Inspectors found male and female staff and governors were being divided in Luton's Rabia Boys and Girls School, said the education watchdog.
Last year, Sir Michael wrote to the education secretary with "serious concerns" about staff segregation.
But the Rabia school has strongly rejected the claims and said it was being "unfairly targeted".
Sir Michael has written to Education Secretary Nicky Morgan advising her that for the initial meeting with inspectors "the school insisted on segregating men and women through the use of a dividing screen across the middle of the room".
"This meeting was not carried out in a religious setting but in a classroom."
Sir Michael said inspectors, visiting the independent school earlier this month, "gathered evidence that male and female staff are segregated during whole-school staff training sessions". [BBC] Read more
