Possible nonconductive role of Geobacter sulfurreducens pilus nanowires in biofilm formation

Abstract

Geobacter sulfurreducens required expression of electrically conductive pili to form biofilms on Fe(III) oxide surfaces, but pili were also essential for biofilm development on plain glass when fumarate was the sole electron acceptor. Furthermore, pili were needed for cell aggregation in agglutination studies. These results suggest that the pili of G. sulfurreducens also have a structural role in biofilm formation.

FIG. 1.

Biofilm formation on Fe(III) oxide coatings by G. sulfurreducens. (A) When provided as a sole electron acceptor for growth, the Fe(III) oxide coating supported the growth of a biofilm, whose biomass increased steadily during the first 72 h, but did not support planktonic growth. OD₆₀₀, optical density at 600 nm. (B to D) Top view (at a 15^o angle) (top panels) and side view (bottom panels) projections generated by CSLM of 48-h wild-type (B) or pilin-deficient mutant (D) biofilms formed on an Fe(III) oxide-coated surface or of a wild-type biofilm on a control coverslip without the Fe(III) oxide coating (C). Green indicates live cells, and red indicates dead cells. Yellow regions are areas where the two dyes overlap. The substratum (coverslip) is at the bottom. Bars, 20 μm.

FIG. 2.

Average biomasses of mature wild-type (WT) and pilin-deficient mutant (PilA−) biofilms formed on Fe(III) oxide coatings [Fe(III)] or glass surfaces. Where indicated, the soluble electron acceptor fumarate also was present in the growth medium. The results are averages for triplicate samples from two independent experiments. OD₆₀₀, optical density at 600 nm.

FIG. 3.

CSLM analyses of wild-type (A and C) and PilA⁻ (B and D) biofilms formed on Fe(III) oxide coatings (A and B) or glass coverslips (C and D) in medium with fumarate. Green indicates live cells, and red indicates dead cells. Yellow indicates dye overlap. The images are three-dimensional top views (top panels) and side views (bottom panels) reconstructed from the fluorescence patterns of the series of two-dimensional optical sections collected by CSLM. Bars, 20 μm.

FIG. 4.

Autoagglutination phenotypes of a wild-type strain (WT), a pilus-deficient mutant (PilA⁻), and a genetically complemented mutant (pRG5-pilA) grown under pilus-inducing conditions (25°C). The results are the averages for triplicate samples from two independent experiments. A600, absorbance at 600 nm.