# This file includes the commands used to read the datafile and perform 
# non-parametric multiple regression analyses using Sex, SkinMass, and Age 
# as response variables and BufotenineQuantity, AlkaloidQuantity, and
# AlkaloidRichness, respectively, as explanatory variables, using package ape
# in R Project 3.2.1, with individual ages estimated using the back-calculation 
# method proposed herein (AgeJeckel), as well as those of Sagor et al. 
# (1998; AgeSagor), Guarino et al. (2003, 2004; AgeGuarino), and Piantoni et al. 
# (2006; AgePiantoni), respectively. See main article and Additional files
# for details and references. 

moreirae = read.table("Additional_file_5.txt", header = TRUE) #read dataframe including all specimens
moreirae
require(ape) #load package ape
lmorigin(BufotenineQuantity~Sex + SkinMass + AgeJeckel, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidQuantity~Sex + SkinMass + AgeJeckel, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidRichness~Sex + SkinMass + AgeJeckel, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)

lmorigin(BufotenineQuantity~Sex + SkinMass + AgeSagor, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidQuantity~Sex + SkinMass + AgeSagor, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidRichness~Sex + SkinMass + AgeSagor, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)

lmorigin(BufotenineQuantity~Sex + SkinMass + AgeGuarino, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidQuantity~Sex + SkinMass + AgeGuarino, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidRichness~Sex + SkinMass + AgeGuarino, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)

lmorigin(BufotenineQuantity~Sex + SkinMass + AgePiantoni, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidQuantity~Sex + SkinMass + AgePiantoni, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidRichness~Sex + SkinMass + AgePiantoni, data=moreirae, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)

moreirae[moreirae$Maturity!="Juvenile", ] -> moreirae_adults #create new dataframe excluding juveniles
moreirae_adults
lmorigin(BufotenineQuantity~Sex + SkinMass + AgeJeckel, data=moreirae_adults, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidQuantity~Sex + SkinMass + AgeJeckel, data=moreirae_adults, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)
lmorigin(AlkaloidRichness~Sex + SkinMass + AgeJeckel, data=moreirae_adults, origin=FALSE, nperm=9999, method=NULL, silent=TRUE)