Variation in mutation, recombination, and transposition rates in Drosophila melanogaster and Drosophila simulans

**Background:** The rates of mutation, recombination, and transposition are fundamental parameters in evolutionary biology, yet direct estimates from wild populations are scarce, even for model species like Drosophila melanogaster. Most prior estimates come from mutation accumulation (MA) lines, which may be biased by selection against deleterious mutations and laboratory adaptation. This study aimed to provide direct, unbiased estimates of these rates and quantify their variation between populations, sexes, and individuals using a parent-offspring sequencing approach in fully wild flies.

**Methods:** The authors sequenced parents and offspring from 18 families of outbred full-sibships: six families from a West African population of D. melanogaster (Zaria, Nigeria), six from a European population of D. melanogaster (Sussex, England), and six from a European population of D. simulans (Gimenells, Spain). Each family comprised two parents and five male F1 offspring (125 flies total; one offspring failed sequencing). DNA was extracted from single flies and sequenced using Illumina NovaSeq (2 × 150 bp paired-end) to a median coverage >30-fold. Reads were mapped to reference genomes (D. melanogaster r6.42; D. simulans GCF_016746395.2). De novo single-nucleotide mutations (SNMs) were identified using GATK with stringent filtering, and candidate mutations were manually inspected in IGV. Callable sites were estimated via simulation of synthetic mutations in raw reads. Recombination breakpoints were inferred by phasing parental haplotypes using offspring data and identifying phase switches. Transposable element (TE) insertions were detected using TEFLoN, with new insertions defined as those heterozygous in a single F1 offspring but absent from all other individuals. Statistical analyses used Bayesian binomial generalized linear mixed models (MCMCglmm) with population and sex as fixed effects and parental ID as a random effect.

**Key Results:** Across 89 offspring, the authors identified 58 SNMs, 286 crossovers, and 89 TE insertions. The overall de novo SNM rate was 3.32 × 10⁻⁹ site⁻¹ gen⁻¹ (95% binomial bounds: 2.52–4.30 × 10⁻⁹). The West African D. melanogaster population had a significantly lower mutation rate (1.67 × 10⁻⁹; 95% HPD CI 0.54–3.14) compared to European D. melanogaster (4.86 × 10⁻⁹; 2.11–8.02; P = 0.035) and European D. simulans (4.51 × 10⁻⁹; 1.94–7.75; P = 0.048). The mutation rate was significantly higher in males than females (5.24 × 10⁻⁹ vs. 2.05 × 10⁻⁹; P = 0.010). Nine short indels were identified, giving an overall indel rate of 5.00 × 10⁻¹⁰ site⁻¹ gen⁻¹ (95% binomial bounds: 2.29–9.50 × 10⁻¹⁰). For recombination, the female recombination rate (excluding inversions) was 3.44 cM/Mb (95% HPD CI 2.72–4.18) in West African D. melanogaster, significantly higher than European D. melanogaster at 2.06 cM/Mb (1.57–2.57; P = 0.004), but not significantly different from European D. simulans at 3.04 cM/Mb (2.45–3.73; P = 0.430). No recombination was detected in males. Large chromosomal inversions significantly suppressed recombination (P < 2 × 10⁻⁴). For TE insertions, the per-copy insertion rate was significantly lower in West African D. melanogaster (8.99 × 10⁻⁵ copy⁻¹ gen⁻¹; 95% HPD CI 5.33–14.21) than in European D. melanogaster (23.36 × 10⁻⁵; 17.04–31.26; P = 0.026), with European D. simulans at an intermediate rate (18.70 × 10⁻⁵; 12.25–27.48). Males tended to have more TE insertions than females, but this was not significant (P = 0.118). The most active TE superfamily in D. melanogaster was CMC-Transib (3.09 × 10⁻³ insertions copy⁻¹ gen⁻¹).

**Clinical Implications:** While this study is not directly clinical, it provides fundamental insights into the rates and variation of mutation, recombination, and transposition in a key model organism. These findings have implications for understanding the mutational burden, genetic load, and evolutionary dynamics that can inform studies of genetic variation, disease modeling, and evolutionary medicine. The demonstration that mutation rates can vary nearly threefold between populations of the same species highlights the importance of using population-specific estimates when calibrating molecular clocks or estimating effective population sizes, which are critical for interpreting genetic diversity in both model organisms and humans.