Comparison between wild type 1026b, DD503 and JW270
From Figs 3 and 5, we observe that 1026b Δasd and Bp82 bear the same biofilm composition while JW270 and E264 are similar despite the fact that JW270 was generated from the DD503 (Fig S1), a derivative of 1026b in which the genes encoding amrAB-oprA antibiotic efflux pump has been deleted [39]. To confirm that the mutation used to generate JW270 may be responsible for its different biofilm composition, we repeated the assays for protein, eDNA and glucose concentration using the wild type 1026b, DD503 and JW270.
1026b and its direct derivative DD503 have the similar protein concentration (Fig 6a, p = 0.006), and eDNA florescence (Fig 6c, p = 0.03) at 72 hours but DD503 shows an increased polysaccharide concentration throughout the experiment time span (Fig 6b). However, once the wcb operon is deleted in DD503 to generate JW270, the polysaccharide and protein concentrations drastically decreases but shows an inverse increase in eDNA. There is a significant difference between the concentrations of all biofilm components when DD503 and JW270 are compared (p = 0.01-0.04). This data implies that the deletion of the wcb cluster may be responsible for the low biofilm polysaccharide and protein concentrations and the overcompensation of eDNA export in JW270 to maintain a stable biofilm. To quickly confirm our hypothesis, we performed an ELISA test using JW270 as test sample, and 1026b Δasd and Bp82 as positive controls. B. thailandensis E264 which lacks the CPS cluster was used as negative control. The ELISA test was performed using an anti-CPS primary antibody, 4B11 [52], and a goat anti-mouse alkaline phosphatase secondary antibody. The results shows reactivity between the 4B11 antibody and 1026b Δasd and Bp82 but not towards JW270 orB. thailandensis E264 (Fig. S2). Taken together, the results demonstrate an inverse relationship between eDNA and CPS I inBurkholderia biofilms.