Cells in shearable and nonshearable regions of Salmonella enterica serovar Enteritidis biofilms are morphologically and physiologically distinct.

Cellular morphology, exopolymer chemistry, and protein expression of shearable and nonshearable fractions of Salmonella enterica serovar Enteritidis biofilms were examined. Biofilms were grown at a laminar flow velocity of 0.07 cm.s-1 for ~120 h, resulting in biofilms with a thickness (mean +/- SD) of 43 +/- 24 microm. An empirically determined shear-inducing flow (1.33 cm.s-1) was then applied for 5 min, effectively reducing biofilm thickness by ~70% and leaving 13 +/- 6 microm of nonshearable material and allowing fractionation of biofilm material into shearable and nonshearable regions. In situ lectin binding analyses revealed that there was no significant difference in the exopolymer glycoconjugate composition of the shearable and nonshearable biofilm zones. Length to width indices of cells from nonshearable and shearable biofilm regions as well as planktonic cells from biofilm effluent and continuous culture were determined to be 3.2, 2.3, 2.2, and 1.7, respectively, indicating that the cells in the shearable fraction were morphologically more similar to planktonic cells than the cells in the nonshearable biofilm fraction. Enhanced expression of proteins involved in cold shock response, adaptation, and broad regulatory functions (CspA, GrcA, and Hns, respectively) in cells from the shearable region as well as protein translation and modification and enhanced expression of protein involved in heat shock response and chaperonin function (DnaK) in cells from the nonshearable region revealed that the physiological status of cells in the two biofilm regions was distinct. This was also reflected in the different morphologies of cells from the two biofilm zones. Stratified patterns of cell metabolism and morphology in biofilms, obtained using shear-induced biofilm fractionation, may yield important information of how cells of deeply embedded biofilm bacteria survive imposed conditions of stress such as treatment with antimicrobial agents or antibiotics.