The Lady Eagles battled back from a set down twice, but couldn't come up with a win on the road over an 11th Region rival.
Lexington Catholic (16-12) used a strong final set to secure a 3-2 win over the Lady Eagles (22-4) on Tuesday in Lexington.
The Lady Knights won by scores of 25-16, 22-25, 25-21, 20-25 and 15-7.
The loss snapped Southern's 13-match winning streak.
The Lady Eagles had not lost since Sept. 12.
Sarah Mitchell had 17 kills and 21 digs for Southern. Julia Bingham had 10 kills and two digs.
Avery St. John had 10 kills and three digs. Alyssa Evans had 10 kills and 17 digs. Hannah Martin had eight kills, three digs and three blocks.
Kyla Atkins had 32 assists, 12 digs, three kills and two aces.
Kennedy Easterling had 24 assists and Autumn Griffith had 11 digs and two aces.
Southern is set to return to action Oct. 15 at home against Bourbon County.