The pontiff traveled to the Shrine of Our Lady of Aparecida by plane and helicopter.
On Monday Pope Francis faced chaotic scenes when crowds mobbed his car as he arrived in the city of Rio de Janeiro.
He is in Brazil to attend the Roman Catholic World Youth Day festival. It is his first trip abroad since becoming head of the Catholic Church in March.
The highlight of the five-day festival will be a prayer service led by Pope Francis on Rio’s iconic Copacabana Beach on Thursday.
The pontiff’s visit is taking place under tight security, after weeks of protests against the government and corruption.
Thousands of pilgrims arrived in the rural town of Aparecida ahead of Wednesday’s Mass, scheduled to take place at 10:30 local time at the basilica holding the shrine.
A homemade explosive device was discovered there on Sunday, but authorities said it was “of low power” and not near the area where the Pope and pilgrims will visit.
The Vatican later insisted that the discovery was no cause for concern.
Pope Francis is expected to fly back to Rio de Janeiro on Wednesday afternoon to visit a hospital and address young people taking part in a drug rehabilitation program.
On Tuesday, around 400,000 Roman Catholics joined a ceremony on Copacabana Beach to mark the opening of World Youth Day.
There was transport chaos shortly before the event when the Rio metro system was brought to a standstill for two hours by a power cut.
Crowds of people hoping to attend the Mass were left stranded, while others crammed onto buses and taxis heading to the beach.
Travel chaos also ensued on Monday when the pontiff’s car became stuck in one of Rio’s infamous traffic jams, after his driver reportedly took the wrong turn and missed lanes that had been cleared by the security services.
Correspondents said Brazil was reviewing security around the pontiff following the incident.
Pope Francis, who is from neighboring Argentina, was officially welcomed at the state governor’s palace by Brazil’s President Dilma Rousseff on Monday.
However, police later fired tear gas to disperse people who were demonstrating outside. They were protesting against the government and also against the cost of the papal visit.
More than a million young Catholics are expected to gather in Rio for World Youth Day, which takes place every two years, and is a celebration of the Catholic faith.
Some 30,000 army and police are on duty throughout the Pope’s visit.
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