While the prime minister did not speak, his brother Casper speaking for the family at the graveside said: "Susan would have wanted us to move forward."
Truck hit car
Susan Tsvangirai was killed in a car crash last Friday in which her husband of 31 years was injured.
Zimbabwe's long history of violence and several suspicious car accidents fuelled speculation that the crash was not an accident. However, Tsvangirai has since ruled out foul play.
Mugabe, at Tuesday's church service, called for an end to political violence and said the crash was an act of God.
The crash occurred near Beatrice, a town about 60km from Harare, the capital.
Tsvangirai was sworn in as prime minister of a national unity government in February after months of political wrangling between Mugabe's Zanu-PF party and the MDC over a disputed presidential election, which the MDC claims Tsvangirai won.
The new administration is tasked with tackling severe food and fuel shortages, record hyperinflation and a cholera outbreak that the World Health Organisation (WHO) says has killed at least 4,000 people.