| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 1 | Lincoln Park |
A | 4:28.62 | FPC Fresh/Soph Meet 2025 |
| 2 | Fort Pierce Westwood |
A | 4:24.25 | Cobra Relays 2025 |
| 3 | Martin County |
A | 4:21.23 | Cobra Relays 2025 |
| 4 | South Fork |
A | 4:12.81 | Cobra Relays 2025 |
| 5 | Treasure Coast | A | DNS | Park Vista #1 2025 |
| 6 | Fort Pierce Central |
A | 4:23.48 | Cobra Relays 2025 |
| 7 | Jupiter |
A | 4:27.25 | Jupiter Warrior Invitational 2025 |