Bioactive glass S53P4 eradicates Staphylococcus aureus in biofilm/planktonic states in vitro

Figures & data

Table 1. Test substance and exposure time.

Test substance	Priming time
BAG-S53P4 < 45 µm, TSB with 1% glucose/1% NaCl
Granule test
BAG - TSB, 100 mg/mL	2 days
Supernatant test
BAG - TSB, 100 mg/mL	2 days
BAG - TSB, 200 mg/mL	2 days
BAG - TSB, 400 mg/mL	2 days
BAG - TSB, 100 mg/mL	14 days
BAG - TSB, 200 mg/mL	14 days
BAG - TSB, 400 mg/mL	14 days
BAG - TSB, 100 mg/mL with ciprofloxacin 5 µg/mL	2 days
BAG - TSB, 200 mg/mL with ciprofloxacin 5 µg/mL	2 days
BAG - TSB, 400 mg/mL with ciprofloxacin 5 µg/mL	2 days
BAG - TSB, 100 mg/mL with ciprofloxacin 5 µg/mL	14 days
BAG - TSB, 200 mg/mL with ciprofloxacin 5 µg/mL	14 days
BAG - TSB, 400 mg/mL with ciprofloxacin 5 µg/mL	14 days
pH-adjusted (NaOH) TSB with 1% glucose/1% NaCl
pH-adjusted TSB, pH 7.0
pH-adjusted TSB, pH 9.7
pH-adjusted TSB, pH 10.7
pH-adjusted TSB, pH 11.7

All TSB with 1% glucose/1% NaCl solutions were from the Norwegian Veterinary Institute, Oslo, Norway.

BAG: bioactive glass; TSB: tryptic soy broth.

Figure 1. Biofilm-oriented antiseptics test (BOAT). Supernatant test: BAG 14-day priming time (a); BAG 2-day priming time (b). BAG concentration from left to right is 400 mg/mL, 200 mg/mL, 100 mg/mL, and 0 mg/mL; 3 wells per concentration. Rows 1–7: strains 1–7; row 8: negative control. Red formazan is a sign of viable cells. (96-well microtiter plate; Nunclon Delta Surface, Thermo Fischer Scientific).

Table 2. A Comparison of BAG effects versus control.

		BOAT			Bactericidal test		Confocal laser scanning microscopy with LIVE/DEAD staining
Test solution	n	Mean	±1 SD	p Value	Surviving strains/total no. strains	p Value	Ratio dead/total cell no.	±1 SD	p Value
Untreated bacterial strains (control)	11	2.884	0.11	–	11/11	–	0.190	0.01	–
Supernatant test
BAG-S53P4 < 45 µm, 100 mg/mL, 2 days	11	2.704	0.49	0.30	11/11	–	0.176	0.12	0.85
BAG-S53P4 < 45 µm, 200 mg/mL, 2 days	11	2.402	0.96	0.15	11/11	–	0.746	0.35	0.07
BAG-S53P4 < 45 µm, 400 mg/mL, 2 days	11	0.789	0.72	<0.001	8/11	0.08	0.931	0.21	0.01
BAG-S53P4 < 45 µm, 100 mg/mL, with ciprofloxacin, 2 days	10	1.635	0.86	0.001	10/10	–	0.811	0.32	0.047
BAG-S53P4 < 45 µm, 200 mg/mL, with ciprofloxacin, 2 days	10	0.754	0.80	<0.001	10/10	–	0.836	0.11	0.002
BAG-S53P4 < 45 µm, 400 mg/mL, with ciprofloxacin, 2 days	10	0.208	0.09	<0.001	4/10	0.01	1.099	0.07	<0.001
BAG-S53P4 < 45 µm, 100 mg/mL, 14 days	11	1.208	1.07	0.001	11/11	–	0.686	0.13	0.008
BAG-S53P4 < 45 µm, 200 mg/mL, 14 days	11	0.173	0.05	<0.001	3/11	0.005	0.958	0.26	0.015
BAG-S53P4 < 45 µm, 400 mg/mL, 14 days	11	0.152	0.03	<0.001	0/11	0.001	1.086	0.05	<0.001
BAG-S53P4 < 45 µm, 100 mg/mL, with ciprofloxacin, 14 days	10	1.163	0.44	0.002	10/10	–	0.871	0.13	0.003
BAG-S53P4 < 45 µm, 200 mg/mL, with ciprofloxacin, 14 days	10	0.257	0.10	<0.001	2/10	0.008	1.099	0.12	0.001
BAG-S53P4 < 45 µm, 400 mg/mL, with ciprofloxacin, 14 days	10	0.207	0.11	<0.001	0/10	0.002	1.129	0.12	0.001
Granule test
BAG-S53P4 < 45 µm, 100 mg/mL	11	N/A	N/A	N/A	0/11	0.001	1.046	0.11	0.001
pH-adjusted TSB
TSB with 1% glucose/1% NaCl, pH 9.7	11	2.915	0.11	0.567	11/11	–	0.236	0.05	0.17
TSB with 1% glucose/1% NaCl, pH 10.7	11	0.749	0.80	<0.001	6/11	0.025	0.731	0.20	0.02
TSB with 1% glucose/1% NaCl, pH 11.7	11	0.129	0.02	<0.001	0/11	0.001	1.170	0.04	<0.001

Significance calculated by paired t test or McNemar asymptotic test.

BAG: bioactive glass; BOAT: biofilm-oriented antiseptics test; N/A: not applicable; SD: standard deviation; TSB: tryptic soy broth.

Figure 2. Confocal laser scanning microscopy (CLSM) stacks of Staphylococcus aureus biofilm. 1) Control; pH-adjusted TSB: 2) pH 9.7; 3) pH 10.7; 4) pH 11.7; 5) Granule test; Supernatant test, 2-day exposure: 6) 100 mg/mL; 7) 100 mg/mL with ciprofloxacin; 8) 200 mg/mL; 9) 200 mg/mL with ciprofloxacin; 10) 400 mg/mL; 11) 400 mg/mL with ciprofloxacin; Supernatant test, 14-day exposure: 12) 100 mg/mL; 13) 100 mg/mL with ciprofloxacin; 14) 200 mg/mL; 15) 200 mg/mL with ciprofloxacin; 16) 400 mg/mL; 17) 400 mg/mL with ciprofloxacin. The units are in µm.