The biofilm inhibitor Carolacton inhibits planktonic growth of virulent pneumococci via a conserved target

Abstract

New antibacterial compounds, preferentially exploiting novel cellular targets, are urgently needed to fight the increasing resistance of pathogens against conventional antibiotics. Here we demonstrate that Carolacton, a myxobacterial secondary metabolite previously shown to damage Streptococcus mutans biofilms, inhibits planktonic growth of Streptococcus pneumoniae TIGR4 and multidrug-resistant clinical isolates of serotype 19A at nanomolar concentrations. A Carolacton diastereomer is inactive in both streptococci, indicating a highly specific interaction with a conserved cellular target. S. mutans requires the eukaryotic-like serine/threonine protein kinase PknB and the cysteine metabolism regulator CysR for susceptibility to Carolacton, whereas their homologues are not needed in S. pneumoniae, suggesting a specific function for S. mutans biofilms only. A bactericidal effect of Carolacton was observed for S. pneumoniae TIGR4, with a reduction of cell numbers by 3 log units. The clinical pneumonia isolate Sp49 showed immediate growth arrest and cell lysis, suggesting a bacteriolytic effect of Carolacton. Carolacton treatment caused a reduction in membrane potential, but not membrane integrity, and transcriptome analysis revealed compensatory reactions of the cell. Our data show that Carolacton might have potential for treating pneumococcal infections.

The Biofilm Inhibitor Carolacton Enters Gram-Negative Cells: Studies Using a TolC-Deficient Strain of Escherichia coli

“…The MIC of carolacton against E. coli TolC was in the same range as that reported by Jansen et al ( 19 ). For S. pneumoniae TIGR4, the MIC of carolacton was determined to be 0.06 µg/ml ( 24 ), similar to the value reported for E. coli TolC. In comparison to E. coli MG1655, E. coli TolC showed a strong increase in sensitivity (≥4-fold) to antibiotics from all functional groups.…”

“…During such screenings, the myxobacterial macrolide ketocarbonic acid carolacton was identified as a biofilm inhibitor ( 19 , 20 ). Its activity against clinically relevant streptococci was later analyzed in great detail ( 20 – 24 ). The exact molecular target of carolacton remains unknown, but the complete loss of biological activity of a carolacton epimer at C-9 [( S ) → ( R )] ( epi -carolacton) in Streptococcus mutans biofilms and planktonically growing Streptococcus pneumoniae cells ( 22 , 24 ) suggests an interaction of carolacton with a conserved cellular target ( 24 ).…”

Deep sequencing of biofilm microbiomes on dental composite materials

“…Carolacton was recently shown to specifically inhibit the essential folate-dependent enzyme FolD/MTHFD, which is present in all bacteria [26]. However, in direct tests, carolacton has only been shown to inhibit certain isolates of S. pneumoniae [27] and does not affect the growth of most bacteria, including S. mutans at neutral or higher pH [10]. Resistance mechanisms have evolved such as the alternative enzyme Fhs, which is especially widespread in anaerobic bacteria [28], and the efflux pumps which most bacteria possess [26,27,29].…”

“…However, in direct tests, carolacton has only been shown to inhibit certain isolates of S. pneumoniae [27] and does not affect the growth of most bacteria, including S. mutans at neutral or higher pH [10]. Resistance mechanisms have evolved such as the alternative enzyme Fhs, which is especially widespread in anaerobic bacteria [28], and the efflux pumps which most bacteria possess [26,27,29]. The strong effect of carolacton on S. mutans biofilm viability was always striking because it only occurs at low pH [30], reflecting the essential role of FolD in acid survival of S. mutans [31].…”

Macrocycles, the edge of drug‐likeness chemical space or Goldilocks zone?

“…The authors show biosynthesis, synthetic methods and describe biological activity. A notable example is carolacton 7 (Figure ) a highly potent bacterial biofilm inhibitor which was synthesized by the Wuest group in 14 steps . Although this is still a little long for a drug candidate, it represents a useful synthetic route that could with adaptations be utilized for analogue preparation.…”