Fourth seed John Isner saw his Rio Open campaign end in the first round after he was beaten 6-7 (5), 7-5, 6-7 (8) by Guido Pella.
It was the second consecutive disappointing performance for the world number 11 who lost to Dusan Lajovic in the second round of the Argentina Open last week.
Neither Pella nor Isner could engineer a break in the first set, and a tie-break was needed to separate the pair, with Pella edging the breaker 7-5.
The second set played out in a similar fashion, with both players holding serve until Isner took a 6-5 lead by breaking Pella in the 11th game before serving out the set to bring himself back into contention.
The American was tested once again in the third set as he was forced to save two match points in the 10th game, but he hung on to force a tie-breaker. However, it was Pella who emerged victorious and will now either face Santiago Giraldo or Gastao Elias in the second round.