Huanglongbing (HLB) disease detection in Cuba in 2006 led to a turn in citrus management in the country, including the production of certified budwood in protected nurseries, eradication of symptomatic plants and old orchards, and chemical control of the vector Diaphorina citri. This dramatic shift in citrus farming could have affected the incidence and composition of citrus tristeza virus (CTV) populations. This work aimed to determine CTV incidence and characterize viral isolates from the main Cuban citrus-producing areas. A survey throughout the country was carried out which resulted in the collection of 27 CTV isolates. Results showed a decrease of CTV incidence in western and central areas of the country, but an increase in the East, compared to previous reports. Bioindexing showed that most of the isolates induced none or mild symptoms. Serological characterization using seven different CTV specific monoclonal antibodies (MAb) showed heterogeneity among the 27 isolates. Six out of twenty-seven were MCA13-reactive, a MAb associated to the presence of severe CTV isolates. Nevertheless, sequence comparison of the p20, p23 and p25 genes evidenced a high nucleotide sequence identity among isolates, and according to the phylogenetic analyses, all the sequences belonged to the CTV T30 genotype. Thus, our results showed a decrease in the CTV incidence in the surveyed areas and the broad dissemination of mild CTV isolates of the T30 genotype, which could be partially explained by the shift in the management programs implemented for CTV and HLB control in Cuba.