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Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19). Sequencing the viral genome as the outbreak progresses is important, particularly in the identification of emerging isolates with different pathogenic potential and to identify whether nucleotide changes in the genome will impair clinical diagnostic tools such as real-time PCR assays. Although single nucleotide polymorphisms and point mutations occur during the replication of coronaviruses, one of the biggest drivers in genetic change is recombination. This can manifest itself in insertions and/or deletions in the viral genome. Therefore, sequencing strategies that underpin molecular epidemiology and inform virus biology in patients should take these factors into account. A long amplicon/read length-based RT-PCR sequencing approach focused on the Oxford Nanopore MinION/GridION platforms was developed to identify and sequence the SARS-CoV-2 genome in samples from patients with or suspected of COVID-19. The protocol, termed Rapid Sequencing Long Amplicons (RSLAs) used random primers to generate cDNA from RNA purified from a sample from a patient, followed by single or multiplex PCRs to generate longer amplicons of the viral genome. The base protocol was used to identify SARS-CoV-2 in a variety of clinical samples and proved sensitive in identifying viral RNA in samples from patients that had been declared negative using other nucleic acid-based assays (false negative). Sequencing the amplicons revealed that a number of patients had a proportion of viral genomes with deletions.
Shona C. Moore; Rebekah Penrice-Randall; Muhannad Alruwaili; Nadine Randle; Stuart Armstrong; Catherine Hartley; Sam Haldenby; Xiaofeng Dong; Abdulrahman Alrezaihi; Mai Almsaud; Eleanor Bentley; Jordan Clark; Isabel García-Dorival; Paul Gilmore; Ximeng Han; Benjamin Jones; Lisa Luu; Parul Sharma; Ghada Shawli; Yani Sun; Qin Zhao; Steven T. Pullan; Daniel P. Carter; Kevin Bewley; Jake Dunning; En-Min Zhou; Tom Solomon; Michael Beadsworth; James Cruise; Derrick W. Crook; David A. Matthews; Andrew D. Davidson; Zana Mahmood; Waleed Aljabr; Julian Druce; Richard Vipond; Lisa Ng; Laurent Renia; Peter J. M. Openshaw; J. Kenneth Baillie; Miles W. Carroll; James Stewart; Alistair Darby; Malcolm Semple; Lance Turtle; Julian A. Hiscox. Amplicon-Based Detection and Sequencing of SARS-CoV-2 in Nasopharyngeal Swabs from Patients With COVID-19 and Identification of Deletions in the Viral Genome That Encode Proteins Involved in Interferon Antagonism. Viruses 2020, 12, 1164 .
AMA StyleShona C. Moore, Rebekah Penrice-Randall, Muhannad Alruwaili, Nadine Randle, Stuart Armstrong, Catherine Hartley, Sam Haldenby, Xiaofeng Dong, Abdulrahman Alrezaihi, Mai Almsaud, Eleanor Bentley, Jordan Clark, Isabel García-Dorival, Paul Gilmore, Ximeng Han, Benjamin Jones, Lisa Luu, Parul Sharma, Ghada Shawli, Yani Sun, Qin Zhao, Steven T. Pullan, Daniel P. Carter, Kevin Bewley, Jake Dunning, En-Min Zhou, Tom Solomon, Michael Beadsworth, James Cruise, Derrick W. Crook, David A. Matthews, Andrew D. Davidson, Zana Mahmood, Waleed Aljabr, Julian Druce, Richard Vipond, Lisa Ng, Laurent Renia, Peter J. M. Openshaw, J. Kenneth Baillie, Miles W. Carroll, James Stewart, Alistair Darby, Malcolm Semple, Lance Turtle, Julian A. Hiscox. Amplicon-Based Detection and Sequencing of SARS-CoV-2 in Nasopharyngeal Swabs from Patients With COVID-19 and Identification of Deletions in the Viral Genome That Encode Proteins Involved in Interferon Antagonism. Viruses. 2020; 12 (10):1164.
Chicago/Turabian StyleShona C. Moore; Rebekah Penrice-Randall; Muhannad Alruwaili; Nadine Randle; Stuart Armstrong; Catherine Hartley; Sam Haldenby; Xiaofeng Dong; Abdulrahman Alrezaihi; Mai Almsaud; Eleanor Bentley; Jordan Clark; Isabel García-Dorival; Paul Gilmore; Ximeng Han; Benjamin Jones; Lisa Luu; Parul Sharma; Ghada Shawli; Yani Sun; Qin Zhao; Steven T. Pullan; Daniel P. Carter; Kevin Bewley; Jake Dunning; En-Min Zhou; Tom Solomon; Michael Beadsworth; James Cruise; Derrick W. Crook; David A. Matthews; Andrew D. Davidson; Zana Mahmood; Waleed Aljabr; Julian Druce; Richard Vipond; Lisa Ng; Laurent Renia; Peter J. M. Openshaw; J. Kenneth Baillie; Miles W. Carroll; James Stewart; Alistair Darby; Malcolm Semple; Lance Turtle; Julian A. Hiscox. 2020. "Amplicon-Based Detection and Sequencing of SARS-CoV-2 in Nasopharyngeal Swabs from Patients With COVID-19 and Identification of Deletions in the Viral Genome That Encode Proteins Involved in Interferon Antagonism." Viruses 12, no. 10: 1164.
Summary LamPORE is a novel diagnostic platform for the detection of SARS-CoV-2 RNA that combines loop-mediated isothermal amplification with nanopore sequencing, which could potentially be used to analyse thousands of samples per day on a single instrument. We evaluated the performance of LamPORE against RT-PCR using RNA extracted from spiked respiratory samples and from stored nose and throat swabs collected at two UK hospitals. The limit of detection of LamPORE was 7-10 genome copies/µl of extracted RNA. This is above the limit achievable by RT-PCR but was not associated with a significant reduction of sensitivity in clinical samples. Positive clinical specimens came mostly from patients with acute symptomatic infection, and among these LamPORE had a diagnostic sensitivity of 99.1% (226/228 [95% CI 96.9–99.9%]). Among negative clinical specimens, including 153 with other respiratory pathogens detected, LamPORE had a diagnostic specificity of 99.6% (278/279 [98.0–100.0%]). Overall, 1.4% (7/514 [0.5–2.9]) of samples produced an indeterminate result on first testing, and repeat LamPORE testing on the same RNA extract had a reproducibility of 96.8% (478/494 [94.8–98.1]). This indicates that LamPORE has a similar performance to RT-PCR for the diagnosis of SARS-CoV-2 infection in symptomatic patients, and offers a promising approach to high-throughput testing.
Leon Peto; Gillian Rodger; Daniel P Carter; Karen L Osman; Mehmet Yavuz; Katie Johnson; Mohammad Raza; Matthew D Parker; Matthew D Wyles; Monique Andersson; Anita Justice; Alison Vaughan; Sarah Hoosdally; Nicole Stoesser; Philippa C Matthews; David W Eyre; Timothy Ea Peto; Miles W Carroll; Thushan I de Silva; Derrick W Crook; Cariad M Evans; Steven T Pullan. Diagnosis of SARS-CoV-2 infection with LamPORE, a high-throughput platform combining loop-mediated isothermal amplification and nanopore sequencing. 2020, 1 .
AMA StyleLeon Peto, Gillian Rodger, Daniel P Carter, Karen L Osman, Mehmet Yavuz, Katie Johnson, Mohammad Raza, Matthew D Parker, Matthew D Wyles, Monique Andersson, Anita Justice, Alison Vaughan, Sarah Hoosdally, Nicole Stoesser, Philippa C Matthews, David W Eyre, Timothy Ea Peto, Miles W Carroll, Thushan I de Silva, Derrick W Crook, Cariad M Evans, Steven T Pullan. Diagnosis of SARS-CoV-2 infection with LamPORE, a high-throughput platform combining loop-mediated isothermal amplification and nanopore sequencing. . 2020; ():1.
Chicago/Turabian StyleLeon Peto; Gillian Rodger; Daniel P Carter; Karen L Osman; Mehmet Yavuz; Katie Johnson; Mohammad Raza; Matthew D Parker; Matthew D Wyles; Monique Andersson; Anita Justice; Alison Vaughan; Sarah Hoosdally; Nicole Stoesser; Philippa C Matthews; David W Eyre; Timothy Ea Peto; Miles W Carroll; Thushan I de Silva; Derrick W Crook; Cariad M Evans; Steven T Pullan. 2020. "Diagnosis of SARS-CoV-2 infection with LamPORE, a high-throughput platform combining loop-mediated isothermal amplification and nanopore sequencing." , no. : 1.
Background The public health impact of Chikungunya virus (CHIKV) is often underestimated. Usually considered a mild condition of short duration, recent outbreaks have reported greater incidence of severe illness, fatality, and longer-term disability. In 2018/19, Eastern Sudan experienced the largest epidemic of CHIKV in Africa to date, affecting an estimated 487,600 people. Known locally as Kankasha, this study examines the clinical characteristics, risk factors, and phylogenetics of the CHIKV epidemic in Kassala City. Methodology/Principal Findings A prospective cohort of 142 cases (102 adults, 40 children) were enrolled at Kassala Teaching Hospital in October 2018. Clinical information, socio-demographic data and sera samples were analysed to confirm diagnosis, characterise illness, and identify the viral strain. CHIKV infection was confirmed by real-time reverse transcription-PCR in 84.5% (120/142) of participants. Nine had concurrent CHIKV/Dengue virus (DENV) infection and 28.8% had a positive Rapid Diagnostic Test for malaria. Five percent had haemorrhagic symptoms including two children with life-threatening haemorrhage. One CHIKV-positive participant died with acute renal injury. Ninety to 120 days post-illness, 63% of those followed-up were still experiencing arthralgia in one or more joints, and 11% remained moderately disabled using Rapid3 assessment. Phylogenetic analysis showed all CHIKV infections belonged to a single clade within the Indian Ocean Lineage (IOL) of the East/Central/South African (ECSA) genotype. History of contact with an infected person was the only socio-demographic factor associated with infection (p=0.01), suggesting that vector transmission in households is important. Conclusions/Significance The epidemic is estimated to have affected ∼ 50% of Kassala City’s population. Substantial vulnerability to CHIKV remains here and elsewhere in Sudan due to widespread Aedes aegypti presence and mosquito-fostering household water storage methods. This study highlights the importance of increasing awareness of the severity and socio-economic impact of CHIKV outbreaks and the need for urgent actions to reduce transmission risk in households. Author summary Chikungunya is an arboviral disease transmitted to humans by infected mosquitoes and characterised by fever and arthralgia. Although it is generally considered a short self-limiting infection, long term sequelae and severe disease are increasingly recognised. In 2018/19, Eastern Sudan experienced the largest epidemic of Chikungunya in Africa to date, affecting approximately 500,000 people. We undertook a prospective hospital-based cohort study of patients presenting with undifferentiated febrile illness in Kassala city, Sudan, supported by next-generation sequencing. We confirmed that CHIKV was the dominant pathogen, with positive CHIKV RT-PCR in 85% of patients presenting during the 7-day study period. Dengue virus was also circulating with nine CHIKV PCR-positive patients co-infected, and we identified high rates of Plasmodium falciparum malaria infection and CHIKV/malaria co-infection. Genetic sequencing confirmed Indian Ocean Lineage of the East/Central/South African CHIKV genotype. A substantial proportion of participants were admitted to hospital including children with haemorrhage, reflecting the severe phenotype linked to this genotype. Increased understanding of the health and economic burden of Chikungunya is needed, and recognition that severe and occasionally fatal infection exists. With widespread presence of Ae. aegypti and household water storage practices that encourage mosquito breeding, timely actions will be essential to prevent further large outbreaks.
Hilary Bower; Mubarak el Karsany; Abd Alhadi Adam Hussein Hussein; Mubarak Ibrahim Idriss; Ma’aaza Abasher AlZain; Mohamed Elamin Ahmed Alfakiyousif; Rehab Mohamed; Iman Mahmoud; Omer Albadri; Suha Abdulaziz Alnour Mahmoud; Orwa Ibrahim Abdalla; Mawahib Eldigail; Nuha Elagib; Ulrike Arnold; Bernardo Gutierrez; Oliver G. Pybus; Daniel P. Carter; Steven T. Pullan; Shevin T. Jacob; Tajeldin Mohammedein Abdallah; Benedict Gannon; Tom E. Fletcher. “Kankasha” in Kassala: a prospective observational cohort study of the clinical characteristics, epidemiology, genetic origin, and chronic impact of the 2018 epidemic of Chikungunya virus infection in Kassala, Sudan. 2020, 1 .
AMA StyleHilary Bower, Mubarak el Karsany, Abd Alhadi Adam Hussein Hussein, Mubarak Ibrahim Idriss, Ma’aaza Abasher AlZain, Mohamed Elamin Ahmed Alfakiyousif, Rehab Mohamed, Iman Mahmoud, Omer Albadri, Suha Abdulaziz Alnour Mahmoud, Orwa Ibrahim Abdalla, Mawahib Eldigail, Nuha Elagib, Ulrike Arnold, Bernardo Gutierrez, Oliver G. Pybus, Daniel P. Carter, Steven T. Pullan, Shevin T. Jacob, Tajeldin Mohammedein Abdallah, Benedict Gannon, Tom E. Fletcher. “Kankasha” in Kassala: a prospective observational cohort study of the clinical characteristics, epidemiology, genetic origin, and chronic impact of the 2018 epidemic of Chikungunya virus infection in Kassala, Sudan. . 2020; ():1.
Chicago/Turabian StyleHilary Bower; Mubarak el Karsany; Abd Alhadi Adam Hussein Hussein; Mubarak Ibrahim Idriss; Ma’aaza Abasher AlZain; Mohamed Elamin Ahmed Alfakiyousif; Rehab Mohamed; Iman Mahmoud; Omer Albadri; Suha Abdulaziz Alnour Mahmoud; Orwa Ibrahim Abdalla; Mawahib Eldigail; Nuha Elagib; Ulrike Arnold; Bernardo Gutierrez; Oliver G. Pybus; Daniel P. Carter; Steven T. Pullan; Shevin T. Jacob; Tajeldin Mohammedein Abdallah; Benedict Gannon; Tom E. Fletcher. 2020. "“Kankasha” in Kassala: a prospective observational cohort study of the clinical characteristics, epidemiology, genetic origin, and chronic impact of the 2018 epidemic of Chikungunya virus infection in Kassala, Sudan." , no. : 1.
Background Understanding how fungi degrade lignocellulose is a cornerstone of improving renewables-based biotechnology, in particular for the production of hydrolytic enzymes. Considerable progress has been made in investigating fungal degradation during time-points where CAZyme expression peaks. However, a robust understanding of the fungal survival strategies over its life time on lignocellulose is thereby missed. Here we aimed to uncover the physiological responses of the biotechnological workhorse and enzyme producer Aspergillus niger over its life time to six substrates important for biofuel production. Results We analysed the response of A. niger to the feedstock Miscanthus and compared it with our previous study on wheat straw, alone or in combination with hydrothermal or ionic liquid feedstock pretreatments. Conserved (substrate-independent) metabolic responses as well as those affected by pretreatment and feedstock were identified via multivariate analysis of genome-wide transcriptomics combined with targeted transcript and protein analyses and mapping to a metabolic model. Initial exposure to all substrates increased fatty acid beta-oxidation and lipid metabolism transcripts. In a strain carrying a deletion of the ortholog of the Aspergillus nidulans fatty acid beta-oxidation transcriptional regulator farA, there was a reduction in expression of selected lignocellulose degradative CAZyme-encoding genes suggesting that beta-oxidation contributes to adaptation to lignocellulose. Mannan degradation expression was wheat straw feedstock-dependent and pectin degradation was higher on the untreated substrates. In the later life stages, known and novel secondary metabolite gene clusters were activated, which are of high interest due to their potential to synthesize bioactive compounds. Conclusion In this study, which includes the first transcriptional response of Aspergilli to Miscanthus, we highlighted that life time as well as substrate composition and structure (via variations in pretreatment and feedstock) influence the fungal responses to lignocellulose. We also demonstrated that the fungal response contains physiological stages that are conserved across substrates and are typically found outside of the conditions with high CAZyme expression, as exemplified by the stages that are dominated by lipid and secondary metabolism.
Jolanda M. Van Munster; Paul Daly; Martin J. Blythe; Roger Ibbett; Matt Kokolski; Sanyasi Gaddipati; Erika Lindquist; Vasanth R. Singan; Kerrie W. Barry; Anna Lipzen; Chew Yee Ngan; Christopher J. Petzold; Leanne Jade G. Chan; Mikko Arvas; Roxane Raulo; Steven T. Pullan; Stéphane Delmas; Igor V. Grigoriev; Gregory A. Tucker; Blake A. Simmons; David B. Archer. Succession of physiological stages hallmarks the transcriptomic response of the fungus Aspergillus niger to lignocellulose. Biotechnology for Biofuels 2020, 13, 69 -16.
AMA StyleJolanda M. Van Munster, Paul Daly, Martin J. Blythe, Roger Ibbett, Matt Kokolski, Sanyasi Gaddipati, Erika Lindquist, Vasanth R. Singan, Kerrie W. Barry, Anna Lipzen, Chew Yee Ngan, Christopher J. Petzold, Leanne Jade G. Chan, Mikko Arvas, Roxane Raulo, Steven T. Pullan, Stéphane Delmas, Igor V. Grigoriev, Gregory A. Tucker, Blake A. Simmons, David B. Archer. Succession of physiological stages hallmarks the transcriptomic response of the fungus Aspergillus niger to lignocellulose. Biotechnology for Biofuels. 2020; 13 (1):69-16.
Chicago/Turabian StyleJolanda M. Van Munster; Paul Daly; Martin J. Blythe; Roger Ibbett; Matt Kokolski; Sanyasi Gaddipati; Erika Lindquist; Vasanth R. Singan; Kerrie W. Barry; Anna Lipzen; Chew Yee Ngan; Christopher J. Petzold; Leanne Jade G. Chan; Mikko Arvas; Roxane Raulo; Steven T. Pullan; Stéphane Delmas; Igor V. Grigoriev; Gregory A. Tucker; Blake A. Simmons; David B. Archer. 2020. "Succession of physiological stages hallmarks the transcriptomic response of the fungus Aspergillus niger to lignocellulose." Biotechnology for Biofuels 13, no. 1: 69-16.
COVID-19 is a complex disease phenotype where the underlying microbiome could influence morbidity and mortality. Amplicon and metagenomic MinION based sequencing was used to rapidly (within 8 hours) identify SARS-CoV-2 and assess the microbiome in nasopharyngeal swabs obtained from patients with COVID-19 by the ISARIC 4C consortium.
Shona C. Moore; Rebekah Penrice-Randal; Muhannad Alruwaili; Xiaofeng Dong; Steven T. Pullan; Daniel P. Carter; Kevin Bewley; Qin Zhao; Yani Sun; Catherine Hartley; En-Min Zhou; Tom Solomon; Michael B. J. Beadsworth; James Cruise; Debby Bogaert; Derrick W T Crook; David A. Matthews; Andrew D. Davidson; Zana Mahmood; Waleed Aljabr; Julian Druce; Richard T. Vipond; Lisa F. P. Ng; Laurent Renia; Peter J. M. Openshaw; J. Kenneth Baillie; Miles W. Carroll; Malcolm G. Semple; Lance Turtle; Julian Alexander Hiscox. Amplicon based MinION sequencing of SARS-CoV-2 and metagenomic characterisation of nasopharyngeal swabs from patients with COVID-19. 2020, 1 .
AMA StyleShona C. Moore, Rebekah Penrice-Randal, Muhannad Alruwaili, Xiaofeng Dong, Steven T. Pullan, Daniel P. Carter, Kevin Bewley, Qin Zhao, Yani Sun, Catherine Hartley, En-Min Zhou, Tom Solomon, Michael B. J. Beadsworth, James Cruise, Debby Bogaert, Derrick W T Crook, David A. Matthews, Andrew D. Davidson, Zana Mahmood, Waleed Aljabr, Julian Druce, Richard T. Vipond, Lisa F. P. Ng, Laurent Renia, Peter J. M. Openshaw, J. Kenneth Baillie, Miles W. Carroll, Malcolm G. Semple, Lance Turtle, Julian Alexander Hiscox. Amplicon based MinION sequencing of SARS-CoV-2 and metagenomic characterisation of nasopharyngeal swabs from patients with COVID-19. . 2020; ():1.
Chicago/Turabian StyleShona C. Moore; Rebekah Penrice-Randal; Muhannad Alruwaili; Xiaofeng Dong; Steven T. Pullan; Daniel P. Carter; Kevin Bewley; Qin Zhao; Yani Sun; Catherine Hartley; En-Min Zhou; Tom Solomon; Michael B. J. Beadsworth; James Cruise; Debby Bogaert; Derrick W T Crook; David A. Matthews; Andrew D. Davidson; Zana Mahmood; Waleed Aljabr; Julian Druce; Richard T. Vipond; Lisa F. P. Ng; Laurent Renia; Peter J. M. Openshaw; J. Kenneth Baillie; Miles W. Carroll; Malcolm G. Semple; Lance Turtle; Julian Alexander Hiscox. 2020. "Amplicon based MinION sequencing of SARS-CoV-2 and metagenomic characterisation of nasopharyngeal swabs from patients with COVID-19." , no. : 1.
Human metapneumovirus (HMPV) infection causes a spectrum of respiratory tract disease, and may be a significant pathogen in the context of immunocompromise. Here, we report direct-from-sample metagenomic sequencing of HMPV using Oxford Nanopore Technology. We applied this sequencing approach to 25 respiratory samples that had been submitted to a clinical diagnostic laboratory in a UK teaching hospital. These samples represented 13 patients under the care of a haematology unit over a 20-day period in Spring 2019 (two sampled twice), and ten other patients elsewhere in the hospital between 2017-2019. We generated HMPV reads from 20/25 samples (sensitivity 80% compared to routine diagnostic testing) and retrieved complete HMPV genomes from 15/20 of these. Consensus sequences from Nanopore data were identical to those generated by Illumina, and represented HMPV genomes from two distinct sublineages, A2b and B2. Sequences from ten haematology patients formed a unique genetic group in the A2b sublineage, not previously reported in the UK. Among these, eight HMPV genomes formed a cluster (differing by ≤3 SNPs), likely to reflect nosocomial transmission, while two others were more distantly related and may represent independent introductions to the haematology unit. Nanopore metagenomic sequencing can be used to diagnose HMPV infection, although more work is required to optimise sensitivity. Improvements in the use of metagenomic sequencing, particularly for respiratory viruses, could contribute to antimicrobial stewardship. Generation of full genome sequences can be used to support or rule out nosocomial transmission, and contribute to improving infection prevention and control practices.
Yifei Xu; Kuiama Lewandowski; Katie Jeffery; Louise O. Downs; Dona Foster; Nicholas D. Sanderson; James Kavanagh; Ali Vaughan; Claudia Salvagno; Richard Vipond; Miles Carroll; Robert Danby; Timothy Peto; Derrick Crook; A. Sarah Walker; Philippa C. Matthews; Steven Pullan. Nanopore metagenomic sequencing to investigate nosocomial transmission of human metapneumovirus from a unique genetic group among haematology patients in the United Kingdom. Journal of Infection 2020, 80, 571 -577.
AMA StyleYifei Xu, Kuiama Lewandowski, Katie Jeffery, Louise O. Downs, Dona Foster, Nicholas D. Sanderson, James Kavanagh, Ali Vaughan, Claudia Salvagno, Richard Vipond, Miles Carroll, Robert Danby, Timothy Peto, Derrick Crook, A. Sarah Walker, Philippa C. Matthews, Steven Pullan. Nanopore metagenomic sequencing to investigate nosocomial transmission of human metapneumovirus from a unique genetic group among haematology patients in the United Kingdom. Journal of Infection. 2020; 80 (5):571-577.
Chicago/Turabian StyleYifei Xu; Kuiama Lewandowski; Katie Jeffery; Louise O. Downs; Dona Foster; Nicholas D. Sanderson; James Kavanagh; Ali Vaughan; Claudia Salvagno; Richard Vipond; Miles Carroll; Robert Danby; Timothy Peto; Derrick Crook; A. Sarah Walker; Philippa C. Matthews; Steven Pullan. 2020. "Nanopore metagenomic sequencing to investigate nosocomial transmission of human metapneumovirus from a unique genetic group among haematology patients in the United Kingdom." Journal of Infection 80, no. 5: 571-577.
The emergence and reemergence of pathogens such as Zika virus, chikungunya virus, and yellow fever virus have drawn attention toward other cocirculating arboviruses in South America. Oropouche virus (OROV) is a poorly studied pathogen responsible for over a dozen outbreaks since the early 1960s and represents a public health burden to countries such as Brazil, Panama, and Peru. OROV is likely underreported since its symptomatology can be easily confounded with other febrile illnesses (e.g., dengue fever and leptospirosis) and point-of-care testing for the virus is still uncommon. With limited data, there is a need to optimize the information currently available. Analysis of OROV genomes can help us understand how the virus circulates in nature and can reveal the evolutionary forces that shape the genetic diversity of the virus, which has implications for molecular diagnostics and the design of potential vaccines.
Bernardo Gutierrez; Emma Wise; Steven Pullan; Christopher H. Logue; Thomas A. Bowden; Marina Escalera-Zamudio; Gabriel Trueba; Marcio R. T. Nunes; Nuno Rodrigues Faria; Oliver G. Pybus. Evolutionary Dynamics of Oropouche Virus in South America. Journal of Virology 2020, 94, 1 .
AMA StyleBernardo Gutierrez, Emma Wise, Steven Pullan, Christopher H. Logue, Thomas A. Bowden, Marina Escalera-Zamudio, Gabriel Trueba, Marcio R. T. Nunes, Nuno Rodrigues Faria, Oliver G. Pybus. Evolutionary Dynamics of Oropouche Virus in South America. Journal of Virology. 2020; 94 (5):1.
Chicago/Turabian StyleBernardo Gutierrez; Emma Wise; Steven Pullan; Christopher H. Logue; Thomas A. Bowden; Marina Escalera-Zamudio; Gabriel Trueba; Marcio R. T. Nunes; Nuno Rodrigues Faria; Oliver G. Pybus. 2020. "Evolutionary Dynamics of Oropouche Virus in South America." Journal of Virology 94, no. 5: 1.
Oropouche virus (OROV) is responsible for outbreaks of Oropouche fever in parts of South America. We recently identified and isolated OROV from a febrile Ecuadorian patient, however, a previously published qRT-PCR assay did not detect OROV in the patient sample. A primer mismatch to the Ecuadorian OROV lineage was identified from metagenomic sequencing data. We report the optimisation of an qRT-PCR assay for the Ecuadorian OROV lineage, which subsequently identified a further five cases in a cohort of 196 febrile patients. We isolated OROV via cell culture and developed an algorithmically-designed primer set for whole-genome amplification of the virus. Metagenomic sequencing of the patient samples provided OROV genome coverage ranging from 68–99%. The additional cases formed a single phylogenetic cluster together with the initial case. OROV should be considered as a differential diagnosis for Ecuadorian patients with febrile illness to avoid mis-diagnosis with other circulating pathogens. Oropouche virus (OROV) causes outbreaks of febrile illness in areas of South and Central America and we recently identified it in Ecuador for the first time, using metagenomic sequencing. The genome sequence data revealed that the Ecuadorian strain of the virus was not detected using a published qRT-PCR, as it differed genetically at the binding site of the reverse primer. To address this, we developed a modified qRT-PCR that showed increased sensitivity for the Ecuadorian strain. This test detected OROV infection in 6 out of 196 febrile patients from Esmeraldas, Ecuador in 2016. OROV was isolated from positive patient samples, viral genome sequences were compared to publicly available OROV sequences. This revealed that the Ecuadorian cases are genetically distinct, suggesting that local transmission of the virus should not be ruled out. This work highlights the need for a better understanding of OROV dynamics in Ecuador and surrounding areas, the importance of considering OROV as a cause of fever in Ecuadorian patients and the possibility of selectively using metagenomic sequencing in parallel to traditional molecular techniques in patient testing.
Emma L. Wise; Sully Márquez; Jack Mellors; Verónica Paz; Barry Atkinson; Bernardo Gutierrez; Sonia Zapata; Josefina Coloma; Oliver Pybus; Simon K. Jackson; Gabriel Trueba; Gyorgy Fejer; Christopher H. Logue; Steven Pullan. Oropouche virus cases identified in Ecuador using an optimised qRT-PCR informed by metagenomic sequencing. PLOS Neglected Tropical Diseases 2020, 14, e0007897 .
AMA StyleEmma L. Wise, Sully Márquez, Jack Mellors, Verónica Paz, Barry Atkinson, Bernardo Gutierrez, Sonia Zapata, Josefina Coloma, Oliver Pybus, Simon K. Jackson, Gabriel Trueba, Gyorgy Fejer, Christopher H. Logue, Steven Pullan. Oropouche virus cases identified in Ecuador using an optimised qRT-PCR informed by metagenomic sequencing. PLOS Neglected Tropical Diseases. 2020; 14 (1):e0007897.
Chicago/Turabian StyleEmma L. Wise; Sully Márquez; Jack Mellors; Verónica Paz; Barry Atkinson; Bernardo Gutierrez; Sonia Zapata; Josefina Coloma; Oliver Pybus; Simon K. Jackson; Gabriel Trueba; Gyorgy Fejer; Christopher H. Logue; Steven Pullan. 2020. "Oropouche virus cases identified in Ecuador using an optimised qRT-PCR informed by metagenomic sequencing." PLOS Neglected Tropical Diseases 14, no. 1: e0007897.
During February 2018–January 2019, we conducted large-scale surveillance for the presence and prevalence of tick-borne encephalitis virus (TBEV) and louping ill virus (LIV) in sentinel animals and ticks in the United Kingdom. Serum was collected from 1,309 deer culled across England and Scotland. Overall, 4% of samples were ELISA-positive for the TBEV serocomplex. A focus in the Thetford Forest area had the highest proportion (47.7%) of seropositive samples. Engorged ticks collected from culled deer within seropositive regions were tested for viral RNA; 5 of 2,041 ticks tested positive by LIV/TBEV real-time reverse transcription PCR, all from within the Thetford Forest area. From 1 tick, we identified a full-length genomic sequence of TBEV. Thus, using deer as sentinels revealed a potential TBEV focus in the United Kingdom. This detection of TBEV genomic sequence in UK ticks has important public health implications, especially for undiagnosed encephalitis.
Maya Holding; Stuart D. Dowall; Jolyon M. Medlock; Daniel Carter; Steven Pullan; James Lewis; Richard Vipond; Mara S. Rocchi; Matthew Baylis; Roger Hewson. Tick-Borne Encephalitis Virus, United Kingdom. Emerging Infectious Diseases 2020, 26, 90 -96.
AMA StyleMaya Holding, Stuart D. Dowall, Jolyon M. Medlock, Daniel Carter, Steven Pullan, James Lewis, Richard Vipond, Mara S. Rocchi, Matthew Baylis, Roger Hewson. Tick-Borne Encephalitis Virus, United Kingdom. Emerging Infectious Diseases. 2020; 26 (1):90-96.
Chicago/Turabian StyleMaya Holding; Stuart D. Dowall; Jolyon M. Medlock; Daniel Carter; Steven Pullan; James Lewis; Richard Vipond; Mara S. Rocchi; Matthew Baylis; Roger Hewson. 2020. "Tick-Borne Encephalitis Virus, United Kingdom." Emerging Infectious Diseases 26, no. 1: 90-96.
Influenza is a major global public health threat as a result of its highly pathogenic variants, large zoonotic reservoir, and pandemic potential. Metagenomic viral sequencing offers the potential for a diagnostic test for influenza virus which also provides insights on transmission, evolution, and drug resistance and simultaneously detects other viruses. We therefore set out to apply the Oxford Nanopore Technologies sequencing method to metagenomic sequencing of respiratory samples.
Kuiama Lewandowski; Yifei Xu; Steven T. Pullan; Sheila F. Lumley; Dona Foster; Nicholas Sanderson; Alison Vaughan; Marcus Morgan; Nicole Bright; James Kavanagh; Richard Vipond; Miles Carroll; Anthony Marriott; Karen E. Gooch; Monique Andersson; Katie Jeffery; Timothy E. A. Peto; Derrick W. Crook; A. Sarah Walker; Philippa C. Matthews. Metagenomic Nanopore Sequencing of Influenza Virus Direct from Clinical Respiratory Samples. Journal of Clinical Microbiology 2019, 58, 1 .
AMA StyleKuiama Lewandowski, Yifei Xu, Steven T. Pullan, Sheila F. Lumley, Dona Foster, Nicholas Sanderson, Alison Vaughan, Marcus Morgan, Nicole Bright, James Kavanagh, Richard Vipond, Miles Carroll, Anthony Marriott, Karen E. Gooch, Monique Andersson, Katie Jeffery, Timothy E. A. Peto, Derrick W. Crook, A. Sarah Walker, Philippa C. Matthews. Metagenomic Nanopore Sequencing of Influenza Virus Direct from Clinical Respiratory Samples. Journal of Clinical Microbiology. 2019; 58 (1):1.
Chicago/Turabian StyleKuiama Lewandowski; Yifei Xu; Steven T. Pullan; Sheila F. Lumley; Dona Foster; Nicholas Sanderson; Alison Vaughan; Marcus Morgan; Nicole Bright; James Kavanagh; Richard Vipond; Miles Carroll; Anthony Marriott; Karen E. Gooch; Monique Andersson; Katie Jeffery; Timothy E. A. Peto; Derrick W. Crook; A. Sarah Walker; Philippa C. Matthews. 2019. "Metagenomic Nanopore Sequencing of Influenza Virus Direct from Clinical Respiratory Samples." Journal of Clinical Microbiology 58, no. 1: 1.
Lorna Neill; Anna M. Checkley; Laura Benjamin; M. Trent Herdman; Daniel Carter; Steven Pullan; Emma Aarons; Katie Griffiths; Bernadette Monaghan; Kushan Karunaratne; Olga Ciccarelli; Jennifer Spillane; David A.J. Moore; Dimitri M. Kullmann. Rhombencephalitis and Myeloradiculitis Caused by a European Subtype of Tick-Borne Encephalitis Virus. Emerging Infectious Diseases 2019, 25, 2317 -2319.
AMA StyleLorna Neill, Anna M. Checkley, Laura Benjamin, M. Trent Herdman, Daniel Carter, Steven Pullan, Emma Aarons, Katie Griffiths, Bernadette Monaghan, Kushan Karunaratne, Olga Ciccarelli, Jennifer Spillane, David A.J. Moore, Dimitri M. Kullmann. Rhombencephalitis and Myeloradiculitis Caused by a European Subtype of Tick-Borne Encephalitis Virus. Emerging Infectious Diseases. 2019; 25 (12):2317-2319.
Chicago/Turabian StyleLorna Neill; Anna M. Checkley; Laura Benjamin; M. Trent Herdman; Daniel Carter; Steven Pullan; Emma Aarons; Katie Griffiths; Bernadette Monaghan; Kushan Karunaratne; Olga Ciccarelli; Jennifer Spillane; David A.J. Moore; Dimitri M. Kullmann. 2019. "Rhombencephalitis and Myeloradiculitis Caused by a European Subtype of Tick-Borne Encephalitis Virus." Emerging Infectious Diseases 25, no. 12: 2317-2319.
The presence of tick-borne encephalitis virus (TBEV) was detected in a questing tick pool in southern England in September 2019. Hitherto, TBEV had only been detected in a limited area in eastern England. This southern English viral genome sequence is distinct from TBEV-UK, being most similar to TBEV-NL. The new location of TBEV presence highlights that the diagnosis of tick-borne encephalitis should be considered in encephalitic patients in areas of the United Kingdom outside eastern England.
Maya Holding; Stuart D Dowall; Jolyon M Medlock; Daniel Carter; Liz McGinley; Mollie Curran-French; Steven Pullan; John Chamberlain; Kayleigh M Hansford; Matthew Baylis; Richard Vipond; Roger Hewson. Detection of new endemic focus of tick-borne encephalitis virus (TBEV), Hampshire/Dorset border, England, September 2019. Eurosurveillance 2019, 24, 1900658 .
AMA StyleMaya Holding, Stuart D Dowall, Jolyon M Medlock, Daniel Carter, Liz McGinley, Mollie Curran-French, Steven Pullan, John Chamberlain, Kayleigh M Hansford, Matthew Baylis, Richard Vipond, Roger Hewson. Detection of new endemic focus of tick-borne encephalitis virus (TBEV), Hampshire/Dorset border, England, September 2019. Eurosurveillance. 2019; 24 (47):1900658.
Chicago/Turabian StyleMaya Holding; Stuart D Dowall; Jolyon M Medlock; Daniel Carter; Liz McGinley; Mollie Curran-French; Steven Pullan; John Chamberlain; Kayleigh M Hansford; Matthew Baylis; Richard Vipond; Roger Hewson. 2019. "Detection of new endemic focus of tick-borne encephalitis virus (TBEV), Hampshire/Dorset border, England, September 2019." Eurosurveillance 24, no. 47: 1900658.
BackgroundUnderstanding how fungi degrade lignocellulose is a cornerstone of improving renewables-based biotechnology, in particular for the production of hydrolytic enzymes. Considerable progress has been made in investigating fungal degradation during time-points where CAZyme expression peaks. However, a robust understanding of the fungal survival strategies over its life time on lignocellulose is thereby missed. Here we aimed to uncover the physiological responses of the biotechnological workhorse and enzyme producerAspergilllus nigerover its life time to six substrates important for biofuel production.ResultsWe analysed the response ofA. nigerto the feedstockMiscanthusand compared it with our previous study on wheat straw, alone or in combination with hydrothermal or ionic liquid feedstock pretreatments. Conserved (substrate-independent) metabolic responses as well as those affected by pretreatment and feedstock were identified via multivariate analysis of genome-wide transcriptomics combined with targeted transcript and protein analyses and mapping to a metabolic model. Initial exposure to all substrates increased fatty acid beta-oxidation and lipid metabolism transcripts. In a strain carrying a deletion of the ortholog of theAspergillus nidulansfatty acid beta-oxidation transcriptional regulatorfarA, there was a reduction in expression of selected lignocellulose degradative CAZyme-encoding genes suggesting that beta-oxidation contributes to adaptation to lignocellulose. Mannan degradation expression was wheat straw feedstock-dependent and pectin degradation was higher on the untreated substrates. In the later life stages, known and novel secondary metabolite gene clusters were activated, which are of high interest due to their potential to synthesize bioactive compounds.ConclusionIn this study, which includes the first transcriptional response of Aspergilli toMiscanthus, we highlighted that life time as well as substrate composition and structure (via variations in pretreatment and feedstock) influence the fungal responses to lignocellulose. We also demonstrated that the fungal response contains physiological stages that are conserved across substrates and are typically found outside of the conditions with high CAZyme expression, as exemplified by the stages that are dominated by lipid and secondary metabolism.
Jolanda M. Van Munster; Paul Daly; Martin J. Blythe; Roger Ibbett; Matt Kokolski; Sanyasi Gaddipati; Erika Lindquist; Vasanth R. Singan; Kerrie W. Barry; Anna M Lipzen; Chew Yee Ngan; Christopher J. Petzold; Leanne Jade G. Chan; Mikko Arvas; Roxane Raulo; Steven T. Pullan; Stéphane Delmas; Igor V. Grigoriev; Gregory A. Tucker; Blake A. Simmons; David B. Archer. Succession of physiological stages hallmarks the transcriptomic response of fungusAspergillus nigerto lignocellulose. 2019, 806356 .
AMA StyleJolanda M. Van Munster, Paul Daly, Martin J. Blythe, Roger Ibbett, Matt Kokolski, Sanyasi Gaddipati, Erika Lindquist, Vasanth R. Singan, Kerrie W. Barry, Anna M Lipzen, Chew Yee Ngan, Christopher J. Petzold, Leanne Jade G. Chan, Mikko Arvas, Roxane Raulo, Steven T. Pullan, Stéphane Delmas, Igor V. Grigoriev, Gregory A. Tucker, Blake A. Simmons, David B. Archer. Succession of physiological stages hallmarks the transcriptomic response of fungusAspergillus nigerto lignocellulose. . 2019; ():806356.
Chicago/Turabian StyleJolanda M. Van Munster; Paul Daly; Martin J. Blythe; Roger Ibbett; Matt Kokolski; Sanyasi Gaddipati; Erika Lindquist; Vasanth R. Singan; Kerrie W. Barry; Anna M Lipzen; Chew Yee Ngan; Christopher J. Petzold; Leanne Jade G. Chan; Mikko Arvas; Roxane Raulo; Steven T. Pullan; Stéphane Delmas; Igor V. Grigoriev; Gregory A. Tucker; Blake A. Simmons; David B. Archer. 2019. "Succession of physiological stages hallmarks the transcriptomic response of fungusAspergillus nigerto lignocellulose." , no. : 806356.
Undifferentiated febrile illness (UFI) is one of the most common reasons for people seeking healthcare in low-income countries. While illness and death due to specific infections such as malaria are often well-quantified, others are frequently uncounted and their impact underappreciated. A number of high consequence infectious diseases, including Ebola virus, are endemic or epidemic in the Federal Republic of Sudan which has experienced at least 12 UFI outbreaks, frequently associated with haemorrhage and high case fatality rates (CFR), since 2012. One of these occurred in Darfur in 2015/2016 with 594 cases and 108 deaths (CFR 18.2%). The aetiology of these outbreaks remains unknown. We report a retrospective cohort study of the 2015/2016 Darfur outbreak, using a subset of 65 of 263 outbreak samples received by the National Public Health Laboratory which met selection criteria of sufficient sample volume and epidemiological data. Clinical features included fever (95.8%), bleeding (95.7%), headache (51.6%) and arthralgia (42.2%). No epidemiological patterns indicative of person-to-person transmission or health-worker cases were reported. Samples were tested at the Public Health England Rare and Imported Pathogens Laboratory using a bespoke panel of likely pathogens including haemorrhagic fever viruses, arboviruses and Rickettsia, Leptospira and Borrelia spp. Seven (11%) were positive for Crimean-Congo haemorrhagic fever virus (CCHFV) by real-time reverse transcription PCR. The remaining samples tested negative on all assays. CCHFV is an important cause of fever and haemorrhage in Darfur, but not the sole major source of UFI outbreaks in Sudan. Prospective studies are needed to explore other aetiologies, including novel pathogens. The presence of CCHFV has critical infection, prevention and control as well as clinical implications for future response. Our study reinforces the need to boost surveillance, lab and investigative capacity to underpin effective response, and for local and international health security. The Federal Republic of Sudan has had at least 12 outbreaks of febrile illness of unknown cause associated with symptoms of haemorrhage and high case fatality rates since 2012. Outbreaks without clear diagnosis are concerning, particularly in countries such as Sudan where a range of high consequence diseases, including viral haemorrhagic fevers, are endemic or epidemic, and local laboratory capacity is limited. We transferred historical samples stored in the National Public Health Authority from one of these outbreaks that occurred in Darfur 2015–2016 to the Public Health England Laboratory at Porton, UK, and tested them against a wide range of infectious diseases to try to identify the cause, and to help the Sudanese Federal Ministry of Health to develop and target their limited laboratory capacity. We found that Crimean-Congo Haemorrhagic Fever was an important cause but not the only source of cases in this outbreak. This has implications for prevention and control as well as for treating cases. Our study also highlighted the need for future studies to explore other possible causes, including new pathogens, and reinforced the need to boost surveillance, lab and investigative capacity for more timely and complete outbreak response.
Hilary Bower; Mubarak El Karsany; Mazza Alzain; Benedict Gannon; Rehab Mohamed; Iman Mahmoud; Mawahib Eldegail; Rihab Taha; Abdalla Osman; Salim Mohamednour; Amanda Semper; Barry Atkinson; Daniel Carter; Stuart Dowall; Jenna Furneaux; Victoria Graham; Jack Mellors; Jane Osborne; Steven T. Pullan; Gillian S. Slack; Tim Brooks; Roger Hewson; Nicholas J. Beeching; Jimmy Whitworth; Daniel G. Bausch; Tom E. Fletcher. Detection of Crimean-Congo Haemorrhagic Fever cases in a severe undifferentiated febrile illness outbreak in the Federal Republic of Sudan: A retrospective epidemiological and diagnostic cohort study. PLOS Neglected Tropical Diseases 2019, 13, e0007571 .
AMA StyleHilary Bower, Mubarak El Karsany, Mazza Alzain, Benedict Gannon, Rehab Mohamed, Iman Mahmoud, Mawahib Eldegail, Rihab Taha, Abdalla Osman, Salim Mohamednour, Amanda Semper, Barry Atkinson, Daniel Carter, Stuart Dowall, Jenna Furneaux, Victoria Graham, Jack Mellors, Jane Osborne, Steven T. Pullan, Gillian S. Slack, Tim Brooks, Roger Hewson, Nicholas J. Beeching, Jimmy Whitworth, Daniel G. Bausch, Tom E. Fletcher. Detection of Crimean-Congo Haemorrhagic Fever cases in a severe undifferentiated febrile illness outbreak in the Federal Republic of Sudan: A retrospective epidemiological and diagnostic cohort study. PLOS Neglected Tropical Diseases. 2019; 13 (7):e0007571.
Chicago/Turabian StyleHilary Bower; Mubarak El Karsany; Mazza Alzain; Benedict Gannon; Rehab Mohamed; Iman Mahmoud; Mawahib Eldegail; Rihab Taha; Abdalla Osman; Salim Mohamednour; Amanda Semper; Barry Atkinson; Daniel Carter; Stuart Dowall; Jenna Furneaux; Victoria Graham; Jack Mellors; Jane Osborne; Steven T. Pullan; Gillian S. Slack; Tim Brooks; Roger Hewson; Nicholas J. Beeching; Jimmy Whitworth; Daniel G. Bausch; Tom E. Fletcher. 2019. "Detection of Crimean-Congo Haemorrhagic Fever cases in a severe undifferentiated febrile illness outbreak in the Federal Republic of Sudan: A retrospective epidemiological and diagnostic cohort study." PLOS Neglected Tropical Diseases 13, no. 7: e0007571.
The Amazon basin is host to numerous arthropod-borne viral pathogens that cause febrile disease in humans. Among these,Oropouche orthobunyavirus(OROV) is a relatively understudied member of the Peribunyavirales that causes periodic outbreaks in human populations in Brazil and other South American countries. Although several studies have described the genetic diversity of the virus, the evolutionary processes that shape the viral genome remain poorly understood. Here we present a comprehensive study of the genomic dynamics of OROV that encompasses phylogenetic analysis, evolutionary rate estimates, inference of natural selective pressures, recombination and reassortment, and structural analysis of OROV variants. Our study includes all available published sequences, as well as a set of new OROV genomes sequences obtained from patients in Ecuador, representing the first set of viral genomes from this country. Our results show that differing evolutionary processes on the three segments that encompass the viral genome lead to variable evolutionary rates and TMRCAs that could be explained by cryptic reassortment. We also present the discovery of previously unobserved putative N-linked glycosylation sites, and codons which evolve under positive selection on the viral surface proteins, and discuss the potential role of these features in the evolution of the virus through a combined phylogenetic and structural approach.
Bernardo Gutierrez; Emma Wise; Steven Pullan; Christopher Logue; Thomas A. A Bowden; Gabriel Trueba; Marcio Nunes; Nuno R. Faria; Oliver G. Pybus. The evolutionary dynamics of Oropouche Virus (OROV) in South America. 2019, 682559 .
AMA StyleBernardo Gutierrez, Emma Wise, Steven Pullan, Christopher Logue, Thomas A. A Bowden, Gabriel Trueba, Marcio Nunes, Nuno R. Faria, Oliver G. Pybus. The evolutionary dynamics of Oropouche Virus (OROV) in South America. . 2019; ():682559.
Chicago/Turabian StyleBernardo Gutierrez; Emma Wise; Steven Pullan; Christopher Logue; Thomas A. A Bowden; Gabriel Trueba; Marcio Nunes; Nuno R. Faria; Oliver G. Pybus. 2019. "The evolutionary dynamics of Oropouche Virus (OROV) in South America." , no. : 682559.
Influenza is a major global public health threat as a result of its highly pathogenic variants, large zoonotic reservoir, and pandemic potential. Metagenomic viral sequencing offers the potential of a diagnostic test for influenza which also provides insights on transmission, evolution and drug resistance, and simultaneously detects other viruses. We therefore set out to apply Oxford Nanopore Technology to metagenomic sequencing of respiratory samples. We generated influenza reads down to a limit of detection of 102-103 genome copies/ml in pooled samples, observing a strong relationship between the viral titre and the proportion of influenza reads (p = 4.7×10-5). Applying our methods to clinical throat swabs, we generated influenza reads for 27/27 samples with high-to-mid viral titres (Cycle threshold (Ct) values 99% complete sequence for all eight gene segments. We also detected Human Coronavirus and generated a near complete Human Metapneumovirus genome from clinical samples. While further optimisation is required to improve sensitivity, this approach shows promise for the Nanopore platform to be used in the diagnosis and genetic analysis of influenza and other respiratory viruses.
Kuiama Lewandowski; Yifei Xu; Steven T. Pullan; Sheila F. Lumley; Dona Foster; Nicholas D Sanderson; Alison Vaughan; Marcus Morgan; Nicole Bright; James Kavanagh; Richard T Vipond; Miles W Carroll; Anthony C. Marriott; Karen E Gooch; Monique Andersson; Katie Jeffery; Timothy E. A. Peto; Derrick W. Crook; A Sarah Walker; Philippa C. Matthews. Metagenomic Nanopore sequencing of influenza virus direct from clinical respiratory samples. 2019, 676155 .
AMA StyleKuiama Lewandowski, Yifei Xu, Steven T. Pullan, Sheila F. Lumley, Dona Foster, Nicholas D Sanderson, Alison Vaughan, Marcus Morgan, Nicole Bright, James Kavanagh, Richard T Vipond, Miles W Carroll, Anthony C. Marriott, Karen E Gooch, Monique Andersson, Katie Jeffery, Timothy E. A. Peto, Derrick W. Crook, A Sarah Walker, Philippa C. Matthews. Metagenomic Nanopore sequencing of influenza virus direct from clinical respiratory samples. . 2019; ():676155.
Chicago/Turabian StyleKuiama Lewandowski; Yifei Xu; Steven T. Pullan; Sheila F. Lumley; Dona Foster; Nicholas D Sanderson; Alison Vaughan; Marcus Morgan; Nicole Bright; James Kavanagh; Richard T Vipond; Miles W Carroll; Anthony C. Marriott; Karen E Gooch; Monique Andersson; Katie Jeffery; Timothy E. A. Peto; Derrick W. Crook; A Sarah Walker; Philippa C. Matthews. 2019. "Metagenomic Nanopore sequencing of influenza virus direct from clinical respiratory samples." , no. : 676155.
The 2018 Nigerian Lassa fever season saw the largest ever recorded upsurge of cases, raising concerns over the emergence of a strain with increased transmission rate. To understand the molecular epidemiology of this upsurge, we performed, for the first time at the epicenter of an unfolding outbreak, metagenomic nanopore sequencing directly from patient samples, an approach dictated by the highly variable genome of the target pathogen. Genomic data and phylogenetic reconstructions were communicated immediately to Nigerian authorities and the World Health Organization to inform the public health response. Real-time analysis of 36 genomes and subsequent confirmation using all 120 samples sequenced in the country of origin revealed extensive diversity and phylogenetic intermingling with strains from previous years, suggesting independent zoonotic transmission events and thus allaying concerns of an emergent strain or extensive human-to-human transmission.
L. E. Kafetzopoulou; S. T. Pullan; P. Lemey; M. A. Suchard; D. U. Ehichioya; M. Pahlmann; A. Thielebein; J. Hinzmann; L. Oestereich; D. M. Wozniak; K. Efthymiadis; D. Schachten; F. Koenig; J. Matjeschk; S. Lorenzen; S. Lumley; Y. Ighodalo; D. I. Adomeh; T. Olokor; E. Omomoh; R. Omiunu; J. Agbukor; B. Ebo; J. Aiyepada; P. Ebhodaghe; B. Osiemi; S. Ehikhametalor; P. Akhilomen; M. Airende; R. Esumeh; E. Muoebonam; R. Giwa; A. Ekanem; G. Igenegbale; G. Odigie; G. Okonofua; R. Enigbe; J. Oyakhilome; E. O. Yerumoh; I. Odia; C. Aire; R. Atafo; E. Tobin; D. Asogun; N. Akpede; P. O. Okokhere; M. O. Rafiu; K. O. Iraoyah; C. O. Iruolagbe; P. Akhideno; C. Erameh; G. Akpede; E. Isibor; D. Naidoo; R. Hewson; J. A. Hiscox; R. Vipond; M. W. Carroll; C. Ihekweazu; P. Formenty; S. Okogbenin; E. Ogbaini-Emovon; S. Günther; S. Duraffour. Metagenomic sequencing at the epicenter of the Nigeria 2018 Lassa fever outbreak. Science 2019, 363, 74 -77.
AMA StyleL. E. Kafetzopoulou, S. T. Pullan, P. Lemey, M. A. Suchard, D. U. Ehichioya, M. Pahlmann, A. Thielebein, J. Hinzmann, L. Oestereich, D. M. Wozniak, K. Efthymiadis, D. Schachten, F. Koenig, J. Matjeschk, S. Lorenzen, S. Lumley, Y. Ighodalo, D. I. Adomeh, T. Olokor, E. Omomoh, R. Omiunu, J. Agbukor, B. Ebo, J. Aiyepada, P. Ebhodaghe, B. Osiemi, S. Ehikhametalor, P. Akhilomen, M. Airende, R. Esumeh, E. Muoebonam, R. Giwa, A. Ekanem, G. Igenegbale, G. Odigie, G. Okonofua, R. Enigbe, J. Oyakhilome, E. O. Yerumoh, I. Odia, C. Aire, R. Atafo, E. Tobin, D. Asogun, N. Akpede, P. O. Okokhere, M. O. Rafiu, K. O. Iraoyah, C. O. Iruolagbe, P. Akhideno, C. Erameh, G. Akpede, E. Isibor, D. Naidoo, R. Hewson, J. A. Hiscox, R. Vipond, M. W. Carroll, C. Ihekweazu, P. Formenty, S. Okogbenin, E. Ogbaini-Emovon, S. Günther, S. Duraffour. Metagenomic sequencing at the epicenter of the Nigeria 2018 Lassa fever outbreak. Science. 2019; 363 (6422):74-77.
Chicago/Turabian StyleL. E. Kafetzopoulou; S. T. Pullan; P. Lemey; M. A. Suchard; D. U. Ehichioya; M. Pahlmann; A. Thielebein; J. Hinzmann; L. Oestereich; D. M. Wozniak; K. Efthymiadis; D. Schachten; F. Koenig; J. Matjeschk; S. Lorenzen; S. Lumley; Y. Ighodalo; D. I. Adomeh; T. Olokor; E. Omomoh; R. Omiunu; J. Agbukor; B. Ebo; J. Aiyepada; P. Ebhodaghe; B. Osiemi; S. Ehikhametalor; P. Akhilomen; M. Airende; R. Esumeh; E. Muoebonam; R. Giwa; A. Ekanem; G. Igenegbale; G. Odigie; G. Okonofua; R. Enigbe; J. Oyakhilome; E. O. Yerumoh; I. Odia; C. Aire; R. Atafo; E. Tobin; D. Asogun; N. Akpede; P. O. Okokhere; M. O. Rafiu; K. O. Iraoyah; C. O. Iruolagbe; P. Akhideno; C. Erameh; G. Akpede; E. Isibor; D. Naidoo; R. Hewson; J. A. Hiscox; R. Vipond; M. W. Carroll; C. Ihekweazu; P. Formenty; S. Okogbenin; E. Ogbaini-Emovon; S. Günther; S. Duraffour. 2019. "Metagenomic sequencing at the epicenter of the Nigeria 2018 Lassa fever outbreak." Science 363, no. 6422: 74-77.
Human metapneumovirus (HMPV) has been recognized as an important pathogen which can cause a spectrum of respiratory tract disease. Here, we report Nanopore metagenomic sequencing of the first full length HMPV genome directly from a throat swab from a UK patient with complex lung disease and immunocompromise. We found a predominance (26.4%) of HMPV reads in the metagenomic sequencing data and consequently assembled the full genome at a high depth of coverage (mean 4,786). Through phylogenetic analyses, we identified this HMPV strain to originate from a unique genetic group in A2b, showing the presence of this group in the UK. Our study demonstrated the effectiveness of Nanopore metagenomic sequencing for diagnosing infectious diseases and recovering complete sequences for genomic characterization, highlighting the applicability of Nanopore sequencing in clinical settings.
Yifei Xu; Kuiama Lewandowski; Sheila Lumley; Nicholas D Sanderson; Alison Vaughan; Richard T Vipond; Miles W Carroll; Katie Jeffery; Dona Foster; A Sarah Walker; Timothy E. A. Peto; Derrick W Crook; Steven T Pullan; Philippa C. Matthews. Nanopore metagenomic sequencing of full length human metapneumovirus (HMPV) within a unique sub-lineage. 2018, 496687 .
AMA StyleYifei Xu, Kuiama Lewandowski, Sheila Lumley, Nicholas D Sanderson, Alison Vaughan, Richard T Vipond, Miles W Carroll, Katie Jeffery, Dona Foster, A Sarah Walker, Timothy E. A. Peto, Derrick W Crook, Steven T Pullan, Philippa C. Matthews. Nanopore metagenomic sequencing of full length human metapneumovirus (HMPV) within a unique sub-lineage. . 2018; ():496687.
Chicago/Turabian StyleYifei Xu; Kuiama Lewandowski; Sheila Lumley; Nicholas D Sanderson; Alison Vaughan; Richard T Vipond; Miles W Carroll; Katie Jeffery; Dona Foster; A Sarah Walker; Timothy E. A. Peto; Derrick W Crook; Steven T Pullan; Philippa C. Matthews. 2018. "Nanopore metagenomic sequencing of full length human metapneumovirus (HMPV) within a unique sub-lineage." , no. : 496687.
Background The recent global emergence and re-emergence of arboviruses has caused significant human disease. Common vectors, symptoms and geographical distribution make differential diagnosis both important and challenging. Aim To investigate the feasibility of metagenomic sequencing for recovering whole genome sequences of chikungunya and dengue viruses from clinical samples. Methods We performed metagenomic sequencing using both the Illumina MiSeq and the portable Oxford Nanopore MinION on clinical samples which were real-time reverse transcription-PCR (qRT-PCR) positive for chikungunya (CHIKV) or dengue virus (DENV), two of the most important arboviruses. A total of 26 samples with a range of representative clinical Ct values were included in the study. Results Direct metagenomic sequencing of nucleic acid extracts from serum or plasma without viral enrichment allowed for virus identification, subtype determination and elucidated complete or near-complete genomes adequate for phylogenetic analysis. One PCR-positive CHIKV sample was also found to be coinfected with DENV. Conclusions This work demonstrates that metagenomic whole genome sequencing is feasible for the majority of CHIKV and DENV PCR-positive patient serum or plasma samples. Additionally, it explores the use of Nanopore metagenomic sequencing for DENV and CHIKV, which can likely be applied to other RNA viruses, highlighting the applicability of this approach to front-line public health and potential portable applications using the MinION.
Liana E. Kafetzopoulou; Kyriakos Efthymiadis; Kuiama Lewandowski; Ant Crook; Dan Carter; Jane Osborne; Emma Aarons; Roger Hewson; Julian A. Hiscox; Miles W. Carroll; Richard Vipond; Steven T. Pullan. Assessment of metagenomic Nanopore and Illumina sequencing for recovering whole genome sequences of chikungunya and dengue viruses directly from clinical samples. Eurosurveillance 2018, 23, 1800228 .
AMA StyleLiana E. Kafetzopoulou, Kyriakos Efthymiadis, Kuiama Lewandowski, Ant Crook, Dan Carter, Jane Osborne, Emma Aarons, Roger Hewson, Julian A. Hiscox, Miles W. Carroll, Richard Vipond, Steven T. Pullan. Assessment of metagenomic Nanopore and Illumina sequencing for recovering whole genome sequences of chikungunya and dengue viruses directly from clinical samples. Eurosurveillance. 2018; 23 (50):1800228.
Chicago/Turabian StyleLiana E. Kafetzopoulou; Kyriakos Efthymiadis; Kuiama Lewandowski; Ant Crook; Dan Carter; Jane Osborne; Emma Aarons; Roger Hewson; Julian A. Hiscox; Miles W. Carroll; Richard Vipond; Steven T. Pullan. 2018. "Assessment of metagenomic Nanopore and Illumina sequencing for recovering whole genome sequences of chikungunya and dengue viruses directly from clinical samples." Eurosurveillance 23, no. 50: 1800228.
Hybrid de novo assembly of Illumina/Nanopore reads produced a complete closed genome sequence of the chromosome and two virulence plasmids of a Bacillus anthracis isolate from a fatal anthrax case in the United Kingdom linked to imported animal skins/drums; this provides a high-quality representative sequence for this lineage.
Steven T. Pullan; Rory W. Miles; Kuiama Lewandowski; Richard Vipond. Closed Genome Sequence Obtained Using Hybrid Nanopore/Illumina Assembly of a Bacillus anthracis Isolate from an Animal-Skin-Drum-Associated Anthrax Case in the United Kingdom. Microbiology Resource Announcements 2018, 7, e00802-18 .
AMA StyleSteven T. Pullan, Rory W. Miles, Kuiama Lewandowski, Richard Vipond. Closed Genome Sequence Obtained Using Hybrid Nanopore/Illumina Assembly of a Bacillus anthracis Isolate from an Animal-Skin-Drum-Associated Anthrax Case in the United Kingdom. Microbiology Resource Announcements. 2018; 7 (13):e00802-18.
Chicago/Turabian StyleSteven T. Pullan; Rory W. Miles; Kuiama Lewandowski; Richard Vipond. 2018. "Closed Genome Sequence Obtained Using Hybrid Nanopore/Illumina Assembly of a Bacillus anthracis Isolate from an Animal-Skin-Drum-Associated Anthrax Case in the United Kingdom." Microbiology Resource Announcements 7, no. 13: e00802-18.