Diagnosing and Tracking COVID-19 Infections Leveraging Next-Gen Sequencing
by Dr. Christiane Scherer and Dr. Andreas Scherer
The COVID-19 pandemic is reaching historic proportions. We are dealing with an infectious disease that is caused by a novel coronavirus we discovered just a few months ago. Since then, it has brought healthcare systems to the brink, it altered how we work, it changed how we socialize, and it impacted the world economy in a major way.
While a global response has been mobilized to defeat the virus, there are currently no good solutions available. The current goal is to reach a sufficiently high level of immunization in the global population and to develop treatment options. In the meantime, we have to be efficient in diagnosing infections, isolating COVID-19 cases, and studying this virus by understanding its subtypes, epidemiology, routes of transmission, and clinical manifestation. In this article, we summarize our current understanding of COVID-19 and the virus SARS-Cov-2 causing it. Most of the papers we reference were published since the beginning of this year. The body of knowledge is quickly expanding and evolving. Next-Generation Sequencing (NGS) can deliver significant insights into this process. This article outlines how NGS can be used for diagnosing and tracking COVID-19 infections in the clinic.
Chapter 1: Introduction
At the end of 2019, a virus appeared somewhere in the Chinese city of Wuhan. For most individuals infected with the virus, symptoms were limited to mild cold and flu-like symptoms, but in a minority of cases, the virus resulted in severe pneumonia and death. It proved to be highly contagious. The disease it caused would soon be coined COVID-19, standing for coronavirus disease of 2019. It quickly emerged as a global phenomenon mobilizing resources in every country to defeat it.
At the time of this writing, we are in the midst of a global pandemic. COVID-19 has arrived in many countries: Asia, Europe, and Latin America. There are cases reported in Australia, the Middle East, Africa, and Canada. The United States was hit especially hard, trying to contain the exponential spread in a country that is based on individual freedom and liberty. As the country adopts social distancing measures, based on federal and state guidance, we are facing the reality that the reported case numbers are climbing undeterred (see Figure 1).
As we are writing this sentence, there have over 4.3M confirmed cases of the coronavirus globally. The death toll is quickly approaching 300,000. This will obviously be out of date by the time you are reading this paragraph. We are dealing with exponentially growing numbers. There are estimations that put the number of infections in the tens of millions and the number of deaths in the hundreds of thousands. The Johns Hopkins University has developed a website, the Coronavirus Resource Center, that gives up-to-date information on this pandemic with updated information multiple times a day.
There are other informative resources that help to quantify the spread of the virus. Dr. Edward Parker, from the Vaccine Centre at the London School of Hygiene & Tropical Medicine, is maintaining a website that allows viewers to visualize current trends with his COVID-19 tracker (see Figure 2).
In addition, the website gives information about other recent outbreaks, such as the epidemic of Severe Acute Respiratory Syndrome (SARS) in 2003, the 2009 Swine Flu Pandemic, and the 2014 Ebola outbreak. The virus behind COVID-19 is called SARS-CoV-2. It is a pathogen that has unique characteristics turning it into a threat to our lives and the global economy. According to Fang et al. 2020, the preliminary estimate of R0, which estimates the expected number of new cases produced by each infected individual in a population, is 2.2 – 3.7 (see also Qun Li et al., 2020). It could be shown that it is able to persist for days on uncleaned surfaces.
Chapter 2: COVID-19 Key Facts
SARS-CoV-2 has just been recently discovered. The knowledge about this virus is fairly new, and certain aspects of it are still under review or in-flux entirely as we learn more about this virus on a daily basis (see Di Wu et al., 2020 and Scherer, 2020a).
The virus has rapidly spread from Wuhan to China’s other areas and reached global proportions as it is now present on all continents except Antarctica. According to the European Centre for Disease Prevention and Control (ECDC), the latest daily risk assessment is moderate to high level. The case fatality rate of the currently reported cases in China is less than 4%, which implies that so far, this novel coronavirus does not seem to cause the high fatality rates previously observed for SARS-CoV and MERS-CoV. However, it has a higher R0 (2.2 -3.7) value than either of these viruses. SARS-CoV has an R0 of 0.67 – 1.23. MERS-CoV has an R0 of 0.29-0.8 (see Trilla, 2020).
Bats and other species can function as so-called reservoir hosts. They have played a critical role in transmitting various viruses, including Ebola. Cui et al. 2019 describes the origins of SARS-CoV and MERS-CoV likely to be in bats as there is a strong genetic overlap between the viruses extracted from bats and their human-transmissible versions.
In fact, recent research showed that SARS- CoV-2 is 96% identical at the whole-genome level to a bat coronavirus. Understanding the origins of a virus in addition to when and how exactly the jump to humans occurred helps us in understanding and eventually controlling its spread.
Route of Transmission
There are a number of ways in which the virus can transmit from human to human. The main transmission route is through droplet infection. Someone comes into direct contact with a carrier when coughing or sneezing. There are also indications that transmission occurs during general social interaction in limited spatial surroundings (e.g., restaurants, school, sports events, etc.). Asymptomatic infections can lead to a wider spread as the hosts are unaware of their ability to transmit.
It is possible that the virus is able to persist for days on uncleaned surfaces. Recently, the new coronavirus was also found in the feces of confirmed patients in Wuhan, Shenzhen, and even in the U.S. Also, neonatal infections (i.e., mother-to-child transmission) have been observed but need to be confirmed (see Fuk- Woo et al., 2020, Phelan et al., 2020, Jin et al., 2020, Shen et al., 2019, Zhu et al., 2020).
Let’s look in further detail at what we know about the clinical manifestation of this novel pathogen.
Incubation Period and Symptoms
A number of publications based on smaller enrollment numbers suggest an incubation period from 1 to 12 days with a mean of 5-7.5 days. In a larger study with 1,099 patients extracting data from laboratory-confirmed cases from 552 hospitals in 30 Chinese provinces, researchers reported that the estimated mean incubation period of a SARS-CoV-2 infection was 4.0 days (Guan et al. 2020).
The same study (Guan et al., 2020) reported the following data. The median age of the patients was 47 years. 41.9 % of the patients were female. 5% were admitted to the I.C.U. 2.3% underwent invasive mechanical ventilation, and 1.4% died. Only 1.9% of the patients had a history of direct contact with wildlife. The most commons symptoms were the following:
- Fever: 43% on admission, 88.7% during hospitalization
- Cough: 8%
- Diarrhea: 8%
Coincidentally, the SARS-CoV-2 infected cases have symptoms like fever, fatigue, dry cough, dyspnea, etc., with or without nasal congestion, runny nose, or other upper respiratory symptoms. There are also reports of loss of smell and taste in otherwise non-symptomatic cases.
From a diagnostic standpoint, the available options are as follows:
Some patients may not present any noteworthy clinical symptoms despite being infected with the virus except perhaps the loss of smell or taste. Patients in severe condition may have shortness of breath, moist rales in lungs, weakened breath sounds, dullness in percussion, and increased or decreased speech tremor.
C.T. Imaging Examination
In the early stage of pneumonia, chest images show multiple small patchy shadows and interstitial changes, remarkable in the lung periphery. Severe cases can further develop to bilateral multiple ground-glass opacity, infiltrating shadows, and pulmonary consolidation, with infrequent pleural effusion. While chest C.T. Scan pulmonary lesions are shown more clearly by C.T. than x-ray examination, including ground-glass opacity and segmental consolidation in bilateral lungs, especially in the lung periphery.
In a study of 41 patients, 40 (98%) had bilateral involvement. The typical findings of chest C.T. images of I.C.U. patients on admission were bilateral multiple lobular and subsegmental areas of consolidation (Figure 3A). The representative chest C.T. findings of non-ICU patients showed bilateral ground-glass opacity and mental areas of consolidation (Figure 3B). Later, chest C.T. images showed bilateral ground- glass opacity, whereas the consolidation had been resolved (see Figure 3). This data was extracted from Huang et al., 2020.
Current diagnostic strategies involve the exclusions of other known viral causes of pneumonia, such as influenza virus, parainfluenza virus, adenovirus, respiratory syncytial virus, rhinovirus, or SARS-CoV. Also, bacterial infections such as mycoplasma pneumonia, chlamydia pneumonia, and bacterial pneumonia should be tested for prior to conducting a COVID-19 test. A variety of specimens such as nasal swabs, nasopharynx or trachea extracts, sputum or lung tissue, blood, and feces are commonly used for testing.
Should those causes be ruled out, samples can then be collected from the upper respiratory tract (oropharyngeal and nasopharyngeal) or lower respiratory tract (endotracheal aspirate, expectorated sputum, or bronchoalveolar lavage). The standard diagnosis is the C.D.C. 2019-nCov Real-Time RT-PCR Diagnostic Panel, a molecular in vitro diagnostic test, based on the widely used nucleic acid amplification technology.
Next-Gen Sequencing (NGS) is an alternative testing paradigm that has significant advantages over the RT-PCR method. We discuss this in detail in the next chapter.
Treatment and Prevention
At this time, there is no vaccine or antiviral treatment for human and animal coronavirus. The World Health Organization (WHO) has announced that a vaccine for SARS- CoV-2 should be available in 18 months. The currently available clinical treatment options essentially focus on dealing with the symptoms arising from the infection. This ranges from bed rest, antiviral therapy, antibiotics application, immunomodulating therapy, organ function support, respiratory support, bronchoalveolar lavage (B.A.L.), blood purification, and extracorporeal membrane oxygenation (ECMO). Prevention is mostly about self-isolation, social distancing, and minimizing the exposure to a potential infection while living a healthy lifestyle.
Development of Future Treatment Options
Kupferschmidt and Cohen (2020) have reviewed four of the most promising therapies that the WHO has identified: an experimental antiviral compound called remdesivir, the malaria medication chloroquine and hydroxychloroquine, a combination treatment consisting of lopinavir and ritonavir and lastly a combination of lopinavir, ritonavir, and interferon-beta. All drugs are being tested, although the WHO opted not to conduct randomized, double-blind studies in the interest of time.
Remdesivir: It was initially tested to treat Ebola with no confirmed efficacy. There is a reported case in the U.S. with a positive health outcome after treatment with the drug. More data is required. It is a drug that is being administered intravenously.
Chloroquine and Hydroxychloroquine: Studies in cell cultures have suggested that there is some effect on SARS-Cov-2 at very high doses close to toxic dose ranges. Multiple smaller studies in various countries such as
China and France have been conducted that showed some encouraging results. However, there is overall insufficient evidence that warrants a broad usage as of today.
Lopinavir and Ritonavir: This is a drug that was approved in 2000 to treat H.I.V. It has shown efficacy in treating MERS virus infections. Rigorous data collection is required. The drug can cause severe liver damage.
Lopinavir, Ritonavir, and Interferon-Beta: This combination is already in trials to treat MERS. It could be potentially helpful to treat a COVID-19 infection, although experts point out that a late application of interferon-beta could actually lead to worse tissue damage.
In the meantime, there is a substantial global effort underway to develop a vaccine that could provide population-level protection against this novel virus. However, there is a major concern. It is a known risk that coronavirus vaccines potentially make the disease worse. The mechanism that causes that risk is not fully understood and is one of the stumbling blocks that has prevented the successful development of a coronavirus vaccine. Normally, researchers would take months to test for the possibility of vaccine enhancement in animals. Given the urgency to stem the spread of the new coronavirus, some drug makers are moving straight into small-scale human tests, without waiting for the completion of such animal tests (see Steenhuysen 2020).
In the U.S., the National Institute of Allergy and Infectious Diseases (NIAID) within the National Institutes of Health (N.I.H.) is overseeing the funding of federal research and response to COVID-19. There are also some companies in the U.S. that are conducting their own COVID-19 research. Internationally, the U.K. Medicines and Healthcare products Regulatory Agency (MHRA) and the European Medicines Agency (E.M.A.) are supporting efforts to develop therapies against COVID-19. In general, it is expected that it takes 12-18 months to develop a vaccine. This tracker lists the major vaccine candidates currently in development.
Chapter 3: Leveraging NGS-Technology in the Fight Against COVID-19
The constellation that made SARS-CoV-2 a pandemic agent lies in its high transmissibility during the incubation period in combination with a mild to an asymptomatic course in most cases of its disease COVID-19 (see Rothe et al. 2020, Chen et al.). As a novel virus, the pathogen meets naive human immune systems worldwide, so that there is no disruption of the
infection chains by a significant proportion of immune individuals in the population. However, the mostly mild course of the disease contrasts with a not inconsiderable proportion that have a severe or critical disease. According to a report on 73,314 cases of the Chinese Centre for Disease Control and Prevention, about 14% of cases are severe with dyspnea and viral pneumonia and 5% critical with a need for intensive care (Wu et al. 2020).
In order to avoid the collapse of health systems, unprecedented, drastic measures are being taken around the world to reduce human contacts to a minimum in order to curb the number of new infections. The downside of these efforts is a freeze in social life with unforeseeable consequences for society. The SARS-CoV-2 pandemic thus presents us with the historic choice of either drastically overburdening our healthcare capacity or driving the global economy into a global recession with far-reaching socio-economic consequences.
With the prospect of such a scenario, the question, “how can we reduce the number of new infections and return to social life,” is crucial to get back to our socio-economic balance.
Answers to this must already be found today, where measures based on contact isolation cannot be consistently implemented. This applies in particular to medical and social community facilities, hospitals, elderly homes, and social housing for people in need. Contact barriers cannot be implemented consistently as personal interactions and assistance are an essential component of the care provided for these facilities.
The role of hospitals in SARS-CoV-2 infection chains has not yet been systematically described, but putting together the facts, it is clear that there is a high potential of
transmission to vulnerable groups of patients who are at risk of severe infection. Systematic testing of patients and staff even without clinical symptoms, depending on a patient group adapted risk assessment, is, therefore, an essential cornerstone for the rapid detection of infections. Especially in the hospital, patients can develop a veiling mix of symptoms due to their underlying disease and in some cases cannot be asked about important symptoms of lighter respiratory infections such as sore throat. However, comprehensive testing alone only makes sense if, preferably, every case leads to an infection-chain search in both patients and employees with the consistent isolation and placing in quarantine critical contacts. A pandemic pathogen provides a unique challenge in tracing the source of an infection. When there are multiple, several competing sources of infection are possible: has patient A really infected employee B or patient C or is there possibly an unrecognized co-patient and employee B has acquired the infection in the home environment? The central hospital hygiene question is always the same: Have I ever seen this pathogen in another case before? In this respect, NGS, together with smart, customized bioinformatics tools, enables us to enter a new dimension of infection surveillance.
With 12 cases of COVID-19 sequenced with NGS, the complete genome and unique variants of each patient’s virus can be compared. With a dendrogram analysis, as shown in Figure 4, clustering is performed on all samples comparing variants by euclidian distance. The visible grouping helps to separate the cases into two main clusters. Patient A and Patient B are unlikely to belong to the same infection chain. With Patient C, no other case is associated. Instead, Patient D-G came from the same long-term care facility, giving a strong hint at an outbreak there.
Together with classic instruments of tracking infection chains such as timelines of the patient’s hospital stay and the identification of all their contacts, hygienists are able to clarify the entire infection process in their facility.
Dendrogram analysis is particularly suitable for distinguishing infections that have been detected in a short period of time. Since it lies in the nature of a virus to acquire new random mutations over time, having complete sequences allows for the identification of known isolates and novel variants to discover unknown isolates. Genetic information also provides insight into the evolutionary lineage of isolates, which can identify a temporal dimension in the relationship between samples. The scientific community publishes SARS-CoV-2 sequences on a regular basis, along with the date and location the sample was collected. Golden Helix’s NGS software is able to determine to what particular previous occurrence a particular sample is most similar. In this way, chains of infection can be presented in a regional, national, or international context beyond their local context (Figure 5). This supports highly specific and accurate contact-tracing and identification of potential hotspots.
With today’s sequencing methods, complete virus genomes can be sequenced very easily from preparations for PCR testing so that the tracking of infection chains at the molecular level is able to be accomplished practically in real-time.
In this way, the comparative analysis of NGS data can help to improve treatment processes in relation to their transmission risk. This can lead to the identification of training deficiencies, e.g., in the use of personal protective equipment. It can also be used to detect infection rates in other facilities: for example, when infections occur in patients who are taken from a particular nursing home or other hospitals.
By sharing this data between institutions (connected hospitals, associated long term facilities, etc.), even novel networks can be created to share data and improve response times to current and future infection events and even the next pandemic.
Clinical Testing with NGS Sequencing
In concert with the rapid-diagnosis capabilities of RT-PCR tests currently in use, the capabilities of NGS machines may be employed to capture the complete genomes of the virus as it spreads and evolves as well as confirm the virus presence. Longer-term, these collected virus samples will provide critical data to understand the evolution of the virus, its biological properties, and to aid in the development of therapeutic drugs and even vaccines.
The key to scaling up any NGS pipeline for clinical diagnosis and rapid turnaround is the automation of as many parts of the bioinformatics process as possible. This not only reduces time-to-result but also removes the potential for human error in otherwise manually performed steps. Today’s state of the art, commercial pipeline tools can be used as the high-throughput automation tool enabling repeatable workflows of the NGS analysis capabilities previously discussed.
With molecular barcoding, the throughput of NGS machines allows for many samples to be multiplexed on a single run. The automated analysis workflow after sequencing run may look like:
- De-multiplex raw reads into per-sample sequence files
- Align sequence reads to the SARS-CoV-2
- Remove duplicate reads
- Call variants from the reference sequence
- Run customized workflow with VSPipeline to:
◆ Call Positive/Negative for SARS-CoV-2 based on coverage analysis
◆ Analysis of strain based on variants
◆ Output PDF clinical report
While these workflows presented here are certainly not comprehensive, they provide examples of common analysis strategies for different use cases of COVID-19 NGS sequencing.
Chapter 4: Summary
COVID-19 challenges the healthcare system, governments, and the global economy in an unprecedented way. As the novel coronavirus SARS-CoV-2 has just recently been discovered, vast resources are being directed towards understanding this new disease and the virus that causes it.
This article outlines the current state of our understanding of COVID-19 and SARS-CoV-2. Chapter 1 gives a brief introduction to this topic. Chapter 2 summarizes key facts about COVID-19; it reviews the epidemiology, reservoir hosts, transmission routes, and clinical manifestation. Chapter 3 answers the question of how Next- Generation Sequencing can be utilized in the clinic for diagnostic and tracking purposes.
By the time you read this sentence, there will be new findings, studies, and research papers. Also, any of the population-level statistics that have been cited will be dwarfed by the numbers that are reported by the time you read these words. But one thing is certain: Next-Gen Sequencing technologies will allow us to gain a deeper understanding of this virus and to develop advanced diagnostic capabilities to help patients and provide us the ability to conduct research at the same time.
Dr. Christiane Scherer is a medical microbiologist and senior hospital hygienist at the Evangelischen Klinikum Bethel, a German hospital with around 1,750 beds in 26 specialist clinics, three institutes and 11 interdisciplinary centers. Since the beginning of the COVID-19 pandemic, she has been part of the hospital’s task force for managing the crisis and is a member of the pandemic working group of the city of Bielefeld. She heads the clinic’s microbiological laboratory, which offers laboratory diagnostics and advice in the fields of serology, bacteriology, parasitology, virology, and molecular biological diagnostics. Dr. Scherer completed her specialist training at the Institute of Medical Microbiology at the University of Essen, where she received her doctorate in the Department of Microbiology. She also holds a Master’s degree in Health Administration from Bielefeld University. Since 2004, she has held courses for medical students, doctors, and nurses, in particular on topics of infection diagnosis, antimicrobial resistance, and infection prevention. She participates in various working groups in the curriculum development of the medical faculty of Bielefeld University, which is in the process of being founded.
Dr. Andreas Scherer is the C.E.O. of Golden Helix. The company has been delivering industry-leading bioinformatics solutions for the advancement of life science research and translational medicine for over two decades. Its innovative technologies and analytic services empower clinicians and scientists at all levels to derive meaning from the rapidly increasing volumes of genomic data produced from next-generation sequencing and microarrays. With its solutions, hundreds of the world’s hospitals, testing labs, academic research organizations, and governments are able to harness the full potential of genomics to identify the cause of disease, develop genomic diagnostics, and advance the quest for personalized medicine. Golden Helix products and services have been cited in thousands of peer-reviewed publications. He is also Managing Partner of Salto Partners, a management consulting firm headquartered in the D.C. metro area. He has extensive experience successfully managing growth as well as orchestrating complex turnaround situations. Dr. Scherer holds a Ph.D. in computer science from the University of Hagen, Germany, and a Master of Computer Science from the University of Dortmund, Germany. He is author and co-author of over 20 international publications and has written books on project management, the Internet, and artificial intelligence. His latest book, “Be Fast Or Be Gone,” is a prizewinner in the 2012 Eric Hoffer Book Awards competition, and has been named a finalist in the 2012 Next Generation Indie Book Awards!.
Adam J Kucharski, Timothy W Russell, Charlie Diamond, Yang Liu, John Edmunds, Sebastian Funk, Rosalind M Eggo, Fiona Sun, Mark Jit, James D Munday, Nicholas Davies, Amy Gimma, Kevin van Zandvoort, Hamish Gibbs, Joel Hellewell, Christopher I Jarvis, Sam Clifford, Billy
J Quilty, Nikos I Bosse, Sam Abbott, Petra Klepac, Stefan Flasche. (2020) Early dynamics of transmission and control of COVID-19: a mathematical modelling study. The Lancet Infectious Diseases.
Alessia Lai, Annalisa Bergna, Carla Acciarri, Massimo Galli, Gianguglielmo Zehender. (2020) Early phylogenetic estimate of the effective reproduction number of SARS‐CoV‐2. Journal of Medical Virology 2020.
Anita Patel, Daniel B. Jernigan, , Fatuma Abdirizak, Glen Abedi, Sharad Aggarwal, Denise Albina, Elizabeth Allen, Lauren Andersen, Jade Anderson, Megan Anderson, Tara Anderson, Kayla Anderson, Ana Cecilia Bardossy, Vaughn Barry, Karlyn Beer, Michael Bell, Sherri Berger, Joseph Bertulfo, Holly Biggs, Jennifer Bornemann, Josh Bornstein, Willie Bower, Joseph Bresee, Clive Brown, Alicia Budd, Jennifer Buigut, Stephen Burke, Rachel Burke, Erin Burns, Jay Butler, Russell Cantrell, Cristina Cardemil, Jordan Cates, Marty Cetron, Kevin Chatham‐Stephens, Kevin Chatham‐Stevens, Nora Chea, Bryan Christensen, Victoria Chu, Kevin Clarke, Angela Cleveland, Nicole Cohen, Max Cohen, Amanda Cohn, Jennifer Collins, Erin Conners, Aaron Curns, Rebecca Dahl, Walter Daley, Vishal Dasari, Elizabeth Davlantes, Patrick Dawson, Lisa Delaney, Matthew Donahue, Chad Dowell, Jonathan Dyal, William Edens, Rachel Eidex, Lauren Epstein, Mary Evans, Ryan Fagan, Kevin Farris, Leora Feldstein, LeAnne Fox, Mark Frank, Brandi Freeman, Alicia Fry, James Fuller, Romeo Galang,
Sue Gerber, Runa Gokhale, Sue Goldstein, Sue Gorman, William Gregg, William Greim, Steven Grube, Aron Hall, Amber Haynes, Sherrasa Hill, Jennifer Hornsby‐Myers, Jennifer Hunter, Christopher Ionta, Cheryl Isenhour, Max Jacobs, Kara Jacobs SliTha, Daniel Jernigan, Michael Jhung, Jamie Jones‐Wormley, Anita Kambhampati, Shifaq Kamili, Pamela Kennedy, Charlotte Kent, Marie Killerby, Lindsay Kim, Hannah Kirking, Lisa Koonin, Ram Koppaka, Christine Kosmos, David Kuhar, Wendi Kuhnert‐Tallman, Stephanie Kujawski, Archana Kumar, Alexander Landon, Leslie Lee, Jessica Leung, Stephen Lindstrom, Ruth Link‐Gelles, Joana Lively, Xiaoyan Lu, Brian Lynch, Lakshmi Malapati, Samantha Mandel, Brian Manns, Nina Marano, Mariel Marlow, Barbara Marston, Nancy McClung, Liz McClure, Emily McDonald, Oliva McGovern, Nancy Messonnier, Claire Midgley, Danielle Moulia, Janna Murray, Kate Noelte, Michelle Noonan‐Smith, Kristen Nordlund, Emily Norton, Sara Oliver, Mark Pallansch, Umesh Parashar, Anita Patel, Manisha Patel, Kristen Pettrone, Taran Pierce, Harald Pietz, Satish Pillai, Lewis Radonovich, Sarah Reagan‐Steiner, Amy Reel, Heather Reese, Brian Rha, Philip Ricks, Melissa Rolfes, Shahrokh Roohi, Lauren Roper, Lisa Rotz, Janell Routh, Senthil Kumar Sakthivel, Luisa Sarmiento, Jessica Schindelar, Eileen Schneider, Anne Schuchat, Sarah Scott, Varun Shetty, Caitlin Shockey, Jill Shugart, Mark Stenger, Matthew Stuckey, Brittany Sunshine, Tamara Sykes, Timothy Uyeki, Grace Vahey, Amy Valderrama, Julie Villanueva, Tunicia Walker, Megan Wallace, Lijuan Wang, John Watson, Angie Weber, Cindy Weinbaum, William Weldon, Caroline Westnedge, Brett Whitaker, Michael Whitaker, Alcia Williams,
Ian Willams, Karen Wong, Amy Xie, Anna Yousef. (2020) Initial public health response and interim clinical guidance for the 2019 novel
coronavirus outbreak — United States, December 31, 2019–February 4, 2020. American Journal of Transplantation 20:3, 889-895.
- Coutard, Valle, X. de Lamballerie, B. Canard, N.G. Seidah, E. Decroly. (2020) The spike glycoprotein of the new coronavirus 2019- nCoV contains a furin-like cleavage site absent in CoV of the same clade. Antiviral Research 176, 104742.
Catrin Sohrabi, Zaid Alsafi, Niamh O’Neill, Mehdi Khan, Ahmed Kerwan, Ahmed Al-Jabir, Christos Iosifidis, Riaz Agha. (2020) World Health Organization declares global emergency: A review of the 2019 novel coronavirus (COVID-19). International Journal of Surgery 76, 71-76.
Chen, T., Wu, D., Chen, H., Yan, W., Yang, D., Chen, G., Ma, K., Xu, D., Yu, H., Wang, H., Wang, T., Guo, W., Chen, J., Ding, C., Zhang, X., Huang, J., Han, M., Li, S., Luo, X., Zhao, J., … Ning, Q. (2020). Clinical characteristics of 113 deceased patients with coronavirus disease 2019: retrospective study. B.M.J. (Clinical research ed.), 368, m1091. https://www.bmj.com/content/368/bmj.m1091
Cui J, Li F, Shi ZL 2019 Origin and evolution of pathogenic coronaviruses, Nat Rev Microbiol 2019 Mar;17(3): 181-192.
Dawei Wang, Bo Hu, Chang Hu, Fangfang Zhu, Xing Liu, Jing Zhang, Binbin Wang, Hui Xiang, Zhenshun Cheng, Yong Xiong, Yan Zhao, Yirong Li, Xinghuan Wang, Zhiyong Peng. (2020) Clinical Characteristics of 138 Hospitalized Patients With 2019 Novel
Coronavirus–Infected Pneumonia in Wuhan, China. JAMA 323:11, 1061.
Di Wu, Tiantian Wu, Qun Liu, Zhicong Yang. (2020) The SARS-CoV-2 outbreak: what we know. International Journal of Infectious Diseases.
Fei Zhou, Ting Yu, Ronghui Du, Guohui Fan, Ying Liu, Zhibo Liu, Jie Xiang, Yeming Wang, Bin Song, Xiaoying Gu, Lulu Guan, Yuan Wei, Hui Li, Xudong Wu, Jiuyang Xu, Shengjin Tu, Yi Zhang, Hua Chen, Bin Cao. (2020) Clinical course and risk factors for mortality of adult inpatients with COVID-19 in Wuhan, China: a retrospective cohort study. The Lancet.
Fuk-Woo CJ, Shuofeng Y, Kin-Hang K et. Al (2020) A familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person: as study of a family cluster. The Lancet. 2020; 515-523.
Guan WJ, Ni ZY, Hu Y et al (2020) Clinical Characteristics of Coronavirus Disease in 2019 in China, N Engl J Med. 2020.
Harald Brüssow. (2020) The Novel Coronavirus – A Snapshot of Current Knowledge. Microbial Biotechnology.
Harding, A. Lanese, N. (2020) The 12 deadliest viruses on Earth, Live Sciences, March 2020.
Hengbo Zhu, Li Wei, Ping Niu. (2020) The novel coronavirus outbreak in Wuhan, China. Global Health Research and Policy 5:1.
Hiroshi Nishiura, Natalie M. Linton, Andrei R. Akhmetzhanov. (2020) Serial interval of novel coronavirus (COVID-19) infections. International Journal of Infectious Diseases 93, 284-286.
Huan Liang, Ganesh Acharya. (2020) Novel corona virus disease (COVID‐19) in pregnancy: What clinical recommendations to follow?. Acta Obstetricia et Gynecologica Scandinavica 99:4, 439-442.
Huang et. al. (2020) Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China, The Lancelet, Vol 395 February 2020.
Huwen Wang, Zezhou Wang, Yinqiao Dong, Ruijie Chang, Chen Xu, Xiaoyue Yu, Shuxian Zhang, Lhakpa Tsamlag, Meili Shang, Jinyan Huang, Ying Wang, Gang Xu, Tian Shen, Xinxin Zhang, Yong Cai. (2020) Phase-adjusted estimation of the number of Coronavirus Disease 2019 cases in Wuhan, China. Cell Discovery 6:1.
Jennifer Harcourt, Azaibi Tamin, Xiaoyan Lu, Shifaq Kamili, Senthil K. Sakthivel, Janna Murray, Krista Queen, Ying Tao, Clinton R. Paden, Jing Zhang, Yan Li, Anna Uehara, Haibin Wang, Cynthia Goldsmith, Hannah A. Bullock, Lijuan Wang, Brett Whitaker, Brian Lynch, Rashi
Gautam, Craig Schindewolf, Kumari G. Lokugamage, Dionna Scharton, Jessica A. Plante, Divya Mirchandani, Steven G. Widen, Krishna Narayanan, Shinji Makino, Thomas G. Ksiazek, Kenneth S. Plante, Scott
- Weaver, Stephen Lindstrom, Suxiang Tong, Vineet D. Menachery, Natalie Thornburg. (2020) Severe Acute Respiratory Syndrome Coronavirus 2 from Patient with 2019 Novel Coronavirus Disease, United States. Emerging Infectious Diseases 26:6.
Jiang, F., Deng, L., Zhang, L. et al. Review of the Clinical Characteristics of Coronavirus Disease 2019 (COVID-19). J GEN INTERN MED (2020).
Joseph T. Wu, Kathy Leung, Mary Bushman, Nishant Kishore, Rene Niehus, Pablo M. de Salazar, Benjamin J. Cowling, Marc Lipsitch, Gabriel M. Leung. (2020) Estimating clinical severity of COVID-19 from the transmission dynamics in Wuhan, China. Nature Medicine 162.
Jun She, Jinjun Jiang, Ling Ye, Lijuan Hu, Chunxue Bai, Yuanlin Song. (2020) 2019 novel coronavirus of Pneumonia in Wuhan, China: emerging attack and management strategies. Clinical and Translational Medicine 9:1.
Kaiyuan Sun, Jenny Chen, Cécile Viboud. (2020) Early epidemiological analysis of the coronavirus disease 2019 outbreak based on crowdsourced data: a population-level observational study. The Lancet Digital Health 2:4, e201-e208.
Kaiyue Diao, Peilun Han, Tong Pang, Yuan Li, Zhigang Yang. (2020) HRCT imaging features in representative imported cases of 2019 novel coronavirus pneumonia. Precision Clinical Medicine 3:1, 9-13.
Kupferschmidt K, Cohen J (2020) WHO launches globar megatrial of the four most promising coronavirus treatments, Science Magazine, March 22, 2020.
Lei Tang, Xiaoyong Zhang, Yvquan Wang, Xianchun Zeng. (2020) Severe COVID-19 Pneumonia: Assessing Inflammation Burden with Volume- rendered Chest CT. Radiology: Cardiothoracic Imaging 2:2, e200044.
Li Heng (2013) Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM, 2013 arXiv:1303.3997
Lin Li, Lixin Qin, Zeguo Xu, Youbing Yin, Xin Wang, Bin Kong, Junjie Bai, Yi Lu, Zhenghan Fang, Qi Song, Kunlin Cao, Daliang Liu, Guisheng Wang, Qizhong Xu, Xisheng Fang, Shiqin Zhang, Juan Xia, Jun Xia. (2020) Artificial Intelligence Distinguishes COVID-19 from Community Acquired Pneumonia on Chest CT. Radiology, 200905.
Mary E Wilson, Lin H Chen. (2020) Travellers give wings to novel coronavirus (2019-nCoV). Journal of Travel Medicine 27:2.
Moritz U. G. Kraemer, Chia-Hung Yang, Bernardo Gutierrez, Chieh-Hsi Wu, Brennan Klein, David M. Pigott, , Louis du Plessis, Nuno R. Faria, Ruoran Li, William P. Hanage, John S. Brownstein, Maylis Layan, Alessandro Vespignani, Huaiyu Tian, Christopher Dye, Oliver G. Pybus, Samuel V. Scarpino. (2020) The effect of human mobility and control measures on the COVID-19 epidemic in China. Science, eabb4218.
Nathan Kapata, Chikwe Ihekweazu, Francine Ntoumi, Tajudeen Raji, Pascalina Chanda-Kapata, Peter Mwaba, Victor Mukonka, Matthew Bates, John Tembo, Victor Corman, Sayoki Mfinanga, Danny Asogun, Linzy Elton, Liã Bárbara Arruda, Margaret J. Thomason, Leonard Mboera, Alexei Yavlinsky, Najmul Haider, David Simons, Lara Hollmann, Swaib A. Lule, Francisco Veas, Muzamil Mahdi Abdel Hamid, Osman Dar, Sarah Edwards, Francesco Vairo, Timothy
- McHugh, Christian Drosten, Richard Kock, Giuseppe Ippolito, Alimuddin Zumla. (2020) Is Africa prepared for tackling the COVID-19 (SARS-CoV-2) Lessons from past outbreaks, ongoing pan-African public health efforts, and implications for the future. International Journal of Infectious Diseases 93, 233-236.
Nian Shao, Min Zhong, Yue Yan, HanShuang Pan, Jin Cheng, Wenbin Chen. (2020) Dynamic models for Coronavirus Disease 2019 and data analysis. Mathematical Methods in the Applied Sciences 50.
Parham Habibzadeh, Emily K. Stoneman. (2020) The Novel Coronavirus: A Bird’s Eye View. The International Journal of Occupational and Environmental Medicine 11:2, 65-71.
Phelan Al, Kat Rebecca, Gostin L (2020) The Novel Coronavirus Origninating in Wuhan China: Challenges for Global Health Governance, Jama 2020.
Poplin et al (2017) Scaling accurate genetic variant discovery to tens of thousands of samples, 2017 bioRxiv
Qianying Lin, Shi Zhao, Daozhou Gao, Yijun Lou, Shu Yang, Salihu S. Musa, Maggie H. Wang, Yongli Cai, Weiming Wang, Lin Yang, Daihai He. (2020) A conceptual model for the coronavirus disease 2019 (COVID-19) outbreak in Wuhan, China with individual reaction and governmental action. International Journal of Infectious Diseases 93, 211-216.
Qingmei Han, Qingqing Lin, Shenhe Jin, Liangshun You. (2020) Coronavirus 2019-nCoV: A brief perspective from the front line. Journal of Infection 80:4, 373-377.
Qun Li, M.Med., Xuhua Guan, Ph.D., Peng Wu, Ph.D., Xiaoye Wang, M.P.H., Lei Zhou, M.Med., Yeqing Tong, Ph.D., Ruiqi Ren, M.Med., Kathy S.M. Leung, Ph.D., Eric H.Y. Lau, Ph.D., Jessica Y. Wong, Ph.D., Xuesen Xing, Ph.D., Nijuan Xiang, M.Med., Yang Wu, M.Sc., Chao Li, M.P.H., Qi Chen, M.Sc., Dan Li, M.P.H., Tian Liu, B.Med., Jing Zhao, M.Sc., Man Liu, M.Sc., Wenxiao Tu, M.Med., Chuding Chen, M.Sc., Lianmei Jin, M.Med., Rui Yang, M.Med., Qi Wang, M.P.H., Suhua Zhou, M.Med., Rui Wang, M.D., Hui Liu, M.Med., Yinbo Luo, M.Sc., Yuan Liu, M.Med., Ge Shao, B.Med., Huan Li, M.P.H., Zhongfa Tao, M.P.H., Yang Yang, M.Med., Zhiqiang Deng, M.Med., Boxi Liu, M.P.H., Zhitao Ma, M.Med., Yanping Zhang, M.Med., Guoqing Shi, M.P.H., Tommy
T.Y. Lam, Ph.D., Joseph T. Wu, Ph.D., George F. Gao, D.Phil., Benjamin J. Cowling, Ph.D., Bo Yang, M.Sc., Gabriel M. Leung, M.D., and Zijian Feng, M.Med, New England Journal of Medicine, March 2020. Vol. 382, No. 13
Rothe, C., Schunk, M., Sothmann, P., Bretzel, G., Froeschl, G., Wallrauch, C., Zimmer, T., Thiel, V., Janke, C., Guggemos, W., Seilmaier, M., Drosten, C., Vollmar, P., Zwirglmaier, K., Zange, S., Wölfel, R., & Hoelscher, M. (2020). Transmission of 2019-nCoV Infection from an Asymptomatic Contact in Germany. The New England journal of medicine, 382(10), 970–971. https://doi.org/10.1056/NEJMc2001468
Roy M Anderson, Hans Heesterbeek, Don Klinkenberg, T Déirdre Hollingsworth. (2020) How will country-based mitigation measures influence the course of the COVID-19 epidemic?. The Lancet 395:10228, 931-934.
Rui Li, Songlin Qiao, Gaiping Zhang. (2020) Analysis of angiotensin- converting enzyme 2 (ACE2) from different species sheds some light on cross-species receptor usage of a novel coronavirus 2019-nCoV. Journal of Infection 80:4, 469-496.
Rui Liu, Huan Han, Fang Liu, Zhihua Lv, Kailang Wu, Yingle Liu, Yong Feng, Chengliang Zhu. (2020) Positive rate of RT-PCR detection of SARS-CoV-2 infection in 4880 cases from one hospital in Wuhan, China, from Jan to Feb 2020. Clinica Chimica Acta 505, 172-175.
Sasmita Poudel Adhikari, Sha Meng, Yu-Ju Wu, Yu-Ping Mao, Rui-Xue Ye, Qing-Zhi Wang, Chang Sun, Sean Sylvia, Scott Rozelle, Hein Raat, Huan Zhou. (2020) Epidemiology, causes, clinical manifestation and diagnosis, prevention and control of coronavirus disease (COVID-19) during the early outbreak period: a scoping review. Infectious Diseases of Poverty 9:1.
Scherer, A. (2020a) “Genetic Analysis of the COVID-19 Virus and Other Pathogens”, ISBN 978-0-9986882-8-2, Golden Helix, 2020
Scherer, A. (2020b) “Leveraging Next-Generation Sequencing Technology in the Fight Against COVID-19”, Clinical Lab Manager, May 4, 2020.
Shen K, Yang Y, Wang T (2020) Diagnosis, treatment and prevention of 2019 novel coronavirus infection in children: experts’ consensus statement. World J Pediatr. 2020Sheng Zhang, MengYuan Diao, Wenbo Yu, Lei Pei, Zhaofen Lin, Dechang Chen. (2020) Estimation of the reproductive number of novel coronavirus (COVID-19) and the probable outbreak size on the Diamond Princess cruise ship: A data-driven analysis. International Journal of Infectious Diseases 93, 201-204.
Sijia Tian, Nan Hu, Jing Lou, Kun Chen, Xuqin Kang, Zhenjun Xiang, Hui Chen, Dali Wang, Ning Liu, Dong Liu, Gang Chen, Yongliang Zhang, Dou Li, Jianren Li, Huixin Lian, Shengmei Niu, Luxi Zhang, Jinjun Zhang. (2020) Characteristics of COVID-19 infection in Beijing. Journal of Infection 80:4, 401-406.
Simiao Chen, Juntao Yang, Weizhong Yang, Chen Wang, Till Bärnighausen. (2020) COVID-19 control in China during mass population movements at New Year. The Lancet395:10226, 764-766.
Steenhuysen, J (2020) AS pressure for coronavirus Vaccine mounts, scientists debate risks of accelerating testing, Reuters.com, Health News, March 11, 2020.
Tang X, Wu C, Lix X, Song Y, Yao X, Wu X et. Al (2020) On the origin and continuing evolution of SARS-CoV-2, Natl Science Rev, 2020.
Tanu Singhal. (2020) A Review of Coronavirus Disease-2019 (COVID-19). The Indian Journal of Pediatrics 87:4, 281-286.
Tian-Mu Chen, Jia Rui, Qiu-Peng Wang, Ze-Yu Zhao, Jing-An Cui, Ling Yin. (2020) A mathematical model for simulating the phase- based transmissibility of a novel coronavirus. Infectious Diseases of Poverty 9:1.
Weilong Shang, Yi Yang, Yifan Rao, Xiancai Rao. (2020) The outbreak of SARS-CoV-2 Pneumonia calls for viral vaccines. npj Vaccines 5:1.
Wen-Chien Ko, Jean-Marc Rolain, Nan-Yao Lee, Po-Lin Chen, Ching-Tai Huang, Ping-Ing Lee, Po-Ren Hsueh. (2020) Arguments in favour of remdesivir for treating SARS-CoV-2 infections. International Journal of Antimicrobial Agents, 105933.
Wu Z, McGoogan JM. (2020). Characteristics of and Important Lessons From the Coronavirus Disease 2019 (COVID-19) Outbreak in China:
Summary of a Report of 72 314 Cases From the Chinese Center for Disease Control and Prevention. JAMA;323(13):1239–1242. doi:10.1001/jama.2020.2648
Xingguang Li, Junjie Zai, Qiang Zhao, Qing Nie, Yi Li, Brian T. Foley, Antoine Chaillon. (2020) Evolutionary history, potential intermediate animal host, and cross‐species analyses of SARS‐CoV‐2. Journal of Medical Virology 6.
Xingguang Li, Wei Wang, Xiaofang Zhao, Junjie Zai, Qiang Zhao, Yi Li, Antoine Chaillon. (2020) Transmission dynamics and evolutionary history of 2019‐nCoV. Journal of Medical Virology 92:5, 501-511.
Yan-Rong Guo, Qing-Dong Cao, Zhong-Si Hong, Yuan-Yang Tan, Shou- Deng Chen, Hong-Jun Jin, Kai-Sen Tan, De-Yun Wang, Yan Yan. (2020) The origin, transmission and clinical therapies on coronavirus disease 2019 (COVID-19) outbreak – an update on the status. Military Medical Research 7:1.
Yi Yang, Weilong Shang, Xiancai Rao. (2020) Facing the COVID‐19 outbreak: What should we know and what could we do?. Journal of Medical Virology.
Yicheng Fang, Huangqi Zhang, Yunyu Xu, Jicheng Xie, Peipei Pang, Wenbin Ji. (2020) C.T. Manifestations of Two Cases of 2019 Novel Coronavirus (2019-nCoV) pneumonia. Radiology 295:1, 208-209.
Ying Liu, Albert A Gayle, Annelies Wilder-Smith, Joacim Rocklöv. (2020) The reproductive number of COVID-19 is higher compared to SARS coronavirus. Journal of Travel Medicine 27:2.
Zachary T. Bloomgarden. (2020) Diabetes and COVID‐19. Journal of Diabetes 12:4, 347-348.
Zhou et al (2020) A pneumonia outbreak associated with a new coronavirus of probable bat origin, Nature 2020 Mar; 579(7798): 2070:273.
Zhu H, Wang L, Fang C. etc al (2020) Clinical analysis of 10 neonates born to mothers with 2019-nCoV Pneumonia, Translational Pediatrics, 2020; 9: 51-60
Zhu et al (2020) A Novel Coronavirus form Patients with Pneumonia in China, 2019. NEJM, January 24, 2020.