Antibiotic-Resistance Monitoring for the 21st Century
How Droplet Digital PCR technology is paving the way for the next generation of antibiotic-resistance monitoring
The issue of antibiotic resistance is a critical concern in clinical settings, with the World Health Organization ranking it among the top 10 global health issues in 2021. The misuse and potential overuse of antibiotics have resulted in widespread and sometimes unchecked antibiotic resistance, costing the United States alone over $4 billion annually. However, methods of assessing antibiotic resistance need to be faster and more consistent. Here, I explore new solutions for monitoring antibiotic resistance, the innovations driving this revolution, and real-life examples of modern antibiotic resistance monitoring.
Classically, researchers assess antibiotic resistance by culturing a pathogen and examining its response to various drugs. Although this method is highly effective, it is time-consuming, taking several days or even weeks. This delay is sometimes too long for patients needing medical attention. More modern approaches to antibiotic resistance detection include molecular-based methods, such as Sanger sequencing and RT-PCR. These techniques monitor antibiotic resistance genes, which indicate a bacterial strain has the genetic capabilities to survive exposure to specific antibiotics. While Sanger sequencing and RT-PCR are faster than bacterial culturing, they can underestimate antibiotic resistance. Additionally, conducting the tests and interpreting the results can require specialized technical expertise and skilled personnel.
Recently, some researchers have adopted a new approach to tracking bacterial resistance: Droplet Digital PCR (ddPCR). Like traditional RT-PCR, ddPCR technology is a rapid, molecular-based test that can detect antibiotic resistance genes quickly. However, ddPCR assays do not require a standard curve, which can introduce user bias and require more technical understanding to analyze. Instead, ddPCR technology partitions samples into 10,000 discrete droplets where individual PCR reactions occur, giving ddPCR assays high precision and sensitivity. Because ddPCR technology requires no standard curve and can be run simply by any lab member, it is also accessible and approachable.
Several examples of recent research illustrate the impact of ddPCR technology on antibiotic resistance detection and monitoring. In the first example, researchers measured the prevalence of antibiotic-resistant genes and the likelihood of those genes transferring to other bacteria, using a ddPCR assay (1) – a proof of concept approach that provides a highly scalable, quantitative, rapid, and less expensive approach to measuring the quantity and mobility of potential antibiotic-resistance genes.
In the second example, researchers focused on sepsis – a dangerous condition that requires rapid treatment with the correct antibiotic. To ensure the fastest and most accurate response, the authors developed a diagnostic ddPCR assay that could identify bacterial and antibiotic resistance genes in a single straightforward and quick step (2). The test was rapidly deployable within a single hour and provided the high sensitivity level needed to support sepsis patients.
Finally, a series of additional publications highlight how ddPCR technology empowers teams to detect antibiotic resistance faster and with great certainty. Researchers have used ddPCR assays to detect antibiotic resistance-related mutations in Mycobacterium tuberculosis (MTB) patients before they received antibiotic treatment, suggesting these mutations may have arisen spontaneously (3). In another MTB-related article,the team used ddPCR assays to simultaneously identify three mutations with high clinical significance throughout MTB treatment, giving patients and physicians greater confidence in their choice of treatment regimen. Beyond MTB, ddPCR assays are also helping teams identify and quantify antibiotic resistance in Helicobacter pylori and Campylobacter jejuni (4) infections, a leading cause of human bacterial gastroenteritis.
Through these examples and more, it’s clear that ddPCR technology is paving the way for the next generation of efficient antibiotic resistance monitoring. With high precision, rapid turnaround time, and unmatched sensitivity, ddPCR technology empowers scientists and clinical teams to detect antibiotic resistance faster and more accurately than ever before.
- MC Barron et al, “Quantification of the mobility potential of antibiotic resistance genes through multiplexed ddPCR linkage analysis,” FEMS Microbiol Ecol, 99, 4 (2023). DOI: 10.1093/femsec/fiad031.
- TJ Abram et al, “Rapid bacterial detection and antibiotic susceptibility testing in whole blood using one-step, high throughput blood digital PCR,” Lab Chip, 20, 3, 477–489 (2019). DOI: 10.1039/c9lc01212e.
- YW Aung et al., “Heteroresistance of Mycobacterium tuberculosis in the Sputum Detected by Droplet Digital PCR,” Biology, 12, 4, 525 (2013). DOI: 10.3390/biology12040525.
- Y Luo et al., “Droplet Digital PCR-Based Detection and Quantification of GyrA Thr-86-Ile Mutation Based Fluoroquinolone-Resistant Campylobacter jejuni,” Microbiol Spectr, 10, 2, e0276921 (2022). DOI: 10.1128/spectrum.02769-21.
Biopharma Segment Manager, Digital Biology Group, Bio-Rad