I don't know if "PreSonus FireStudio" is the correct driver choice. It should be "StudioLive RM/RML 16AI" (or 32AI), but I am no sure anymore how this was handled with FireWire mixers. To me "Playback Device" and "Recording Device" in your picture look like a selection of a stereo input and output audio interface. May I ask which mixer/interface you can see in UC Control? This selection (the mixer) should be seen in the settings of Studio One.
And, have you loaded the template for your mixer when you created the song you are working on? The template is called "StudioLive 16.4.2AI and RM 16 Session" and it provides all necessary routing. Channel 1 (as example) is routed to track 1 and vice versa. In this setting the stereo track "Main L/R" is the input (!) of "Main L/R" of the rack mixer, which means that you can record a stereo track of your mix on the rack mixer. You can check in Studio One's settings, to which FireWire streams Main L/R is routed and check in you rack mixers settings, if they correspond.
Studio One's stereo output "2TrackIn L/R" is not active in this scenario, because it functions as sum of a mix done in Studio One. But because the tracks are routed to their respective channels on the rack mixer, there is no "mix" happening in Studio One. You would either need to route a track to "2TrackIn L/R" or create an effect bus, which automatically is routed to "2TrackIn L/R". On the rack mixer this stereo sum comes in via "Tape In/Digital In". You will have to select "Digital" as input source.
Hope this helps. It's not really clear what your question is. This answer is presenting a basic way to connect Studio One and a rack mixer. And remember that if you want to listen to recorded tracks, you have to select "Digital" as input source for the respective channels on your rack mixer. This toggling between analog and digital inputs will always be needed, because the rack mixer needs to know WHERE a signal is coming from.
