DeChambeau holed 143 feet of putts in a 63 that moved him to 11 under but he had Van Rooyen breathing down his neck after the South African equalled the course record at Chapultepec Golf Club with a 62.
On his 30th birthday, Van Rooyen made nine birdies in a blemish free effort to get to ten under alongside Patrick Reed after being three over after seven holes on day one.
American Reed matched countryman DeChambeau with a 63 but incredibly surpassed his putting performance - holing 150 feet and eight inches.
World Number One Rory McIlroy was then at eight under as he birdied three of his last five holes to rescue a 69.
Van Rooyen announced himself to the world with a top ten at the 2019 US PGA Championship and later that year he would claim a maiden European Tour win en route to a top ten finish on the Race to Dubai Rankings Presented by Rolex.
His 62 ties Retief Goosen for the lowest round ever shot by a South African in a WGC, and life is good for Van Rooyen as he enters his fourth decade.
"The first thing I told my wife this morning: 'man, I feel really old'," he said. "But I guess I'm still quite young and what a way to kick off the 30th and shoot nine under. A little bit of a gift to myself.
"Retief is obviously someone I've always looked up to, watched him win his Majors and win a lot of really nice events when I was a kid, so it's good to be in that category.
"I've been working really hard on the game, as we all do, and things came together nicely today, so I'm happy."
DeChambeau won last season's Dubai Desert Classic after making his Ryder Cup debut in 2018 but the American is still searching for his first World Golf Championships victory.
"It was a lot of fun," he said after coming home in 29. "Surely when you have days like that you can't help but smile out there? Making those putts the way I did, striking the ball the way I did, it's surely a joy out there.
"The confidence has got to be high. It is high and it's a lot of fun to see putts finally going in. Overall everything is going pretty well, firing on all cylinders."
American Thomas sandwiched a bogey on the 14th with birdies on the par five 11th and 15th and then looked like he would run away with it as some stunning iron play made it four birdies in a row.
When he drove the short first and holed from 35 feet for an eagle, he led by three but DeChambeau set about reeling him in.
The 26-year-old birdied the two par fives on the back nine with a dropped shot on the 14th but then made four birdies in a row from the 18th - all from within 15 feet.
A wonderful approach to the fifth put him in a share of the lead as Thomas bogeyed the fourth, and DeChambeau holed a 23 footer on the sixth before finding the cup from twice that distance on the next to get to 11 under.
Van Rooyen got up and down on the driveable first and second before taking advantage of the par five sixth.
A third chip-in of the week on the ninth then sparked a run of four birdies in a row to catapult him up the leaderboard.
A birdie on the par five fifth had him within two of the lead and a gain on the last from six feet made him the nearest challenger.
Reed - the 2018 Masters Tournament winner - made four birdies in a row from the third but dropped a shot on the eighth to turn in 32 before sandwiching long putts on the 11th and 17th with further birdies on the 12th, 14th and 15th.
Japan's Matsuyama turned in 32 with four birdies and then birdied five of six holes from the first before dropping shots on the seventh and eighth.
Northern Irishman McIlroy had to scramble for par on his first four holes and dropped shots on the ninth and 13th but made gains on the 12th, 14th, 16th and 17th.