Persistent anthrax as a major driver of wildlife mortality in a tropical rainforest

Abstract

Anthrax is a globally important animal disease and zoonosis. Despite this, our current knowledge of anthrax ecology is largely limited to arid ecosystems, where outbreaks are most commonly reported1,2,3. Here we show that the dynamics of an anthrax-causing agent, Bacillus cereus biovar anthracis, in a tropical rainforest have severe consequences for local wildlife communities. Using data and samples collected over three decades, we show that rainforest anthrax is a persistent and widespread cause of death for a broad range of mammalian hosts. We predict that this pathogen will accelerate the decline and possibly result in the extirpation of local chimpanzee (Pan troglodytes verus) populations. We present the epidemiology of a cryptic pathogen and show that its presence has important implications for conservation.

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Figure 1: Bcbva occurrence and study sampling sites in sub-Saharan Africa.
Figure 2: Bcbva cases in TNP.
Figure 3: Phylogenomic tree of Bcbva isolates.
Figure 4: Proportions of simulated chimpanzee communities surviving 150 years with and without presence of anthrax.

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Acknowledgements

We thank the authorities in Côte d’Ivoire for long-term support, especially the Ministry of the Environment and Forests, the Ministry of Research, the directorship of the TNP and the CSRS in Abidjan; and national authorities from all other countries for providing permissions for our research (MINFoF, MINRESI, the Service de la Conservation de la Réserve du Dja, Cameroon, in Central African Republic; the Ministère des Eaux et Fôrets, Chasse et Peche and the Ministère de l’Education Nationale, de l’Alphabetisation, de l’Enseignement Superieur, et de la Recherche, the Agence Nationale des Parcs Nationaux, Gabon; Centre National de la Recherche Scientifique et Technologique, Gabon; Direction des Eaux, Forêts et Chasses, Senegal; Forestry Development Authority, Liberia; Institut Congolais pour la Conservation de la Nature, Democratic Republic of the Congo; Ministère de l’Agriculture de l’Elevage et des Eaux et Forêts, Guinea; Instituto da Biodiversidade e das Áreas Protegidas (IBAP), Guinea-Bissau; Ministère de la Recherche Scientifique, Democratic Republic of the Congo; Ministère de le Recherche Scientifique et Technologique, Democratic Republic of the Congo; Nigeria National Park Service, Nigeria, Uganda National Council for Science and Technology, Ugandan Wildlife Authority, Uganda). We thank the WWF Central African Republic, T. Börding, T. Hicks, Y. Moebius, V. Sommer, K. Zuberbühler and M. Peeters for their logistical support; the field assistants A. Henlin, K. Albrechtova and A. Lang for the collection of samples in TNP; and the field assistants from all other sites for their support; S. Becker, T. Franz, S. Howaldt, A. Lander, P. Lochau, H. Nattermann and A. Schneider for the laboratory work; J. Hinzmann, A. Nitsche and J. Tesch for sequencing; P. Wojciech Dabrowski and T. Semmler from RKI, as well as G. Hamilton at Glasgow Polyomics, for bioinformatic support; and M. Kovacev-Wegener for administrative support. We thank the German Research Council DFG KL 2521/1-1 and the Sonnenfeld-Stiftung for funding; and the Max-Planck-Society and Krekeler Foundation for funding of the Pan African Programme.

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C.H., F.Z., A.A., S.A., M.A., G.B., K.C., T.D., P.D., K.D., H.E., P.F., Y.G.Y., A.G., A.-C.G., S.McG., J.H., S.J., J.J., J.K., K.La., J.L., K.Le., F.L., V.L., T.L., S.Ma., A.M., S.Me., M.M., J.v.S., E.T. and D.W. collected flies, bones and associated field data. Necropsies on wildlife that was found dead were performed by F.Z., K.N., A.B., E.C.-H., A.D., P.F., S.A.L., T.L., S.Me., S.N., H.D.N. and F.H.L. and laboratory analyses were performed by C.H., F.Z., K.N., S.D., R.G., K.M.-R., K.M., S.Me., H.D.N., A.S., U.T., S.R.K., L.H.W., S.C.-S. and F.H.L. The data were analysed by C.H., F.Z., R.B., H.K., R.M. and S.C.-S. and the manuscript was prepared by C.H., F.Z., R.B., H.K., R.M., J.F.G., S.C.-S. and F.H.L. The manuscript was revised and approved by all authors. The study was supervised by C.B., R.M.W., S.C.-S. and F.H.L.

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Correspondence to Fabian H. Leendertz.

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Extended data figures and tables

Extended Data Figure 1 Necropsies performed since 1996.

The total number of necropsies performed per year in TNP from 1996 to 2015. Grey bars indicate the number of Bcbva-positive necropsies and are annotated with the associated proportion. In the years 2003 and 2010 only limited veterinary service was available at TNP owing to political insecurity in the region.

Extended Data Figure 2 Geographic location of Bcbva-positive carcasses in TNP.

Necropsies that tested Bcbva-positive in TNP since 2001. GPS data was available for 70 of all necropsies that tested positive (n = 81).

Extended Data Figure 3 Effect of mammalian DNA content on anthrax positivity in flies.

Shown is the probability of Bcbva positivity (PA) as a function of the amount of mammalian DNA (copies) found in a fly. The amount of mammal DNA was binned (bin width of 0.25) and the area of the points depicts the number of flies (range, 1–206) in the respective bins. The dashed line indicates the fitted model and the dotted lines the 95% confidence interval.

Extended Data Figure 4 Effect of season on anthrax positivity in flies.

The probability of Bcbva positivity (PA) over the course of a year (binned in 10-day periods) is shown. The area of the points depicts the number of flies in the respective 10-day period. The dashed line indicates the fitted model and the dotted lines the 95% confidence interval.

Extended Data Figure 5 Maximum clade credibility tree based on chromosomal sequences of Bcbva isolates from TNP (Côte d’Ivoire, n = 124) and Grebo (Liberia, n = 2).

One sequence per host (mammals or flies, two divergent isolates for fly 600) was included and the final alignment of variant sites measured 298 bp. The size of the nodes represents posterior probability values. The location of the root received a posterior probability of 1.

Extended Data Figure 6 Maximum likelihood tree for sub-Saharan Bcbva strains.

Maximum likelihood tree based on chromosomal sequences of Bcbva strains from Côte d’Ivoire, Cameroon, Central African Republic and Liberia. The alignment of variant sites measured 1,016 bp. Bootstrap values are shown above the branches and the scale bar indicates substitution per chromosomal site. The tree was rooted using TempEst version 1.5.

Extended Data Figure 7 Fly snapshot sampling scheme.

For the fly snapshot, flies were caught following a 2 × 2-km grid system within and outside the research area within 19 days. In total 908 snapshot flies were analysed.

Extended Data Figure 8 Genetic and geographic distances of Bcbva isolates from the fly snapshot.

a, Maximum likelihood tree based on chromosomal sequences of Bcbva isolates from the 19-day fly snapshot. Each dot represents one fly isolate. Colours were chosen to illustrate the distribution of genetically clustering isolates on the map presented in b. The final alignment of variant sites measured 123 bp. Bootstrap values are shown above all internal branches. The tree was rooted using the ‘best-fit’ option in Path-O-Gen version 1.2. The scale bar shows substitutions per site. b, Geographic origin of Bcbva isolates collected during the fly snapshot. Colours correspond to maximum likelihood tree in a. Large circles represent two isolates.

Extended Data Figure 9 Box plot of genetic and mean geographic distances.

Bcbva isolates from TNP were binned by relative genetic distance (bin size = 0.03, approximately 2.5 SNPs).The two most genetically distant isolates received a value of 1 and all other distances were scaled accordingly. Diamonds indicate the geographic distance means of the groups. To examine variation within genetic lineages, we analysed isolates with low genetic distance (maximum relative genetic distance <0.5, marked in blue) and their mean geographic distance. For low genomic distances, the linear regression of geographic distances on genetic distances has an R2 of 0.72 and a slope coefficient that differs significantly from zero (Student’s t-test, P = 4 × 10−5).

Extended Data Figure 10 Fly species composition based on generalized mixed Yule-coalescent model (GMYC) analysis.

ac, Fly species composition for three sites with known Bcbva occurrence: TNP, Côte d’Ivoire (a); Dja Faunal Reserve, Cameroon (b); Dzanga-Sangha Protected Areas, Central African Republic (c). The proportions of flies per site (%) belonging to a single fly species identified with GMYC models are shown. Different colours indicate different taxonomic fly families.

Supplementary information

Supplementary Information

This file contains a detailed method section as well as additional tables (Tables S1-10) and figures (Fig. S1-8). (PDF 2638 kb)

Supplementary Table 1

This file contains results that were derived from the analyses of flies caught in TNP analyzed in this study. The file includes results from PCR and culture as well as flymeal analysis results for a selection of flies. (XLSX 148 kb)

Supplementary Table 2

This file contains results of fly meal analysis with taxonomic assignment at genus level. The file provides the number of sequences per amplicon assigned at genus level. (XLSX 97 kb)

Supplementary Table 3

This file contains results of fly meal analysis with taxonomic assignment at order level. The file provides the number of sequences per amplicon assigned at order level. (XLSX 24 kb)

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Hoffmann, C., Zimmermann, F., Biek, R. et al. Persistent anthrax as a major driver of wildlife mortality in a tropical rainforest. Nature 548, 82–86 (2017). https://doi.org/10.1038/nature23309

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