Activity and stability of a recombinant plasmid-borne tce degradative pathway in biofilm cultures

Abstract

The activity and stability of the TCE degradative plasmid TOM31c in the transconjugant host Burkholderia cepacia 17616 was studied in selective and non-selective biofilm cultures. The activity of plasmid TOM31c in biofilm cultures was measured by both TCE degradative studies and the expression of the Tom pathway. Plasmid loss was measured using continuous flow, rotating annular biofilm reactors, and various analytical and microbiological techniques. The probability of plasmid loss in the biofilm cultures was determined using a non-steady-state biofilm plasmid loss model that was derived from a simple mass balance, incorporating results from biofilm growth and plasmid loss studies. The plasmid loss model also utilized Andrew's inhibition growth kinetics and a biofilm detachment term.

Results from these biofilm studies were compared to similar studies performed on suspended cultures of Burkholderia cepacia 17616-TOM31c to determine if biofilm growth has a significant effect on either plasmid retention or Tom pathway expression (i.e., TCE degradation rates). Results show that the activity and expression of the Tom pathway measured in biofilm cultures was significantly less than that found in suspended cultures at comparable growth rates. The data obtained from these studies fit the plasmid loss model well, providing plasmid loss probability factors for biofilm cultures that were equivalent to those previously found for suspended cultures. The probability of plasmid loss in the B. cepacia 17616-TOM31c biofilm cultures was equivalent to those found in the suspended cultures. The results indicate that biofilm growth neither helps nor hinders plasmid stability. In both the suspended and the biofilm cultures, plasmid retention and expression could be maintained using selective growth substrates and/or an appropriate plasmid-selective antibiotic.