Skip to main content
. 2021 Sep 2;24(9):103071. doi: 10.1016/j.isci.2021.103071

Figure 5.

Figure 5

Overexpression of HfsH increases biofilm formation in C. crescentus but not H. baltica

(A) Schematic representations of cross-complementation constructs of the hfsH gene from C. crescentus (hfsHCC) and H. baltica (hfsHHB) under native holdfast synthesis (Phfs) or xylose inducible (Pxyl) promoters. Native promoters were fused to foreign hfsH genes (PhfsCC and PxylCC from C. crescentus, or PhfsHB and PxylHB from H. baltica) and assembled into the pMR10 plasmid.

(B) Quantification of short-term adhesion (12 h) by the crystal violet assay. Data are expressed as a mean percent of WT crystal violet staining from three biological replicates with four technical replicates each. Error is expressed as the standard error of the mean.

(C) Representative images showing merged phase and fluorescence channels of C. crescentus and H. baltica strains. Holdfasts are labeled with AF488-WGA (green). PhfsCC-hfsHHB, C. crescentus ΔhfsH cross-complemented with HfsH from H. baltica under the control of the hfs promoter; PhfsHB-hfsHCC, H. baltica ΔhfsH cross-complemented with HfsH from C. crescentus under the control of the hfs promoter. Exponential planktonic cultures were used to quantify the percentage of predivisional cells with holdfast. Scale bar, 2 μm. Data are expressed as the mean of three independent biological replicates with four technical replicates each. Error bars represent the standard error of the mean.

(D) Images showing merged phase and fluorescence channels of C. crescentus and H. baltica strains bound to glass slides. Holdfast is labeled with AF488-WGA (green). Exponential cultures were incubated on the glass slides for 1 h, unbound cells were washed off, and AF488-WGA was added to label bound holdfast. Scale bar, 2 μm.

(E) Merged phase and fluorescence channels of H. baltica ΔhfsH and C. crescentus ΔhfsH strains bound to glass slides. Holdfast is labeled with AF488-WGA (green). Exponential cultures were incubated on the glass slides for 1 h, unbound cells were washed off, and AF488-WGA was added to label bound holdfast. Strains carry native or cross-complemented HfsH under the control of the xylose inducible promoter for overexpression. Scale bar, 2 μm.