Telomere-to-telomere genome assembly uncovers Wolbachia-driven recurrent male bottleneck effect and selection in a sawfly

Abstract

Wolbachia, a widespread endosymbiotic bacterium, profoundly impacts insect hosts by distorting reproduction and population dynamics. Despite extensive laboratory research, its long-term effects on host evolution in nature remain poorly understood, especially the genomic consequences linked to disruptions in sex determination and reproductive processes. We present the first telomere-to-telomere (T2T) genome assembly of the sawfly Analcellicampa danfengensis and the complete genome of its symbiotic Wolbachia. Comparative population genomics across six Analcellicampa species revealed that Wolbachia-infected populations show starkly different demographic signals. While uninfected populations show similar demographic signals for both sexes, infected populations exhibit a lower apparent effective population size (Ne) in males, which may reflect a recurrent male bottleneck effect driven by Wolbachia-induced male scarcity. Genomic scans identified positively selected genes associated with reproductive functions, sensory perception, neural development, and longevity, suggesting that Wolbachia likely manipulates critical host pathways to promote its transmission. These findings provide direct genomic insights into Wolbachia as an evolutionary force, highlighting specific host genes and regions under selection resulting from these altered evolutionary dynamics. This work provides deeper insights into host-endosymbiont coevolution and has important implications for evolutionary theory and pest management strategies.

Fig. 1. Assembly and key characterization of the A.danfengensis T2T genome, including centromeric characterization, symbiotic Wolbachia and mitochondrial DNA.

a Genome-wide Hi-C contact matrix of A.danfengensis. Red color intensity in the heatmap shows frequency of interaction between two loci at 25 kb resolution. b Distribution of genomic features of the A.danfengensis genome. Tracks are aggregated in 100-kb bins as follows: I, gene density; II, TE density; III, TR density; IV, GC content; V, SNPs density; VI, the 9 chromosomes (chr1–chr9). c BUSCO completeness scores for the host genome using insecta and hymenoptera databases, and for Wolbachia genome using rickettsiales database. d Features of the three largest centromeres: ① Chr1 (~4 Mb); ② Chr3 (~4 Mb); ③ Chr8 (~3.5 Mb). e Circular map of the wAnd genome. The outermost tracks 1 and 2 represent the positions of the CDS, tRNA, rRNA, and tmRNA genes on the positive and negative strands. Tracks 3 and 4 tracks show the GC content and GC skew, respectively. f Features of the mitochondrial genome.

Fig. 2. Population structure, phylogeny, and demographic history of Analcellicampa.

a Sampling sites. The bar charts indicate the species and number of samples collected at each sampling location. Colors correspond to those in (d). b Maximum-likelihood tree depicting the evolutionary relationships among genus Analcellicampa and outgroup. c ADMIXTURE analysis with K = 3, 7 and 8. Colors in each column represent ancestry proportion. d Nucleotide diversity (π), nucleotide differences (dxy) across the six species. The value in each circle represents a measure of nucleotide diversity for each species; values in red on each line indicate pairwise population nucleotide differences between species. e Patterns of LD (linkage disequilibrium) decay across the genome in different geographic populations. r², Pearson’s correlation coefficient. f Genomic similarity of six species of Analcellicampa to the AD reference genome. Chromosomes are indicated by different colors along the left y axis. Identical score (IS) values are shown for SNPs within each 50-kb window across the genome. g Estimated split times between each species after 50 bootstraps. The widths indicate probability densities. h Dynamic of N_e inferred by PSMC. The vertical lines indicate divergence times corresponding to those shown in (g). The marked pentagrams indicate species fully infected with Wolbachia.

Fig. 3. Phylogenetic relationships of Wolbachia-infected host and their Wolbachia.

a Phylogeny of host. b Phylogeny of Wolbachia. The numbers at the major branches are bootstrap values.

Fig. 4. Contrasting demographic signals inferred from male and female genomes over time, estimated using PSMC and SMC++.

a N_e dynamics of females and males in three geographically distinct AD. b, c N_e dynamic of females and males in AX and AM, with Wolbachia-uninfected individuals used for the analysis.

Fig. 5. Selective sweep and enrichment analysis of the Wolbachia-infected AD.

a Distribution of the log₂(π-_AD/π-_AX) and XP-CLR values were calculated in 10 kb sliding windows with 5 kb steps. The horizontal and vertical lines represent threshold lines of the top 5% of the XP-CLR and π ratio values, respectively. Points (red) located in the top right sector represent selective signatures in AD. Yellow and black bins in the histograms of XP-CLR (right) and π ratio (top) represent levels respectively higher and lower than the threshold line. b Significant GO enrichment of PSGs associated with Wolbachia-induced reproductive manipulations.