three different systems operating in Denmark for the treatment of domestic wastewater (horizontal flow constructed wetlands (HFCW), vertical flow constructed wetlands (VFCW) and biofilters (BF)) was analysed using endpoint PCR followed by Denaturing Gradient Gel Electrophoresis (DGGE). Further sequencing of the most representative bacterial bands revealed that diverse and distinct bacterial communities were found in each system unit, being γ-Proteobacteria and Bacteroidetes present mainly in all of them, while Firmicutes was observed in HFCW and BF. Members of the Actinobacteria group, although found in HFCW and VFCW, seemed to be more abundant in BF units. Finally, some representatives of α, ß and δ-Proteobacteria, Acidobacteria and Chloroflexi were also retrieved from some samples. On the other hand, a lower archaeal diversity was found in comparison with the bacterial population. Cluster analysis of the DGGE bacterial band patterns showed that community structure was related to the design of the treatment system and the organic matter load, while no clear relation was established between the microbial assemblage and the wastewater influent.

Identification of neotropical felid faeces using RCP-PCR.

Faeces similarity among sympatric felid species has generally hampered their use in distributional, demographic and dietary studies. Here, we present a new and simple approach based on a set of species-specific primers, for the unambiguous identification of faeces from sympatric neotropical felids (i.e. puma, jaguar, jaguarundi and ocelot/ margay). This method, referred to as rapid classificatory protocol-PCR (RCP-PCR), consists of a single-tube multiplex PCR yielding species-specific banding patterns on agarose gel. The method was optimized with samples of known origin (14 blood and 15 fresh faeces) and validated in faecal samples of unknown origin (n = 138), for some of which (n = 40) we also obtained species identification based on mtDNA sequencing. This approach proved reliable and provides high identification success rates from faeces. Its simplicity and cost effectiveness should facilitate its application for routine surveys of presence and abundance of these species.

A new microtitre plate screening method for evaluating the viability of aerobic respiring bacteria in high surface biofilms.

AIMS: It is difficult to determine the effects of bactericidal compounds against bacteria in a biofilm because classical procedures for determining cell viability require several working days, multiple complicated steps and are frequently only applicable to cells in suspension. We attempt to develop a compact, inexpensive and versatile system to measure directly the extent of biofilm formation from water systems and to determine the viability of respiring bacteria in high surface biofilms. METHODS AND RESULTS: It has been reported that the reduction of tetrazolium sodium salts, such as XTT (sodium 3,3'-[1-[(phenylamino)carbonyl]-3,4-tetrazolium]Bis(4-methoxy)-6-nitro)benzene sulfonic acid hydrate), during active bacterial metabolism can be incorporated into a colorimetric method for quantifying cell viability. XTT is reduced to a soluble formazan compound during bacterial aerobic metabolism such that the amount of formazan generated is proportional to the bacterial biomass. CONCLUSIONS: We show here, for the first time, that this colorimetric approach can be used to determine the metabolic activity of adherent aerobic bacteria in a biofilm as a measure of cell viability. This technique has been used to estimate viability and proliferation of bacteria in suspension, but this is the first application to microbial communities in a real undisturbed biofilm. SIGNIFICANCE AND IMPACT OF THE STUDY: This simple new system can be used to evaluate the complex biofilm community without separating the bacteria from their support. Thus, the results obtained by this practice may be more representative of the circumstances in a natural system, opening the possibility to multiple potential applications.

Size exclusion behavior of hydroxypropylcellulose beads with temperature-dependent porosity.

Beads prepared from a thermosensitive polymer, hydroxypropylcellulose, exhibit temperature-dependent porosity. At temperatures below 40 degrees C the beads are swollen having large pores, while at temperatures above 45 degrees C the beads are in a shrunken state having smaller pores. In the presence of 1 M NaCl the transition temperature decreased to about 30 degrees C. In a swollen state the size of pore is large enough to accommodate lysozyme (mol. mass 14400) and alpha-chymotrypsin (mol. mass 21600) but not bovine serum albumin (mol. mass 67000). When the beads are shrunken, all the proteins are eluted from the column packed with hydroxypropylcellulose beads in the volume close to the void volume of the column.