A space test monkey named Bingo crash-lands in the barnyard. The animals decide to let him stay in the barnyard, but Bingo is unsatisfied with the fact that Otis is the leader, so he tries to get rid of Otis so he can become leader.

When Otis gets a hypnosis kit, he accidentally hypnotizes himself so that he tries to destroy the farmer whenever he hears the sound of a bell. Now, the other animals must try to protect the farmer while trying to cure Otis at the same time. But every time Abby tries to stop Otis from hurting the farmer, he thinks she is hurting him.

Otis is embarrassed when Abby repeatedly beats him in the Barnyard Olympics, so he tries to find Abby's weaknesses and use them to his advantage.

Otis is excited because favorite country music star – Stumpity Joe – is coming to town. However, when trying to speed up crop collecting for the farmer to leave, he accidentally injures the farmer. Now, the animals must do all of the farming, or else the farm can go bankrupt. But several things had gone wrong because the animals aren't accurate farmers themselves. Otis milks himself wrinkly with the milk machine, Bessy and Abby refuse to use the milk machine on themselves, Pig eats all of the corn they picked (making him very fat), and Freddy goofs with the energy drink for the hens by putting dynamite in it, which he claims it is pepper, making the eggs the hens lay explode on contact.

When Duke tries crazy methods for the sheep to be safe during bear season, they escape and now Otis and Duke must go on a cross country trip to get them before they escape to New Zealand. Meanwhile, Abby and Bessy dress Pig, Freddy, and Peck as sheep. But it backfired when the farmer thought Pig had the "nervous woolies" when Pig accidentally said "Beeeee" and "Boooo" instead of "Baaaa".

When the veterinarian stops by, she mentions that her van, the "Big Steer", won't last a week. However, Otis mistakenly thinks the veterinarian was referring to him and begins to think that he only has a week to live. To relieve himself, Otis tries to do everything he has ever dreamed of. When he returns, the animals are happy that he was gone so he abandons them. When he tried to do a jump over a canyon, he fell to the bottom and got injured. After that, the animals apologize to him for kicking him out of the barnyard.

Otis' favorite circus is coming to town, but the gang accidentally injures the main lead. Feeling guilty, Otis offers to take over and along with his friends, but Abby is too afraid to do her part because of an incident that happened long ago: She had dropped her partner when trying to catch him. Abby refuses to do the part because of being afraid to drop Otis. But without Abby, the show ends up being a disaster

Otis' negligence causes Pip to get captured by Snotty Boy, who plans on feeding him to his new pet snake (who talks exactly like him).

Pip's mousehole is destroyed by Otis accidentally. He moves in with the others, but they're all unused to each other's habits. After hearing a misunderstanding that is impossible to share a home with Pip, he runs away and finds a bee hive. Otis and his friends must search for him and return him to the barnyard.

When the gang gets caught by Snotty Boy in a game of "Dungeons and Barn Animals", they make Snotty believe he is in the mythical land of Barnia, and now the animals must force him out before he takes over the barn.

When it's discovered that Pig has a beautiful singing voice, Otis and the gang enter Pig in a singing competition to win the farmer a golden tractor. When Mrs. Beady loses, she is out to prove that Pig is a talking barnyard animal. Meanwhile, Pig's over-practicing causes him to lose his voice. So Otis tries to use lip synching but he doesn't know the lyrics in the song, so Pig is booed for singing terrible lyrics. In the end, Pig did win a prize (a tiny tote bag), but they couldn't keep the tractor they stole because it was illegal to steal a grand prize without approval of winning.

Otis becomes afraid when he receives a letter from his childhood bully, Krauser Krebs, saying that he's coming to visit the barnyard, so his friends try to help him defend his fear of the bully.

Peck takes a fake potion to get enough courage to speak to his crush, Hannah, but he ends up being so confident that he challenges Root, a rooster much bigger than him, to a duel.

When Freddy gets depressed over his lack of talent, Otis and the gang decide to make him take up ventriloquism. The dummy that they make, Mr. Jinx, comes alive, however, and tries to kill them. It turns out that a colony of termites has possessed Mr. Jinx to get revenge on them for using their tree to create Mr. Jinx.

Otis' new Kobe cow neighbors begin to think that he's the reincarnation of the legendary Sun Cow when they discover Otis has a fifth udder (which was actually a bee sting). Otis is glad he has that status until he realizes that the cow neighbours actually serve "Sun Cows" at a steak restaurant and they're planning to do the same to Otis.

While Duke is at the vet after being sprayed by Pig’s skunk, a mischievous mutt named Baxter sneaks out and goes to the barnyard while disguised to look like Duke for what his life is.

Otis uses Crazy Louie – a loopy groundskeeper at a high-end golf course and apparently his friend – to let him and the animals through the back gate and play a round. When Pig throws a club and accidentally hits Louie in the appendix, though, a guilt-ridden Otis decides to win the big golf tournament to get a new appendix.

Abby is excited that her cousin, Veronica, is coming to the barnyard for a visit, but it turns out Veronica, despite appearing to be nice on the outside, is actually using her beauty to manipulate the animals into doing things to please her.

Pig writes a love letter to his favorite pastry treat, but Otis finds it and thinks Abby wrote it to him.

When responsibility is thrust upon Otis to watch the three kids of the barnyard, Joey the calf, Macy the lamb, and Boyle the chick, after Abby and Bessy read story time, Otis loses them upon taking them around the big city when Otis and the gang had to remove a chip from Freddy's skin to keep Abby and Bessy from finding out where they are. So Otis and the gang stage a fake robbery to reunite with the kids after a cop finds them and about to take them to the station. But soon the kids call Abby & Bessy to pick them up. On the way back to the barnyard in a helicopter, Bessy tells them about Otis being a lair and after each line, she punches Otis and with him asking for a parachute to bail the helicopter.

The animals buy a security system called The Barn Buddy (voiced by Gilbert Gottfried) for the barnyard, but it turns out that the system is doing more harm than good.

The animals find an old safe in the barn that belonged to Everett's former owner Nebraska Schwartz. Everett doesn't remember the lock combination to open the safe, so Otis helps Everett recreate past memories of Everett's adventures with Nebraska Schwartz to help Everett recall the number.

The animals liberate a show whale from a water show at a fair, but Otis has a lot more to go when the animals are being constantly eaten by this whale.

The animals must stand in for injured football players in a game and the coach’s mother to like him after becoming the mascots for the team.

n a bid to collect the most Halloween candy and win a prize, Otis tops off the animals' trick-or-treat candy with a handful of candy from a hollow tree that legend says belongs to Hockey Mask Bob who always gets his candy stolen and vows to return for revenge and reclaim his candy.

After Freddy's recipe for a tofrooster that is going to be cooked on the show, things get worse when Chef Big Bones (voiced by Jim Cummings) insists on using a real rooster, and captures Peck, so Otis disguises himself as a chef to save Peck from being cooked.

Bessy is constantly rude towards the animals, even though she is barely seen. The animals wonder what she does when they don't see her and try to spy on her.

After Otis accidentally breaks up Peck's body with a hammer, he rebuilds Peck by adding robot parts to him so he can be a big help around the barnyard, but things go awry when Peck's circuits get overloaded after Otis puts pizza inside him and he goes out of control.

The animals successfully prank Mrs. Beady into taking a holiday. Now, they no longer have to worry about getting caught by her, but now Otis experiences withdrawal symptoms because he is no longer able to prank her.

When a beautiful she-donkey named Prunella takes an interest in Pip, Bessy suspects ulterior motives and follows them on their dates, causing the other animals to accuse her of being jealous.

A group of Canadian Crows want to steal the corn from the barnyard, but after repeated failed attempts, they decide to give the animals a fake treasure map to distract them from guarding the corn.

An ancient cave cow comes to attack the barn animals! Meanwhile, scientists kidnap Otis, thinking he's really Cave Cow.

The leprechaun steals the farmer's potatoes after Otis sprayed the potatoes, Otis and pals must stop him before the farmer goes bankrupt and the animals for less outlet. Unfortunately, also, the leprechaun turns Freddy and Peck into shamrocks and Pig into a corn beef sandwich head. Luckily, Otis gets an idea to cut out the middle man meaning to find the leprechaun's gold instead of catching him to surrender.

Pig's mole subjects (from when he was Emperor of the Mole People for 45 minutes) have sought him out to return as their Emperor and defend them from Eric the Terrible, a verbally abusive worm and with the mole people around the barnyard, they can't get Pig to hang out with them because they keep shocking them when they are near Pig.

The animals call Dr. Filly to settle a recent persistent argument between Freddy and Peck, but instead of making things better, Dr. Filly seems to make it worse when other animals begin disliking each other. They were almost going to leave (except Bessy) until Otis reveals that "Dr. Filly" is an imposter donkey named Chip who likes to impersonate Dr. Filly.

In the series finale, Otis orders a satellite dish online in order to watch monkey boxing from Rio. Mrs. Beady sees this and believes that he is secretly signaling Aliens. Otis finds out and tries to prank Mrs. Beady with a fake alien invasion broadcast. This broadcast is sent to real aliens who are attracted to Earth and plan to conquer and destroy the planet. The animals must stop and stall them while they figure out their weakness.