DOI: 10.5061/dryad.gf1vhhmnq
title: Impacts of flowering density on pollen dispersal and gametic diversity are scale dependent

description: Pollen dispersal is a key evolutionary and ecological process, but the
 degree to which variation in the density of concurrently flowering
 conspecific plants (i.e., co-flowering density) shapes pollination
 patterns remains understudied. We monitored co-flowering density and
 corresponding pollination patterns of the insect-pollinated palm
 Oenocarpus bataua in northwestern Ecuador and found that the influence of
 co-flowering density on these patterns was scale-dependent: high
 neighborhood densities were associated with reductions in pollen dispersal
 distance and gametic diversity of progeny arrays, whereas we observed the
 opposite pattern at the landscape scale. In addition, neighborhood
 co-flowering density also impacted forward pollen dispersal kernel
 parameters, suggesting that low neighborhood densities encourage pollen
 movement and may promote gene flow and genetic diversity. Our work reveals
 how co-flowering density at different spatial scales influences pollen
 movement, which in turn informs our broader understanding of the
 mechanisms underlying patterns of genetic diversity and gene flow within
 populations of plants.
description:          We recorded monthly phenological state for all adults
 (n=181) within a core 130-ha plot by visiting each tree within the study
 plot and recording the number of reproductive structures. In 2015, we
 surveyed a 250 m buffer zone around the 130-ha plot and recorded 60
 additional adults, all mapped and genotyped, but for which phenology was
 not recorded. This yielded 241 geo-located and genotyped candidate
 fathers, of which 181 also had monthly phenology data for analysis. To
 collect progeny for genetic analysis, we collected ripe fruits directly
 from the infructescences of <em>O. bataua </em>individuals; we
 refer to these trees as ‘maternal’ trees and pollen sources as ‘paternal’
 trees, noting that the same individual could be both. We randomly sampled
 maternal trees with complete phenology records. We germinated the seeds in
 a nursery and collected and stored a tissue sample from the first leaf of
 each seedling. For each maternal tree, we used monthly phenology data to
 calculate the co-flowering density in the time frame that the maternal
 tree was flowering. We calculated density at two spatial scales: the
 ‘landscape’ scale, defined as the entire 130-ha study plot, and
 ‘neighborhood’ scale, defined as the average effective pollination
 neighborhood (A<sub>ep</sub>) of all progeny arrays included
 in this study, which was a circular area with a radius of ~ 320 m and an
 area of 33.40 ha. If the boundary of the neighborhood area extended beyond
 that of the study plot, neighborhood co-flowering density was estimated
 using the subset of individuals within the neighborhood area for which
 phenology data were available.        We extracted
 genomic DNA from 962 offspring leaf samples representing 43 progeny arrays
 collected from 35 maternal trees. All samples were genotyped using 11
 microsatellite loci through Polymerase Chain Reaction, following
 established protocols. We genotyped the 181 adults (pollen sources and
 maternal trees) in the study plot and the 60 additional adults in the 250
 m buffer around the study plot using equivalent methods, totaling 241
 genotyped parental trees. We used offspring and adult genotypes in the
 program CERVUS v. 3.0.3 (Marshall <em>et al.,</em> 1998) to
 assign paternity with the aim of (1) calculating the average
 A<sub>ep</sub>, (2) quantifying the ‘observed’ distance pollen
 dispersed from paternal sources to maternal trees, and (3) estimating the
 paternal contribution to the genetic diversity of each progeny array. We
 used critical trio (Δ) values with at least 80% confidence and the
 following simulation parameters used previously for parentage analysis in
 our study area.
description: Folders and Files: Folders are .zip files
   1. Folder name - Distance calculation: the
 files in this folder were used to calculate the distance that pollen
 dispersed from CERVUS assigned paternal trees to maternal trees.
 1a.
 File name - data.csv. This is the data to be read into R. 
 1ai.
 Variables: MotherID is the genetic sample code for the maternal
 tree.  
 UTM1 is the first coordinate for the mother tree. 
 UTM2
 is the second coordinate for the maternal tree. Candidate father is the
 assigned paternal tree in CERVUS.  UTM1 is the first
 coordinate for the paternal tree.  UTM2 is the second
 coordinate for the paternal tree.  1b. File name -
 AssignedFathers.csv. This file is the list of CERVUS assigned fathers
 (column 3) for each offspring (column 1), corresponding to each maternal
 tree (column 2).  1c. File name -
 code.R. This is the annotated R script.  1d. File name -
 output.csv. This is the output from the R script. You should get the
 maternal tree id/progeny array (column 2), the mean pollen dispersal
 distance (column 3), 
 standard deviation (column 4), and variance (column
 5).   2. Folder name -
 Modeling_Figures1-3: This is the files in this folder were used to
 complete the main modeling results and generate figures 1-3 in the
 manuscript.  2a. File name -
 data.csv. File with the summary data for each progeny array. 
 2ai.
 Variables: Tag is the unique maternal ID tag name.
 ID
 is the non-unique maternal ID. Date is the month that
 the maternal tree was in flower with 1 being September 2011 and 48 is
 August 2015 UTM1 is the first
 coordinate for each maternal tree UTM2 is the second
 coordinate for each maternal tree  x is the longitude for
 each maternal tree y is the longitude for
 each maternal tree   Alphak_50 is the alpha
 diversity measure for male gametes for each progeny array
 mean
 is the mean pollen dispersal distance for each maternal tree
 
 Log_Alpha is log transformed Alphak_50 
 
 Ndensity is estimated neighborhood density
 
 Ldensity is estimated landscape density
 
 Mean_CI is the 95% confidence interval for average pollen
 dispersal distance Adiv_CI is the 95% CI
 for alpha diversity  Radius is the radius
 of the neighborhood area for each maternal tree 
 
 AepNDensity is the estimated density of the neighborhood
 density in the neighborhood area (Aep) 2b. File name -
 LD_high.csv. Observed pollen dispersal distances for offspring fertilized
 during high landscape density scenarios.  2c. File name -
 LD_low.csv. Observed pollen dispersal distances for offspring fertilized
 during low landscape density scenarios. 2d. File name -
 ND.high.csv. Observed pollen dispersal distances for offspring fertilized
 during high neighborhood density scenarios. 2e. File name -
 ND.low.csv. Observed pollen dispersal distances for offspring fertilized
 during low neighborhood density scenarios. 2f. File name -
 all_dists.csv. Observed pollen dispersal distances for offspring
 fertilized during all density scenarios.  2g. File name -
 Stats_Figure1-3.R. Script for modeling and making figures in R. 
 2h.
 File name - Figure1.tiff. Output for figure 1. 2i. File name -
 Figure1.tiff. Output for figure 2. 2j. File name -
 Figure3.tiff. Output for figure 3.  
 3. Folder name - Pollen_kernel_simulations.
 3a.
 File name - Adult_Genotypes_UTMs.csv. This file contains all adult O.
 bataua genotypes and coordinate information for each adult (UTM1,
 UTM2).  
 3ai. Variables: ID is the genetic sample ID for each
 individual Field_Tag is the field
 ID for each individual UTM1 is first
 coordinate UTM2 is the second
 coordinate Mother? Is a yes/no
 detailing if that individual is a maternal tree
 The
 following columns are microsatellite alleles. 3b. File name -
 allelePmiDiversity.R. This script is used to calculate allelic diversity
 of the pollen pool. Modified from the DispersalDiversity R
 package. 3c. File name -
 Flowering_IDS.csv. This file contains the Field tag of all flowering
 maternal trees in the study (Array_Maternal_ID).
 3d.
 File name - Paternal_IDS.csv. This file contains the Field tags of all
 potential flowering trees (Pat_ID) for each flowering maternal tree
 (Mat_ID) in the study. 3e. File name -
 pmiDiversity.R This script is used to calculate allelic diversity of the
 pollen pool. Modified from the DispersalDiversity R package.
 3f.
 File name - Pollen_Kernel_simulations_2022_01_13_annotated.Rmd. This is an
 R markdown document that contains the code needed to run the pollen
 simulation analyses presented in the results.
   4. Folder name - NmpiAnalysis. 
 4a.
 Folder name - Data. Tab-separated text files containing genotype data in
 NMpi format for each co-flowering density scenario. Used in the Nmpi
 analysis. 4b. File name -
 error_template.txt. Basic template for genotyping errors used in Nmpi
 analysis. 4c. Folder name -
 info_files. Contains the info files required in NMpi analysis for each
 co-flowering density scenario and pollen dispersal kernel. Each
 combination has 2000 info files with randomly generated initial
 values. 
 4d. File name - info_template.txt. Basic template for
 info.txt file used in Nmpi analysis. 4e. File name -
 NMPi_analysis 2022_01_13.Rmd. Annotated R markdown file for generating
 info.txt files and running Nmpi analysis from the command line. Note that
 the Nmpi analysis will only run on Windows computers.
 4f.
 File name - NMpi.exe. NMpi executable called from NMPi_analysis
 2022_01_13.Rmd. See Chybicki 2018 for description: DOI:
 10.1111/1755-0998.12710 4g. Folder name -
 Results. Tab separated files containing the collated results of the Nmpi
 analysis for each co-flowering density scenario + pollen dispersal kernel
 combination. These values were summarized and collated using the windowed
 version of Nmpi. 4h. File name -
 selection_template.txt. Basic template for selection.txt used in Nmpi
 analysis 4i. Folder name -
 temp. Temporary directory to hold the intermediate files of the Nmpi
 analysis. 4j. File name -
 wNMpi.exe. Executable to open windowed version of Nmpi. See Chybicki 2018
 for description: DOI: 10.1111/1755-0998.12710
   5. Folder name - Data. 
 5a.
 File name - Adult_Genotypes_UTMs.csv. This file contains all adult
 O.bataua genotypes and coordinate information for each adult (UTM1,
 UTM2). 
 5ai. Variables: ID is the genetic sample ID for each
 individual Field_Tag is the field
 ID for each individual UTM1 is first
 coordinate UTM2 is the second
 coordinate Mother? Is a yes/no
 detailing if that individual is a maternal tree
 The
 following columns are microsatellite alleles.  5b. File name -
 Offspring_Genotypes.csv. This is the file that contain all offspring
 genotypes use in the analyses.  5bi. Variables:
 Field_Number is the offspring field ID  Mother_Tag is the
 field ID for the maternal tree Mother_ID is the
 genetic sample ID of the mother tree Failed_alleles are the
 number of alleles for that individual that were not genotyped
 The
 remaining columns are the genotypes for each allele.