DOI: 10.5061/dryad.6t1g1jx1k
title: Male mating success evolves in response to increased levels of male-male competition

description: Male-biased operational sex ratios can increase male-male competition and
 can potentially select for both increased pre-and post-copulatory male
 success. In the present study, using populations of Drosophila
 melanogaster evolved under male-biased (M) or female-biased (F) sex
 ratios, we asked whether (a) male mating success can evolve (b) males are
 better at mating females that they have co-evolved with (c) males mating
 success is affected by female mating status and (d) male mating success is
 correlated with their courtship effort. We directly competed M and F males
 for mating with (a) virgin ancestral (common) females, (b) virgin females
 from the M and F populations, and (c) singly mated females from the M and
 F populations. We also assessed the courtship frequency of the males when
 paired with mated M or F females. Our results show that M males, evolving
 under an increased level of male-male competition, have higher mating
 success than F males irrespective of the female evolutionary history.
 However, the difference in mating success is more pronounced if the
 females had mated before. M males also have a higher courtship frequency
 than F males, but we did not find any correlation between mating success
 and courtship frequency. 
description: This file ("dataset_mating success.xlsx") was generated
 in 2015 by Tejinder Singh Chechi and others at the Evolutionary Biology
 Lab, IISER Mohali. GENERAL INFORMATION
 Author Information  A. Principal Investigator
 Contact Information  Name: Prof. N. G. Prasad
  Institution: Indian Institute of Science Education and Research,
 Mohali  Address: IISER Mohali, Sector 81, Knowledge
 City, SAS Nagar, Punjab - 140306, India.  Email:
 prasad@iisermohali.ac.in   B.
 Associate or Co-investigator Contact Information Name:
 Tejinder Singh Chechi Institution: Indian Institute of
 Science Education and Research, Mohali Address: IISER
 Mohali, Sector 81, Knowledge City, SAS Nagar, Punjab - 140306,
 India. Email: chechi.tj@gmail.com
   Duration of data collection:
 2016-17 Geographic location of data collection: Mohali,
 Punjab, India    Information about
 funding sources that supported the collection of the data: IISER Mohali,
 Govt. of India.  
 <strong>METHODOLOGICAL INFORMATION</strong>
 Details of fly populations Manipulating levels
 of male-male competition.  The flies used in this study
 were derived from a large outbred Drosophila melanogaster stock population
 called LH. Flies from the LHst stock were used to create 3 replicate sets
 of populations subjected to an experimental evolution regimes of either
 elevated or reduced levels of male-male competition (Nandy et al. 2013b).
 Detailed description of selection regimes is provided in the supplementary
 material (supplementary method 1). In brief, male-male
 competition levels were manipulated by changing the sex ratio - M (3:1
 male biased, elevated levels of male-male competition) and F (1:3 female
 biased, reduced levels of male-male competition). There are three
 independent replicate populations belonging to both M (M1-3) and F (F1-3)
 regimes. Flies from the selection populations follow a 14-day discrete
 generation cycle similar to their ancestral stock. On the 9-10th day post
 egg collection, flies are collected as virgins and held at a density of 8
 males or females per vial containing 2mL of food. On day 12, these flies
 are combined into fresh food vials supplemented with measured amount of
 live yeast paste (0.47mg per female), at a density of 24 males and 8
 females for M population and 24 females and 8 males for F population.
 These populations are maintained at a Ne &gt; 450 (Nandy et al.
 2013b). Males and females from M1-3 and F1-3 replicate populations were
 used in this study.  Male mating success competing for
 virgin ancestral females To assay male mating success
 with virgin ancestral females in a competitive scenario, 12 day old (post
 egg collection) flies were used. One virgin male from M, one virgin male
 from F and one virgin female from LHst were transferred to a fresh food
 vial. Males were colored with either pink or green, fluorescent dust
 (Day-Glo Corp.) prior to the assay to allow observers to visually
 distinguish between them. Vials were observed and the male which mated
 successfully was recorded, along with the latency to mate and duration of
 copulation. 80 vials were setup for each replicate line of M and F
 population. In total, 240 such vials were setup and the data from 227
 vials (in which successful matings were observed) was used for further
 analysis. Male mating success competing for virgin
 females from selected populations To assay male mating
 success with virgin females from M and F populations in a competitive
 scenario, a similar design as previous assay was used. We combined one M1
 male and F1 male with one M1 or F1 female in a single vial. A similar
 design was followed for all the replicate populations (M1-3 and F1-3). All
 flies were 12 day old (post egg collection). 80 vials per replicate line
 were setup for F female, and likewise 80 vials per replicate line for M
 female. In total, 480 such vials (160, for each replicate of M1-3 and F1-3
 population) were setup and the data from 456 vials (in which successful
 matings were observed) was used for further analysis. 
 Male mating success competing for singly mated females from
 selected populations  To assay male mating success with
 singly mated females from M and F populations under direct competition, M
 and F females were first mated to common ancestral LHst males. 8 M or F
 females were combined with 10 LHst males into fresh food vials containing
 2mL of food. Vials were observed till all mating pairs were formed, and
 males were separated under light CO2 anesthesia, after all matings ended
 and females were held in vials for 1-2 hours. Singly mated females from M
 or F populations were then combined into fresh food vials with one virgin
 M male and one virgin F male. Males were colored with either pink or green
 fluorescent dust, with reciprocal coloration for each treatment
 combination. Unlike virgin females, singly mated females are less
 receptive and latency to mate with them is much higher. All mating trial
 vials were observed for 8-9 hours in the day-light period spanning from
 3.5 hours to 11.5 hours from time of lights on. 80 vials per replicate
 line were setup for F females, and likewise 80 vials per replicate line
 for M females. In total, 480 such vials (160 vials, for each replicate of
 M1-3 and F1-3 population) were set up and the data from 290 vials (in
 which successful matings were observed) was used for further analysis.
 Males and females from the same replicate line were used in the assay.
 Since the replicate lines are independent, mating assays were distributed
 across three days, with each replicate line handled on a different
 day. M and F male courtship frequency towards singly
 mated females The courtship data was recorded from the
 same vials used in the previous experimental setup used to measure mating
 success competing for singly mated females from selection populations. At
 three different time points during mating observations 5 hours from light
 on, 7.5 hours from light on and 9 hours from lights on, vials were scanned
 every minute for an hour, to record courtship activity. Every minute, each
 vial was scanned for presence or absence of courtship activity. If any of
 the males in a vial display courtship behavior (orientation, chasing, wing
 flapping, licking and attempted copulation) it was recorded as one
 courtship bout, entered with the color identity of the male, or else zero
 activity for the vial was recorded. From this data of total courtship
 bouts in each vial, proportion of courtship bouts performed by each male
 in a vial were calculated.
description: DATA & FILE OVERVIEW File List:
 "Experiment Data.xlsx" Note: This file
 contains four separate tabs: Tab 1. "mating
 success_LHst_virgin"  Tab 2. "mating
 success_virgin" Tab 3. "mating
 success_non-virgin" Tab 4.
 "courtship_data"   Tab 1.
 "mating success_LHst_virgin"  1. Number of
 variables: 7 2. Number of cases/rows: 227
 3. Variable List:  Block (3
 independent replicates of each selection regime; "Block 1",
 "Block 2" or "Block 3") Mating
 latency (Latency to mating from start of observation to start of mating,
 measured in minutes) Copulation duration (copulation
 duration measured in minutes) Male color (two colors
 used to distinguish males; green (g) or pink (p))
 Male ID(male successful in obtaining a mating; M male (M) or F
 male (F))   Tab 2.
 "mating success_virgin"  1. Number of
 variables: 7 2. Number of cases/rows: 456
 3. Variable List:  Block (3
 independent replicates of each selection regime; "Block 1",
 "Block 2" or "Block 3") Female ID
 (Females from two selection regimes used in the experiment; "M
 Female" and "F Female") Mating latency
 (Latency to mating from start of observation to start of mating, measured
 in minutes) Copulation duration (copulation duration
 measured in minutes) Male color (two colors used to
 distinguish males; green "g" or pink "p")
 Male ID(male successful in obtaining a mating; M male
 "M" or F male "F") 
   Tab 3. "mating
 success_non-virgin" 1. Number of variables:
 7 2. Number of cases/rows: 290 3.
 Variable List:  Block (3
 independent replicates of each selection regime; "Block 1",
 "Block 2" or "Block 3") Female ID
 (Females from two selection regimes used in the experiment; "M
 Female" and "F Female") Mating latency
 (Latency to mating from start of observation to start of mating, measured
 in hours) Copulation duration (copulation duration
 measured in hours) Male color (two colors used to
 distinguish males; green "g" or pink "p")
 Male ID(male successful in obtaining a mating; M male
 "M" or F male "F") 
   Tab 4. "courtship_data"
 1. Number of variables: 12 2. Number of
 cases/rows: 431 3. Variable List:  Block (3
 independent replicates of each selection regime; "Block 1",
 "Block 2" or "Block 3")
 Female(Females from two selection regimes used in the
 experiment; "M Female" and "F Female")
 Total bouts(total number of courtship bouts observed)
 Prop M courtship( proportion of courtship bouts from M
 male) Prop F males(proportion of courtship bouts by F
 male) Winner(Male that obtained successful
 mating) Prop_M_Std(Normalised proportion of courtship
 bouts by M male) Prop_F_Std(Normalised proportion of
 courtship bouts by M male)