Chelsea midfielder Jorginho and Brescia star Sandro Tonali will require coronavirus tests before linking up with Roberto Mancini's Italy squad.
The Italian Football Federation (FIGC) said on Thursday that both will have tests once their "respective isolation requirements are complete", and only then will they be clear to join Mancini's group at Coverciano from Saturday.
The extended Italy squad includes uncapped trio Alessandro Bastoni of Inter and Sassuolo duo Manuel Locatelli and Francesco Caputo, while captain Giorgio Chiellini is back after missing a year of international football due to injury.
Tonali's Brescia team-mate Mario Balotelli has not been called up.
Italy face Bosnia-Herzegovina in Florence on September 4 and the Netherlands three days later in Amsterdam as they get their Nations League campaign underway.
Italy squad in full:
Alessio Cragno (Cagliari), Gianluigi Donnarumma (Milan), Alex Meret (Napoli), Salvatore Sirigu (Torino); Francesco Acerbi (Lazio), Alessandro Bastoni (Inter), Cristiano Biraghi (Inter), Leonardo Bonucci (Juventus), Mattia Caldara (Atalanta), Giorgio Chiellini (Juventus), Danilo D'Ambrosio (Inter), Giovanni Di Lorenzo (Napoli), Alessandro Florenzi (Roma), Gianluca Mancini (Roma), Luca Pellegrini (Juventus), Leonardo Spinazzola (Roma); Nicolo Barella (Inter), Giacomo Bonaventura, Gaetano Castrovilli (Fiorentina), Bryan Cristante (Roma), Jorginho (Chelsea), Roberto Gagliardini (Inter), Manuel Locatelli (Sassuolo), Lorenzo Pellegrini (Roma), Stefano Sensi (Inter), Sandro Tonali (Brescia), Nicolo Zaniolo (Roma); Andrea Belotti (Torino), Federico Bernardeschi (Juventus), Francesco Caputo (Sassuolo), Federico Chiesa (Fiorentina), Stephan El Shaarawy (Shanghai Shenua), Ciro Immobile (Lazio), Lorenzo Insigne (Napoli), Moise Kean (Everton), Kevin Lasagna (Udinese), Riccardo Orsolini (Bologna).