Rapid detection and respirometric profiling of aerobic bacteria on panels of selective media

Abstract

Aims: To evaluate high-throughput optical oxygen microrespirometry for selective detection and predictive identification of aerobic bacteria. Methods and Results: Using GreenLight probe, standard 384-well plates and time-resolved fluorescence reader, a representative panel of 16 partially selective media and 9 aerobic bacteria (Escherichia coli, Bacillus cereus, Staphylococcus aureus, Campylobacter jejuni, Yersinia enterocolitica, Pseudomonas aeruginosa, Streptococcus pyogenes, Salmonella typhimurium and Listeria innocua) were analysed. For each medium, bacterial strain and dilution, growth profiles were recorded, from which calibrations, doubling/generation times and growth patterns in different media were determined. Analytical performance, selectivity and general usability of the method were assessed, and mixed bacterial cultures were analysed. Conclusion: The microrespirometry platform facilitates simple, real-time detection and predictive identification of aerobic bacteria by looking at the patterns of their growth and respiration in several media and determining their growth and doubling times. Significance and Impact of the Study: The new screening method can be used for routine microbiological analysis and testing of aerobic bacterial cultures as well as complex food, environmental and clinical samples.

Biological detection by optical oxygen sensing

“…Doubling times can be determined by analysing the sample at two or more dilutions, and used for predictive identification of microbial species. 137 Yeast and fungi are also detectable using this method, though their respiration can be significantly slower than for bacteria. 138 Metabolic and chemical interactions between different organisms is an emerging area, for example in gut health, (neuro)gastroenterology, studies of biogeochemical communities in soil.…”

“…Nonetheless, in a recent study, a panel of 9 common species of aerobic bacteria was investigated by high-throughput microrespirometry in 384-well plates with 16 partially selective media. 137 For each medium and bacterial strain, growth profiles were recorded at different dilutions, and standard curves, doubling times and growth patterns in different media determined. Selective, sensitive and rapid (one day experiment) determination of bacteria in pure cultures and simple mixtures was thus demonstrated as a proof of concept.…”

“…For example, they can be used as part of the pre-enrichment and/ or selective enrichment steps, to identify negative samples and exclude them from further testing, or combined with traditional tests with chromogenic metabolic substrates. 137 Coupling with optical sensors for pH, CO 2 or ammonia, which operate in a similar format as the O 2 sensor 104 can also improve the selectivity of such microbial assays.…”

Bacterial Respiration Used as a Proxy to Evaluate the Bacterial Load in Cooling Towers

“…By measuring the consumption or production of a metabolite, these types of measurements link metabolic activity to abundance. Bacterial respiration is an established method in monitoring food spoilage [31][32][33] as well as within environmental analysis [34][35][36], but has so far not been intensively studied as an option within cooling tower monitoring. In situ measurements showed that the reservoir water was fully air-saturated, implying that measuring oxygen consumption, and therefore aerobic metabolism, should be a good proxy for the total bacterial load.…”

A sensor‐based system for rapid on‐site testing of microbial contamination in meat samples and carcasses

“…Optical O 2 sensing is based on dynamic (collisional) quenching of phosphorescence, which produces robust changes in sensor signal upon transition from high (air‐saturated) to low (depleted by respiration) levels of O 2 (Papkovsky & Dmitriev, 2013). This change in phosphorescence over time can be monitored and has been used to study pure bacterial cultures (Jasionek et al, 2013), crude food homogenates (Hempel et al, 2011), small organisms (O'Mahony et al, 2005) and antimicrobial agents (Elisseeva et al, 2020). The benefits of optical oxygen respirometry is that it provides contactless high throughput analysis of microbial growth through oxygen consumption/respiration, and this allows for significant automation and sensitivity, simple set‐up and quantitative real‐time readout (Elisseeva et al, 2020; Papkovsky & Dmitriev, 2013).…”