diff --git a/tests/test_env.py b/tests/test_env.py
index 04f7733..eb883b5 100644
--- a/tests/test_env.py
+++ b/tests/test_env.py
@@ -779,11 +779,20 @@ def child(pytester: pytest.Pytester, mocker: MockerFixture) -> Path:
     [pytest.param(False, id="omitted"), pytest.param(True, id="empty")],
 )
 @pytest.mark.parametrize(
-    "extend",
-    [pytest.param(False, id="config-only"), pytest.param(True, id="cli-extend")],
+    "scenario",
+    [
+        pytest.param((False, False), id="config-only-unset"),
+        pytest.param((False, True), id="config-only-set"),
+        pytest.param((True, False), id="cli-extend-unset"),
+        pytest.param((True, True), id="cli-extend-set"),
+    ],
 )
 def test_native_env_files_override_ini(
-    pytester: pytest.Pytester, config_name: str, empty_env_files: bool, extend: bool
+    pytester: pytest.Pytester,
+    mocker: MockerFixture,
+    config_name: str,
+    empty_env_files: bool,
+    scenario: tuple[bool, bool],
 ) -> None:
     (pytester.path / "fallback.env").write_text("INHERITED=from_ini\n", encoding="utf-8")
     (pytester.path / "cli.env").write_text("FROM_CLI=from_cli\n", encoding="utf-8")
@@ -795,21 +804,49 @@ def test_native_env_files_override_ini(
     else:
         native += '\n[pytest]\nenv_files = ["fallback.env"]\n'
     (pytester.path / config_name).write_text(native, encoding="utf-8")
-    expected_env: Final[tuple[str | None, str | None]] = (
+    pass_python_env: Final[bool] = scenario[1]
+    pythonpath_marker: Final[str] = str(pytester.path / "pythonpath_marker")
+    python_env: Final[dict[str, str]] = (
+        {
+            "PYTHONPATH": os.pathsep.join(filter(None, (os.environ.get("PYTHONPATH"), pythonpath_marker))),
+            "PYTHONDONTWRITEBYTECODE": "1",
+        }
+        if pass_python_env
+        else {}
+    )
+    extend: Final[bool] = scenario[0]
+    expected_env: Final[tuple[str | None, str | None, bool, str | None]] = (
         None if empty_env_files else "from_ini",
         "from_cli" if extend else None,
+        pass_python_env,
+        python_env.get("PYTHONDONTWRITEBYTECODE"),
     )
     pytester.makepyfile(
         test_env=dedent(f"""\
             import os
 
             def test_env() -> None:
-                assert (os.environ.get("INHERITED"), os.environ.get("FROM_CLI")) == {expected_env!r}
+                assert (
+                    os.environ.get("INHERITED"),
+                    os.environ.get("FROM_CLI"),
+                    {pythonpath_marker!r} in os.environ["PYTHONPATH"].split(os.pathsep),
+                    os.environ.get("PYTHONDONTWRITEBYTECODE"),
+                ) == {expected_env!r}
         """),
     )
-    new_env: Final[dict[str, str]] = {"PYTEST_DISABLE_PLUGIN_AUTOLOAD": "1", "PYTEST_PLUGINS": "pytest_env.plugin"}
-    with mock.patch.dict(os.environ, new_env, clear=True):
-        pytester.runpytest_subprocess(*(["--envfile", "+cli.env"] if extend else [])).assert_outcomes(passed=1)
+    system_env: Final[dict[str, str]] = {key: os.environ[key] for key in ("SYSTEMROOT", "WINDIR") if key in os.environ}
+    mocker.patch.dict(os.environ, python_env, clear=True)
+    mocker.patch.dict(
+        os.environ,
+        {
+            **system_env,
+            **{key: os.environ[key] for key in ("PYTHONPATH", "PYTHONDONTWRITEBYTECODE") if key in os.environ},
+            "PYTEST_DISABLE_PLUGIN_AUTOLOAD": "1",
+            "PYTEST_PLUGINS": "pytest_env.plugin",
+        },
+        clear=True,
+    )
+    pytester.runpytest_subprocess(*(["--envfile", "+cli.env"] if extend else [])).assert_outcomes(passed=1)
 
 
 @pytest.mark.parametrize(