Quorum quenching effect of cyclodextrins on the pyocyanin and pyoverdine production of Pseudomonas aeruginosa

Abstract

Various virulence determinants in Pseudomonas aeruginosa are regulated by the quorum sensing (QS) network producing and releasing signalling molecules. Two of these virulence determinants are the pyocyanin and pyoverdine, which interfere with multiple cellular functions during infection. The application of QS-inhibiting agents, such as cyclodextrins (CDs), appears to be a promising approach. Further to method development, this research tested in large-volume test systems the effect of α- and β-CD (ACD, BCD) at 1, 5, and 10 mM concentrations on the production of pyocyanin in the P. aeruginosa model system. The concentration and time-dependent quorum quenching effect of native CDs and their derivatives on pyoverdine production was tested in a small-volume high-throughput system. In the large-volume system, both ACD and BCD significantly inhibited pyocyanin production, but ACD to a greater extent. 10 mM ACD resulted in 58% inhibition, while BCD only ~40%. Similarly, ACD was more effective in the inhibition of pyoverdine production; nevertheless, the results of RMANOVA demonstrated the significant efficiency of both ACD and BCD, as well as their derivatives. Both the contact time and the cyclodextrin treatments significantly influenced pyoverdine production. In this case, the inhibitory effect of ACD after 48 h at 12.5 mM was 57%, while the inhibitory effect of BCD and its derivatives was lower than 40%. The high-level significant inhibition of both pyocyanin and pyoverdine production by ACD was detectable. Consequently, the potential value of CDs as QS inhibitors and the antivirulence strategy should be considered. KEYPOINTS: • Applicability of a simplified method for quantification of pyocyanin production was demonstrated. • The cyclodextrins significantly affected the pyocyanin and pyoverdine production. • The native ACD exhibited the highest attenuation in pyoverdine production.

Keywords: Pseudomonas aeruginosa; Cyclodextrins; Pyocyanin; Pyoverdine; Quorum quenching; Quorum sensing.

Fig. 1

Fluorescence intensity of the P. aeruginosa PAO1 cell suspension and its supernatant containing pyoverdine measured at different excitation and emission wavelength combinations

Fig. 2

Fluorescence intensity of the P. aeruginosa DSM 1117 cell suspension and its supernatant containing pyocyanin measured at different excitation and emission wavelength combinations

Fig. 3

Absorbance of the chloroform extracted P. aeruginosa DSM 1117 supernatant containing pyocyanin at 520 nm

Fig. 4

Modulation of relative pyocyanin production normalized to cell density by ACD and BCD parent CD molecules in the P. aeruginosa DSM 1117 large-volume model system. Statistical significance (p < 0.05) is marked by lowercase letters, where a indicates the smallest value. Values signed with the same letter indicate that there was no significant difference between them. Statistical analysis was carried out distinctively for ACD and BCD datasets. Data represent averages of three replicates

Fig. 5

Effect of increasing concentrations of ACD (a), QAACD (b), HPACD (c), and ACDPS (d) on pyoverdine pigment production of P. aeruginosa PAO1 after 24- and 48-h exposure time normalized to cell density. Statistical significance (p < 0.05) is marked by lowercase letters, where a indicates the smallest value. Values signed with the same letter indicate that there was no significant difference between them. Data represent averages of five replicates

Fig. 6

Effect of increasing concentrations of BCD (a), QABCD (b), HPBCD (c), BCDPS (d), RAMEB (e), and SBEBCD (f) on pyoverdine pigment production of P. aeruginosa PAO1 after 24- and 48-h exposure time normalized to cell density. Statistical significance (p < 0.05) is marked by lowercase letters, where a indicates the smallest value. Values signed with the same letter indicate that there was no significant difference between them. Data represent averages of five replicates