Molecular probe on pathogensAugust 7, 2019
Modern clinical practices in pathogen detection are shifting from microbiology tests to molecular systems. Increasingly, this shift in molecular diagnostics away from laborious and time- consuming microbial assays is re-writing even gold-standard pathogen detection. However, microbiology will always play an important role in complementing and supplementing newer molecular methods. The increase in demand for molecular assays is primarily driven by the need for faster results. Usually, molecular diagnostics have faster turnaround times, resulting in clinically actionable results in as little as a few hours. The fast turnaround for results plays an important role in specific conditions like antibiotic resistance crisis due to prolonged exposure to empiric broad-spectrum antibiotics. In such instances, microbial methods typically take several days to generate the results, whereas the ever-evolving molecular assays allow clinical laboratories to deliver test reports within a day. This approach can ideally give proper identification of pathogen, followed by specific analyses of resistance markers which can enable faster targeted therapy for the patients without complicating the existing resistance. The discovery of molecular markers has also led to the development of better test panels for antibiotic resistance, co-infections and other co-morbidities. The past two decades have witnessed a rapid development of multiplex assay systems which are highly sensitive and quantitative. A few of the new technologies include point-of-care (POC) and low-cost DNA amplification devices like polymerase chain reaction (PCR) machines. Significant technology consolidation is happening quickly with manufacturers creating automated laboratory molecular tests with several add-on features. These advances will improve clinical outcomes and satisfy doctors and laboratories. In India, with its large patient numbers with high prevalence of infections including hospital-acquired infections, these technologies will play a major role in healthcare.
One of the recent studies done by World Health Organization (WHO), along with the United Nations Children’s Fund (UNICEF), World Bank and United Nations Development Program (UNDP), evaluated several technologies for infectious diseases, including those for low-resource settings. The three broad categories of technologies were 1) classical assays like microscopy and culture 2) biochemical, immunological and calorimetric assays 3) advanced biotechnology technologies like sequencing, genotyping, microarray and other nanotechnology-based assays. Each of these assay systems have their own advantages and disadvantages or limitations. However, given limitations such as resources, new technologies may give a better handle on managing the early diagnosis and clinical implementations. For example, take the case of tuberculosis (TB), which, according to a WHO global report (http://apps.who.int/iris/bitstream/ 10665/75938/1/9789241564502_eng.pdf), infected 8.7 million in 2011, out of which 1.4 million died. A lack of adequate diagnostic methods, especially in cases of co-infection of HIV and TB, is a major concern, along with drug resistance. Even though traditional methods continue to be useful, the implementation of new technologies can help tackle these diseases better, as in the case of improved light emitting diode microscopy as recommended by WHO. In cases of multidrug resistance and drug susceptibility tests, several immunological techniques like enzyme-linked immunosorbent assay (ELISA), were not really successful due to several reasons. New and advanced, DNA-based molecular line probe assay (LPA), which is a polymerase chain reaction (PCR) based reverse hybridization, was very specific (>99%) and sensitive (>97%). This technology also does not need a viable pathogen, making it safer to perform the assay without the need for extensive infrastructure. The major limitation is that one can perform LPA on a single mutation, whereas no single mutation can be attributed to drug resistance. This suggests that there is a need for complementary technologies. Real-time PCR (RTPCR) is also one of the advanced, rapid DNA-based assay system. Newer methods that are fully automated and easy-to-use have helped make drug resistance results available with almost 100% specificity within 3-5 hours. Next Generation Sequencing (NGS) has revolutionized several assays and is now in several clinical practices. This technology, however, needs a larger clinical validation, more so in countries like India, with Indian bio-specimens.
Newer techs on horizon
Other major challenges in tackling infectious diseases are bio-specimen collection systems, pre-processing and assay readiness. Clinical laboratories in majority of developing countries or in low resource areas are not equipped to handle the highly pathogenic or virulent strains. So, sample collection systems are crucial. Technical advances in these aspects have been very slow and depend on high-class and costly sample collection systems. One major difficulty in this area is the specific infrastructure needed for pre-processing and post-processing assays. If the sample collection system can neutralize the bio-specimen, these steps could be potentially made simpler and easier to perform. An important step towards developing technologies which are tailored for this requirement are projects promoted by Indian Council of Medical Research and Department of Biotechnology, Government of India. We may soon see a new horizon of technological advances being made in India which can revolutionize this aspect of healthcare.
The author is medical scientist and former director of SGRF, Bangalore