The Impact of Noncontact, Nonthermal, Low-Frequency Ultrasound on Bacterial Counts in Experimental and Chronic Wounds

Thomas Serena, MD, FACS; S. Kwon Lee, MS, FACS, FCCWS; Kan Lam, BS, RLAT; Paul Attar, PhD; Patricio Meneses, PhD; and William Ennis, DO

Pre- and post-treatment tissue biopsies were obtained by following a specified protocol and sent to an independent central laboratory (LabCorp, Burlington, NC) for quantitative analysis.

     Data collection and analysis. Data collection and documentation were performed by investigative site personnel. The study sponsor’s clinical personnel monitored data abstraction, completion of study documentation, and correctness of case report forms. Descriptive statistics were performed to summarize and compare baseline data with outcomes following 2 weeks of noncontact ultrasound therapy. Bacteria quantities were summarized in CFU/g of tissue.


     Depth of penetration. Nile red dye penetrated into the intact porcine skin samples 2.0 mm to 2.5 mm with noncontact ultrasound therapy compared with 0.05 mm to 0.07 mm with the sham therapy. The majority of the dye was concentrated in the epidermis and stratum corneum for both treatments. In wounded skin samples, Nile red dye penetrated 3 mm to 3.5 mm with noncontact ultrasound therapy and 0.35 mm to 0.50 mm with sham therapy. With noncontact ultrasound therapy, the dye penetrated into the reticular dermis but not into the underlying subcutaneous fat (see Table 1).

     In vitro bacteria reduction. Following the noncontact ultrasound treatment, the percentage of dead bacteria was 33%, 40%, and 27% for P. aeruginosa, E. coli, and E. faecalis, respectively (see Figures 2, 3a,b). The sham treatment resulted in no dead organisms (0%). Using this treatment protocol, noncontact ultrasound had little or no effect on MRSA (1% increase of live bacteria) and S. aureus (0% change).

     Scanning electron microscopic images of E. faecalis showed structural changes to the round shape of the bacteria with distinct cell wall punctures or wall destruction after noncontact ultrasound treatment compared with intact cell walls in sham-treated controls (see Figure 4).

     In vivo bacteria reduction. Histologically, no differences were noted between the noncontact ultrasound and sham treatment groups with regard to edema, granulation tissue formation, or the presence of eschar. All animals remained healthy and gained weight during the 7-day study. Both the noncontact ultrasound therapy and silver antimicrobial dressing resulted in an overall reduction of bacterial counts (see Table 2). Overall bacterial colony counts increased consistently over time in the moist control group; whereas, bacterial counts in sham therapy-treated wounds decreased or increased at various treatment times. However, the patterns for P. aeruginosa and S. aureus were different. Although noncontact ultrasound therapy resulted in reduced bacterial counts of both organisms during days 3 to 7, during that time, bacterial counts of the Gram-negative bacteria P. aeruginosa changed from 8 ± 0.73 to 5.8 ± 0.74 in the ultrasound, and from 5.7 ± 2. to 4.7 ± 0.85 in the silver antimicrobial dressing group. Given the small sample sizes (as low as n = 2), statistical comparisons between treatment groups were not performed.

     Clinical bacteria reduction study. Of the 18 patients with Stage III pressure wounds enrolled between November 2006 and April 2007, 11 completed baseline and post-treatment biopsies and were considered evaluable for the effectiveness analysis.


1. Mertz PM, Ovington LG. Wound healing microbiology. Dermatol Clin. 1993;11(4):739–747.
2. Ennis WJ, Meneses P. Wound healing at the local level: the stunned wound. Ostomy Wound Manage. 2000;46(1A suppl):39S–48S.
3. Carignan A, Allard C, Pepin J,Cossette B, Nault V, Valiquette L. Risk of Clostridium difficile infection after perioperative antibacterial prophylaxis before and during an outbreak of infection due to a hypervirulent strain. Clin Infect Dis. 2008;46(12):1838–1843.
4. Costerton JW. Cystic fibrosis pathogenesis and the role of biofilms in persistent infection. Trends Microbiol. 2001;9(2):50–52.
5. Robson MC, Edstrom LE, Krizek TJ, Groskin MG. The efficacy of systemic antibiotics in the treatment of granulating wounds. J Surg Res. 1974;16(4):299–306.
6. Ennis WJ, Mozen FPN, Massey J, Conner-Kerr T, Meneses P. Ultrasound therapy for recalcitrant diabetic foot ulcers: results of a randomized, double-blind, controlled, multicenter trial. Ostomy Wound Manage. 2005;51(8):24–39.
7. Ennis WJ, Valdes W, Gainer M, Meneses P. Evaluation of clinical effectiveness of MIST ultrasound therapy for the healing of chronic wounds. Adv Skin Wound Care. 2006;19(8):437–446.
8. Kavros SJ, Miller JL, Hanna SW. Treatment of ischemic wounds with noncontact, low-frequency ultrasound: the Mayo clinic experience, 2004-2006. Adv Skin Wound Care. 2007;20(4):221–226.
9. Kavros SJ, Liedl DA, Boon AJ, Miller JL, Hobbs JA, Andrews KL. Expedited wound healing with noncontact, low-frequency ultrasound therapy in chronic wounds: a retrospective analysis. Adv Skin Wound Care. 2008;21(9): 416–423.
10. Johns LD. Nonthermal effects of therapeutic ultrasound: the Frequency Resonance Hypothesis. J Athl Train. 2002;37(3):293–299.
11. Bertuglia S. Mechanisms by which low-intensity ultrasound improve tolerance to ischemia-reperfusion injury. Ultrasound Med Biol. 2007;33(5):663–671.
12. Waldrop K, Serfass A. Clinical effectiveness of noncontact, low-frequency, nonthermal ultrasound in burn care. Ostomy Wound Manage. 2008;54(6):66–69.
13. Young SR, Dyson M. The effect of therapeutic ultrasound on angiogenesis. esis. Ultrasound Med Biol. 1990;16(3):261–269.
14. Unger P. Low-frequency, noncontact, nonthermal ultrasound therapy; a review of the literature. Ostomy Wound Manage. 2008;54(1):57–60.
15. Mukherjee S, Raghuraman H, Chattopadhyay A. Membrane localization and dynamics of Nile Red: effect of cholesterol. Biochim Biophys Acta. 2007;1768(1):59–66.
16. Duc Q, Breetveld M, Middelkoop E, Scheper RJ, Ulrich MM, Gibbs S. A cytotoxic analysis of antiseptic medication on skin substitutes and autograft. Br J Dermatol. 2007;157(1):33–40.
17. Poon VK, Burd A. In vitro cytotoxity of silver: implication for clinical wound care. Burns. 2004;30(2):140–147.
18. Kavros SJ, Schenck EC. Use of noncontact low-frequency ultrasound in the treatment of chronic foot and leg ulcerations: a 51-patient analysis. J Am Podiatr Med Assoc. 2007;97(2):95–101.
19. Whitney J, Phillips L, Aslam R, et al. Guidelines for the treatment of pressure ulcers. Wound Repair Regen. 2006;14(6):663–679.
20. Serena TE, Robson MC, Cooper DM, Ignatius J. Lack of reliability of clinical/visual assessment of chronic wound infection: the incidence of biopsy-proven infection in venous leg ulcers. Wounds. 2006;18(7):197–202.
21. Field FK, Kerstein MD. Overview of wound healing in a moist environment. Am J Surg. 1994;167(1A suppl):2S–6S.

Post new comment

  • Web page addresses and e-mail addresses turn into links automatically.
  • Lines and paragraphs break automatically.
  • Use to create page breaks.

More information about formatting options

Enter the characters shown in the image.