The vine cactus Selenicereus megalanthus, known as “yellow pitaya”, produces attractive tasty fruits and has considerable economic potential as a new exotic fruit crop. This study explored the effects of different pollination treatments on fruit set and fruit quality characteristics in 14 clones of S. megalanthus. Hand-pollinated flowers set more viable seeds per fruit (213–215) and produced heavier fruits (213–225 g) with higher total soluble solids (TSS) contents (15.3–15.8 °Brix) than self-pollinated (bagged) or bee-pollinated flowers (70–100 seeds per fruit; fruit weight 86–87 g with 14.3–14.7% TSS content). Thus, the effectiveness of spontaneous self-pollination or bee pollination was low. Similar fruit set and seed set were obtained for both hand self-pollinated and hand cross-pollinated flowers, indicating full self-compatibility of all tested clones. Our results indicate that, to obtain high quality and high yield, commercial growers should hand-pollinate flowers, possibly using self-pollen. An orchard of one of the clones examined here, could thus be planted with a single clone, without requiring other clones for cross-pollination. These results are discussed in the context of our earlier (1994) study using only two clones of S. megalanthus.