Michio Kimura

Diagnostic Problems Caused by HBsAg Mutants – A Consensus Report of an Expert Meeting

A panel of 16 experts from 9 countries convened on April 14 at Schloss Reinhartshausen near Wiesbaden in Germany, to discuss the diagnostic significance of mutants, variants and genotypes of hepatitis B virus (HBV). Since the description of Australia antigen in 1965 and the subsequent observation that it was the envelope of the HBV and now designated hepatitis B surface antigen (HBsAg), this lipoprotein has been a mainstay in the diagnosis of HBV infections. HBsAg tests are used routinely in the diagnosis of acute and chronic liver disease, the screening of blood or organ donors and the surveillance of persons at risk to acquire or to transmit HBV. Current immunoassays for HBsAg are very specific and sensitive (both 199%) and are usually able to detect !0.5 ng HBsAg/ml serum. Their performance is validated in extensive trials before licensing and their detection limit is assayed with an International Standard for HBsAg. An immanent problem of virology is the variability of viruses. Due to the low fidelity of the viral nucleic acid polymerases and the high replication rates, virtually all nucleotide positions of a viral genome can be mutated within a relatively short time. However, the viability of the virus and its adaptation to the host allow the selection and outgrowth of only a limited number of mutants. The survival strategy of HBV in the population is mainly based on induction of immune tolerance and persistence of high viremia. In this state of infection the existing viral genome is favored whereas mutants are less fit and selected against or may be subject to an immune reaction and preferably eliminated. In fact, most of the HBsAg carriers in Germany have a very similar S gene sequence. However, under selective pressure the virus can express many different viable HBsAg mutants. Summary of the Presentations

Replication of the insect Chilo iridescent virus (CIV) in a poikilothermic vertebrate cell line.

Chilo iridescent virus (CIV) increased in titer in a vertebrate poikilothermic ‘C-type’-carrying cell line 4 days following inoculation. Cytoplasmic DNA synthesis, cytoplasmic array of virions typical of the iridovirus group, and CIV particles were observed budding from the plasma membrane.

Ferritin in insect vectors of the maize streak disease agent: electron microscopy and electron microprobe analysis.

Crystalline accumulations of electron-dense particles were observed in the gut of Cicadulina mbila leafhoppers, following feeding on leaves of maize streak diseased plants. Microcrystals were detected in the gut lumen and epithelial cells of infective insects, but not in controls fed on healthy plants. The crystals appeared to consist of numerous spherical electron-dense particles, approximately 9.5–10.5 nm in diameter, with cores of 6 nm. The orientation of electron-dense particles within the typical crystalline structure formed a zig-zag pattern with an angle of 120°. Although the morphological appearance of the particles was similar to the one of the presumptive maize streak virus, it was determined by electron microprobe analysis that the particles contained iron. Single particles were also observed in the vicinity of crystals. Based on the electron microprobe analysis data, as well as the ultrastructural characteristics and visualization in nonstained preparations, the crystals are believed to be comprised of ferritin. A hypothesis is presented that ferritin in leafhopper vectors of the maize streak disease agent might be derived from phytoferritin present in diseased plants and that it is temporarily stored in the gut cells of infective insects.

High sustained virologic response rate after interferon monotherapy in Japanese hepatitis C patients with a low HCV RNA titer and/or HCV genotype 2. A prospective study.

Objective: Hepatitis C virus (HCV) RNA titer and HCV genotype are considered to be major determinants of the outcome of interferon monotherapy. To clarify whether interferon monotherapy is really effective in patients with the appropriate viral parameters, we prospectively examined these parameters and treated the patients with interferon monotherapy. Methods: Sixty-four patients with an HCV RNA titer <100 kIU/ml and/or HCV genotype 2 were enrolled in the study. Eighteen patients with an HCV RNA titer >100 kIU/ml and genotype 1 were also enrolled as controls. All patients were treated with 10 megaunits of interferon-α2b every day for 2 weeks and then 3 times a week for 24 weeks. Results: Of the 64 patients with either HCV RNA <100 kIU/ml and/or genotype 2, seven dropped out from the study. Of the remaining 57 who completed the treatment, 48 (84%) showed a virologic sustained response. In contrast, only 4 of the 18 patients (22%) with HCV RNA >100 kIU/ml and genotype 1 were virologic sustained responders (p < 0.001). Conclusion: Our current study showed that the patients with HCV RNA <100 kIU/ml and/or HCV genotype 2 are good candidates for interferon monotherapy.

Efficacy of lamivudine or entecavir against virological rebound after achieving HBV DNA negativity in chronic hepatitis B patients.

Nucleos(t)ide analogues (NAs) lead to viral suppression and undetectable hepatitis B virus (HBV) DNA in some individuals infected with HBV, but the rate of virological rebound has been unknown in such patients. We examined the prevalence of virological rebound of HBV DNA among NA-treated patients with undetectable HBV DNA. We retrospectively analyzed 303 consecutive patients [158 entecavir (ETV)- and 145 lamivudine (LAM)-treated] who achieved HBV DNA negativity, defined as HBV DNA < 3.7 log IU/mL for at least 3 months. They were followed up and their features, including their rates of viral breakthrough, were determined. Viral rebound after HBV DNA negativity was not observed in the ETV-group. Viral rebound after HBV DNA negativity occurred in 38.7% of 62 HBe antigen-positive patients in the LAM-group. On multivariate analysis, age was an independent factor for viral breakthrough among these patients (P = 0.035). Viral rebound after HBV DNA negativity occurred in 29.1% of 79 HBe antigen-negative patients in the LAM-group. Differently from LAM, ETV could inhibit HBV replication once HBV DNA negativity was achieved. In contrast, LAM could not inhibit HBV replication even if HBV negativity was achieved in the early phase. Attention should be paid to these features in clinical practice.

Large regenerative nodule perfused by the portal vein

Abstract  In a 42‐year‐old Japanese woman with esophageal varices, abdominal ultrasound (US) demonstrated a hyperechoic lesion 3 cm in diameter in segment 4 (S4). This nodular lesion had high intensity on T1‐weighted magnetic resonance imaging (MRI), low intensity on T2, and very high intensity on superparamagnetic iron oxide (SPIO) enhanced MRI. Angiography showed sparse distribution of arterial branches and dense distribution of portal branches in S4. Meandering, thin arteries were seen in the peripheral area of the right lobe. The second branches of the portal vein were hardly visualized anywhere in the liver. Computed tomography arterioportography (CTAP) revealed portal blood flow dominance in this nodular lesion. There was no evidence of ischemic liver damage, such as thromboembolic episodes, laboratory data of liver damage, coagulation abnormalities etc. Therefore this abnormality was more likely to be caused by anomalous changes than thrombotic changes. Needle biopsy revealed no atypical cells. Two years later, the nodule size was reduced to 1.9 cm, showing its benign nature. Based on these findings, this lesion was classified as a new type of large regenerative nodule (LRN) associated with anomalies in the portal veins and arteries. This is the first report of an LRN of this size in which portal vein perfusion was dominant. Moreover, this lesion was difficult to differentiate from hepatocellular carcinoma (HCC) by imaging. Analysis of the images and pathological features of this case would contribute to a better understanding of the pathogenesis of nodular lesions of the liver.