Cytonuclear Epistasis Controls the Density of Symbiont Wolbachia pipientis in Nongonadal Tissues of Mosquito Culex quinquefasciatus

Abstract

Wolbachia pipientis, a bacterial symbiont infecting arthropods and nematodes, is vertically transmitted through the female germline and manipulates its host's reproduction to favor infected females. Wolbachia also infects somatic tissues where it can cause nonreproductive phenotypes in its host, including resistance to viral pathogens. Wolbachia-mediated phenotypes are strongly associated with the density of Wolbachia in host tissues. Little is known, however, about how Wolbachia density is regulated in native or heterologous hosts. Here, we measure the broad-sense heritability of Wolbachia density among families in field populations of the mosquito Culex pipiens, and show that densities in ovary and nongonadal tissues of females in the same family are not correlated, suggesting that Wolbachia density is determined by distinct mechanisms in the two tissues. Using introgression analysis between two different strains of the closely related species C. quinquefasciatus, we show that Wolbachia densities in ovary tissues are determined primarily by cytoplasmic genotype, while densities in nongonadal tissues are determined by both cytoplasmic and nuclear genotypes and their epistatic interactions. Quantitative-trait-locus mapping identified two major-effect quantitative-trait loci in the C. quinquefasciatus genome explaining a combined 23% of variance in Wolbachia density, specifically in nongonadal tissues. A better understanding of how Wolbachia density is regulated will provide insights into how Wolbachia density can vary spatiotemporally in insect populations, leading to changes in Wolbachia-mediated phenotypes such as viral pathogen resistance.

Keywords: Culex quinquefasciatus; QTL mapping; Wolbachia density; Wolbachia pipientis; cytonuclear epistasis.

Figure 1

Heritability analysis of C. pipiens. Rank ordering of field-collected C. pipiens families by mean Wolbachia density in (A) whole females, with most (>99%) of the measured Wolbachia from the ovary, and (B) ovariectomized females, with the measured Wolbachia from nongonadal tissues. The mean (±SE) for six siblings from each family is shown. Levels of broad-sense heritability are shown with SE estimates. (C) Pearson’s correlation test of Wolbachia densities in ovary vs. nongonadal tissues. Data used for calculations were originally reported in Micieli and Glaser (2014).

Figure 2

Introgression analysis of C. quinquefasciatus. Reciprocal introgression backcrosses were performed between the Ben95 (Ben) and Arg12 (Arg) strains of C. quinquefasciatus. Wolbachia densities were measured in (A) whole females, with most (>99%) of the measured Wolbachia from the ovary, and (B) ovariectomized females, with the measured Wolbachia from nongonadal tissues. Female mosquitoes containing either Ben95-derived (•) or Arg12-derived (×) cytoplasmic genotypes are indicated. Measurements were made on mosquitoes with the parental, F1, and F5 nuclear genotypic background, as indicated at the top of (A). The cytoplasmic and nuclear genotypes for the mosquitoes collected for each sample are indicated across the bottom of (B). Means are indicated by a horizontal line.

Figure 3

C. quinquefasciatus genetic linkage maps. The SNP-based linkage maps reported here (right) are compared to microsatellite-based linkage maps reported previously (left)(Hickner et al. 2013). Lines between the maps connect markers located on the same genomic scaffolds shared between the two maps. Total chromosome lengths in centimorgan and the positions of QTL 1 and QTL 2 are indicated.

Figure 4

A QTL scan for Wolbachia density in nongonadal tissues. The genome was scanned for QTL using the SNP genotypes and Wolbachia-density measurements made for 91 F2 females from the mapping population. (A) Two major-effect QTL were identified on LG 2. The 0.1 (red) and 0.05 (blue) whole-genome significance thresholds are shown. (B) Genetic effect analysis with means and SEs shown for Wolbachia density in nongonadal tissue measured in mosquitoes homozygous and heterozygous for QTL alleles from the indicated parental strain.