Cl-compound specific isotope analysis and assessment of functional genes for monitoring monochlorobenzene (MCB) biodegradation under aerobic conditions.

A laboratory approach was adopted in this study to explore the potential of Cl-CSIA in combination with C-CSIA and Biological Molecular Tools (BMTs) to estimate the occurrence of monochloroenzene (MCB) aerobic biodegradation. A new analytical method for Cl-CSIA of MCB was developed in this study. This methodology using a GC-IRMS allowed to determine δCl values within an internal error of ±0.3‰. Samples from a heavily MCB contaminated site were collected and MCB aerobic biodegradation microcosms with indigenous cultures in natural and enhanced conditions were set up. The microcosms data show a negligible fractionation for C associated to MCB mass decrease of >95% over the incubation time. Conversely, an enrichment factor of -0.6±0.1‰ was estimated for Cl, which is a reflection of a secondary isotope effect. Moreover, the dual isotope approach showed a pattern for aerobic degradation which differ from the theoretical trend for reductive dehalogenation. Quantitative Polymerase Chain Reaction (qPCR) results showed a significant increase in todC gene copy number with respect to its initial levels for both natural attenuation and biostimulated microcosms, suggesting its involvement in the MCB aerobic degradation, whereas phe gene copy number increased only in the biostimulated ones. Indeed, Cl fractionation in combination with the dual carbon‑chlorine isotope approach and the todC gene copy number represent valuable indicators for a qualitative assessment of MCB aerobic biodegradation in the field.