DOI: 10.5061/dryad.j3tx95xnp
title: Ecological shifts underlie parallels between ontogenetic and evolutionary allometries in parrotfishes

description: During ontogeny, animals often undergo significant shape and size changes,
 coinciding with ecological shifts. This is evident in parrotfishes
 (Eupercaria: Labridae), which experience notable ecological shifts during
 development, transitioning from carnivorous diets as larvae and juveniles
 to herbivorous and omnivorous diets as adults, using robust beaks and
 skulls for feeding on coral skeletons and other hard substrates. These
 ontogenetic shifts mirror their evolutionary history, as parrotfishes are
 known to have evolved from carnivorous wrasse ancestors. Parallel shifts
 at ontogenetic and phylogenetic levels may have resulted in similar
 evolutionary and ontogenetic allometric trajectories within parrotfishes.
 To test this hypothesis, using micro-CT scanning and 3D geometric
 morphometrics, we analyze the effects of size on the skull shape of the
 striped parrotfish Scarus iseri and compare its ontogenetic allometry to
 the evolutionary allometries of 57 parrotfishes and 162 non-parrotfish
 wrasses. The young S. iseri have skull shapes resembling non-parrotfish
 wrasses and grow towards typical adult parrotfish forms as they mature.
 There was a significant relationship between size and skull shapes, and
 strong evidence for parallel ontogenetic and evolutionary slopes in
 parrotfishes. Our findings suggest that morphological changes associated
 with the ecological shift characterizing interspecific parrotfish
 evolution are conserved in their intraspecific ontogenies.
description: To investigate growth allometry, we examined the ontogenetic
 series of <em>S. iseri</em>, encompassing 54 individuals with
 a total length ranging from 1.75 to 33.5 cm.  For skull
 shape comparison analyses among <em>S. iseri</em>, other
 parrotfishes, and non-parrotfishes wrasses, our sample of adults comprises
 336 individuals (160 adult parrotfishes and 176 adult non-parrotfish
 wrasses) from 217 labrid fishes (Table S1). From this dataset, we compared
 the evolutionary allometry of Scarines reef clade (57 species) to
 ontogenetic allometry of <em>S. iseri</em>. The ontogenetic
 series of <em>S. iseri</em> was obtained through sampling
 carried out in Belize in 2023 (IACUC protocols: UT Austin/AUP-2021-00064
 and SERC/10-15-15-SJB; Collection permits: BF000005-16 and BF0042-22) with
 additional individuals from the Field Museum of Natural History (FMNH).
 The remaining species were obtained through several museums (Table
 S1). We assessed the three-dimensional skull shape of
 each labrid species through micro-computed tomography (μCT) scans.
 Specimens were scanned at Rice University, the University of Texas Austin,
 the University of Minnesota, and the University of Washington Friday
 Harbor Labs in conjunction with the scanAllFishes and oVert initiatives.
 The scans were segmented in Amira v2.0.0 (Thermo Fisher Scientific,
 Waltham, MA) to isolate the skull while eliminating scales and debris. The
 isolated skulls were transformed into 3D meshes and imported into
 Checkpoint (Stratovan, Davis, CA). We followed the landmark scheme of
 [19,28], however, three additional landmarks detailing points along the
 pectoral girdle were incorporated [27]. The characterization of shape
 variation involved 200 3D points, 83 landmarks, and 117 semi-landmarks.
 These points comprehensively sampled the pharyngeal jaws, oral jaws,
 neurocranium, nasals, hyomandibula, operculum, hyoid apparatus, and
 pectoral girdle. All points were exclusively situated on the left side of
 the skull. To accommodate for the rotation and
 translation inherent in highly kinetic articulating components of the fish
 skull), we initially aligned each dataset through Generalized Procrustes
 analysis (GPA) using the <em>gpagen</em> function.
 Subsequently, a local superimposition was conducted to standardize the
 positioning of diverse skull elements [38,39]. To analyze the shape
 evolution of individual bones, we partitioned our extensive skull shape
 dataset into smaller datasets for each bone, and the raw coordinates were
 individually superimposed.  In this context, size was quantified as
 centroid size (CS), which represents the square root of the summed squared
 distances of each landmark to the centroid. After the local
 superimposition, we computed the average shapes and centroid sizes for the
 adult specimens of each species. All geometric morphometric analyses were
 carried out using the <em>geomorph </em>package, version 4.0.4
 [40] in R, version 4.3.2 [41].
description: # Ecological shifts underlie parallels between ontogenetic and
 evolutionary allometries in the striped parrotfish Continued methods
 **(a)   Ontogenetic allometry of *Scarus iseri*** To analyze the shape
 variation of entire skull and individual bones of *S. iseri* across
 development, we applied Principal Component Analysis (PCA) using
 *gm.prcomp* function in *geomorph* package (Adams et al., 2022). To
 estimate coefficients of ontogenetic allometry and test the effects of
 size on skull and bone shapes (superimposed coords), we used Analysis of
 Variance, fitting a model with one main effect, the factor “Size”,
 measured as ln-transformed CS (LCS). The ANOVA was performed using the
 *procD.lm* function in *geomorph* (Collyer and Adams, 2018; 2021). To
 better visualize these relationships, we predicted individual shapes using
 group-specific regression models, and the resulting shapes for all samples
 were plotted using a PCA. We plotted PC1 scores against size. Allometric
 axes were then plotted on the first PC, along with scores relative to the
 size axis, using the *plotAllometry* function in *geomorph* with the
 method set to "RegScore" **(b)  Ontogenetic and evolutionary
 allometry comparisons** To compare the ontogenetic allometric coefficients
 of *S. iseri* and the evolutionary allometry of parrotfishes, we conducted
 pairwise analyses involving vectors of allometric coefficients. The growth
 allometries were estimated using OLS, while evolutionary allometries were
 estimated using PGLS. Following the methodology outlined by Zelditch and
 Swiderski (2023), we executed pairwise comparisons by extracting fitted
 values from the OLS and PGLS models. These values were then combined, and
 a subsequent ANCOVA was performed using the *procD.lm* function in the
 *geomorph* package. This ANCOVA involved regressing the fitted values
 against size. Finally, pairwise comparisons between vectors were carried
 out, utilizing the *pairwise* function in RRPP. To explore evolutionary
 allometry, we used a recently published phylogenetic hypothesis involving
 410 species of labrid fishes (Larouche et al., 2023). The tree was pruned
 down to include the 217 taxa from which morphometric data were collected,
 a process accomplished using the *drop.tip* function in the *ape* package
 version 5.0 (Paradis and Schliep, 2019). To derive coefficients for
 evolutionary allometry and assess the evolutionary allometry of
 parrotfishes, we employed Phylogenetic Analysis of Covariance (PGLS). The
 model encompassed one principal effect: the factor "Size,"
 measured as the natural logarithm-transformed centroid size
 (ln-transformed CS or LCS). We used a Type II Sums of Squares for
 statistical tests of model terms (Adams and Collyer, 2018). In line with
 the analysis of growth allometry, tests of statistical significance were
 conducted through permutations of the residuals from the reduced model.
 The statistical significance of the slopes was determined by comparing it
 to the distribution of random values. The phylogenetic ANCOVA was fitted
 using the *procD.pgls* function in *geomorph*, and pairwise comparisons
 between ontogenetic and evolutionary allometries were conducted using the
 *pairwise* function in RRPP. Beyond assessing the null hypothesis that the
 angles differ no more than expected by chance, we also examined the
 hypothesis that they are no more similar than anticipated by chance.
 Allometric axes were then plotted on the first PC, along with scores
 relative to the size axis, using the *plotAllometry* function in
 *geomorph* with the method set to "Predline." Given the species
 non-independence we reduced the ordinary least squares (OLS) residuals
 through PCA up to the first 12 principal components. Subsequently, we
 estimated Pagels λ for both multivariate datasets, utilizing the
 *transformPhylo.ML* function in the *motmot* package version 2.1.3
 (Puttick et al., 2019). Finally, we transformed the phylogeny by the λ
 value using the *rescale* function in the *geiger* package version 2.07
 (Pennell et al., 2014). ## Description of the data and file structure We
 have submitted our museum specimen information (`Table_S1.docx`), raw data
 (landmark coordinate files in a 2d array format  for all labrids 
 - `merged_labrid_coords.csv`, just for parrotfishes -
 `merged_labrid_coords_PARROTS.csv`, just for wrasses species
 -`merged_labrid_coords_WRASSES.csv`, and just for Scarus iseri
 - `merged_scarus_coords.csv`), slide landmarks for entire skull and each
 individual bone (`skull_slider_2.csv`, `angular_slide.csv`,
 `cer_slide.csv`, `dentary_slide.csv`, `hyo_slide.csv`, `nasal_slide.csv`,
 `neuro_slide_2.csv`, `pjaw_slide.csv`, `premax_slide.csv`,
 `uro_slide.csv`), raw data for group information (`ontogeny_groups.csv`,
 `Scarines_groups.csv,` `parrots_groups.csv`, `wrasses_groups.csv` and
 `Scarus_LS.csv`),  raw trimmed phylogenetic trees
 (`Labrid82ModTimeTree.tre`), and R script (`Parrotfish_ontogeny.R`). *
 Table S1: Species list used in our study (word format): species, sample
 number and origins (Museum or Fieldwork) * *Species*: Currently valid
 scientific name * *n*: sample number * *Source/Museum*: Fieldwork Belize,
 museums (BMNH, ANSP, FMNH, KAUM, UF, UWFC, AM, USNM, AMNH, JFBM, UVI), or
 Uncat (species not deposited in a museum) * *Catalog number*: voucher
 number associated to the museum or Uncat (species not deposited in a
 museum) * **Ontogeny - S. iseri shape dataset**
 (*merged_scarus_coords.csv*): the superimposed landmarks coordinates of 54
 *S. iseri* individuals (csv file) used for ontogenetic analysis (Step 1 of
 the R script) * *Species* (categorical variable): *Scarus iseri*
 specimens  * *V1-V600* (numerical variable): landmark coordinates in a 2d
 array formart * **Evolutionary allometry - adult parrotfish shape
 dataset** (*merged_labrid_coords_PARROTS.csv*): the superimposed landmarks
 coordinates of 91 parrotfishes (csv file) used for comparing ontogenetic
 allometry of *S. iseri* to the evolutionary allometries of Scarinini (Step
 2 of the R script) * *Species* (categorical variable): Currently valid
 scientific name * *V1-V600* (numerical variable): landmark coordinates in
 a 2d array formart * **All Labridae evolutionary allometry - adult labrids
 shape dataset** (*merged_labrid_coords.csv*): the superimposed landmarks
 coordinates of 315 individuals (csv file) used for comparing ontogenetic
 allometry of *S. iseri* to the other Labrids (Step 3 of the R script) *
 *Species* (categorical variable): Currently valid scientific name *
 *V1-V600* (numerical variable): landmark coordinates in a 2d array format
 - **Wrasses evolutionary allometry - adult labrids shape dataset**
 (*merged_labridcoords*_WRASSES.csv): the superimposed landmarks
 coordinates of 176 individuals (csv file) used for comparing ontogenetic
 allometry of *S. iseri* to the other Labrids (Step 3 of the R script) *
 *Species* (categorical variable): Currently valid scientific name *
 *V1-V600* (numerical variable): landmark coordinates in a 2d array format
 * **Landmarks definition**: landmarks and semilandmarks definition used
 for whole skull and each bone ((**skull_slider_2.csv, angular_slide.csv,
 cer_slide.csv, dentary_slide.csv**, **hyo_slide.csv**,
 **nasal_slide.csv**, **neuro_slide_2.csv**, **pjaw_slide.csv**,
 **premax_slide.csv**, **uro_slide.csv**) * *Before, Slide,
 Ater*: positions and links of  landmarks and semilandmarks * Bones code:
 Angular (ang), Ceratohyal (cer), Dentary (dent), Hyomandibula (hyo), Nasal
 (nasal), Neurocranium (neuro), Pharyngeal jaw (pjaw), Premaxilla (premax),
 Urohyal (uro)  * **Labridae Tree** (*Labrid82ModTimeTree.tre*): the
 phylogeny (TRE format) * Phylogenetic tree with 412 tips and 411 internal
 nodes. * Classifiers (**ontogeny_group.cvs**): Table containing the name
 of the species indicating the classification according to two taxonomic
 groups * *Specimen* (categorical variable): individual species' name
 * *Species* (categorical variable): Currently valid scientific name *
 *Group* (categorical variable):  1 = wrasses species; 2 = parrotfishes
 species; * *Group_2* (categorical variable): 1 = wrasses species; 2 =
 parrotfishes species; 3 = *S. iseri* individuals) * Classifiers
 (**Scarus_LS.cvs**): Table containing S. iseri individuals with total
 length (millimeters) and size class codes. * *Species* (categorical
 variable): individual species' name and its identification number *
 *TL* (numerical variable):  total length (millimeters)  * *Size_code*
 (categorical variable): 1 = &gt; 2.2 cm; 2 = 2.3 - 3.5 cm; 3 = 3.6 - 5.5
 cm; 4 = 5.6 - 6.9 cm; 5 = 7.0 - 10.7 cm; 6 = 10.8 - 12.2 cm; 7 = 12.3 -
 14.9 cm; 8 = 15.0 - 20.3 cm; 9 = 28.9 - 33.7 cm) * Classifiers
 (**Scarins_group.cvs**): Table containing all parrotfishes used in the
 study and their classifications between adults Scarines and *S.
 iseri* individuals from ontogeny series * *Species* (categorical
 variable): Currently valid scientific name * *Group* (categorical
 variable):  Scarines (adult parrotfishes) or Scarus_ontogeny (*S.
 iseri* individuals from ontogeny series) - Classifiers
 (**wrasses_group.cvs**): Table containing all wrasses used in the study
 and their classifications between adults wrasses and *S.
 iseri* individuals from ontogeny series * *Species* (categorical
 variable): Currently valid scientific name * *Group* (categorical
 variable): Wrasses (adult wrasses species) or Scarus_ontogeny (*S.
 iseri* individuals from ontogeny series) ## Code/Software R is required to
 run *Parrotfish_ontogeny.R*; the script was created using version 4.3.2.\
 Annotations are provided throughout the script through: \#STEP 1 - *Scarus
 iseri* ontogeny analysis \#STEP 2 - Comparing ontogenetic allometry of *S.
 iseri* to the evolutionary allometries of Scarinini and all Labrids \#STEP
 3 - Comparing skull shape of *S. iseri* to the other Labrids ###
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