I was assuming that on the trail -- you drink from the water bottles first -- and then replenish from the 4L bladder when needed? It so, it doesn't hurt to treat all the water simultaneously. After all, if the water is cold, it can take up to 4 hours to properly treat against protozoa cysts (giardia, crypto, etc.)! Longer treatment time never hurts.
OTOH, say you are nearing camp... you fill your two water bottles for drinking and you fill the 4L bladder for cooking and washing. In this case, it may be wasteful to treat the 4L bladder. Perhaps better to leave the 4L bladder untreated -- use the water for cooking and washing -- and whatever you have left -- treat only if you plan to drink later or the next day.