Comparative Genomics of “ Dehalococcoides ethenogenes ” 195 and an Enrichment Culture Containing Unsequenced “ Dehalococcoides ” Strains

Abstract

Tetrachloroethene (PCE) and trichloroethene (TCE) are prevalent groundwater contaminants that can be completely reductively dehalogenated by some "Dehalococcoides" organisms. A Dehalococcoides-organism-containing microbial consortium (referred to as ANAS) with the ability to degrade TCE to ethene, an innocuous end product, was previously enriched from contaminated soil. A whole-genome photolithographic microarray was developed based on the genome of "Dehalococcoides ethenogenes" 195. This microarray contains probes designed to hybridize to >99% of the predicted protein-coding sequences in the strain 195 genome. DNA from ANAS was hybridized to the microarray to characterize the genomic content of the ANAS enrichment. The microarray results revealed that the genes associated with central metabolism, including an apparently incomplete carbon fixation pathway, cobalamin-salvaging system, nitrogen fixation pathway, and five hydrogenase complexes, are present in both strain 195 and ANAS. Although the gene encoding the TCE reductase, tceA, was detected, 13 of the 19 reductive dehalogenase genes present in strain 195 were not detected in ANAS. Additionally, 88% of the genes in predicted integrated genetic elements in strain 195 were not detected in ANAS, consistent with these elements being genetically mobile. Sections of the tryptophan operon and an operon encoding an ABC transporter in strain 195 were also not detected in ANAS. These insights into the diversity of Dehalococcoides genomes will improve our understanding of the physiology and evolution of these bacteria, which is essential in developing effective strategies for the bioremediation of PCE and TCE in the environment.Chlorinated solvents, such as tetrachloroethene (PCE) and trichloroethene (TCE), are prevalent groundwater contaminants. Microorganisms from various genera, including Dehalobacter (17), Desulfitobacterium (11, 12), Desulfomonile (8), Desulfuromonas (22), Enterobacter (34), and Sulfurospirillum (26, 32), are able to dechlorinate PCE and TCE to predominantly dichloroethene (DCE). To date, the only microorganisms known to completely dechlorinate PCE and TCE to the innocuous end product ethene are "Dehalococcoides" bacteria (28).The sequencing and annotation of the genome of "Dehalococcoides ethenogenes" 195, the first Dehalococcoides bacterium sequenced, provided insights into its unique physiology (33). The subsequent sequencing of the genome of Dehalococcoides sp. strain CBDB1, which has different dechlorination abilities than strain 195 (1, 4), allowed a genetic comparison of two functionally different Dehalococcoides organisms (23). That study revealed a high degree of gene sequence conservation and synteny within central metabolism and informationprocessing systems ("housekeeping" genes). However, these genomes appear to be punctuated by regions of high plasticity, many of which contained putative reductive dehalogenase (RD) genes and integrated genetic elements (IEs), suggesting a high degree of evolutionary variability (23). Many coding regi...

Identification and transcriptional analysis of trans-DCE-producing reductive dehalogenases in Dehalococcoides species

“…The dceA7 gene (100% identity to DET0088) is truncated, and the 3 0 anchoring protein is replaced by a hypothetical protein. Nevertheless, recent studies have reported that the expression of DET0088 was upregulated during PCE dechlorination in Alameda Naval Air Station enrichments and mixed cultures containing strain 195 (Rahm et al, 2006;West et al, 2008). Also, the dceA7 gene was identified using the newly designed primer pairs, RDH F1C and RDH R1C, revealing inadequacies of the previous degenerate primer pair.…”

Metagenomic analysis of a stable trichloroethene-degrading microbial community

“…For metagenomic sequencing, samples from the same time point (27 h) from four different feeding cycles were pooled to collect enough material for sequencing. Total nucleic acids were extracted from frozen filters using a modified version of the bead beating and phenol extraction method described previously (West et al, 2008).…”

“…This culture was derived from sediment collected at the Alameda Naval Air Station and is referred to as ANAS (Richardson et al, 2002). The phylogenetic composition of ANAS has been studied using clone libraries (Richardson et al, 2002;Lee et al, 2006), and the Dhc strains in ANAS have been analyzed using quantitative PCR and whole-genome microarrays West et al, 2008;Lee et al, 2011). ANAS contains two Dhc strains, which have recently been isolated Lee et al, 2011).…”

Overview of organohalide-respiring bacteria and a proposal for a classification system for reductive dehalogenases

“…Several molecular tools have been developed for examining a broad range of rdhA genes in a given sample (site, isolate or mixed culture), including both microarray-based methods and PCR-based protocols [60][61][62][63]. Identification of novel reductive dehalogenase genes has primarily come from PCR studies [32,33,[35][36][37]39,40,42] and subsequently from genome and metagenome sequencing efforts [6,13,14,41,63], with each new organism or environment sampled increasing the known diversity of putative rdhA genes.…”