This investigation aims to study the reproductive traits, including sex ratio, spawning season and pattern, batch fecundity (F), and length at first maturity (Lm), of the commercial catfish Mystus mysticetus living in the freshwater of the Mekong and Chao Phraya basins. A collection of 596 individuals was caught using trawl nets from Cai Rang, Can Tho (CT) and Long Phu, Soc Trang (ST) from October 2020 to September 2021. The results showed a female-biased sex ratio (0.64 males: 1.00 females) and a pattern of multispawner as ripe ovaries contained different developmental stages of oocytes. A combined analysis of gonadal occurrence frequency and gonadosomatic index showed that this fish spawned during the wet season with a peak in June-July at CT and September-October at ST. The F ranged from 4,470–22,457 eggs/female (CT) to 4,104–23,651 eggs/female (ST) and these values increased with fish size. The egg diameter ranged from 0.63 ± 0.06 to 0.64 ± 0.06 SD mm, and Lm was higher in males (11.6 at CT and 11.2 at ST) than in females (10.9 at CT and 10.4 at ST). To conclude, it is recommended that the fish should be caught at length > Lm and avoid fishing in the main spawning period in order to exploit this fish resource sustainably.