Pupal remodeling and the evolution and development of alternative male morphologies in horned beetles

Abstract

Background: How novel morphological traits originate and diversify represents a major frontier in evolutionary biology. Horned beetles are emerging as an increasingly popular model system to explore the genetic, developmental, and ecological mechanisms, as well as the interplay between them, in the genesis of novelty and diversity. The horns of beetles originate during a rapid growth phase during the prepupal stage of larval development. Differential growth during this period is either implicitly or explicitly assumed to be the sole mechanism underlying differences in horn expression within and between species. Here I focus on male horn dimorphisms, a phenomenon at the center of many studies in behavioral ecology and evolutionary development, and quantify the relative contributions of a previously ignored developmental process, pupal remodeling, to the expression of male dimorphism in three horned beetle species.

Results: Prepupal growth is not the only determinant of differences in male horn expression. Instead, following their initial prepupal growth phase, beetles may be extensively remodeled during the subsequent pupal stage in a sex and size-dependent manner. Specifically, male dimorphism in the three Onthophagus species studied here was shaped not at all, partly or entirely by such pupal remodeling rather than differential growth, suggesting that pupal remodeling is phylogenetically widespread, evolutionarily labile, and developmentally flexible.

Conclusion: This study is the first to document that male dimorphism in horned beetles is the product of two developmentaly dissociated processes: prepupal growth and pupal remodeling. More generally, adult morphology alone appears to provide few clues, if any, as to the relative contributions of both processes to the expression of alternative male morphs, underscoring the importance of developmental studies in efforts aimed at understanding the evolution of adult diversity patterns.

Figure 2

Ontogenetic changes in allometric scaling between body size and thoracic horn length in male Onthophagus nigriventris. (A) Scaling relationship between body size (presented as standard deviations away from mean) and horn length in male pupae (●) and corresponding adults (○). Allometries differ significantly both in amplitude and slope. (B) Absolute (■ right) and relative (□ left) loss of pupal horn length as a function of adult male body size. Both absolute and relative horn loss decline drastically with adult size.(C) Log-log plot of pupal against adult horn length. Gray line indicates expectation if adult horn length is a direct reflection of pupal horn length (y-intercept = 0, slope = 1). Regression analysis shows that the y-intercept is significantly different from 0 (indicating pupal remodeling) and the slope is significantly greater than 1 (indicating that remodeling occurs to a greater degree in minor compared to major male morphs). Red lines indicate 99% confidence intervals.

Figure 3

Ontogenetic changes in allometric scaling between body size and head horn length in male Onthophagus taurus. (A) Scaling relationship between body size (presented as standard deviations away from mean) and horn length in male pupae (●) and corresponding adults (○). There are no significant differences between pupal and adult allometries. (B) Absolute (■ right) and relative (□ left) loss of pupal horn length as a function of adult male body size. Relative horn loss declines steadily with adult size, whereas absolute horn loss first increases and reaches a maximum in medium-sized males before declining again to near zero values in large males. (C) Log-log plot of pupal against adult horn length. Gray line indicates expectation if adult horn length is a direct reflection of pupal horn length (y-intercept = 0, slope = 1). Regression analysis shows that the y-intercept is significantly different from 0 (indicating pupal remodeling) and the slope is significantly greater than 1 (indicating that remodeling occurs to a greater degree in minor compared to major male morphs). Red lines indicate 99% confidence intervals.

Figure 4

Ontogenetic changes in allometric scaling between body size and thoracic horn length in male Onthophagus binodis. (A) Scaling relationship between body size (presented as standard deviations away from mean) and horn length in male pupae (●) and corresponding adults (○). Pupal and adult allometries differ significantly in y-intercept but not slope. (B) Absolute (■ right) and relative (□ left) loss of pupal horn length as a function of adult male body size. Relative horn loss exhibits a marginally significant negative correlation with adult size, whereas absolute horn loss increases significantly with adult size. (C) Log-log plot of pupal against adult horn length. Gray line indicates expectation if adult horn length is a direct reflection of pupal horn length (y-intercept = 0, slope = 1). Regression analysis shows that the y-intercept is significantly different from 0, but that the slope is indistinguishable from 1 (indicating that remodeling occurs similarly for all males regardless of size). Red lines indicate 99% confidence intervals.

Figure 5

Dorsal view of a male O. binodis as (A) pupa and (B) corresponding adult. Note presence of pronounced lateral concavities, or depressions, in the prothorax of the adult but not pupae (marked by arrows on left side of the adult). a indicates pupal and adult horn length measurements used in this study, which failed to reveal a male dimorphism in this species. b indicates adult horn length measurement used by previous studies, which relies on μ as the posterior landmark, marked by the posterior-most edge of the two lateral prothoracic depressions. (C) Use of μ as posterior landmark, and b as a measure of horn length, recovers a male horn dimorphism in O. binodis adults similar to what has been documented in earlier studies. (D) Scaling relationship between c (which measures the proportion of the prothorax that does not participate in concavity formation) and adult male size. c initially increases with male size and then declines rapidly in large males, suggesting that large males devote a disproportionately larger fraction of their lateral prothorax toward exposing the medial thoracic horn (solid line = log normal regression: r² = 0.42, p < 0.0001; in contrast a linear regression (not shown) fails to yield a significant fit; r² = 0.06; p = ns). Combined, these results are consistent with the hypothesis that dimorphic remodeling of the lateral prothorax during the pupal stage, rather than dimorphic growth of actual horn tissue during the prepupal stage, generates male dimorphism in O. binodis. To increase statistical power data shown in (C) and (D) include an additional 23 male O. binodis reared under identical conditions excpet that no measurements were taken during the pupal stage.

Figure 1

Species used in the present study. (A) Onthophagus nigriventris, (B) O. taurus, and (C) O. binodis. Shown for each species are large horned (major) males (top) and small, hornless (minor) males (bottom) as pupae (left) and corresponding adults (right). Arrows highlight lateral concavity in adult, but not pupal, O. binodis referred to in text.