Circulating immune complexes and mutations of HBsAg are associated with the undetectable HBsAg in anti-HBs and HBeAg positive occult hepatitis B virus infection

Abstract

IntroductionOccult hepatitis B virus infection (OBI) is an HBsAg negative state in HBV infection with usually inactive HBV replication. However, there were a minority of individuals with positive HBeAg and anti-HBs among OBI blood donors and few studies have focused on this unusual serological pattern.Methods2022 plasma of blood donors that preliminary screened reactive for HBV DNA and non-reactive for HBsAg were collected from 16 provinces in China from 2015 to 2018. HBV DNA and HBsAg in these samples were retested using the Cobas TaqScreen MPX test and ARCHITECT HBsAg Quantitative II assay. Lumipulse HBsAg-HQ assay and polyethylene glycol (PEG)-double precipitation following HCl and trypsin digestion were performed to detect HBsAg from HBsAg-anti-HBs circulating immune complexes (CICs).Results1487 of 2022 samples were positive for Cobas HBV DNA test and non-reactive for ARCHITECT HBsAg assay, while 404 of them were positive using Lumipulse HBsAg-HQ assay. 10 HBsAg-/anti-HBs+/HBeAg+ OBI blood donor samples were further dissociated and HBsAg-CICs were detected in 7 samples. Sequencing analysis showed that D44N, N98T, G73S, Del 56-116, and I161T occurred in the pre-S region, and immune escape mutations such as P127T, F134L, G145R, V168A, and I126T/S in the S region were found.DiscussionIn conclusion, there were a minority of HBsAg-/anti-HBs+/HBeAg+ individuals in OBI blood donors. The undetectable HBsAg in these individuals was mainly due to HBsAg-CICs. Immune escape-associated mutations also happened under the host’s selective pressure. HBsAg dissociation methods or Lumipulse HBsAg-HQ assay is recommended to distinguish these individuals.

Frequency of HBsAg variants in occult hepatitis B virus infected patients and detection by ARCHITECT HBsAg quantitative

“…The "a" determinant is the immunogenic region of HBsAg (Huang et al, 2017), and its second loop structure is the major antigenic determinant of HBsAg (Zhang et al, 2017). Mutations of a single sequence site or multiple adjacent ones in this region can cause changes in the conformational epitope of HBsAg protein, reducing the antigenicity of HBsAg, thereby affecting the binding reaction of HBsAg with the corresponding antibody (Yan et al, 2022). In addition, genetic mutations in the MHR will also affect the expression and secretion of viral HBsAg, making it lower than the detection limit of clinical HBsAg test reagents or causing the viral load in the circulation to be at a low level, thereby leading to missed detection of HBsAg (Qi et al, 2023).…”