The influence of the lipopolysaccharide (LPS) chain length on bacterial adhesion was investigated by measuring the collision efficiencies of three Escherichia coli K12 strains, each having a different length LPS, to silica glass beads in column tests (macroscale tests). Nanoscale interactions between the bacteria and a silicon nitride tip were probed utilizing atomic force microscopy (AFM). Adhesion results based on column tests indicated that collision efficiencies of the three bacteria were not consistently correlated to LPS length. Under conditions of low ionic strength (1 mM NaCl), collision efficiencies increased with LPS length for the three strains of E. coli. However, if cells were fixed with glutaraldehyde (2.5%), the strain with the shortest LPS chain had the greatest adhesion, while the bacterium with the mid-length LPS had the least adhesion to glass beads. Collision efficiencies increased when the solution ionic strength was increased from 1 to 100 mM as expected, and in most cases glutaraldehyde treatment also increased adhesion. AFM force curves failed to distinguish the adhesion characteristics of these bacteria measured in column tests, as all AFM force curves on the bacteria were identical. Changes in adhesion were also not predictable by more conventional measurements of bacterial properties based on ζ potential or contact angle. These results suggest that the LPS molecule length is not the sole determinant of adhesion of the three E. coli strains in porous media and that AFM force curve analysis, zeta potential, or contact angle data cannot yet be used to fully predict adhesion of these three strains to glass beads.

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For biofuel applications, synthetic endoglucanase E1 and xylanase (Xyn10A) derived from Acidothermus cellulolyticus were transiently expressed in detached whole sunflower (Helianthus annuus L.) leaves using vacuum infiltration. Three different expression systems were tested, including the constitutive CaMV 35S-driven, CMVar (Cucumber mosaic virus advanced replicating), and TRBO (Tobacco mosaic virus RNA-Based Overexpression Vector) systems. For 6-day leaf incubations, codon-optimized E1 and xylanase driven by the CaMV 35S promoter were successfully expressed in sunflower leaves. The two viral expression vectors, CMVar and TRBO, were not successful although we found high expression in Nicotiana benthamiana leaves previously for other recombinant proteins. To further enhance transient expression, we demonstrated two novel methods: using the plant hormone methyl jasmonic acid in the agroinfiltration buffer and two-phase optimization of the leaf incubation temperature. When methyl jasmonic acid was added to Agrobacterium tumefaciens cell suspensions and infiltrated into plant leaves, the functional enzyme production increased 4.6-fold. Production also increased up to 4.2-fold when the leaf incubation temperature was elevated above the typical temperature, 20C, to 30C in the late incubation phase, presumably due to enhanced rate of protein synthesis in plant cells. Finally, we demonstrated co-expression of E1 and xylanase in detached sunflower leaves. To our knowledge, this is the first report of (co)expression of heterologous plant cell wall-degrading enzymes in sunflower.