Home 5 Clinical Diagnostic Insider 5 Advancements in Antimicrobial Susceptibility Testing: Part 2

Advancements in Antimicrobial Susceptibility Testing: Part 2

by | Jan 24, 2024 | Clinical Diagnostic Insider, Emerging Tests-dtet

Several methods are being developed that improve the ability to rapidly identify organisms and report susceptibility results.

With the exception of marginal improvements, the current standard method for antimicrobial susceptibility testing has remained stagnant for decades. Emerging technologies are being applied to microbiological practices to quickly change that paradigm. Patients and laboratory personnel alike stand to benefit significantly during this evolution.

Genotypic and phenotypic technologies

Antimicrobial susceptibility testing (AST) has long been performed by phenotypic assessment. The phenotype are those characteristics that cover morphology, biochemical and physiologic properties that ultimately determine how an organism will behave and react to an antimicrobial.

More recently, there has been a notable shift toward complementary genotypic AST assessment. Genotypic assessment has become popularized through the use of PCR technology. The use of next-generation sequencing (NGS) has also been introduced into the foray though adoption is limited due to the high complexity, technical expertise, and laborious procedures needed.

Genotypic AST provides rapid determination of common genetically-based resistance traits, generating clinically actionable information. Phenotypic determination, often resulting days later, is still needed as not all resistance characteristics are genetically encoded.

What’s notable is that the current evolution of AST is coming predominantly in the form of phenotypic determination with potential to displace today’s phenotypic and genotypic options. Here are some of the key technologies that lab leaders may want to keep an eye on:


Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) took the microbiology world by storm just over a decade ago. MALDI focuses on using proteomics to determine the identification of an organism. It offers a rapid and cost-effective replacement to traditional biochemical microbial identification.1

MALDI is now being assessed for phenotypic and genotypic AST. One such method includes using the proteomic profile of well-known resistant clonal groups, such as Methicillin-resistant Staphylococcus aureus (MRSA) and Bacteroides fragilis.2 Other studies have shown the ability to detect plasmids encoding for antibiotic resistance traits, like the blaKPC plasmid that confers production of a carbapenemase.3

However, MALDI for AST requires further study, with current AST application only reported in research settings.

Fluorescence in situ hybridization

Newest to the field of FDA approved AST comes the Accelerate Pheno system from Accelerate Diagnostics. This system utilizes fluorescence in situ hybridization for organism identification combined with direct microscopic colony assessment for morpho-kinetic changes. For positive blood cultures, the system can provide organism identification in as little as 90 minutes and AST within seven hours.4

Rapid determination of organism ID and AST leads to timely escalation and/or de-escalation of antibiotics. This is a drastic improvement to current methods requiring 60 minutes for genotypic assessment and an additional 18-24 hours for phenotypic AST.

Limitations of the Accelerate Pheno System include the limited targets available in the select target organisms. Target limitations can cause false negative findings, identification to only the genus level, and limitations on panel antibiotics. Additionally, polymicrobial infections can prove problematic for AST determination, requiring further phenotypic AST of additional organisms.5 Finally, some very major errors have been noted in other studies, therefore drug/bug combinations require extra scrutiny or confirmatory testing via a different method.

Microfluidics and microscopy

Perhaps the most surprising new advancement in AST comes from the deployment of microfluidics and microscopy technologies.

Pattern Bioscience is charting new territory with their Digital Culture technology.6 With precise microfluidics, individual bacterial cells combine with chamber specific reagents. This enables single cell microscopy measurement of each droplet combined with artificial intelligence to assess the microbial response. With both metabolic indicators and antimicrobials present in the various droplets, simultaneous metabolic identification and phenotypic susceptibility can be determined by the AI.

Several vendors are now competing in the space of microfluidics with digital microscopy. QuantaMatrix offers blood culture medium inoculated into a 96-well plate combined with agar, broth, and direct automated microscopic assessment.7 Another promising microfluidic technology utilizes measurement of dissolved oxygen levels within chambers containing an antibiotic.8

Much of the benefit being brought forth with microfluidics is the advancement in rapid AST, with these newer systems allowing AST results in two to six hours from positive blood culture.

What’s next?

A new evolution in the advancement of rapid AST is on the horizon. PCR has heralded the current era of genotypic AST but its time in the spotlight may be passing. Though limitations in each new phenotypic technology exist, microfluidics with digital microscopy boasts a most promising future.


    1. https://journals.asm.org/doi/10.1128/jcm.01452-16

    1. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8388893/

    1. https://www.frontiersin.org/articles/10.3389/fcimb.2020.572909/full

    1. https://acceleratediagnostics.com/technology/mca/?ref=product

    1. https://journals.asm.org/doi/10.1128/jcm.01166-17

    1. https://pattern.bio/pattern-bioscience-selected-as-winner-of-adlm-disruptive-technology-award/

    1. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9229829/

    1. https://pubs.acs.org/doi/10.1021/acssensors.1c00020

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