Setting them all at once to the same line makes them run behind eachother. I dont know the effects ingame (as i think the average time of travelling is counted, not the longest single waiting time. So the time for 1 train to complete its line devided by the number of trains running that line. Doesnt matter what the time between those trains is).
But i dont like to see 3 busses or something running behind eachother. its ineffective as the last will just be empty.