Bacterial diversity of lactate-and ethanol-utilizing sulfate-reducing fluidized-bed reactor (FBR) communities was investigated with culture-independent methods. The FBRs were fed for 500 days with synthetic mineral processing wastewater containing sulfate, zinc and iron with hydraulic retention time of 16^24 h. Sodium lactate or ethanol was used as electron donor for microbial sulfate reduction. For microbial characterization, 16S rRNA gene clone libraries and denaturing gradient gel electrophoresis (DGGE) fingerprinting were employed. The FBR communities were diverse and contained many previously undescribed bacteria. The clone library indicated significant differences between bacterial communities of the two reactors. Most notable was the large number of Proteobacterium sequences retrieved from the ethanol-fed reactor, whereas in the lactate-fed reactor, sequences clustering with Nitrospira phylum were most abundant. Ethanol-utilizing FBR culture was more diverse than the lactate-utilizing one. Some sequences from each reactor were closely related to known sulfate reducers, such as Desulfobacca acetoxidans, Desulforhabdus amnigenus, and species of Desulfovibrio. DGGE profiling showed some changes in the bacterial communities over 393 days of continuous FBR operation. This study showed that it is possible to maintain diverse sulfate-reducing consortia using simple electron donors, lactate or ethanol in an open engineered ecosystem.

Rapid detection of pathogenic Naegleria fowler in water distribution networks is critical for water utilities. Current detection methods rely on sampling drinking water followed by culturing and molecular identification of purified strains. This culture-based method takes an extended amount of time (days), detects both nonpathogenic and pathogenic species, and does not account for N. fowleri cells associated with pipe wall biofilms. In this study, a total DNA extraction technique coupled with a real-time PCR method using primers specific for N. fowleri was developed and validated. The method readily detected N. fowleri without preculturing with the lowest detection limit for N. fowleri cells spiked in biofilm being one cell (66% detection rate) and five cells (100% detection rate). For drinking water, the detection limit was five cells (66% detection rate) and 10 cells (100% detection rate). By comparison, culture-based methods were less sensitive for detection of cells spiked into both biofilm (66% detection for <10 cells) and drinking water (0% detection for <10 cells). In mixed cultures of N. fowleri and nonpathogenic Naegleria, the method identified N. fowleri in 100% of all replicates, whereastests with the current consensus primers detected N. fowleri in only 5% of all replicates. Application of the new method to drinking water and pipe wall biofilm samples obtained from a distribution network enabled the detection of N. fowleri in under 6 h, versus 3+ daysforthe culture based method. Further, comparison of the real-time PCR data from the field samples and the standard curves enabled an approximation of N. fowleri cells in the biofilm and drinking water. The use of such a method will further aid water utilities in detecting and managing the persistence of N. fowleri in water distribution networks.

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Fluorescent in situ hybridization (FISH) using 16S and 23S rRNA-targeted probes together with construction of an archaeal 16S ribosomal DNA (rDNA) clone library was used to characterize the microbial populations of an anaerobic baffled reactor successfully treating industrial dye waste. Wastewater produced during the manufacture of food dyes containing several different azo and other dye compounds was decolorized and degraded under sulfidogenic and methanogenic conditions. Use of molecular methods to describe microbial populations showed that a diverse group of Bacteria and Archaea was involved in this treatment process. FISH enumeration showed that members of the gamma subclass of the class Proteobacteria and bacteria in the Cytophaga-Flexibacter-Bacteroides phylum, together with sulfate-reducing bacteria, were prominent members of a mixed bacterial population. A combination of FISH probing and analysis of 98 archaeal 16S rDNA clone inserts revealed that together with the bacterial population, a methanogenic population dominated by Methanosaeta species and containing species of Methanobacterium and Methanospirillum and a relatively unstudied methanogen, Methanomethylovorans hollandica, contributed to successful anaerobic treatment of the industrial waste. We suggest that sulfate reducers, or more accurately sulfidogenic bacteria, together with M. hollandica contribute considerably to the treatment process through metabolism of dye-associated sulfonate groups and subsequent conversion of sulfur compounds to carbon dioxide and methane.