Assessing inhibitory activity of probiotic culture supernatants against Pseudomonas aeruginosa: a comparative methodology between agar diffusion, broth culture and microcalorimetry

Cell free supernatants (CFS) obtained from probiotic species are routinely used to preliminary investigate the antimicrobial activity of potential probiotic isolates by the agar well diffusion and broth culture. Both methods have documented limitations. In this work, the potential of isothermal microcalorimetry (IMC), a technique based on the measurement of heat produced by growing bacteria was used to investigate the antimicrobial effects of two commercial probiotic species (Lactobacillus acidophilus LA-5((R)) and Bifidobacterium lactis BB-12((R))) and one reference strain (Bifidobacterium bifidum ATCC 11863) against Pseudomonas aeruginosa NCIMB 8628 using unmodified, neutralised and heat-treated CFS. P. aeruginosa was inhibited in growth by the unmodified CFS of all the species. No inhibitory activity was recorded for neutralised CFS of all the species using the agar well diffusion assay. Plate count during co-incubation of P. aeruginosa with the neutralised CFS of all the species showed no inhibition. However, IMC data showed significant inhibition with neutralised CFS obtained from the two Bifidobacterium species suggesting presence of other non-acidic inhibitory compounds in the CFS. The results in this work demonstrated that IMC has potential in probiotic bioassay as it has the capability to record in real-time and capture even subtle effects, which could be unnoticed with traditional assays.