Thus, sample size was mainly determined by the huge amount of data to process and the cost of UDPS. In conclusion, the results of this study provide further evidence of the utility of UDPS for investigating the evolution of the HBV QA. In addition, they provide confirmatory data for previous findings in studies with lower analytical coverage MK-1775 Wee1 inhibitor indicating greater QA variability in HBeAg-negative than HBeAg-positive patients. Our results show that high complexity in the preCore region is associated with low viral replication, in keeping with the key role of this region in HBV replication, and suggest an enhanced immune response in HBeAg-negative patients, probably related to the lack of HBeAg immunomodulatory activity. In the same direction, the positive selection of Core variants in HBeAgnegative and fluctuating status can be understood as a potential mechanism to escape the host immune system by nucleocapsid sequence changes. Finally, the strong negative correlation of QA evolution in the treatment-free period and under treatment shows the importance of studying the QA before treating patients, as a potential predictive factor of HBV evolution in cases of NUC nonresponse. With the consolidation of next-generation sequencing methods that enable the reproduction of viral haplotype study, QA complexity parameters could be useful for clinical management of HBV infection. Non-alcoholic fatty liver disease is defined as the accumulation of liver fat exceeding 5% of hepatocytes in the absence of significant alcohol intake, viral infection, or any other specific etiology of liver disease. NAFLD has an increasing prevalence worldwide and is now the leading cause of liver diseases in Western countries. The prevalence rate of NAFLD is reported to be 14–44% in the general population in Europe or the US and even 42.6–69.5% in people with type 2 diabetes. Patients with NAFLD, particularly those with non-alcoholic steatohepatitis, have a higher prevalence and incidence of clinically manifested cardiovascular disease as well and a 10-fold increased liver-related mortality owing to liver cirrhosis and hepatocellular carcinoma. In a Danish study, after adjustment for sex, diabetes and cirrhosis at baseline, NAFLD-associated age-adjusted standardized mortality ratios were 2.3 for all causes, 19.7 for hepatobiliary disease, and 2.1 for cardiovascular disease. Due to the lack of effective therapeutic measures and due to the epidemic of obesity and metabolic syndrome, NAFLD is projected to become the leading indication for liver transplantation in the next several years. The progression of NAFLD, from hepatic lipid overload, steatosis, to non-alcoholic steatohepatitis and to its complications liver fibrosis, cirrhosis or hepatocellular carcinoma, is causally linked to a massive inflammatory response in the liver. However, despite the fact that the extent of hepatic inflammation is the predominant factor determining disease progression in NAFLD, no specific anti-inflammatory interventional approaches have entered clinical practice yet.