diff --git a/moto/moto_server/threaded_moto_server.py b/moto/moto_server/threaded_moto_server.py
index af4f347968ea..9a6ea62db36f 100644
--- a/moto/moto_server/threaded_moto_server.py
+++ b/moto/moto_server/threaded_moto_server.py
@@ -1,11 +1,26 @@
 from threading import Event, Thread
 from typing import Optional
 
-from werkzeug.serving import BaseWSGIServer, make_server
+from werkzeug.serving import BaseWSGIServer, WSGIRequestHandler, make_server
 
 from .werkzeug_app import DomainDispatcherApplication, create_backend_app
 
 
+class _MotoRequestHandler(WSGIRequestHandler):
+    def handle_expect_100(self) -> bool:
+        """Suppress the duplicate ``100 Continue`` from BaseHTTPRequestHandler.
+
+        Werkzeug's ``run_wsgi()`` already sends its own ``100 Continue``
+        when it sees the ``Expect`` header.  The default
+        ``BaseHTTPRequestHandler.parse_request()`` sends a second one,
+        resulting in two ``100 Continue`` responses on the wire.
+
+        Returning ``True`` without writing anything prevents the duplicate
+        while letting werkzeug send exactly one ``100 Continue`` later.
+        """
+        return True
+
+
 class ThreadedMotoServer:
     def __init__(
         self, ip_address: str = "0.0.0.0", port: int = 5000, verbose: bool = True
@@ -21,7 +36,13 @@ def __init__(
     def _server_entry(self) -> None:
         app = DomainDispatcherApplication(create_backend_app)
 
-        self._server = make_server(self._ip_address, self._port, app, True)
+        self._server = make_server(
+            self._ip_address,
+            self._port,
+            app,
+            threaded=True,
+            request_handler=_MotoRequestHandler,
+        )
         self._server_ready_event.set()
         self._server.serve_forever()
 
diff --git a/moto/server.py b/moto/server.py
index 133e909952fd..ca0d348827b7 100644
--- a/moto/server.py
+++ b/moto/server.py
@@ -9,6 +9,7 @@
 # Expose ThreadedMotoServer as a public symbol according to PEP-561
 from moto.moto_server.threaded_moto_server import (
     ThreadedMotoServer as ThreadedMotoServer,
+    _MotoRequestHandler,
 )
 from moto.moto_server.werkzeug_app import (
     DomainDispatcherApplication,
@@ -83,6 +84,7 @@ def main(argv: Optional[list[str]] = None) -> None:
         threaded=True,
         use_reloader=args.reload,
         ssl_context=ssl_context,
+        request_handler=_MotoRequestHandler,
     )