Mitochondrial heteroplasmy and the evolution of insecticide resistance: non-Mendelian inheritance in action

Abstract

Genes encoded by mitochondrial DNA (mtDNA) exist in large numbers per cell but can be selected very rapidly as a result of unequal partitioning of mtDNA between germ cells during embryogenesis. However, empirical studies of this "bottlenecking" effect are rare because of the apparent scarcity of heteroplasmic individuals possessing more than one mtDNA haplotype. Here, we report an example of insecticide resistance in an arthropod pest (Tetranychus urticae) being controlled by mtDNA and on its inheritance in a heteroplasmic mite strain. Resistance to the insecticide bifenazate is highly correlated with remarkable mutations in cytochrome b, a mitochondrially encoded protein in the respiratory pathway. Four sites in the Q(o) site that are absolutely conserved across fungi, protozoa, plants, and animals are mutated in resistant mite strains. Despite the unusual nature of these mutations, resistant mites showed no fitness costs in the absence of insecticide. Partially resistant strains, consisting of heteroplasmic individuals, transmit their resistant and susceptible haplotypes to progeny in highly variable ratios consistent with a sampling bottleneck of approximately 180 copies. Insecticide selection on heteroplasmic individuals favors those carrying resistant haplotypes at a frequency of 60% or more. This combination of factors enables very rapid evolution and accounts for mutations being fixed in most field-collected resistant strains. The results provide a rare insight into non-Mendelian mechanisms of mitochondrial inheritance and evolution, relevant to anticipating and understanding the development of other mitochondrially encoded adaptations in arthropods. They also provide strong evidence of cytochrome b being the target site for bifenazate in spider mites.

Fig. 1.

Cytb resistance mutations in the mitochondrial genome. (A) Linear representation of the mitochondrial genome of T. urticae (13,103 bp) showing the position and orientation of the protein-encoding genes (n = 13) and the ribosomal RNA genes (n = 2). (B) Sequence alignment of conserved Q_o pocket residues positioned on the cytochrome b of T. urticae with those of S. cerevisiae (ABS28693), P. falciparum (NP_059668), Venturia inaequalis (AAC03553), Arabidopsis thaliana (CAA47966), Drosophila melanogaster (CAB91062), Gallus gallus (AAO44995), and Homo sapiens (AAX15094). Fully conserved residues in the alignment are marked in black. Point mutations linked to bifenazate resistance in T. urticae are indicated by triangles.

Fig. 2.

Inheritance of mtDNA heteroplasmy for the P262T mutation in the HOL3 strain of T. urticae. Frequencies of resistant haplotypes are shown as corresponding relative peak heights of nucleotides on forward and reverse strands. (A) Frequency of resistant haplotypes in 25 randomly selected individual mites on collection from the field. (B–F) Frequency of resistant haplotypes in single mothers (shown by arrows) and in eight of their first-generation offspring. Dotted lines show the estimated mutation frequency (60%) needed to survive field-applied concentrations of bifenazate.

Fig. 3.

Establishing a link between phenotype and genotype. The relationship between frequency of the resistant haplotype in single founder females from the HOL3 strain of T. urticae and susceptibility of third-generation siblings to 100 mg of active ingredient (a.i.) per liter of bifenazate is shown.