Molecular mechanisms of genetic adaptation to xenobiotic compounds.

Microorganisms in the environment can often adapt to use xenobiotic chemicals as novel growth and energy substrates. Specialized enzyme systems and metabolic pathways for the degradation of man-made compounds such as chlorobiphenyls and chlorobenzenes have been found in microorganisms isolated from geographically separated areas of the world. The genetic characterization of an increasing number of aerobic pathways for degradation of (substituted) aromatic compounds in different bacteria has made it possible to compare the similarities in genetic organization and in sequence which exist between genes and proteins of these specialized catabolic routes and more common pathways. These data suggest that discrete modules containing clusters of genes have been combined in different ways in the various catabolic pathways. Sequence information further suggests divergence of catabolic genes coding for specialized enzymes in the degradation of xenobiotic chemicals. An important question will be to find whether these specialized enzymes evolved from more common isozymes only after the introduction of xenobiotic chemicals into the environment. Evidence is presented that a range of genetic mechanisms, such as gene transfer, mutational drift, and genetic recombination and transposition, can accelerate the evolution of catabolic pathways in bacteria. However, there is virtually no information concerning the rates at which these mechanisms are operating in bacteria living in nature and the response of such rates to the presence of potential (xenobiotic) substrates. Quantitative data on the genetic processes in the natural environment and on the effect of environmental parameters on the rate of evolution are needed.     

Environmental genomics: exploring the unmined richness of microbes to degrade xenobiotics

Increasing pollution of water and soils by xenobiotic compounds has led in the last few decades to an acute need for understanding the impact of toxic compounds on microbial populations, the catabolic degradation pathways of xenobiotics and the set-up and improvement of bioremediation processes. Recent advances in molecular techniques, including high-throughput approaches such as microarrays and metagenomics, have opened up new perspectives and pointed towards new opportunities in pollution abatement and environmental management. Compared with traditional molecular techniques dependent on the isolation of pure cultures in the laboratory, microarrays and metagenomics allow specific environmental questions to be answered by exploring and using the phenomenal resources of uncultivable and uncharacterized micro-organisms. This paper reviews the current potential of microarrays and metagenomics to investigate the genetic diversity of environmentally relevant micro-organisms and identify new functional genes involved in the catabolism of xenobiotics.     

Functional gene diversity analysis in BTEX contaminated soils by means of PCR-SSCP DNA fingerprinting: comparative diversity assessment against bacterial isolates and PCR-DNA clone libraries

Developments in molecular biology based techniques have led to rapid and reliable tools to characterize microbial community structures and to monitor their dynamics under in situ conditions. However, there has been a distinct lack of emphasis on monitoring the functional diversity in the environment. Genes encoding catechol 2,3-dioxygenases (C23O), as key enzymes of various aerobic aromatic degradation pathways, were used as functional targets to assess the catabolic gene diversity in differentially BTEX contaminated environments by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP). Site specific PCR-SSCP fingerprints were obtained, showing that gene diversity experienced shifts correlated to temporal changes and levels of contamination. PCR-SSCP enabled the recovery of predominant gene polymorphs, and results closely matched with the information retrieved from random sequencing of PCR-DNA clone libraries. A new method for isolating strains capable of growing on BTEX compounds was developed to diminish preselection or enrichment bias and to assess the function of predominant gene polymorphs. C23O abundance in isolates correlated with the levels of BTEX pollution in the soil samples analysed. Isolates harbouring C23O genes, identical to the gene polymorph predominant in all contaminated sites analysed, showed an unexpected benzene but not toluene mineralizing phenotype whereas isolates harbouring a C23O gene variant differing by a single point mutation and observed in highly polluted sites only, were capable, among some other isolates, to mineralize benzene and toluene, indicating a catabolically determined sharing of carbon sources on-site. The PCR-SSCP technique is thus a powerful tool for assessing the diversity of functional genes and the identification of predominant gene polymorphs in environmental samples as a prerequisite to understand the functioning of microbial communities.     

细菌降解多环芳烃上游途径的遗传学研究进展

多环芳烃是一类毒性较大的环境污染物。微生物降解和转化是消除此类污染物的理想方法,已发现多种细菌具有这种功能。主要针对细菌在多环芳烃降解中上游途径的代谢酶及基因簇的组成进行综述,阐述了酶的遗传学特点,并探讨了PAHs代谢基因的进化。这有助于了解PAHs的细菌降解机制,并为有效实施生物修复提供理论依据。     

Insights into environmental bioremediation by microorganisms through functional genomics and proteomics

Environmental pollutants in the soil are a major concern worldwide. Bioremediation mediated by microorganisms is a highly promising technology that is environmentally friendly, safe, and effective. However, incomplete biological information regarding the cellular responses in many microbial communities restricts progress in the site-specific mineralization process. The application of proteomics in environmental bioremediation research provides a global view of the protein compositions of the microbial cells and offers a promising approach to address the molecular mechanisms of bioremediation. With the combination of proteomics, functional genomics provide an insight into global metabolic and regulatory networks that can enhance the understanding of gene functions. This article deals with the applications of functional genomics and proteomics to dissect the cellular responses to environmental stimuli, such as stress response, induction and expressions of regulatory proteins/enzymes in response to aromatic hydrocarbons and heavy metals. An understanding of the growth conditions governing the expression of the proteome (for example, enzymes and regulatory proteins of aromatic hydrocarbon degradation, energy generation pathways, transport and stress-related proteins) in a specific environment is essential for developing rational strategies for successful bioremediation.     

Emerging high-throughput approaches to analyze bioremediation of sites contaminated with hazardous and/or recalcitrant wastes

Sustainable development requires the promotion of environmental management and a constant search for new technologies to treat a wide range of aquatic and terrestrial habitats contaminated by increasing anthropogenic activities. Bioremediation, i.e. the elimination of natural or xenobiotic pollutants by living organisms, is an environmentally friendly and cost-effective alternative to physico-chemical cleanup options. However, the strategy and outcome of bioremediation in open systems or confined environments depend on a variety of physico-chemical and biological factors that need to be assessed and monitored. In particular, microorganisms are key players in bioremediation applications, yet their catabolic potential and their dynamics in situ remain poorly characterized. To perform a comprehensive assessment of the biodegradative potential of a contaminated site and efficiently monitor changes in the structure and activities of microbial communities involved in bioremediation processes, sensitive, fast and large-scale methods are needed. Over the last few years, the scientific literature has revealed the progressive emergence of genomic high-throughput technologies in environmental microbiology and biotechnology. In this review, we discuss various high--throughput techniques and their possible--or already demonstrated-application to assess biotreatment of contaminated environments.     

Microarray-based analysis of microbial functional diversity along an oil contamination gradient in oil field

To understand better the in situ microbial functional diversity under oil contamination stress, soils were sampled along a contamination gradient at an oil field in north-east China. Microbial community functional structure was examined with a functional gene array, termed GeoChip. Multivariate statistical analysis and meta-analysis were conducted to study the functional gene responses to oil concentrations. The total functional gene abundance and diversity decreased along the gradient of increasing contamination. The overall abundance of soil bacteria, archaea and fungi decreased to 10%, 40% and 80% of those in the pristine soil. Several functional genes in the families pgl, rbcL, nifH and nor and those encoding cellulase, laccase, chitinase, urease and key enzymes in metabolizing organic compounds were significantly decreased with oil contamination, especially under high contamination stress. However, a few genes encoding key enzymes for catechol, protocatechuate, and biphenyl degradation and in the gene families of nir, rbcL and pgl showed a significant increase at a medium level of oil contamination. Oil content and soil available nitrogen were found to be important factors influencing the microbial community structure. The results provide an insight into microbial functional diversity in oil-contaminated soils, providing potential information for on-site management and remediation measures.     

Anaerobic biodegradation of aromatic hydrocarbons: pathways and prospects

Aromatic hydrocarbons contaminate many environments worldwide, and their removal often relies on microbial bioremediation. Whereas aerobic biodegradation has been well studied for decades, anaerobic hydrocarbon biodegradation is a nascent field undergoing rapid shifts in concept and scope. This review presents known metabolic pathways used by microbes to degrade aromatic hydrocarbons using various terminal electron acceptors; an outline of the few catabolic genes and enzymes currently characterized; and speculation about current and potential applications for anaerobic degradation of aromatic hydrocarbons.     

Assessment and modification of degenerate qPCR primers that amplify functional genes from etheneotrophs and vinyl chloride-assimilators

Aims: Degenerate qPCR primer sets that target the functional genes etnC and etnE in etheneotrophs and vinyl chloride‐assimilating bacteria were assessed and modified in an effort to improve performance. Methods and Results: Functional gene abundance in four pure cultures was estimated by qPCR using novel (MRTC and MRTE) and existing (RTC and RTE) degenerate primer sets and compared to abundances estimated with nondegenerate gene‐specific primers (GSPs). Functional gene abundance in groundwater DNA extracted from several contaminated sites was also estimated with MRTC and MRTE primers. Conclusions: MRTC primers displayed significantly improved etnC quantification in both pure cultures and environmental samples. Significance and Impact of the Study: Application of MRTC and MRTE primer sets will enhance microbial ecology studies involving etheneotrophs and qPCR analyses that support vinyl chloride bioremediation strategies.