I have a web site running on Windows Server 2012 R2, IIS 8.5.9600.16384. The site has several virtual directories that point to a file share from which documents (mostly PDFs) are retrieved. All of these virtual directories are using a custom identity for the "Connect As..." option, and the "Test Setting..." button returns successful results.
Ever since we went live, users will occasionally get 500 errors when attempting to access files on these servers through the virtual directories. I am able to duplicate the problem at that moment. The weird thing is that restarting IIS completely fixes the problem for a time. At some later point (usually a day or two), the problem returns. It seems to impact the various virtual directories randomly; while one is failing, the others may or may not work, but restarting fixes them all.
I have no idea how to even go about troubleshooting this. Any ideas?