Package 'SpotSweeper' reference manual

Title:	Spatially-aware quality control for spatial transcriptomics
Description:	Spatially-aware quality control (QC) software for both spot-level and artifact-level QC in spot-based spatial transcripomics, such as 10x Visium. These methods calculate local (nearest-neighbors) mean and variance of standard QC metrics (library size, unique genes, and mitochondrial percentage) to identify outliers spot and large technical artifacts. Scales linearly with the number of spots and is designed to be used with 'SpatialExperiment' objects.
Authors:	Michael Totty [aut, cre] , Boyi Guo [aut]
Maintainer:	Michael Totty <[email protected]>
License:	MIT + file LICENSE
Version:	1.1.0
Built:	2024-09-30 04:54:07 UTC
Source:	https://github.com/bioc/SpotSweeper

human DLPFC dataset with a technical artifact (hangnail).

Description

The DLPFC_artifact dataset is a SpatialExperiment object containing a single-sample subset of the human dorsolateral prefrontal cortex (DLPFC) dataset from Hukki-Myers et al. 2023. This particular sample ('Br2743_ant') is included to demonstrate the identification and removal of technical artifacts within spatial transcriptomics data. The dataset serves as an example for artifact detection using the 'SpotSweeper' workflow.

Usage

data(DLPFC_artifact)
data(DLPFC_artifact)

Format

An SpatialExperiment object.

Source

spatialLIBD

References

Hukki-Myers et al. (2023) bioRxiv (bioRxiv)

Examples

data(DLPFC_artifact)
data(DLPFC_artifact)

Identify and annotate artifacts in spatial transcriptomics data

Description

This function identifies and annotates potential artifacts in spatial transcriptomics data. Artifacts are detected based on local mito variance, and the results are added to the original SpatialExperiment (sce) object.

Usage

findArtifacts(
  spe,
  mito_percent = "expr_chrM_ratio",
  mito_sum = "expr_chrM",
  samples = "sample_id",
  n_rings = 5,
  log = TRUE,
  name = "artifact",
  var_output = TRUE
)
findArtifacts(
  spe,
  mito_percent = "expr_chrM_ratio",
  mito_sum = "expr_chrM",
  samples = "sample_id",
  n_rings = 5,
  log = TRUE,
  name = "artifact",
  var_output = TRUE
)

Arguments

`spe`	A SingleCellExperiment object.
`mito_percent`	The column name representing the mitochondrial percent. Default is 'expr_chrM_ratio'.
`mito_sum`	The column name representing sum mitochondrial expression. Default is 'expr_chrM'.
`samples`	The column name representing sample IDs. Default is 'sample_id'.
`n_rings`	The number of rings for local mito variance calculation. Default is 5.
`log`	Logical, indicating whether to log1p transform mito_percent. Default is TRUE.
`name`	Prefix for the local variance column names. Default is 'artifact'.
`var_output`	Logical, indicating whether to include local variances in the output. Default is TRUE.

Value

Returns the modified SingleCellExperiment object with artifact annotations.

Examples

library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

data(DLPFC_artifact)
spe <- DLPFC_artifact

# find artifacts
spe <- findArtifacts(spe,
    mito_percent = "expr_chrM_ratio",
    mito_sum = "expr_chrM",
    n_rings = 2, # 5 recommended, using 1 for time
    name = "artifact"
)


library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

data(DLPFC_artifact)
spe <- DLPFC_artifact

# find artifacts
spe <- findArtifacts(spe,
    mito_percent = "expr_chrM_ratio",
    mito_sum = "expr_chrM",
    n_rings = 2, # 5 recommended, using 1 for time
    name = "artifact"
)

localOutliers Function

Description

This function detects local outliers in spatial transcriptomics data based on standard quality control metrics, such as library size, unique genes, and mitochondrial ratio. Local outliers are defined as spots with low/high quality metrics compared to their surrounding neighbors, based on a modified z-score statistic.

Usage

localOutliers(
  spe,
  metric = "detected",
  direction = "lower",
  n_neighbors = 36,
  samples = "sample_id",
  log = TRUE,
  cutoff = 3
)
localOutliers(
  spe,
  metric = "detected",
  direction = "lower",
  n_neighbors = 36,
  samples = "sample_id",
  log = TRUE,
  cutoff = 3
)

Arguments

`spe`	SpatialExperiment object
`metric`	colData QC metric to use for outlier detection
`direction`	Direction of outlier detection (higher, lower, or both)
`n_neighbors`	Number of nearest neighbors to use for outlier detection
`samples`	Column name in colData to use for sample IDs
`log`	Logical indicating whether to log1p transform the features (default is TRUE)
`cutoff`	Cutoff for outlier detection (default is 3)

Value

SpatialExperiment object with updated colData containing outputs

Examples

library(SpotSweeper)
library(SpatialExperiment)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metrics for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

# Identifying local outliers using SpotSweeper
spe <- localOutliers(spe,
                     metric = "sum",
                     direction = "lower",
                     log = TRUE
)

library(SpotSweeper)
library(SpatialExperiment)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metrics for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

# Identifying local outliers using SpotSweeper
spe <- localOutliers(spe,
                     metric = "sum",
                     direction = "lower",
                     log = TRUE
)

localVariance Function

Description

This function does calculates the local variance based on kNN.

Usage

localVariance(
  spe,
  n_neighbors = 36,
  metric = c("expr_chrM_ratio"),
  samples = "sample_id",
  log = FALSE,
  name = NULL
)
localVariance(
  spe,
  n_neighbors = 36,
  metric = c("expr_chrM_ratio"),
  samples = "sample_id",
  log = FALSE,
  name = NULL
)

Arguments

`spe`	SpatialExperiment object with the following columns in colData: sample_id, sum_umi, sum_gene
`n_neighbors`	Number of nearest neighbors to use for variance calculation
`metric`	metric to use for variance calculation
`samples`	Column in colData to use for sample ID
`log`	Whether to log1p transform the metric
`name`	Name of the new column to add to colData

Value

SpatialExperiment object with metric variance added to colData

Examples


# for more details see extended example in vignettes
library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# show column data before SpotSweepR
colnames(colData(spe))

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metric for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

spe <- localVariance(spe,
    metric = "subsets_Mito_percent",
    n_neighbors = 36,
    name = "local_mito_variance_k36"
    )

plotQC(spe, metric="local_mito_variance_k36")

# for more details see extended example in vignettes
library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# show column data before SpotSweepR
colnames(colData(spe))

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metric for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

spe <- localVariance(spe,
    metric = "subsets_Mito_percent",
    n_neighbors = 36,
    name = "local_mito_variance_k36"
    )

plotQC(spe, metric="local_mito_variance_k36")

Plot QC metrics for a Single Sample in a SpatialExperiment object

Description

This function generates a plot for a specified sample within a SpatialExperiment object, highlighting outliers based on a specified metric. The plot visualizes the metric of interest and indicates outliers with a distinct color.

Usage

plotQC(
  spe,
  sample_id = "sample_id",
  sample = unique(spe$sample_id)[1],
  metric = "detected",
  outliers = NULL,
  point_size = 2,
  colors = c("white", "black"),
  stroke = 1
)
plotQC(
  spe,
  sample_id = "sample_id",
  sample = unique(spe$sample_id)[1],
  metric = "detected",
  outliers = NULL,
  point_size = 2,
  colors = c("white", "black"),
  stroke = 1
)

Arguments

`spe`	A SpatialExperiment object containing the data to be plotted.
`sample_id`	A character string specifying the column name in `colData(spe)` that contains unique sample identifiers. Default is "sample_id".
`sample`	A character string or numeric value specifying the sample to be plotted. By default, it plots the first unique sample found in `spe$sample_id`.
`metric`	A character string specifying the metric to be visualized in the plot. This metric should be a column name in `colData(spe)`.
`outliers`	A character string specifying the column name in `colData(spe)` that indicates whether a data point is considered an outlier. Default is NULL.
`point_size`	A numeric value specifying the size of the points in the plot. Default is 2.
`colors`	A character vector specifying the colors to be used for the gradient scale. If length is 2, the gradient will be a single color gradient.
`stroke`	A numeric value specifying the border thickness for outlier points. Default is 1.

Value

The function returns a plot object created by make_escheR and modified with additional layers for visualizing the specified metric and outliers. The plot is not explicitly printed by the function and should be printed by the caller.

Examples

library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metrics for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

# Identifying local outliers using SpotSweeper
spe <- localOutliers(spe,
                     metric = "sum",
                     direction = "lower",
                     log = TRUE
)

plotQC(spe, metric="sum", outliers="sum_outliers")

library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metrics for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

# Identifying local outliers using SpotSweeper
spe <- localOutliers(spe,
                     metric = "sum",
                     direction = "lower",
                     log = TRUE
)

plotQC(spe, metric="sum", outliers="sum_outliers")

Plot Outlier Metrics to PDF

Description

This function generates a PDF file containing plots for each sample in the SpatialExperiment object, highlighting outliers based on specified metrics. Each plot visualizes outlier metrics for a single sample, allowing for easy comparison and analysis across samples.

Usage

plotQCpdf(
  spe,
  sample_id = "sample_id",
  metric = "detected",
  outliers = "local_outliers",
  colors = c("white", "black"),
  stroke = 1,
  point_size = 2,
  width = 5,
  height = 5,
  fname
)
plotQCpdf(
  spe,
  sample_id = "sample_id",
  metric = "detected",
  outliers = "local_outliers",
  colors = c("white", "black"),
  stroke = 1,
  point_size = 2,
  width = 5,
  height = 5,
  fname
)

Arguments

`spe`	A SpatialExperiment object containing the data to be plotted.
`sample_id`	A character string specifying the column name in `colData(spe)` that contains unique sample identifiers. Default is 'sample_id'.
`metric`	A character string specifying the metric to be visualized in the plot. This metric should be a column name in `colData(spe)`.
`outliers`	A character string specifying the column name in `colData(spe)` that indicates whether a data point is considered an outlier. Default is local_outliers'.
`colors`	A character vector specifying the colors to be used for the gradient scale. If length is 2, the gradient will be a single color gradient
`stroke`	A numeric value specifying the border thickness for outlier points. Default is 1.
`point_size`	A numeric value specifying the size of the points in the plot. Default is 2.
`width`	A numeric value indicating the width of the plot. Default is 5.
`height`	A numeric value indicating the height of the plot. Default is 5.
`fname`	A character string specifying the path and name of the output PDF file.

Value

ggplot object if specified. Generates a plot otherwise.

Examples

library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

tempFilePath <- file.path(tempdir(), "examplePlot.pdf")

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metrics for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

# Identifying local outliers using SpotSweeper
spe <- localOutliers(spe,
                     metric = "sum",
                     direction = "lower",
                     log = TRUE
)

plotQCpdf(spe,
          metric="sum",
          outliers="sum_outliers",
          fname=tempFilePath)

library(SpotSweeper)
library(SpatialExperiment)
library(escheR)

# load example data
spe <- STexampleData::Visium_humanDLPFC()

tempFilePath <- file.path(tempdir(), "examplePlot.pdf")

# change from gene id to gene names
rownames(spe) <- rowData(spe)$gene_name

# drop out-of-tissue spots
spe <- spe[, spe$in_tissue == 1]
spe <- spe[, !is.na(spe$ground_truth)]

# Identifying the mitochondrial transcripts in our SpatialExperiment.
is.mito <- rownames(spe)[grepl("^MT-", rownames(spe))]

# Calculating QC metrics for each spot using scuttle
spe <- scuttle::addPerCellQCMetrics(spe, subsets = list(Mito = is.mito))
colnames(colData(spe))

# Identifying local outliers using SpotSweeper
spe <- localOutliers(spe,
                     metric = "sum",
                     direction = "lower",
                     log = TRUE
)

plotQCpdf(spe,
          metric="sum",
          outliers="sum_outliers",
          fname=tempFilePath)

Package 'SpotSweeper'

Help Index

human DLPFC dataset with a technical artifact (hangnail).

Description

Usage

Format

Source

References

Examples

Identify and annotate artifacts in spatial transcriptomics data

Description

Usage

Arguments

Value

See Also

Examples

localOutliers Function

Description

Usage

Arguments

Value

Examples

localVariance Function

Description

Usage

Arguments

Value

Examples

Plot QC metrics for a Single Sample in a SpatialExperiment object

Description

Usage

Arguments

Value

Examples

Plot Outlier Metrics to PDF

Description

Usage

Arguments

Value

Examples