Laser speckle imaging for visualization of hidden effects for early detection of antibacterial susceptibility in disc diffusion tests

Abstract

Rapid identification of effective antibiotic treatment is crucial for increasing patient survival and preventing the formation of new antibiotic-resistant bacteria due to preventative antibiotic use. Currently utilized “gold standard” methods require 16–24 h to determine the most appropriate antibiotic for the patient’s treatment. The proposed technique of laser speckle imaging with subpixel correlation analysis allows for identifying dynamics and changes in the zone of inhibition, which are impossible to observe with classical methods. Furthermore, it obtains the resulting zone of inhibition diameter earlier than the disk diffusion method which is recommended by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). These results could improve mathematical models of changes in the diameter of the zone of inhibition around the disc containing the antimicrobial agent, thereby speeding up and facilitating epidemiological analysis.

Tools for classification of growing/non-growing bacterial colonies using laser speckle imaging

“…Moreover, like previous experiments, the bacterial activity displayed a trend over time-peaking at around 4-5 h and almost ceasing by 19 h. More details on the analysis of the zone of inhibition using subpixel correlation methods can be found in the cited literature (Balmages et al, 2023c;Lihacova et al, 2023).…”

“…From previous work, it is known that bacterial activity is detectable 3-4 h post-inoculation using the subpixel correlation method (Balmages et al, 2021b). Therefore, the antibiotic disk was introduced into the Petri dish 4-4.5 h after bacterial inoculation (Balmages et al, 2023c). This approach allowed for realtime monitoring of the antibiotic's effect on an already growing bacterial population.…”

Development of Rapid Disk Diffusion Device Using Laser Speckle Formation Technology for Rapid Antimicrobial Susceptibility Testing

“…Thus, it is possible to precisely establish the form and position of the disk, allowing accurately quantify the diameter of the inhibitory zone. This undoubtedly represents an improvement above the previous similar methods [ 10 ]. Besides, the method is not influenced by the mechanical movements as image shift does not interfere with spatial correlation [ 17 ].…”

“…This would enhance treatment efficacy and impede the emergence of AMR. Furthermore, previous investigations on using laser speckle time series images for the dynamic visualization of inhibition zones by antimicrobials demonstrated the ability to detect these areas earlier than conventional approaches [ 10 ]. In these investigations, laser was employed to illuminate the sample dish and capture the speckle pattern that is reflected from the surface of the dish for measurements.…”

Determination of operating parameters of fungal growth signals analyzed by laser speckle contrast imaging

“…Laser speckle imaging used to identify submicron objects in detail and proposed additional sensitive subpixel correlation method allows detection of small changes, and not observed by the standard method phenomena. This non-invasive, non-contact optical method has been used in our previous studies to describe microbial growth activity [2] and detect hidden effects in bacterial behavior [3,4]. Agar media with inoculated bacteria (such as: E.coli, S.aureus, V.natriegens, etc.)…”