Diagnostic Technology: Molecular or Not Molecular?

May 5, 2017

Asking the above question is like asking whether the computer has changed the world, whether you love microbes, or whether Louis Pasteur had something to do with the glass of wine in your hand: there is a clear, right answer. Molecular technologies have not only revolutionized the field of microbiology, they have also transformed medicine, patient care, infection control and prevention, and overall hospital operation. There is no doubt that laboratories are adopting more and more molecular technologies to provide better quality of service, and, in particular, to diagnose emerging infectious diseases.

The Emergence of Multidrug-Resistant Organisms

Soon after the introduction of the first effective antimicrobials—the sulfonamides and penicillin, in the 1930s and 40s—scientists and clinicians realized that pathogens can develop tolerance or resistance to these therapeutic agents. The competition between humans (to discover new antimicrobial agents) and pathogens (to develop drug resistance) has persisted since then. In recent decades, pathogens have evolved into multidrug-resistant strains, and have become a global public health and economic burden. Just to name a few examples: methicillin resistance caused by mecA and mecC genes; vancomycin resistance caused by vanA and vanB genes; carbapenem resistance caused by carbapenemases KPC, NDM, OXA, IMP, VIM; emergence of highly resistant Candida auris and ciprofloxacin-resistant Shigella ... the list goes on and on. Multidrug-resistant organisms (MDROs) are often associated with increased lengths of stay, costs, and mortality.

In addition to the direct issue of patient care, antibiotic-resistant infections confer a financial cost. The overall economic burden of antibiotic resistance was estimated to be at least €1.5 billion in 2007 in Europe and $55 billion in 2000 in the US, including patient and hospital costs. The development of multidrug resistance is mostly attributable to the widespread availability and overuse of antimicrobial agents among humans and animals. Imperfect diagnostic technology for new or emerging organisms and resistance mechanisms, as well as suboptimal infection control, contribute to the spread of MDROs. Furthermore, rapid globalization with international population mobility has directly contributed to the emergence and spread of diseases and multidrug-resistant organisms (MDROs). There is an urgent need to better detect and control the spread of MDROs.

How Can Molecular Technologies Help?

The advantages of molecular technologies are high sensitivity/specificity, fast turnaround time (TAT), and high throughput. Life-threatening conditions such as bloodstream infection (BSI) and meningitis can be diagnosed in a timely manner. Pathogens are identified from direct specimens or positive blood culture samples using the sample-to-result approach. To ensure prompt optimization of antibiotic regimen and infection control measures, a technologist may report out a positive nucleic acid amplification test (NAAT, a molecular test) result as a critical value while awaiting the growth of the pathogen on culture media as confirmation. This is particularly important for infections caused by MDROs that necessitate contact/droplet precautions. Let’s take methicillin-resistant S. aureus (MRSA) as an example. The TATs for MRSA detection using conventional workflow and specific chromogenic medium are 48 hours and 24 hours, respectively, from a positive blood culture. With NAATs, the TAT time is reduced to 1 – 2.5 hours, resulting in improved clinical outcomes and reduced hospital cost.

NAATs can also be used to diagnose infections that are of epidemiological importance and that require isolation precautions. A negative result of NAAT targeting Mycobacterium tuberculosis from a sputum specimen is highly predictive of 3 negative acid-fast smear results, thus allowing earlier removal of patients from airborne isolation precautions. The high negative predictive value of NAAT for detection of toxigenic Clostridium difficile reduces the use of empiric antibiotics by 54%, and may allow removal of patients with diarrhea from isolation precautions. Importantly, these measures are most effective when accompanied with the implementation of an antibiotic stewardship program, as recently highlighted by the Infectious Diseases Society of America and the Society for Healthcare Epidemiology of America. Effective notification of the critical result to the care provider, as well as education on the utility and limitations of new technologies, have also been shown to be critical to ensure optimal use of the information by care providers. The partnership between the laboratory, infectious diseases specialists, pharmacists, and infection preventionists is essential to make the best use of molecular technologies to improve patient care.

Bacterial infections aren’t the only infectious diseases to benefit from molecular testing. Conventional viral culture for detecting influenza virus and other respiratory viruses, such as respiratory syncytial virus (RSV), adenovirus, and parainfluenza virus, is laborious, slow, and not sensitive. The cytopathic effect of conventional culture that indicates the presence of virus may take several days. The long TAT of this methodology contradicts the optimal administration window of antiviral drugs: oseltamivir should be taken within 48 hours of flu-like symptom onset. The patient may also be unnecessarily admitted to the hospital while awaiting the test result. Rapid influenza diagnostic tests (RIDTs) based on antigen detection, on the other hand, have a fast TAT, but their sensitivity is only about 50–70%. When influenza is prevalent (40%), the negative predictive value (NPV) of these RIDT is only about 70 – 80%. In other words, a negative test result does not necessarily rule out infection. NAATs have significantly shortened the time to diagnose respiratory viral diseases. The targets of such assays range from influenza virus alone to a panel of viruses that commonly cause respiratory illness. Some of the NAATs that can be performed as point-of-care tests have turnaround times as short as 30 minutes,  with sensitivity and specificity outperforming those of conventional viral culture and rapid antigen assay. Early detection of influenza by NAAT is associated with significantly lower odds ratios for admission, length of stay, antimicrobial duration, and number of chest radiographs. In addition to seasonal influenza, the investigation of recent viral disease outbreaks has also relied upon the application of NAATs. Cross-reactivity between closely related virus species may hinder the use of enzyme immunoassay (EIA) to detect antibodies for disease diagnosis. NAATs are sensitive and specific to diagnose infections by Ebola virus and Zika virus, especially soon after symptom onset when the viral load is high. For infectious gastroenteritis, certain NAATs are now able to detect astrovirus and sapovirus, two pathogens previously not commonly tested. Early diagnosis of a viral etiology helps reduce the unnecessary use of antibiotics, and it  is anticipated that more viral targets will be included in future molecular assays.

Limitations of Molecular Technologies

While molecular technologies improve diagnostic performance, turnaround time, and patient outcomes, it is also important to understand their limitations. NAATs can only detect the targets designed as part of the assay – a new test would need to be developed to detect any new/rare strains or resistance markers. For example, the outbreak of carbapenemase-producing Klebsiella pneumoniae associated with endoscopic retrograde cholangiopancreatography was due to OXA-232 , a carbapenemase that was detected by whole genome sequencing but is not targeted by existing diagnostic NAATs. A positive result of a NAAT for toxigenic C. difficile might indicate colonization rather than infection;  misinterpretation of the result could lead to unnecessary treatment. High-resolution molecular sequencing technology, if not used carefully, may create nomenclatural instability and confusion. Finally, the adoption of molecular technologies should not replace the conventional culture system, as the isolation of pathogens is still critical for conducting epidemiological studies and for optimizing antimicrobial therapy in patients.

The above post reflects the thoughts of its author, Dr. Jacky Chow, and not the American Society for Microbiology.

Author: Siu-Kei (Jacky) Chow

Siu-Kei (Jacky) Chow
Siu-Kei (Jacky) Chow is the Technical Director of Infectious Diseases Diagnostics at MultiCare Health System in Washington. He is a diplomate of the American Board of Medical Microbiology. He is interested in emerging infectious diseases, antimicrobial resistance, workflow optimization, and drawings of microorganisms.