Point-of-care fluorescence imaging reveals extent of bacterial load in diabetic foot ulcers

Abstract

Elevated levels of bacteria, including biofilm, increase the risk of chronic wound infection and inhibit healing. Addressing asymptomatic high bacterial loads is challenged by a lack of clinical terminology and diagnostic tools. This post-hoc multicenter clinical trial analysis of 138 diabetic foot ulcers investigates fluorescence (FL)-imaging role in detecting biofilm-encased and planktonic bacteria in wounds at high loads. The sensitivity and specificity of clinical assessment and FL-imaging were compared across bacterial loads of concern (10⁴ -10⁹ CFU/g). Quantitative tissue culture confirmed the total loads. Bacterial presence was confirmed in 131/138 ulcers. Of these, 93.9% had loads >10⁴ CFU/g. In those wounds, symptoms of infection were largely absent and did not correlate with, or increase proportionately with, bacterial loads at any threshold. FL-imaging increased sensitivity for the detection of bacteria across loads 10⁴ -10⁹ (P < .0001), peaking at 92.6% for >10⁸ CFU/g. Imaging further showed that 84.2% of ulcers contained high loads in the periwound region. New terminology, chronic inhibitory bacterial load (CIBL), describes frequently asymptomatic, high bacterial loads in diabetic ulcers and periwound tissues, which require clinical intervention to prevent sequelae of infection. We anticipate this will spark a paradigm shift in assessment and management, enabling earlier intervention along the bacterial-infection continuum and supporting improved wound outcomes.

Keywords: bacterial load; diabetic foot; fluorescence imaging; infection; wound healing.

FIGURE 1

(A) Prevalence of red and/or cyan fluorescence and delayed healing at each log increase in bacterial load (10⁴–>10⁸ CFU/g). (B) Proportion of DFUs exhibiting delayed healing that were positive for red and/or cyan fluorescence at each bacterial threshold. Delayed healing = delayed healing beyond expectation, identified if the wound area had not reduced by at least 30% during the prior 4 weeks of care

FIGURE 2

(A) Receiver operator characteristic (ROC) curves for each CSS in the IWGDF diabetic foot infection classification (pain, erythema, local warmth, local swelling, and purulent discharge) and delayed healing beyond expectation plotted across each of the bacterial load thresholds assessed (>10⁴–>10⁸ CFU/g). (B) ROC curve of combined IWGDF criteria (2 or more criteria considered positive for infection) compared with combined IWGDF+FL and FL alone at >10⁴ CFU/g. Inset shows combined (2 or more) IWGDF ROC curve. Diagonal grey dotted line denotes ‘line of chance’

FIGURE 3

Fluorescence imaging of DFUs shows high prevalence of asymptomatic bacterial loads and high prevalence of bacterial loads in the periwound area. Clinicians classified wounds as positive (CSS+) or negative (CSS‐) for clinical signs and symptoms of infection. Bacterial loads in all cases shown exceeded 10⁵ CFU/g (confirmed by microbiological analysis). In each of these cases, red or cyan fluorescence indicative of elevated bacterial load (>10⁴ CFU/g) was detected (red arrows)

FIGURE 4

Chronic inhibitory bacterial load on the bacterial‐infection continuum. Based on the International Wound Infection Institute (IWII) 2022 wound infection continuum. ‘CSS’ denotes clinical signs and symptoms of chronic wound infection, as identified by the IWII (e.g., delayed healing, hypergranulation, erythema, local warmth, bleeding, swelling)