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. 

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.

METHODOLOGICAL INFORMATION

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 > 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.

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)