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Preponderance of enterovirus C in RD-L20B-cell-culture-negative stool samples from children diagnosed with acute flaccid paralysis in Nigeria

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Abstract

Recently, a reverse transcriptase semi-nested polymerase chain reaction (RT-snPCR) assay was recommended by the WHO for direct detection of enteroviruses in clinical specimens. In this study, we use this assay and a modification thereof to screen acute flaccid paralysis (AFP) samples that had previously tested negative for enteroviruses by the RD-L20B algorithm. Thirty paired stool suspensions collected in 2015 as part of the national AFP surveillance program in different states of Nigeria were analyzed in this study. The samples had previously tested negative for enteroviruses in the polio laboratory in accordance with the WHO-recommended RD-L20B-cell-culture-based algorithm. Two samples that had previously been found to contain enteroviruses were included as positive controls. All samples were subjected to RNA extraction, the RT-snPCR assay and a modified version of the RT-snPCR. All amplicons were sequenced, and enteroviruses were identified using the enterovirus genotyping tool and phylogenetic analysis. Amplicons were recovered from the two controls and 50% (15/30) of the samples screened. Fourteen were successfully typed, of which, 7.1% (1/14), 21.4% (3/14), 64.3% (9/14) and 7.1% (1/14) were enterovirus (EV) -A, EV-B, EV-C and a mixture of EV-B and C (EV-C99 and E25), respectively. The two controls were identified as EV-C99 and coxsackievirus (CV) -A1, both of which belong to the species Enterovirus C. In one sample, poliovirus serotype 2 was detected and found to have the VP1 ILE143 variation and was therefore identified as a vaccine strain. The results of this study showed that significant proportion of enterovirus infections (including some with Sabin PV2) are being missed by the RD-L20B-cell-culture-based algorithm, thus highlighting the value of the RT-snPCR assay and its modifications. The circulation and preponderance of EV-C in Nigeria was also confirmed.

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Acknowledgements

We thank the WHO National Polio Laboratory in Ibadan, Nigeria, for providing the anonymous samples analyzed in this study.

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Authors and Affiliations

  1. Department of Virology, College of Medicine, University of Ibadan, Ibadan, Oyo State, Nigeria

    J. A. Adeniji, A. O. Oragwa, U. E. George, U. I. Ibok, T. O. C. Faleye & M. O. Adewumi

  2. WHO National Polio Laboratory, University of Ibadan, Ibadan, Oyo State, Nigeria

    J. A. Adeniji

  3. Department of Veterinary Microbiology and Pathology, Faculty of Veterinary Medicine, University of Jos, Jos, Plateau State, Nigeria

    A. O. Oragwa

  4. Department of Microbiology, Faculty of Science, Ekiti State University, Ado-Ekiti, Ekiti State, Nigeria

    T. O. C. Faleye

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  1. J. A. Adeniji
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  2. A. O. Oragwa
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  3. U. E. George
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  4. U. I. Ibok
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  5. T. O. C. Faleye
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  6. M. O. Adewumi
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Contributions

1. Study design (JAA, MOA, TOCF). 2. Sample collection, laboratory and data analysis (all authors). 3. Wrote, revised, read and approved the final draft of the manuscript (all authors).

Corresponding author

Correspondence to M. O. Adewumi.

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Conflict of interest

The authors declare that no conflict of interest exists.

Ethical approval

There was no contact with human participants by any of the authors, and the article does not contain any information that can be used to associate the enterovirus types analyzed in this study to any individual.

Funding

This study was funded by contributions from the authors.

Electronic supplementary material

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705_2017_3466_MOESM1_ESM.docx

Supplementary Table 1: Estimation of sensitivity of the RD and L20B cell lines used in the WHO-accredited polio laboratory (polio lab) in Ibadan, Nigeria. These sensitivity data were generated following the method described in the guidelines for poliovirus surveillance (WHO, 2004) and is for the period (August 2015) during which the negative samples analyzed in this study were screened by the polio lab. Hence, these data were not generated by the authors of this study but were generously provided on retrospective request from the archives of the polio lab. (DOCX 11 kb)

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Adeniji, J.A., Oragwa, A.O., George, U.E. et al. Preponderance of enterovirus C in RD-L20B-cell-culture-negative stool samples from children diagnosed with acute flaccid paralysis in Nigeria. Arch Virol 162, 3089–3101 (2017). https://doi.org/10.1007/s00705-017-3466-2

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