Panayiotou and his co-accused, Nenembe and Sibeko, are facing charges of conspiring, kidnapping, robbing and killing Jayde Panayiotou.
A fourth suspect, Sizwezakhe Vumazonke, the alleged hitman, died before the trial began.
Jayde, a teacher, was kidnapped outside her townhouse complex in Kabega Park, Port Elizabeth, while waiting for her lift to Riebeek College Girls High School, on the morning of Tuesday, April 21, 2015.
She was found shot dead in a veld outside KwaNobuhle township, Uitenhage, the following day.
The State is trying to prove that Panayiotou ordered Jayde's murder.
At their previous appearance in June, the State called witnesses to testify on Panayiotou's possible motives for having his wife murdered, and about his affair with his employee, Chanelle Coutts.
Clarishka Kapp, a friend of Coutts, said Panayiotou had told her he was worried about losing Coutts when he married Jayde.
The State also called Vumazonke's aunt, a traditional healer, who testified to having seen her nephew the day of the murder.
The aunt had previously made a statement that Vumazonke had confessed to her that he was involved in the murder.
However, the confession was ruled inadmissible due to being hearsay evidence, as Vumazonke's death meant he could not challenge his aunt's version of events.