It is possible that Peach's horn in Mario Kart 8/Deluxe is a reference to the first four notes of the main melody of the Peach Gardens track from Mario Kart DS (Mario Kart 8 Deluxe Booster Course Pass version used in the post, but the original track has the same notes).
Even if the reference is not intentional (since Peach Gardens was not part of Mario Kart 8 originally), the coincidence of a Peach-themed track starting off with a similar 4-note progression is slightly notable.
Cover of a 1988 issue of the Japanese Famicom Hisshoubon magazine, featuring original pixel art of Super Mario Bros. 3.
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Full 1991 Japanese commercial for Kirin Lemon soda, featuring a Super Mario World cross-promotion. Note how despite creating a variety of Super Mario World-inspired scenes with drawings and claymation, no assets from the game were used, and the level layouts are original designs as well.
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In Super Mario RPG, Fireworks are an expensive item that can be bought from a mole in Moleville for 500 coins (half of Mario's maximum coin carrying capacity). The game never suggests these are used for anything except being part of a trade sequence where they can be exchanged for a Shiny Stone, which can open the secret Culex boss battle or itself be further exchanged.
However, a secret use for these is that - independent of how many Mario actually has in his inventory - the game keeps track how many of them were purchased on that save file. Depending on the number of Fireworks Mario has purchased before beating the final boss, the shape of the fireworks in the credits will change between a Mushroom, a Fire Flower and a Star.
The same is also true for the Nintendo Switch version.
In Super Mario Bros. Wonder, the zero-gravity space section during the Wonder Effect of the Cosmic Hoppos level is reached by jumping on a special Hoppo that bounces Mario many screens up into the sky.
The combination of the Bubble Flower and the Boosting Spin Jump badge allows Mario to gain arbitrary height by blowing a bubble, spinning, bouncing off the bubble, and repeating this indefinitely. This can actually be used to very slowly manually make one's way to space, which has been sped up in the footage.
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Concept artwork for the 2023 Super Mario Bros. movie.
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Unused sleeping animation found in the files of Wario Land 4, likely intended to be used when Wario is left idle for long enough, similar to how Mario falls asleep in many different games under the same circumstances.
Interestingly, in the finished game, Wario does the opposite of sleeping when left idle - beginning to do exercise with dumbbells or a jumping rope instead. It is possible that this more standard sleeping animation was created early on in development before the designers decided that Wario's personality would be more suited for being always active.
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Top: unique illustration of a glitch found in Super Mario Bros.: The Lost Levels whereby Mario is immune to Hammer Bros.' hammers as long as he is touching the left side of the screen, seen in a 1986 issue of the Japanese Family Computer Magazine.
Bottom: game footage of the glitch in action, for reference. Note that the glitch also exists in Super Mario Bros., but since Hammer Bros. do not usually move to the left in the same way in that game (only doing this if Mario spends too much time lingering on the same screen), the glitch is far less useful.
Top: in Super Smash Bros. Ultimate, Mario and Luigi's stage entrance animations involve them jumping out of Warp Pipes. It is difficult to notice during regular gameplay unless comparing them directly, but Mario's pipe is a darker green than Luigi's.
Bottom: however, internally, the textures for the two pipes are actually swapped in their brightness relative to the objects as they are seen in-game. On the left is Mario's pipe texture, which is light green but is made darker at runtime with a shader. On the right is Luigi's pipe texture, which is dark green but made much lighter with a shader.
It is unknown why the developers chose such a bizarre way of implementing the pipe colors.
The Game & Watch Gallery 2 version of Helmet includes a unique Easter egg whereby a different character is unlocked for playing badly enough.
If the player gets a Game Over with a score of less than 100, and selects Retry on the Game Over screen, Mario will be replaced with Wario. Note that there is no other way to play as Wario, so a player that is too good at the game might never discover this.
Top: in the preview image for the Bowser Jr.'s playroom field in Mario Super Sluggers, a cloud can be seen to the top right of the Bowser Jr. decoration.
Middle: however, in-game, there are no clouds at all on the walls of this field. At first this may appear to be simply a case of an outdated preview image: as often happens in development, screenshots are taken of areas in a certain stage, but then the area is changed while the screenshot is never updated, creating the discrepancy.
Bottom: the truth, however, is much more bizarre. The clouds are actually there in-game, but can only be seen if the camera is moved outside the field itself. Note how only the clouds on the near side (seen from outside) are visible, and not the far side (seen normally, from inside).
This is due to the clouds being mistakenly coded to be inside-out (in technical terms, their culling is set to "backface", which is the opposite of how most regular objects are rendered). This strange oversight results in clouds that can not be seen during regular gameplay and require clipping out of bounds to discover.
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During the final battle against Bowser in Super Mario 64, a rare glitch may occur if Bowser is thrown offstage in a specific location where he already made part of the arena fall off.
After jumping back up from the abyss and making another part of the arena fall, Bowser will miss his follow-up jump to the safe part of the arena. Since Bowser's code only checks if he should jump back up after being thrown offstage by Mario instead of a case where he somehow falls through a fault of his own, he will simply never come back up.
This particular match against Bowser will become unwinnable due to Bowser being missing, and Mario must die to continue regular gameplay.