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\begin{document}
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% figures saved in ./figs; create if necessary below
<<echo=FALSE>>=
  figdir <- 'figs'
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@ 
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\title{Taxonomic classification using \shellcommand{pplacer},
  \Rpackage{clst}, and \Rpackage{clstutils}}
\author{Noah Hoffman}
\maketitle

\tableofcontents

\section{Introduction}

This vignette assumes that you have already created a reference
package, and have used it to run \shellcommand{pplacer} against an
alignment of reference sequences. For instruction on performing the
above operations, see the documentation for \shellcommand{pplacer}
here: \url{http://matsen.fhcrc.org/pplacer/}

\section{Input files}

The following input is required (file names defined as variables in parentheses):

\begin{enumerate}
  \item a reference package (\code{refpkg})
  \item a pplacer file created using the same reference package (\code{placefile})
  \item a file providing distances between nodes in the reference tree (\code{distfile})
\end{enumerate}

Note that \code{distfile} is generated from \code{placefile} using
\shellcommand{placeutil} (distributed with \shellcommand{pplacer}).

<<data>>=
library(clstutils)

expand <- function(fname){
  orig.dir <- getwd()
  destdir <- tempdir()
  setwd(destdir)
  archive <- system.file('extdata','vaginal_16s.refpkg.tar.gz', package='clstutils')
  system(sprintf('tar --no-same-owner -xzf "%s"', archive))
  setwd(orig.dir)
  file.path(destdir, fname)
}

refpkg <- expand('vaginal_16s.refpkg')
placefile <- system.file('extdata','merged.json', package='clstutils')
distfile <- system.file('extdata','merged.distmat.bz2', package='clstutils')
@

\section{Reading the input}

Classification requires a matrix representation of distances between
``objects'' being classified, in this case sequences in a phylogenetic
tree. \Rfunction{treeDists} returns a list containing matrix
representations of distances between internal and terminal edges
(\code{\$dists} and \code{\$paths}), and \code{\$dmat}, a square
matrix of distances between terminal edges. 

<<treeDists>>=
treedists <- treeDists(distfile=distfile, placefile=placefile)
@

We also need a description of the taxonomy of the reference
sequences. This is read from the reference package using
\Rfunction{taxonomyFromRefpkg}. The \Rfunarg{seqnames} argument
ensures that the output is arranged in an order compatible with
\code{treedists}. We indicate that the most specific rank that we want
to consider is ``species'' using \Rfunarg{lowest\_rank}.

<<taxonomyFromRefpkg>>=
taxdata <- taxonomyFromRefpkg(refpkg, seqnames=rownames(treedists$dmat), lowest_rank='species')
@

\section{Classification of a single sequence}

Given the distances and taxonomic information describing the reference
tree, the only additional data required to perform classification is
the position of a sequence placed onto a tree. At a minimum, this
consists of a data.frame with columns \code{at}, \code{edge}, and
\code{branch}. This data will be used to generate a vector of branch
lengths between the query sequence and each of the reference sequences
on the tree.

<<>>=
placetab <- data.frame(at=49, edge=5.14909e-07, branch=5.14909e-07)
@ 

The function \Rfunction{classifyPlacements} is a wrapper around
\Rpackage{clst}::\Rfunction{classifyIter}. The output is a
\Robject{data.frame} describing the taxonomic assignment, along with a
description of the confidence of the classification. See the man page
for \Rpackage{clst}::\Rfunction{classify} for details on the output.

<<>>=
cdata <- classifyPlacements(taxdata, treedists, placetab)
cdata
@

% \section{Classification of multiple sequences}

% Placement information for multiple sequences may be contained in a
% placefile. This information can be read directly from a single
% placefile using \Rfunction{placeData}.

% <<>>=
% placetab <- placeData(placefile)
% head(placetab)
% @ 

% Note that pplacer also provides tax\_id assignments using a different
% method. Also, a list of possible placements are provided for each
% sequence accompanied by the probability of each. We will use only the
% first placement (ie, the top ``hit'') for each sequence.

% <<>>=
% tophits <- subset(placetab, hit==1)
% set.seed(125)
% cdata <- classifyPlacements(taxdata, treedists, 
%                             tophits[sample(nrow(tophits),10),])
% cdata
% @ 

\end{document}
