Simple, reusable, solid-state system for measuring total (aerobic) viable count, TVC, using the micro-respirometry method (μRM)

Liquid micro-respirometry, Liq-μRM, is a popular, more rapid alternative method for measuring Total Viable Count of aerobes, i.e., TVC, units: colony forming units (CFU)/mL, to the traditional, time-consuming, plate counting method. Liq-μRM is based on monitoring the consumption of dissolved O₂ in a liquid growth medium as a function of incubation time. However, in Liq-μRM, the O₂ indicator is difficult to make, non-reusable and not well suited for use in well-plates, due to agitation issues. In this work a new, Solid-State micro-respirometry method, i.e., SS-μRM, is described based on a solid, rather than liquid, growth medium, in which the consumption of O₂ in the headspace is monitored, using a 3D printed O₂ indicator set in the lid of a small Petri dish or well plate containing the solid growth medium. The performance of the SS-μRM in a small Petri dish is compared with that of a commercial Liq-μRM system, using Escherichia coli, i.e., E. coli, as the test bacterium over the range 10¹ – 10⁸ CFU/mL and demonstrated to be equivalent. The same system is used to measure the TVC of Pseudomonas aeruginosa, P. aeruginosa, and Enterobacter cloacae, E. cloacae, over the range 10¹ – 10⁸ CFU/mL. When used to measure the TVC of E. coli, in a well plate, SS-μRM is shown to be insensitive to agitation, in contrast to Liq-μRM. The ability of the SS-μRM to address the main concerns regarding Liq-μRM are discussed.