DOI: 10.5061/dryad.4f4qrfjdc
title: Relaxed feeding constraints facilitate the evolution of mouthbrooding in Neotropical cichlids

description: Multifunctionality is often framed as a core constraint of phenotypic
 evolution. Mouthbrooding, a form of parental care where offspring develop
 inside a parent’s mouth, increases multifunctionality by adding a major
 function (reproduction) to a structure already serving other vital
 functions (feeding and respiration). Despite increasing
 multifunctionality, mouthbrooding has evolved repeatedly from other forms
 of parental care in at least 7 fish families. We hypothesized that
 mouthbrooding is more likely to evolve in lineages with feeding
 adaptations that are already advantageous for mouthbrooding. We tested
 this hypothesis in Neotropical cichlids, where mouthbrooding has evolved
 4–5 times, largely within winnowing clades, providing several pairwise
 comparisons between substrate brooding and mouthbrooding sister taxa. We
 found that the mouthbrooding transition rate was 15 times higher in
 winnowing than in non-winnowing clades, and that mouthbrooders and
 winnowers overlapped substantially in their buccal cavity morphologies,
 which is where offspring are incubated. Species that exhibit one or both
 of these behaviors had larger, more curved buccal cavities, while species
 that exhibit neither behavior had narrow, cylindrical buccal cavities.
 Given the results we present here, we propose a new model for the
 evolution of mouthbrooding, integrating the roles of multifunctional
 morphology and the environment.
description: <strong>Phylogeny: </strong>We used a 559-species
 Neotropical cichlid phylogeny trimmed from the cichlid phylogeny presented
 in McGee et al. (2020). <strong>Behavioral
 classifications: </strong>We classified every species in this tree
 for which data was available as either a winnower or a non-winnower, and
 as either a mouthbrooder or a substrate brooder, so that every species was
 categorized as one of four possible states: 1) mouthbrooding and
 winnowing; 2) mouthbrooding and non-winnowing; 3) substrate brooding and
 winnowing; or 4) substrate brooding and non-winnowing. Parental care data
 were gathered from Stawikowski & Werner (1998), Goodwin et al.
 (1998), and López-Fernández et al. (2012), and winnowing data from
 López-Fernández et al. (2014), Weller et al. (2017), and López-Fernández
 et al. (2012). Because winnowing and mouthbrooding encompass a range of
 behaviors, we classified species inclusively for each category: a species
 was classified in a behavioral category if individuals exhibit the
 behavior to any degree that it could impact their morphology. The
 variations of each behavior (for example, mouthbrooding eggs or
 mouthbrooding larvae) probably pose slightly different constraints, but
 because we lacked sufficient phylogenetic and behavioral data to
 differentiate these fine-scaled differences, here we focus on their
 aggregate effects. Any species in which at least one sex orally incubates
 the offspring during at least one developmental period (eggs, larvae, or
 juveniles) was classified as a mouthbrooder. Similarly, any species which
 feeds by orally sifting substrate for invertebrates or edible detritus was
 classified as a winnower, even if it also feeds using other strategies. If
 we could not reliably determine whether or not a species is a
 mouthbrooder--for example, if it was most commonly reported as substrate
 brooding and only rarely reported as mouthbrooding in aquaria (Staeck
 2015), or if mouthbrooding and substrate brooding behaviors seemed highly
 variable even in captivity (Römer et al. 2017, Breeze 2007), these species
 were conservatively classified as substrate brooders (4 species total). We
 also ran our comparative analyses below on versions of the dataset with 1)
 these species classified as mouthbrooders, and 2) excluded from the
 dataset altogether, to assess the sensitivity of our results to these
 ambiguous cases.
 <strong>Dissections</strong>: Dissection images
 include 41 species across 20 genera representing the major radiations of
 Neotropical cichlids, mostly from the University of Michigan Museum of
 Zoology and the Field Museum of Natural History. Minimally, we sampled to
 the resolution of the node representing the most recent common ancestor
 between mouthbrooding and non-mouthbrooding taxa for each clade of
 mouthbrooding species, so that we sampled the immediate substrate brooding
 sister taxon for each mouthbrooding taxon. We selected only specimens that
 were fixed in closed-mouth, neutral positions (i.e. no opercular flaring
 or hyoid depression) for analysis. Each specimen was dissected using a
 modified version of the procedure described by Ridewood (1904) to remove
 the right suspensorium and oral jaw elements, exposing the buccal cavity
 and branchial arches. First, we made a midline cut on an
 anterior-posterior axis separating the premaxillary bones, and continued
 this cut by bisecting the parasphenoid along the midline. We separated the
 circumorbital bones from around the orbit before cutting the levator
 operculi. Ventrally, we separated the basihyal from its articulation with
 the urohyal and basibranchials, then cut the branchiostegal membrane from
 the urohyal through the mandibular symphysis. We were then able to remove
 the entire right cheek intact, including the opercular series,
 suspensorium, adductor mandibulae, nasal and circumorbital bones, and oral
 jaws. We photographed specimens before and after
 dissection on either a Leica or Zeiss stereomicroscope to capture both
 external and buccal cranial morphologies. We based our external landmarks
 on those in Weller (2017), including curves for the rostrum, eye, lower
 jaw, operculum, and adductor mandibulae complex. For the buccal cavity, we
 chose landmarks describing structures that make up the borders of the
 cavity, including the vomer, parasphenoid, basibranchial series, hyoid,
 and lower jaw. A full list of fixed and sliding landmarks is given in the
 supplemental information. <strong>Landmarks:
 </strong>Photographs were scaled and landmarked using
 <em>StereoMorph</em> (Olsen 2015) then imported into
 <em>geomorph</em> using the readland.shapes function, which
 converts <em>StereoMorph</em> curves into sliding
 semilandmarks (Adams et al. 2013).
description: Consult the README.md file for the specific contents of each code
 and datafile and how to run them.