For each species we used separate single-factor (two treatment levels: protein deprived and protein fed) multivariate analyses of variance (MANOVAs) and Fisher’s Least Significant Difference post-hoc analyses to determine whether the mean (± 1 standard error): (1) mechanical properties (ultimate strength, extensibility, toughness, and Young’s modulus), (2) nanostructures (L, 2θ FWHM of the (200) and (120) diffraction peaks and amorphous halo, I200/ I120, Xc, and fc), (3) mole compositions of glutamine, serine, proline, glycine, and alanine, and (4) spidroin (MaSp1a, MaSp1b, MaSp2a, and MaSp2b) expression. We used additional univariate (treatment) ANOVAs to individually compare the MaSp1a, MaSp1b, MaSp2a, and MaSp2b 2-ΔΔCT values between treatments for each species. We log10 or arcsine (amino acid composition data) transformed any data that failed Levene’s heterogeneity tests.