In one of his guest appearances on Retsutalk, Jon mentions that all three guys are sharing a room together at the time causing Slowbeef and Diabetus to make a few comments along this line. Jon doesn't even bother to make a counter arguement, claiming he knows it wont do a thing at that point.
Later in Brawl, he asked if since Tim was playing Princess Zelda, did that make Josh a prince since Tim was his "wife". Apparently because of his infatuation with this song, Chugga ships Josh with Tim now.
If you pay close attention to their second New Super Mario Bros. Wii video, you can hear LOTS of talk like this.
In the 3rd vid Chugga says he can't live without Jon. Admittedly it's because he tends to die when Jon does, though.
When they reach the flagpole depending on where they landed, they'll go "I'm on top/bottom" or complain that they wanted to be on top.
Josh: *to Jon, after Jon nearly throws him off a cliff* You just almost fucked me.
Episode 11. Jon has invinciblity, Chugga doesn't:
Chugga: Can I have a hug so I can have star power?
Chugga: *fake sad* You never hug me anymore!
Jon: Because you don't deserve it anymore.
Jon: *overdramatic* I NEVER LOVED YOU TO BEGIN WITH!
Chugga: *fake sobbing* You did not just say that! My life no longer has meaning!
Near the end of episode 10, if you listen carefully, you can hear Jon calling the face Chugga is making "adorable".
Chugga has a screenshot of one of the enemies (looking like he's) sucking off Diddy Kong. Jon questions this.
NCS, in a Final Fantasy IV video, mentions a conversation he and Chugga had while they were staying with Jon in Canada... in bed.
In Episode 8, Chugga insisting that Jon pointed at him and winked, while Jon denies it by saying he was 'just blinking'.
In episode 9 during the 2 dark Samus fight, Chugga seems like he wants to see Jon naked.
Chugga says that Zero Suit Samus's name is technically inaccurate, as she's actually wearing a suit. Proton Jon replies to this with "It's not a birthday suit." Chugga brings up that it's Jon's birthday. Hilarity Ensues.
Jon: Are you certain you're straight? Because you keep saying things which a straight person should not be saying. Chugga: I didn't mean it like that! I meant you threw the bomb at the... Jon: And you wonder why there's slash.
During Minigame Coaster, Chugga accidentally kicks Tim's leg and feels something in his pocket. He asks if it's his phone, but it's actually his wallet, and when Jon realizes what's going on he has this line:
Within all the screaming at the end of the tournament, we hear this little gem?
Jon: Emile why are you humping him? Stop that!
New Super Luigi U
In Episode three, Chugga compares Jon to Viridi, while his intention was saying Jon was being Tsundere, Chugga quickly realized that not only have people compared him to Pit, but thanks to his still-recent LP of Uprising, it was well-known at this point he openly shipped Pit/Viridi.
In Episode 8, Chugga says that he will suck all of the emotion out of the other Guys' cockles. He amends this to "the cockles of your heart," but not before Jon gets in a Big "WHAT?!".
In the ninth episode, the guys play another one-up minigame, where Chugga assumes they'll only end up with a handful of one-up mushrooms. When they leave with six extra lives, Chugga gives us a brilliant moment of innuendo:
Chugga: Oh, two or three-up my ass! Jon: ... you might wanna rephrase that.