Abstract
Successful treatments against bacterial infections depend on antimicrobial susceptibility testing (AST). However, conventional AST requires more than 24 h to obtain an outcome, thereby contributing to high patient mortality. An antibiotic therapy based on experiences is therefore necessary for saving lives and escalating the emergence of multidrug-resistant pathogens. Accordingly, a fast and effective drug screen is necessary for the appropriate administration of antibiotics. The mixed pathogenic nature of infectious diseases emphasizes the need to develop an assay system for polymicrobial infections. On this basis, we present a novel technique for simultaneous and quantitative monitoring of co-cultured microorganisms by coupling optical diffusometry with bead-based immunoassays. This simple integration simultaneously achieves a rapid AST analysis for two pathogens. Triple color particles were simultaneously recorded and subsequently analyzed by functionalizing different fluorescent color particles with dissimilar pathogen-specific antibodies. Results suggested that the effect of the antibiotic, gentamicin, on co-cultured Pseudomonas aeruginosa and Staphylococcus aureus was effectively distinguished by the proposed technique. This study revealed a multiplexed and time-saving (within 2 h) platform with a small sample volume (~0.5 μL) and a low initial bacterial count (50 CFU per droplet, ~105 CFU/mL) for continuously monitoring the growth of co-cultured microorganisms. This technique provides insights into timely therapies against polymicrobial diseases in the near future.
Highlights
In clinical practices, the polymicrobial nature of many diseases or infections was confirmed and gained attention from the community[8,9,10]
Our previous study showed that the diffusometry-enabled antimicrobial susceptibility testing (AST) was convenient and reliable[23]
We further expanded the technique to AST for co-cultured bacteria by drawing on the features of bead-based immunoassays
Summary
The polymicrobial nature of many diseases or infections was confirmed and gained attention from the community[8,9,10]. Numerous techniques based on morphological analysis[13], fluorescence intensity[14,15], asynchronous magnetic bead rotation[16,17], dielectrophoresis[18], Raman-enhanced spectroscopy[19,20], atomic force microscopy[21] and surface plasmon resonance imaging[22] are developed to improve AST These techniques use direct quantification factors (including counting by image analysis[13], fluorescence intensity[14,15] and bacterial proliferation volume16) and indirect factors (including morphology[13,18] medium viscosity[16], bacteria-disrupted secretion[19,20] and bacterial fluctuations originating from metabolism21,22) to efficiently determine the antibiotic susceptibility profiles of bacteria.
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