Antibiotic Susceptibility Testing Techniques for Live Biotherapeutic Products

Antibiotic resistance is defined as the genetic ability of bacteria to code for resistance genes that fake the survival inhibition effects of potential antibiotics. It can enter the bacterial genome through natural recombination and integration, or it can be acquired through horizontal gene mutation events. Antimicrobial susceptibility testing is widely used clinically to determine antibiotic resistance profiles of bacterial isolates, guide antibiotic treatment decisions, and predict therapeutic outcomes. Creative Biolabs is one of the USA’s leading providers to pharma, bioscience and contract research organisations for flexible and professional contract development of live biotherapeutic products (LBP). We offer our technical expertise and experience to provide traceable, customizable solutions for antibiotic susceptibility testing (AST).

Overview of AST

Minimum inhibitory concentrations (MICs) of various AST are categorized by various international agencies. These MIC guidelines determine susceptibility to antibiotics. The Clinical and Laboratory Standards Institute (CLSI) provides the most popular guidelines based on pharmacokinetic-pharmacodynamic (PK-PD) characteristics and resistance mechanisms. In the presence of antibiotics, antibiotic sensitivity is determined by monitoring bacterial growth (or growth inhibition), metabolism, and viability, or by using molecular assays to identify resistant biomarkers. Currently, microbiological techniques, such as biometrics, and screening methods, including disk diffusion, antibacterial gradient diffusion, broth dilution, and cell culture, are some of the common methods used by health care settings to measure antibiotic effectiveness.

Highlights of different techniques for AST applications.Fig.1 Highlights of different techniques for AST applications. (Behera, 2019)

Antibiotic Susceptibility Testing Techniques at Creative Biolabs

Phenotypic methods, through their fundamental interaction with the physical environment, provide more avenues for emerging engineering techniques to apply to them to improve their efficiency and versatility. Traditional AST is based on bacteria growing on solid agar plates or liquid media, with or without antibiotics. Phenotypic methods are by far the most commonly used and include microbroth dilution, antibiotic gradient, and disc diffusion. The test is performed on an agar plate by determining the inhibition zone after culture, while in the liquid-based method, changes in optical density are measured.

Molecular tests for AST rely on the identification of biomarkers indicative of bacterial genera, species, and strains or antibiotic resistance, including genetic materials, proteins, enzymes, and metabolites. Antibiotic resistance genes (e.g., mecA gene for MRSA; vanA/B genes for VRE; TEM, SHV, OXA, and CTX-M gene families for ESBLs; and KPC, NDM, OXA-48, VIM, and IMP genes for CREs), as well as antibiotic-responsive mRNA transcripts, have been widely used as markers for rapid AST.

With the increasing clinical demand for rapid AST, various new AST techniques based on optical imaging, micro-channel resonators, and other biosensors have been pursued. The imaging-based AST technologies have greatly reduced the time and comparable accuracy and reasonable reproducibility compared to traditional and commercial methods.

Regulation of defined LBPs is concerned with the process as well as specific components of the product, increasing oversight of the characteristics of the component such as the genetic identity of the strain and the potential for virulence or antibiotic resistance genes to be transferred to other members of the microbiome. Creative Biolabs is a premier provider of customized solutions to LBP development. We have extensive experience in assessing the safety of LBP candidate strains. Please contact us to discuss your requirements and pricing information.


  1. Behera, B.; et al. Emerging technologies for antibiotic susceptibility testing. Biosensors and Bioelectronics. 2019, 142: 111552.

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