Persistence of infectious hepadnavirus in the offspring of woodchuck mothers recovered from viral hepatitis

Abstract

Mother-to-child transmission is an important route for hepatitis B virus (HBV) dissemination. It has been established that HBV traces persist for years after complete clinical recovery from hepatitis B. Similarly, resolution of hepatitis caused by HBV-related woodchuck hepatitis virus (WHV) is followed by occult lifelong carriage of pathogenic virus. In this study, we documented that WHV persisting after termination of acute hepatitis is transmittable to newborns as an asymptomatic long-term infection. All 11 offspring from 4 dams studied carried transcriptionally active WHV genomes for 3.5 years after birth without immunovirological markers of infection. WHV genomes and mRNA were detected both in the liver and lymphoid tissue in the majority of offspring; WHV covalently closed circular DNA was detected in some samples. In 4 offspring, however, the virus was restricted to the lymphatic system. In the circulation, WHV DNA-reactive particles were DNase resistant and of comparable size and density to complete virions. Importantly, the virus in offspring with or without hepatic WHV DNA expression was infectious to WHV-naive woodchucks. Finally, offspring challenged with WHV were not protected against reinfection. These findings show that mothers with occult hepadnaviral carriage transmit pathogenic virus to their offspring, inducing a persistent infection invariably within the lymphatic system but not always in the liver.

Persistence of isolated antibodies to woodchuck hepatitis virus core antigen is indicative of occult infection

“…30 For PCR, WHV DNA was amplified using three sets of direct or nested oligonucleotide primers specific for WHV core (C), envelope (surface, S) and X gene sequences, as reported. 30,31 For PCR detecting WHV cccDNA, mung bean nuclease treatment, primers and conditions previously established were applied. 32,33 The presence of WHV mRNA was examined using total RNA extracted with Trizol (Invitrogen Life Technologies, Burlington, Ontario, Canada).…”

“…Reverse transcription (RT) reaction was done as reported. 30,31 In all instances, DNA and RNA isolations were carried out in parallel with mock samples containing TE buffer (1 mmol EDTA in 10 mmol/L Tris-HCl buffer, pH 8.0) instead of nucleic acid. In addition, DNA or RNA from serum, PBMC, or liver tissue of a healthy animal and from a woodchuck with WHsAgpositive chronic WHV hepatitis were included as negative and positive controls, respectively.…”

“…In the case of RT-PCR, test RNA samples not treated with RT were included to rule out potential WHV DNA carryover. 30,31 The final PCR contamination controls consisted of water added to the direct or nested PCR cocktails instead of template DNA, cDNA or direct PCR product. Nucleic acid hybridization (NAH), i.e., Southern blot analysis, of the PCR products to a rWHV DNA probe was routinely used to verify specificity of detection and validity of con-trols.…”

“…The sensitivity of the nested PCR/NAH was Ͻ 10 vge/mL and that for WHV cccDNA Ϸ10 2 vge/mL, as reported. [31][32][33] Electron Microscopy. Four mL of autopsy sera from 6/F and 10/M (Table 1) were centrifuged at 200,000g for 18 hours at 5°C in an SW50.1 Beckman rotor and the resulting pellets examined by electron microscopy after negative staining with 1% phosphotungstic acid.…”

“…30 In contrast, offspring born to woodchuck mothers convalescent from WHV hepatitis carry comparable levels of infectious virus in the absence of anti-WHc. 31 Similarly, animals inoculated with small amounts of WHV (Ͻ10 3 vge/mL) develop an anti-WHc-negative infection with low-level viremia. 32,33 In this study, the significance of the sole detection of anticore antibodies as an indicator of a low-rate hepadnavirus replication was explored.…”