Weston McKennie became the latest player to test positive for coronavirus.
The Juventus midfielder produced the result in the latest round of staff testing.
Juventus' first-team squad have now entered a voluntary period of isolation.
His result comes just 24 hours after Cristiano Ronaldo tested Covid-19 positive.
Juventus' squad have entered a period of voluntary isolation after Weston McKennie tested positive for Covid-19, just 24 hours after Cristiano Ronaldo produced a positive test while on international duty with Portugal.
United States international McKennie, who joined the club this summer from German side Schalke, was the only player to turn up a positive result in the latest round of testing.
As a precautionary measure ahead of a weekend game against Crotone, Andrea Pirlo's first-team squad will now isolate.
A Juventus statement read: 'Juventus Football Club announces that, during the checks provided for by the protocol in force, the player Weston McKennie has tested positive for Covid 19,' the official statement reads.
'In compliance with the regulations and the protocol, the team entered into fiduciary isolation this evening.
'This procedure will allow all persons negative to the controls to carry out regular training and match activities, but will not allow contact with people outside of the group.
'The Club is in constant contact with the competent health authorities.'
Juventus last match descended into chaos when Napoli were left unable to travel to Turin due to the commands of a local health authority.
Pirlo still named a starting 11 and players were seen at the stadium, with the expectation that thee champions would be awarded a walkover 3-0 win. The outcome of that was result was delivered on Wednesday.
Juventus themselves announced that two members of staff had coronavirus in the week leading up to the Napoli fixture and with McKennie's positive test, coupled with Ronaldo's, the club are looking to protect the squad.
It remains unclear what impact, if any, it will have on their game away to Crotone on Saturday night.
Ronaldo returned to Italy on board his private jet on Wednesday to spend the quarantine in his luxurious home in Turin. Upon arriving back in Italy he is required to observe a period of isolation and so will not participate in upcoming matches against Crotone in Serie A and Dynamo Kiev in the Champions League.
The presence of Ronaldo for the blockbuster Champions League match on October 28 against Lionel Messi's Barcelona is also under discussion.
It is reported he will need to produce a negative Covid-19 test by October 21 to be considered eligible to face his long-time Argentine rival.
Katia Aveiro, Ronaldo's sister, took aim at Covid-19 testing on her Instagram page following the Juventus attacker's positive result.
'If Cristiano Ronaldo has to wake the world up, I have to say he really is a prophet from God. Thank you she wrote.
'I believe that today thousands of people will stop believing as much in this pandemic, in the tests and in the measures taken like me ... the biggest fraud I have ever seen since I was born. Open your eyes please.'