Chlorhexidine Hydrochloride

ST11 is the most common lineage among carbapenem-resistant Klebsiella pneumoniae (CRKP) infections in Asia. Diverse morphotypes resulting from genetic mutations are associated with significant differences in microbial characteristics among K. pneumoniae isolates. Here, we investigated the genetic determinants and critical characteristics associated with distinct morphotypes of ST11 CRKP. An ST11-KL47 CRKP isolate carrying a pLVPK-like virulence plasmid was isolated from a patient with a bloodstream infection; the isolate had the "mcsw" morphotype. Two distinct morphotypes ("ntrd" and "msdw") were derived from this strain during in vitro passage. Whole genome sequencing was used to identify mutations that cause the distinct morphotypes of ST11 CRKP. Transmission electron microscopy, antimicrobial susceptibility tests, growth assays, biofilm formation, virulence assays, membrane permeability assays, and RNA-seq analysis were used to investigate the specific characteristics associated with different morphotypes of ST11 CRKP. Compared with the parental mcsw morphotype, the ntrd morphotype resulted from mutation of genes involved in capsular polysaccharide biosynthesis (wza, wzc, and wbaP), a result validated by gene knockout experiments. This morphotype showed capsule deficiency and lower virulence potential, but higher biofilm production. By contrast, the msdw morphotype displayed competition deficiency and increased susceptibility to chlorhexidine and polymyxin B. Further analyses indicated that these characteristics were caused by interruption of the sigma factor gene rpoN by insertion mutations and deletion of the rpoN gene, which attenuated membrane integrity presumably by downregulating the phage shock protein operon. These data expand current understanding of genetic, virulence, and antimicrobial resistance characteristics associated with distinct morphotypes in ST11 CRKP.

This research first investigated the effect of mesoporous silica nanoparticles (nMS) carrying chlorhexidine and silver (nMS-nAg-Chx) on periodontitis-related biofilms. This study aimed to investigate (1) the antibacterial activity on Porphyromonas gingivalis (P. gingivalis) biofilm; (2) the suppressing effect on virulence of P. gingivalis biofilm; (3) the regulating effect on periodontitis-related multispecies biofilm.

Silver nanoparticles (nAg) and chlorhexidine (Chx) were co-loaded into nMS to form nMS-nAg-Chx. Inhibitory zone test and minimum inhibitory concentration (MIC) against P. gingivalis were tested. Growth curves, crystal violet (CV) staining, live/dead staining and scanning electron microscopy (SEM) observation were performed. Biofilm virulence was assessed. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and Quantitative Real Time-PCR (qPCR) were performed to validate the activity and composition changes of multispecies biofilm (P. gingivalis, Streptococcus gordonii and Streptococcus sanguinis).

nMS-nAg-Chx inhibited P. gingivalis biofilm dose-dependently (p<0.05), with MIC of 18.75 µg/mL. There were fewer live bacteria, less biomass and less virulence in nMS-nAg-Chx groups (p<0.05). nMS-nAg-Chx inhibited and modified periodontitis-related biofilms. The proportion of pathogenic bacteria decreased from 16.08 to 1.07% and that of helpful bacteria increased from 82.65 to 94.31% in 25 μg/mL nMS-nAg-Chx group for 72 h.

nMS-nAg-Chx inhibited P. gingivalis growth, decreased biofilm virulence and modulated periodontitis-related multispecies biofilms toward healthy tendency. pH-sensitive nMS-nAg-Chx inhibit the pathogens and regulate oral microecology, showing great potential in periodontitis adjunctive therapy.