Over the past two decades, there has been an explosion in biomedical information and technology. Those of us who started our careers in the late 1990s and early 2000s must have found it exhilarating to witness this extra-ordinary change in every field of medicine, and this has translated into improved patient care. At the forefront of this Biomedical research revolution is Molecular Diagnostics, which has transformed many diagnostic algorithms across disciplines like Infectious diseases and Oncology; and opened other avenues of laboratory medicine like Proteomics & Genomics. As a representative of the 90s generation, I present this ‘walk down the memory lane’ with considerable excitement for these are heady times for laboratory medicine as a science, and Molecular Microbiology in particular.
While molecular diagnostic methods for diagnosing infectious diseases are gradually gaining popularity in India, it would be surprising for many of us to know that PCR as a technology has been in existence for more than 40 years. PCR technique was invented in 1983 by Dr Kary Mullis, for which he earned the Nobel Prize in the year 1993. It was in the year 1997, that Specialty Ranbaxy Limited (later known as SRL Diagnostics and now known as Agilus Diagnostics) offered the first ‘commercial’ PCR test in India. They offered TB-PCR to begin with and later added HIV, HBV and HCV PCRs.
In the 2000s, other referral labs followed suit and the PCR set-ups at the time, were prized possessions of a few large laboratories. The specimens for the PCR tests were transported from across the country to these referral centres in large cities and only a few labs could negotiate this logistical challenge. The years 2000 to 2010 also saw the advent of the Real time PCR machines (RTPCR as we know them) and commercial IVD approved PCR kits giving a further boost to the diagnostic applications, while the conventional gel-doc PCRs and ‘home-brew’ assays were restricted only to research laboratories. The test menu gradually expanded to include Cytomegalovirus (CMV), Dengue, Leptospira, to name a few, and PCR played a significant role in the H1N1 limited pandemic of 2009-10.
In the following decade (2010-19), Point-of-Care (POC) PCR platforms were introduced, further opening the door for wider acceptance and application of RT-PCR technology. Today, no diagnostic algorithm of Tuberculosis (TB) is complete without involving either of these POC platforms: GeneXpert CB-NAAT (Cepheid Diagnostics) and TrueNAT (Molbio Diagnostics). The availability of POC PCR platforms has breathed fresh life into the TB screening programs enabling the government to set ambitious targets for TB eradication. The challenge for wider applicability of the PCR platform across various infectious diseases persisted due to their perceived lack of commercial viability. The need of the hour, at that time, was to popularise the PCR technology amongst clinicians to increase the prescription base and for the diagnostic industry to add more PCR kits to the test menu for a viable purchase proposal.
Come 2020 and the pandemic changed it all!! There was an explosion of RT-PCR Laboratories across the country. They helped us negotiate the pandemic and popularised the RT-PCR like never before. Today, even a layperson has heard about RT-PCR, so-much-so that RTPCR is at the risk of being slotted as a ‘COVID test’ (Just like ELISA was synonymous with an HIV antibody test in the 1990s). The pandemic also highlighted to the policy makers and administrators the hitherto unexplored topic of ‘cost per reportable result’ in a batch processing technology which could be multiple times more than the ‘cost per test’ if the number of tests processed per batch is low. Post-Covid, there remains a network of under-utilised RT-PCR infrastructure across the country with challenges of test menu expansion due to lack of adequate test numbers for batch testing.
Thus, molecular diagnosis of infectious diseases is at a crossroads. On one hand, there is a rapidly expanding market for POC platforms, which are designed for faster results in low resource settings that receive around 1-2 tests at a time, but upscaling to multiple parameters and high-volume testing is difficult. On the other hand, there is an organically expanding market of ‘Syndromic PCR’ or Multiplex PCR that can detect multiple pathogens in one go. The need for syndromic PCR is especially felt in Critical care areas like ICUs & NICUs where time is of essence and clinicians would need multiple answers from a single specimen (for e.g., a respiratory syndromic panel detecting 20-30 probable respiratory pathogens from a sputum specimen in a single test or a Meningo-encephalitis panel detecting a plethora of probable pathogens from a CSF sample).
The Film Array technology is at the forefront of these multiplex PCR solutions with its first-mover’s advantage; but being cost intensive, has limited its reach to microbiology departments of large hospitals in metro cities. Post-Covid, the unutilised capacity on open RTPCR platforms is gradually being serviced by a slew of multiplex PCR kits from a variety of Indian manufacturers. Also in the fray, are specialised referral laboratories like MedGenome & Rivaara Labs that are tapping into this ‘latent demand’ of syndromic PCRs. The cost benefit ratio of this high value tests not only tilts the balance in favour of better and faster patient care; but also has cascading benefits on antimicrobial stewardship. Since more patients are administered appropriate antibiotics earlier and therefore discharged earlier, the hospital bed occupancy improves as well.
The advent of these Syndromic PCR options for Infectious Disease syndromes has opened a Pandora’s box with respect to existing diagnostic algorithms based on microscopy, culture, antibiotic sensitivity, and serology tests. While culture remains the gold standard in diagnosing Infectious Diseases, the inherent limitations of culture, like prior antibiotic usage and the fastidious nature of certain microbes, can be bridged using these Syndromic PCR panels. After all, when was the last time our clinician colleagues have seen, on a sputum culture report, a fastidious organism like Hemophilus influenzae? Or when did serology reports diagnose (in time), agents of atypical pneumonia like Legionella, Mycoplasma or Chlamydia? The traditional concept of implicating only one pathogen in an infection is being challenged as well. When interpreting a report with multiple pathogens for certain specimen types like sputum, the dilemma in the minds of clinicians is whether the reported pathogen is normal flora or a true pathogen. Co-infection with multiple pathogens is a reality that has been unearthed by molecular tests; one can find numerous reference articles on the internet depicting the same. The dilemma in minds of Clinical Microbiologists whilst reporting more than one pathogen, one of which is having a higher Ct value (>30), is whether to underplay this as a probable contaminant or report this as a probable pathogen that may be missed on culture due to phenotypic factors at play. Such a situation would call for a detailed discussion with the treating clinicians, making them aware of the alternate pathogen that may raise its ugly head as one treats the first.
In the fast-paced world of biomedical research, many new concepts and prototypes are being evaluated. A few amongst these technologies are able to sustain and create a niche for themselves. The thumb rule is simple, if it bridges the gap between the need and reality, the technology is here to stay. One such promising technology that has already started to make in-roads and has the capability to further revolutionise diagnostic algorithms in Infectious Disease diagnostics is the NextGen Sequencing (NGS). A major advantage of NGS over PCR is that it is not dependent on target-specific primers and can identify from a single specimen, all pathogens / organisms that are available in its genome library through Artificial Intelligence (AI) based algorithms. As always, there will be pros and cons with every novel technology that is introduced in the diagnostic settings. Prudent use of the tests by clinicians coupled with innovative interpretative strategies by clinical microbiologists to make the reports palatable & integral to clinical settings, will determine the true potential of these newer technologies.
Last but not the least, I would like to address the often-asked question, ‘Will the clinical microbiologist be redundant in the onslaught of AI powered newer molecular techniques?’ Well, as a representative of the 90s generation, I would like to highlight that the advent of computers at that time, did not take away jobs, but redefined many professions. Clinical microbiologists are thus uniquely placed in the midst of these developments. The researchers and applied science professionals can deliver novel and reproducible technologies, but it is the Clinical Microbiologists who can interpret the results of these technologies in view of the patient’s clinical scenario.
Heady times indeed for Molecular Microbiology!
