16S ribosomal RNA gene (rDNA) amplicon analysis remains the standard approach for the cultivation-independent investigation of microbial diversity. The accuracy of these analyses depends strongly on the choice of primers. The overall coverage and phylum spectrum of 175 primers and 512 primer pairs were evaluated in silico with respect to the SILVA 16S/18S rDNA non-redundant reference dataset (SSURef 108 NR). Based on this evaluation a selection of 'best available' primer pairs for Bacteria and Archaea for three amplicon size classes (100-400, 400-1000, ≥ 1000 bp) is provided. The most promising bacterial primer pair (S-D-Bact-0341-b-S-17/S-D-Bact-0785-a-A-21), with an amplicon size of 464 bp, was experimentally evaluated by comparing the taxonomic distribution of the 16S rDNA amplicons with 16S rDNA fragments from directly sequenced metagenomes. The results of this study may be used as a guideline for selecting primer pairs with the best overall coverage and phylum spectrum for specific applications, therefore reducing the bias in PCR-based microbial diversity studies.

Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies

Fouling in membrane bioreactors used in wastewater treatment

Physico-chemical treatment techniques for wastewater laden with heavy metals

Recent advances in membrane bioreactors (MBRs): membrane fouling and membrane material.

We report the design and evaluation of PCR primers 63f and 1387r for amplification of 16S rRNA genes from bacteria. Their specificity and efficacy were tested systematically with a variety of bacterial species and environmental samples. They were found to be more useful for 16S rRNA gene amplification in ecological and systematic studies than PCR amplimers that are currently more generally used.

Design and Evaluation of Useful Bacterium-Specific PCR Primers That Amplify Genes Coding for Bacterial 16S rRNA

Teaching the MDGs: the Role of the Nurse in Multidsciplinary Project Based Instruction

The microbiological quality of drinking water is assessed using culture-based methods that are highly selective and that tend to underestimate the densities and diversity of microbial populations inhabiting distribution systems. In order to better understand the effect of different disinfection treatments on the bacteria in water distribution systems (WDS) we have used 16S rDNA PCR-based techniques. 16S rDNA clone libraries were developed using DNA extracted from samples collected at different times and locations in a metropolitan distribution system derived from different source waters. Since it is possible for DNA to persist in the environment after cell death we also used rRNA (instead of DNA) as the target molecule to study the active bacterial fraction in drinking water biofilms. Phylogenetic analyses and sequence comparisons with existing databases revealed that αand β-Proteobacteria were among the most predominant bacterial groups identified in both biofilm and planktonic communities. Clones closely related to Mycobacterium spp. and Legionella spp. were also obtained in these studies. While members of the Mycobacterium and Legionella genera are known to be pathogenic, the public health relevance of these clones has yet to be determined. Differences in biofilm community structure between disinfection treatments were evident as biofilms exposed to chloramine primarily consisted of Mycobacterium spp. and Dechloromonas spp. In contrast, a variety of αand βProteobacteria dominated the DNA-based clone libraries of biofilm receiving no disinfectant and biofilm exposed to chlorine. Sequence analysis of RNA-based clones derived from biofilms exposed to chlorine suggested that the active bacterial fraction consisted of a few dominant bacterial groups related to Nevskia ramosa, a member of the γ-Proteobacteria group frequently found in the air-water interphase (i.e., neuston). Some of the bacterial groups identified in the latter clone libraries are affiliated with yet to be cultured organisms suggesting that a significant fraction of drinking water communities cannot be studied using culturing methods. The results from these studies suggest that we have a limited understanding of the molecular diversity and population dynamics of WDS microbial communities. Future studies will focus on expanding sequencing databases to more accurately characterize potential differences between disinfection treatments and to better understand the microbial diversity in drinking water systems.

Molecular diversity of drinking water microbial communities: a phylogenetic approach

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NSF CCLI: Developing a Molecular Biology Lab Course in Environmental Engineering and Science

Influence of Loading/Decanting Patterns on Sludge Properties and Membrane Fouling in Membrane Bioreactors Treating Synthetic Early Planetary Base Wastewater

In this paper, we discuss the development of a novel method of bacteria detection using a microwave resonant cavity. The approach is rapid, on the order of minutes, and provides an alternative to the slower fluorescence-based assay currently used with culture-based biochips.

Daniel B. Oerther

Papers

Teaching the MDGs: the Role of the Nurse in Multidsciplinary Project Based Instruction

Molecular diversity of drinking water microbial communities: a phylogenetic approach

NSF CCLI: Developing a Molecular Biology Lab Course in Environmental Engineering and Science

Influence of Loading/Decanting Patterns on Sludge Properties and Membrane Fouling in Membrane Bioreactors Treating Synthetic Early Planetary Base Wastewater

Rapid Detection of Microorgansims Using Microwaves