Silver nanoparticles (AgNPs) are potential antimicrobials agents, which can be considered as an alternative to antibiotics for the treatment of infections caused by multi-drug resistant bacteria. The antimicrobial effects of double and triple combinations of AgNPs, visible blue light, and the conventional antibiotics amoxicillin, azithromycin, clarithromycin, linezolid, and vancomycin, against ten clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) were investigated. The antimicrobial activity of AgNPs, applied in combination with blue light, against selected isolates of MRSA was investigated at 1/2–1/128 of its minimal inhibitory concentration (MIC) in 24-well plates. The wells were exposed to blue light source at 460 nm and 250 mW for 1 h using a photon emitting diode. Samples were taken at different time intervals, and viable bacterial counts were determined. The double combinations of AgNPs and each of the antibiotics were assessed by the checkerboard method. The killing assay was used to test possible synergistic effects when blue light was further combined to AgNPs and each antibiotic at a time against selected isolates of MRSA. The bactericidal activity of AgNPs, at sub-MIC, and blue light was significantly (p < 0.001) enhanced when both agents were applied in combination compared to each agent alone. Similarly, synergistic interactions were observed when AgNPs were combined with amoxicillin, azithromycin, clarithromycin or linezolid in 30–40 % of the double combinations with no observed antagonistic interaction against the tested isolates. Combination of the AgNPs with vancomycin did not result in enhanced killing against all isolates tested. The antimicrobial activity against MRSA isolates was significantly enhanced in triple combinations of AgNPs, blue light and antibiotic, compared to treatments involving one or two agents. The bactericidal activities were highest when azithromycin or clarithromycin was included in the triple therapy compared to the other antibiotics tested. A new strategy can be used to combat serious infections caused by MRSA by combining AgNPs, blue light, and antibiotics. This triple therapy may include antibiotics, which have been proven to be ineffective against MRSA. The suggested approach would be useful to face the fast-growing drug-resistance with the slow development of new antimicrobial agents, and to preserve last resort antibiotics such as vancomycin.

(1) Background: Hyaluronic acid (HA) is a polyanionic mucopolysaccharide extensively used in biomedical and cosmetic industries due to its unique rheological properties. Recombinant HA production using other microbial platforms has received increasing interest to avoid potential toxin contamination associated with its production by streptococcal fermentation. In this study, the Gram-negative strains Escherichia coli (pLysY/Iq), E. coli Rosetta2, E. coli Rosetta (DE3) pLysS, E. coli Rosetta2 (DE3), E. coli Rosetta gammiB(DE3)pLysS, and the Gram-positive Bacillus megaterium (MS941) were investigated as new platforms for the heterologous production of HA. (2) Results: The HA biosynthesis gene hasA, cloned from Streptococcus equi subsp. zoopedemicus, was ligated into plasmid pMM1522 (MoBiTec), resulting in pMM1522 hasA, which was introduced into E. coli Rosetta-2(DE3) and B. megaterium (MS941). The initial HA titer by the two hosts in the LB medium was 5 mg/L and 50 mg/L, respectively. Streptococcal hasABC and hasABCDE genes were ligated into plasmid pP_T7 (MoBiTec) and different E. coli host strains were then transformed with the resulting plasmids pP_T7hasABC and pP_T7hasABCDE. For E. coli Rosetta-gamiB(DE3)pLysS transformed with pP_T7hasABC, HA production was 500 ± 11.4 mg/L in terrific broth (TB) medium. Productivity was slightly higher (585 ± 2.9 mg/L) when the same host was transformed with pP_T7 carrying the entire HA operon. We also transformed B. megaterium (MS941) protoplasts carrying T7-RNAP with pP_T7hasABC and pP_T7hasABCDE. In comparison, the former plasmid resulted in HA titers of 2116.7 ± 44 and 1988.3 ± 19.6 mg/L in LB media supplemented with 5% sucrose and A5 medium + MOPSO, respectively; the latter plasmid boosted the titer final concentration further to reach 2476.7 ± 14.5 mg/L and 2350 ± 28.8 mg/L in the two media, respectively. The molecular mass of representative HA samples ranged from 10⁵ − 10⁶ Daltons (Da), and the polydispersity index (PDI) was <2. Fourier transform infrared spectroscopy (FTIR) spectra of the HA product were identical to those obtained for commercially available standard polymers. Finally, scanning electron microscopic examination revealed the presence of extensive HA capsules in E. coli Rosetta-gamiB(DE3)pLysS, while no HA capsules were produced by B. megaterium. (3) Conclusions: Our results suggested that Gram-positive bacteria are probably superior host strains for recombinant HA production over their Gram-negative counters. The titers and the molecular weight (MW) of HA produced by B. megaterium were significantly higher than those obtained by different E. coli host strains used in this study.