diff --git a/nf_core/__init__.py b/nf_core/__init__.py
index 42c3f188f3..e333335280 100644
--- a/nf_core/__init__.py
+++ b/nf_core/__init__.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Main nf_core module file.
 
 Shouldn't do much, as everything is under subcommands.
diff --git a/nf_core/bump_version.py b/nf_core/bump_version.py
index 53766678b0..1f38775bb6 100644
--- a/nf_core/bump_version.py
+++ b/nf_core/bump_version.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Bumps the version number in all appropriate files for
 a nf-core pipeline.
 """
diff --git a/nf_core/create.py b/nf_core/create.py
index ca06bd9c36..ed23c8487c 100644
--- a/nf_core/create.py
+++ b/nf_core/create.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Creates a nf-core pipeline matching the current
 organization's specification based on a template.
 """
diff --git a/nf_core/download.py b/nf_core/download.py
index 923d953fae..5e7f4edb2f 100644
--- a/nf_core/download.py
+++ b/nf_core/download.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Downloads a nf-core pipeline to the local file system."""
 
 from __future__ import print_function
diff --git a/nf_core/launch.py b/nf_core/launch.py
index 53fce1f43f..03fcb6bce8 100644
--- a/nf_core/launch.py
+++ b/nf_core/launch.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Launch a pipeline, interactively collecting params """
 
 from __future__ import print_function
diff --git a/nf_core/licences.py b/nf_core/licences.py
index 2e65462a1d..3b188861eb 100644
--- a/nf_core/licences.py
+++ b/nf_core/licences.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Lists software licences for a given workflow."""
 
 from __future__ import print_function
diff --git a/nf_core/lint/__init__.py b/nf_core/lint/__init__.py
index 5ea01142e5..a0de378d82 100644
--- a/nf_core/lint/__init__.py
+++ b/nf_core/lint/__init__.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Linting policy for nf-core pipeline projects.
 
 Tests Nextflow-based pipelines to check that they adhere to
diff --git a/nf_core/lint/actions_awsfulltest.py b/nf_core/lint/actions_awsfulltest.py
index e021ebd384..e8e1c951b1 100644
--- a/nf_core/lint/actions_awsfulltest.py
+++ b/nf_core/lint/actions_awsfulltest.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import yaml
diff --git a/nf_core/lint/actions_awstest.py b/nf_core/lint/actions_awstest.py
index 4f27cbd765..ccdf0abf6a 100644
--- a/nf_core/lint/actions_awstest.py
+++ b/nf_core/lint/actions_awstest.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import yaml
diff --git a/nf_core/lint/actions_ci.py b/nf_core/lint/actions_ci.py
index 4d6b0e6dfe..b4c10f3a74 100644
--- a/nf_core/lint/actions_ci.py
+++ b/nf_core/lint/actions_ci.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 import re
 
diff --git a/nf_core/lint/actions_schema_validation.py b/nf_core/lint/actions_schema_validation.py
index 7ded008cfc..9d49b84c6b 100644
--- a/nf_core/lint/actions_schema_validation.py
+++ b/nf_core/lint/actions_schema_validation.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import glob
 import logging
 import os
diff --git a/nf_core/lint/files_exist.py b/nf_core/lint/files_exist.py
index 2a1dee62c9..eb8c04916a 100644
--- a/nf_core/lint/files_exist.py
+++ b/nf_core/lint/files_exist.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import logging
 import os
 
diff --git a/nf_core/lint/files_unchanged.py b/nf_core/lint/files_unchanged.py
index 8b63a5769e..cadced5483 100644
--- a/nf_core/lint/files_unchanged.py
+++ b/nf_core/lint/files_unchanged.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import filecmp
 import logging
 import os
diff --git a/nf_core/lint/merge_markers.py b/nf_core/lint/merge_markers.py
index 75fbf931bf..f33a5095d8 100644
--- a/nf_core/lint/merge_markers.py
+++ b/nf_core/lint/merge_markers.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import fnmatch
 import io
 import logging
diff --git a/nf_core/lint/modules_json.py b/nf_core/lint/modules_json.py
index 3e97eed043..dd0a59d558 100644
--- a/nf_core/lint/modules_json.py
+++ b/nf_core/lint/modules_json.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 from pathlib import Path
 
 from nf_core.modules.modules_json import ModulesJson
diff --git a/nf_core/lint/modules_structure.py b/nf_core/lint/modules_structure.py
index 830e0d1f16..f0e13e0346 100644
--- a/nf_core/lint/modules_structure.py
+++ b/nf_core/lint/modules_structure.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import logging
 import os
 from pathlib import Path
diff --git a/nf_core/lint/multiqc_config.py b/nf_core/lint/multiqc_config.py
index 37580a1f11..3378efce5f 100644
--- a/nf_core/lint/multiqc_config.py
+++ b/nf_core/lint/multiqc_config.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import yaml
diff --git a/nf_core/lint/nextflow_config.py b/nf_core/lint/nextflow_config.py
index 635e33cfb5..cf0fbf4d40 100644
--- a/nf_core/lint/nextflow_config.py
+++ b/nf_core/lint/nextflow_config.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import logging
 import os
 import re
diff --git a/nf_core/lint/pipeline_name_conventions.py b/nf_core/lint/pipeline_name_conventions.py
index e1ecad0be2..7fb6ffca0f 100644
--- a/nf_core/lint/pipeline_name_conventions.py
+++ b/nf_core/lint/pipeline_name_conventions.py
@@ -1,6 +1,3 @@
-#!/usr/bin/env python
-
-
 def pipeline_name_conventions(self):
     """Checks that the pipeline name adheres to nf-core conventions.
 
diff --git a/nf_core/lint/pipeline_todos.py b/nf_core/lint/pipeline_todos.py
index 91a7cf6307..890e227fa1 100644
--- a/nf_core/lint/pipeline_todos.py
+++ b/nf_core/lint/pipeline_todos.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import fnmatch
 import io
 import logging
diff --git a/nf_core/lint/readme.py b/nf_core/lint/readme.py
index 99def0a204..a85a0979d3 100644
--- a/nf_core/lint/readme.py
+++ b/nf_core/lint/readme.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 import re
 
diff --git a/nf_core/lint/schema_description.py b/nf_core/lint/schema_description.py
index 3a670e5f70..ca22f266ab 100644
--- a/nf_core/lint/schema_description.py
+++ b/nf_core/lint/schema_description.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import nf_core.schema
 
 
diff --git a/nf_core/lint/schema_lint.py b/nf_core/lint/schema_lint.py
index f7c1b11048..178063d5dd 100644
--- a/nf_core/lint/schema_lint.py
+++ b/nf_core/lint/schema_lint.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import logging
 
 import nf_core.schema
diff --git a/nf_core/lint/schema_params.py b/nf_core/lint/schema_params.py
index 6b32535738..9280fe4703 100644
--- a/nf_core/lint/schema_params.py
+++ b/nf_core/lint/schema_params.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import nf_core.schema
 
 
diff --git a/nf_core/lint/template_strings.py b/nf_core/lint/template_strings.py
index 17886cec3f..436abe7b2b 100644
--- a/nf_core/lint/template_strings.py
+++ b/nf_core/lint/template_strings.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import io
 import mimetypes
 import re
diff --git a/nf_core/lint/version_consistency.py b/nf_core/lint/version_consistency.py
index 89a8751af6..fa5b50de01 100644
--- a/nf_core/lint/version_consistency.py
+++ b/nf_core/lint/version_consistency.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 
diff --git a/nf_core/list.py b/nf_core/list.py
index 62c925ee47..b4353050e6 100644
--- a/nf_core/list.py
+++ b/nf_core/list.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Lists available nf-core pipelines and versions."""
 
 from __future__ import print_function
diff --git a/nf_core/modules/create.py b/nf_core/modules/create.py
index a0e0fa66e4..1d18fee5d5 100644
--- a/nf_core/modules/create.py
+++ b/nf_core/modules/create.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 The ModuleCreate class handles generating of module templates
 """
diff --git a/nf_core/modules/lint/__init__.py b/nf_core/modules/lint/__init__.py
index 10063f90c5..73c7f57d2b 100644
--- a/nf_core/modules/lint/__init__.py
+++ b/nf_core/modules/lint/__init__.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 Code for linting modules in the nf-core/modules repository and
 in nf-core pipelines
diff --git a/nf_core/modules/lint/main_nf.py b/nf_core/modules/lint/main_nf.py
index e396e7e813..f0098c8ab7 100644
--- a/nf_core/modules/lint/main_nf.py
+++ b/nf_core/modules/lint/main_nf.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 Lint the main.nf file of a module
 """
diff --git a/nf_core/modules/lint/meta_yml.py b/nf_core/modules/lint/meta_yml.py
index 77df16deb2..d6ec296999 100644
--- a/nf_core/modules/lint/meta_yml.py
+++ b/nf_core/modules/lint/meta_yml.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 from pathlib import Path
 
 import yaml
diff --git a/nf_core/modules/lint/module_deprecations.py b/nf_core/modules/lint/module_deprecations.py
index f7e8761c75..aa4cabd7b2 100644
--- a/nf_core/modules/lint/module_deprecations.py
+++ b/nf_core/modules/lint/module_deprecations.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 import logging
 import os
 
diff --git a/nf_core/modules/lint/module_todos.py b/nf_core/modules/lint/module_todos.py
index 90af44987e..8b685a7172 100644
--- a/nf_core/modules/lint/module_todos.py
+++ b/nf_core/modules/lint/module_todos.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 import logging
 
 from nf_core.lint.pipeline_todos import pipeline_todos
diff --git a/nf_core/modules/lint/module_version.py b/nf_core/modules/lint/module_version.py
index ccf1c4d91e..3f68395031 100644
--- a/nf_core/modules/lint/module_version.py
+++ b/nf_core/modules/lint/module_version.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 Verify that a module has a correct entry in the modules.json file
 """
diff --git a/nf_core/modules/modules_test.py b/nf_core/modules/modules_test.py
index 94dfd344c6..d1f47dcff9 100644
--- a/nf_core/modules/modules_test.py
+++ b/nf_core/modules/modules_test.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 The ModulesTest class runs the tests locally
 """
diff --git a/nf_core/modules/test_yml_builder.py b/nf_core/modules/test_yml_builder.py
index 13d415a2b0..854700bb89 100644
--- a/nf_core/modules/test_yml_builder.py
+++ b/nf_core/modules/test_yml_builder.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 The ModulesTestYmlBuilder class handles automatic generation of the modules test.yml file
 along with running the tests and creating md5 sums
diff --git a/nf_core/refgenie.py b/nf_core/refgenie.py
index e8d421d176..a10e4fecdf 100644
--- a/nf_core/refgenie.py
+++ b/nf_core/refgenie.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 Update a nextflow.config file with refgenie genomes
 """
diff --git a/nf_core/schema.py b/nf_core/schema.py
index 04e18409cb..0be825fea2 100644
--- a/nf_core/schema.py
+++ b/nf_core/schema.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Code to deal with pipeline JSON Schema """
 
 from __future__ import print_function
diff --git a/nf_core/subworkflows/create.py b/nf_core/subworkflows/create.py
index 2440b2ad8d..755db20127 100644
--- a/nf_core/subworkflows/create.py
+++ b/nf_core/subworkflows/create.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 The SubworkflowCreate class handles generating of subworkflow templates
 """
diff --git a/nf_core/subworkflows/test_yml_builder.py b/nf_core/subworkflows/test_yml_builder.py
index 271dd7edd5..4d3162bd7c 100644
--- a/nf_core/subworkflows/test_yml_builder.py
+++ b/nf_core/subworkflows/test_yml_builder.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 The ModulesTestYmlBuilder class handles automatic generation of the modules test.yml file
 along with running the tests and creating md5 sums
diff --git a/nf_core/sync.py b/nf_core/sync.py
index 5aabd8eb51..2384ad4de7 100644
--- a/nf_core/sync.py
+++ b/nf_core/sync.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Synchronise a pipeline TEMPLATE branch with the template.
 """
 
diff --git a/nf_core/utils.py b/nf_core/utils.py
index 57907109f9..f5a464b3c2 100644
--- a/nf_core/utils.py
+++ b/nf_core/utils.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """
 Common utility functions for the nf-core python package.
 """
diff --git a/tests/lint/actions_awsfulltest.py b/tests/lint/actions_awsfulltest.py
index 234628227d..30293e31a4 100644
--- a/tests/lint/actions_awsfulltest.py
+++ b/tests/lint/actions_awsfulltest.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import yaml
diff --git a/tests/lint/actions_awstest.py b/tests/lint/actions_awstest.py
index 45d7e66c3d..0e19f781aa 100644
--- a/tests/lint/actions_awstest.py
+++ b/tests/lint/actions_awstest.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import yaml
diff --git a/tests/lint/actions_ci.py b/tests/lint/actions_ci.py
index 81eb26f22c..b6e58889f4 100644
--- a/tests/lint/actions_ci.py
+++ b/tests/lint/actions_ci.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import yaml
diff --git a/tests/lint/actions_schema_validation.py b/tests/lint/actions_schema_validation.py
index d71603a56e..48bb07e4dd 100644
--- a/tests/lint/actions_schema_validation.py
+++ b/tests/lint/actions_schema_validation.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import yaml
diff --git a/tests/lint/files_exist.py b/tests/lint/files_exist.py
index 02686e9add..4e5e4d3c2b 100644
--- a/tests/lint/files_exist.py
+++ b/tests/lint/files_exist.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import nf_core.lint
diff --git a/tests/lint/merge_markers.py b/tests/lint/merge_markers.py
index 6ea2882417..be0d076757 100644
--- a/tests/lint/merge_markers.py
+++ b/tests/lint/merge_markers.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python
-
 import os
 
 import nf_core.lint
diff --git a/tests/test_bump_version.py b/tests/test_bump_version.py
index 96eb1240a0..658a2339d4 100644
--- a/tests/test_bump_version.py
+++ b/tests/test_bump_version.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Some tests covering the bump_version code.
 """
 import os
diff --git a/tests/test_cli.py b/tests/test_cli.py
index f991681b0f..92e6e4150e 100644
--- a/tests/test_cli.py
+++ b/tests/test_cli.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Tests covering the command-line code.
 
 Most tests check the cli arguments are passed along and that some action is
diff --git a/tests/test_create.py b/tests/test_create.py
index 5f732fc180..baac509d74 100644
--- a/tests/test_create.py
+++ b/tests/test_create.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Some tests covering the pipeline creation sub command.
 """
 import os
diff --git a/tests/test_download.py b/tests/test_download.py
index feb486e2c8..4577a83992 100644
--- a/tests/test_download.py
+++ b/tests/test_download.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Tests for the download subcommand of nf-core tools
 """
 
diff --git a/tests/test_launch.py b/tests/test_launch.py
index 4d4ed79649..de597127a2 100644
--- a/tests/test_launch.py
+++ b/tests/test_launch.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Tests covering the pipeline launch code.
 """
 
diff --git a/tests/test_licenses.py b/tests/test_licenses.py
index a2dde1639b..4fb58a107c 100644
--- a/tests/test_licenses.py
+++ b/tests/test_licenses.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Some tests covering the pipeline creation sub command.
 """
 # import json
diff --git a/tests/test_lint.py b/tests/test_lint.py
index db9bf0fc36..5012d20683 100644
--- a/tests/test_lint.py
+++ b/tests/test_lint.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """Some tests covering the linting code.
 """
 import fnmatch
diff --git a/tests/test_list.py b/tests/test_list.py
index 2acef31a76..70af3fada5 100644
--- a/tests/test_list.py
+++ b/tests/test_list.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Tests covering the workflow listing code.
 """
 
diff --git a/tests/test_refgenie.py b/tests/test_refgenie.py
index 9314b44eef..1ff2683416 100644
--- a/tests/test_refgenie.py
+++ b/tests/test_refgenie.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Tests covering the refgenie integration code
 """
 
diff --git a/tests/test_schema.py b/tests/test_schema.py
index 10313c81f3..d3b4fda817 100644
--- a/tests/test_schema.py
+++ b/tests/test_schema.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Tests covering the pipeline schema code.
 """
 
diff --git a/tests/test_sync.py b/tests/test_sync.py
index 0cedf51698..f0f6c7edca 100644
--- a/tests/test_sync.py
+++ b/tests/test_sync.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Tests covering the sync command
 """
 
diff --git a/tests/test_utils.py b/tests/test_utils.py
index b43d6e8ff2..2ab5b64bfc 100644
--- a/tests/test_utils.py
+++ b/tests/test_utils.py
@@ -1,4 +1,3 @@
-#!/usr/bin/env python
 """ Tests covering for utility functions.
 """
 
diff --git a/tests/utils.py b/tests/utils.py
index 65c7d48758..8db4cd7494 100644
--- a/tests/utils.py
+++ b/tests/utils.py
@@ -1,5 +1,3 @@
-#!/usr/bin/env python3
-# -*- coding: utf-8 -*-
 """
 Helper functions for tests
 """