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The 37-year-old chased down Spaniard Margarita Victoria Garcia, who had attacked with 45km to go.
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With 7km to go Garcia was caught by the imperious Van Vleuten, who then dropped her rival and rode out for the win, following up on her victory in the same race last season. Michal Kwiatkowski and Fabian Cancellara have won the men's race on multiple occasions but not back-to-back.
'This rider can do no wrong' - Van Vleuten seals Strade Bianche win
The victory is Van Vleuten’s fourth since the re-start and the road world champion has picked up where she left off before the season was suspended.
"Usually I try to think opposite. Everything that is hard for me is good for me. I love the heat, it's good for me. We knew there would be possibilities for attacks and Amanda Spratt did a really good job," Van Vleuten said afterwards.
“So many emotions today. I thought I came here for nothing because, for my team-mate was doing well, but for myself I cannot make my own attack. Suddenly, the girls were away and Mavi Garcia, a really strong rider, which was beautiful for the race, and I had to go full-in that attack.
"There were a lot of emotions. The Piazza del Campo is the most beautiful finish for me to win. A lot of emotions.”
Van Vleuten didn't come to Strade Bianche expecting to win