When One's Not Enough: Colony Pool-Seq Outperforms Individual-Based Methods for Assessing Introgression in Apis mellifera mellifera

Abstract

The human management of honey bees (Apis mellifera) has resulted in the widespread introduction of subspecies outside of their native ranges. One well known example of this is Apis mellifera mellifera, native to Northern Europe, which has now been significantly introgressed by the introduction of C lineage honey bees. Introgression has consequences for species in terms of future adaptive potential and long-term viability. However, estimating introgression in colony-living haplodiploid species is challenging. Previous studies have estimated introgression using individual workers, individual drones, multiple drones, and pooled workers. Here, we compare introgression estimates via three genetic approaches: SNP array, individual RAD-seq, and pooled colony RAD-seq. We also compare two statistical approaches: a maximum likelihood cluster program (ADMIXTURE) and an incomplete lineage sorting model (ABBA BABA). Overall, individual approaches resulted in lower introgression estimates than pooled colonies when using ADMIXTURE. However, the pooled colony ABBA BABA approach resulted in generally lower introgression estimates than all three ADMIXTURE estimates. These results highlight that sometimes one individual is not enough to assess colony-level introgression, and future studies that do use colony pools should not be solely dependent on clustering programs for introgression estimates.

Keywords: ABBA BABA; ADMIXTURE; Apis mellifera; RAD-seq; SNP array; colony; introgression; pool-seq.

Figure 1

Overview of South West England RAD-seq samples and subspecies standards bioinformatics workflows. Samples were processed based on the sampling approach.

Figure 2

The principles of the ABBA BABA approach. Allele patterns of derived (“B”) and ancestral (“A”) alleles across the P1, P2, P3, and Po (outgroup) groups are illustrated. The ABBA pattern is when P2 and P3 share the derived allele, while P1 has the ancestral allele from the outgroup. The BABA pattern is when P1 and P3 share the derived allele, while P2 shares the ancestral allele with the outgroup. Here, pooled colonies were tested using A. m. mellifera as P1 (Mel), South West England as P2 (SW), and a c-lineage (C) colony in the P3 position, and the outgroup Po, represented by A. cerana (O).

Figure 3

The f statistic compares the ABBA BABA counts to a scenario of complete admixture (b). Here, to simulate complete admixture, the P2 population (a) has been replaced with another P3 population, labelled P3a (b).

Figure 4

ADMIXTURE for K = 2 of honey bees in the South West of England (labelled SW) using (A) SNP array, (B) individual RAD-seq and (C) pooled colony RAD-seq. The blue cluster represents the C-lineage honey bees, A. m. carnica and A. m. ligustica (labelled Car and Lig, respectively), the red cluster represents A. m. mellifera (Mel). (D) Comparison of RAD-seq Q values to SNP array Q values for membership to the C lineage cluster, including an R² value (denoted as R^2).

Figure 5

C-Lineage proportion estimated by ADMIXTURE at K = 2 using the SNP array (80 SNPs); individual ddRAD-seq (23,916 SNPs) and pooled colony RAD-seq (158,496 SNPs) across the 17 samples examined in all three methods.

Figure 6

Average f statistics (proportion of admixture) calculated for pooled colonies. Trios tested for introgression between the South West and two different C-lineage subspecies. Shown here are trios consisting of A. m. mellifera; South West; A. m. carnica (Orange circle), and A. m. mellifera; South West; A. m. ligustica (blue triangle). A red X indicates colonies that had a non-significant result. Error bars represent the standard deviation of f for each South West colony estimate, calculated with every combination of A. m. mellifera and A. m. carnica or A. m. ligustica colony.