Dehalorespiration is an environmental-friendly bacterial anaerobic respiration that couples reductive dechlorination of chlorinated organic pollutants with energy conservation. Tetrachloroethene (PCE) respiration is encoded by the pce gene cluster in Desulfitobacterium hafniense strain TCE1, a cluster embedded in the composite transposon Tn-Dha1. The objective of this study was to characterize the stability and/or mobility of the pce transposon. Starting from an inoculum of strain TCE1 routinely growing on PCE as sole electron acceptor, we cultivated it on fumarate instead of PCE, and successively sub-cultivated it on fumarate for more than 30 times. We followed the dehalorespiration process using a multi-level approach (protein and enzymatic activity, mRNA and DNA) revealing a striking loss of the pce gene cluster in strain TCE1 population between sub-cultivations 6 and 12. Using a PCR-based strategy, we were able to identify two genetic processes, namely transposition of Tn-Dha1 (and of single copies of the insertion sequence ISDha1 flanking the transposon at both ends), and homologous recombination across ISDha1. Both phenomona take place simultaneously and have detrimental effects on dehalorespiration. Analysis of the intervening sequence during ISDha1 excision allowed us to further consider the transposition mechanism.