Maybe a new mutant created by the Foot Clan. This mutant could be specifically designed to target the sister. Since she's new to the team, she might not be as experienced in fighting, making her an easy target for this new villain. The new mutant could have powers that are the opposite of the sister's, if she has any special powers. For example, if the sister has the power of healing, the new mutant could have the power to spread disease or cause harm in some way.
It could be Shredder as always. He might see the sister as a new threat or a way to get to the turtles. He could try to kidnap her to lure the turtles into a trap.
Krang from Dimension X could also be a great villain. He might be interested in the human sister because she has some unique knowledge or ability that he wants to use for his own evil plans in taking over Earth. He could send his minions to capture her and the Turtles would have to journey to Dimension X to save her.
Perhaps the sister was created by the same mutagen as the turtles but was separated from them. When she finally finds her brothers, there could be some initial distrust and confusion. But as they get to know each other, she becomes an important part of the team. She might have a different personality compared to the turtles, like being more diplomatic, which could help in dealing with some of the more complex situations they face, such as negotiating with other mutants or humans who are not immediately hostile.
Perhaps a new villain emerges. A powerful sorcerer who is angry that the turtles have been turned human by a magic that he believes should be under his control. So he tries to either turn them back into turtles in a more twisted way or simply destroy them in their human forms because he sees them as abominations.
April O'Neil could also be a main character. She might be the one who discovers the mecha technology or helps the turtles figure out how to use it. Her role as a human ally would be important in a story like this.
Perhaps there are new mutants created by some mad scientist who are bent on destruction. These mutants could be stronger and more cruel than the usual villains the Turtles face. They might have abilities that are specifically designed to counter the Turtles' skills.
The main characters are, of course, the four Ninja Turtles: Leonardo, the leader who is disciplined and strategic; Raphael, the tough - as - nails and hot - tempered one; Donatello, the intelligent and tech - savvy turtle; and Michelangelo, the fun - loving and laid - back member of the group. Also, their sensei, Splinter, a rat who trained them in the ways of ninjutsu, is an important character.
You can try looking for them at your local comic book store. They might have a good selection.
A powerful warlord who wants to conquer both Ceez's world and the ninja world. He has an army of dark warriors and uses dark magic to try and achieve his goals. He sees Ceez and Ninja as the biggest threats to his plans, so he constantly tries to eliminate them.
Maybe the human sister is a scientist who is working on a project that could potentially help the Ninja Turtles in their fight against evil. But the bad guys find out about it and try to stop her. Or she could be an artist who uses her art to inspire the Turtles and give them new ideas for their strategies. She might also have a secret past that is somehow connected to the mutagen that created the Turtles.
A jealous witch who was the one who originally cast the curse on the Beast could be the main villain. She might see the little sister's presence as a threat to her plan of keeping the Beast trapped forever. Maybe she tries to trick the little sister into making things worse for the Beast.