Ghana sped into a 2-0 lead with goals from the Lyon forward Ernest Nuamah and Félix Afena-Gyan.
But Gambia hit back on Tuesday night with two strikes before half-time at the Accra Sports Stadium.
Ebrima Colley put Gambia ahead midway through the second half before the 26-year-old struck again in stoppage-time to wrap up the points in the Group C tie.
The defeat – following on from the 2-0 loss in Côte d'Ivoire last week – left Ghana bottom of a four-team group with no points after two of the six qualifying games.
'I'll be the first one to resign'
"If you believe that with the coach resigning, you can change the situation of football in Ghana, I'll be the first one to resign," said Queiroz during a fractious interchange with Ghanaian reporters after the game.
"I don't have that belief but that is a decision for the board of the football federation to analyse."
Ghana's football chiefs recalled 73-year-old Queiroz to lead the country's 2027 Cup of Nations campaign after he stepped down following their run to the last 32 at the 2026 World Cup in the United States, Mexico and Canada.
They will attempt to relaunch their Cup of Nations campaign with two games against Somalia in November while Gambia and Cote d'Ivoire slug it out at the top of the group.
"We cannot undo the result against Gambia," added Queiroz who apologised to fans for his side's performance.
"But what we can do is to be better prepared, better organised for the next games and win the next games. This is what we have to do."
Nigeria crash in Guinea-Bissau
Nigeria coach Eric Chelle was also on the defensive after his side slumped to a 3-0 defeat in Guinea-Bissau in Group L which contains co-hosts Tanzania as well as Madagascar.
Under competition rules, only one other team will progress from a pool containing a co-host.
Franculino and Mama Baldé, who were both on target in Guinea-Bissau's win over Tanzania on 25 September, continued their fine form to give the hosts a 2-0 lead. Álvaro Djaló added the gloss in second-half stoppage-time.
"Guinea-Bissau were more motivated and more aggressive than us," Chelle conceded.
"They put more intensity into everything and we never found the solution. But this is not the end. You know, we have a lot of games to play so we have to stay focused, calm. It was a bad day for us but I'm still confident about my team and my players."
Guinea-Bissau set the pace
Guinea-Bissau lead the pool with a maximum six points while Nigeria lie second with three. Madagascar and Tanzania both have one point following their 1-1 draw in Meknes.
Elsewhere, in Group A, Morocco, who were awarded the 2025 championships despite losing in the final 1-0 to Senegal, made it two wins out of two following a 2-0 victory against Lesotho.
Brahim Diaz struck on the stroke of half time and Azzedine Ounahi secured the victory eight minutes from time.
Senegal, who are appealing to the Court of Arbitration for Sport against the decision of the Confederation of African Football to strip them of the title, won 1-0 in Ethiopia to furnish new boss Patrick Vieira witha first victory.
However, there was more disappointment for Cape Verde and their new coach Humberto Bettencourt.
Following an opening day defeat in Mali, Bettencourt's side went down 2-1 at home to Rwanda and prop up Group K where Mali set the pace with six points after a 1-0 win over Liberia.



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