The 15-player squad will be led by Laura Wolvaardt and is made up largely of the players who have helped the team to consecutive runners-up places at the T20 World Cup.
South Africa have also reached the semi-finals of the last two 50-over global finals, but a first major trophy continues to elude them.
Van Niekerk, 32, who retired from international cricket in March 2023 having been left out of the squad for the T20 World Cup that year, had hoped for a return having continued to play domestically.
But coach Mandla Mashimbyi suggested last week it was highly unlikely. Van Niekerk is married to key all-rounder Marizanne Kapp, who has been selected.
South Africa will tour Pakistan from September 16-22 for three One Day Internationals in preparation for the World Cup, where they play their first fixture against England in Guwahati on October 3.
South Africa 2025 World Cup squad:
Laura Wolvaardt (captain), Anneke Bosch, Tazmin Brits, Nadine de Klerk, Annerie Dercksen, Sinalo Jafta, Marizanne Kapp, Ayabonga Khaka, Masabata Klaas, Suné Luus, Karabo Meso, Nonkululeko Mlaba, Tumi Sekhukhune, Nondumiso Shangase, Chloé Tryon.