 <p>Where you have:</p> <pre><code>&lt;Directory /home/bitnami/public_html/http/flasktest1&gt; WSGIProcessGroup flaskapp WSGIApplicationGroup %{GLOBAL} Order deny,allow Allow from all &lt;/Directory&gt; </code></pre> <p>it should be:</p> <pre><code>&lt;Directory /home/bitnami/public_html/http&gt; WSGIProcessGroup flaskapp WSGIApplicationGroup %{GLOBAL} Order deny,allow Allow from all &lt;/Directory&gt; </code></pre> <p>as you are neither running your app in daemon mode or in the main interpreter because of the directives being in the wrong context.</p> <p>That Directory directive then conflicts with one for same directory above so merge them.</p> <p>If using mod_wsgi 3.0 or later count instead perhaps drop that second Directory block and use:</p> <pre><code>WSGIDaemonProcess flaskapp threads=5 WSGIScriptAlias /flasktest1 /home/bitnami/public_html/wsgi/flasktest1.wsgi process-group=flaskapp application-group=%{GLOBAL} </code></pre> <p>Note that processes=1 has been dropped as that is the default and setting it implies other things you likely don't want. You also don't need to set user/group as it will automatically run as the Apache user anyway.</p>