The Fecal Virome of Children with Hand, Foot, and Mouth Disease that Tested PCR Negative for Pathogenic Enteroviruses

Linsuwanon, Piyada; Poovorawan, Yong; Li, Linlin; Deng, Xutao; Vongpunsawad, Sompong; Delwart, Eric

doi:10.1371/journal.pone.0135573

Cited by 19 publications

(9 citation statements)

References 105 publications

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“…The clinical phenotype resulting from viral infections is determined by complex interactions of multiple factors [ 49 ]. Besides the virus, host factors, the intestinal virome, coinfection with more than two EV types, and the environment can also influence HFMD outcome [ 6 , 50 – 55 ]. No coinfection was found in patients with severe HFMD based on the traditional experimental methods used in this study.…”

Section: Discussionmentioning

confidence: 99%

Molecular epidemiology of enteroviruses associated with severe hand, foot and mouth disease in Shenzhen, China, 2014-2018

Chen

Yang

et al. 2020

Arch Virol

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In this study, we investigated the epidemiology and molecular characteristics of enteroviruses associated with severe hand, foot and mouth disease (HFMD) in Shenzhen, China, during 2014-2018. A total of 137 fecal specimens from patients with severe HFMD were collected. Enterovirus (EV) types were determined using real-time reverse transcription polymerase chain reaction (RT-PCR), RT nested PCR, and sequencing. Sequences were analyzed using bioinformatics programs. Of 137 specimens tested, 97 (70.8%), 12 (8.8%), and 10 (7.3%) were positive for EV-A71, coxsackievirus A6 (CVA6), and CVA16, respectively. Other pathogens detected included CVA2 (2.9%, 4/137), CVA10 (2.9%, 4/137), CVA5 (0.7%, 1/137), echovirus 6 (E6) (0.7%, 1/137) and E18 (0.7%, 1/137). The most frequent complication in patients with proven EV infections was myoclonic jerk, followed by aseptic encephalitis, tachypnea, and vomiting. The frequencies of vomiting and abnormal eye movements were higher in EV-A71-infected patients than that in CVA6-infected or CVA16-infected patients. Molecular phylogeny based on the complete VP1 gene revealed no association between the subgenotype of the virus and disease severity. Nevertheless, 12 significant mutations that were likely to be associated with virulence or the clinical phenotype were observed in the 5’UTR, 2A pro , 2C, 3A, 3D pol and 3’UTR of CVA6. Eight significant mutations were observed in the 5’UTR, 2B, 3A, 3D pol and 3’UTR of CVA16, and 10 significant mutations were observed in the 5’UTR, VP1, 3A and 3C pro of CVA10. In conclusion, EV-A71 is still the main pathogen causing severe HFMD, although other EV types can also cause severe complications. Potential virulence or phenotype-associated sites were identified in the genomes of CVA6, CVA16, and CVA10. Electronic supplementary material The online version of this article (10.1007/s00705-020-04734-z) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Molecular epidemiology of enteroviruses associated with severe hand, foot and mouth disease in Shenzhen, China, 2014-2018

Chen

Yang

et al. 2020

Arch Virol

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“…Exploring the use of next-generation sequencing based assay as a diagnostic tool was an objective in many recent reports [ 30 – 32 ]. In addition, next-generation sequencing has been shown to be able to establish the diagnostics in swabs from HFMD patients that were enterovirus specific PCR negative [ 33 ]. Similarly, our assay could sequence and provide correct serotype information of the targeted enteroviruses in all tested samples with Cp values between 20.9 and 33.2, although we did not test our assay on samples with lower viral load (i.e.…”

Section: Discussionmentioning

confidence: 99%

Development and evaluation of a non-ribosomal random PCR and next-generation sequencing based assay for detection and sequencing of hand, foot and mouth disease pathogens

Nguyen

Tran

Van

et al. 2016

Virol J

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BackgroundHand, foot and mouth disease (HFMD) has become a major public health problem across the Asia-Pacific region, and is commonly caused by enterovirus A71 (EV-A71) and coxsackievirus A6 (CV-A6), CV-A10 and CV-A16. Generating pathogen whole-genome sequences is essential for understanding their evolutionary biology. The frequent replacements among EV serotypes and a limited numbers of available whole-genome sequences hinder the development of overlapping PCRs for whole-genome sequencing.We developed and evaluated a non-ribosomal random PCR (rPCR) and next-generation sequencing based assay for sequence-independent whole-genome amplification and sequencing of HFMD pathogens. A total of 16 EV-A71/CV-A6/CV-A10/CV-A16 PCR positive rectal/throat swabs (Cp values: 20.9–33.3) were used for assay evaluation.ResultsOur assay evidently outperformed the conventional rPCR in terms of the total number of EV-A71 reads and the percentage of EV-A71 reads: 2.6 % (1275/50,000 reads) vs. 0.1 % (31/50,000) and 6 % (3008/50,000) vs. 0.9 % (433/50,000) for two samples with Cp values of 30 and 26, respectively. Additionally the assay could generate genome sequences with the percentages of coverage of 94–100 % of 4 different enterovirus serotypes in 73 % of the tested samples, representing the first whole-genome sequences of CV-A6/10/16 from Vietnam, and could assign correctly serotyping results in 100 % of 24 tested specimens. In all but three the obtained consensuses of two replicates from the same sample were 100 % identical, suggesting that our assay is highly reproducible.ConclusionsIn conclusion, we have successfully developed a non-ribosomal rPCR and next-generation sequencing based assay for sensitive detection and direct whole-genome sequencing of HFMD pathogens from clinical samples.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-016-0580-9) contains supplementary material, which is available to authorized users.

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“…Such a case has been made for the marine vesiviruses in studies that have reported a higher prevalence of vesivirus antibodies in certain patient groups with hepatic disease ( 36 , 37 ). It is noteworthy that virus discovery efforts by deep sequencing have not detected vesivirus RNA in human clinical samples ( 49 ), but vesivirus sequences have been detected with these techniques in sea lions and other wild animals ( 50 ). These data suggest that species tropism, largely determined by receptor specificity, is responsible for the host range of these viruses and that interspecies transmission is a rare event.…”

Section: Discussionmentioning

confidence: 99%

Identification of Human Junctional Adhesion Molecule 1 as a Functional Receptor for the Hom-1 Calicivirus on Human Cells

Sosnovtsev

Sandoval-Jaime

Parra

et al. 2017

mBio

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The Hom-1 vesivirus was reported in 1998 following the inadvertent transmission of the animal calicivirus San Miguel sea lion virus to a human host in a laboratory. We characterized the Hom-1 strain and investigated the mechanism by which human cells could be infected. An expression library of 3,559 human plasma membrane proteins was screened for reactivity with Hom-1 virus-like particles, and a single interacting protein, human junctional adhesion molecule 1 (hJAM1), was identified. Transient expression of hJAM1 conferred susceptibility to Hom-1 infection on nonpermissive Chinese hamster ovary (CHO) cells. Virus infection was markedly inhibited when CHO cells stably expressing hJAM were pretreated with anti-hJAM1 monoclonal antibodies. Cell lines of human origin were tested for growth of Hom-1, and efficient replication was observed in HepG2, HuH7, and SK-CO15 cells. The three cell lines (of hepatic or intestinal origin) were confirmed to express hJAM1 on their surface, and clustered regularly interspaced short palindromic repeats/Cas9-mediated knockout of the hJAM1 gene in each line abolished Hom-1 propagation. Taken together, our data indicate that entry of the Hom-1 vesivirus into these permissive human cell lines is mediated by the plasma membrane protein hJAM1 as a functional receptor.

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The Fecal Virome of Children with Hand, Foot, and Mouth Disease that Tested PCR Negative for Pathogenic Enteroviruses

Cited by 19 publications

References 105 publications

Molecular epidemiology of enteroviruses associated with severe hand, foot and mouth disease in Shenzhen, China, 2014-2018

Molecular epidemiology of enteroviruses associated with severe hand, foot and mouth disease in Shenzhen, China, 2014-2018

Development and evaluation of a non-ribosomal random PCR and next-generation sequencing based assay for detection and sequencing of hand, foot and mouth disease pathogens

Identification of Human Junctional Adhesion Molecule 1 as a Functional Receptor for the Hom-1 Calicivirus on Human Cells

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