‍ ObjectiveTo investigate the serum level of HBV RNA in untreated or treatment-experienced patients with chronic hepatitis B （CHB） and the correlation between serum HBV RNA level and the duration of antiviral therapy with nucleos（t）ide analogues （NAs）. MethodsA total of 300 patients with CHB who attended Department of Infectious Diseases in The First Affiliated Hospital of Shihezi University School of Medicine from February to July， 2022， were enrolled as subjects. Related clinical data were collected， and according to the duration of antiviral therapy， they were divided into untreated group with 73 patients， treatment duration ≤1 year group with 91 patients， and treatment duration >1 year group with 136 patients. Serum HBV RNA load， HBV DNA load， and HBsAg concentration were measured for all patients. The Mann-Whitney U test was used for comparison of continuous data between two groups， and the Kruskal-Wallis H test was used for comparison between multiple groups， further pairwise comparison using Bonferroni method； the chi-square test was used for comparison of categorical data； a Spearman correlation analysis was used to investigate the degree of correlation between various indicators. ResultsThe positive rate of HBeAg was 18.3%， and among the patients with negative HBV DNA， the patients with positive HBV RNA accounted for 44.1% （86/195）. There was a significant difference in the distribution of the serum levels of HBV RNA， HBV DNA， and HBsAg between the positive HBeAg group and the negative HBeAg group （Z=10.740， 6.300， and 7.280， all P<0.05）. There was a significant difference in the distribution of DNA level between the untreated group and the treatment duration ≤1 year group （P<0.05）； there was a significant difference in the distribution of HBV RNA and HBV DNA levels between the untreated group and the treatment duration >1 year group （P<0.05）； there was a significant difference in the distribution of HBV RNA， HBV DNA， and HBsAg levels between the treatment duration ≤1 year group and the treatment duration >1 year group （P<0.05）. The correlation analysis between the duration of antiviral therapy and the levels of HBV RNA， HBV DNA， and HBsAg showed that the duration of antiviral therapy had an extremely weak negative correlation with the levels of HBV RNA and HBsAg （r=-0.247 and -0.138， both P<0.05） and a strong negative correlation with the level of HBV DNA （r=-0.771， P<0.001）. There was a low degree of correlation between the serum level of HBV RNA and the serum levels of HBV DNA and HBsAg （r=0.360 and 0.442， both P<0.001）. Further stratified analysis showed that in the untreated group， there was a strong positive correlation between HBV RNA and HBV DNA （r=0.752， P<0.001） and a moderate positive correlation between HBV RNA and HBsAg （r=0.559， P<0.001）； in the treatment duration ≤1 year group， there was a low degree of positive correlation between HBV RNA and HBV DNA/HBsAg （r=0.396 and r=0.388， both P<0.001）； in the treatment duration >1 year group， there was a low degree of positive correlation between HBV RNA and HBsAg （r=0.352， P<0.001）. ConclusionSerum HBV RNA is negatively correlated with the duration of treatment with NAs， and the correlation of HBV RNA with HBV DNA and HBsAg gradually decreases with the increase in the duration of treatment. Therefore， it can be used as a supplementary indicator for monitoring the level of virologic response in CHB patients to a certain extent， with a relatively high accuracy in reflecting the level of viral replication in untreated patients.