All my car radios can read flash drives these days - MP3 and WAV for sure, some accept some of the more obscure encoding techniques...
There is no standard, AFAIK, for how the directory (folder) structure is navigated.
Mine, however, all seem to employ a "depth-first" search pattern if there are multiple folders.
It will start with "the first" (by name? by the order they're written to the drive? -- there are differences and no standards...) folder.
If there are folders there, it will pick the first, and open that. And so on until there are no more folders - just music tracks.
When it's played all those, it goes back up a level, and enters the next folder, repeating the process.
If the directory structure is really complicated and there are lots of files, it can take a -long- time for the player to read the while structure when you plug one in. 16GB can be a lot of files - and some car players (probably just older ones these days) don't supply enough power to the drive to run bigger drives. 4GB is still a lot, and they're cheap.
So I usually just use a 1-level system - one folder per album - max.
Right now I'm using the 'immersion method' on the expected setlist for an upcoming blues jam.
I searched my main music drive for all the occurrences of the tracks I want and just stuck them (20 or so) in the root.
And it just repeats those 20 tracks while I drive to and from work...
If you're not trying to play them in a car radio, nevermind...