# Advance Tree Annotation

####
Guangchuang Yu (guangchuangyu@gmail.com) and Tommy Tsan-Yuk Lam (ttylam@hku.hk)
School of Public Health, The University of Hong Kong

#### 2017-03-17

# Visualize tree with associated matrix

The `gheatmap` function is designed to visualize phylogenetic tree with heatmap of associated matrix.

In the following example, we visualized a tree of H3 influenza viruses with their associated genotype.

    beast_file <- system.file("examples/MCC_FluA_H3.tree", package="ggtree")
    beast_tree <- read.beast(beast_file)

    genotype_file <- system.file("examples/Genotype.txt", package="ggtree")
    genotype <- read.table(genotype_file, sep="\t", stringsAsFactor=F)
    p <- ggtree(beast_tree, mrsd="2013-01-01") + geom_treescale(x=2008, y=1)
    p <- p + geom_tiplab(size=3)
    gheatmap(p, genotype, offset = 2, width=0.5)

The width parameter is to control the width of the heatmap. It supports another parameter offset for controlling the distance between the tree and the heatmap, for instance to allocate space for tip labels.

For time-scaled tree, as in this example, it’s more often to use x axis by using `theme_tree2`. But with this solution, the heatmap is just another layer and will change the `x` axis. To overcome this issue, we implemented `scale_x_ggtree` to set the x axis more reasonable.

    p <- ggtree(beast_tree, mrsd="2013-01-01") + geom_tiplab(size=3, align=TRUE) + theme_tree2()
    pp <- (p + scale_y_continuous(expand=c(0, 0.3))) %>%
        gheatmap(genotype, offset=4, width=0.5, colnames=FALSE) %>%
            scale_x_ggtree()
    pp + theme(legend.position="right")

# Visualize tree with multiple sequence alignment

With `msaplot` function, user can visualize multiple sequence alignment with phylogenetic tree, as demonstrated below:

    fasta <- system.file("examples/FluA_H3_AA.fas", package="ggtree")
    msaplot(ggtree(beast_tree), fasta)

    ## Warning: Ignoring unknown aesthetics: x, y

A specific slice of the alignment can also be displayed by specific window parameter.

    msaplot(ggtree(beast_tree), fasta, window=c(150, 200)) + coord_polar(theta='y')

    ## Warning: Ignoring unknown aesthetics: x, y

# Annotate a phylogenetic tree with insets

`ggtree` provides a function, `inset`, for adding subplots to a phylogenetic tree. The input is a tree graphic object and a named list of ggplot graphic objects (can be any kind of charts), these objects should named by node numbers. To facilitate adding bar and pie charts (e.g. summarized stats of results from ancestral reconstruction) to phylogenetic tree, ggtree provides `nodepie` and `nodebar` functions to create a list of pie or bar charts.

## Annotate with bar charts

    set.seed(2015-12-31)
    tr <- rtree(15)
    p <- ggtree(tr)

    a <- runif(14, 0, 0.33)
    b <- runif(14, 0, 0.33)
    c <- runif(14, 0, 0.33)
    d <- 1 - a - b - c
    dat <- data.frame(a=a, b=b, c=c, d=d)
    ## input data should have a column of `node` that store the node number
    dat$node <- 15+1:14

    ## cols parameter indicate which columns store stats (a, b, c and d in this example)
    bars <- nodebar(dat, cols=1:4)

    inset(p, bars)

The sizes of the insets can be ajusted by the paramter width and height.

    inset(p, bars, width=.06, height=.1)

Users can set the color via the parameter color. The x position can be one of ‘node’ or ‘branch’ and can be adjusted by the parameter hjust and vjust for horizontal and vertical adjustment respecitvely.

    bars2 <- nodebar(dat, cols=1:4, position='dodge',
                     color=c(a='blue', b='red', c='green', d='cyan'))
    p2 <- inset(p, bars2, x='branch', width=.06, vjust=-.3)
    print(p2)

## Annotate with pie charts

Similarly, users can use `nodepie` function to generate a list of pie charts and place these charts to annotate corresponding nodes. Both `nodebar` and `nodepie` accepts parameter alpha to allow transparency.

    pies <- nodepie(dat, cols=1:4, alpha=.6)
    inset(p, pies)

    inset(p, pies, hjust=-.06)

## Annotate with other types of charts

The `inset` function accepts a list of ggplot graphic objects and these input objects are not restricted to pie or bar charts. They can be any kinds of charts and hybrid of these charts.

    pies_and_bars <- bars2
    pies_and_bars[9:14] <- pies[9:14]
    inset(p, pies_and_bars)

    d <- lapply(1:15, rnorm, n=100)
    ylim <- range(unlist(d))
    bx <- lapply(d, function(y) {
        dd <- data.frame(y=y)
        ggplot(dd, aes(x=1, y=y))+geom_boxplot() + ylim(ylim) + theme_inset()
    })
    names(bx) <- 1:15
    inset(p, bx, width=.06, height=.2, hjust=-.05)

After annotating with insets, users can further annotate the tree with another layer of insets.

    p2 <- inset(p, bars2, x='branch', width=.03, vjust=-.4)
    p2 <- inset(p2, pies, x='branch', vjust=.4)
    bx2 <- lapply(bx, function(g) g+coord_flip())
    inset(p2, bx2, width=.4, height=.06, vjust=.04, hjust=p2$data$x[1:15]-4) + xlim(NA, 4.5)

## Annotate with image files

    imgfile <- tempfile(, fileext=".png")
    download.file("https://avatars1.githubusercontent.com/u/626539?v=3&u=e731426406dd3f45a73d96dd604bc45ae2e7c36f&s=140", destfile=imgfile, mode='wb')
    img <- list(imgfile, imgfile)
    names(img) <- c("18", "22")
    inset(p, img)

# Plot tree with associated data

For associating phylogenetic tree with different type of plot produced by user’s data, `ggtree` provides `facet_plot` function which accepts an input `data.frame` and a ‘geom’ function to draw the input data. The data will be displayed in an additional panel of the plot.

    tr <- rtree(30)

    d1 <- data.frame(id=tr$tip.label, val=rnorm(30, sd=3))
    p <- ggtree(tr)

    p2 <- facet_plot(p, panel="dot", data=d1, geom=geom_point, aes(x=val), color='firebrick')
    d2 <- data.frame(id=tr$tip.label, value = abs(rnorm(30, mean=100, sd=50)))

    facet_plot(p2, panel='bar', data=d2, geom=geom_segment, aes(x=0, xend=value, y=y, yend=y), size=3, color='steelblue') + theme_tree2()

# Tree annotation with Phylopic

PhyloPic is a database that stores reusable silhouette images of organisms. `ggtree` supports downloading images from PhyloPic and annotating phylogenetic tree with the downloaded images.

    pp <- ggtree(tree) %>% phylopic("79ad5f09-cf21-4c89-8e7d-0c82a00ce728", color="steelblue", alpha = .3)
    print(pp)

    pp %>% phylopic("67382184-5135-4faa-8e98-eadff02c3e8a", color="#86B875", alpha=.8, node=4) %>%
         phylopic("d3563b54-780f-4711-a49a-7ea051e9dacc", color="darkcyan", alpha=.8, node=17, width=.2)