Label-free Optical Biosensor to Monitor Antibiotic Resistance in Bacterial Pathogens

Introduction: Antibiotics are used for treatment of microbial diseases in humans and animals, and as growth promoting agents during animal meat production. Extensive and indiscriminate use of antibiotics resulted in the emergence of antibiotic resistance in microbes, and contributes to a large number of nosocomial or community acquired infections and mortalities. Current molecular and immunological assays to screen antibiotic resistance in pathogens are arduous, need technical expertise and resource to perform the analysis. We used light scattering device designated BARDOT (bacterial rapid detection using optical scattering technology) as a rapid on-plate antibiotics resistance screening tool.

Purpose: To test BARDOT as a screening tool for antibiotic resistance in bacterial pathogens.

Methods: BARDOT uses a red diode laser (635 nm, 1 mW) to illuminate bacterial colonies of 1.2 ± 0.1 mm diameter to generate the signature scatter patterns. Cultures of Salmonella, Staphylococcus, Enterococcus, Klebsiella and Acinetobacter (total strains, n = 15) were plated with and without antibiotics on nutrient agar plate (BHI, TSA, LB, and NB) for inclusive growth of all pathogens. Scatter patterns of colonies were acquired and images were analyzed using quantitative image classifier. The results were represented as positive predictive value (PPV). Quantitative reverse transcriptase PCR (qRT-PCR) was performed to confirm the expression of antibiotic resistance gene (aadA, mecA) in a dose dependent manner.

Results: BARDOT generated differentiating scatter pattern for bacterial colony grown on antibiotic containing nutrient agar plate compared to agar media without antibiotics in a dose dependent manner. Scatter pattern profile of colonies on plate containing antibiotic or without antibiotic were differentiated with >90% PPV. qRT-PCR results verified increased expression of antibiotic resistance and corroborated with the observed scatter patterns for the antibiotic treatment in a dose dependent manner. 

Significance: BARDOT could be used as a real-time, label-free on-plate colony screening tool for antibiotic resistance in bacterial pathogens.