Now, return to the Virtual Media Manager by pressing Ctrl+D, select the CD/DVD Images tab once more, then select the missing image file within the list and click the "Remove" button at the top of the window. When prompted to confirm, click "Remove" to delete the image listing.
And there you have it. VirtualBox will no longer nag you about an image file in Virtual Media Manager that you deleted. If anyone has any other interesting tricks for dealing with image files in VirtualBox, please opine in the comments section below.
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Images by Matthew Nawrocki for TechRepublic.