Figure 7.

Expression of the AlgU/T regulon in a ΔalgL strain background rescues Pseudomonas aeruginosa lethality during alginate production.A, Western blot analysis of whole cell lysates expressing Alg44, AlgD, or AlgL after 1 h induction with 0% or 0.5% (w/v) L-arabinose using poly-clonal Alg protein specific antibodies. B, growth curves of PAO1 ΔwspF P_BADalg (parental), PAO1 ΔwspF P_BADalg ΔalgL (ΔalgL), PAO1 ΔwspF P_BADalg ΔalgL::algL (ΔalgL::algL), and PAO1 ΔpelA ΔpslBCD mucA22 ΔalgL ΔalgD::algD (mucA22 ΔalgL ΔalgD::algD). After reaching an OD₆₀₀ of approximately 0.5, cells were induced to produce alginate with 0.5% (w/v) L-arabinose. Values shown represent averages of two biological replicates with two technical replicates for each biological replicate. C, quantification of alginate produced over the course of 22 h by parental and mucA22 ΔalgL ΔalgD::algD with addition of 0.5% (w/v) L-arabinose to the growth media. Values represent all technical replicates across four separate experiments. Red lines represent the median. Statistical analysis was carried out using a Mann–Whitney test: ∗∗∗∗ indicates p < 0.0001. D, chemiluminescence detection of a dot blot on secreted and periplasmic fraction samples from P. aeruginosa strains using a monoclonal alginate-reactive antibody.