Effect of antimicrobial peptides on planktonic growth, biofilm formation and biofilm-derived bacterial viability of Streptococcus pneumoniae

Abstract

Streptococcus pneumoniae is a leading cause of pneumonia mortality globally. Pneumococcal disease is often associated with prolonged colonisation of hosts and this process is facilitated by biofilm formation that is largely resistant to conventional antibiotics. We investigated the effects of antimicrobial peptides (AMPs) lysozyme, lactoferrin, LL37 and a combination of all three on planktonic growth, biofilm formation and biofilm-derived bacterial viability by S. pneumoniae, serotype 23F. Planktonic growth and biofilm-derived bacterial viability were determined using standard colony-forming techniques, while biofilm formation was measured using a crystal violet based spectrophotometric method. Relative to controls, lysozyme significantly reduced biofilm formation (0.08 OD vs. 0.10 OD at 570 nm, p = 0.01), while LL37 and the AMP combination increased biofilm formation (0.14 OD vs. 0.10 OD at 570 nm, p = 0.01). The combination of AMPs significantly decreased planktonic growth (1.10 × 10⁸ colony-forming units per millilitres [CFU/mL] vs. 2.13 × 10⁸ CFU/mL, p = 0.02). Biofilm-derived bacterial viability was greatly reduced by exposure to a combination of AMPs (1.05 × 10⁵ CFU/mL vs. 1.12 × 10⁶ CFU/mL, p = 3.60 × 10⁻⁸). Streptococcus pneumoniae displays marked resistance to the individual AMPs. A combination of lysozyme, lactoferrin and LL37 effectively inhibited planktonic growth and biofilm-derived bacterial viability; however, persister cell growth was still evident after exposure.