England seamer Kate Cross has been ruled out of the one-off Women's Ashes Test starting at the Melbourne Cricket Ground on Thursday.
Captain Heather Knight confirmed that Cross, 33, had not sufficiently recovered from a back injury that she suffered against South Africa in December.
Cross was not part of the T20 side for the Ashes series but missed the three one-day internationals which opened the tour with the same issue.
"Kate is pretty much close to full fitness now, but just not quite at the pace and the performance that we know she can reach at her best," Knight told BBC Sport.
"A Test match is four days, so she will miss out and it is a real shame for her.
"She's a really key part of our group, she loves playing Test match cricket and she's been so desperate to play. We're all really disappointed for her that she hasn't been able to make it."
It is a significant blow for England as Knight will remain without her most experienced seamer.
Knight did not reveal the England team as they were still deciding on the bowling attack, but confirmed that they would be playing an extra batter in Sophia Dunkley.
Lauren Bell, Lauren Filer and all-rounder Nat Sciver-Brunt are likely to form the seam attack.
England have lost all six white-ball matches on the tour so far and are fighting to avoid a 16-0 clean sweep, which has never been done before in the multi-format series.
"There's a lot of hurt and disappointment around how this tour has gone," Knight added.
"We haven't shown what we are capable of as a team, but we want to do that this week and show our character to the fans that are here and watching back home."
A Test win is worth four points and a draw two each.
England also went winless in the 2022 series in Australia but they finished with four points overall because of a drawn Test and two T20 washouts.
Knight would not be drawn into the debate surrounding her future as captain, and did not confirm whether she would remain in the role after the conclusion of the series.
She has led the side since 2016 after the retirement of Charlotte Edwards and has worn the heavy burden of England's disappointing performances on the tour.
The Melbourne Test will be her 199th game in charge in all formats.
"I'm just concentrating on this week," said Knight.
"There's been lot of frustration around this tour that we haven't performed how we want to, and you always take that on your shoulders as the leader because you're at the helm of it all.
"This week is a chance for us to show our best cricket and, whatever happens after that, we'll see."
On Tuesday all-rounder and vice-captain Sciver-Brunt said she would be interested in taking on the captaincy if the opportunity arose, but no timeframe has been given around the decision, with England out of action until May after the Ashes concludes.
There is also speculation surrounding the future of head coach Jon Lewis, who has also made headlines in the Australian media for his comments around "climate benefits" which contribute to the opponents' superiority in athleticism.