A calf injury kept Messi out of action in the early weeks of the campaign, though he came off the bench against Granada on Saturday.
Barca lost that match 2-0 and have made their worst start to a LaLiga season for 25 years as a result, taking seven points from five games.
🔵🔴 Barça XI#BarçaVillarreal— FC Barcelona (@FCBarcelona) September 24, 2019
Under pressure coach Ernesto Valverde has responded by thrusting Messi into his starting lineup, the captain replacing Carles Perez in Barca's front three.
Messi - crowned FIFA's Best Men's Player on Monday - will make his 400th LaLiga start for Barca and he becomes only the second non-Spanish player, after Donato, to reach that milestone.
He will be joined in attack by Luis Suarez and Antoine Griezmann with prodigy Ansu Fati included on the bench with fit again France forward Ousmane Dembele for Tuesday's game.
Ivan Rakitic drops out of the XI after making his first start of the season against Granada, with Sergio Busquets and Arthur coming into midfield alongside Sergi Roberto after Frenkie de Jong was also left out for the first time since joining from Ajax.
Barcelona are unbeaten in their last 20 LaLiga games against Villarreal and have won their last nine LaLiga home games despite their poor start to the 2019-20 campaign.