diff --git a/tests/conftest.py b/tests/conftest.py
index a6458d7..1e46d61 100644
--- a/tests/conftest.py
+++ b/tests/conftest.py
@@ -1,7 +1,11 @@
 """Shared fixtures for EvoScientist tests."""
 
+from pathlib import Path
+
 import pytest
 
+_NONEXISTENT_DOTENV = str(Path(__file__).with_name(".pytest-dotenv-does-not-exist"))
+
 
 @pytest.fixture(autouse=True)
 def _reset_tool_selection_state():
@@ -164,3 +168,24 @@ def restore_model_passthrough_patch():
         yield
     finally:
         _reset()
+
+
+@pytest.fixture(autouse=True)
+def _isolate_dotenv(monkeypatch):
+    """Keep the developer's real .env out of the test environment.
+
+    ``get_effective_config`` runs ``load_dotenv(find_dotenv(usecwd=True),
+    override=True)``, so any test that loads config injects the repo's
+    real .env into ``os.environ`` for the rest of the pytest process.
+    An empty-valued line like ``MINIMAX_BASE_URL=`` then makes
+    ``os.environ.get(key, default)`` return "" instead of the default,
+    breaking unrelated tests later in the run (see issue #322).
+
+    Pointing ``find_dotenv`` at a fixed path that does not exist makes
+    ``load_dotenv`` a no-op without creating a temporary directory for
+    every test.
+    """
+    monkeypatch.setattr(
+        "EvoScientist.config.settings.find_dotenv",
+        lambda *args, **kwargs: _NONEXISTENT_DOTENV,
+    )
diff --git a/tests/test_config.py b/tests/test_config.py
index df7a7a7..522d964 100644
--- a/tests/test_config.py
+++ b/tests/test_config.py
@@ -53,11 +53,6 @@ def temp_config_dir(tmp_path, monkeypatch):
     """Use a temporary directory for config during tests."""
     config_dir = tmp_path / "evoscientist"
     monkeypatch.setenv("XDG_CONFIG_HOME", str(tmp_path))
-    # Prevent load_dotenv from loading the project's real .env file
-    monkeypatch.setattr(
-        "EvoScientist.config.settings.find_dotenv",
-        lambda *a, **k: str(tmp_path / ".env"),
-    )
     # Also clear any API keys from environment
     for key in [
         "ANTHROPIC_API_KEY",
@@ -930,3 +925,30 @@ def test_scheduler_config_defaults_and_env(monkeypatch):
     assert eff2.memory_skill_synthesis_mode == MemorySkillSynthesisMode.AUTO
     assert eff2.memory_skill_synthesis_cadence == MemorySkillSynthesisCadence.MONTHLY
     assert eff2.memory_skill_synthesis_time == "04:30"
+
+
+# =============================================================================
+# Dotenv isolation (issue #322)
+# =============================================================================
+
+
+class TestDotenvIsolation:
+    def test_env_file_not_leaked_into_process_env(self, tmp_path, monkeypatch):
+        """A .env in cwd must not leak into os.environ during tests.
+
+        Without the suite-wide ``_isolate_dotenv`` fixture,
+        ``get_effective_config`` loads the developer's real .env with
+        ``override=True``; an empty-valued line like ``MINIMAX_BASE_URL=``
+        then poisons ``os.environ.get(key, default)`` lookups for every
+        test that runs afterwards in the same process.
+        """
+        monkeypatch.setenv("XDG_CONFIG_HOME", str(tmp_path))
+        repro_dir = tmp_path / "repro"
+        repro_dir.mkdir()
+        (repro_dir / ".env").write_text("MINIMAX_BASE_URL=\n")
+        monkeypatch.chdir(repro_dir)
+        monkeypatch.delenv("MINIMAX_BASE_URL", raising=False)
+
+        get_effective_config()
+
+        assert "MINIMAX_BASE_URL" not in os.environ