Horizontal gene transfer (HGT) as a mechanism of disseminating RDX-degrading activity among Actinomycete bacteria

Abstract

Aims: Hexahydro‐1,3,5‐trinitro‐1,3,5,‐triazine (RDX) is a cyclic nitramine explosive that is a major component in many high‐explosive formulations and has been found as a contaminant of soil and groundwater. The RDX‐degrading gene locus xplAB, located on pGKT2 in Gordonia sp. KTR9, is highly conserved among isolates from disparate geographical locations suggesting a horizontal gene transfer (HGT) event. It was our goal to determine whether Gordonia sp. KTR9 is capable of transferring pGKT2 and the associated RDX degradation ability to other bacteria. Methods and Results: We demonstrate the successful conjugal transfer of pGKT2 from Gordonia sp. KTR9 to Gordonia polyisoprenivorans, Rhodococcus jostii RHA1 and Nocardia sp. TW2. Through growth and RDX degradation studies, it was demonstrated that pGKT2 conferred to transconjugants the ability to degrade and utilize RDX as a nitrogen source. The inhibitory effect of exogenous inorganic nitrogen sources on RDX degradation in transconjugant strains was found to be strain specific. Conclusions: Plasmid pGKT2 can be transferred by conjugation, along with the ability to degrade RDX, to related bacteria, providing evidence of at least one mechanism for the dissemination and persistence of xplAB in the environment. Significance and Impact of Study: These results provide evidence of one mechanism for the environmental dissemination of xplAB and provide a framework for future field relevant bioremediation practices.

The essential role of nitrogen limitation in expression of xplA and degradation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in Gordonia sp. strain KTR9

“…Indeed, the successful conjugal transfer of the xplA-containing plasmid, pGKT2, along with its ability to degrade RDX to Gordonia polyisoprenivorans, Rhodococcus jostii RHA1, and Nocardia sp. TW2 was demonstrated in vitro (Jung et al 2011). Interestingly, the inhibitory effects of exogenous inorganic nitrogen sources on RDX degradation in the transconjugant strains were also observed, similar to strain KTR9, suggesting a more global Actinomycete control mechanism for regulating xplA gene expression.…”

“…2c). A possible explanation for this inefficient system is that the xplAB genes were recently acquired by KTR9 via horizontal gene transfer in the RDX-contaminated environment from which KTR9 was isolated (Jung et al 2011). Indeed, the successful conjugal transfer of the xplA-containing plasmid, pGKT2, along with its ability to degrade RDX to Gordonia polyisoprenivorans, Rhodococcus jostii RHA1, and Nocardia sp.…”

“…(SethSmith et al 2008). The xplAB genes share remarkably high identity among these genera, and evidence suggests that their distribution is the result of recent horizontal transfer (Andeer et al 2009;Jung et al 2011;Schoenmuth et al 2013), perhaps in response to selection in RDX-contaminated environments.…”

Analysis of the xplAB -Containing Gene Cluster Involved in the Bacterial Degradation of the Explosive Hexahydro-1,3,5-Trinitro-1,3,5-Triazine

“…The highly conserved nature of this gene cluster between two different genera of actinomycetes indicates that genes have been moved by horizontal gene transfer. Conjugal transfer of RDX degradation among actinomycete bacteria in vitro has been reported (28). The environment has been exposed to RDX, a synthetic compound, for less than a century, and the evolution of its RDX-metabolizing activity is unclear.…”

Dissemination of Bacterial Fluoroquinolone Resistance in Two Multidrug-Resistant Enterobacteriaceae

“…A positive transconjugant clone from each strain was further validated by Southern hybridization and through assessment of the antimicrobial response. A DIG-High Prime-labeled probe (Roche, Indianapolis, Ind., USA) of a qnrS PCR amplicon was generated and used to probe genomic preparations of transconjugants and donor that had been transferred from gels of pulsed field gel electrophoresis to nylon membranes, as detailed previously [Indest et al, 2010;Jung et al, 2011]. The conditions for the pulsed field gel electrophoresis were as follows: 0.5× TBE at 5 V/cm with a switch time of 30-90 s at a 120° angle for 24 h at 10 ° C for total DNA and 0.5× TBE at 5 V/cm with a switch time of 30-60 s at a 120° angle for 21 h at 10 ° C for 40 U I-CeuI/plug overnight digests of DNA (New England Biolabs, Ipswich, Mass., USA).…”