| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 2 | DeSoto County |
A | Trejo-Balderas | Rios | Johnson | Martinez | 5:41.51 | Lake Placid Invitational 2026 |
| 3 | LaBelle |
A | Chairez | Deas | Johnson | Jackson | 4:54.20 | Lake Placid Invitational 2026 |
| 4 | Sebring |
A | Madison | Brito | Holding | Melendez | 4:27.61 | Avon Park Red Devil Invitational |
| 5 | Hardee |
A | Manriquez | Herrera | Villa | Nunez | 4:52.00 | |
| 6 | Avon Park |
A | Tate | Glabach | Virkler | Chamberlain | 5:00.56 | 2026 Titan Invitational |