Warning: beware of spoilers due to the nature of the game.
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Main Cast
Mario
The titular plumber hero. He gets a letter from Bowser, who is up to his usual tricks and has abducted the Princess, so it's up to him to save her.
- Adaptational Angst Upgrade: While in the original games, Mario has never had any angst, here, he's developed depression and has been trying to come to terms with his brother's death.
King Bowser
The Koopa King who has once again abducted Princess Peach, and sends Mario a letter letting him know this. He sends his minions to obstruct Mario and awaits at the end of his castle.
- Big Bad Ensemble: With the Shadow Mario/Luigi. He has once again kidnapped Princess Peach, and sends his minions to stop Mario from saving her.
- Disc-One Final Boss: Mario believes that defeating him is all he needs to save the princess, and he is seemingly the boss of Bowser's Castle, the last level. But he goes down relatively easily in a battle reminiscent of his fights in the original Super Mario Bros., and afterwards Mario still has to go through the Dark World version to fight the other villain, The Shadow.
Luigi (Unmarked Spoilers)
Mario's beloved brother. The Best ending reveals that he died for some unexplained reason, and his death is what sends Mario into his depression-fueled nightmares.
- Dead All Along: At first, Luigi just seems to be absent from the game. The Best Ending implies, via a grave marked with his initial, that he's been dead from the start.
- Plot-Triggering Death: His death is what caused Mario to lapse into depression, and the entire game is a nightmare born of that depression.
- Posthumous Character: He died some time ago for some unexplained reason.
The Shadow (Unmarked Spoilers)
- "It's-a me, Mario!"
A shadowy version of Mario and/or Luigi who stalks the plumber throughout his journey. He is the one responsible for turning the Mushroom Kingdom into a demented hellish version of itself. He is implied in the Best Ending to be a living embodiment of Mario's depression due to losing his brother, Luigi.
- Background Boss: It stays in the background as it rains fire, lasers, spikes, and such on you.
- Big Bad Ensemble: With Bowser. He is a Humanoid Abomination warping the Mushroom Kingdom into a nightmarish hellscape and stalking Mario. It is revealed to be an embodiment of his depression from the death of his brother Luigi.
- Creepy Long Fingers: Like the tall Marios, it has giant claws that it uses to attack Mario, and can shoot lasers out of them.
- Early-Bird Cameo: It can be seen hiding in both its Mario and Luigi forms several times in the first four levels, watching Mario behind a warp pipe or through a wall. It also serves as a Jump Scare in the end of the Ghost House, jumping at the player.
- Humanoid Abomination: He resembles Mario and/or Luigi, but is something that can warp the Mushroom Kingdom into a hellish Dark World full of blood, creepy Marios, and monsters.
- King Mook: Essentially one for the tall Marios. Like them, it is a Mario monster with Creepy Long Fingers.
- Mental Monster: It is hinted in the Best Ending that it represents Mario's depression over losing his brother Luigi.
- One-Winged Angel: In both the Normal and Best endings, it grows from its normal Mario and Luigi forms into giant versions of them.
- Scary Teeth: It has long, needle-like teeth that it shows off in both fights.
- True Final Boss: Its Giant Luigi form, a harder version of the Giant Mario form, must be beaten to get the Best Ending, and is only available if you get all five Green Stars, then take them to the secret door in Bowser's Castle.
- Red Eyes, Take Warning: The Shadow Mario has bright red eyes, which are especially visible during the battle against it.
Others
Bosses
The leaders of Bowser's troops who show up to fight Mario. They consist of a giant Piranha Plant and its Dark World version, a Chain Chomp, a King Dry Bones, a Giant Blooper, and King Bob-omb.
- Background Boss: The big Piranha Plants, King Dry Bones, and Giant Blooper attack from the background, the former two only surfacing to show their weak point.
- Giant Hands of Doom: King Dry Bones attacks with these, which it uses to slam into Mario and slide across the boss room.
- King Mook: All of them except the Chain Chomp are big versions of regular enemies.
- Optional Boss: King Bob-omb appears as the boss of an easily-missable Secret Level.
- Turns Red: After hitting the Chain Chomp twice, it will turn into a Dark World version- while the pattern of its attacks don't change, now you also have to hide from the giant Mario head in the background and time your jumps carefully to avoid being spotted by it.
The Mario Monsters
A collection of beasts that resemble Mario, but are more monstrous in nature. They consist of a giant Mario head that can kill Mario just by looking at him; tall Marios with creepy long legs and fingers; and white-headed Marios who appeared to have drowned and been eaten by Unagi, but are still capable of moving around.
- Back for the Finale: The giant Mario head makes a brief return in the Dark World Bowser's Castle as another obstacle, while the tall Marios return as enemies.
- Creepy Long Fingers: The tall Marios have these, which they use to skewer and rip Mario apart should they catch him.
- Evil Doppelgänger: They all resemble Mario, but are monstrous versions of him that attack him.
- Invincible Minor Minion: The tall Marios cannot be killed at all, and will instantly skewer Mario if they catch him.
The Puppet
- "IT WAS NOT YO-"
A giant puppet-like creature that appears in the Ghost House.
- Giant Space Flea from Nowhere: The other entities all have some connection to the plot or Mario, but this thing does not seem to have any connection whatsoever. It just shows up, creeps the player out for three scenes, then vanishes.
- Perverse Puppet: It at least looks like a creepy, giant puppet, and while it never outright attacks Mario, it doesn't seem to be particularly nice either.
- The Unfought: Though it might look like this thing will be the Ghost House boss, it is actually King Dry Bones who serves that role, while this thing never fights Mario.
Unagi (Unmarked Spoilers)
A giant eel that appears in the Underwater Level and swallows Mario.
- Flying Seafood Special: Though first encountered in the water, it somehow flies up in the air during a storm, as does its brethren.
- Sea Serpents: Giant eels based on the Unagi from various Mario games, it takes their creepiness from the series and milks it for all its worth, depicting them as terrifying predators that eat Mario alive.
