Fernando Gonzalez defeated qualifier Martin Klizan 6-2, 6-4 in his return from an eight-month injury layoff because of hip surgery.
The big-serving Isner had 18 aces but Mello converted the only break point of the first set to take control of the match.
He then saved four of the five break points he faced in the second before prevailing in the tiebreaker against last year's finalist.
Gonzalez, 30, dominated the first set, although both players struggled to hold serve in the second. Klizan double-faulted on Gonzalez's second match point.
Gonzalez will play Feliciano Lopez, who upset eighth-seeded Juan Monaco 6-4, 7-5.
ESTORIL OPEN: In Oeiras, Portugal, former U.S. Open champion Juan Martin del Potro got by Pedro Sousa 6-2, 3-6, 6-3.
Del Potro, who was making his season debut on clay, broke his opponent's serve in the third and seventh games to take the first set in under half an hour.
BMW Open: In Munich, third-seeded Marin Cilic edged Simone Bolelli 7-5, 7-6 (4) to advance to the second round.
Cilic was the runner-up last year and used his 11 aces to help win the close match.
Fifth-seeded Florian Mayer eased past qualifier Steve Darcis 6-1, 6-2.
BARCELONA OPEN: In Barcelona, Spain, Sara Errani beat Maria-Teresa Torro-Flor 6-1, 6-0.
Alberta Brianti topped third-seeded Tsvetana Pironkova 6-1, 6-2, while Mirjana Lucic won 7-6 (4), 6-3 over No. 4 Ekaterina Makarova.