Geographic population structure of the honeybee microsporidian parasite Vairimorpha (Nosema) ceranae in the South West Indian Ocean

Abstract

The microsporidian Vairimorpha (Nosema) ceranae is one of the most common parasites of the honeybee. A single honeybee carries many parasites and therefore multiple alleles of V. ceranae genes that seem to be ubiquitous. As a consequence, nucleotide diversity analyses have not allowed discriminating genetic structure of parasite populations. We performed deep loci-targeted sequencing to monitor the haplotype frequencies of genome markers in isolates from discontinuous territories, namely the tropical islands of the South West Indian Ocean. The haplotype frequency distribution corroborated the suspected tetraploidy of the parasite. Most major haplotypes were ubiquitous in the area but with variable frequency. While oceanic isolates differed from European and Asian outgroups, parasite populations from distinct archipelagoes also differed in their haplotype distribution. Interestingly an original and very divergent Malagasy isolate was detected. The observed population structure allowed formulating hypotheses upon the natural history of V. ceranae in this oceanic area. We also discussed the usefulness of allelic distribution assessment, using multiple informative loci or genome-wide analyses, when parasite population is not clonal within a single host.

Figure 1

Map of the South West Indian Ocean (SWIO) showing the studied islands. Color-codes indicate the grouping of isolates throughout the forthcoming figures and tables.

Figure 2

Distribution of haplotype frequencies, showing the number of values with a frequency from 0.01 to 1, with increasing intervals of 0,01 (n = 10,942 values). Haplotypes whose frequency was below and above 12% were classified as “rare” (yellow) or “major” (blue), respectively. Peaks at frequency close to 0.25, 0.5 and 0.75 are indicated by stars.

Figure 3

Cumulated frequencies of the 636 haplotypes from 83 markers in isolates (a) and their distribution in selected categories (b). Since markers were independently studied, the cumulative frequency that equaled 1 for each marker reached 83 over the dataset. A haplotype was considered present with a frequency higher than 1% in one isolate and ubiquitous with a frequency higher than 1% in all isolates. Haplotypes with a frequency higher than 12% in at least one isolate were considered as “major”. Haplotypes with a frequency higher than 1% in at least one isolate but never higher than 12% were considered as “rare”. If rare haplotypes were numerous (b), their cumulated frequency was low (a). Ubiquitous major haplotypes appear in blue, including those that are frequent in all isolates (dark blue). Shared major haplotypes, defined as present in at least two groups, appear in green. Isolate-specific major haplotypes are in purple. Ubiquitous rare haplotypes are in yellow. Shared rare haplotypes in orange. SWIO-specific, group-specific and isolate-specific rare haplotypes are in dark red, red and pink respectively. There was no SWIO-specific or group-specific major haplotype.

Figure 4

Principal component analyses (PCAs) of the N. ceranae haplotypes frequencies in A. mellifera isolates from the South West Indian Ocean (SWIO). Honeybees were sampled in La Réunion (blue), Mauritius and Rodrigues (green), the Comoros archipelago (orange), the Seychelles archipelago (pink) and Madagascar (red). Outgroup from Europe (grey) and from A. ceranae and A. dorsata in Thailand (black) were included. For details on the isolates, see Table 1. (a) PCA of all the 636 haplotypes, whatever their frequency, representing 83 coding and non-coding genetic markers from the 7 considered groups. The three first components represented 52.7% of the variance. (b) PCA (53.7% of the variance represented) of the 636 haplotypes from the SWIO isolates, i.e. without the outgroups. (c) PCA (53.7% of the variance) of the 305 major haplotypes from the SWIO isolates. (d) PCA (57.3% of the variance) of the 274 haplotypes, whatever their frequency, representing 43 coding markers from the SWIO isolates. (e) PCA (56.1% of the variance) of the 362 haplotypes, whatever their frequency, representing 29 non-coding markers from the SWIO isolates. (f) PCA (68.4% of the variance) of the 142 haplotypes, whatever their frequency, representing 11 microsatellites markers from the SWIO isolates.