The Tykes suffered a shock 2-0 loss at Coventry City on Sunday to remain sixth, while Huddersfield strengthened their survival hopes with a win by the same scoreline at Nottingham Forest.
Valerien Ismael's side have been in exceptional form since the end of January, winning 11, drawing four and losing just two of their last 17 league matches.
However, Barnsley's loss at St Andrew's Stadium at the weekend, combined with a point for Reading against Cardiff City, leaves them looking over their shoulder at the chasing pack.
Only Reading, Cardiff and Millwall can catch Barnsley in the final playoff spot, though the Royals - four points worse off - are the Tykes' only realistic concern.
The defeat against Coventry ended a club-record run of six successive away league victories, and Ismael will now be after a swift response from his side.
Huddersfield, by comparison, are winless in five at the John Smith's Stadium, having drawn four in a row on home soil before losing 2-1 to Bournemouth last week.
The Terriers followed up that loss with a 2-0 victory at Forest on Saturday, however, to move eight points clear of the relegation zone with four games to go.
However, that gap could potentially be just two points should 22nd-placed Rotherham United win their two games in hand.
Carlos Corberan's side could do with getting another win on the board to all but secure safety, but they have only won back-to-back matches on two occasions this season.
Town were beaten 2-1 by Barnsley in December's reverse match at Oakwell, meanwhile, and are aiming to avoid three losses in a row in this fixture for the first time since 1999.
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Huddersfield are light in terms of attacking options after losing Fraizer Campbell to injury against Forest at the weekend.
The one-time Manchester United striker joins Yaya Sanogo on the sidelines, although the latter does have a slight chance of playing a part on Wednesday.
Defender Michal Helik scored twice for Barnsley in their Boxing Day win over Huddersfield - the last Tykes player to score in both league games against the Terriers in a season was Mike Sheron in 1999-00.
Huddersfield Town possible starting lineup:
Schofield; Edmonds-Green, Keogh, Sarr, Toffolo; Bacuna, Hogg; Holmes, O'Brien, Rowe; Koroma
Barnsley possible starting lineup:
Collins; Andersen, Helik, Sibbick; Brittain, Palmer, Mowatt, Styles; Chaplin, Dike, Frieser
We say: Huddersfield Town 0-2 Barnsley
Barnsley will be eager to show that their loss to Coventry was just a blip, having won their previous six away matches before the trip to St Andrew's.
Like their opponents, Huddersfield are also in need of a victory, but they have not won back-to-back matches since early December and we cannot see that changing here.