Morphometric Inferences on Sibling Species and Sexual Dimorphism in Neochlamisus Bebbianae Leaf Beetles: Multivariate Applications of the Thin-plate Spline

—Nominally polyphagous species of herbivorous insects sometimes are comprised of multiple morphologically similar biological species with more specialized appetites. When meristic morphological traits cannot be found to distinguish such suspected sibling species, molecular data are increasingly sought as a source of evidence. A role for morphology in distinguishing such taxa might be reclaimed, however, by recent advances in geometric morphometric methods, such as the statistical analysis of partial-warp scores from the thin-plate spline. We employed this method to detect and characterize subtle shape differences among populations and between sexes of the nominal leaf beetle species Neochlamisus bebbianae. Using the thin-plate spline, the shapes of specimens from seven beetle populations collected from five host plants in five eastern North American localities were calculated. These shapes were analyzed by MANOVA, revealing significant variation in both uniform and nonuniform components of shape among test populations. Significant sexual dimorphism in size, shape, and allometric relationships were also documented across these populations. More interestingly, our study provided evidence of sibling species where traditional taxonomic approaches have failed. Individual MANOVAs revealed significant shape variation between sympatric populations from different host plants in each of three localities. Because these sympatric shape differences were significant when adjusted for size, they cannot be attributed to allometric consequences of size variation among test populations. Because certain beetle populations differed significantly in size and shape when reared in a common environment, these morphometric traits may have a genetic basis. Together, these results are consistent with an earlier suggestion that N. bebbianae represents a complex of host-specific races or sibling species, a hypothesis that has received additional support from recent studies on host use traits, sex ratios, and mitochondrial DNA. In sum, these analyses demonstrate the power and utility of the thinplate spline as a morphological means of discriminating among closely related and anatomically homogeneous taxa. [Chrysomelidae; host races; morphometrics; Neochlamisus; relative warps; sexual dimorphism; sibling species; thin-plate spline.] When separate biological species are exacters, he inferred that interbreeding did tremely similar in morphology, documentnot occur between such sympatric popuing their existence is often difficult and lations, which he accordingly diagnosed as many complexes of such cryptic or sibling separate "phytophagic species." Walsh's species go undetected, languishing under approach has since been used to ferret out single nominal species names (Mayr, many host-specific biological species from 1942). This is especially true when musewithin single nominal species assumed to um specimens and traditional taxonomic have more generalized feeding habits approaches provide the only available ev(Brown, 1958, 1959; Diehl and Bush, 1984). idence on biological variation. ObservaThe recognition of cryptic biological spetions on behavioral or ecological polymorties is of taxonomic significance and is also phisms, however, may hint at such sibling vitally important for evolutionary biolospecies and have proven especially inforgists, ecologists, and applied biologists mative in systematic studies of herbivowho often assume the evolutionary and rous insect taxa. In 1864, Benjamin Walsh genetic coherence of the nominal species described slight but consistent morphologthat they study. Data on "intraspecific" ical differences between nominally conspedifferentiation in host use, patterns of ecocific populations of insects associated with logical specialization, or the efficacy of bidifferent host plants at single localities, ological control agents, for example, may From the strict segregation of these chardemand quite different interpretations if