DOI: 10.5061/dryad.b8gtht7bz
title: Data: the complex ecology of genitalia: gonopodium length and allometry in the Trinidadian guppy

description: Male genitalia present an extraordinary pattern of rapid divergence in
 animals with internal fertilization, which is usually attributed to sexual
 selection. However, the effect of ecological factors on genitalia
 divergence could also be important, especially so in animals with
 non-retractable genitalia because of their stronger interaction with the
 surrounding environment in comparison to animals with retractable
 genitalia. Here we examine the potential of a pervasive ecological factor
 (predation) to influence the length and allometry of the male genitalia in
 guppies. We sampled guppies from pairs of low-predation (LP) and
 high-predation (HP) populations in seven rivers in Trinidad, and measured
 their body and gonopodium length. A key finding was that that HP adult
 males do not have consistently longer gonopodia than do LP adult males, as
 had been described in previous work. However, we did find such divergence
 for juvenile males: HP juveniles have longer gonopodia than do LP
 juveniles. We therefore suggest that an evolutionary trend toward the
 development of longer gonopodia in HP males (as seen in the juveniles) is
 erased after maturity owing to the higher mortality of mature males with
 longer gonopodia. Beyond these generalities, gonopodium length and
 gonopodium allometry were remarkably variable among populations even
 within a predation regime, thus indicating strong context dependence to
 their development/evolution. Our findings highlight the complex dynamics
 of genitalia evolution in Trinidadian guppies.
description: <i>Sampling and fish
 care</i> We
 sampled juvenile and adult male guppies in one low and one high predation
 locality in each of seven different streams in the Northern Mountain
 Range, Trinidad. These localities were classified as low and high
 predation localities based on the absence or presence of piscivorous fish,
 respectively (Endler, 1978; Gotanda & Hendry, 2014; Kelly, Godin,
 & Abdallah, 2000; Reznick & Endler, 1982). All fish were
 transported to our laboratory at the William Beebe Tropical Research
 Station in Trinidad, acclimatized for 30 min, transferred to 20 L
 aquariums, and immediately treated for bacterial, fungal, and parasitic
 infections with Polyguard<sup>TM</sup> (Seachem Laboratories,
 Inc.). Fish were fed live brine shrimp or flake food if they remained more
 than 24 hours in the laboratory, but most fish were released back to their
 original site the day after processing (details of processing below). All
 fish were kept at 20 – 24°C and on a natural 12:12 (light:dark)
 photoperiod. All fish handling was in accordance with McGill Animal Use
 Protocol No. 4570. <i>Measurements and
 maturity status</i> Body length (from snout to caudal
 peduncle) and gonopodium length (from base of gonopodium to distal tip,
 excluding the hood – see Kelly, Godin, & Abdallah, 2000) were
 obtained from digital photographs using the software ImageJ (Abràmoff,
 Magalhães, & Ram, 2004). We first anesthetized the fish with an
 aqueous solution of tricaine methanesulfonate (MS-222) and
 NaHCO<sub>3</sub> and then placed them on their right side on
 a white background containing a ruler. We then photographed the left side
 of each fish with a Nikon D300 digital camera equipped with a 60mm macro
 lens, with illumination provided by two full spectrum fluorescent lights
 and a Nikon speedlight commander Kit R1C1 flash.             The development stage
 of males was determined based on the stage of development of the hood, a
 sensory protuberance in the gonopodium (Houde, 1997), which was visualized
 under a Leica ES2 stereomicroscope before the photographs were taken.
 Males were categorized as mature when the hood extended beyond the distal
 tip of the gonopodium (the hook), and immature when the hood was shorter
 than the gonopodium (Houde, 1997). Furthermore, we visually classified the
 development of the gonopodium into three different stages (Fig. 1): (1)
 early-stage, when the differentiation of the anal fin into the gonopodium
 is ongoing and it bears a wide base, forming a triangular shape – not
 shown in Figure 1; (2) Advanced stage, including the sub-stages
 “<i>Hood not developed</i>”, when the gonopodium has developed
 a thinner base – what remains henceforth – and has acquired an appearance
 of a fully developed gonopodium, similar to the “<i>Hood
 developing</i>” sub-stage and the “<i>Final stage</i>”
 of development, as described below; and “<i>Hood
 developing</i>”, representing the phase in which the hood has just
 started to develop until the phase in which it has acquired a
 filament-like shape but is still shorter than the gonopodium; and, finally
 (3) Final stage, when the hood is fully developed and longer than the hook
 – typical from adult males. Although we initially distinguished the
 sub-stages “<i>Hood not developed</i>” from “<i>Hood
 developing</i>”, we subsequently grouped them together as “Advanced
 stage” because there was no apparent difference in the allometric growth
 between these stages. We removed early-stage juveniles from the
 statistical analysis due to small sample sizes; therefore, we refer to
 advanced stage juveniles simply as juveniles henceforth.