Anaerobic Methyl tert -Butyl Ether-Degrading Microorganisms Identified in Wastewater Treatment Plant Samples by Stable Isotope Probing

Abstract

<jats:title>ABSTRACT</jats:title><jats:p>Anaerobic methyl<jats:italic>tert</jats:italic>-butyl ether (MTBE) degradation potential was investigated in samples from a range of sources. From these 22 experimental variations, only one source (from wastewater treatment plant samples) exhibited MTBE degradation. These microcosms were methanogenic and were subjected to DNA-based stable isotope probing (SIP) targeted to both bacteria and archaea to identify the putative MTBE degraders. For this purpose, DNA was extracted at two time points, subjected to ultracentrifugation, fractioning, and terminal restriction fragment length polymorphism (TRFLP). In addition, bacterial and archaeal 16S rRNA gene clone libraries were constructed. The SIP experiments indicated bacteria in the phyla<jats:named-content content-type="genus-species">Firmicutes</jats:named-content>(family<jats:named-content content-type="genus-species">Ruminococcaceae</jats:named-content>) and<jats:named-content content-type="genus-species">Alphaproteobacteria</jats:named-content>(genus<jats:named-content content-type="genus-species">Sphingopyxis</jats:named-content>) were the dominant MTBE degraders. Previous studies have suggested a role for<jats:named-content content-type="genus-species">Firmicutes</jats:named-content>in anaerobic MTBE degradation; however, the putative MTBE-degrading microorganism in the current study is a novel MTBE-degrading phylotype within this phylum. Two archaeal phylotypes (genera<jats:named-content content-type="genus-species">Methanosarcina</jats:named-content>and<jats:named-content content-type="genus-species">Methanocorpusculum</jats:named-content>) were also enriched in the heavy fractions, and these organisms may be responsible for minor amounts of MTBE degradation or for the uptake of metabolites released from the primary MTBE degraders. Currently, limited information exists on the microorganisms able to degrade MTBE under anaerobic conditions. This work represents the first application of DNA-based SIP to identify anaerobic MTBE-degrading microorganisms in laboratory microcosms and therefore provides a valuable set of data to definitively link identity with anaerobic MTBE degradation.</jats:p>

The key microorganisms for anaerobic degradation of pentachlorophenol in paddy soil as revealed by stable isotope probing

“…T-RFLP fingerprinting of densityresolved DNA fractions was done with primers 27F-FAM (5 -AGAGTTTGATCMTGGCTCAG-3 , 5 end-labeled with carboxyfluorescein) and 1492R (5 -GGTTACCTTGTTACGACTT-3 ) for bacteria and A109F-FAM (5 -ACKGCTCAGTAACACGT-3 , 5 end-labeled with carboxyfluorescein) and A934R (5 -GTGCTCCCCCGCCAATTCCT-3 ) for archaea; the purified PCR products were digested with HaeIII, AluI and RsaI (New England Biolabs) and the data were analyzed with GeneScan software, all as described previously [39][40][41].…”

Using DNA‐Stable Isotope Probing to Identify MTBE‐ and TBA‐Degrading Microorganisms in Contaminated Groundwater

“…In a DNA-SIP study of MTBE degradation in methanogenic microcosms, 18 clones enriched with 13C were related to Sphingopyxis of the Sphingomonadaceae family (Sun et al 2012). The Sphingomonas genus in this family has also been associated with MTBE degradation.…”

Biodegradation of gasoline ether oxygenates

“…Historically, mixed results have been obtained in studies of anaerobic ether oxygenate and tertiary alcohol biodegradation [2] and this trend has continued to the present. In some recent studies, little or no biodegradation has been observed over long periods [3,4]. In other cases, rapid anaerobic biodegradation has been observed [5].…”

“…A laboratory-based 13 C-DNA-stable isotope probing study recently identified Firmicutes as well as Sphigopyxis strains as dominant MTBE-degrading organisms in methanogenic wastewater treatment plant samples [4]. A study using 13 C 4 -TBA has also characterized aerobic TBA-metabolizing strains in Bio-GAC reactor samples.…”