Modelling hospital disinfectant against multi-drug-resistant dry surface biofilms grown under artificial human sweat

Abstract

Dry surface biofilms (DSB) have been found abundantly across hospital surfaces within intensive care units and may explain how nosocomial pathogens can remain virulent and persist on surfaces for extended periods. Testing standards governing the performance of disinfectant products employ planktonic models under routine growth conditions, which are known to be less tolerant than their biofilm counterpart. To evaluate biofilm models cultured under artificial human sweat (AHS), a source of nutrient expected on touch surfaces, to assess the antimicrobial performance of common cleaning agents, including a quaternary ammonium, hydrogen peroxide and active chlorine. 5 single-species biofilms, using pathogenic bacteria such as Acinetobacter baumannii, Pseudomonas aeruginosa, Staphylococcus aureus and Enterococcus faecalis, were generated on stainless steel substrates using a sedimentation protocol under both AHS and nutrient rich conditions for a direct comparison of phenotypic tolerance. The biofilm models were grown over 5 days followed by desiccation cycles, before being submerged into the disinfectant solutions for up to 25 minutes. Epifluorescence (EF) microscopy using LIVE/DEAD™ stain was used to visualise microcolony viability. The results revealed biofilms cultured under AHS exhibited a greater antimicrobial tolerance and reduced speed of kill for all cleaning agents compared with the routine media; an average reduction of 72.4% vs 96.9%, respectively. EF microscopy revealed traces of viable bacteria across all coupons after disinfection indicating a potential opportunity for regrowth and recontamination. The notable difference in biocidal performance between the two growth conditions highlights potential pitfalls within current antimicrobial test standards, and the importance of accurate representation of the microbial challenge.