The Women's Super League side produced an impressive display to beat Barcelona on matchday one, while St Polten were unable to take anything from their away meeting with Hammarby.
Match preview
St Polten are featuring in the final 16 for the fourth time in five seasons after successfully negotiating their way through qualifying.
The Austrian champions ousted Neftci Baki and Vllaznia in the first stage of qualifying, before they cruised to an 8-0 aggregate win over ZNK Mura to seal their spot in the main draw.
Unfortunately for St Polten, their run of European wins came to an end with a 2-0 away defeat in their group opener against Hammarby.
As a result of that loss, St Polten already know they will have to produce something special to reach the knockout rounds, with their next four European matches set to take place against Man City and Barcelona, before they wrap up the group stage with an away clash with Hammarby.
Having failed to win their last 10 group games, St Polten will head into Wednesday's contest with hopes of claiming their first victory in the main draw since beating Slavia Prague in November 2022.
Man City will carry significant momentum into the midweek fixture after kicking off the campaign with seven wins and one draw in all competitions.
Gareth Taylor's side recorded two of those victories in their qualifying tie against Paris FC, before they recorded another win in last week's home game against three-time European champions Barcelona.
Man City produced an impressive display to claim a statement win over the Catalan giants, with Naomi Layzell and Khadija Shaw grabbing a goal apiece in a memorable 2-0 victory at the Joie Stadium.
Shaw went on to play a key role in Sunday's WSL game against Liverpool, scoring a second-half brace to help her side come from behind to clinch a 2-1 victory at Anfield.
The striker's stoppage-time winner moved Man City onto 10 points, leaving them top of the WSL table with a one-point advantage over Chelsea and Manchester United, who both have a game in hand.
The WSL leaders will now turn their focus to Wednesday's away trip, where they will attempt to claim a sixth consecutive victory in all competitions.
St Polten Women Women's Champions League form:
W
W
W
W
L
St Polten Women form (all competitions):
Manchester City Women Women's Champions League form:
W
W
W
Manchester City Women form (all competitions):
D
W
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W
Team News
The Austrian champions remain without the services of defender Ella Touon, who is currently sidelined with a knee injury.
Tea Vracevic could return to the St Polten backline after dropping to the bench for Sunday's league win over Dornbirn.
St Polten boss Celia Brancao may also be tempted to recall Sophie Hillebrand and Sarah Mattner-Trembleau.
As for the visitors, they are unable to call upon Risa Shimizu and Sandy MacIver due to anterior crucial ligament injuries.
Layzell is set to miss a second straight game after she picked up an injury in the closing stages of the 2-0 win over Barcelona.
Vivianne Miedema was absent for Sunday's league clash against Liverpool, and the Netherlands international will have to be assessed ahead of the midweek trip to Austria.
Shaw will be looking to continue her fine run of form, having scored six goals in her last five appearances in all competitions.
St Polten Women possible starting lineup:
Schluter, Krizaj, Vracevic, Klein, Wenger; Mikolajova, Hillebrand, Dubcova; Madl, Mattner-Trembleau, Pekel
Manchester City Women possible starting lineup:
Yamashita; Casparij, Aleixandri, Greenwood, Ouahabi; Park, Hasegawa, Roord; Fowler, Hemp, Shaw