Several duck Tembusu virus (DTMUV) clusters have been identified since its first emergence in 2010. However, the pathogenesis evaluation of DTMUV has been restricted to cluster 2.2 Chinese DTMUVs. In this study, the pathogenesis of a cluster 2.1 Thai DTMUV was investigated in three ages of Cherry Valley ducks (1-, 4- and 27-week-old). In each age, 35 ducks were inoculated with a cluster 2.1 Thai DTMUV and evaluated for clinical signs, virus distribution and shedding, pathology and serological response. Our results demonstrated that all duck ages were susceptible to Thai DTMUV; however, Thai DTMUV induced greater disease severity in younger ducks (1- and 4-week-old) when compared to older ducks (27-week-old) reflected by higher morbidity and mortality rates, and higher degree of pathological severity. Corresponding to these results, longer-term viremia, higher levels of viral loads in tissues and lower neutralizing antibody titers were also observed in younger ducks compared to those in older ducks. However, it should be noted that a significant drop in egg production was found in older ducks, which also indicates the susceptibility to Thai DTMUV in older ducks. Interestingly, prolonged shedding period with high viral loads was observed in older ducks even without showing clinical signs, suggesting the potential role of the older ducks as the carriers of Thai DTMUV. This finding highlights the importance of monitoring DTMUV and preventing the transmission of DTMUV in adult ducks. Overall, this study provides insights into the pathogenesis and infection dynamics of a cluster 2.1 Thai DTMUV in ducks.