Add P-values to GGPLOT Facets with Different Scales

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27 Jun

Add P-values to GGPLOT Facets with Different Scales

Alboukadel

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FAQ

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This article describes how to compute and automatically add p-values onto ggplot facets with different scales using the ggpubr and the rstatix R packages. For multipanel plots with approximately similar y-axis scales on each panel, you can follow steps described in this article: How to Add P-values to GGPLOT Facets.

Here, we’ll We’ll use a demo data for creating panels of plots with very different y scales. You will learn how correctly auto-compute the y positions of the p-values when the facet scales is set to free option. Examples are shown for box plots and bar plots.



Contents:

Prerequisites

Read the following related article: How to Add P-values to GGPLOT Facets.

Make sure you have installed the following R packages:

  • tidyverse for data manipulation and visualization
  • ggpubr for creating easily publication ready plots
  • rstatix provides pipe-friendly R functions for easy statistical analyses.

Start by loading the following required packages:

library(ggpubr)
library(rstatix)

Data preparation

# Transform `dose` into factor variable
df <- ToothGrowth
df$dose <- as.factor(df$dose)
# Add a random grouping variable
df$group <- factor(rep(c("grp1", "grp2"), 30))
# Add some extremely high values in column 1 at rows c(1, 3, 5).
df[c(1, 3, 5),  1] <- c(500, 495, 505)
head(df, 3)
##     len supp dose group
## 1 500.0   VC  0.5  grp1
## 2  11.5   VC  0.5  grp2
## 3 495.0   VC  0.5  grp1

Statistical tests

Facet by the supp and group variables, and compare the levels of the dose variable on the x-axis. The Tukey post hoc test is used for the pairwise comparisons.

stat.test <- df %>%
  group_by(group, supp) %>%
  tukey_hsd(len ~ dose) 
stat.test 
## # A tibble: 12 x 11
##   supp  group term  group1 group2 null.value estimate conf.low conf.high    p.adj p.adj.signif
## * <fct> <fct> <chr> <chr>  <chr>       <dbl>    <dbl>    <dbl>     <dbl>    <dbl> <chr>       
## 1 OJ    grp1  dose  0.5    1               0     6.32    0.275      12.4 0.0403   *           
## 2 OJ    grp1  dose  0.5    2               0    11.3     5.26       17.3 0.000852 ***         
## 3 OJ    grp1  dose  1      2               0     4.98   -1.06       11.0 0.112    ns          
## 4 VC    grp1  dose  0.5    1               0  -286.   -548.        -23.5 0.0328   *           
## 5 VC    grp1  dose  0.5    2               0  -276.   -539.        -14.0 0.0389   *           
## 6 VC    grp1  dose  1      2               0     9.46 -253.        272.  0.995    ns          
## # … with 6 more rows

Facet with fixed scales

# Create bar plots with significance levels
# Hide ns (non-significant)
# Add 15% space between labels and the plot top border
stat.test <- stat.test %>% add_xy_position(x = "dose", fun = "mean_se")
ggbarplot(
  df, x = "dose", y = "len", fill = "#00AFBB",
  add = "mean_se", facet = c("supp", "group")
  ) +
  stat_pvalue_manual(stat.test, hide.ns = TRUE, tip.length = 0, step.increase = 0) +
  scale_y_continuous(expand = expansion(mult = c(0.05, 0.15)))

Facet with free scales

Facet wrap

You need to specify the option scales = "free" in both the add_xy_position() and in the ggbarplot() functions.

stat.test <- stat.test %>% 
  add_xy_position(x = "dose", fun = "mean_se", scales = "free")
ggbarplot(
  df, x = "dose", y = "len", fill = "#00AFBB",
  add = "mean_se", facet.by = c("supp", "group")
  ) +
  facet_wrap(vars(supp, group), scales = "free") +
  stat_pvalue_manual(stat.test, hide.ns = TRUE, tip.length = 0) +
  scale_y_continuous(expand = expansion(mult = c(0.05, 0.15)))

Facet grid

When the scales = "free" argument is added in facet grid, plots on the same row cannot have different y-axis. Similarly, there can be only single x-axis for each column. Using facet_wrap(), each plot is displayed independently, so it can “free” its x-axis and y-axis.

Facet grid is useful when you want to relatively compare the plots within a category, which can be accomplished by setting the same axis scales. Meanwhile, facet wrap is more useful for plots that are more independent between one another.

There are two possible solutions to customize the y position of significance levels.

Solution 1: Using the option step.increase

The default of the function add_xy_position() is to automatically compute a global step increase value between brackets. This calculation assumes that the y scales of plot panels are fixed.

In the situation, where you want free scales, you can:

  1. Set the option step.increase to 0 when calling the function add_xy_position().
  2. Specify only the option step.increase in the function stat_pvalue_manual(). In this case, the step.increase will be adapted to each plot panel.
stat.test <- stat.test %>% 
  add_xy_position(x = "dose", fun = "mean_se", step.increase = 0)
bp <- ggbarplot(
  df, x = "dose", y = "len", fill = "#00AFBB", add = "mean_se",
  facet.by = c("supp", "group"), scales = "free"
  ) 
bp +
  stat_pvalue_manual(stat.test, hide.ns = TRUE, tip.length = 0, step.increase = 0.2) +
  scale_y_continuous(expand = expansion(mult = c(0.05, 0.15)))

Solution 2: Using the option scales = “free”.

In facet grid, the scales of the generated plot panels are not completely free. Consequently you will need more customization to adapt the look of the significance level position. You will have to play with the options step.increase and bracket.nudge.y in the stat_pvalue_manual() function.

# Default plot
stat.test <- stat.test %>% 
  add_xy_position(x = "dose", fun = "mean_se", scales = "free")
bp <- ggbarplot(
  df, x = "dose", y = "len", fill = "#00AFBB", add = "mean_se",
  facet.by = c("supp", "group"), scales = "free"
  ) 
bp +
  stat_pvalue_manual(stat.test, hide.ns = TRUE, tip.length = 0) +
  scale_y_continuous(expand = expansion(mult = c(0.05, 0.15)))

# Increase the step between the brackets
bp +
  stat_pvalue_manual(
    stat.test, hide.ns = TRUE, tip.length = 0,
    step.increase = 0.1
    ) +
  scale_y_continuous(expand = expansion(mult = c(0.05, 0.15)))

# Nudge down the brackets
# Specify the option bracket.nudge.y for each of the panel
# Should be of the same length as the number of comparisons
bracket.nudge.y <- c(
  -2, -3,               # Panel 1: grp1/OJ
  -100, -160,           # Panel 2: grp1/VC
  -10, -11,             # Panel 3: grp2/OJ
  -250, -250, -250      # Panel 4: grp2/VC
  )
bp +
  stat_pvalue_manual(
    stat.test, hide.ns = TRUE, tip.length = 0,
    step.increase = 0.09, bracket.nudge.y = bracket.nudge.y
    ) +
  scale_y_continuous(expand = expansion(mult = c(0.05, 0.15)))

Conclusion

This article describes how to add p-values onto ggplot facets with different y scales. See other related frequently questions: ggpubr FAQ.



 
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Comments ( 12 )

  • Venus
    18 Jul 2020

    This will be very helpful. However, the rstatix version 0.6.0 I just installed returns “unused argument (scales = “free”)” error message. Is there a newer version that takes the scales argument?

    Reply
    • Kassambara
      20 Jul 2020

      This option is available in the latest dev version (not on CRAN yet), which can be installed as follow: devtools::install_github(“kassambara/rstatix”)

      Reply
      • Daniel Petersheim
        27 Aug 2020

        Hi,
        the new scales = “free” argument seems to be a very welcome new feature. However, when I try to install the latest version from
        devtools::install_github(“kassambara/rstatix”), I get the following message:

        Error: Failed to install ‘rstatix’ from GitHub:
        (converted from warning) installation of package ‘/var/folders/fl/lrh0tkf16_j29g5lgw6vsvpw0000gn/T//RtmpI4YwFc/fileb2e03c48f23b/rstatix_0.6.0.999.tar.gz’ had non-zero exit status

        Any suggestions would be highly appreciated.

        Thanks a lot 🙂

        Daniel

        Reply
  • Pumbaa
    19 Jul 2020

    With the v0.6.0 rstatix I just installed (I’m using R-3.6.1), the “facet with free scales” example code does not work. The line stat.test <- stat.test %>% add_xy_position(x = "dose", fun = "mean_se", scales = "free") returns error message:
    Error in add_y_position(., fun = fun, step.increase = step.increase, stack = stack, : unused argument (scales = "free")

    Reply
    • Kassambara
      20 Jul 2020

      This option is available in the latest dev version (not on CRAN yet), which can be installed as follow: devtools::install_github(“kassambara/rstatix”)

      Reply
      • Pumbaa
        20 Jul 2020

        Thanks. I thought my first post didn’t go through. Please delete the first duplicate.

        Reply
      • Pumbaa
        20 Jul 2020

        Thanks. One of the dependencies of the rstatix installation, broom v0.7.0, seemed to have incompatibility issue and causing problems on other functions. Is v0.7.0 needed? Is there a way to specify using lower versions like 0.5.2? Thanks!

        Reply
  • Pumbaa
    20 Jul 2020

    Thanks. One of the dependencies of the rstatix installation, broom v0.7.0, seemed to have incompatibility issue and causing problems on other functions. Is v0.7.0 needed? Is there a way to specify using lower versions like 0.5.2? Thanks!

    Reply
  • Ezequiel
    08 Dec 2022

    Hello! because I used scale_x_discrete to re order the factors on the x axis, now the Tukey results are not in the right order when using stat_pvalue_manual. Is there a solution to plot in the right order? Thanks a lot!

    Reply
    • Alboukadel
      08 Dec 2022

      Hello!

      To avoid this issue, the easiest solution is to modify your data by defining the grouping variable level order, using R code like this:

      my_data[["my_grouping_variable"]] = factor( my_data[["my_grouping_variable"]], levels = c("grp1", "grp2", "grp3"))

      Then, use the data to do statistical tests and plots. In this case, you don’t need scale_x_discrete().

      Reply
  • Dqt
    22 Feb 2023

    Hi!

    How can I add add p-values generated elsewhere to a facet_wrap with different scales? How can I set the y.position to be 1.1 times bigger than the maximum value in each panel?

    The tutorials show how to do that with p-values obtained from the rstatix package, but I want to use already computed p-values.

    Thank you very much!

    Reply
  • Angela
    25 Mar 2023

    Hi, tried running this with my data:

    stat.test % add_xy_position(x = “dose”, fun = “mean_se”)

    where dose in my case is “A” vs “B” for 11 species (supp) at 4 timepoint categories (group)

    and I get this error message:

    Error in `tidyr::unnest()`:
    ! Can’t duplicate names between the affected columns and the original data.
    ✖ These names are duplicated:
    ℹ `variable`, from `data`.
    ℹ Use `names_sep` to disambiguate using the column name.
    ℹ Or use `names_repair` to specify a repair strategy.
    Run `rlang::last_error()` to see where the error occurred.

    have you seen this before?

    Reply

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