Abstract
Bacterial keratitis is a serious and vision-threatening condition in veterinary and human patients, one that often requires culture and susceptibility testing to adjust therapy and improve clinical outcomes. The present study challenges the antimicrobial susceptibility testing (AST) paradigm in ophthalmology, enabling more accurate in vitro-to-in vivo translation by incorporating factors normally present during host-pathogen interactions in clinical patients. Thirty bacteria (10 Staphylococcus pseudintermedius, 10 Streptococcus canis, 10 Pseudomonas aeruginosa) were isolated from canine patients with infectious keratitis. For each isolate, commercial plates (Sensititre™ JOEYE2) were used to assess the minimal inhibitory concentration (MIC) of 17 different antibiotics in the absence (0% albumin, control) or presence of canine albumin (0.01–2%). For Staphylococcus pseudintermedius, the experiment was repeated with actual tear fluid collected from canine eyes with ocular surface inflammation. Kruskal-Wallis, Wilcoxon signed rank test and Spearman's correlation tests were used for statistical analysis. Clinical outcomes were unfavorable in selected canine patients with bacterial keratitis (e.g., globe perforation, graft dehiscence) despite standard AST (i.e., 0% albumin in test medium) confirming that most corneal infections (93%) were susceptible to ≥1 topical antibiotics used at the initial visit. Albumin levels ≥0.05% increased MICs in a dose-dependent, bacteria-specific, and antibiotic-specific manner. No significant differences (P = 1.000) were noted in MICs of any antibiotic whether albumin or tear fluid was added to the Mueller-Hinton broth. Percent protein binding inherent to each antibiotic was associated with clinical interpretations (Spearman's rho = −0.53, P = 0.034) but not changes in MICs. Albumin in tears impacted the efficacy of selected ophthalmic antibiotics as only the unbound portion of an antibiotic is microbiologically active. The present findings could improve decision making of clinicians managing bacterial keratitis, reduce development of antimicrobial resistance, influence current guidelines set by the Clinical and Laboratory Standards Institute, and serve as a reference for bacteriological evaluations across medical fields and across species.
Highlights
Antimicrobial susceptibility testing (AST) determines the lowest concentration of an antimicrobial agent that prevents visible growth of a microorganism using an agar or broth dilution susceptibility test
Albumin solutions—The impact of albumin on minimal inhibitory concentration (MIC) was dependent on the protein concentration, the antibiotic, and the bacterial isolate
Antimicrobial therapy must be started on the basis of clinical and laboratory evaluation; clinical success can be hindered by several challenges such as the rise of antimicrobial resistance [6, 20,21,22] or progression of keratomalacia despite the appropriate use of antibiotics [23,24,25]
Summary
Antimicrobial susceptibility testing (AST) determines the lowest concentration of an antimicrobial agent that prevents visible growth of a microorganism using an agar or broth dilution susceptibility test. This information is used to define the isolate as either susceptible, intermediate or resistant based on clinical guidelines set by the Clinical and Laboratory Standards Institute (CLSI) [1]. Differences between in vitro testing and in vivo clinical responses to antimicrobial agents are well-documented in the scientific literature [3,4,5], contributing in part to suboptimal clinical outcomes and the emergence of multi-drug resistance [6]. The presence of albumin in tears could impact the antimicrobial efficacy of a given antibiotic as only the unbound fraction of an antibiotic is known to be microbiologically active [12, 13]
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