Wildlife in Cameroon harbor diverse coronaviruses, including many closely related to human coronavirus 229E

  1. Nkom F Ntumvi
  2. Valantine Ngum Ndze
  3. Amethyst Gillis
  4. Joseph Le Doux Diffo
  5. Ubald Tamoufe
  6. Jean-Michel Takuo
  7. Moctar M M Mouiche
  8. Julius Nwobegahay
  9. Matthew LeBreton
  10. Anne W Rimoin
  11. Bradley S Schneider
  12. Corina Monagin
  13. David J McIver
  14. Sanjit Roy
  15. James A Ayukekbong
  16. Karen E Saylors
  17. Damien O Joly
  18. Nathan D Wolfe
  19. Edward M Rubin
  20. Christian E Lange

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Abstract

Zoonotic spillover of animal viruses into human populations is a continuous and increasing public health risk. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) highlights the global impact of emergence. Considering the history and diversity of coronaviruses (CoVs), especially in bats, SARS-CoV-2 will likely not be the last to spillover from animals into human populations. We sampled and tested wildlife in the Central African country Cameroon to determine which CoVs are circulating and how they relate to previously detected human and animal CoVs. We collected animal and ecological data at sampling locations and used family-level consensus PCR combined with amplicon sequencing for virus detection. Between 2003 and 2018, samples were collected from 6,580 animals of several different orders. CoV RNA was detected in 175 bats, a civet, and a shrew. The CoV RNAs detected in the bats represented 17 different genetic clusters, coinciding with alpha (n = 8) and beta (n = 9) CoVs. Sequences resembling human CoV-229E (HCoV-229E) were found in 40 Hipposideridae bats. Phylogenetic analyses place the human-derived HCoV-229E isolates closest to those from camels in terms of the S and N genes but closest to isolates from bats for the envelope, membrane, and RNA-dependent RNA polymerase genes. The CoV RNA positivity rate in bats varied significantly (P < 0.001) between the wet (8.2 per cent) and dry seasons (4.5 per cent). Most sampled species accordingly had a wet season high and dry season low, while for some the opposite was found. Eight of the suspected CoV species of which we detected RNA appear to be entirely novel CoV species, which suggests that CoV diversity in African wildlife is still rather poorly understood. The detection of multiple different variants of HCoV-229E-like viruses supports the bat reservoir hypothesis for this virus, with the phylogenetic results casting some doubt on camels as an intermediate host. The findings also support the previously proposed influence of ecological factors on CoV circulation, indicating a high level of underlying complexity to the viral ecology. These results indicate the importance of investing in surveillance activities among wild animals to detect all potential threats as well as sentinel surveillance among exposed humans to determine emerging threats.

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  1. Version published to 10.1093/ve/veab110
  2. ScreenIT

    SciScore for 10.1101/2021.09.03.458874: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    EthicsIACUC: Specimen collection was approved by an Institutional Animal Care and Use Committee (IACUC) of UC Davis (#16067, # 17803, and # 19300), JHU (ACR #2007-110-03, #2007-110-02 and #2007-110-11) and UCLA (Protocols # FS03M221 and FS06H205) and the Government of Cameroon.
    Field Sample Permit: Sample collection staff were trained in safe collection techniques in collaboration with representatives from Ministry of Fisheries Livestock Animal Industries (MINEPIA), the Ministry of Forestry and Wildlife (MINFOF), and the Ministry of Environment Nature Protection and Sustainable Development MINEPDED) and wore dedicated clothing, N95 masks, nitrile gloves, and protective eyewear during animal capture, handling, and sampling.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    Amplified DNA was extracted using either the QIAquick Gel Extraction Kit (Qiagen) or the Wizard SV Gel and PCR Cleanup System (Promega) and sent for commercial Sanger sequencing at either GATC, First Base or Macrogen.
    Macrogen
    suggested: (Macrogen, RRID:SCR_014454)
    All results from sequencing were analyzed in the Geneious 7.1 software, and primer trimmed consensus sequences compared to the GenBank database (BLAST N, NCBI).
    Geneious
    suggested: (Geneious, RRID:SCR_010519)
    , ClustalW Alignment).
    ClustalW
    suggested: (ClustalW, RRID:SCR_017277)
    Bayesian phylogeny of the polymerase gene fragment was inferred using MrBayes (version 3.2) with the following parameters: Datatype=DNA, Nucmodel=4by4, Nst=1, Coavion=No, # States=4, Rates=Equal, 2 runs, 4 chains of 10,000,000 generations.
    MrBayes
    suggested: (MrBayes, RRID:SCR_012067)
    The first 10% of the trees were discarded and the remaining ones combined using TreeAnnotator (version 2.5.1; http://beast.bio.ed.ac.uk) and displayed with FIGTREE (1.4.4; http://tree.bio.ed.ac.uk/) [Bouckaert 2019].
    http://beast.bio.ed.ac.uk
    suggested: (BEAST, RRID:SCR_010228)
    FIGTREE
    suggested: (FigTree, RRID:SCR_008515)
    All statistical data analyses were conducted using the statistical software SPSS (Statistical Package for the Social Sciences) version 26.
    SPSS
    suggested: (SPSS, RRID:SCR_002865)
    Statistical Package for the Social Sciences
    suggested: (SPSS, RRID:SCR_002865)

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • No funding statement was detected.
    • No protocol registration statement was detected.

    Results from scite Reference Check: We found no unreliable references.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  3. Version published to 10.1101/2021.09.03.458874v1 on bioRxiv