Saints went into this afternoon's game unbeaten and breathing down the necks of National Group 2 leaders Manchester United, while the Potters were rooted to the foot of the table without a point.
But, despite dominating proceedings, Hunter's team – which included new goalkeeper Artur Boruc – were beaten courtesy of Jordan Richardson's 25th-minute strike against the run of play.
Southampton had the opportunity to level in second-half stoppage time when they were awarded a spot-kick, but top scorer Sam Hoskins saw his effort turned away by Stoke's impressive goalkeeper Daniel Bachmann.
"If you look at the statistics of the game, we definitely out-passed them and got into their final third lots of times but we need to be more ruthless and play with more guile," Hunter said at full time.
"From Artur right the way up to their last third, we were composed and did OK but to lose one-nil and miss a penalty is something we all need to learn from.
"The players are very disappointed, and we'll get back on the training pitch and work as hard as we normally do.
"There were some positives – I thought both the young centre-backs (Jordan Turnbull and Bevis Mugabi) were very composed, so that's a plus – but there's lots of things we need to work on."
Saints U21 0-1 Stoke U21: Report
Hunter made six changes to his starting line-up for what was Saints' first Barclays U21 Premier League game in over a fortnight.
Boruc replaced the injured Cody Cropper between the sticks, while the absences of Jack Stephens and Calum Chambers – currently away with England's Under-19s – opened up starting spots for Matt Young and Bevis Mugabi.
Luke Shaw was named as captain on his first appearance since a recent illness, while Steve De Ridder was also called into Under-21 action for the first time.
The first chance of the game went the way of the visitors as a cross from the right found midfielder George Rigg, but his shot was excellently turned away by Boruc.
Saints quickly found their feet, though, and almost went ahead when De Ridder's shot rebounded kindly into the path of Corby Moore, but the midfielder's drive flew inches wide.
Hoskins – who had netted five times in the U21s' previous four games – then had an opportunity to add to his healthy goalscoring account when a Moore corner found him at the back post but the striker's volley went just off target.
Instead, having survived some heavy pressure from the hosts, it was Stoke who took the lead when Richardson turned his man excellently before shooting beyond Boruc and inside the far post.
Saints responded well, with De Ridder drawing a good save from Bachmann before Mugabi nodded inches wide from another dangerous Moore corner.
Mugabi had another opportunity shortly after the break when some neat interplay in the Potters' box allowed him to loop a shot towards goal only for it to drop the wrong side of the crossbar.
Hoskins' frustrating afternoon continued as he turned another De Ridder cross wide, before a rampaging run by Shaw ended with his toe-poked shot being deflected wide by a superb save from Bachmann.
Hunter brought on Alberto Seidi and Omar Rowe in place of Jake Sinclair and De Ridder, but the clock continued to run down without them breaching the Stoke defence.
But they were handed a lifeline in injury time when Jordan Keane was penalised for handball, only for Hoskins' penalty to be parried away by the unbeatable Bachmann as Saints' frustrating afternoon was compounded.
Saints U21s: Boruc, Young, Mugabi, Turnbull, Shaw, Robinson, Moore, Reeves, Sinclair (Seidi 61), Hoskins, De Ridder (Rowe 82). Subs not used: Johns (GK), McCarthy, Curtis.
Stoke U21s: Bachmann, Glasgow, Campbell, Clarkson, Keane, O'Reilly, Hall (Richards 73), Rigg, Obiake (Alabi 33 (Rossi 80)), Richardson, Musungu. Subs not used: Eve (GK), Coulson.
Goal: Richardson 25
Bookings: Alabi 66, O'Reilly 69
Full highlights of this afternoon's game plus video interviews with Martin Hunter and midfielder Ben Reeves will be on Saints Player in the near future