1. There are old asexuals in low sex/infection lakes # Asexual lineages Background All else being equal, asexual females will produce twice as many daughters as sexual females. Only females contribute directly to population growth; therefore, the production of males by sexual females creates a two-fold cost of sexual reproduction that in theory should result in the elimination of sex (Fig. 1) (Maynard Smith 1978). Sex is extremely common, however, meaning that sexual reproduction must confer profound advantages or that asexual reproduction has sever disadvantages. System Potamopyrgus antipodarum is a good system to use when studying the maintenance of sex because sexuals and asexuals often coexist in natural populations. There is wide across-population variation in the relative frequency of sexual individuals (Lively 1992; Fig. 2), which indicates that sex is favored in some environments while asexuals are favored in others. Materials and Methods Used field-collected snails from 14 New Zealand lakes (Fig. 6). Prior knowledge of mtDNA phylogeography and asexual lineage age distribution (Neiman et al. 2005) were used to select lakes that best captured genetic variation within P. antipodarum. 50 female snails from each high-sex lake and 30 female snails from each low-sex lake had their lengths measured, body dissected and the number of brooded embryos were counted. Used more female snails from high-sex lakes to account for the fact some of the females will be sexual. Heads of snails were frozen for flow cytometry to distinguish between diploid (sexual) and triploid (asexual) snails. I will compare the reproductive output of asexual females from low and high-sex lakes. Expected Results I predict that there will be lower embryo production by females from high-sex vs. low-sex lakes (Fig. 7). Could Mutation Accumulation in Potamopyrgus antipodarum Help to Favor Sex? = Male = Female Fig 1. The two-fold cost of sexual reproduction. Only females make offspring, and thus only females can directly contribute to the rate of population growth. Meaning that an asexual population (left) will grow much more quickly than a sexual population (right). # of Embryos Fig. 7. Predicted differences in embryo production in asexual female P. antipodarum from high- vs. low-sex lakes if mutation accumulation negatively affects asexual phenotype. Asexual Lineage “Quality” Mutation accumulation Implications and Future Directions If our predictions are met, they will provide indirect evidence that accelerated mutation accumulation in P. antipodarum could contribute to asexual lineage extinction and could help explain the maintenance of sex in this species. A different outcome would suggest that either our methods of measuring mutation accumulation were not adequate or mutation accumulation does not favor sex. If we find evidence for low phenotypic quality in snails from low-sex lakes, future research should be directed at estimating, for example, whether more harmful mutations are present in the nuclear genome of new vs. old asexual lineages when compared to sexuals. In addition, since many traits contribute to fitness, more traits should be studied. Acknowledgments I would like to thank ICRU and Carver Trust for the funding for my research. I would also like to thank Jukka Jokela, Kayla King, Kirsten Klappert, Curt Lively, and Dorota Paczesniak for snail collection. Finally I thank Laura Heiberger, Bryce Rasmussen and the rest of the Neiman Lab for their help in freezing and embryo counting. References Cited Fig. 5. Above is a histogram showing the distribution of asexual P. antipodarum lineage ages (Neiman et al. 2005). The blue bars represent young asexual lineages ( /= 5 mutations from the closest sexual relative). The central figure (modified from Lively 1992) shows how these two types of asexual lineages are distributed across low-sex (bottom, blue) and high-sex (top, red) lakes (Lively and Jokela 2002). The circles on the New Zealand map indicate the lakes that were sampled, with white circles indicating lakes in which old lineages were found. Fig. 4. This gene genealogy shows evidence for accelerated mutation accumulation in the mitochondrial genomes of asexual (black) vs. sexual (blue) lineages of P. antipodarum. The genealogy represents genetic variation at the 8330 non-synonymous sites in the ~14 kilobase mitochondrial genome of P. antipodarum. The accumulation of non-synonymous substitutions in asexual P. antipodarum was at a significantly higher rate. This is reflected in the longer length of branches leading to asexual vs. sexual lineages. The increased mutation load of asexual lineages is especially evident in the old asexual lineages (marked with “*”). The sexual P. estuarinus was used as the outgroup. Figure modified from Neiman et al. (2010). Fig. 6 This figure shows the location of the high-sex (blue)and low-sex (red) lakes in New Zealand. Katelyn Larkin, Francesca Baglivi, and Maurine Neiman Department of Biology, University of Iowa, Iowa City, IA, USA Expectations If accelerated mutation accumulation has phenotypic consequences that could play a role in asexual extinction, asexual lineages of P. antipodarum from high-sex regions should have: Relatively high reproductive output. Relatively high population growth rate (Fig. 3). Lively CM, Jokela J. 2002. Temporal and Spatial Distributions of Parasites and Sex in a Freshwater Snail. Evo. Eco. Res. 4: 219-225. Lynch M, Gabriel W. 1990. Mutation Load and the Survival of Small Populations. Evolution 44: 1725-1737. Maynard Smith J. 1978. The Evolution of Sex. Cambridge: Cambridge University Press. 1: 2-5. Muller HJ. 1964. The Relation of Recombination to Mutational Advance. Mut. Res 1: 2-9. Neiman M. Hehman G. Miller J. Logsdon J. Taylor D. 2010. Accelerated Mutation Accumulation in Asexual Lineages of a Freshwater Snail. MBE v27 4:954-963. Neiman M, Jokela J, Lively CM. 2005. Variation in Asexual Lineage Age in Potamopyrgus antipodarum, a New Zealand Snail. Evolution 59: 1945-1952. Fig. 3. Predicted relationship between asexual lineage quality (e.g. reproductive output, population growth rate) from high- vs. low-sex lakes if mutation accumulation has phenotypic consequences that could contribute to asexual lineage extinction and the maintenance of sex. High- sex Low- sex Does mutation accumulation contribute to the maintenance of sex? Theory suggests that sexual lineages are better able to clear harmful genetic mutations than asexual counterparts. Asexual reproduction does not involve genetic recombination, meaning that asexual organisms cannot produce offspring with fewer mutations than they themselves have (Muller 1964). Mutations are inevitable. Asexual lineages should eventually be driven extinct by mutation accumulation (Muller 1964, Lynch and Gabriel 1990). If mutation accumulation is relevant to the maintenance of sex, asexual lineages with a higher mutation load should also have lower absolute fitness. Old asexual lineages of P. antipodarum (derived over 500,000 years ago from sexual ancestors) also have more of these mutations than do more recently derived asexual lineages (Neiman et al. 2010). Old asexual lineages are also confined to low-sex lakes, suggesting that extinction happens more rapidly in lakes where sex is maintained at a high relative frequency (Neiman et al. 2005). (Fig. 5) Pattern suggests more intense selection against asexuals in high-sex lakes. Fig. 2. The frequencies of male and female P. antipodarum vary between lakes. Each of the pie diagrams represents a single lake in New Zealand. The white slices represent male frequencies while the black indicates female frequencies. This shows the wide variation between the relative frequency of sexuals and asexuals. Low-Sex High-Sex Evidence Accelerated mutation accumulation in the mtDNA of asexual vs. sexual P. antipodarum (Fig. 4). ****** **** * * * # Mixed or asexual lineages # Mutations from closest sexual relative (asexual lineage age) Fig. 8. Preliminary data showing the mean embryo number for all snails collected in both low and high- sex lakes. The data collected thus far shows a large variation in the mean number of embryos produced (Fig. 8). This preliminary data shows the embryo output for all the snails dissected, including sexuals. There is a noticeable trend indicating low-sex lakes have a higher mean, but the large variation between lakes encourages me to continue with experiment. Mean embryo number (95% CI)