- The Trix (Seasons 1,2,3,5) are the main antagonists of the series. They managed to take away Bloom's power, The Dragon Flame (Fire) until Bloom managed to believe in herself and use all of the power that was still in her and destroy The Trix. They were arrested and sent to the Fortress of Roccaluce in the end of the first season, but they escaped with the help of Lord Darkar in the beginning of the second season. He gave them their Gloomix which multiplies their power. In the third season, they worked with Valtor. He gave them a power-up called Disenchantix, which is the opposite of Enchantix. They made a comeback in the second film as minions of the Ancestral Witches. The Trix returned in the fifth season and gained a power-up. They also gained Dark Sirenix that Tritannus stole from Daphne and gave them.
- Icy : The leader and the oldest of the trio, her name refers to her powers of ice. She enjoys torturing pixies and despises Bloom and her friends, therefore she is most competitive with the girls for the whole series. She is the most powerful of the three. In comparison to her sisters, she is more aggressive than Darcy, but calmer than Stormy. She has white/gray hair, and has a blue ribbon in her hair that ties it into a pony tail with three ice blue diamonds on it and a blue outfit with an 'I' in the middle and dark blue boots. In season two, Darkar gives Icy a light blue necklace(Gloomix) that makes a perfect shape around her neck then makes a wider circle shape and ends with a diamond. Icy is also given the form Disenchantix in season 3, as do Stormy and Darcy. Her boyfriend is Tritannus. She is voiced in the original Italian version by Tatiana Dessi. She was voiced by Lisa Ortiz in the 4kids dub, She is currently voiced by Larisa Oleynik in the Nickelodeon dub.
- Darcy: Darcy's powers works with all forms of darkness, including illusions, some say she has mesmatizing powers as her name suggests ("Darcy" meaning "Dark One"). She enjoys tormenting fairies and is less aggressive than her sisters. Darcy hypnotized Riven, getting him to turn on the fairies, for most of season one. In season two, Darkar gives Darcy a bracelet (Gloomix) that goes on the back of her hand to her elbow, which is purple on her left hand.She has long, brown hair and light brown bangs and a purple outfit with a 'D' on it, she is sometimes seen with glasses. She is voiced in the original Italian version by Federica De Bortoli, and was voiced by Caren Manuel in the 4kids dub, and is currently voiced by Jennifer Cody in the Nickelodeon dub.
- Stormy (Season 1–3, 5): The youngest of the trio, her powers involve lightning, storms, and weather. She is the most hot-tempered of the group and is prone to violent outbursts. She often does not think before she does something. Stormy is Musa's rival for most of the series. In season two, Darkar gives Stormy a bracelet (Gloomix) that goes on the back of her hand all the way up to her elbow, which is violet on her right hand. She is voiced in the original Italian version by Valeria Vidali. She was voiced by Suzy Myers in the 4kids dub, She is currently voiced by Kimberly Brooks in the Nickelodeon dub.
- Lord Darkar, also called The Shadow Phoenix is the primary antagonist of the second season. He is an ancient being from the beginning of when all things were created. The Shadow Phoenix is the diametrical opposite of the Dragon, and like the Dragon, he is very powerful. The dragon is the source of energy and light while the Shadow Phoenix absorbs energy and magic, making his essence the opposite of the Dragon's Flame.
He is the cause of the destruction of Domino, since the Ancestral Witches were his servers who acted upon his orders. After the Ancestral Witches were defeated, the Lord of the Templars of Roccaluce defeated Lord Darkar and sent him into a deep slumber from which he woke up at the beginning of season 2, possibly by the reappearance of the Dragon Flame in the Magic Dimension. He bears many similarities to Sauron from J. R. R. Tolkien's legendarium and may have been inspired by him. He kidnapped the pixies, rescued the Trix from their prison and used the Codex and Bloom's powers to open a portal to the Realm of the Relix. The Shadow Phoenix nearly succeeded in his plot to claim the ultimate power and rule the magical dimension, but in the end, Darkar was defeated by all six Winx members, with a Charmix convergence, which destroyed him for good.
- Dark Bloom is the evil doppelgänger of Bloom after being turned evil by Darkar's shadow magic. In her first appearance, she destroyed half of Alfea's secret archives, defeated the rest of the Winx Club and sent a piece of the codex to Lord Darkar. In the last episode of season 2, she entered the realm of Realix to claim the Ultimate Power, initially with the intention of handing it to Lord Darkar, but after Sky confessed his love for her, the small drop of good in her was strong enough for her to regain control of herself and she used her healing powers to cure herself and the others of the dark spell. She then returned the Ultimate Power and helped the other fairies defeat Darkar. She does not speak much in her last appearance, although she does laugh occasionally and chant when opening Realix. Darkar continually flirts with her throughout the last few episodes of season two and stated she was his natural consort, which annoys the Trix even more after Darkar chose Dark Bloom as his main witch. Her outfit is black-navy.
- Keborg is a bat-like creature that serves Lord Darkar as a spy.
- Valtor is a powerful wizard who aspires to be the greatest wizard of all. He turns people into monsters that willingly serve him and places a mark that looks like a "V" on other people so they too will serve him. By doing so he hopes to be able to conquer each planet or realm. He was created from the Dragon's Flame by the Three Ancient Witches. He was with them when they destroyed Domino, but was sent to the Omega Dimension as punishment. Seventeen years later, the Trix are sent there as well, where they freed him from his prison. They escape and he begins conquering Andros, Aisha's home planet. Valtor then moves on to Solaria where he restores his lost powers with the planet's second sun. In their youth, he and Ms. Griffin were magical companions. During that time he meets Bloom's parents, Queen Marion and King Oritel of Domino, who are both stronger than he is. However, he tricks them and supposedly kills them the day Domino is destroyed. He is shown to make those who defeat him miserable. He hunts down Faragonda, Griffin and Saladin, imprisoning them.
In the second-to-last episode of season three, Valtor transforms into a demon and unleashes a spell known as the Spell of the Elements, sending elemental attacks in four directions: water (flooding) to Cloudtower, wind (tornadoes) to Redfountain, fire (forever-burning, impossible to put out) to Alfea and earth (earthquakes) to Magix. However, the Winx manage to release all the spells that Valtor had stolen. In the final episode, Valtor is defeated by Bloom in his demon form from the inside out and banished to the Realm of Oblivion along with the Three Ancient Witches. He functions as the primary villain of season three. Sean Schemmel voices Valtor in the 4Kids version and Guido Di Naccio in the original version. In the 4Kids version, his name is changed from Valtor to Baltor. In the Nickelodeon dub, he was voiced by Josh Keaton.
- Three Ancestral Witches trapped Bloom's parents, put a curse on Daphane and destroyed Domino. They are the Trix's ancestors.
- Mandragora is an evil sorceress and the one of the main antagonists of Secret of the Lost Kingdom.
- The Wizards Of The Black Circle, Ogron, Gantlos, Duman, and Anagan, are the four main villains of the fourth season.
- Ogron can absorb and deflect magic.
- Duman is a shape-shifter.
- Anagan can create shock waves.
- Gantlospossesses super speed.
- At some point, they had developed a way to resist fairy magic, which allowed them to capture the fairies of Earth, and steal all of the magic from the planet. Throughout the first half of the season, they battle the Winx, and almost every time the fairies barely make it out alive. However, as time went on the Winx became more accustomed to fighting them, and eventually were able to give them a real fight. They are defeated in the middle of the season by Bloom, with help from Roxy, who used her Believix powers to convince all of the humans in the area to believe that fairies exist. This rendered the Wizards' abilities useless, and made them vulnerable to Bloom's attack. In the beginning of the second half of the season, it is revealed that they are still alive, but their powers are weaker than before and they must hide in the sewers. Later on in the season, the Fairy Hunters claim to be good when Duman falls ill. Duman is taken care of by Sibylla, the Fairy of Justice afterwards, though his power goes out of control and he has to be killed. While all four Hunters seem to share certain powers, like opening a Dark Gate to capture fairies and cast dark enchantments, they also have individual powers. Patrizio Prata as Ogron, Christian Iansante as Gantlos, Davide Lepore as Duman and Andrea Lavagnino as Anagan play The Fairy Hunters in the original version. In the Rai English version, Terrence Scammell as Ogron, Mark Camacho as Gantlos, Simon Peacock as Duman, and Thor Bishopric as Anagan.
- Tritannus is the main antagonist for Season 5. He is Aisha's cousin. He was once a normal merman who was sent to prison after trying to kill his twin brother Crown Prince Nereus. He was later mutated into a horrible monster due to toxic pollution. He currently seeks to rule the Infinite Ocean and absorb more pollution to become more powerful. He stole Daphne's Sirenix powers and gave it to the Trix. He and Icy are known to flirt with each other often, much to the annoyance of Stormy and Darcy.