Genomic evaluation of clinical samples and microbiomes have improved our understanding of the role of pathogens in clinical microbiology. Being highly sensitive, next generation sequencing (NGS) is the gold standard in pathogen identification and can overcome several limitations of current day diagnostics (1). A genomics-based approach for pathogen detection has the potential to be a one-test solution to identify several pathogens simultaneously. The utility of implementing genomics in infectious disease and particularly in critically ill patients has been explored but challenges remain (2). A diagnostic solution for the detection of pathogens needs to consider:<ol><li>Turnaround-time (TAT)</li><li>Cost</li><li>Comprehensivity</li></ol>A workflow that combined target enrichment, long-read sequencing and automated bioinformatics (Figure 1) could address these challenges and was demonstrated to simultaneously identify bacteria, fungi and antibiotic resistance genes (ARGs). This strategy could overcome the issues associated with host background DNA and error rates of long read sequencing to provide accurate detection of pathogens. When tested across standard organisms, clinical isolates and blood samples from patients with suspected bloodstream infections this workflow could successfully and reproducibly identify the relevant pathogen and ARG. A blinded comparison of various clinical isolates revealed a concordance of 0.83 at species level and 0.90 at the genus level compared to standard diagnostic methods. This test could be implemented on-site as a single assay and a TAT of 12 hrs was consistently achieved (3).Figure 1. Workflow for the simultaneous detection of bacteria, fungi and ARGs <img src='https://haystackanalytics.in/assets/blog/infexn-workflow.png'></img>Current methods used in clinical microbiology lack sensitivity and often rely on culture increasing TAT. Studies have already established an improved sensitivity and specificity of NGS compared with standard-of-care tests (4,5). Thus, a workflow for the direct-from-sample, on-site sequencing combined with automated genomics was demonstrated to be implementable, reproducible and feasible. Such an approach can be a viable option as the primary screening tool in clinical settings and promote a precision approach to antibiotic prescribing.REFERENCES:<ol><li>Gu W, Miller S, Chiu CY. Clinical Metagenomic Next-Generation Sequencing for Pathogen Detection. Annu Rev Pathol. 2019 Jan 24;14:319-338. doi:10.1146/annurev-pathmechdis-012418-012751. )</li><li>Rossen JWA, Friedrich AW, Moran-Gilad J; ESCMID Study Group for Genomic and Molecular Diagnostics (ESGMD). Practical issues in implementing whole-genome-sequencing in routine diagnostic microbiology. Clin Microbiol Infect. 2018 Apr;24(4):355-360. doi: 10.1016/j.cmi.2017.11.001. </li><li>Kuruwa S, Zade A, Shah S, Moidu R, Lad S, Chande C, Joshi A, Hirani N, Nikam C, Bhattacharya S, Poojary A, Kapoor M, Kondabagil K, Chatterjee A. An integrated method for targeted Oxford Nanopore sequencing and automated bioinformatics for the simultaneous detection of bacteria, fungi and ARG. J Appl Microbiol. 2024 Feb 12:lxae037. doi: 10.1093/jambio/lxae037. </li><li>Grumaz S, Grumaz C, Vainshtein Y, Stevens P, Glanz K, Decker SO, Hofer S, Weigand MA, Brenner T, Sohn K. Enhanced Performance of Next-Generation Sequencing Diagnostics Compared With Standard of Care Microbiological Diagnostics in Patients Suffering From Septic Shock. Crit Care Med. 2019 May;47(5):e394-e402. doi: 10.1097/CCM.0000000000003658. </li><li>Zhang Y, Lu X, Tang LV, Xia L, Hu Y. Nanopore-Targeted Sequencing Improves the Diagnosis and Treatment of Patients with Serious Infections. mBio. 2023 Feb 28;14(1):e0305522. doi: 10.1128/mbio.03055-22.</li></ol>

Beyond Preventive Healthcare Screening

An integrated method for targeted Oxford Nanopore sequencing and automated bioinformatics

We all aspire to optimal health, which is the most critical aspect not just to achieve our aspirations and goals but also to have a quality of life. That simply mandates that we shift from the reactive to a proactive framework. Preventive healthcare screening is the first step in that direction. The preventive screening tests often bring to fore an underlying issue whose symptoms have not yet manifested, so it results in early detection, reduced healthcare costs, and treatment outcomes, and longevity. Almost 70% of the healthcare decisions are based on preventive screening, followed by diagnostic testing.Most preventive tests include evaluating the blood parameters which can identify organ specific aberrations (e.g cholesterol levels, hemoglobin levels, liver enzymes, thyroid hormone levels, etc), additionally can also involve other screening methods like X-rays, mammograms, vision and hearing tests. The tests are also tailored to address the needs of various age-groups and demographics. The results of the preventive screening tests help clinicians/health-care providers to suggest specific recommendations for lifestyle modification and/or therapeutic interventions. Though preventive screening healthcare tests help to a large extent, we need to go one level higher if we are to be truly proactive, we need to evaluate our inherent risks based on genetic screening. Since we not only inherit the genetic material from our parents but along with it the risks of our family and previous generations. We know for a fact that the risks of several diseases such as heart ailments, hypertension, diabetes runs in families. Genetic healthcare screening provides valuable insights into an individual's genetic risk profile, enabling personalized healthcare interventions. By integrating preventive healthcare screening with genetic screening, individuals can benefit from a comprehensive approach to health management. This synergy allows for the development of personalized healthcare plans tailored to an individual's genetic predispositions and lifestyle factors. The genetic screening tests are truly predictive and personalized in nature. They can help identify predictive and predispositional testing (e.g risk of diabetes, risk of cancers), intolerances (gluten, lactose etc), sensitivities (caffeine), genetic risk for deficiencies (Vitamins, minerals), response to macronutrients. Genetic screening tests can also help to know if you are a carrier for certain disorders which has implications for reproductive choices and decisions, identify about the variations in an individual’s DNA which influence how you will metabolize the specific drugs. In conclusion the future of healthcare lies in and precision health initiatives. Preventive healthcare screening and genetic screening are powerful tools for promoting long-term health and well-being. By integrating genetic insights into healthcare decisions, individuals can take proactive steps to prevent disease and optimize their quality of life. Remember, your health is your most valuable asset—invest in it wisely.

Precision Medicine: As genomic sequencing becomes more affordable and accessible, the focus is shifting towards personalized medicine. This involves tailoring medical treatment and prevention strategies to individual genetic profiles. Integrating genomic data with clinical information leads to more targeted therapies, improved diagnosis, and better patient outcomes. Advancements in single-cell sequencing, spatial transcriptomics, and long-read sequencing enhance our understanding of genetic variation and its implications for health and disease.Multi-Omics Integration: Integrating data from multiple 'omics' disciplines such as genomics, transcriptomics, proteomics, metabolomics, and epigenomics provides a more comprehensive view of biological systems. This holistic approach allows researchers to unravel complex interactions between genes, proteins, and metabolites, leading to insights into disease mechanisms and identifying new therapeutic targets. Advanced computational methods and machine learning algorithms are being developed to analyze and interpret multi-omics data, driving forward the field of systems biology.Single-Cell Genomics: Traditional genomic techniques often provide average measurements across populations of cells, masking cellular heterogeneity. Single-cell genomics technologies enable the analysis of individual cells, uncovering rare cell types, cellular states, and dynamic processes such as development, differentiation, and disease progression. Emerging methods like single-cell RNA sequencing (scRNA-seq), single-cell ATAC-seq, and spatial transcriptomics are revolutionizing our understanding of cellular diversity and function in various biological contexts.Long-Read Sequencing and Structural Variation: While short-read sequencing technologies have dominated the genomics landscape, long-read sequencing platforms like Pacific Biosciences (PacBio) and Oxford Nanopore are gaining traction due to their ability to generate longer sequencing reads. Long-read sequencing is valuable for resolving complex genomic regions, detecting structural variations, and improving genome assembly quality. As these technologies become more cost-effective and scalable, they are expected to play a crucial role in deciphering the 'dark matter' of the genome, including repetitive regions and structural variations.Gene Editing Therapies: Advances in gene editing technologies like CRISPR/Cas9 can correct disease-causing genetic mutations directly. These therapies hold promise for treating a wide range of genetic disorders, including rare diseases that have been historically challenging to treat.Overall, the next significant advancements in genomic technologies will likely arise from the convergence of these trends, leading to more personalized, comprehensive, and precise approaches to understanding the complexities of the genome and its impact on human health and biology.

The next big thing in genomic technologies 

In a large country like India with a population of more than 1.4 billion, access to quality healthcare continues to pose an enormous challenge for the masses.&nbsp;The paucity of &nbsp;hospital beds, lack of paramedical staff, &nbsp;broken supply chains, dearth of trained epidemiologists, and shortage of medical supplies during the Covid-19 crisis exposed our grossly inadequate public healthcare systems like never before. On the flip side, however, it also resulted in some breakthrough medical interventions. Under normal circumstances, vaccine development would usually take anywhere between 10-15 years, but India was swift to respond to the situation and scaled up its efforts by developing covid vaccines in less than a year, highlighting the ability of our healthcare community to achieve impossible targets through integration and collaboration.While we found ourselves in dire straits due to the rampant spread of the novel coronavirus, the pandemic has derailed us from the path to fight many other diseases that could impair our healthcare systems, if not addressed in due time.Tuberculosis, a bacterial infectious disease that directly affects the lungs, has been a subject of concern worldwide and affects millions of people across the country. India reports an alarming chunk of about 3 million TB cases every year, with a mortality number of almost half a million in 2019. In 2020, India alone accounted for 26% of the TB cases across the globe. In addition to this, there are more than a million &lsquo;missing&rsquo; cases every year that go unreported because they largely remain undiagnosed or inadequately diagnosed. The Global Tuberculosis Report published by the World Health Organization (WHO) &nbsp;in October 2021, revealed that the Covid-19 pandemic and its impact on our healthcare systems had &lsquo;reversed years of progress in providing essential TB services and reducing TB disease burden.&rsquo;&nbsp;The National Strategic Plan (NSP) 2017-2025 formulated by the Govt. of India, is a flagship public health initiative built on the success and learnings of the last NSP and serves as a comprehensive framework for all the stakeholders who are working towards eliminating TB from India. The plan proposes the adoption of bold strategies coupled with the availability of resources to realize its vision of a &lsquo;TB free India&rsquo; by 2030, in line with the UN Sustainable Development Goals (SDGs).While India has been making consistent efforts to ramp up its services by treating more than 10 million TB patients under RNTCP, we are far from meeting the 2030 SDGs and 2035 End TB targets. Inadequate diagnosis and treatment, outdated and ineffective diagnostic tools like sputum-smear microscopy, and multidrug resistant tuberculosis (MDR TB) which account for about 1% of new TB cases; continue to be some of the major challenges in controlling the spread of TB. &nbsp;In addition to this, regulating and monitoring the spread of TB is a Herculean task in a densely populated country such as ours, and the ongoing pandemic has further aggravated the situation by diverting the majority of our healthcare staff and diagnostic services towards Covid management.&nbsp;Recent research indicates that 75% of MDR TB cases globally were not treated accurately and contributed to active transmission of MDR TB in the community (Farhat et al 2019). In some sense, as the world is effectively treating drug susceptible forms of TB, the community transmission of drug resistant forms can snowball into a difficult to treat epidemic.&nbsp;This poses a crucial question before us: How can we effectively combat the spread of tuberculosis and mitigate the damage caused to our healthcare systems? Although prior techniques have had limited impact in controlling the spread, comprehensively-deployed interventions based on the four strategic pillars of the &lsquo;Detect-Treat-Prevent-Build&rsquo; (DTPB) model can allow us to design and to bring about a rapid transformation in the prevention and treatment of tuberculosis.<h3>A case for Whole Genome Sequencing&nbsp;</h3>For several years, clinical microbiology heavily relied on the use of conventional phenotypic and genotypic methods for the identification and treatment of infections. Not only are these methods extremely time consuming and expensive, the lack of technological infrastructure has also prevented us from distinguishing highly genetically related strains. The need of the hour is to build solutions that are affordable and accessible and can revolutionize TB diagnostics while facilitating personalized drug therapy by employing a decentralized approach to diagnosis and treatment.&nbsp;Whole genome sequencing (WGS) is a technique that allows us to determine the entirety of an organism&rsquo;s DNA in a cost-effective way through a single comprehensive test. The use of WGS technology for the diagnosis of tuberculosis has been gaining traction across the world as it enables an &lsquo;all-in-one&rsquo; approach by providing epidemiological data and comprehensive information on genetic background and expected sensitivity of the strains.While the use of Whole Genome Sequencing (WGS) for the treatment of tuberculosis has been deployed in countries such as the UK, Spain, &amp; Italy, there have been little efforts to expand the scope for this approach in India where we have merely laid the groundwork for its use.Given the tremendous success of WGS based prediction of drug resistance in a single step, national programs must consider a change in strategy, moving away from sequential in vitro diagnostics, to large scale comprehensive screening using WGS.&nbsp;These platforms are &nbsp;cost effective and can provide unprecedented asset utilization, wherein each installation can perform &gt;600,000 tests per year. Tentatively, 10 such installations can effectively cover the entire estimated TB caseload of India, at a cost comparable to current CBNAAT tests. Consequently, only a very small select number of cases will need culture DST to resolve outcome based discrepancies.&nbsp;The expansion of genome sequencing based tests in India, which are extremely cost effective and provide comprehensive information in a quick turnaround time of less than 12 days, will enable us to combat diagnostic inaccuracies and delays that impede appropriate drug administration. By investing in sequencing technologies and ramping up the infrastructure and expertise for &nbsp;genome based tests, we can create a unique opportunity by establishing it as the gold standard in TB diagnostics and provide accessible healthcare to the masses; an attempt to truly revolutionize and re-envision the future of healthcare in our country as we strive to achieve our 2025 end TB targets. &nbsp;

Data Scientist working on Genomics and Metagenomics, with special focus on Infectious diseases, Virology and disease epidemiology

Can India eliminate Tuberculosis by 2025: A roadmap for inclusive diagnostics

&lsquo;So much done; yet, so much to achieve!,&rsquo; Tuberculosis (TB) is still looming as a major global concern, being the 2nd leading infectious killer after COVID-19 (above HIV/AIDS). Almost 1.5 million people lost their lives to TB in 2020 and an estimated 10 million fell ill worldwide; including 5.6 million men, 3.3 million women and 1.1 million children. Individuals infected with TB bacteria, have a 5-10% lifetime risk of falling ill with TB, and immunocompromised individuals (people with HIV, malnutrition, diabetes, or using tobacco) have greater risk of contracting the disease [1].More than 95% of the cases and higher mortality are witnessed in developing countries, affecting people of all age, reducing the productivity of the nation. India leads amongst the countries accounting for 2/3rd of the global TB cases, followed by China, Indonesia, the Philippines, Pakistan, Nigeria, Bangladesh and South Africa. Growing Multidrug-resistant TB (MDR-TB) cases have only increased the disease burden worldwide [1]. These patients do not respond to isoniazid and rifampicin (the 2 most-effective first-line anti-TB drugs). Treating MDR-TB is complex and quite expensive, primarily due to its association with important adverse events in patients. The success rate of treatment lowers from 83% in patients having newly diagnosed or relapse non-DR-TB receiving first-line therapy to 54% in MDR-TB patients, to 30% in extensively DR-TB (XDR-TB- showing resistance to at least one fluoroquinolone and one second-line injectable agent) patients [2].&nbsp;Early and comprehensive detection of TB and MDR-TB is the call of the hour and over the recent decades, many techniques have been devised to tackle this issue. The gold standard for TB diagnosis is culture &ndash;drug sensitivity test (CDST); however, due to its time-consuming nature, several major alternatives were devised like line-probe assays, real-time PCRs, GenXpert (Xpert) [2]. The line-probe assays, although simple, quick and easy-to-perform, requires expensive equipment, whereas real-time PCRs despite of lesser contamination chances and rapid results, require specific equipment and skilled technicians [2]. Xpert MTB/RIF, which is a rapid molecular-based test endorsed by WHO, based on cartridge-based assay has revolutionized the TB diagnosis and can detect MTB complex and RIF resistance, using semi-quantitative nested real-time PCR for amplifying hotspot region of the rpoB gene. It has remarkable sensitivity (100%) and specificity (99%) for smear-positive samples and 67% sensitivity and 99% specificity for smear-negative samples, as compared to CDST. Xpert has been very valuable in tackling TB burden in low- and middle-income countries, with turnaround time (TAT) being &lt;2hours [2] , with Xpert Ultra, adding more value, thereby increasing the sensitivity, especially in paucibacillary TB including tuberculous pleurisy, tuberculous meningitis and paediatric TB [3] as well as in pulmonary and extra-pulmonary TB [4]. XpertUltra is a 10-channel upgradation of classic Xpert MTB/RIF, with information regarding second-line injectable drugs (SLIDs) and 98-100% specificity[5] however most of the SLIDs are not recommended in the current treatment protocols longer. Despite these advantages, mutations linked with isoniazid resistance could be missed by Xpert and thus warrants other tests CDST for confirming. The higher cost also makes it unaffordable in many high-burden countries, along with need for constant electric supply and sensitivity to heat and dust adding to the challenges [2].Some of these issues can be effectively addressed by using sequencing technology, especially next generation sequencing (NGS), which can rapidly analyze the entire genome, allow species identification, screen all mutations, detecting drug resistance and predicting evolution of organism. Whole Genome Sequencing (WGS) screens the entire genome using high-end sequencing technique, spanning entire genome, to trace the culprit gene/mutation causing TB or drug resistance. It is more advantageous, as it gives greater genome coverage (not only the target region), &nbsp;increased epidemiological information and understanding of new resistance mechanism for both existing and novel drugs [2]. WGS can assist in efficient prediction of treatment outcomes and resistance profiles of all anti-TB drugs in a single run with rapid results [6]. Poly-microbial infections (mixed and co-infections) can cause misidentification of strains and underestimation of disease transmission, resulting in poor treatment outcomes [7]. Susceptibility profile of drugs like pyrazinamide can be achieved with WGS, which can help in shortening the treatment regimen. WGS can provide high resolution in investigating disease transmission outbreaks, influence early decision of optimum drug therapy and improve patient outcomes [8]. WGS was found to be 7% more cost-effective and consistent in performance as compared to DST [9, 10]. &nbsp;With the advent of WGS, there has been a revolutionary change in TB diagnosis, which can facilitate rapid and timely diagnosis and thereby appropriate treatment for the patients. Disruption in the transmission outbreaks can speed-up the path towards TB elimination, a goal definitely nurtured by all!Reference:<ol><li>W.H.O. Tuberculosis. 2021.[Internet]. https://www.who.int/news-room/fact-sheets/detail/tuberculosis.&nbsp;</li><li>Nyugen TNA, Anton-Le Berre V, Ba&ntilde;uls AL, Nguyen TVA. Molecular Diagnosis of Drug-Resistant Tuberculosis; A Literature Review. Front Microbiol. 2019 Apr; 10: 794. &nbsp;201. doi: 10.3389/fmicb.2019.00794.</li><li>Jiang J, Yang J, Shi Y, Jin Y, Tang S, Zhang N, et al. Head-to-head comparison of the diagnostic accuracy of Xpert MTB/RIF and Xpert MTB/RIF Ultra for tuberculosis: a meta-analysis. Infectious Diseases. 2020;52(11): 763-775. http://doi.org/10.1080/23744235.2020.1788222.&nbsp;</li><li>Zhang M, Xue M, He J. Diagnostic accuracy of the new Xpert MTB/RIF Ultra for tuberculosis disease: A preliminary systematic review and meta-analysis. Int J Infect Dis. 2020; 90:35-45. doi: 10.1016/j.ijid.2019.09.016.</li><li>Penn-Nicholson A, Georghiou SB, Ciobanu N, Kazi M, Bhalla M, David A, et al. Detection of isoniazid, fluoroquinolone, ethionamide, amikacin, kanamycin, and capreomycin resistance by the Xpert MTB/XDR assay: a cross-sectional multicentre diagnostic accuracy study. Lancet Infect Dis. 2021; S1473-3099(21)00452-7. doi: 10.1016/S1473-3099(21)00452-7.</li><li>He G, Chen X, Chen J, Zhang W. Prediction of treatment outcomes for multidrug-resistant tuberculosis by whole-genome sequencing. Int J Infect Dis. 2020 Jul; 96:68-72. doi: 10.1016/j.ijid.2020.04.043.</li><li>Advani J, Verma R, Chatterjee O, et al. Whole Genome Sequencing of Mycobacterium tuberculosis Clinical Isolates From India Reveals Genetic Heterogeneity and Region-Specific Variations That Might Affect Drug Susceptibility. Front Microbiol. 2019 Feb 26; 10:309. doi: 10.3389/fmicb.2019.00309. &nbsp;</li><li>Katale BZ, Mbelele PM, Lema NA, Campino S, Mshana SE, Rweyemamu MM, et al. Whole genome sequencing of Mycobacterium tuberculosis isolates and clinical outcomes of patients treated for multidrug-resistant tuberculosis in Tanzania. BMC Genomics (2020) 21:174 https://doi.org/10.1186/s12864-020-6577-1.&nbsp;</li><li>World Health Organization. Meeting report of the WHO expert consultation on the definition of extensively drug-resistant tuberculosis. 2020.</li><li>CRyPTIC Consortium and the 100,000 Genomes Project, et al. Prediction of susceptibility to first-line tuberculosis drugs by DNA sequencing. N Engl J Med. 2018 Oct 11;379(15):1403-1415. doi: 10.1056/NEJMoa1800474.</li></ol>

Tuberculosis (TB) is still looming as a major global concern, being the 2nd leading infectious killer after COVID-19 (above HIV/AIDS).

Current scenario of TB diagnosis and what the future holds?

Tuberculosis (TB) is a preventable, treatable, and curable disease. However, in 2020, 9.9 million people were estimated to have developed tuberculosis, and 1.5 million people were estimated to have died from it. Whereas in India, 2.6 million were diagnosed with TB and 436,000 succumbed to TB in 2019. The COVID-19 pandemic has substantially reduced access to services for the diagnosis and treatment of TB, resulting in an increase in deaths and a reversal in global progress. [1] Presently, TB incidence is falling at a rate of 2% per year, hampered mainly by the rearing pandemic of drug-resistant tuberculosis (DR-TB). Particularly concerning is multi-drug resistant TB (MDR-TB), defined as resistance towards isoniazid (INH) and rifampicin (RIF). [2] The World Health Organization (WHO) targeted to reduce worldwide TB incidence by 90% by 2035. [1] Early initiation of effective treatment,based on susceptibility patterns of the Mycobacterium tuberculosis complex (MTBC), is considered key to successful TB control in countries with a high incidence of DR-TB. Worldwide, MDR-TB treatment outcomes are poor, with cure rates of less than 60% (2), due to the lack of comprehensive drug susceptibility Testing (DST). This leads to the inadequate anti-TB activity of the provided regimens (3–5), unlike regimens advised based on DST results, to ensure optimal results. (6) In addition to resistance to the established regimens, resistance to the new DR-TB drugs is increasing. On World TB Day, the Academy of Advanced Medical Education, Thyrocare Technologies Limited and HaystackAnalytics - IITB, along with expert pulmonologists and renowned physicians from India, convened for an advisory board meeting in Delhi to discuss the role of Whole Genome Sequencing (WGS) in the diagnosis and management of TB. Objectives and specific topics relating to WGS in MDR-TB were discussed. Each expert shared their views, which led to a group discussion committed to putting the patient first and increasing their collective efforts, recognizing the possibility of accomplishing the goal. The organizations involved in the discussion declared their commitment to collaborate to tackle DR-TB detection efficiently. They advocate strengthening access to WGS TB services, controlling and preventing TB, improving surveillance and drug resistance management, and investing in research and development. This round table serves as a framework to build on and ensure that the goal of ending TB is achievable with WGS services wherever needed. Post-discussion, a uniform consensus was said to have been established if more than 80% of the board members agreed to the statement. Consensus statements : <ol> <li>Patients with MDR-TB and XDR-TB will benefit from WGS.</li> <li>Patients with relapse or reinfection will benefit from WGS.</li> <li> WGS of MTBC may be beneficial when results of other molecular tests are inconclusive or indeterminate.</li> <li> WGS may be beneficial when phenotypic DST is not available for newer drugs like bedaquiline and delamanid).</li> <li>WGS is beneficial to detect disease transmission (identify cluster and strain).</li> <li>WGS in TB is an important part of epidemiological surveys.</li> <li>WGS provides additional information such as, drug susceptibility profile and MTBC mutation information.</li> <li>Research needs to be encouraged to definitely observe the benefits of WGS in TB patients.</li> <li>Due to lack of data from the Indian population, WGS is not recommended for routine use. Further, research and clinical outcome data in the Indian setting should be promoted.</li> <li>WGS is useful in certain patient profiles: <ol> <li>Failure of MDR-TB or XDR-TB treatment</li> <li>Treatment of DR-TB is not effective</li> <li>Past history of DR-TB</li> <li>Inconclusive results with other molecular tests</li> </ol> </li> </ol> References:<ol><li>WHO Global Tuberculosis Report 2021 India Tb Report 2020</li><li>Uplekar M, Weil D, Lonnroth K, Jaramillo E, Lienhardt C, Dias HM, et al. WHO’s new end TB strategy. Lancet Lond Engl. 2015 May 2;385(9979):1799–801.</li><li>Nahid P, Mase SR, Migliori GB, Sotgiu G, Bothamley GH, Brozek JL, et al. Treatment of Drug-Resistant Tuberculosis. An Official ATS/CDC/ERS/IDSA Clinical Practice Guideline. Am J Respir Crit Care Med. 2019 Nov 15;200(10):e93–142.</li><li>Impact of rapid drug susceptibility testing for tuberculosis: program experience in Lima, Peru - PubMed [Internet]. [cited 2022 Mar 25]. Available from: https://pubmed.ncbi.nlm.nih.gov/22990138/</li><li>Seung KJ, Keshavjee S, Rich ML. Multidrug-Resistant Tuberculosis and Extensively Drug-Resistant Tuberculosis. Cold Spring Harb Perspect Med. 2015 Sep;5(9):a017863.</li><li>Rodrigues C, Jani J, Shenai S, Thakkar P, Siddiqi S, Mehta A. Drug susceptibility testing of Mycobacterium tuberculosis against second-line drugs using the Bactec MGIT 960 System. Int J Tuberc Lung Dis Off J Int Union Tuberc Lung Dis. 2008 Dec;12(12):1449–55.</li></ol>

Genomic revolution: Transforming tuberculosis diagnosis and treatment with the use of Whole Genome Sequencing

Genomic revolution: Transforming tuberculosis diagnosis and treatment with the use of Whole Genome Sequencing - A consensus statement

Medical Tourism or Health Tourism has been gaining precedence across the world, and an increasing &nbsp;emphasis on individual wellbeing, high-quality affordable medical services, and &nbsp;easier cross-border travel, has led India to become a popular hub for overseas patients.&nbsp;It is no revelation that over the past few decades, India&rsquo;s medical sector has grown by leaps and bounds due to its highly skilled medical specialists and staff, advanced medical infrastructure and facilities powered by emerging technology, and low-cost treatment options.&nbsp;Both public and private healthcare players, supported by the government and favourable National Health policies, have led the country to realize its potential to grow as the hub of medical tourism. India&#39;s efforts to &nbsp;supply vaccines to many countries across the world in the aftermath of the Covid-19 pandemic, has &nbsp;also furthered our vision of &nbsp;becoming &lsquo;One Nation, One Health System&rsquo;.While global medical tourism, valued at USD 4.0 billion in 2021, &nbsp;is expected to grow at a compound annual growth rate (CAGR) of 32.51% by 2030, India is making relentless efforts to ramp up its healthcare infrastructure and medical &nbsp;services to meet the growing demands of the changing world.<h3>What makes India a preferred destination for Medical Tourists&nbsp;</h3>With relaxed visa norms and a huge influx of patients from countries like Bangladesh, Kenya, Iraq, Oman, Tanzania, Nigeria, Afghanistan, Maldives etc, &nbsp;the country has witnessed &nbsp;over 40% growth in Foreign Tourist Arrivals (FTAs). The number of tourists visiting on medical visas has increased from 495,056 in 2017 to almost 697,000 in 2019. The reasons for the growing popularity of India as a medical hub are manifold. Unlike most developed countries like the U.S which have extremely high treatment costs, quality healthcare services in India can be obtained at a cheaper rate. Since many patients cannot afford to pay for medical insurance in their own countries, &nbsp;travelling to India to obtain medical services allows them to avail world class medical services at almost 50% less rates. This helps patients with chronic medical conditions who have to wait for longer durations to avail medical services in their native countries, gain access to prompt treatment at reasonable costs.&nbsp;The availability of world-class amenities and renowned doctors who are pioneers in their fields has also been a major enabling factor in India&rsquo;s growing popularity as a preferred destination for overseas patients. Lakhs of patients visit India every year to avail medical treatment, especially in the departments of cardiology, neurology, and orthopaedics. Treatments for cardiovascular diseases, eye surgery, hip replacement, kidney ailments etc are among the &nbsp;most commonly sought after treatments by foreign patients.&nbsp;Given that almost 70% of today&#39;s medical decisions are based on laboratory results, the availability of CAP and NABL accredited diagnostic labs equipped with state-of-the-art technology &nbsp;to generate rapid results, has also enabled these patients to opt for specialised surgical treatments &nbsp;and other forms of medical care in the country. These factors also play a crucial role in minimising unnecessary expenditures for medical tourists, by reducing the overall duration of their stay.<h3>How Genomics can be the game changer in India&rsquo;s Medical Tourism expansion strategy</h3>As India strives &nbsp;to become the leader in global healthcare delivery, it can only be made possible when we expand the use of technological advancements and next generation sequencing techniques like Whole Genome Sequencing (WGS) to provide correct treatment options for patients. &nbsp;The success of the Human Genome Project, a 13 year long project aimed at determining the DNA sequence of the entire human genome, has enabled scientists and researchers to identify advanced techniques and tools to deliver new age precision diagnostics.&nbsp;It is integral to remember that while the country gains popularity as the preferred destination for medical tourists, our priorities should first be targeted at meeting the &nbsp;clinical needs and demands of our &nbsp;own population of &nbsp;over 1.38 billion. The use of &nbsp;Whole Genome Sequencing &nbsp;(WGS) &nbsp;can help improve not just curative healthcare delivery for overseas patients, but also help adopt &nbsp;preventive health care practices &nbsp;for the masses. &nbsp;The use of Whole Genome Sequencing can be especially effective for treatments in oncology, pharmacology, &nbsp;genetic disorders, infectious and rare diseases. Given that the multiple test paradigm is experimental and ineffective with almost 68% patients going undiagnosed, patients no longer need to depend on &nbsp;time consuming tests to access accurate results for a range of medical conditions. &nbsp;With the availability of the Universal Infectious Diseases test in India that deploys Next Generation Sequencing technology, anyone suffering from a fever or infection of unknown origin can gain access to a single test that gathers genomic data to identify existing and emerging infections. &nbsp;By estimating health parameters such as lifetime risk of a disease, responses to drug and adverse reactions, genomic tests can prove to be effective screening tools for such patients and help the medical staff identify and manage existing conditions and symptoms by &nbsp;designing strategies that can reduce disease risk and delays.&nbsp;With costs that are almost 60-80% lower than their own countries, both medical tourists and native patients can benefit immensely from such cost effective and prompt medical treatment options, which were hitherto unavailable. As we expand the use of Whole Genome Sequencing in India, we can not only allow for greater detection of health problems for the masses, but also generate genomic information that can aid the translation of applications to design targeted therapeutic strategies for medical tourists.Genomics can pave the way for new age precision medicine, as India caters to the needs of overseas patients, by providing them high quality, timely and accurate treatment, while reducing their exposure to avoidable drug toxicity. &nbsp;

How India can emerge as a hub for Medical Tourism with Next Generation Sequencing

How is NGS changing the paradigm of infectious disease diagnosis?Decisions on the choice of the right empiric therapy have been challenging owing to the growing burden of antibiotic resistance and the inherent limitations of current-day diagnostics. Blanket empiric therapy with broad-spectrum antimicrobials adds to the cost, toxic side-effects and selection pressure of de novo resistant microbes. Early detection and identification of bloodstream pathogens along with resistance patterns will improve population-level outcomes. The existing gold standard of microbiological diagnosis is hindered by inefficient turnaround time, poor sensitivity (10-30%) and failure to detect fastidious and non-culturable organisms (1). Molecular diagnostic tests based on PCR technologies are hypothesis-driven, with a high dependence on the primer selection. This allows identification of only a limited range of pathogens; thus failing to detect unknown pathogens which can lead to ambiguous results and confound clinical decision making (2). Through NGS technologies currently may not replace traditional microbiological diagnostics, it definitely is changing the face of it (3), with the scope of unbiased and data-driven detection of multiple classes of organisms on a single platform. The development of real-time sequencing platforms such as Oxford Nanopore sequencing has scaled resolution, accuracy, and shorter turnaround time compared to currently available commercial techniques. Routine implementation of NGS in the clinical setting will markedly improve point-of-care pathogen detection (4).Accurate, comprehensive and rapid diagnostics (5,6) can thus aid initiation of optimal therapy, better patient outcomes and shorter hospital stay. A comprehensive and scalable diagnostic technology with improved sensitivity and specificity for pathogen detection is essential to monitor the extent of the problem, develop feasible local diagnostic algorithms and databases that might aid in clinical decision making, limit the emergence of resistant pathogens and alleviate the fatality rate.REFERENCES : <ol><li>Cao XG, Zhou SS, Wang CY, Jin K, Meng HD. The diagnostic value of next-generation sequencing technology in sepsis. Front Cell Infect Microbiol. 2022 Sep 14;12:899508.</li> <li>Grumaz C, Hoffmann A, Vainshtein Y, Kopp M, Grumaz S, Stevens P, et al. Rapid Next-Generation Sequencing-Based Diagnostics of Bacteremia in Septic Patients. J Mol Diagn [Internet]. 2020 Mar 1 [cited 2023 Dec 4];22(3):405–18.Available from: https://pubmed.ncbi.nlm.nih.gov/32146977/</li> <li> Betz BL, Mroz P, Billington C, Hilt EE, Ferrieri P. Next Generation and Other Sequencing Technologies in Diagnostic Microbiology and Infectious Diseases. Genes 2022, Vol 13, Page 1566 [Internet]. 2022 Aug 31 [cited 2023 Dec 12];13(9):1566. Available from: https://www.mdpi.com/2073-4425/13/9/1566/htm</li> <li> Li JY, Shen GG, Liu TG, Tang L V, Xia LH, Hu Y. Nanopore-targeted sequencing for simultaneous diagnosis of suspected sepsis and early targeted therapy. Ann Transl Med [Internet]. 2021 [cited 2023 Dec 12];9(23):1749. Available from: https://dx.doi.org/10.21037/atm-21-2923</li> <li>Liu J, Zhang Q, Dong YQ, Yin J, Qiu YQ. Diagnostic accuracy of metagenomic next-generation sequencing in diagnosing infectious diseases: a meta-analysis. Scientific Reports 2022 12:1 [Internet]. 2022 Dec 5 [cited 2023 Dec 12];12(1):1–10.Available from: https://www.nature.com/articles/s41598-022-25314-y</li><li>Hong M, Peng D, Fu A, Wang X, Zheng Y, Xia L, et al. The application of nanopore targeted sequencing in the diagnosis and antimicrobial treatment guidance of bloodstream infection of febrile neutropenia patients with</li> <li>hematologic disease. J Cell Mol Med [Internet]. 2023 Feb 1 [cited 2023 Dec 12];27(4):506–14. Available from:https://onlinelibrary.wiley.com/doi/full/10.1111/jcmm.17651</;i></ol>

How is NGS changing the paradigm of infectious disease diagnosis?

Clinicians worldwide rely heavily (almost 70%) on the laboratory results, for diagnosing the illness, recommending the appropriate treatment and follow-ups. Hence it becomes an onus on laboratorians like us, to give an early and precise diagnosis. With highly infectious diseases like Tuberculosis (TB), it is all the more important to diagnose the patient at the earliest, for initiating appropriate treatment, thereby breaking the chain of spread.&nbsp;Despite remarkable advances made over the recent decades, to control this debilitating and life-threatening disease, gaps still exist in its complete eradication. Approximately 3 million people with TB are deemed &lsquo;missing&rsquo; due to under-diagnosis and under-reporting to national TB surveillance programs. In order to achieve the goal of TB eradication by 2030, the World Health Organization (WHO), appeals to trace these missing millions. The picture is more complex, due to the emergence and rapid spread of multi-drug-resistant TB (MDR-TB) strains. &nbsp; Being a laboratorian, we always strive to give the required diagnostic information by developing and utilizing tests with high sensitivity and specificity. Sensitivity of a test is the probability (in percentage) that the sample tests positive given the patient has the disease, whereas specificity is the ability of the test to detect a true negative case. Initially the diagnosis of TB was mostly carried out by smear microscopy and imaging techniques which were not as effective as the gold-standard culture drug- susceptibility test (DST). However, DST being time-consuming, molecular assays which are based on the DNA (genetic material) isolated from the TB bacilli like line probe assay (LPA) and GenXpert, TruNAATs and sequencing were introduced in the recent decades. GenXpert, a revolutionary method, facilitated in rapid detection of presence of TB causing pathogen Mycobacterium tuberculosis (MTB) and also identified the presence of rifampicin drug resistance. However, since MTB managed to acquire resistance towards other drugs other assays such as LPA were utilized as it could detect the resistance towards first-line and second &ndash;line anti-TB drugs. The LPA&rsquo;s are based on binding of the mycobacterial DNA to probes which were specific to certain regions only and so there was always the possibility of missing some resistant strains which were harbored in the patient. Taken as a whole, these diagnostic tests though very sensitive and specific, lacked completeness and thus have limited use. Over the period, the TB pathogen still manages to evolve resistance towards most of the drugs being used against it and this poses a great dilemma to the laboratorians as well as the clinicians, not to forget the patients dealing with it first-hand! These issues led to the introduction of sequencing method into TB diagnosis.&nbsp;What do we know about sequencing?Since the discovery of DNA, way back in 1953, research and diagnostic fields have progressed immensely. The nucleotide sequence of DNA often called the alphabet of life (adenine, guanine, cytosine, thymine) carries the information regarding the gene, that can give better understanding of its functions. Any change from the normal sequence, gives rise to mutation and these occurring in the MTB genome, causing emergence of various drug-resistance. Identifying these mutations is vital for understanding how the disease progresses. &nbsp; &nbsp;The most recent development in sequencing method called as next generation sequencing (NGS), which is based on the same premise of DNA sequencing albeit in a high through-put manner (that means it can sequence several genes as well as several samples at the same time) has rapidly moved into our laboratories.&nbsp;NGS can facilitate rapid diagnosis compared to the traditional practices and provide detailed and accurate sequence information. Using this technology, the entire genome of the Mycobacterium or the whole genome can be scanned to look for mutations in all the genes through which the TB drugs act (WGS is used as an acronym for whole genome sequencing). &nbsp;WGS can rapidly and comprehensively identify the culprit and the underlying drug-resistance mechanism for both existing and novel drugs and that too in a single run! Since the entire DNA is being sequenced, the WGS also gives me really important information if there are more than one pathogen known as co-infection or if there are two different types of the Mycobacteriun strain itself (mixed infections) and it could be that one of them is sensitive and one is resistant in the same individual. Some of the previously mentioned tests are unable to pick this and often can result in poor treatment outcomes as optimally effective drug will not be given to this individual. &nbsp;As a laboratorian I now rely on TB-whole genome sequencing to give rapid, comprehensive, detailed diagnosis and drug resistance profile. &nbsp;This way I am contributing to achieving our goal to ENDTB soon and I foresee this reality as more and more laboratories are adopting this technology to give crucial information about drug resistance in TB patients.&nbsp; &nbsp;

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