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Spoiler warning! This section of the article contains spoilers, or hints about the game's storyline or progression.
You might want to skip down to the next heading if you do not want facts about the game's storyline or plot revealed to you.
Surprisingly, Bowser is not the villain of the game.
The second main villain of the game, he has many minions under him. He is an interdimensional villain who has acquired the Dark Prognosticus and plans to use it to destroy all worlds and make his own from scratch. His real name is actually Count Blumiere and his true love is Tippi, also known as Lady Timpani. Their real names are revealed at the end of the game. He is only fought once in Castle Bleck.
Count Bleck's assistant. She has hypnotic and brainwashing powers. She often stumbles over her sentences, adding unnecessary words such as "Um" or "Yeah".
A strong, Scottish, but very unintelligent minion of Count Bleck. He is sometimes late for meetings. He is fought many times on Mario's journey, once in Yold Desert, once in The Land of the Cragnons, once in Floro Caverns and once in Castle Bleck.
A sadistic shapeshifting cute girl who transforms into a giant spider (known to her and the player as True Mimi) in boss fights. She is a minion of Count Bleck. She constantly changes dresses through every scene she appears, but is known for wearing a yellow dress with white polka-dots.
An evil magician, also the true main villain, who controls his own dimension (Dimension D) and can create many copies of himself. He often adds similes to his sentence, and almost always smiles. He dresses like a jester. He turned into Super Dimentio once but Mario, Peach and Bowser defeated him.
Is he Luigi? He builds giant robots in order to fight the heroes. He probably became a minion of Count Bleck through Nastasia's brainwashing. He acts cocky and gives the Heroes of Light nicknames (Mario is Mr. Jumpsallthetime, for example. This indicates that Mr. L may have some memory of being Luigi).
Mr. L's giant robot. Another piece of evidence that he is Luigi. All you need to do is take Peach and use her umbrella by using it and gliding over Brobot. In another situation you use Mario and the funny alien who has to go to the bathroom and you have to shoot Brobot. Just keep away from the rocks, and you'll be OK.