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Srinivasa VR, Griffith MP, Sundermann AJ, Mills E, Raabe NJ, Waggle KD, Shutt KA, Phan T, Wang-Erickson AF, Snyder GM, Van Tyne D, Pless LL, Harrison LH. Genomic Epidemiology of Healthcare-Associated Respiratory Virus Infections. MEDRXIV : THE PREPRINT SERVER FOR HEALTH SCIENCES 2025:2025.04.20.25325828. [PMID: 40313286 PMCID: PMC12045434 DOI: 10.1101/2025.04.20.25325828] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 05/03/2025]
Abstract
Background Respiratory virus transmission in healthcare settings is not well understood. To investigate the transmission dynamics of common healthcare-associated respiratory virus infections, we performed retrospective whole genome sequencing (WGS) surveillance at one pediatric and two adult teaching hospitals in Pittsburgh, PA. Methods From January 2, 2018, to January 4, 2020, nasal swab specimens positive for rhinovirus, influenza, human metapneumovirus (HMPV), or respiratory syncytial virus (RSV) from patients hospitalized for ≥3 days were sequenced on Illumina platform. High-quality genomes were assessed for genetic relatedness using ≤3 single nucleotide polymorphisms (SNPs) cut-off, except for rhinovirus (10 SNPs). Patient health records were reviewed for genetically related clusters to identify epidemiological connections. Results We collected 436 viral specimens from 359 patients: rhinovirus (n=291), influenza (n=50), HMPV (n=47), and RSV (n=48). Of these, 55% (197/359 patients) were from pediatric hospital and 45% from adult hospitals. Patients ranged in age from 14 days to 93 years, 61% were male, and 74% were white. WGS was performed on 61.2% (178/291) rhinovirus, 78% (39/50) influenza, 92% (44/48) RSV, and all HMPV specimens. Among high-quality genomes, we identified 14 genetically related clusters involving 36 patients, ranging in size from 2-5 patients. We identified common epidemiological links for 53% (19/36) of clustered patients; 63% (12/19) patients had same-unit stay, 26% (5/19) had overlapping hospital stays, and 11% (2/19) shared common provider. On average, genetically related clusters spanned 16 days (range:0-55 days). Conclusion WGS offered insights into respiratory virus transmission dynamics. These advancements could potentially improve infection prevention and control strategies, leading to enhanced patient safety and healthcare outcomes.
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Affiliation(s)
- Vatsala Rangachar Srinivasa
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
- Department of Epidemiology, School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA
| | - Marissa P. Griffith
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Alexander J. Sundermann
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Department of Epidemiology, School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA
| | - Emma Mills
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Nathan J. Raabe
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
- Department of Epidemiology, School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA
| | - Kady D. Waggle
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Kathleen A. Shutt
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Tung Phan
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Anna F. Wang-Erickson
- Department of Epidemiology, School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA
- Department of Pediatrics, Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
- Institute of Infection, Inflammation, and Immunity in Children (i4Kids), Pittsburgh, PA
| | - Graham M. Snyder
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
- Department of Infection Control and Hospital Epidemiology, UPMC, Pittsburgh, PA
| | - Daria Van Tyne
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Lora Lee Pless
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Lee H. Harrison
- Microbial Genomic Epidemiology Laboratory, Center for Genomic Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA
- Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
- Department of Epidemiology, School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA
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Goya S, Wendm ST, Xie H, Nguyen TV, Barnes S, Shankar RR, Sereewit J, Cruz K, Pérez-Osorio AC, Mills MG, Greninger AL. Genomic Epidemiology and Evolution of Rhinovirus in Western Washington State, 2021-2022. J Infect Dis 2025; 231:e154-e164. [PMID: 38963827 PMCID: PMC11793040 DOI: 10.1093/infdis/jiae347] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2024] [Revised: 06/25/2023] [Accepted: 07/02/2024] [Indexed: 07/06/2024] Open
Abstract
BACKGROUND Human rhinoviruses (RVs) primarily cause the common cold, but infection outcomes vary from subclinical to severe cases, including asthma exacerbations and fatal pneumonia in individuals who are immunocompromised. To date, therapeutic strategies have been hindered by the high diversity of serotypes. Global surveillance efforts have traditionally focused on sequencing VP1 or VP2/VP4 genetic regions, leaving gaps in our understanding of RV genomic diversity. METHODS We sequenced 1078 RV genomes from nasal swabs of symptomatic and asymptomatic individuals to explore viral evolution during 2 epidemiologically distinct periods in Washington State: when the COVID-19 pandemic affected the circulation of other seasonal respiratory viruses except for RV (February-July 2021) and when the seasonal viruses reemerged with the severe outbreak of respiratory syncytial virus and influenza (November-December 2022). We constructed maximum likelihood and BEAST phylodynamic trees to characterize intragenotype evolution. RESULTS We detected 99 of 168 known genotypes and observed intergenotypic recombination and genotype cluster swapping from 2021 to 2022. We found a significant association between the presence of symptoms and viral load but not with RV species or genotype. Phylodynamic trees, polyprotein selection pressure, and Shannon entropy revealed cocirculation of divergent clades within genotypes with high amino acid constraints throughout the polyprotein. CONCLUSIONS Our study underscores the dynamic nature of RV genomic epidemiology within a localized geographic region, as >20% of existing genotypes within each RV species cocirculated each studied month. Our findings also emphasize the importance of investigating correlations between RV genotypes and serotypes to understand long-term immunity and cross-protection.
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Affiliation(s)
- Stephanie Goya
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Seffir T Wendm
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Hong Xie
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Tien V Nguyen
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Sarina Barnes
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Rohit R Shankar
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Jaydee Sereewit
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Kurtis Cruz
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Ailyn C Pérez-Osorio
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Margaret G Mills
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
| | - Alexander L Greninger
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington
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Liu W, Zhang E, Li W, Lv R, Lin Y, Xu Y, Li J, Lai Y, Jiang Y, Lin S, Wang X, Zhou P, Song Y, Shen W, Sun Y, Li Y. Advances and challenges of mpox detection technology. BIOSAFETY AND HEALTH 2024; 6:260-269. [PMID: 40078738 PMCID: PMC11895016 DOI: 10.1016/j.bsheal.2024.09.005] [Citation(s) in RCA: 5] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2024] [Revised: 09/05/2024] [Accepted: 09/08/2024] [Indexed: 03/14/2025] Open
Abstract
Mpox is a zoonotic disease caused by the monkeypox virus (MPXV). Diagnosing and treating the disease has become a global health concern requiring close attention to its spread to non-endemic regions. Clinical diagnosis is based on laboratory test results. Conventional detection techniques include real-time quantitative polymerase chain reaction (qPCR), genome sequencing, antigen and antibody identification, and virus isolation. Nevertheless, these methods fall short of rapidly and efficiently identifying MPXV, as they require specialized training, specific laboratory environments, and professional-grade equipment. Emerging technologies offer complementary advantages to meet diverse diagnostic needs, including various point-of-care testing (POCT) approaches and integrating biosensors with rapid detection techniques. This review discusses prospective future research avenues for MPXV detection, examining the advances and challenges of various detection techniques which may contribute to the ongoing elimination of mpox human-to-human transmission and serves as a reference for developing effective prevention and control strategies.
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Affiliation(s)
- Wenjing Liu
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
| | - Erxin Zhang
- School of Pharmacy, China Pharmaceutical University, Nanjing 211198, China
| | - Wei Li
- School of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Nanjing 211198, China
| | - Ruichen Lv
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
| | - Yanfeng Lin
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
| | - Yingjia Xu
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
| | - Jiameng Li
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
| | - Yuzhen Lai
- School of Life Science and Technology, China Pharmaceutical University, Nanjing 211198, China
| | - Yuxin Jiang
- School of Public Health, Nanjing Medical University, Nanjing 211100, China
| | - Sijia Lin
- School of Pharmacy, China Pharmaceutical University, Nanjing 211198, China
| | - Xueqin Wang
- School of Life Science and Technology, China Pharmaceutical University, Nanjing 211198, China
| | - Peize Zhou
- School of Engineering, China Pharmaceutical University, Nanjing 211198, China
| | - Yue Song
- School of Public Health, Nanjing Medical University, Nanjing 211100, China
| | - Wanpeng Shen
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
| | - Yiqian Sun
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
| | - Yuexi Li
- Nanjing Bioengineering (Gene) Technology Center for Medicines, Nanjing 210002, China
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Quarton S, Livesey A, Jeff C, Hatton C, Scott A, Parekh D, Thickett D, McNally A, Sapey E. Metagenomics in the Diagnosis of Pneumonia: Protocol for a Systematic Review. JMIR Res Protoc 2024; 13:e57334. [PMID: 39293053 PMCID: PMC11447427 DOI: 10.2196/57334] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/14/2024] [Revised: 07/16/2024] [Accepted: 07/18/2024] [Indexed: 09/20/2024] Open
Abstract
BACKGROUND Causative pathogens are currently identified in only a minority of pneumonia cases, which affects antimicrobial stewardship. Metagenomic next-generation sequencing (mNGS) has potential to enhance pathogen detection due to its sensitivity and broad applicability. However, while studies have shown improved sensitivity compared with conventional microbiological methods for pneumonia diagnosis, it remains unclear whether this can translate into clinical benefit. Most existing studies focus on patients who are ventilated, readily allowing for analysis of bronchoalveolar lavage fluid (BALF). The impact of sample type on the use of metagenomic analysis remains poorly defined. Similarly, previous studies rarely differentiate between the types of pneumonia involved-community-acquired pneumonia (CAP), hospital-acquired pneumonia (HAP), or ventilator-associated pneumonia (VAP)-which have different clinical profiles. OBJECTIVE This study aims to determine the clinical use of mNGS in CAP, HAP, and VAP, compared with traditional microbiological methods. METHODS We aim to review all studies (excluding case reports of a series of fewer than 10 people) of adult patients with suspected or confirmed pneumonia that compare metagenomic analysis with traditional microbiology techniques, including culture, antigen-based testing, and polymerase chain reaction-based assays. Relevant studies will be identified through systematic searches of the Embase, MEDLINE, Scopus, and Cochrane CENTRAL databases. Screening of titles, abstracts, and subsequent review of eligible full texts will be done by 2 separate reviewers (SQ and 1 of AL, CJ, or CH), with a third clinician (ES) providing adjudication in case of disagreement. Our focus is on the clinical use of metagenomics for patients with CAP, HAP, and VAP. Data extracted will focus on clinically important outcomes-pathogen positivity rate, laboratory turnaround time, impact on clinical decision-making, length of stay, and 30-day mortality. Subgroup analyses will be performed based on the type of pneumonia (CAP, HAP, or VAP) and sample type used. The risk of bias will be assessed using the QUADAS-2 tool for diagnostic accuracy studies. Outcome data will be combined in a random-effects meta-analysis, and where this is not possible, a narrative synthesis will be undertaken. RESULTS The searches were completed with the assistance of a medical librarian on January 13, 2024, returning 5750 records. Screening and data extraction are anticipated to be completed by September 2024. CONCLUSIONS Despite significant promise, the impact of metagenomic analysis on clinical pathways remains unclear. Furthermore, it is unclear whether the use of this technique will alter depending on whether the pneumonia is a CAP, HAP, or VAP or the sample type that is collected. This systematic review will assess the current evidence base to support the benefit of clinical outcomes for metagenomic analysis, depending on the setting of pneumonia diagnosis or specimen type used. It will identify areas where further research is needed to advance this methodology into routine care. TRIAL REGISTRATION PROSPERO CRD42023488096; https://tinyurl.com/3suy7cma. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID) DERR1-10.2196/57334.
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Affiliation(s)
- Samuel Quarton
- National Institute for Health Research Birmingham Biomedical Research Centre, Birmingham, United Kingdom
- Institute of Inflammation and Ageing, University of Birmingham, Birmingham, United Kingdom
| | - Alana Livesey
- National Institute for Health Research Welcome Trust Clinical Research Facility, University Hospitals Birmingham, Birmingham, United Kingdom
| | - Charlotte Jeff
- National Institute for Health Research Birmingham Biomedical Research Centre, Birmingham, United Kingdom
| | - Christopher Hatton
- National Institute for Health Research Midlands Patient Safety Research Collaboration, University of Birmingham, Birmingham, United Kingdom
| | - Aaron Scott
- National Institute for Health Research Birmingham Biomedical Research Centre, Birmingham, United Kingdom
| | - Dhruv Parekh
- National Institute for Health Research Birmingham Biomedical Research Centre, Birmingham, United Kingdom
| | - David Thickett
- Institute of Inflammation and Ageing, University of Birmingham, Birmingham, United Kingdom
| | - Alan McNally
- National Institute for Health Research Birmingham Biomedical Research Centre, Birmingham, United Kingdom
| | - Elizabeth Sapey
- National Institute for Health Research Birmingham Biomedical Research Centre, Birmingham, United Kingdom
- National Institute for Health Research Midlands Patient Safety Research Collaboration, University of Birmingham, Birmingham, United Kingdom
- National Institute for Health Research Midlands Applied Research Collaborative, University of Birmingham, Birmingham, United Kingdom
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Quarton S, Livesey A, Pittaway H, Adiga A, Grudzinska F, McNally A, Dosanjh D, Sapey E, Parekh D. Clinical challenge of diagnosing non-ventilator hospital-acquired pneumonia and identifying causative pathogens: a narrative review. J Hosp Infect 2024; 149:189-200. [PMID: 38621512 DOI: 10.1016/j.jhin.2024.02.029] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2023] [Revised: 02/12/2024] [Accepted: 02/21/2024] [Indexed: 04/17/2024]
Abstract
Non-ventilated hospital-acquired pneumonia (NV-HAP) is associated with a significant healthcare burden, arising from high incidence and associated morbidity and mortality. However, accurate identification of cases remains challenging. At present, there is no gold-standard test for the diagnosis of NV-HAP, requiring instead the blending of non-specific signs and investigations. Causative organisms are only identified in a minority of cases. This has significant implications for surveillance, patient outcomes and antimicrobial stewardship. Much of the existing research in HAP has been conducted among ventilated patients. The paucity of dedicated NV-HAP research means that conclusions regarding diagnostic methods, pathology and interventions must largely be extrapolated from work in other settings. Progress is also limited by the lack of a widely agreed definition for NV-HAP. The diagnosis of NV-HAP has large scope for improvement. Consensus regarding a case definition will allow meaningful research to improve understanding of its aetiology and the heterogeneity of outcomes experienced by patients. There is potential to optimize the role of imaging and to incorporate novel techniques to identify likely causative pathogens. This would facilitate both antimicrobial stewardship and surveillance of an important healthcare-associated infection. This narrative review considers the utility of existing methods to diagnose NV-HAP, with a focus on the significance and challenge of identifying pathogens. It discusses the limitations in current techniques, and explores the potential of emergent molecular techniques to improve microbiological diagnosis and outcomes for patients.
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Affiliation(s)
- S Quarton
- National Institute for Health Research Birmingham Biomedical Research Centre, University of Birmingham, Birmingham, UK.
| | - A Livesey
- National Institute for Health Research/Wellcome Trust Clinical Research Facility, University Hospitals Birmingham, Birmingham, UK
| | - H Pittaway
- Queen Elizabeth Hospital Birmingham, University Hospitals Birmingham, Birmingham, UK
| | - A Adiga
- Warwick Hospital, South Warwickshire University NHS Foundation Trust, Warwick, UK
| | - F Grudzinska
- Institute of Inflammation and Ageing, University of Birmingham, Birmingham, UK
| | - A McNally
- Institute of Microbiology and Infection, University of Birmingham, Birmingham, UK
| | - D Dosanjh
- Institute of Inflammation and Ageing, University of Birmingham, Birmingham, UK
| | - E Sapey
- National Institute for Health Research Birmingham Biomedical Research Centre, University of Birmingham, Birmingham, UK; National Institute for Health Research Midlands Patient Safety Research Collaboration, University of Birmingham, Birmingham, UK; National Institute for Health Research Midlands Applied Research Collaborative, University of Birmingham, Birmingham, UK
| | - D Parekh
- Institute of Inflammation and Ageing, University of Birmingham, Birmingham, UK
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Cao Y, Wang B, Wang Y, Wang Y, Huai W, Bao X, Jin M, Jin Y, Jin Y, Zhang Z, Shan J. Construction of a postoperative infection outbreak investigation form: A tool for early detection and control measures. Am J Infect Control 2024; 52:588-594. [PMID: 38142776 DOI: 10.1016/j.ajic.2023.12.011] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/29/2023] [Revised: 12/18/2023] [Accepted: 12/18/2023] [Indexed: 12/26/2023]
Abstract
BACKGROUND To develop an investigation form for postoperative infection outbreak (PIO), and to identify sources of the outbreak in the early stage. METHODS After an exhaustive literature review, we used the Delphi method to determine the indicators and relative risk scores of the assessment tools through 2 rounds of specialist consultation and overall consideration of the opinions and suggestions of 20 specialists. RESULTS A total of 203 studies of PIO were eligible for inclusion. The mean authority coefficient (Cr) was 0.87. Kendall's W coefficient of the specialist consultation was 0.704 after 2 rounds of consultation (P < .005), suggesting that the specialists had similar opinions. Based on 4 primary items and 19 secondary items of the source of PIO, and tripartite distribution characteristics of infected patients, we constructed the PIO investigation form. CONCLUSIONS The PIO investigation form can be used in the investigation of the early-stage cluster of cases, it's a prerequisite for taking effective control measures, avoiding PIO occurrence. However, the effect of the investigation form needs to be further evaluated.
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Affiliation(s)
- Yulong Cao
- Department of Hospital-Acquired Infection Control, Peking University People's Hospital, Beijing, China
| | - Bin Wang
- Department of Neurosurgery, Peking University People's Hospital, Beijing, China
| | - Yanbin Wang
- Department of Obstetrics and Gynecology, Peking University People's Hospital, Beijing, China
| | - Yan Wang
- Department of Nephrology, Peking University People's Hospital, Beijing, China
| | - Wei Huai
- Department of Emergency, Peking University Third Hospital, Beijing, China
| | - Xiaoyuan Bao
- Medical Information Center, Peking University Health Science Center, Beijing, China
| | - Meng Jin
- Medical Information Center, Peking University Health Science Center, Beijing, China
| | - Yicheng Jin
- School of General Studies, Columbia University, New York, USA
| | - Yixi Jin
- Khoury College of Computer Science, Northeastern University, Seattle, USA
| | - Zexin Zhang
- Graduate School of Medicine Faculty of Medicine, Kyoto University, Kyoto, Japan
| | - Jiao Shan
- Department of Hospital-Acquired Infection Control, Beijing Jishuitan Hospital, Capital Medical University, Beijing, China.
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Wong SC, Yip CCY, Chen JHK, Yuen LLH, AuYeung CHY, Chan WM, Chu AWH, Leung RCY, Ip JD, So SYC, Yuen KY, To KKW, Cheng VCC. Investigation of air dispersal during a rhinovirus outbreak in a pediatric intensive care unit. Am J Infect Control 2024; 52:472-478. [PMID: 37972820 DOI: 10.1016/j.ajic.2023.11.003] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2023] [Revised: 11/03/2023] [Accepted: 11/04/2023] [Indexed: 11/19/2023]
Abstract
BACKGROUND While airborne transmission of rhinovirus is recognized in indoor settings, its role in hospital transmission remains unclear. METHODS We investigated an outbreak of rhinovirus in a pediatric intensive care unit (PICU) to assess air dispersal. We collected clinical, environmental, and air samples, and staff's surgical masks for viral load and phylogenetic analysis. Hand hygiene compliance and the number of air changes per hour in the PICU were measured. A case-control analysis was performed to identify nosocomial rhinovirus risk factors. RESULTS Between March 31, 2023, and April 2, 2023, three patients acquired rhinovirus in a cubicle (air changes per hour: 14) of 12-bed PICU. A portable air-cleaning unit was placed promptly. Air samples (72,000 L in 6 hours) from the cohort area, and outer surfaces of staff's masks (n = 8), were rhinovirus RNA-negative. Hand hygiene compliance showed no significant differences (31/34, 91.2% vs 33/37, 89.2%, P = 1) before and during outbreak. Only 1 environmental sample (3.8%) was positive (1.86 × 103 copies/mL). Case-control and next-generation sequencing analysis implicated an infected staff member as the source. CONCLUSIONS Our findings suggest that air dispersal of rhinovirus was not documented in the well-ventilated PICU during the outbreak. Further research is needed to better understand the dynamics of rhinovirus transmission in health care settings.
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Affiliation(s)
- Shuk-Ching Wong
- Infection Control Team, Queen Mary Hospital, Hong Kong West Cluster, Hong Kong Special Administrative Region, China
| | - Cyril C-Y Yip
- Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China
| | - Jonathan H-K Chen
- Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China
| | - Lithia L-H Yuen
- Infection Control Team, Queen Mary Hospital, Hong Kong West Cluster, Hong Kong Special Administrative Region, China
| | - Christine H-Y AuYeung
- Infection Control Team, Queen Mary Hospital, Hong Kong West Cluster, Hong Kong Special Administrative Region, China
| | - Wan-Mui Chan
- State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China
| | - Allen W-H Chu
- State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China
| | - Rhoda C-Y Leung
- State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China
| | - Jonathan D Ip
- State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China
| | - Simon Y-C So
- Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China
| | - Kwok-Yung Yuen
- State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; Centre for Virology, Vaccinology and Therapeutics, Hong Kong Science and Technology Park, Hong Kong Special Administrative Region, China; Department of Clinical Microbiology and Infection Control, The University of Hong Kong-Shenzhen Hospital, Shenzhen, China
| | - Kelvin K-W To
- State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; Centre for Virology, Vaccinology and Therapeutics, Hong Kong Science and Technology Park, Hong Kong Special Administrative Region, China; Department of Clinical Microbiology and Infection Control, The University of Hong Kong-Shenzhen Hospital, Shenzhen, China.
| | - Vincent C-C Cheng
- Infection Control Team, Queen Mary Hospital, Hong Kong West Cluster, Hong Kong Special Administrative Region, China; Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China.
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Mwakibete L, Greening SS, Kalantar K, Ahyong V, Anis E, Miller EA, Needle DB, Oglesbee M, Thomas WK, Sevigny JL, Gordon LM, Nemeth NM, Ogbunugafor CB, Ayala AJ, Faith SA, Neff N, Detweiler AM, Baillargeon T, Tanguay S, Simpson SD, Murphy LA, Ellis JC, Tato CM, Gagne RB. Metagenomics for Pathogen Detection During a Mass Mortality Event in Songbirds. J Wildl Dis 2024; 60:362-374. [PMID: 38345467 DOI: 10.7589/jwd-d-23-00109] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2023] [Accepted: 01/02/2024] [Indexed: 04/06/2024]
Abstract
Mass mortality events in wildlife can be indications of an emerging infectious disease. During the spring and summer of 2021, hundreds of dead passerines were reported across the eastern US. Birds exhibited a range of clinical signs including swollen conjunctiva, ocular discharge, ataxia, and nystagmus. As part of the diagnostic investigation, high-throughput metagenomic next-generation sequencing was performed across three molecular laboratories on samples from affected birds. Many potentially pathogenic microbes were detected, with bacteria forming the largest proportion; however, no singular agent was consistently identified, with many of the detected microbes also found in unaffected (control) birds and thus considered to be subclinical infections. Congruent results across laboratories have helped drive further investigation into alternative causes, including environmental contaminants and nutritional deficiencies. This work highlights the utility of metagenomic approaches in investigations of emerging diseases and provides a framework for future wildlife mortality events.
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Affiliation(s)
| | - Sabrina S Greening
- Department of Pathobiology, Wildlife Futures Program, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
| | | | - Vida Ahyong
- Chan Zuckerberg Biohub, San Francisco, California 94158, USA
| | - Eman Anis
- Department of Pathobiology, Wildlife Futures Program, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
- Department of Pathobiology, PADLS New Bolton Center, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
| | - Erica A Miller
- Department of Pathobiology, Wildlife Futures Program, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
| | - David B Needle
- New Hampshire Veterinary Diagnostic Lab, University of New Hampshire, Durham, New Hampshire 03824, USA
| | - Michael Oglesbee
- Infectious Diseases Institute, The Ohio State University, Columbus, Ohio 43210, USA
| | - W Kelley Thomas
- Hubbard Center for Genome Studies, University of New Hampshire, Durham, New Hampshire 03824, USA
| | - Joseph L Sevigny
- Hubbard Center for Genome Studies, University of New Hampshire, Durham, New Hampshire 03824, USA
| | - Lawrence M Gordon
- Hubbard Center for Genome Studies, University of New Hampshire, Durham, New Hampshire 03824, USA
| | - Nicole M Nemeth
- Southeastern Cooperative Wildlife Disease Study and Department of Pathology, College of Veterinary Medicine, University of Georgia, Athens, Georgia 30602, USA
- Department of Pathology, College of Veterinary Medicine, University of Georgia, Georgia 30602, USA
| | - C Brandon Ogbunugafor
- Department of Ecology and Evolutionary Biology, Yale University, New Haven, Connecticut 06511, USA
| | - Andrea J Ayala
- Department of Ecology and Evolutionary Biology, Yale University, New Haven, Connecticut 06511, USA
| | - Seth A Faith
- Infectious Diseases Institute, The Ohio State University, Columbus, Ohio 43210, USA
| | - Norma Neff
- Chan Zuckerberg Biohub, San Francisco, California 94158, USA
| | | | - Tessa Baillargeon
- New Hampshire Veterinary Diagnostic Lab, University of New Hampshire, Durham, New Hampshire 03824, USA
| | - Stacy Tanguay
- New Hampshire Veterinary Diagnostic Lab, University of New Hampshire, Durham, New Hampshire 03824, USA
| | - Stephen D Simpson
- Hubbard Center for Genome Studies, University of New Hampshire, Durham, New Hampshire 03824, USA
| | - Lisa A Murphy
- Department of Pathobiology, Wildlife Futures Program, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
- Department of Pathobiology, PADLS New Bolton Center, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
| | - Julie C Ellis
- Department of Pathobiology, Wildlife Futures Program, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
| | - Cristina M Tato
- Chan Zuckerberg Biohub, San Francisco, California 94158, USA
| | - Roderick B Gagne
- Department of Pathobiology, Wildlife Futures Program, University of Pennsylvania School of Veterinary Medicine, New Bolton Center, Kennett Square, Pennsylvania 19348, USA
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9
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Craney A, Miller S. Present and Future Non-Culture-Based Diagnostics: Stewardship Potentials and Considerations. Clin Lab Med 2024; 44:109-122. [PMID: 38280793 DOI: 10.1016/j.cll.2023.10.003] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/29/2024]
Abstract
The medical microbiologist plays a key role in the transition from culture-based to molecular test methods for diagnosis of infectious diseases. They must understand the scientific and technical bases underlying these tests along with their associated benefits and limitations and be able to educate administrators and patient providers on their proper use. Coordination of testing practices between clinical departments and the spectrum of public health and research laboratories is essential to optimize health care delivery.
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Affiliation(s)
- Arryn Craney
- Center for Infectious Disease Diagnostics and Research, Diagnostic Medicine Institute, Geisinger Health System, 100 North Academy Avenue, Danville, PA 17822, USA
| | - Steve Miller
- Delve Bio, Inc. and Department of Laboratory Medicine, University of California San Francisco, 953 Indiana Street, San Francisco, CA 94107, USA.
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10
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Makhsous N, Goya S, Avendaño CC, Rupp J, Kuypers J, Jerome KR, Boeckh M, Waghmare A, Greninger AL. Within-Host Rhinovirus Evolution in Upper and Lower Respiratory Tract Highlights Capsid Variability and Mutation-Independent Compartmentalization. J Infect Dis 2024; 229:403-412. [PMID: 37486790 PMCID: PMC10873175 DOI: 10.1093/infdis/jiad284] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2023] [Revised: 07/13/2023] [Accepted: 07/17/2023] [Indexed: 07/26/2023] Open
Abstract
BACKGROUND Rhinovirus (RV) infections can progress from the upper (URT) to lower (LRT) respiratory tract in immunocompromised individuals, causing high rates of fatal pneumonia. Little is known about how RV evolves within hosts during infection. METHODS We sequenced RV complete genomes from 12 hematopoietic cell transplant patients with infection for up to 190 days from both URT (nasal wash, NW) and LRT (bronchoalveolar lavage, BAL). Metagenomic and amplicon next-generation sequencing were used to track the emergence and evolution of intrahost single nucleotide variants (iSNVs). RESULTS Identical RV intrahost populations in matched NW and BAL specimens indicated no genetic adaptation is required for RV to progress from URT to LRT. Coding iSNVs were 2.3-fold more prevalent in capsid over nonstructural genes. iSNVs modeled were significantly more likely to be found in capsid surface residues, but were not preferentially located in known RV-neutralizing antibody epitopes. Newly emergent, genotype-matched iSNV haplotypes from immunocompromised individuals in 2008-2010 could be detected in Seattle-area community RV sequences in 2020-2021. CONCLUSIONS RV infections in immunocompromised hosts can progress from URT to LRT with no specific evolutionary requirement. Capsid proteins carry the highest variability and emergent mutations can be detected in other, including future, RV sequences.
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Affiliation(s)
- Negar Makhsous
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, USA
| | - Stephanie Goya
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, USA
| | - Carlos C Avendaño
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, USA
| | - Jason Rupp
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, USA
| | - Jane Kuypers
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, USA
| | - Keith R Jerome
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA
| | - Michael Boeckh
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA
- Department of Medicine, University of Washington, Seattle, USA
| | - Alpana Waghmare
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA
- Department of Pediatrics, University of Washington, Seattle, USA
| | - Alexander L Greninger
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA
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11
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Makhsous N, Goya S, Avendaño C, Rupp J, Kuypers J, Jerome KR, Boeckh M, Waghmare A, Greninger AL. Within-host rhinovirus evolution in upper and lower respiratory tract highlights capsid variability and mutation-independent compartmentalization. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.05.11.540440. [PMID: 37214809 PMCID: PMC10197658 DOI: 10.1101/2023.05.11.540440] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/24/2023]
Abstract
Background Human rhinovirus (HRV) infections can progress from the upper (URT) to lower (LRT) respiratory tract in immunocompromised individuals, causing high rates of fatal pneumonia. Little is known about how HRV evolves within hosts during infection. Methods We sequenced HRV complete genomes from 12 hematopoietic cell transplant patients with prolonged infection for up to 190 days from both URT (nasal wash, NW) and LRT (bronchoalveolar lavage, BAL) specimens. Metagenomic (mNGS) and amplicon-based NGS were used to study the emergence and evolution of intra-host single nucleotide variants (iSNVs). Results Identical HRV intra-host populations in matched NW and BAL specimens indicated no genetic adaptation is required for HRV to progress from URT to LRT. Microbial composition between matched NW and BAL confirmed no cross-contamination during sampling procedure. Coding iSNVs were 2.3-fold more prevalent in capsid over non-structural genes, adjusted for length. iSNVs modeled onto HRV capsid structures were significantly more likely to be found in surface residues, but were not preferentially located in known HRV neutralizing antibody epitopes. Newly emergent, serotype-matched iSNV haplotypes from immunocompromised individuals from 2008-2010 could be detected in Seattle-area community HRV sequences from 2020-2021. Conclusion HRV infections in immunocompromised hosts can progress from URT to LRT with no specific evolutionary requirement. Capsid proteins carry the highest variability and emergent mutations can be detected in other, including future, HRV sequences.
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Affiliation(s)
- Negar Makhsous
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, 98102, USA
| | - Stephanie Goya
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, 98102, USA
| | - Carlos Avendaño
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, 98102, USA
| | - Jason Rupp
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, 98102, USA
| | - Jane Kuypers
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, 98102, USA
| | - Keith R. Jerome
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, 98102, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, 98109, USA
| | - Michael Boeckh
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, 98109, USA
- Department of Medicine, University of Washington, Seattle, 98102, USA
| | - Alpana Waghmare
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, 98109, USA
- Department of Pediatrics, University of Washington, Seattle, 98105, USA
| | - Alexander L Greninger
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, 98102, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, 98109, USA
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12
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Saiman L, Prill MM, Wilmont S, Neu N, Alba L, Hill-Ricciuti A, Larson E, Whitaker B, Lu X, Garg S, Gerber SI, Kim L. Surveillance for Acute Respiratory Illnesses in Pediatric Chronic Care Facilities. J Pediatric Infect Dis Soc 2023; 12:49-52. [PMID: 36219180 PMCID: PMC11262614 DOI: 10.1093/jpids/piac109] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/30/2022] [Accepted: 10/10/2022] [Indexed: 11/13/2022]
Abstract
Overall, 119 (33%) of 364 pediatric chronic care facility residents experienced 182 acute respiratory illnesses (ARIs) that met the surveillance definition which led to 31 (17%) emergency room visits, 34 (19%) acute care hospitalizations, and/or 25 (14%) ICU admissions. Continued PCR-positivity was observed in 35% of ARIs during follow-up testing.
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Affiliation(s)
- Lisa Saiman
- Department of Pediatrics, Columbia University Irving Medical Center, New York, New York, USA
- Department of Infection Prevention & Control, New York-Presbyterian Hospital, New York, New York, USA
| | - Mila M. Prill
- Division of Viral Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Sibyl Wilmont
- Department of Pediatrics, Columbia University Irving Medical Center, New York, New York, USA
| | - Natalie Neu
- Department of Pediatrics, Columbia University Irving Medical Center, New York, New York, USA
| | - Luis Alba
- Department of Pediatrics, Columbia University Irving Medical Center, New York, New York, USA
| | - Alexandra Hill-Ricciuti
- Department of Pediatrics, Columbia University Irving Medical Center, New York, New York, USA
| | - Elaine Larson
- Columbia University School of Nursing, New York, New York, USA
| | - Brett Whitaker
- Division of Viral Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Xiaoyan Lu
- Division of Viral Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Shikha Garg
- Influenza Division, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
- United States Public Health Service, Rockville, Maryland, USA
| | - Susan I. Gerber
- Division of Viral Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Lindsay Kim
- Division of Viral Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
- United States Public Health Service, Rockville, Maryland, USA
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13
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Wong TY, Horspool AM, Russ BP, Ye C, Lee KS, Winters MT, Bevere JR, Miller OA, Rader NA, Cooper M, Kieffer T, Sourimant J, Greninger AL, Plemper RK, Denvir J, Cyphert HA, Barbier M, Torrelles JB, Martinez I, Martinez-Sobrido L, Damron FH. Evaluating Antibody Mediated Protection against Alpha, Beta, and Delta SARS-CoV-2 Variants of Concern in K18-hACE2 Transgenic Mice. J Virol 2022; 96:e0218421. [PMID: 35080423 PMCID: PMC8941865 DOI: 10.1128/jvi.02184-21] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2021] [Accepted: 01/02/2022] [Indexed: 12/04/2022] Open
Abstract
SARS-CoV-2 variants of concern (VoC) are impacting responses to the COVID-19 pandemic. Here, we utilized passive immunization using human convalescent plasma (HCP) obtained from a critically ill COVID-19 patient in the early pandemic to study the efficacy of polyclonal antibodies generated to ancestral SARS-CoV-2 against the Alpha, Beta, and Delta VoC in the K18 human angiotensin converting enzyme 2 (hACE2) transgenic mouse model. HCP protected mice from challenge with the original WA-1 SARS-CoV-2 strain; however, only partially protected mice challenged with the Alpha VoC (60% survival) and failed to save Beta challenged mice from succumbing to disease. HCP treatment groups had elevated receptor binding domain (RBD) and nucleocapsid IgG titers in the serum; however, Beta VoC viral RNA burden in the lung and brain was not decreased due to HCP treatment. While mice could be protected from WA-1 or Alpha challenge with a single dose of HCP, six doses of HCP could not decrease mortality of Delta challenged mice. Overall, these data demonstrate that VoC have enhanced immune evasion and this work underscores the need for in vivo models to evaluate future emerging strains. IMPORTANCE Emerging SARS-CoV-2 VoC are posing new problems regarding vaccine and monoclonal antibody efficacy. To better understand immune evasion tactics of the VoC, we utilized passive immunization to study the effect of early-pandemic SARS-CoV-2 HCP against, Alpha, Beta, and Delta VoC. We observed that HCP from a human infected with the original SARS-CoV-2 was unable to control lethality of Alpha, Beta, or Delta VoC in the K18-hACE2 transgenic mouse model of SARS-CoV-2 infection. Our findings demonstrate that passive immunization can be used as a model to evaluate immune evasion of emerging VoC strains.
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Affiliation(s)
- Ting Y. Wong
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Alexander M. Horspool
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Brynnan P. Russ
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Chengjin Ye
- Host-Pathogen Interactions and Population Health Programs, Texas Biomedical Research Institute, San Antonio, Texas, USA
| | - Katherine S. Lee
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Michael T. Winters
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
| | - Justin R. Bevere
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Olivia A. Miller
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Nathaniel A. Rader
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Melissa Cooper
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Theodore Kieffer
- Department of Pathology, Anatomy and Laboratory Medicine, West Virginia University School of Medicine, Morgantown, West Virginia, USA
| | - Julien Sourimant
- Institute for Biomedical Sciences, Georgia State University, Atlanta, Georgia, USA
| | - Alexander L. Greninger
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, Washington, USA
| | - Richard K. Plemper
- Institute for Biomedical Sciences, Georgia State University, Atlanta, Georgia, USA
| | - James Denvir
- Department of Biomedical Sciences, Marshall University, Huntington, West Virginia, USA
| | - Holly A. Cyphert
- Department of Biological Sciences, Marshall University, Huntington, West Virginia, USA
| | - Mariette Barbier
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
| | - Jordi B. Torrelles
- Host-Pathogen Interactions and Population Health Programs, Texas Biomedical Research Institute, San Antonio, Texas, USA
| | - Ivan Martinez
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- West Virginia University Cancer Institute, Morgantown, West Virginia, USA
| | - Luis Martinez-Sobrido
- Host-Pathogen Interactions and Population Health Programs, Texas Biomedical Research Institute, San Antonio, Texas, USA
| | - F. Heath Damron
- Department of Microbiology, Immunology, and Cell Biology, West Virginia University, Morgantown, West Virginia, USA
- Vaccine Development Center at West, Virginia University Health Sciences Center, Morgantown, West Virginia, USA
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14
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Zhan L, Huang K, Xia W, Chen J, Wang L, Lu J, Wang J, Lin J, Wu W. The Diagnosis of Severe Fever with Thrombocytopenia Syndrome Using Metagenomic Next-Generation Sequencing: Case Report and Literature Review. Infect Drug Resist 2022; 15:83-89. [PMID: 35046673 PMCID: PMC8760998 DOI: 10.2147/idr.s345991] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2021] [Accepted: 12/21/2021] [Indexed: 12/15/2022] Open
Abstract
Background Severe fever with thrombocytopenia syndrome (SFTS) is an infectious disease caused by a bunyaviridae virus. Its main clinical manifestation is fever with thrombocytopenia, which may be accompanied by other clinical symptoms. Here, we report a patient diagnosed with SFTS using metagenomic next‑generation sequencing (mNGS). Case Presentation A 56-year-old female patient was hospitalized with intermittent diarrhea and fever. She visited a local clinic for treatment, but instead of improving, the symptoms progressed to unconsciousness. Diagnosis Using mNGS, we isolated the bunyaviridae virus and several other pathogens from the patient’s blood samples to confirm the diagnosis. Interventions The patient was treated with symptomatic and supportive therapy, including intravenous human γ-globulin (20 g/d), platelet transfusion, platelet elevation (subcutaneous injection of recombinant human thrombopoietin, 15,000 IU), white blood cell elevation (subcutaneous injection of recombinant human granulocyte colony-stimulating factor, 200 ug, qd); and antibiotic (cefoperazone sodium and tazobactam sodium, 2 g, q8h), antiviral (ganciclovir, 250 mg, q12h), and antifungal therapy (voriconazole for injection, 0.2 g, q12h). After ten days of treatment, the patient’s condition gradually improved. Conclusion Compared to traditional detection methods, mNGS has many advantages. It can quickly identify the pathogen when the patient’s clinical manifestations are complex and difficult to diagnose, resulting in the formulation of an effective treatment.
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Affiliation(s)
- Liying Zhan
- Department of Critical Care Medicine, Renmin Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
| | - Kai Huang
- Department of Gastroenterology, Zhongnan Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
| | - Wenfang Xia
- Department of Critical Care Medicine, Renmin Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
| | - Jingdi Chen
- Department of Orthopedics, The Airborne Military Hospital, Wuhan, Hubei, People’s Republic of China
| | - Lu Wang
- Department of Critical Care Medicine, Renmin Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
| | - Jiaming Lu
- Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
| | - Jing Wang
- Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
| | - Jun Lin
- Department of Gastroenterology, Zhongnan Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
- Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
| | - Wei Wu
- Department of Critical Care Medicine, Renmin Hospital of Wuhan University, Wuhan, Hubei, People’s Republic of China
- Correspondence: Wei Wu; Jun Lin Email ;
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15
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Greninger AL, Zerr DM. NGSocomial Infections: High-Resolution Views of Hospital-Acquired Infections Through Genomic Epidemiology. J Pediatric Infect Dis Soc 2021; 10:S88-S95. [PMID: 34951469 PMCID: PMC8755322 DOI: 10.1093/jpids/piab074] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/05/2023]
Abstract
Hospital outbreak investigations are high-stakes epidemiology. Contacts between staff and patients are numerous; environmental and community exposures are plentiful; and patients are highly vulnerable. Having the best data is paramount to understanding an outbreak in order to stop ongoing transmission and prevent future outbreaks. In the past 5 years, the high-resolution view of transmission offered by analyzing pathogen whole-genome sequencing (WGS) is increasingly part of hospital outbreak investigations. Concerns over speed and actionability, assay validation, liability, cost, and payment models lead to further opportunities for work in this area. Now accelerated by funding for COVID-19, the use of genomics in hospital outbreak investigations has firmly moved from the academic literature to more quotidian operations, with associated concerns involving regulatory affairs, data integration, and clinical interpretation. This review details past uses of WGS data in hospital-acquired infection outbreaks as well as future opportunities to increase its utility and growth in hospital infection prevention.
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Affiliation(s)
- Alexander L Greninger
- Department of Laboratory Medicine and Pathology, University of Washington Medical Center, Seattle, Washington, USA,Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA,Corresponding Author: Alexander L. Greninger MD, PhD, MS, MPhil, 1616 Eastlake Ave East Suite 320, Seattle, WA 98102, USA. E-mail:
| | - Danielle M Zerr
- Department of Pediatrics, University of Washington Medical Center, Seattle, Washington, USA,Division of Infectious Diseases, Seattle Children’s Hospital, Seattle, Washington, USA
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16
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Schuele L, Cassidy H, Peker N, Rossen JWA, Couto N. Future potential of metagenomics in clinical laboratories. Expert Rev Mol Diagn 2021; 21:1273-1285. [PMID: 34755585 DOI: 10.1080/14737159.2021.2001329] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/19/2022]
Abstract
INTRODUCTION Rapid and sensitive diagnostic strategies are necessary for patient care and public health. Most of the current conventional microbiological assays detect only a restricted panel of pathogens at a time or require a microbe to be successfully cultured from a sample. Clinical metagenomics next-generation sequencing (mNGS) has the potential to unbiasedly detect all pathogens in a sample, increasing the sensitivity for detection and enabling the discovery of unknown infectious agents. AREAS COVERED High expectations have been built around mNGS; however, this technique is far from widely available. This review highlights the advances and currently available options in terms of costs, turnaround time, sensitivity, specificity, validation, and reproducibility of mNGS as a diagnostic tool in clinical microbiology laboratories. EXPERT OPINION The need for a novel diagnostic tool to increase the sensitivity of microbial diagnostics is clear. mNGS has the potential to revolutionise clinical microbiology. However, its role as a diagnostic tool has yet to be widely established, which is crucial for successfully implementing the technique. A clear definition of diagnostic algorithms that include mNGS is vital to show clinical utility. Similarly to real-time PCR, mNGS will one day become a vital tool in any testing algorithm.
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Affiliation(s)
- Leonard Schuele
- University of Groningen, University Medical Center Groningen, Department of Medical Microbiology and Infection Prevention, Groningen, the Netherlands
| | - Hayley Cassidy
- University of Groningen, University Medical Center Groningen, Department of Medical Microbiology and Infection Prevention, Groningen, the Netherlands
| | - Nilay Peker
- University of Groningen, University Medical Center Groningen, Department of Medical Microbiology and Infection Prevention, Groningen, the Netherlands
| | - John W A Rossen
- University of Groningen, University Medical Center Groningen, Department of Medical Microbiology and Infection Prevention, Groningen, the Netherlands.,Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Natacha Couto
- University of Groningen, University Medical Center Groningen, Department of Medical Microbiology and Infection Prevention, Groningen, the Netherlands.,The Milner Centre for Evolution, Department of Biology and Biochemistry, University of Bath, Bath, United Kingdom
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17
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Horspool AM, Ye C, Wong TY, Russ BP, Lee KS, Winters MT, Bevere JR, Kieffer T, Martinez I, Sourimant J, Greninger A, Plemper RK, Denvir J, Cyphert HA, Torrelles J, Martinez-Sobrido L, Damron FH. SARS-CoV-2 B.1.1.7 and B.1.351 variants of concern induce lethal disease in K18-hACE2 transgenic mice despite convalescent plasma therapy. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2021:2021.05.05.442784. [PMID: 33972945 PMCID: PMC8109207 DOI: 10.1101/2021.05.05.442784] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
SARS-CoV-2 variants of concern (VoCs) are impacting responses to the COVID-19 pandemic. Here we present a comparison of the SARS-CoV-2 USA-WA1/2020 (WA-1) strain with B.1.1.7 and B.1.351 VoCs and identify significant differences in viral propagation in vitro and pathogenicity in vivo using K18-hACE2 transgenic mice. Passive immunization with plasma from an early pandemic SARS-CoV-2 patient resulted in significant differences in the outcome of VoC-infected mice. WA-1-infected mice were protected by plasma, B.1.1.7-infected mice were partially protected, and B.1.351-infected mice were not protected. Serological correlates of disease were different between VoC-infected mice, with B.1.351 triggering significantly altered cytokine profiles than other strains. In this study, we defined infectivity and immune responses triggered by VoCs and observed that early 2020 SARS-CoV-2 human immune plasma was insufficient to protect against challenge with B.1.1.7 and B.1.351 in the mouse model.
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18
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Gil P, Dupuy V, Koual R, Exbrayat A, Loire E, Fall AG, Gimonneau G, Biteye B, Talla Seck M, Rakotoarivony I, Marie A, Frances B, Lambert G, Reveillaud J, Balenghien T, Garros C, Albina E, Eloit M, Gutierrez S. A library preparation optimized for metagenomics of RNA viruses. Mol Ecol Resour 2021; 21:1788-1807. [PMID: 33713395 DOI: 10.1111/1755-0998.13378] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2019] [Revised: 02/23/2021] [Accepted: 02/25/2021] [Indexed: 11/28/2022]
Abstract
Our understanding of the viral communities associated to animals has not yet reached the level attained on the bacteriome. This situation is due to, among others, technical challenges in adapting metagenomics using high-throughput sequencing to the study of RNA viromes in animals. Although important developments have been achieved in most steps of viral metagenomics, there is yet a key step that has received little attention: the library preparation. This situation differs from bacteriome studies in which developments in library preparation have largely contributed to the democratisation of metagenomics. Here, we present a library preparation optimized for metagenomics of RNA viruses from insect vectors of viral diseases. The library design allows a simple PCR-based preparation, such as those routinely used in bacterial metabarcoding, that is adapted to shotgun sequencing as required in viral metagenomics. We first optimized our library preparation using mock viral communities and then validated a full metagenomic approach incorporating our preparation in two pilot studies with field-caught insect vectors; one including a comparison with a published metagenomic protocol. Our approach provided a fold increase in virus-like sequences compared to other studies, and nearly-full genomes from new virus species. Moreover, our results suggested conserved trends in virome composition within a population of a mosquito species. Finally, the sensitivity of our approach was compared to a commercial diagnostic PCR for the detection of an arbovirus in field-caught insect vectors. Our approach could facilitate studies on viral communities from animals and the democratization of metagenomics in community ecology of viruses.
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Affiliation(s)
- Patricia Gil
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Virginie Dupuy
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Rachid Koual
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Antoni Exbrayat
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Etienne Loire
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Assane G Fall
- Laboratoire National de l'Elevage et de Recherches Vétérinaires, Institut Sénégalais de Recherches Agricoles (ISRA), Dakar-Hann, Senegal
| | - Geoffrey Gimonneau
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France.,Laboratoire National de l'Elevage et de Recherches Vétérinaires, Institut Sénégalais de Recherches Agricoles (ISRA), Dakar-Hann, Senegal
| | - Biram Biteye
- Laboratoire National de l'Elevage et de Recherches Vétérinaires, Institut Sénégalais de Recherches Agricoles (ISRA), Dakar-Hann, Senegal
| | - Momar Talla Seck
- Laboratoire National de l'Elevage et de Recherches Vétérinaires, Institut Sénégalais de Recherches Agricoles (ISRA), Dakar-Hann, Senegal
| | - Ignace Rakotoarivony
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | | | | | | | - Julie Reveillaud
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France
| | - Thomas Balenghien
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Claire Garros
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Emmanuel Albina
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
| | - Marc Eloit
- Pathogen Discovery Laboratory, Institut Pasteur, Paris, France.,The OIE Collaborating Centre for Detection and Identification in Humans of Emerging Animal Pathogens, Institut Pasteur, Paris, France.,École nationale vétérinaire d'Alfort, Maisons-Alfort, France
| | - Serafin Gutierrez
- ASTRE, Cirad, INRAE, University of Montpellier, Montpellier, France.,Cirad, UMR ASTRE, Montpellier, F-34398, France
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19
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Nakamichi K, Shen JZ, Lee CS, Lee A, Roberts EA, Simonson PD, Roychoudhury P, Andriesen J, Randhawa AK, Mathias PC, Greninger AL, Jerome KR, Van Gelder RN. Hospitalization and mortality associated with SARS-CoV-2 viral clades in COVID-19. Sci Rep 2021; 11:4802. [PMID: 33637820 PMCID: PMC7910290 DOI: 10.1038/s41598-021-82850-9] [Citation(s) in RCA: 43] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2020] [Accepted: 01/20/2021] [Indexed: 02/07/2023] Open
Abstract
The COVID-19 epidemic of 2019-20 is due to the novel coronavirus SARS-CoV-2. Following first case description in December, 2019 this virus has infected over 10 million individuals and resulted in at least 500,000 deaths world-wide. The virus is undergoing rapid mutation, with two major clades of sequence variants emerging. This study sought to determine whether SARS-CoV-2 sequence variants are associated with differing outcomes among COVID-19 patients in a single medical system. Whole genome SARS-CoV-2 RNA sequence was obtained from isolates collected from patients registered in the University of Washington Medicine health system between March 1 and April 15, 2020. Demographic and baseline clinical characteristics of patients and their outcome data including their hospitalization and death were collected. Statistical and machine learning models were applied to determine if viral genetic variants were associated with specific outcomes of hospitalization or death. Full length SARS-CoV-2 sequence was obtained 190 subjects with clinical outcome data. 35 (18.4%) were hospitalized and 14 (7.4%) died from complications of infection. A total of 289 single nucleotide variants were identified. Clustering methods demonstrated two major viral clades, which could be readily distinguished by 12 polymorphisms in 5 genes. A trend toward higher rates of hospitalization of patients with Clade 2 infections was observed (p = 0.06, Fisher's exact). Machine learning models utilizing patient demographics and co-morbidities achieved area-under-the-curve (AUC) values of 0.93 for predicting hospitalization. Addition of viral clade or sequence information did not significantly improve models for outcome prediction. In summary, SARS-CoV-2 shows substantial sequence diversity in a community-based sample. Two dominant clades of virus are in circulation. Among patients sufficiently ill to warrant testing for virus, no significant difference in outcomes of hospitalization or death could be discerned between clades in this sample. Major risk factors for hospitalization and death for either major clade of virus include patient age and comorbid conditions.
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Affiliation(s)
- Kenji Nakamichi
- Department of Ophthalmology, University of Washington School of Medicine, 325 9th Avenue, Campus Box 359608, Seattle, WA, 98104, USA
| | - Jolie Z Shen
- University of Washington School of Medicine, Seattle, WA, USA
| | - Cecilia S Lee
- Department of Ophthalmology, University of Washington School of Medicine, 325 9th Avenue, Campus Box 359608, Seattle, WA, 98104, USA
| | - Aaron Lee
- Department of Ophthalmology, University of Washington School of Medicine, 325 9th Avenue, Campus Box 359608, Seattle, WA, 98104, USA
| | - Emma A Roberts
- University of Washington School of Medicine, Seattle, WA, USA
| | - Paul D Simonson
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA, USA
| | - Pavitra Roychoudhury
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA, USA
| | - Jessica Andriesen
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA
| | - April K Randhawa
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA
| | - Patrick C Mathias
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA, USA
| | - Alex L Greninger
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA
| | - Keith R Jerome
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA
| | - Russell N Van Gelder
- Department of Ophthalmology, University of Washington School of Medicine, 325 9th Avenue, Campus Box 359608, Seattle, WA, 98104, USA.
- Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA, USA.
- Department of Biological Structure, University of Washington, Seattle, WA, USA.
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20
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Li N, Cai Q, Miao Q, Song Z, Fang Y, Hu B. High-Throughput Metagenomics for Identification of Pathogens in the Clinical Settings. SMALL METHODS 2021; 5:2000792. [PMID: 33614906 PMCID: PMC7883231 DOI: 10.1002/smtd.202000792] [Citation(s) in RCA: 147] [Impact Index Per Article: 36.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/31/2020] [Revised: 10/24/2020] [Indexed: 05/25/2023]
Abstract
The application of sequencing technology is shifting from research to clinical laboratories owing to rapid technological developments and substantially reduced costs. However, although thousands of microorganisms are known to infect humans, identification of the etiological agents for many diseases remains challenging as only a small proportion of pathogens are identifiable by the current diagnostic methods. These challenges are compounded by the emergence of new pathogens. Hence, metagenomic next-generation sequencing (mNGS), an agnostic, unbiased, and comprehensive method for detection, and taxonomic characterization of microorganisms, has become an attractive strategy. Although many studies, and cases reports, have confirmed the success of mNGS in improving the diagnosis, treatment, and tracking of infectious diseases, several hurdles must still be overcome. It is, therefore, imperative that practitioners and clinicians understand both the benefits and limitations of mNGS when applying it to clinical practice. Interestingly, the emerging third-generation sequencing technologies may partially offset the disadvantages of mNGS. In this review, mainly: a) the history of sequencing technology; b) various NGS technologies, common platforms, and workflows for clinical applications; c) the application of NGS in pathogen identification; d) the global expert consensus on NGS-related methods in clinical applications; and e) challenges associated with diagnostic metagenomics are described.
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Affiliation(s)
- Na Li
- Department of Infectious DiseasesZhongshan HospitalFudan UniversityShanghai200032China
| | - Qingqing Cai
- Genoxor Medical Science and Technology Inc.Zhejiang317317China
| | - Qing Miao
- Department of Infectious DiseasesZhongshan HospitalFudan UniversityShanghai200032China
| | - Zeshi Song
- Genoxor Medical Science and Technology Inc.Zhejiang317317China
| | - Yuan Fang
- Genoxor Medical Science and Technology Inc.Zhejiang317317China
| | - Bijie Hu
- Department of Infectious DiseasesZhongshan HospitalFudan UniversityShanghai200032China
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21
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Nakamichi K, Shen JZ, Lee CS, Lee AY, Roberts EA, Simonson PD, Roychoudhury P, Andriesen JG, Randhawa AK, Mathias PC, Greninger A, Jerome KR, Van Gelder RN. Outcomes associated with SARS-CoV-2 viral clades in COVID-19. MEDRXIV : THE PREPRINT SERVER FOR HEALTH SCIENCES 2020:2020.09.24.20201228. [PMID: 32995827 PMCID: PMC7523168 DOI: 10.1101/2020.09.24.20201228] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 04/16/2023]
Abstract
Background The COVID-19 epidemic of 2019-20 is due to the novel coronavirus SARS-CoV-2. Following first case description in December, 2019 this virus has infected over 10 million individuals and resulted in at least 500,000 deaths world-wide. The virus is undergoing rapid mutation, with two major clades of sequence variants emerging. This study sought to determine whether SARS-CoV-2 sequence variants are associated with differing outcomes among COVID-19 patients in a single medical system. Methods Whole genome SARS-CoV-2 RNA sequence was obtained from isolates collected from patients registered in the University of Washington Medicine health system between March 1 and April 15, 2020. Demographic and baseline medical data along with outcomes of hospitalization and death were collected. Statistical and machine learning models were applied to determine if viral genetic variants were associated with specific outcomes of hospitalization or death. Findings Full length SARS-CoV-2 sequence was obtained 190 subjects with clinical outcome data. 35 (18.4%) were hospitalized and 14 (7.4%) died from complications of infection. A total of 289 single nucleotide variants were identified. Clustering methods demonstrated two major viral clades, which could be readily distinguished by 12 polymorphisms in 5 genes. A trend toward higher rates of hospitalization of patients with Clade 2 was observed (p=0.06). Machine learning models utilizing patient demographics and co-morbidities achieved area-under-the-curve (AUC) values of 0.93 for predicting hospitalization. Addition of viral clade or sequence information did not significantly improve models for outcome prediction. Conclusion SARS-CoV-2 shows substantial sequence diversity in a community-based sample. Two dominant clades of virus are in circulation. Among patients sufficiently ill to warrant testing for virus, no significant difference in outcomes of hospitalization or death could be discerned between clades in this sample. Major risk factors for hospitalization and death for either major clade of virus include patient age and comorbid conditions.
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22
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Beyond personal protective equipment: adjunctive methods for control of healthcare-associated respiratory viral infections. Curr Opin Infect Dis 2020; 33:312-318. [PMID: 32657968 DOI: 10.1097/qco.0000000000000655] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
PURPOSE OF REVIEW Prevention of nosocomial transmission of respiratory viruses is a priority in all healthcare settings and often achieved with the use of personal protective equipment. Several adjunctive infection prevention methods are in common use but their effectiveness in reducing healthcare-associated respiratory viral infections is unclear. In this review, recent advances regarding the effectiveness of several adjunctive infection prevention methods to reduce healthcare-associated respiratory viral infections are discussed. RECENT FINDINGS Training and education on hand hygiene guidelines, mandatory influenza vaccination for healthcare personnel, access to paid sick leave to reduce ill presenteeism, cohorting of patients with the same infection or clinical syndrome, neuraminidase inhibitor chemoprophylaxis during influenza outbreaks, and enhanced visitor restrictions in pediatric hospitals all have shown some degree of effectiveness in observational or quasi-experimental studies. SUMMARY Most of the studies evaluating the effect of adjunctive infection prevention methods on healthcare-associated respiratory viral infections are observational or quasi-experimental and are often combined with other interventions. Therefore, it is difficult to determine the precise effectiveness or efficacy of these interventions and more controlled trials are needed. Multimodal infection prevention policies are likely to be most effective in reducing healthcare-associated respiratory viral infections.
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23
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Ciccozzi M, Lai A, Zehender G, Borsetti A, Cella E, Ciotti M, Sagnelli E, Sagnelli C, Angeletti S. The phylogenetic approach for viral infectious disease evolution and epidemiology: An updating review. J Med Virol 2019; 91:1707-1724. [PMID: 31243773 DOI: 10.1002/jmv.25526] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2019] [Accepted: 06/24/2019] [Indexed: 12/16/2022]
Abstract
In the last decade, the phylogenetic approach is recurrent in molecular evolutionary analysis. On 12 May, 2019, about 2 296 213 papers are found, but typing "phylogeny" or "epidemiology AND phylogeny" only 199 804 and 20 133 are retrieved, respectively. Molecular epidemiology in infectious diseases is widely used to define the source of infection as so as the ancestral relationships of individuals sampled from a population. Coalescent theory and phylogeographic analysis have had scientific application in several, recent pandemic events, and nosocomial outbreaks. Hepatitis viruses and immunodeficiency virus (human immunodeficiency virus) have been largely studied. Phylogenetic analysis has been recently applied on Polyomaviruses so as in the more recent outbreaks due to different arboviruses type as Zika and chikungunya viruses discovering the source of infection and the geographic spread. Data on sequences isolated by the microorganism are essential to apply the phylogenetic tools and research in the field of infectious disease phylodinamics is growing up. There is the need to apply molecular phylogenetic and evolutionary methods in areas out of infectious diseases, as translational genomics and personalized medicine. Lastly, the application of these tools in vaccine strategy so as in antibiotic and antiviral researchers are encouraged.
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Affiliation(s)
- Massimo Ciccozzi
- Unit of Medical Statistics and Molecular Epidemiology, University Campus Bio-Medico of Rome, Rome, Italy
| | - Alessia Lai
- Department of Biomedical and Clinical Sciences 'L. Sacco', University of Milan, Milan, Italy
| | - Gianguglielmo Zehender
- Department of Biomedical and Clinical Sciences 'L. Sacco', University of Milan, Milan, Italy
| | - Alessandra Borsetti
- National HIV/AIDS Research Center, Istituto Superiore di Sanità, Roma, Italy
| | - Eleonora Cella
- Unit of Medical Statistics and Molecular Epidemiology, University Campus Bio-Medico of Rome, Rome, Italy
| | - Marco Ciotti
- Laboratory of Molecular Virology, Polyclinic Tor Vergata Foundation, Rome, Italy
| | - Evangelista Sagnelli
- Department of Mental Health and Public Medicine, Section of Infectious Diseases, University of Campania Luigi Vanvitelli, Naples, Italy
| | - Caterina Sagnelli
- Department of Mental Health and Public Medicine, Section of Infectious Diseases, University of Campania Luigi Vanvitelli, Naples, Italy
| | - Silvia Angeletti
- Unit of Clinical Laboratory Science, University Campus Bio-Medico of Rome, Rome, Italy
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24
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Kroneman A, de Sousa R, Verhoef L, Koopmans MPG, Vennema H, On Behalf Of The HAVNet Network. Usability of the international HAVNet hepatitis A virus database for geographical annotation, backtracing and outbreak detection. ACTA ACUST UNITED AC 2019; 23. [PMID: 30229723 PMCID: PMC6144472 DOI: 10.2807/1560-7917.es.2018.23.37.1700802] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023]
Abstract
HAVNet is an international laboratory network sharing sequences and corresponding metadata on hepatitis A virus in an online database. Aim: We give an overview of the epidemiological and genetic data and assess the usability of the present dataset for geographical annotation, backtracing and outbreak detection. Methods: A descriptive analysis was performed on the timeliness, completeness, epidemiological data and geographic coverage of the dataset. Length and genomic region of the sequences were reviewed as well as the numerical and geographical distribution of the genotypes. The geographical signal in the sequences was assessed based on a short common nt stretch using a 100% identity analysis. Results: The 9,211 reports were heterogeneous for completeness and timeliness, and for length and genomic region of the sequences. Some parts of the world were not represented by the sequences. Geographical differences in prevalence of HAV genotypes described previously could be confirmed with this dataset and for a third (1,075/3,124) of the included sequences, 100% identity of the short common sequence coincided with an identical country of origin. Conclusion: Analysis of a subset of short, shared sequences indicates that a geographical annotation on the level of individual countries is possible with the HAVNet data. If the current incompleteness and heterogeneity of the data can be improved on, HAVNet could become very useful as a worldwide reference set for geographical annotation and for backtracing and outbreak detection.
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Affiliation(s)
- Annelies Kroneman
- National Institute for Public Health and the Environment (RIVM), Bilthoven, the Netherlands
| | - Rita de Sousa
- National Institute of Health Dr. Ricardo Jorge, Lisbon, Portugal
| | - Linda Verhoef
- Office for Risk Assessment and Research, Netherlands Food and Consumer Product Safety Authority (NVWA), Utrecht, the Netherlands (current affiliation).,National Institute for Public Health and the Environment (RIVM), Bilthoven, the Netherlands
| | | | - Harry Vennema
- National Institute for Public Health and the Environment (RIVM), Bilthoven, the Netherlands
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25
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Casto AM, Adler AL, Makhsous N, Crawford K, Qin X, Kuypers JM, Huang ML, Zerr DM, Greninger AL. Prospective, Real-time Metagenomic Sequencing During Norovirus Outbreak Reveals Discrete Transmission Clusters. Clin Infect Dis 2019; 69:941-948. [PMID: 30576430 PMCID: PMC6735836 DOI: 10.1093/cid/ciy1020] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2018] [Accepted: 11/29/2018] [Indexed: 12/11/2022] Open
Abstract
BACKGROUND Norovirus outbreaks in hospital settings are a common challenge for infection prevention teams. Given the high burden of norovirus in most communities, it can be difficult to distinguish between ongoing in-hospital transmission of the virus and new introductions from the community, and it is challenging to understand the long-term impacts of outbreak-associated viruses within medical systems using traditional epidemiological approaches alone. METHODS Real-time metagenomic sequencing during an ongoing norovirus outbreak associated with a retrospective cohort study. RESULTS We describe a hospital-associated norovirus outbreak that affected 13 patients over a 27-day period in a large, tertiary, pediatric hospital. The outbreak was chronologically associated with a spike in self-reported gastrointestinal symptoms among staff. Real-time metagenomic next-generation sequencing (mNGS) of norovirus genomes demonstrated that 10 chronologically overlapping, hospital-acquired norovirus cases were partitioned into 3 discrete transmission clusters. Sequencing data also revealed close genetic relationships between some hospital-acquired and some community-acquired cases. Finally, this data was used to demonstrate chronic viral shedding by an immunocompromised, hospital-acquired case patient. An analysis of serial samples from this patient provided novel insights into the evolution of norovirus within an immunocompromised host. CONCLUSIONS This study documents one of the first applications of real-time mNGS during a hospital-associated viral outbreak. Given its demonstrated ability to detect transmission patterns within outbreaks and elucidate the long-term impacts of outbreak-associated viral strains on patients and medical systems, mNGS constitutes a powerful resource to help infection control teams understand, prevent, and respond to viral outbreaks.
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Affiliation(s)
- Amanda M Casto
- Department of Medicine, University of Washington, Seattle
| | - Amanda L Adler
- Seattle Children’s Hospital, University of Washington, Seattle
| | - Negar Makhsous
- Department of Laboratory Medicine, University of Washington, Seattle
| | | | - Xuan Qin
- Department of Medicine, University of Washington, Seattle
| | - Jane M Kuypers
- Department of Laboratory Medicine, University of Washington, Seattle
| | - Meei-Li Huang
- Department of Laboratory Medicine, University of Washington, Seattle
| | - Danielle M Zerr
- Seattle Children’s Hospital, University of Washington, Seattle
- Department of Pediatrics, University of Washington, Seattle
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26
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Molecular and epidemiologic investigation of a rhinovirus outbreak in a neonatal intensive care unit. Infect Control Hosp Epidemiol 2018; 40:245-247. [PMID: 30516128 DOI: 10.1017/ice.2018.311] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022]
Abstract
We performed a molecular and epidemiologic study of a healthcare-associated rhinovirus outbreak to better understand transmission in neonatal intensive care settings. Sequencing of the 7 outbreak strains revealed 4 distinct clades, indicating multiple sources. A single clade infected 3 patients in adjacent rooms, suggesting horizontal transmission. We observed 1 rhinovirus-associated death.
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27
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Greninger AL, Naccache SN. Metagenomics to Assist in the Diagnosis of Bloodstream Infection. J Appl Lab Med 2018; 3:643-653. [PMID: 31639732 DOI: 10.1373/jalm.2018.026120] [Citation(s) in RCA: 30] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2018] [Accepted: 11/08/2018] [Indexed: 12/13/2022]
Abstract
BACKGROUND Metagenomic next-generation sequencing (mNGS) has emerged as a promising technology that enables pan-pathogen detection from any source. However, clinical utility and practical integration into the clinical microbiology work flow and a bloodstream infection detection algorithm are currently uncharted. In the context of bloodstream infections, the challenges associated with blood culture, including sensitivity, postantibiotic treatment, attaining sufficient volumes sufficient volumes, and turnaround time, are well-known. Molecular assays have helped expedite turnaround time, especially when performed directly from positive culture media bottles. mNGS offers an unbiased but more complex version of molecular testing directly from sample, but it is unclear how and if it should be implemented in the clinical microbiology laboratory today. CONTENT Here we map out the potential utility and application of mNGS tests to infectious disease diagnostics from blood sources, including intrinsic limitations of the methodology in diagnosing bloodstream infections and sepsis vs DNAemia, current barriers to integration into routine workup, and milestones that may need to be met before implementation. SUMMARY Polymerases and pores move faster than bugs divide, so the thermodynamics of mNGS adoption for bloodstream infection is favorable. Nonetheless, considerable activation barriers exist that will slow this likely diagnostic transition. We eagerly await the manufacturer who designs an integrated sample-to-answer box to do for mNGS what has been done for other aspects of molecular detection.
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Affiliation(s)
| | - Samia N Naccache
- Department of Laboratory Medicine, Children's Hospital Los Angeles, Los Angeles, CA
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28
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Das S, Dunbar S, Tang YW. Laboratory Diagnosis of Respiratory Tract Infections in Children - the State of the Art. Front Microbiol 2018; 9:2478. [PMID: 30405553 PMCID: PMC6200861 DOI: 10.3389/fmicb.2018.02478] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/18/2018] [Accepted: 09/28/2018] [Indexed: 12/13/2022] Open
Abstract
In the pediatric population, respiratory infections are the most common cause of physician visits. Although many respiratory illnesses are self-limiting viral infections that resolve with time and supportive care, it can be critical to identify the causative pathogen at an early stage of the disease in order to implement effective antimicrobial therapy and infection control. Over the last few years, diagnostics for respiratory infections have evolved substantially, with the development of novel assays and the availability of updated tests for newer strains of pathogens. Newer laboratory methods are rapid, highly sensitive and specific, and are gradually replacing the conventional gold standards, although the clinical utility of these assays is still under evaluation. This article reviews the current laboratory methods available for testing for respiratory pathogens and discusses the advantages and disadvantages of each approach.
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Affiliation(s)
- Shubhagata Das
- Global Scientific Affairs, Luminex Corporation, Austin, TX, United States
| | - Sherry Dunbar
- Global Scientific Affairs, Luminex Corporation, Austin, TX, United States
| | - Yi-Wei Tang
- Department of Laboratory Medicine, Memorial Sloan Kettering Cancer Center, Weill Medical College of Cornell University, New York, NY, United States.,Department of Pathology and Laboratory Medicine, Memorial Sloan Kettering Cancer Center, Weill Medical College of Cornell University, New York, NY, United States
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29
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Ogimi C, Xie H, Leisenring WM, Kuypers JM, Jerome KR, Campbell AP, Englund JA, Boeckh M, Waghmare A. Initial High Viral Load Is Associated with Prolonged Shedding of Human Rhinovirus in Allogeneic Hematopoietic Cell Transplant Recipients. Biol Blood Marrow Transplant 2018; 24:2160-2163. [PMID: 30009982 PMCID: PMC6239940 DOI: 10.1016/j.bbmt.2018.07.006] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2018] [Accepted: 07/05/2018] [Indexed: 02/05/2023]
Abstract
We examined prolonged shedding of rhinovirus after stem cell transplantation. The median shedding duration of rhinovirus was similar between species. Initial high viral load was a risk factor for prolonged shedding of rhinovirus. Recent data suggest human rhinovirus (HRV) is associated with lower respiratory tract infection and mortality in hematopoietic cell transplant (HCT) recipients. Examining risk factors for prolonged viral shedding may provide critical insight for the development of novel therapeutics and help inform infection prevention practices. Our objective was to identify risk factors for prolonged shedding of HRV post-HCT. We prospectively collected weekly nasal samples from allogeneic HCT recipients from day 0 to day 100 post-transplant, and performed real-time reverse transcriptase PCR (December 2005 to February 2010). Subjects with symptomatic HRV infection and a negative test within 2 weeks of the last positive were included. Duration of shedding was defined as time between the first positive and first negative samples. Cycle threshold (Ct) values were used as a proxy for viral load. HRV species were identified by sequencing the 5′ noncoding region. Logistic regression analyses were performed to evaluate factors associated with prolonged shedding (≥21 days). We identified 38 HCT recipients with HRV infection fulfilling study criteria (32 adults, 6 children). Median duration of shedding was 9.5 days (range, 2 to 89 days); 18 patients had prolonged shedding. Among 26 samples sequenced, 69% were species A, and species B and C accounted for 15% each; the median shedding duration of HRV did not differ among species (P = .17). Bivariable logistic regression analyses suggest that initial high viral load (low Ct value) is associated with prolonged shedding. HCT recipients with initial high viral loads are at risk for prolonged HRV viral shedding.
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Affiliation(s)
- Chikara Ogimi
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington; Department of Pediatrics, University of Washington, Seattle, Washington; Pediatric Infectious Diseases Division, Seattle Children's Hospital, Seattle, Washington
| | - Hu Xie
- Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington
| | - Wendy M Leisenring
- Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington
| | - Jane M Kuypers
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington; Department of Laboratory Medicine, University of Washington, Seattle, Washington
| | - Keith R Jerome
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington; Department of Laboratory Medicine, University of Washington, Seattle, Washington
| | - Angela P Campbell
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington; Department of Pediatrics, University of Washington, Seattle, Washington; Pediatric Infectious Diseases Division, Seattle Children's Hospital, Seattle, Washington
| | - Janet A Englund
- Department of Pediatrics, University of Washington, Seattle, Washington; Pediatric Infectious Diseases Division, Seattle Children's Hospital, Seattle, Washington
| | - Michael Boeckh
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington; Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington; Department of Medicine, University of Washington, Seattle, Washington
| | - Alpana Waghmare
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, Washington; Department of Pediatrics, University of Washington, Seattle, Washington; Pediatric Infectious Diseases Division, Seattle Children's Hospital, Seattle, Washington
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Viral Entry Properties Required for Fitness in Humans Are Lost through Rapid Genomic Change during Viral Isolation. mBio 2018; 9:mBio.00898-18. [PMID: 29970463 PMCID: PMC6030562 DOI: 10.1128/mbio.00898-18] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
Human parainfluenza viruses cause a large burden of human respiratory illness. While much research relies upon viruses grown in cultured immortalized cells, human parainfluenza virus 3 (HPIV-3) evolves in culture. Cultured viruses differ in their properties compared to clinical strains. We present a genome-wide survey of HPIV-3 adaptations to culture using metagenomic next-generation sequencing of matched pairs of clinical samples and primary culture isolates (zero passage virus). Nonsynonymous changes arose during primary viral isolation, almost entirely in the genes encoding the two surface glycoproteins-the receptor binding protein hemagglutinin-neuraminidase (HN) or the fusion protein (F). We recovered genomes from 95 HPIV-3 primary culture isolates and 23 HPIV-3 strains directly from clinical samples. HN mutations arising during primary viral isolation resulted in substitutions at HN's dimerization/F-interaction site, a site critical for activation of viral fusion. Alterations in HN dimer interface residues known to favor infection in culture occurred within 4 days (H552 and N556). A novel cluster of residues at a different face of the HN dimer interface emerged (P241 and R242) and imply a role in HPIV-3-mediated fusion. Functional characterization of these culture-associated HN mutations in a clinical isolate background revealed acquisition of the fusogenic phenotype associated with cultured HPIV-3; the HN-F complex showed enhanced fusion and decreased receptor-cleaving activity. These results utilize a method for identifying genome-wide changes associated with brief adaptation to culture to highlight the notion that even brief exposure to immortalized cells may affect key viral properties and underscore the balance of features of the HN-F complex required for fitness by circulating viruses.IMPORTANCE Human parainfluenza virus 3 is an important cause of morbidity and mortality among infants, the immunocompromised, and the elderly. Using deep genomic sequencing of HPIV-3-positive clinical material and its subsequent viral isolate, we discover a number of known and novel coding mutations in the main HPIV-3 attachment protein HN during brief exposure to immortalized cells. These mutations significantly alter function of the fusion complex, increasing fusion promotion by HN as well as generally decreasing neuraminidase activity and increasing HN-receptor engagement. These results show that viruses may evolve rapidly in culture even during primary isolation of the virus and before the first passage and reveal features of fitness for humans that are obscured by rapid adaptation to laboratory conditions.
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Goya S, Valinotto LE, Tittarelli E, Rojo GL, Nabaes Jodar MS, Greninger AL, Zaiat JJ, Marti MA, Mistchenko AS, Viegas M. An optimized methodology for whole genome sequencing of RNA respiratory viruses from nasopharyngeal aspirates. PLoS One 2018; 13:e0199714. [PMID: 29940028 PMCID: PMC6016902 DOI: 10.1371/journal.pone.0199714] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2018] [Accepted: 06/12/2018] [Indexed: 11/25/2022] Open
Abstract
Over the last decade, the number of viral genome sequences deposited in available databases has grown exponentially. However, sequencing methodology vary widely and many published works have relied on viral enrichment by viral culture or nucleic acid amplification with specific primers rather than through unbiased techniques such as metagenomics. The genome of RNA viruses is highly variable and these enrichment methodologies may be difficult to achieve or may bias the results. In order to obtain genomic sequences of human respiratory syncytial virus (HRSV) from positive nasopharyngeal aspirates diverse methodologies were evaluated and compared. A total of 29 nearly complete and complete viral genomes were obtained. The best performance was achieved with a DNase I treatment to the RNA directly extracted from the nasopharyngeal aspirate (NPA), sequence-independent single-primer amplification (SISPA) and library preparation performed with Nextera XT DNA Library Prep Kit with manual normalization. An average of 633,789 and 1,674,845 filtered reads per library were obtained with MiSeq and NextSeq 500 platforms, respectively. The higher output of NextSeq 500 was accompanied by the increasing of duplicated reads percentage generated during SISPA (from an average of 1.5% duplicated viral reads in MiSeq to an average of 74% in NextSeq 500). HRSV genome recovery was not affected by the presence or absence of duplicated reads but the computational demand during the analysis was increased. Considering that only samples with viral load ≥ E+06 copies/ml NPA were tested, no correlation between sample viral loads and number of total filtered reads was observed, nor with the mapped viral reads. The HRSV genomes showed a mean coverage of 98.46% with the best methodology. In addition, genomes of human metapneumovirus (HMPV), human rhinovirus (HRV) and human parainfluenza virus types 1–3 (HPIV1-3) were also obtained with the selected optimal methodology.
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Affiliation(s)
- Stephanie Goya
- Ricardo Gutiérrez Children’s Hospital, Ciudad Autónoma Buenos Aires, Argentina
- Consejo Nacional de Investigaciones Científicas y Tecnológicas (CONICET), Buenos Aires, Argentina
| | - Laura E. Valinotto
- Ricardo Gutiérrez Children’s Hospital, Ciudad Autónoma Buenos Aires, Argentina
- Consejo Nacional de Investigaciones Científicas y Tecnológicas (CONICET), Buenos Aires, Argentina
| | - Estefania Tittarelli
- Ricardo Gutiérrez Children’s Hospital, Ciudad Autónoma Buenos Aires, Argentina
- Consejo Nacional de Investigaciones Científicas y Tecnológicas (CONICET), Buenos Aires, Argentina
| | - Gabriel L. Rojo
- Ricardo Gutiérrez Children’s Hospital, Ciudad Autónoma Buenos Aires, Argentina
| | - Mercedes S. Nabaes Jodar
- Ricardo Gutiérrez Children’s Hospital, Ciudad Autónoma Buenos Aires, Argentina
- Ministerio de Salud de la Ciudad de Buenos Aires, Buenos Aires, Argentina
| | - Alexander L. Greninger
- Department of Laboratory Medicine, University of Washington, Seattle, Washington, United States of America
| | - Jonathan J. Zaiat
- Consejo Nacional de Investigaciones Científicas y Tecnológicas (CONICET), Buenos Aires, Argentina
- Argentine Bioinformatic Platform (BIA), Buenos Aires, Argentina
| | - Marcelo A. Marti
- Consejo Nacional de Investigaciones Científicas y Tecnológicas (CONICET), Buenos Aires, Argentina
- Argentine Bioinformatic Platform (BIA), Buenos Aires, Argentina
| | - Alicia S. Mistchenko
- Ricardo Gutiérrez Children’s Hospital, Ciudad Autónoma Buenos Aires, Argentina
- Comisión de Investigaciones Científicas (CIC), Buenos Aires, Argentina
| | - Mariana Viegas
- Ricardo Gutiérrez Children’s Hospital, Ciudad Autónoma Buenos Aires, Argentina
- Consejo Nacional de Investigaciones Científicas y Tecnológicas (CONICET), Buenos Aires, Argentina
- * E-mail:
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Abstract
PURPOSE OF REVIEW With the advent of novel massively parallel sequencing technologies and bioinformatic processing capabilities, clinical applications of metagenomic studies are rapidly being integrated into medicine. Through this paper, we hope to introduce this powerful new tool to clinicians caring for children. RECENT FINDINGS Very few studies have looked at metagenomic applications in children. The ability to perform these types of massive sequencing projects was not possible as little as 7 years ago. SUMMARY Metagenomics is defined as the study of all genetic material within a given sample. Novel sequencing and analysis approaches allow for unbiased assays to identify pathogens missed by targeted sequencing and culture methods. Although not widely available yet, metagenomic studies have been used to diagnose pediatric infections, identify resistance genes in clinical samples, and characterize outbreaks. Although cost and turnaround time have limited its application in clinical laboratories to date, novel platforms and increasing comfort with these techniques continue to push diagnostic metagenomics into clinical pediatric medicine. Much work in this field is yet to be done. That being said, we feel that pediatric clinicians will be using metagenomic techniques in the care of children with increasing frequency in the near future.
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Greninger AL. A decade of RNA virus metagenomics is (not) enough. Virus Res 2018; 244:218-229. [PMID: 29055712 PMCID: PMC7114529 DOI: 10.1016/j.virusres.2017.10.014] [Citation(s) in RCA: 103] [Impact Index Per Article: 14.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2017] [Revised: 10/14/2017] [Accepted: 10/17/2017] [Indexed: 12/16/2022]
Abstract
It is hard to overemphasize the role that metagenomics has had on our recent understanding of RNA virus diversity. Metagenomics in the 21st century has brought with it an explosion in the number of RNA virus species, genera, and families far exceeding that following the discovery of the microscope in the 18th century for eukaryotic life or culture media in the 19th century for bacteriology or the 20th century for virology. When the definition of success in organism discovery is measured by sequence diversity and evolutionary distance, RNA viruses win. This review explores the history of RNA virus metagenomics, reasons for the successes so far in RNA virus metagenomics, and methodological concerns. In addition, the review briefly covers clinical metagenomics and environmental metagenomics and highlights some of the critical accomplishments that have defined the fast pace of RNA virus discoveries in recent years. Slightly more than a decade in, the field is exhausted from its discoveries but knows that there is yet even more out there to be found.
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Affiliation(s)
- Alexander L Greninger
- Virology Division, Department of Laboratory Medicine, University of Washington, Seattle, WA, United States; Fred Hutchinson Cancer Research Center, Seattle, WA, United States
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Shean RC, Greninger AL. Private collection: high correlation of sample collection and patient admission date in clinical microbiological testing complicates sharing of phylodynamic metadata. Virus Evol 2018; 4:vey005. [PMID: 29511571 PMCID: PMC5829646 DOI: 10.1093/ve/vey005] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Infectious pathogens are known for their rapid evolutionary rates with new mutations arising over days to weeks. The ability to rapidly recover whole genome sequences and analyze the spread and evolution of pathogens using genetic information and pathogen collection dates has lead to interest in real-time tracking of infectious transmission and outbreaks. However, the level of temporal resolution afforded by these analyses may conflict with definitions of what constitutes protected health information (PHI) and privacy requirements for de-identification for publication and public sharing of research data and metadata. In the United States, dates and locations associated with patient care that provide greater resolution than year or the first three digits of the zip code are generally considered patient identifiers. Admission and discharge dates are specifically named as identifiers in Department of Health and Human Services guidance. To understand the degree to which one can impute admission dates from specimen collection dates, we examined sample collection dates and patient admission dates associated with more than 270,000 unique microbiological results from the University of Washington Laboratory Medicine Department between 2010 and 2017. Across all positive microbiological tests, the sample collection date exactly matched the patient admission date in 68.8% of tests. Collection dates and admission dates were identical from emergency department and outpatient testing 86.7% and 96.5% of the time, respectively, with >99% of tests collected within 1 day from the patient admission date. Samples from female patients were significantly more likely to be collected closer to admission date that those from male patients. We show that PHI-associated dates such as admission date can confidently be imputed from deposited collection date. We suggest that publicly depositing microbiological collection dates at greater resolution than the year may not meet routine Safe Harbor-based requirements for patient de-identification. We recommend the use of Expert Determination to determine PHI for a given study and/or direct patient consent if clinical laboratories or phylodynamic practitioners desire to make these data available.
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Affiliation(s)
- Ryan C Shean
- Department of Laboratory Medicine, University of Washington, 1616 Eastlake Avenue East, Suite 320, Seattle, WA 98102, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, 1100 Eastlake Avenue East, Seattle, WA 98102, USA
| | - Alexander L Greninger
- Department of Laboratory Medicine, University of Washington, 1616 Eastlake Avenue East, Suite 320, Seattle, WA 98102, USA
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, 1100 Eastlake Avenue East, Seattle, WA 98102, USA
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