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. 2023 Sep 13:12:e66550.
doi: 10.7554/eLife.66550.

Mosquito community composition shapes virus prevalence patterns along anthropogenic disturbance gradients

Kyra Hermanns  1 , Marco Marklewitz  1 , Florian Zirkel  2 , Anne Kopp  1 , Stephanie Kramer-Schadt  3   4 , Sandra Junglen  1
Affiliations

Mosquito community composition shapes virus prevalence patterns along anthropogenic disturbance gradients

Kyra Hermanns et al. Elife. .

Abstract

Previously unknown pathogens often emerge from primary ecosystems, but there is little knowledge on the mechanisms of emergence. Most studies analyzing the influence of land-use change on pathogen emergence focus on a single host-pathogen system and often observe contradictory effects. Here, we studied virus diversity and prevalence patterns in natural and disturbed ecosystems using a multi-host and multi-taxa approach. Mosquitoes sampled along a disturbance gradient in Côte d'Ivoire were tested by generic RT-PCR assays established for all major arbovirus and insect-specific virus taxa including novel viruses previously discovered in these samples based on cell culture isolates enabling an unbiased and comprehensive approach. The taxonomic composition of detected viruses was characterized and viral infection rates according to habitat and host were analyzed. We detected 331 viral sequences pertaining to 34 novel and 15 previously identified viruses of the families Flavi-, Rhabdo-, Reo-, Toga-, Mesoni- and Iflaviridae and the order Bunyavirales. Highest host and virus diversity was observed in pristine and intermediately disturbed habitats. The majority of the 49 viruses was detected with low prevalence. However, nine viruses were found frequently across different habitats of which five viruses increased in prevalence towards disturbed habitats, in congruence with the dilution effect hypothesis. These viruses were mainly associated with one specific mosquito species (Culex nebulosus), which increased in relative abundance from pristine (3%) to disturbed habitats (38%). Interestingly, the observed increased prevalence of these five viruses in disturbed habitats was not caused by higher host infection rates but by increased host abundance, an effect tentatively named abundance effect. Our data show that host species composition is critical for virus abundance. Environmental changes that lead to an uneven host community composition and to more individuals of a single species are a key driver of virus emergence.

Keywords: arbovirus; community composition; disease ecology; ecology; epidemiology; global health; insect-specific virus; mosquito; viruses.

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Conflict of interest statement

KH, MM, FZ, AK, SK, SJ No competing interests declared

Figures

Figure 1.
Figure 1.. Study sites and mosquito distribution.
(A) Study site location and overview over the five habitat types. The different habitat types along the anthropogenic disturbance gradient are depicted by photos and drawings. (B) Alluvial plot showing the distribution of the main mosquito species groups and main virus families to the five respective habitats. CulAnn: Culex annulioris; CulDec: Culex decens; CulNeb: Culex nebulosus; Cul_spec: other Culex species; Ano_spec: Anopheles species; Ura_spec: Uranotaenia species; Coq_spec: Coquillettidia species; others: all other grouped species (see main text). Bunya: order Bunyavirales containing Phenuiviridae, Peribunyaviridae, and Phasmaviridae; Flavi: Flaviviridae; Toga: Togaviridae; Rhabdo: Rhabdoviridae; Reo: Reoviridae; Iflavi: Iflaviridae; Meso: Mesoniviridae.
Figure 1—figure supplement 1.
Figure 1—figure supplement 1.. Biodiversity analyses.
Mosquito species richness per habitat. Rarefaction curves (with 95% confidence intervals) show the observed (solid lines) and interpolated (dotted line) mosquito diversity in the five habitats.
Figure 1—figure supplement 2.
Figure 1—figure supplement 2.. Hierarchical cluster analysis based on associations between mosquito species. Main mosquito groups were identified as CulAnn, CulDec, CulNeb, Cul_spec, Ura_spec, Ano_spec, and Coq_spec. CulAnn: Culex annulioris; CulDec: Culex decens; CulNeb: Culex nebulosus; Cul_spec: other Culex species; Ano_spec: Anopheles species; Ura_spec: Uranotaenia species; Coq_spec: Coquillettidia species, others: all other grouped species. PF: primary forest; SF: secondary forest; A: agriculture; C: camp; V: village.
Figure 2.
Figure 2.. Phylogenetic analyses of detected bunyaviruses.
Phylogenetic trees were inferred with PhyML (LG substitution model) based on MAFFT-E protein alignments covering the conserved RdRp motifs of the families Phenuiviridae (A), Peribunyaviridae (B), and Phasmaviridae (C). The alignment length was approximately 290, 300, and 690 amino acids, respectively. Novel viruses from this study are indicated in red, and previously published viruses detected in our data set are indicated in blue. Sample origin from the different habitat types is indicated by colored circles while no detection is indicated by gray circles. Live virus isolates are marked with a blue virion. PF: primary forest; SF: secondary forest; A: agriculture; C: camp; V: village.
Figure 2—figure supplement 1.
Figure 2—figure supplement 1.. Phylogenetic analyses of detected phasmaviruses.
Phylogenetic trees were inferred with PhyML (WAG (A) and LG (B) substitution model) based on MAFFT-E protein alignments covering the glycoproteins (A) and nucleocapsid (B) proteins of the family Phasmaviridae. The alignment length was approximately 1700 and 280 amino acids, respectively. Novel viruses from this study are indicated in red, and previously published viruses detected in our data set are indicated in blue.
Figure 3.
Figure 3.. Phylogenetic analyses of detected rhabdoviruses and iflaviruses.
Phylogenetic trees were inferred with PhyML (LG substitution model) based on MAFFT-E protein alignments covering the conserved RdRp motifs of the families Rhabdoviridae (A) and Iflaviridae (B). The alignment length was approximately 270 and 250 amino acids, respectively. Novel viruses from this study are indicated in red, and detected virus-like sequences are indicated in gray. Sample origin from the different habitat types is indicated by colored circles while no detection is indicated by gray circles. Live virus isolates are marked with a blue virion. PF: primary forest; SF: secondary forest; A: agriculture; C: camp: V: village.
Figure 4.
Figure 4.. Phylogenetic analyses of detected flaviviruses and orbiviruses.
Phylogenetic trees were inferred with PhyML (GTR substitution model) based on MAFFT-E nucleotide alignments covering the conserved RdRp motifs of the genus Flavivirus (A) and with PhyML (LG substitution model) based on a MAFFT-E protein alignment of the polymerase of the genus Orbivirus (B). The alignment length was approximately 1170 nucleotides and 1190 amino acids, respectively. Novel viruses from this study are indicated in red, previously published viruses detected in our data set are indicated in blue, and detected virus-like sequences are indicated in gray. Sample origin from the different habitat types is indicated by colored circles while no detection is indicated by gray circles. Live virus isolates are marked with a blue virion. PF: primary forest; SF: secondary forest; A: agriculture; C: camp; V: village.
Figure 4—figure supplement 1.
Figure 4—figure supplement 1.. Phylogenetic analyses of the detected Wanken orbivirus (WKOV) and members of the genus Orbivirus.
Phylogenetic trees were inferred with PhyML (LG substitution model) based on MAFFT-E protein alignments of VP3 (A), VP4 (B), VP7 (C), and VP5 (D). The alignment length was approximately 890, 640, 350, and 510 amino acids, respectively. WKOV is indicated in red.
Figure 4—figure supplement 2.
Figure 4—figure supplement 2.. Phylogenetic analyses of detected reoviruses, alphaviruses, and mesoniviruses.
Phylogenetic trees were inferred with PhyML (LG substitution model) based on a MAFFT-E protein alignment of the polymerase of members of the family Reoviridae (A) or with PhyML (GTR substitution model) based on MAFFT-E nucleotide alignments covering the E2-6K-E1 region of members of the genus Alphavirus (B) of the polymerase of members of the family Mesoniviridae (C). The alignment length was approximately 290 amino acids and 2720 and 7430 nucleotides, respectively. Novel viruses from this study are indicated in red, previously published viruses detected in our data set are indicated in blue, and detected virus-like sequences are indicated in gray. Sample origin from the different habitat types is indicated by colored circles while no detection is indicated by gray circles. Live virus isolates are marked with a blue virion. PF: primary forest; SF: secondary forest; A: agriculture; C: camp; V: village.
Figure 4—figure supplement 3.
Figure 4—figure supplement 3.. Potential non-retroviral integrated RNA virus sequences (NIRVS).
(A) PCR amplicons of the generic rhabdovirus PCR assay. RNA/DNA extracts without reverse transcription were used for the PCR. Nested PCR amplicons of Cimo rhabdovirus I and two rhabdovirus-like NIRVS were visualized by ethidium bromide-stained agarose gel electrophoresis. Amplicons with the expected size of 260 bp are framed by a blue box. (B) Schematic representation of selected flavivirus-like NIRVS. Stop codons are indicated by an asterisk, deletions are shown as light gray boxes, frame shifts are indicated by overlapping blue boxes, and insertions with similarity to insect genes are shown as waved lines.
Figure 5.
Figure 5.. Temperature-dependent replication of novel virus isolates.
C6/36 cells were infected with an MOI of 0.1 with Sefomo virus (A), Mikado virus (B), Sassandra virus (C), and Tafomo virus (D). Replication was measured for 3 dpi at 28, 30, 32, and 34°C.
Figure 6.
Figure 6.. Spearman’s rank correlation rho for the most abundant viruses.
Only significant correlations are shown. AnthroDist refers to the gradient of anthropogenic disturbance from low to high. PF: primary forest; SF: secondary forest; A: agriculture; C: camp; V: village.
Figure 6—figure supplement 1.
Figure 6—figure supplement 1.. Ordination plot of the principal coordinate analysis (PCoA) showing that the viral community primarily partitions by Culex nebulosus (CulNeb), Uranotaenia species (Ura_spec), and Culex decens (CulDec).
Figure 6—figure supplement 2.
Figure 6—figure supplement 2.. Ordination plot of the principle coordinate analysis (PCoA) showing that the viral community primarily partitions by Culex nebulosus (CulNeb), Uranotaenia species (Ura_spec) and Culex decens (CulDec) as shown in Figure 6—figure supplement 1, which can be related to their main habitats in primary forest, agriculture and villages.
CulAnn: Culex annulioris, Cul_spec: other Culex species; Ano_spec: Anopheles species; Ura_spec: Uranotaenia species.
Figure 7.
Figure 7.. Richness and cumulative minimum infection rate (MIR) across all tested virus taxa.
The number of distinct viruses (A) and the cumulated MIR per 1000 mosquitoes (B) were calculated for all habitat types and for the complete data set. Different virus taxa are shown in different colors. PF: primary forest; SF: secondary forest; A: agriculture; C: camp; V: village.
Figure 7—figure supplement 1.
Figure 7—figure supplement 1.. Cumulative minimum infection rate (MIR) per virus taxon.
The cumulated MIR per 1000 mosquitoes was calculated for the analyzed taxa Phenuiviridae (A), Phasmaviridae (B), Reoviridae (C), Alphavirus (D), Flavivirus (E), Peribunyaviridae (F), Rhabdoviridae (G), Mesoniviridae (H), and Iflaviridae (I), as well as for all detected viruses (J) in the different habitat types and for the complete data set. The different viruses or taxa are shown in different colors.
Figure 8.
Figure 8.. Prevalence patterns of selected viruses along the disturbance gradient.
For all viruses that were detected in >10 pools, Gouléako virus (GOLV) (A), Herbert virus (HEBV) (B), Ferak virus (FERV) (C), Cimo rhabdovirus I (D), Cavally virus (CAVV) (E), Cimo phenuivirus II (F), Jonchet virus (G), Spilikins virus (H), and Cimo flavivirus I (I), the minimum infection rate (MIR) and maximum likelihood estimation (MLE) per 1000 mosquitoes of the whole data set were calculated for all habitat types (left graphs for A–E). The abundance of the main mosquito host species was plotted. The five viruses GOLV (A), HEBV (B), FERV (C), Cimo rhabdovirus I (D), and CAVV (E) occurred frequently enough in their main mosquito host species (>10 positive pools) to analyze their prevalence in these species. For these viruses, the MIR and MLE per 1000 mosquitoes of the respective species were calculated for all habitat types (right graphs). Significant differences in the infection probability with the most abundant viruses in the different habitats are shown in Supplementary file 2.
Figure 9.
Figure 9.. Schematic presentation of the abundance effect.
Infection rates are shown for Gouléako virus (GOLV) in all sampled mosquitoes (representing minimum infection rate (MIR) and maximum likelihood estimation (MLE) values as shown in Figure 8) and only in the main mosquito host species, Culex nebulosus. The abundance of the main mosquito host species, Culex nebulosus is indicated by a gray line.
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