Background: Brucellosis is an endemic disease in Latin America and other countries. Serologic cross-reaction between cholera and Brucella infection is well recognized. Since the introduction to cholera in 1991 in Latin America, interpreting serologic tests has become potentially problematic. This study attempts to evaluate this problem. Methods: Tube agglutination tests were performed to detect Brucella antibodies in 44 Peruvian adult patients with moderate to severe diarrhea due to Vibrio cholerae O1 El Tor infection. These patients had no prior history and no clinical evidence of brucellosis. Results: False positive reactions were observed in 43.2% and 15.9% of the patients when cut-off points of ≥ 1/80 and ≥ 1/160 titer, respectively, were selected. These false positive reactions occurred within 4 to 14 days after the onset of diarrhea. The cross-reactivity decreased at the end of the fifth week (only 8.33% had a positive value at the fifth week). Conclusions: Physicians should be alert to the false positive reaction to Brucella in patients with diarrhea. This is relevant to the evaluation of febrile illness in patients coming from developing countries where they could have been exposed to cholera.