Proteomic Analysis Reveals Differential Protein Expression by Bacillus cereus during Biofilm Formation

Abstract

Bacillus cereus, a dairy-associated toxigenic bacterium, readily forms biofilms on various surfaces and was used to gain a better understanding of biofilm development by gram-positive aerobic rods. B. cereus DL5 was shown to readily adapt to an attached mode of growth, with dense biofilm structures developing within 18 h after inoculation when glass wool was used as a surface. Two-dimensional gel electrophoresis (2DE) revealed distinct and reproducible phenotypic differences between 2-and 18-h-old biofilm and planktonic cells (grown both in the presence and in the absence of glass wool). Whereas the 2-h-old biofilm proteome indicated expression of 15 unique proteins, the 18-h-old biofilm proteome contained 7 uniquely expressed proteins. Differences between the microcolony (2-h) proteome and the more developed biofilm (18-h) proteome were largely due to up-and down-regulation of the expression of a multitude of proteins. Selected protein spots excised from 2DE gels were subjected to N-terminal sequencing and identified with high confidence. Among the proteins were catabolic ornithine carbamoyltransferase and L-lactate dehydrogenase. Interestingly, increased levels of YhbH, a member of the sigma 54 modulation protein family which is strongly induced in response to environmental stresses and energy depletion via both B and H , could be observed within 2 h in both attached cells and planktonic cultures growing in the presence of glass wool, indicating that this protein plays an important role in regulation of the biofilm phenotype. Distinct band differences were also found between the extracellular proteins of 18-h-old cultures grown in the presence and in the absence of glass wool.

Protein Expression by Streptococcus mutans during Initial Stage of Biofilm Formation

“…These data confirm those obtained with Pseudomonas spp. and B. cereus showing that surface adherence results in altered protein expression (6,23,32). An altered RRS of 33 protein spots, 27% of the number of protein spots analyzed, in these 2-h-old biofilms is a proportion similar to that in a 3-day-old S. mutans biofilm, where 19% (135 of 694) of the analyzed protein spots were modulated (34).…”

“…Alginate production (algC) was also up-regulated within 15 min of cell attachment to a glass surface (9). It has been shown similarly that in Bacillus cereus 15 uniquely expressed proteins were synthesized within 2 h after surface attachment, compared to only 7 uniquely expressed proteins in 18-h-old biofilm compared to planktonic controls (23).…”

Modulation of virulence factors in Francisella tularensis determines human macrophage responses

“…YhbH has been implicated in virulence of Bacillus cereus for its roles in sporulation [24] and biofilm formation [25]. In E. coli, YhbH is known to play a role in quorum sensing [26] and stabilizing ribosomes [36].…”

“…One of the proteins identified was intracellular growth locus C (IglC), a known F. tularensis virulence factor [13][14][15][16][17][18]. Also included were aconitase, which is part of the TCA cycle and has been associated with Staphylococcus aureus virulence [23], and a homolog of a σ 54 modulation protein (YhbH), which has known functions in sporulation [24], biofilm formation [25], and quorum sensing [26]. Other identified proteins included fumerate hydratase, a protein involved in the TCA cycle, and a heat shock protein (Hsp20).…”

“…6). We selected acnA, iglC, and yhbH for this test as each has been associated with bacterial virulence [14,17,[23][24][25][26]. The abundance of these genes' transcripts was measured by quantitative PCR using 16S rRNA as an internal reference.…”