Genotypic Methods for Antibiotic Susceptibility Testing

Genotypic susceptibility testing is the detection of resistance genes expressed in a specific organism by molecular methods. The antimicrobial sensitivity profile of the strain(s) present in the LBP is of the highest importance. For antibiotic sensitivity testing (AST), it is recommended to complement molecular methods with culture-based methods. Creative Biolabs is one of the USA's leading providers to pharma, bioscience, and contract research organizations for flexible and professional contract development of live biotherapeutic products (LBP). We offer our technical expertise and experience to provide traceable, customizable solutions for AST services with genotypic methods.

Overview of Genotypic AST

Genotypic ASTs are based on the detection of a specific genetic marker (plasmids, genes, or mutations) associated with resistance phenotypes by using common genetic tools (e.g., sequence-specific amplification by PCR, padlock probe-mediated rolling circle amplification, or whole-genome sequencing. Rapid detection of the presence (or absence) of a resistance gene may allow improved antimicrobial therapy independent of phenotypic susceptibility results. Currently, rather than replacing phenotypic susceptibility testing, genotypic testing supplements it. Molecular genotypic AST is the effective direct method that eliminates tedious bacterial cultures, long incubation, chances of contamination, and the spreading of deadly infections. The application of genotypic techniques to AST has been used to both help improve the time to results and, in some cases, the accuracy of AST. To date, however, molecular testing has been able to provide only a qualitative, not a quantitative, readout of resistance.

The basic workflow of molecular-based techniques for antimicrobial susceptibility testing.Fig.1 The basic workflow of molecular-based techniques for antimicrobial susceptibility testing. (Gajic, 2022)

Genotypic Methods for AST at Creative Biolabs

PCR, DNA microarray and DNA chips, and loop-mediated isothermal amplification (LAMP) are some of the genotypic techniques for the detection of antibiotic resistance.

With increased knowledge of the genetic basis of antibiotic resistance, PCR-based approaches have been developed for detecting the presence of genetic determinants of resistance to a variety of antibiotics for many different bacterial species. The main advantage of PCR-based methods is that they can be performed in a relatively short time and, in some cases, pure cultures are not required to use clinical samples. Thus, PCR has the potential to significantly reduce turnaround time and rapidly provide information on antibiotic resistance.

DNA microarrays are promising techniques for screening susceptibility. Microarray technology addresses the limitations of traditional drug resistance gene detection techniques because of its simplicity, specificity, and high throughput, allowing multiple specific genes to be detected in a single reaction.

SmartChip is the most frequently used platform for the analysis and profiling of ARGs. The SmartChip nano will platform can be used for large-scale gene expression studies, especially in the field of antimicrobial resistance. This technique has proven to be very cost-effective, as nano upgrade reactions can save large amounts of consumables and reagents and also allow more efficient use of existing samples.

Advantages and Disadvantages

Genotypic methods are generally attributed to the rapid, direct, sensitive, and specific detection of resistance genes, but they also suffer from drawbacks. These drawbacks include: (1) the individual antimicrobial agents to be tested need a specific assay for detection, (2) only potential/key resistance genes can be detected, which are often not relevant due to coincidental mutations, (3) the genetic mechanism/profile for the resistance of all bacteria is not yet defined, (4) the occurrence of false-positive results due to contamination of the test sample might be expected, (5) they require expensive reagents and machinery with specific maintenance conditions, and most importantly (6) all the tools have a prerequisite of skilled personnel.

Creative Biolabs specializes in custom services for LBP development. Our scientists are highly qualified with extensive experience and exceptional skills in the fields of probiotics and LBPs. Our goal is to provide our customers with affordable development services with reliable results. If you are interested in our AST services with genotypic methods, please contact us for further discussion.

Reference

  1. Gajic, I.; et al. Antimicrobial Susceptibility Testing: A Comprehensive Review of Currently Used Methods. Antibiotics. 2022, 11(4): 427.

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