Enrichment of a dioxin-dehalogenating Dehalococcoides species in two-liquid phase cultures.
Environ Microbiol, 2008/10;10(10):2670-83.
Bunge M[1], Wagner A, Fischer M, Andreesen JR, Lechner U
Affiliations
PMID: 18627411DOI: 10.1111/j.1462-2920.2008.01688.x
Impact factor: 5.476
Abstract
Enrichment cultures capable of reductively dechlorinating 1,2,4-trichlorodibenzo-p-dioxin (1,2,4-TrCDD) were shown to dechlorinate 1,2,3-trichlorobenzene (1,2,3-TrCB) to 1,3-dichlorobenzene. To test if this activity can be used to enrich for dioxin-dechlorinating bacteria, a two-liquid phase cultivation with 200 mM 1,2,3-TrCB dissolved in hexadecane was established. During the dechlorination of 1,2,3-TrCB, the number of 1,2,4-TrCDD-dechlorinating bacteria increased by four orders of magnitude, eventually accounting for 11% of the total cell number. Characterization of the bacterial communities of the initial dioxin-dechlorinating culture and of the trichlorobenzene enrichments by restriction fragment length polymorphism (RFLP) analysis of cloned 16S rRNA genes revealed a proportional increase of nine different sequence types, one representing a Dehalococcoides strain. Inhibition of methanogens further enhanced the rate of chlorobenzene dehalogenation and also resulted in a rapid dechlorination of 1,2,3,4-tetrachlorodibenzo-p-dioxin that was applied via a hexadecane phase. The further enrichment was monitored by terminal RFLP, quantitative real-time PCR and microscopy, and aimed at the reduction of the accompanying non-dehalogenating populations by using different combinations of electron donors and the application of antibiotics. Hydrogen as the sole electron donor proved to be less efficient due to the co-enrichment of acetogens. The novel Dehalococcoides strain DCMB5 was enriched up to 50% by the cultivation with organic acids, hydrogen and vancomycin, and was finally purified by conventional isolation techniques.
MeSH terms
Anti-Bacterial Agents; Carboxylic Acids; Chlorobenzenes; Chloroflexi; Colony Count, Microbial; Culture Media; DNA Fingerprinting; DNA, Bacterial; DNA, Ribosomal; Hydrogen; Metabolic Networks and Pathways; Microscopy; Molecular Sequence Data; Molecular Structure; Phylogeny; Polychlorinated Dibenzodioxins; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; RNA, Ribosomal, 16S; Sequence Analysis, DNA; Vancomycin
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