Sperm of some species form motile, coordinated groups as they migrate through the female 49 reproductive tract to the site of fertilization. This collective motion is predicted to improve sperm 50 swimming performance and therefore may be beneficial in a competitive context, but limited 51 evidence supports this theory. Here we examine sperm aggregates across closely-related species of 52 Peromyscus mice that naturally vary by mating system, and thus sperm competition intensity. We 53 find that phylogenetic history predicts the likelihood that sperm will aggregate, and that relative 54 testis size is negatively associated with variation in number of aggregated cells, suggesting that 55 sperm competition has a stabilizing effect on sperm group size. Moreover, we show that 56 aggregates are kinematically beneficial for some species but costly for others, and these 57 differences are largely dependent on the orientation and composition of sperm within the groups.
58In addition, when we compared sperm of the two sister-species that aggregate most frequently, we 59 find that sperm from the species that evolved under intense sperm competition forms aggregates 60 with more efficient geometry more frequently than sperm from its monogamous congener. These 61 results are consistent with the prediction that sperm aggregation evolved to improve motility in a 62 competitive context; however, when monogamy evolved secondarily, relaxed sexual selection 63 allowed for less efficient strategies to persist. Together, our findings in Peromyscus reveal that 64 collective sperm behavior is likely to evolve rapidly and is shaped by changes in the selective 65 regime. 66 67 widely across taxa (7). In mammals alone, sperm of some species assemble during epididymal 100 transport and are molecularly "glued" to one another as bi-flagellate pairs in grey short-tailed 101 opossums (25), as bundles of roughly 100 cells in monotremes (26), or as organized rouleaux of 102 five or more cells in guinea pigs (Cavia porcellus, 24). Conversely, mammalian sperm may also 103 assemble after ejaculation to form variably-sized groups. For instance, sperm may form temporary 104 clusters of up to sixteen cells in bulls (Bos Taurus, 15) or more fixed groups of up to 30 cells in 105 house mice (5), 50 cells in the Norway rat (5), or thousands of cells in the wood mouse (Apodemus 106 sylvaticus, 4) whereby the hook-shaped heads interlock or attach to the flagella of other sperm. For 107 these latter species in which sperm groups form after ejaculation, both single and aggregated 108 sperm typically co-occur, thus allowing for direct comparisons between collective and solitary 109 sperm movements within an ejaculate while controlling for within-male variability.
110Closely-related species in the rodent genus, Peromyscus, produce sperm that naturally vary in 111 their collective behavior. Sperm of some species assemble temporary groups after ejaculation by 112 adhering to one another at their head region (27) and disassemble prior to fertilization (10). In P. 1...