from os.path import expanduser, isdir, isfile, sep, splitext import click from numpy import ndarray, unique from scipy.sparse.csc import csc_matrix @click.command() @click.argument("data", nargs=1, metavar="", required=True) @click.option("--layout", "-l", default=["umap", "tsne"], multiple=True, type=click.Choice(["umap", "tsne"]), help="Layout algorithm", show_default=True) @click.option("--recipe", "-r", default="none", type=click.Choice(["none", "seurat", "zheng17"]), help="Preprocessing to run.", show_default=True) @click.option("--output", "-o", default="", help="Save a new file to filename.", metavar="") @click.option("--plotting", "-p", default=False, is_flag=True, help="Whether to generate plots.", show_default=True) @click.option("--sparse", default=False, is_flag=True, help="Whether to force sparsity.", show_default=True) @click.option("--overwrite", default=False, is_flag=True, help="Allow file overwriting.", show_default=True) @click.option("--set-obs-names", default="", help="Named field to set as index for obs.", metavar="") @click.option("--set-var-names", default="", help="Named field to set as index for var.", metavar="") @click.option("--make-obs-names-unique", default=True, is_flag=True, help="Ensure obs index is unique.", show_default=True) @click.option("--make-var-names-unique", default=True, is_flag=True, help="Ensure var index is unique.", show_default=True) def prepare(data, layout, recipe, output, plotting, sparse, overwrite, set_obs_names, set_var_names, make_obs_names_unique, make_var_names_unique): """Preprocesses data for use with cellxgene. This tool runs a series of scanpy routines for preparing a dataset for use with cellxgene. It loads data from different formats (h5ad, loom, or a 10x directory), runs dimensionality reduction, computes nearest neighbors, computes a layout, performs clustering, and saves the results. Includes additional options for naming annotations, ensuring sparsity, and plotting results.""" # collect slow imports here to make CLI startup more responsive click.echo("[cellxgene] Starting CLI...") import matplotlib matplotlib.use("Agg") import scanpy.api as sc # scanpy settings sc.settings.verbosity = 0 sc.settings.autosave = True # check args if sparse and not recipe == "none": raise click.UsageError("Cannot use a recipe when forcing sparsity") output = expanduser(output) if not output: click.echo("Warning: No file will be saved, to save the results of cellxgene prepare include " "--output to save output to a new file") if isfile(output) and not overwrite: raise click.UsageError(f"Cannot overwrite existing file {output}, try using the flag --overwrite") def load_data(data): if isfile(data): name, extension = splitext(data) if extension == ".h5ad": adata = sc.read_h5ad(data) elif extension == ".loom": adata = sc.read_loom(data) else: raise click.FileError(data, hint="does not have a valid extension [.h5ad | .loom]") elif isdir(data): if not data.endswith(sep): data += sep adata = sc.read_10x_mtx(data) else: raise click.FileError(data, hint="not a valid file or path") if not set_obs_names == "": if set_obs_names not in adata.obs_keys(): raise click.UsageError(f"obs {set_obs_names} not found, options are: {adata.obs_keys()}") adata.obs_names = adata.obs[set_obs_names] if not set_var_names == "": if set_var_names not in adata.var_keys(): raise click.UsageError(f"var {set_var_names} not found, options are: {adata.var_keys()}") adata.var_names = adata.var[set_var_names] if make_obs_names_unique: adata.obs_names_make_unique() if make_var_names_unique: adata.var_names_make_unique() if not adata._obs.index.is_unique: click.echo("Warning: obs index is not unique") if not adata._var.index.is_unique: click.echo("Warning: var index is not unique") return adata def make_sparse(adata): if (type(adata.X) is ndarray) and sparse: adata.X = csc_matrix(adata.X) def run_recipe(adata): if recipe == "seurat": sc.pp.recipe_seurat(adata) elif recipe == "zheng17": sc.pp.recipe_zheng17(adata) else: sc.pp.filter_cells(adata, min_genes=5) sc.pp.filter_genes(adata, min_cells=25) if sparse: sc.pp.scale(adata, zero_center=False) else: sc.pp.scale(adata) def run_pca(adata): if sparse: sc.pp.pca(adata, svd_solver="arpack", zero_center=False) else: sc.pp.pca(adata, svd_solver="arpack") def run_neighbors(adata): sc.pp.neighbors(adata) def run_louvain(adata): try: sc.tl.louvain(adata) except ModuleNotFoundError: click.echo("\nWarning: louvain module is not installed, no clusters will be calculated. " "To fix this please install cellxgene with the optional feature louvain enabled: " "`pip install cellxgene[louvain]`") def run_layout(adata): if len(unique(adata.obs["louvain"].values)) < 10: palette = "tab10" else: palette = "tab20" if "umap" in layout: sc.tl.umap(adata) if plotting: sc.pl.umap(adata, color="louvain", palette=palette, save="_louvain") if "tsne" in layout: sc.tl.tsne(adata) if plotting: sc.pl.tsne(adata, color="louvain", palette=palette, save="_louvain") def show_step(item): names = { "make_sparse": "Ensuring sparsity", "run_recipe": f"Running preprocessing recipe \"{recipe}\"", "run_pca": "Running PCA", "run_neighbors": "Calculating neighbors", "run_louvain": "Calculating clusters", "run_layout": "Computing layout" } if item is not None: return names[item.__name__] steps = [make_sparse, run_recipe, run_pca, run_neighbors, run_louvain, run_layout] click.echo(f"[cellxgene] Loading data from {data}, please wait...") adata = load_data(data) click.echo("[cellxgene] Beginning preprocessing...") with click.progressbar(steps, label="[cellxgene] Progress", show_eta=False, item_show_func=show_step) as bar: for step in bar: step(adata) # saving if not output == "": click.echo(f"[cellxgene] Saving results to {output}...") adata.write(output) click.echo("[cellxgene] Success!")