Fruit flies attack numerous crops, including cherry tomatoes (Solanum lycopersicum var. cerasiforme). The potential presence of the immature stages of fruit fly species inside tomatoes during export hinders their international market access. Therefore, phytosanitary treatment must be performed before export to prevent fruit fly species from entering countries where they are not naturally found. We developed a phytosanitary cold disinfestation treatment protocol to eliminate oriental fruit fly (Bactrocera dorsalis Hendel), melon fly (Zeugodacus cucurbitae Coquillett), and pumpkin fruit fly (Zeugodacus tau Walker) concealed inside cherry tomatoes without causing critical damage to the fruit. We determined that the third instar of Z. cucurbitae exhibited the highest cold tolerance among the various development stages of the three fruit fly species. Thus, we performed a small-scale disinfestation test on Z. cucurbitae in two cultivars of tomato. We achieved complete disinfestation after 15 days of cold treatment at 1°C-1.5°C. The confirmatory test revealed the elimination of more than 80,000 treated third instar of Z. cucurbitae in each tomato variety. The developed phytosanitary cold treatment allows the tomatoes to retain their commercial value. This study provides a standard phytosanitary cold treatment protocol for cherry tomatoes, ensuring the disinfestation of fruit flies before their export to international markets.