The prisoners were kept in constant isolation for 14 days at HMP Birmingham.
An inspection also found all inmates were kept in their cells for 23 and a half hours at the height of the pandemic between March and July.
Prison governors have been forced to take extreme measures to try and prevent the spread of Covid, but the lenght of time prisoners have had to spend in isolation has raised concerns about the impact on their health.
HMP Birmingham, which has around 1,000 inmates, had three confirmed coronavirus cases, all at the start of the pandemic.
The jail's monitoring board praised efforts to stop the virus spreading but said the treatment of those suspected of having the virus was "extreme and unacceptable over a long period".
However, the report added: "It needs to be considered in the context of the prison having only three confirmed prisoner cases of Covid-19. Men generally accepted the restricted regime as keeping them safe, and commensurate with some of the restrictions in the outside community."
The monitoring board said the longer the crisis had gone on, the more some prisoners' willingness to comply had begun to wane.
"Frustrations have increased as the COVID-19 regime restrictions have continued over months," the report said, adding verbal attacks and spitting towards officers had increased.
Bosses at HMP Birmingham were praised for their efforts in helping to reduce drug use and violence, following damning criticism which saw the prison taken back into state control in 2018.
The report said: "From June 2019 to March 2020, the Board judged the prison to be much safer than before but still subject to some serious assaults and incidents of violence."
It continued: "It was confirmed that positive and covert action was being taken to tackle the ingress of drugs, and this has been independently verified by security staff."
The Express & Star revealed earlier this month how prisoners were still being kept locked up for up to 23 hours a day in some jails, raising questions about the long-term management of offenders.