Multiplex PCR performed using ompA, csuE, and bla OXA-51-like as target genes [24] confirmed these differences (data not shown). Biofilm formation by A. baumannii clinical isolates The A. baumannii isolates belonging to the SMAL

clone were tested for click here their ability to form biofilm, measured as surface adhesion to polystyrene microtiter plates. Biofilm growth is considered an important factor for host colonization [25, 26] and for resistance to Tucidinostat datasheet environmental and cellular stresses [11]. Ability to form biofilm, measured as surface adhesion to polystyrene microtiter plates, was very similar for all A. baumannii isolates tested (data not shown); results shown throughout the paper refer to the A. baumannii isolate described in Line 22 of Table 1. This isolate PND-1186 mouse was considered

representative of the A. baumannii SMAL clone since it belongs to the main genotypic subgroup of the SMAL clone (Figure 1) and since it was the first A. baumannii to be isolated in this survey. Surface adhesion to microtiter plates by A. baumannii SMAL clone was determined in various growth conditions, comparing two growth temperatures (30°C vs. 37°C), and different growth media: the rich peptone-based LB medium, LB medium diluted 1:4 (LB1/4), the M9Glu/sup medium [[27], described in Methods], and the M9Suc/sup in which 0.2% sucrose was added as main carbon source instead of glucose. LB1/4 was tested since it was shown to promote production of adhesion factors in other Gram negative bacteria, such as Escherichia coli [28]. We found that biofilm formation by A. baumannii SMAL was strongly affected both by growth media and by temperature: indeed, while surface adhesion was very poor in LB medium at either 30°C or 37°C, it was clearly stimulated by growth in LB1/4, although only

at 30°C. Finally, growth in M9Glu/sup resulted in efficient surface adhesion both at 30°C and at 37°C, while growth mafosfamide in sucrose-based medium (M9Suc/sup) resulted in much lower levels (Figure 2A). The observation that growth temperature affects biofilm formation in the LB1/4, but not in sugar-based media such as M9Glu/sup, would suggest that this process could be mediated by different mechanisms and by different adhesion factors. Figure 2 A. Surface adhesion to polystyrene microtiter plates by A. baumannii SMAL clone. Black bars bacterial cultures grown in LB medium; light grey bars LB1/4 medium; white bars M9Glu/sup; dark grey bars M9Suc/sup. B. Binding of Calcofluor to A. baumannii SMAL clone grown in solid media. C. Inhibition of A. baumannii biofilm formation by cellulase treatment: circles, M9Glu/sup medium; diamonds, M9Suc/sup medium; squares, LB1/4 medium. The horizontal dotted line indicates the 50% inhibition mark. IC50′s values are indicated by vertical dotted lines. A major adhesion factor characterized in A. baumannii is represented by the csu pili described in the A. baumannii strain ATCC 19606 [17].