Fertilization modes and the evolution of sperm characteristics in marine fishes: Paired comparisons of externally and internally fertilizing species

Abstract

Fertilization mode may affect sperm characteristics, such as morphology, velocity, and motility. However, there is little information on how fertilization mode affects sperm evolution because several factors (e.g., sperm competition) are intricately intertwined when phylogenetically distant species are compared. Here, we investigated sperm characteristics by comparing seven externally and four internally fertilizing marine fishes from three different groups containing close relatives, considering sperm competition levels. The sperm head was significantly slenderer in internal fertilizers than in external fertilizers, suggesting that a slender head is advantageous for swimming in viscous ovarian fluid or in narrow spaces of the ovary. In addition, sperm motility differed between external and internal fertilizers; sperm of external fertilizers were only motile in seawater, whereas sperm of internal fertilizers were only motile in an isotonic solution. These results suggest that sperm motility was adapted according to fertilization mode. By contrast, total sperm length and sperm velocity were not associated with fertilization mode, perhaps because of the different levels of sperm competition. Relative testis mass (an index of sperm competition level) was positively correlated with sperm velocity and negatively correlated with the ratio of sperm head length to total sperm length. These findings suggest that species with higher levels of sperm competition have faster sperm with longer flagella relative to the head length. These results contradict the previous assumption that the evolution of internal fertilization increases the total sperm length. In addition, copulatory behavior with internal insemination may involve a large genital morphology, but this is not essential in fish, suggesting the existence of various sperm transfer methods. Although the power of our analyses is not strong because of the limited number of species, we propose a new scenario of sperm evolution in which internal fertilization would increase sperm head length, but not total sperm length, and change sperm motility.

Keywords: copulation; external fertilization; genital morphology; internal fertilization; marine fish; sperm competition.

FIGURE 1

Interfamilial relationship of phylogeny of (a) Ovalentaria, (b) Scorpaenoidei, and (c) Gasterosteoidei. Each phylogenetic tree was based on Near et al. (2013), Smith et al. (2018) and Kawahara et al. (2009), respectively. Outlined taxa show the families used in this study.

FIGURE 2

Sperm morphology of species with external and species with internal insemination. (a) Amphiprion clarkii. (b) Chromis notata. (c) Pomacentrus nagasakiensis. (d) Ditrema temmincki temmincki. (e) Dendrochirus zebra. (f) Paracentropogon rubripinnis. (g) Sebastes cheni. (h) Sebastiscus marmoratus. (i) Aulorhynchus flavidus. (j) Hypoptychus dybowskii. (k) Aulichthys japonicus. Scale bars indicate 5 mm.

FIGURE 3

Differences in sperm morphological characteristics and sperm velocity (mean ± SD) of three groups with different fertilization modes. Total sperm length in Group I (a), Group II (b), and Group III (c). Head ratio in Group I (d), Group II (e), and Group III (f). Midpiece ratio in Group I (g), Group II (h), and Group III (i). Sperm velocity in Group I (j), Group II (k), and Group III (l). White bars and black bars indicate species with external fertilization and internal insemination, respectively. Estimated relative sperm competition levels (ESCL) are also shown under the bar: Low (L), medium (M), and high (H). Ac, Amphiprion clarkii; Af, Aulorhynchus flavidus; Aj, Aulichthys japonicus; Cn, Chromis notata; Dtt, Ditrema temmincki temmincki; Dz, Dendrochirus zebra; Hd, Hypoptychus dybowskii; Pn, Pomacentrus nagasakiensis; Pr, Paracentropogon rubripinnis; Sc, Sebastes cheni; Sm, Sebastiscus marmoratus. Different letters indicate significant differences between species (LMMs with sequential Bonferroni correction, p < .05).

FIGURE 4

Relationship between relative testes mass (RTM) and sperm components. (a) Correlation between RTM and the ratio of sperm head length to total sperm length (PGLS, intercept = 0.08, slope = −0.061). (b) Correlation between RTM and velocity (PGLS, intercept = 82.49, slope = 92.97). Regression lines are from PGLS analyses (see Table 4 for statistics).

FIGURE 5

Genital morphology of species with external fertilization and species with internal insemination. (a) Amphiprion clarkii. (b) Chromis notata. (c) Pomacentrus nagasakiensis. (d) Ditrema temmincki temmincki. (E) Dendrochirus zebra. (f) Paracentropogon rubripinnis. (g) Sebastes cheni. (h) Sebastiscus marmoratus. (i) Aulorhynchus flavidus. (j) Hypoptychus dybowskii. (k) Aulichthys japonicus. Scale bars indicate 5 mm; arrows indicate the positions of genitalia.