Ntcp is the key host factor limiting HBV infection in cynomolgus and rhesus macaques and in pigs. In rodents (mouse, rat) and dogs, transfer of hNTCP supports viral entry but additional host factors are required for the establishment of HBV infection. This finding paves the way for the development of macaques and pigs as immunocompetent animal models to study HBV infection in vivo, immunological responses against the virus and viral pathogenesis. (Hepatology 2017;66:703-716).

Objective To investigate the positive impact of e-aid cognitive behavioural therapy on the sleep quality, anxiety, and depression of nurses on site during the COVID-19 pandemic. Methods Nurses on site at the Tianjin Medical University General Hospital Airport Site experiencing insomnia, anxiety and depression during the COVID-19 prevention and control period, from February 2020 to April 2021, were selected and divided into either an e-aid cognitive behavioural therapy (eCBT-I) group or a control group using a randomized grouping method. The eCBT-I group was given standard eCBT-I for 6 weeks; the control group did not get any intervention. The Pittsburgh Sleep Quality Index (PSQI) and the Insomnia Severity Index (ISI) were used to evaluate the sleep quality of the subjects. The Generalized Anxiety Disorder 7-item (GAD-7) and the Patient Health Questionnaire (PHQ-9) were used to assess the subjects’ anxiety and depression. Changes in sleep quality, anxiety and depression before and after treatment were compared between the two groups. Results Of 118 nurses randomized, the PSQI and ISI scores within the eCBT-I group ( n =60) were significantly lower after treatment (5.9 ± 3.9, 6.7 ± 4.5) than before treatment (10.4 ± 3.5, 12.4 ± 4.7) ( p <0.05). Compared to the scores of the control group ( n =58) (9.1 ± 3.9, 10.6 ± 4.1), the PSQI and ISI scores in the eCBT-I group (5.9 ± 3.9, 6.7 ± 4.5) were lower after treatment ( p <0.05). The GAD-7 and PHQ-9 scores in the eCBT-I group were all lower after treatment (3.7±3.4, 4.2±4.1) than before treatment (6.7±4.9, 7.7±5.1) ( p <0.05). Compared with subjects in the control group (7.1±5.6, 7.3±5.1), subjects in the eCBT-I group (3.7±3.4, 4.2±4.1) had lower scores on the GAD-7 and PHQ-9 scales after treatment ( p <0.05). Conclusion eCBT-I improved the sleep quality of frontline nurses during the COVID-19 prevention and control period and relieved anxiety and depression.

Persistence of the human hepatitis B virus (HBV) requires the maintenance of covalently closed circular (ccc)DNA, the episomal genome reservoir in nuclei of infected hepatocytes. cccDNA elimination is a major aim in future curative therapies currently under development. In cell culture based in vitro studies, both hybridization-and amplification-based assays are currently used for cccDNA quantification. Southern blot, the current gold standard, is time-consuming and not practical for a large number of samples. PCR-based methods show limited specificity when excessive HBV replicative intermediates are present. We have recently developed a real-time quantitative PCR protocol, in which total cellular DNA plus all forms of viral DNA are extracted by silica column. Subsequent incubation with T5 exonuclease efficiently removes cellular DNA and all non-cccDNA forms of viral DNA while cccDNA remains intact and can reliably be quantified by PCR. This method has been used for measuring kinetics of cccDNA accumulation in several in vitro infection models and the effect of antivirals on cccDNA. It allowed detection of cccDNA in non-human cells (primary macaque and swine hepatocytes, etc.) reconstituted with the HBV receptor, human sodium taurocholate cotransporting polypeptide (NTCP).Here we present a detailed protocol of this method, including a work flowchart, schematic diagram and illustrations on how to calculate "cccDNA copies per (infected) cell".