diff --git a/packages_rs/nextclade-cli/src/cli/nextalign_cli.rs b/packages_rs/nextclade-cli/src/cli/nextalign_cli.rs
index fda391619..56e5cf606 100644
--- a/packages_rs/nextclade-cli/src/cli/nextalign_cli.rs
+++ b/packages_rs/nextclade-cli/src/cli/nextalign_cli.rs
@@ -84,7 +84,15 @@ pub enum NextalignOutputSelection {
 pub struct NextalignRunArgs {
   /// Path to a FASTA file with input sequences
   #[clap(value_hint = ValueHint::FilePath)]
-  pub input_fasta: Vec<PathBuf>,
+  pub input_fastas: Vec<PathBuf>,
+
+  /// REMOVED. Use positional arguments instead.
+  ///
+  /// Example: nextalign run -D dataset/ -O out/ seq1.fasta seq2.fasta
+  #[clap(long, short = 'i', visible_alias("sequences"))]
+  #[clap(value_hint = ValueHint::FilePath)]
+  #[clap(hide_long_help = true, hide_short_help = true)]
+  pub input_fasta: Option<PathBuf>,
 
   /// Path to a FASTA file containing reference sequence.
   ///
@@ -124,6 +132,12 @@ pub struct NextalignRunArgs {
   #[clap(value_hint = ValueHint::FilePath)]
   pub genes: Option<Vec<String>>,
 
+  /// REMOVED. Use `--output-all` instead
+  #[clap(long)]
+  #[clap(value_hint = ValueHint::DirPath)]
+  #[clap(hide_long_help = true, hide_short_help = true)]
+  pub output_dir: Option<PathBuf>,
+
   /// Produce all of the output files into this directory, using default basename and predefined suffixes and extensions. This is equivalent to specifying each of the individual `--output-*` flags. Convenient when you want to receive all or most of output files into the same directory and don't care about their filenames.
   ///
   /// Output files can be optionally included or excluded using `--output-selection` flag.
@@ -252,7 +266,7 @@ fn generate_completions(shell: &str) -> Result<(), Report> {
 /// Get output filenames provided by user or, if not provided, create filenames based on input fasta
 pub fn nextalign_get_output_filenames(run_args: &mut NextalignRunArgs) -> Result<(), Report> {
   let NextalignRunArgs {
-    input_fasta,
+    input_fastas: input_fasta,
     output_all,
     ref mut output_basename,
     ref mut output_errors,
@@ -349,6 +363,47 @@ At least one of the following flags is required:
   Ok(())
 }
 
+const ERROR_MSG_INPUT_FASTA_REMOVED: &str = r#"The argument `--input-fasta` (alias: `--sequences`, `-i`) is removed in favor of positional arguments.
+
+Try:
+
+  nextalign run -r ref.fasta -m genemap.gff -O out/ seq1.fasta seq2.fasta
+
+                                                       ^          ^
+                                              one or multiple positional arguments
+                                                with paths to input fasta files
+
+
+When positional arguments are not provided, nextalign will read input fasta from standard input.
+
+For more information, type
+
+  nextalign run --help"#;
+
+const ERROR_MSG_OUTPUT_DIR_REMOVED: &str = r#"The argument `--output-dir` is removed in favor of `--output-all`.
+
+When provided, `--output-all` allows to write all possible outputs into a directory.
+
+The defaut base name of the files can be overriden with `--output-basename` argument.
+
+The set of output files can be restricted with `--output-selection` argument.
+
+For more information, type:
+
+  nextalign run --help"#;
+
+pub fn nextalign_check_removed_args(run_args: &mut NextalignRunArgs) -> Result<(), Report> {
+  if run_args.input_fasta.is_some() {
+    return make_error!("{ERROR_MSG_INPUT_FASTA_REMOVED}");
+  }
+
+  if run_args.output_dir.is_some() {
+    return make_error!("{ERROR_MSG_OUTPUT_DIR_REMOVED}");
+  }
+
+  Ok(())
+}
+
 pub fn nextalign_parse_cli_args() -> Result<NextalignArgs, Report> {
   let mut args = NextalignArgs::parse();
 
@@ -369,6 +424,7 @@ pub fn nextalign_parse_cli_args() -> Result<NextalignArgs, Report> {
       generate_completions(shell).wrap_err_with(|| format!("When generating completions for shell '{shell}'"))?;
     }
     NextalignCommands::Run(ref mut run_args) => {
+      nextalign_check_removed_args(run_args)?;
       nextalign_get_output_filenames(run_args).wrap_err("When deducing output filenames")?;
     }
   }
diff --git a/packages_rs/nextclade-cli/src/cli/nextalign_loop.rs b/packages_rs/nextclade-cli/src/cli/nextalign_loop.rs
index 8b3223d5f..e1d48b1c0 100644
--- a/packages_rs/nextclade-cli/src/cli/nextalign_loop.rs
+++ b/packages_rs/nextclade-cli/src/cli/nextalign_loop.rs
@@ -23,7 +23,7 @@ pub fn nextalign_run(args: NextalignRunArgs) -> Result<(), Report> {
   info!("Command-line arguments:\n{args:#?}");
 
   let NextalignRunArgs {
-    input_fasta,
+    input_fastas: input_fasta,
     input_ref,
     genes,
     input_gene_map,
@@ -38,6 +38,7 @@ pub fn nextalign_run(args: NextalignRunArgs) -> Result<(), Report> {
     jobs,
     in_order,
     alignment_params: alignment_params_from_cli,
+    ..
   } = args;
 
   let mut alignment_params = AlignPairwiseParams::default();
diff --git a/packages_rs/nextclade-cli/src/cli/nextclade_cli.rs b/packages_rs/nextclade-cli/src/cli/nextclade_cli.rs
index 8be024706..19b1aa8f1 100644
--- a/packages_rs/nextclade-cli/src/cli/nextclade_cli.rs
+++ b/packages_rs/nextclade-cli/src/cli/nextclade_cli.rs
@@ -230,7 +230,15 @@ pub enum NextcladeOutputSelection {
 pub struct NextcladeRunArgs {
   /// Path to a FASTA file with input sequences
   #[clap(value_hint = ValueHint::FilePath)]
-  pub input_fasta: Vec<PathBuf>,
+  pub input_fastas: Vec<PathBuf>,
+
+  /// REMOVED. Use positional arguments instead.
+  ///
+  /// Example: nextclade run -D dataset/ -O out/ seq1.fasta seq2.fasta
+  #[clap(long, short = 'i', visible_alias("sequences"))]
+  #[clap(value_hint = ValueHint::FilePath)]
+  #[clap(hide_long_help = true, hide_short_help = true)]
+  pub input_fasta: Option<PathBuf>,
 
   /// Path to a directory or a zip file containing a dataset.
   ///
@@ -318,6 +326,12 @@ pub struct NextcladeRunArgs {
   #[clap(value_hint = ValueHint::FilePath)]
   pub genes: Option<Vec<String>>,
 
+  /// REMOVED. Use `--output-all` instead
+  #[clap(long)]
+  #[clap(value_hint = ValueHint::DirPath)]
+  #[clap(hide_long_help = true, hide_short_help = true)]
+  pub output_dir: Option<PathBuf>,
+
   /// Produce all of the output files into this directory, using default basename and predefined suffixes and extensions. This is equivalent to specifying each of the individual `--output-*` flags. Convenient when you want to receive all or most of output files into the same directory and don't care about their filenames.
   ///
   /// Output files can be optionally included or excluded using `--output-selection` flag.
@@ -508,7 +522,7 @@ fn generate_completions(shell: &str) -> Result<(), Report> {
 /// Get output filenames provided by user or, if not provided, create filenames based on input fasta
 pub fn nextclade_get_output_filenames(run_args: &mut NextcladeRunArgs) -> Result<(), Report> {
   let NextcladeRunArgs {
-    input_fasta,
+    input_fastas: input_fasta,
     output_all,
     output_basename,
     output_ndjson,
@@ -642,6 +656,46 @@ At least one of the following flags is required:
   Ok(())
 }
 
+const ERROR_MSG_INPUT_FASTA_REMOVED: &str = r#"The argument `--input-fasta` (alias: `--sequences`, `-i`) is removed in favor of positional arguments.
+
+Try:
+
+  nextclade run -D dataset/ -O out/ seq1.fasta seq2.fasta
+
+                                        ^          ^
+                               one or multiple positional arguments
+                                 with paths to input fasta files
+
+When positional arguments are not provided, nextclade will read input fasta from standard input.
+
+For more information, type:
+
+  nextclade run --help"#;
+
+const ERROR_MSG_OUTPUT_DIR_REMOVED: &str = r#"The argument `--output-dir` is removed in favor of `--output-all`.
+
+When provided, `--output-all` allows to write all possible outputs into a directory.
+
+The defaut base name of the files can be overriden with `--output-basename` argument.
+
+The set of output files can be restricted with `--output-selection` argument.
+
+For more information, type
+
+  nextclade run --help"#;
+
+pub fn nextclade_check_removed_args(run_args: &mut NextcladeRunArgs) -> Result<(), Report> {
+  if run_args.input_fasta.is_some() {
+    return make_error!("{ERROR_MSG_INPUT_FASTA_REMOVED}");
+  }
+
+  if run_args.output_dir.is_some() {
+    return make_error!("{ERROR_MSG_OUTPUT_DIR_REMOVED}");
+  }
+
+  Ok(())
+}
+
 pub fn nextclade_parse_cli_args() -> Result<NextcladeArgs, Report> {
   let mut args = NextcladeArgs::parse();
 
@@ -662,6 +716,7 @@ pub fn nextclade_parse_cli_args() -> Result<NextcladeArgs, Report> {
       generate_completions(shell).wrap_err_with(|| format!("When generating completions for shell '{shell}'"))?;
     }
     NextcladeCommands::Run(ref mut run_args) => {
+      nextclade_check_removed_args(run_args)?;
       nextclade_get_output_filenames(run_args).wrap_err("When deducing output filenames")?;
     }
     _ => {}
diff --git a/packages_rs/nextclade-cli/src/cli/nextclade_loop.rs b/packages_rs/nextclade-cli/src/cli/nextclade_loop.rs
index 1cb85fac7..5854dad96 100644
--- a/packages_rs/nextclade-cli/src/cli/nextclade_loop.rs
+++ b/packages_rs/nextclade-cli/src/cli/nextclade_loop.rs
@@ -58,7 +58,7 @@ pub fn nextclade_run(args: NextcladeRunArgs) -> Result<(), Report> {
   info!("Command-line arguments:\n{args:#?}");
 
   let NextcladeRunArgs {
-    input_fasta,
+    input_fastas: input_fasta,
     output_all,
     output_basename,
     output_selection,