Turn multiple PNG images into a H264 encoded 1280x720 animation for YouTube HD upload

• ffmpeg or mencoder can be used to create videos out of image sequences. I found this tutorial on it after a quick google. Their codecs are (mostly) built-in. The apps work on Windows, Mac and Linux.
• VirtualDub will allow you to import PNG image sequences, and encode them into videos. If you have the right codecs installed you can create H.264 files. You will need a separate application to convert it into the MP4 container. It does provide a command-line application. VDub is Windows-only.

And please don't forget that YouTube is in the habit of re-encoding its videos even if they're in the right format, so regardless you may just lose quality anyway. Thankfully it's not too much for live-action videos, but videos like 2D animations (anime, cartoons and the like) do suffer.

Take a look at WinFF, a great GUI frontend to make using FFMpeg easier. You can add all your images in batch and choose the output format. If YouTube is your target, they recommend using H.264, MPEG-2 or MPEG-4 as the format as they work best with their conversion system quality wise, so don't just limit yourself to H.264 as the others will turn out well too and there are more tools available for those formats.

mencoder 'mf://*.png' -mf fps=25 -o output.mp4 -ovc lavc -lavcopts vcodec=libx264

Or something like that.