Llantwit Major were beaten 2-0 at home to Cymru South newcomers Cardiff Draconians on Saturday.
Karl Lewis' men slipped to their first defeat in four league games when they took on the Gabalfa club, who have remained unbeaten throughout September.
Major fell behind after 16 minutes when Sam Cawley volleyed home over keeper Ryley Allen, before Dan Bowen doubled the Draconians' lead just four minutes later from Harry Treharne's cross.
Both Josh Jones and Kofi Rowe came close to getting one back, but the hosts seldom posed much threat as the Draconians held out to take all three points from Windmill Lane.
Llantwit drop to 11th in the table as attention to next Saturday's visit to Baglan Dragons - the second clash between the two teams in just three weeks, following Llantwit's Welsh Cup win last weekend.
Cambrian United retain a four-point lead at the top of the table following their 1-0 win at Baglan Dragons, while Treflin remain in second after a 5-0 demolition of hosts Ynyshir Albions.
Third-placed Caerau Ely also won 5-0 on the road at Treowen Stars, with two goals from Oliver Redhead and one apiece for former Llantwit striker Matty Kimmins, Zack Maher and sub Liam Bishop.
Newport City's lenghty unbeaten run was finally ended with a 1-0 defeat at Carmarthen while Aberystwyth Town won 1-0 at Afan Lido and bottom club Cwmbran Celtic drew 2-2 with Trethomas Bluebirds.
JD Cymru South
Ammanford 1-1 Pontypridd United (Friday)
Baglan Dragons 0-1 Cambrian United
Carmarthen Town 1-0 Newport City
Cwmban Celtic 2-2 Trethomas Bluebirds (Friday)
Afan Lido 0-1 Aberystwyth Town
Llantwit Major 0-2 Cardiff Draconians
Treowen Stars 0-5 Caerau Ely
Ynyshir Albions 0-5 Trefelin BGC
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