The visualization of biofilms in chronic diabetic foot wounds using routine diagnostic microscopy methods.
Diabetic foot wounds are commonly colonised by taxonomically diverse microbial communities and may additionally be infected with specific pathogens. Since biofilms are demonstrably less susceptible to antimicrobial agents than are planktonic bacteria, and may be present in chronic wounds, there is increasing interest in their aetiological role. In the current investigation, the presence of structured microbial assemblages in chronic diabetic foot wounds is demonstrated using several visualization methods. Debridement samples, collected from the foot wounds of diabetic patients, were histologically sectioned and examined using bright-field, fluorescence, and environmental scanning electron microscopy and assessed by quantitative differential viable counting. All samples (n = 26) harboured bioburdens in excess of 5 log₁₀ CFU/g. Microcolonies were identified in 4/4 samples by all three microscopy methods, although bright-field and fluorescence microscopy were more effective at highlighting putative biofilm morphology than ESEM. Results in this pilot study indicate that bacterial microcolonies and putative biofilm matrix can be visualized in chronic wounds using fluorescence microscopy and ESEM, but also using the simple Gram stain.
Manchester Pharmacy School, The University of Manchester, Oxford Road, Manchester M13 9PT, UK.
Angela
0000-0003-0519-4556
A
Oates
Department of Medicine Manchester Royal Infirmary, The University of Manchester, Oxford Road, Manchester M13 9PT, UK.
Frank L
FL
Bowling
Department of Medicine Manchester Royal Infirmary, The University of Manchester, Oxford Road, Manchester M13 9PT, UK.
Andrew J M
AJ
Boulton
ConvaTec Ltd., Deeside CH5 2NU, UK.
Philip G
PG
Bowler
ConvaTec Ltd., Deeside CH5 2NU, UK.
Daniel G
DG
Metcalf
Manchester Pharmacy School, The University of Manchester, Oxford Road, Manchester M13 9PT, UK.
Andrew J
AJ
McBain
D018441:Biofilms / Q000254:growth & development*; D015169:Colony Count, Microbial; D003646:Debridement; D017719:Diabetic Foot / Q000150:complications*; D004755:Enterobacteriaceae / Q000254:growth & development / Q000302:isolation & purification / Q000502:physiology / Q000648:ultrastructure; D005840:Gentian Violet / Q000737:chemistry; D006090:Gram-Negative Bacteria / Q000254:growth & development / Q000302:isolation & purification / Q000502:physiology* / Q000648:ultrastructure; D016905:Gram-Negative Bacterial Infections / Q000150:complications / Q000175:diagnosis* / Q000382:microbiology / Q000473:pathology; D006094:Gram-Positive Bacteria / Q000254:growth & development / Q000302:isolation & purification / Q000502:physiology* / Q000648:ultrastructure; D016908:Gram-Positive Bacterial Infections / Q000150:complications / Q000175:diagnosis* / Q000382:microbiology / Q000473:pathology; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D050296:Microbial Viability; D008855:Microscopy, Electron, Scanning; D010619:Phenazines / Q000737:chemistry; D010865:Pilot Projects; D013194:Staining and Labeling; D040141:Staphylococcaceae / Q000254:growth & development / Q000302:isolation & purification / Q000502:physiology / Q000648:ultrastructure; D013289:Streptococcaceae / Q000254:growth & development / Q000302:isolation & purification / Q000502:physiology / Q000648:ultrastructure
C035863:Gram's stain; D010619:Phenazines; D005840:Gentian Violet