Tatsunori Nakano

A case of subclinical hepatitis E virus (HEV) infection was detected by nucleic acid amplification test on blood donation. The patient was followed-up until day 220 after the blood donation but showed no symptoms throughout the observation period. Aspartate aminotransferase and alanine aminotransferase levels reached the maximum values on day 37 with a slight increase but remained in normal ranges from day 67 to 220. The quantity of HEV RNA at the initial examination on day 13 was 1.1 × 102 copies/mL, which increased to 2.8 × 103 copies/mL by day 37. It was not detected from day 67 to 220. Immunoglobulin G class antibody to HEV (anti-HEV IgG) was below the cut-off value until day 37 and exceeded the cut-off value to positive on day 67, accompanied by normalization of liver function and negative conversion of HEV RNA. Thereafter, the titer decreased gradually, falling below the cut-off value on day 163, and continuing negative until day 220. Although the persistent duration of anti-HEV IgG positive is believed to be generally long, it was within only 126 days for this subclinical case. Further investigation is needed to determine whether short-term positivity for anti-HEV IgG is typical in subclinical HEV infection.

岐阜県近郊で発生したE型肝炎13例のE型肝炎ウイルス(HEV)の感染源・感染経路を、問診と原因HEVの遺伝子配列の解析から推測した。全員孤発例であった。2例は生のブタ内臓肉、1例は調理したブタ内臓肉、7例は調理したブタ肉、2例は調理不十分なブタ肉の喫食歴があった。HEVゲノムのORF2領域の412塩基長の配列をもとに近隣結合法で系統樹を作成した。1例はORF1領域を解析した。原因HEVの遺伝子型は、3aが5例、3bが6例、3fが1例、4iが1例であった。13例中2例は原因HEV株に近縁株の登録はなかったが、11例には97%以上の相同性を持つ株が見つかり、地域内拡散8例、国内広域拡散2例、国際拡散1例の3つのパターンに類別された。

AIM: To evaluate the clinical and molecular characteristics of hepatitis E virus (HEV) infection in Mie Prefecture, Japan, from 2004 through 2018. METHODS: The clinical information of hepatitis E cases was collected from 21 medical institutions in Mie Prefecture. The nucleotide sequences of infecting HEV strains were determined for cases with available serum samples. The origins or transmission routes were inferred from phylogenetic analyses of the nucleotide sequences. RESULTS: Fifty-three patients were diagnosed with HEV infection. The number of cases increased each year through 2012 and then decreased. Analyses of the clinical characteristics of the cases indicated that even mild cases were detected in the latter 10 years of the study. Nucleotide sequence analyses were undertaken on 38 of the 53 cases. The HEV subtype 3e (HEV-3e) strains identified for 13 cases were closely related to a swine HEV-3e strain that was isolated from the liver of a pig bred in Mie Prefecture. The number of cases infected with the indigenous Mie HEV-3e strains increased until 2012 but have not been reported since 2014. In the latter half of the study, cases involving various HEV strains of different genotypes and subtypes emerged. CONCLUSIONS: The disappearance of indigenous Mie HEV-3e strains appeared to be the primary cause for the decrease in hepatitis E cases in Mie Prefecture. The disappearance might have been associated with improved hygienic conditions on pig farms or the closure of contaminated farms. The results suggest that indigenous HEV strains can be eradicated by appropriate management.

著者らは三重県と岐阜県を中心にE型肝炎の発生状況を調査し、患者の喫食歴などを問診するとともに、原因となったHEVの遺伝子配列を決定し、症例相互の原因HEV株の関係や、データベースから探究した近縁株との関係を調べることで、感染源・感染経路の解明に努めている。今回、問診では互いに接点がないと思われたが、発症時期と遺伝子解析結果から感染源が同一と推測された3組6症例が存在したので、各症例の概要を報告した。6例とも同一経済圏内の発生であり、感染にはブタ肉などの食品の流通や人の往来が関与していると考えられた。

症例は73歳男性で、進行胃癌+肝転移に対する化学療法にて薬剤性間質性肺炎を発症し、プレドニゾロンを12日間投与され軽快傾向で退院した。定期受診にて急性肝障害が指摘され緊急入院となり、グリチルリチン製剤投与にて経過観察中IgAクラスE型肝炎ウイルス陽性が判明し急性E型肝炎と診断した。血小板減少を伴ったが、入院13日目には肝障害が改善したため退院し、血小板数も正常に復した。退院後52日目に再入院して胃癌に対しニボルマブを投与したところ血小板が0.2×10^4/μlまで減少し、血小板輸血とデキサメサゾン投与を行ったが、再入院15日目に上部消化管出血のため死亡した。骨髄穿刺検査の結果と経過から、ニボルマブ投与による免疫性の血小板減少症が考えられた。

Hepatitis E virus subtype 3f (HEV-3f) strains are usually isolated in Europe and Thailand. Recently, HEV-3f strains were detected from six acute hepatitis E patients in Japan, none of whom had a history of travel to endemic areas. We inferred the origin and transmission route of the six HEV-3f strains. A time-scaled phylogenetic tree of the six strains with reference strains was constructed using a Bayesian statistical inference framework. The time-scaled tree indicated that the six strains independently derived from similar European strains between 2008 and 2014. The pattern suggested recent inflow of multiple HEV-3f strains from Europe to Japan. Japan imports a substantial amount of pork from European countries every year. The emergence of acute hepatitis cases caused by HEV-3f strains in Japan, in patients with no history of travel abroad, might be influenced by the increased opportunities to consume pork products imported from European countries.

Hepatitis E virus (HEV) causes acute or chronic hepatitis in humans worldwide and can be transmitted via the fecal-oral route. Four HEV strains (HE-JA14-2173, HE-JA15-1335, HE-JA15-1920 and HE-JA16-0610) obtained from patients with imported (from Pakistan or India) or autochthonous acute hepatitis E in Japan were most closely related to the Nepalese and Mongolian genotype 1 HEV strains of unassigned subtype within the partial ORF2 sequence. To investigate whether a putative novel subtype (1g) of genotype 1 can be assigned, full-length genomic sequences were determined for the four HEV strains. They shared 95.4-99.2% nucleotide identity over the entire genome, and differed by 6.3-11.7% from the reported HEV strains of subtypes 1a-1f and by only 0.6-4.7% from a Mongolian genotype 1 HEV strain (MNE15-072) of unassigned subtype. A phylogenetic analysis showed that the four HEV strains obtained in the present study formed a cluster with MNE15-072, with a bootstrap value of 100%. Although the p-distance between subtypes 1a and 1f was 0.048-0.083, these five strains showed a higher nucleotide p-distance value of 0.068-0.138 with the genotype 1 HEV strains of subtypes 1a-1f. A BLAST search revealed the presence of candidate members of subtype 1g HEV in at least five other countries, including France, Israel, the Netherlands, Portugal, and the UK, sharing identities of 95.4-99.6% with the HE-JA16-0610 strain within the common sequence of 294-867 nucleotides. These results support the assignment of a new subtype 1g within genotype 1 and suggest a global distribution of subtype 1g strains. Subtype 1g strains found in Europe can be imported from Asia. Further studies are needed to confirm the global distribution of HEV subtype 1g.

バイタル変化や消化器症状、胃瘻部位のトラブルなどにより経腸栄養の中止が必要となることがある。今回、我々は、Stevens-Johnson症候群と考えられる症例において、反復する高熱や頻脈、振戦、および、遷延する低血圧の出現により、経腸栄養の再開時期を躊躇した。これらの症状は、Epstein-Barr virus(EBV)の再燃が関与したと考えられた。EBVのような日和見病原体によって発症する症状により、胃瘻からの経腸栄養の継続が困難となることがあり注意が必要である。(著者抄録)

In 2015, we saw a 67-year-old Japanese man with autochthonous acute hepatitis E living in the northern district of Mie prefecture, Japan. He had no history of travel abroad and blood transfusion. A rare HEV strain of subgenotype 3f was recovered from his serum. He was a job hunter engaged in wild boar hunting and processed captured boars with his naked hands. Although subgenotype 3b HEV strain was recovered from stored meat of captured wild boars, it seemed likely that he was infected with 3f HEV via exposure to meat and viscera of HEV-infected wild boars. Up to the present, at least four cases of autochthonous acute hepatitis E infected with 3f HEV have been identified in four prefectures in Japan. Further studies are warranted to clarify whether 3f HEV strains have been indigenized and domestic pigs and wild boars are infected with 3f HEV strains in Japan.

We experienced a case of acute hepatitis E who developed persistent jaundice for as long as six months. The phylogenetic tree analysis indicated that the hepatitis E virus (HEV) isolate obtained from the patient lived in Gifu was included in the HEV subtype 3f (HEV- 3f) cluster and the isolate clustered with two HEV-3f isolates previously reported in Japan. Interestingly, our patient and the two others had no history of travel abroad, e.g. to Europa, where HEV-3f strains are endemic. The source and the route of the HEV-3f infection are unknown and the specific symptoms of HEV- 3f infection are not well-known in Japan, either. We should reveal the characteristics of HEV-3f infection in Japan by accumulating cases.

In 2015, we have seen two cases of sporadic acute hepatitis E in northern and central district of Mie prefecture, Japan. Case No.1 was a 28-year-old Nepalese man and diagnosed with imported hepatitis E, while case No. 2 was a 30-year-old Japanese man, who had no history of travel abroad within one year before the disease onset and genotype 1a strain was recovered from his serum regardless of staying in Japan at the incubation period. The HEV strains recovered from these two patients were 93.2% identical to each other in the 412-nucleotide sequence within ORF2 and did not belong to the same cluster. Case No. 1 was typical imported infectious hepatitis E disease, however, case No. 2 did not have any evidence of imported infectious disease. This result suggests that genotyping of hepatitis E virus strain is necessary for a more accurate analysis even if a patient has no history of going or living abroad.

Background & Aims: Hepatitis E virus (HEV) genotype 4 has mainly been isolated from sporadic hepatitis cases and swine in Asian countries. We analysed the origin and global dispersal history of genotype 4 using a Bayesian phylogeographical approach. Methods: The 412-nucleotide sequences of open reading frame 2 of genotype 4 (47 Japanese, 40 Chinese, 1 Indian, 8 Indonesian, 1 Korean, 1 Taiwanese, 2 Danish and 2 Italian), of which sampling date and location were known, were collected. Evolutionary rate, divergence time, demographic growth and phylogeography were co-estimated in the Bayesian statistical inference framework implemented in the BEAST package to model spatial dispersal on a time-scaled genealogy. Results: The most probable origin of genotype 4 was Japan and the time of origin was 1909 (95% highest posterior density, 1871-1940). Seven lineages of genotype 4 migrated from Japan to China. The analysis also showed the migration of genotype 4 from Japan or China to India and Indonesia and from China to Indonesia, Taiwan, Korea and a few European countries. Conclusions: Swine trade between countries coincided with the migration time and direction of genotype 4 in some cases and was considered the primary cause of dispersal. However, there was no clear cause of dispersal for some cases, for which no records of pig trade were found. Future research should analyse additional nucleotide sequences paired with epidemiological data from various countries to improve our understanding of HEV dispersal.

In Gifu city, Japan, an acute hepatitis E case occurred in 2012 (previously reported in Kanzo 55: 713-716, 2014) and another hepatitis E case occurred in 2014. Both cases developed hepatitis E after eating raw pig liver and heart at the same restaurant. The patients had no other clear risk factors for hepatitis E virus (HEV) infection. The two HEV strains were totally different, suggesting that the pig farms which provided the pig products were contaminated by different HEV strains. Japanese government issued an order forbidding restaurants to serve raw pig offal in June 2015. However, the sources and the routes of HEV infection remain unknown in most hepatitis E cases in Japan. We must still be cautious for HEV infection.

An HCV outbreak occurred in 2012 in China, affecting hundreds of patients. We characterized HCV subtype 2a and 6a sequences from 60 and 102 patients, respectively, and co-analyzed them with 82 local controls and 103 calibrating references. The close grouping of the patients' sequences contrasted sharply with the diversity of local controls. Scaled by the calibrating references, the emergence of patients' isolates was estimated at 2-5 years before sampling. In contrast, the controls intermingled with the calibrating references that were much older. For both subtypes, the major and minor clusters could be defined, with the closeness to indicate linked transmission. Conclusion: HCV sequences from the study patients grouped into three subtype 2a and two subtype 6a clusters, in addition to three 6a solitary branches, representing descendants of eight earlier strains that were distinct and otherwise sporadic. Due to iatrogenic transmission through reusing needles, five strains were highly selected and preferentially spread. (C) 2015 Published by Elsevier Inc.

宮崎県内で同時期に2例のE型急性肝炎症例を経験した。2例に交友関係、共通の生活歴はなかった。両症例のE型肝炎ウイルス(HEV)株は、遺伝子配列の解析から近縁ではあるが同一株ではなく、感染源は異なると考えられた。2例のHEV株はともに12年前に同県内の飼育ブタから検出された遺伝子型3型株の子孫と考えられ、このウイルスが宮崎県内で存続していることが証明された。1例は急性末梢性顔面神経麻痺を合併した。(著者抄録)

We experienced six cases of hepatitis E within 18 months around Gifu city. The phylogenetic analysis indicated that the 6 hepatitis E virus (HEV) strains were included in different lineages, respectively. One case showed marked symptoms of hepatitis and was infected with a genotype 4 Aichi/Shizuoka strain which is ordinary isolated from wild animal. The case had history of consuming raw pig liver. The fact may indicate that Aichi/Shizuoka strains are confined not only to wild animal but also to pig bred around Gifu city. Other 5 cases had no clear risk factor of acquiring HEV and did not show obvious symptoms of hepatitis. HEV infection may occur more commonly than we think by various HEV lineages from unknown reservoirs.

Aim: To characterize hepatitis E in Mie prefecture and to investigate whether raw pig liver sold as food in Mie is contaminated with hepatitis E virus (HEV) strains similar to those recovered from patients. Methods: Seventeen patients with sporadic acute hepatitis E treated from 2004 to 2012 were studied. A total of 243 packages of raw pig liver from regional grocery stores were tested for the presence of HEV RNA. The partial genomic sequences of human and swine HEV isolates were determined and subjected to the phylogenetic analyses. Results: The HEV isolates recovered from the 17 patients segregated into genotype 3 (n = 15) and genotype 4 (n = 2), and 15 genotype 3 isolates further segregated into 3e (n = 11) and 3b (n = 4). Pig liver specimens from 12 (4.9%) of the 243 packages had detectable HEV RNA. All 12 swine HEV isolates were grouped into genotype 3 (3a or 3b). Although no 3e strains were isolated from pig liver specimens, two 3b swine strains were 99.5-100% identical to two HEV strains recovered from hepatitis patients, within 412-nt partial sequences. Conclusion: The 3e HEV was prevalent among hepatitis E patients. HEV RNA was detected in approximately 5% of pig liver sold as food. The presence of identical HEV strains between hepatitis patients and pig liver indicated that pigs play an important role as reservoirs for HEV in humans in Mie. Further studies are needed to clarify the source of 3e HEV in the animal and environmental reservoirs.

The source of European-type subgenotype 3e/3sp hepatitis E virus (HEV) strain that is prevalent in hepatitis E patients in Mie, Japan, remained unknown. The present analyses of stored RNA samples that had been extracted from liver tissues obtained from 85 slaughtered pigs in Mie in 2007, revealed the presence of two HEV strains (swJ-Mie07-37L and swJ-Mie07-80L). The swJ-Mie07-80L strain was classified into subgenotype 3b/3jp, while swJ-Mie07-37L belonged to 3e/3sp and shared high nucleotide sequence identities of 98.399.2% with the 3e/3sp strains recovered from hepatitis E patients in Mie during 2004-2012. These results suggest that pigs are important reservoirs for HEV infection even in Mie, although the hepatitis E patients did not recall consuming raw pig liver/intestine before the disease onset.

We experienced a case of hepatitis E in Mie prefecture infected with genotype 4 hepatitis E virus (HEV) strain endemic in Aichi and Shizuoka prefectures (Aichi/Shizuoka strain). The HEV isolate obtained from the patient clustered with other Aichi/Shizuoka strains with 100% of bootstrap value in the phylogenetic tree, and was more than 97.8% identical to other Aichi/Shizuoka strains, in the 412-nucleotide sequence within the ORF2 region. Interestingly, the patient had no history of consuming wild animal meat although all other reported Aichi/Shizuoka strains were recovered from meat from wild animals or humans after eating wild animal meat. The finding indicated that the origin of Aichi/Shizuoka strains is confined not only to wild animal meat but also to other unknown reservoirs. © 2014 The Japan Society of Hepatology.

We investigated the migration patterns of hepatitis C virus (HCV) in China. Partial E1 and/or NS5B sequences from 411 volunteer blood donors sampled in 17 provinces and municipalities located in five large regions, the north-northeast, northwest, southwest, central south, and southeast, were characterized. The sequences were classified into eight subtypes ( 1a, n = 3; 1b, n = 183; 2a, n = 83; 3a, n = 30; 3b, n = 44; 6a, n = 55; 6n, n = 10; 6v, n = 1) and a new subtype candidate. Bayesian evolutionary analysis by sampling trees of the E1 sequences of the five major subtypes revealed distinct migration patterns. Subtype 1b showed four groups: one is prevalent nationwide with possible origins in the north-northeast; two are locally epidemic in the central south and northwest, respectively, and have spread sporadically to other regions; and the fourth one is likely linked to the long-distance dispersion among intravenous drug users from the northwest. Subtype 2a showed two groups: the larger one was mainly restricted to the northwest and seemed to show a trend toward migration via the Silk Road; the smaller one was geographically mixed and may represent descendants of those that spread widely during the contaminated plasma campaign in the 1990s. Subtype 3a exhibited three well-separated geographic groups that may be epidemically unrelated: one showed origins in the northwest, one showed origins in the southwest, and the other showed origins in the central south. In contrast, subtype 3b had a mixture of geographic origins, suggesting migrations from the southwest to the northwest and sporadically to other regions. Structurally resembling the tree for subtype 3a, the tree for subtype 6a showed four groups that may indicate migrations from the central south to southeast, southwest, and northwest. Strikingly, no subtype 6a strain was identified in the north-northeast.

We compared the clinical features and laboratory data of 13 cases with acute hepatitis E and 61 cases with drug-induced liver injury (DILI) in our hospital. Out of 13 hepatitis E cases, 12 had no history of eating or undercooked raw meat/viscera and remained unknown for the origin and route of hepatitis E virus (HEV) infection. More than half of acute hepatitis E and DILI did not exhibit jaundice during the clinical course. All 13 cases of acute hepatitis E had a high score of ≥5 points by the DDW-J 2004 scale. Without HEV screening, eight cases of acute hepatitis E might have been misdiagnosed as DILI. The DDW-J 2004 scoring scale, current diagnostic scale for DILI, cannot discriminate acute hepatitis E cases from DILI cases because the scoring scale has no support for HEV infection. We propose that HEV-IgA test should be included in the current diagnostic scale for drug-induced liver injury for differential diagnosis of acute HEV infection to provide more accurate diagnosis of DILI in Japan. © 2014 The Japan Society of Hepatology.

A 67-year-old male living in Tsu city, Mie prefecture, Japan was referred to our hospital for further examination of acute liver injury and was diagnosed as having clinical hepatitis E virus (HEV) infection in January 2010. The HEV strain (HE-JA11-1701) isolated from the patient belonged to genotype 3 and European-type subgenotype 3e. It was presumed that the patient had been infected from a wild boar (Sus scrofa leucomystax) because he consumed meat/viscera from a wild boar that he had captured himself as a hunter approximately 2 months before disease onset. A specimen of the boar meat/viscera that the patient had ingested was not available. However, the HE-JA11-1701 strain was 99.8% identical within the 412-nucleotide sequence of the open reading frame 2 region to a HEV strain (JBOAR012-Mie08) that had been recovered from a wild boar captured near the patient's hunting area in 2008. A phylogenetic analysis confirmed that the two HEV strains had a close genetic relationship and were segregated into subgenotype 3e, supported by a high bootstrap value of 99%. Of note, the HE-JA11-1701 and JBOAR012-Mie08 strains were remotely related to the 3e strains reported in Japan and European countries, with a nucleotide difference of 7.9-13.9%, reinforcing the uniqueness of the 3e strains obtained in the present study. These results strongly support our speculation that the patient developed acute hepatitis E via consumption of HEV-infected boar meat/viscera. Genetic analyses of HEV strains are useful for tracing infectious sources in sporadic cases of acute hepatitis E.

In this study, we characterized the full-length genome sequences of seven hepatitis C virus (HCV) isolates belonging to genotype 1. These represent the first complete genomes for HCV subtypes 1e, 1h, 1l, plus one novel variant that qualifies for a new but unassigned subtype. The genomes were characterized using 19-22 overlapping fragments. Each was 9400-9439 nt long and contained a single ORF encoding 3019-3020 amino acids. All viruses were isolated in the sera of seven patients. residing in, or originating from, Cameroon. Predicted amino acid sequences were inspected and unique patterns of variation were noted. Phylogenetic analysis using full-length sequences provided evidence for nine genotype 1 subtypes, four of which are described for the first time here. Subsequent phylogenetic analysis of 141 partial NS5B sequences further differentiated 13 subtypes (1a-1m) and six additional unclassified lineages within genotype 1. As a result of this study, there are now seven HCV genotype 1 subtypes (1a-1c, 1e, 1g, 1h, 1l) and two unclassified genotype 1 lineages with full-length genomes characterized. Further analysis of 228 genotype 1 sequences from the HCV database with known countries is consistent with an African origin for genotype 1, and with the hypothesis of subsequent dissemination of some subtypes to Asia, Europe and the Americas.

Nucleotide sequences of hepatitis E virus (HEV) isolates infecting wild boars in Mie prefecture, which is located in the central region of Japan and is far from the most prevalent regions of HEV infection in Japan, were determined and characterised. Among 144 serum samples of wild boars captured in Mie prefecture, 7 were positive for HEV-RNA. The nucleotide sequence of nearly the entire genome was determined for 4 of the 7 positive samples. Phylogenetic tree analyses indicated that 6 samples were subtype 3e and 1 was subtype 3a among the 7 isolates. We identified the indigenization of subtype 3e isolates in Japanese wild boars. Furthermore, 5 subtype 3e isolates were closely related and were located in the peripheral branch of subtype 3e isolates from European countries in the phylogenetic tree. The structure indicated that the ancestor of the 5 subtype 3e isolates originated in Europe. The phylogenetic structure and coalescent analyses suggested that the subtype 3e isolates entered Japan from Europe by importation of large-race pigs around 1966. The results also indicated that several lineages of subtype 3e expanded to a wide area of Japan around 1992 and 1 of the lineages was indigenized in wild boars in Mie prefecture between 1992 and 2009. The appearance of a wild boar cluster in the peripheral branch in the phylogenetic lineage may indicate the direction of gene flow of HEV subtype 3e from swine to wild boars. Clarification of the transmission direction or route should be helpful to prevent a future endemic or epidemic of HEV infection. (C) 2013 Elsevier B.V. All rights reserved.

74歳男性。糖尿病および慢性腎不全により血液透析を行っていた。今回、糖尿病に対しビルダグリプチン(VG)投与を開始後、腹部膨満感と便秘症状が出現、更に腹痛および吐気も伴った。画像所見にて小腸および大腸ガスと半結腸主体の便の停滞が認められたため、内服薬を中止し、絶飲食を行い、あわせて補腋を開始したところ、腹痛や腹満は消失し、排便回数や排便量が増加傾向となった。そこで、VG内服の再開を開始したが、腹部症状が増悪したため、再び絶飲食、内服薬中止、末梢輸液を行った。その結果、症状は改善し、経口摂取を再開した。尚、今までの経緯よりVG再開後に便秘症状が再発したことから、同薬剤が便秘の原因の被疑薬と考え、内服を中止したところ、腹痛・腹満・食事摂取量・排便回数・排便量・排便性状は安定した。

In Mie prefecture in Japan, 12 cases of sporadic hepatitis E occurred from 2004 to 2011. Mie prefecture is located in the central region of Japan, far from the most prevalent regions of hepatitis E virus (HEV) infection in Japan, the north and northeastern part. These 12 cases did not have any common risk factors of HEV infection. We analyzed the molecular epidemiology of the cases in Mie prefecture. We obtained the nucleotide sequences of the HEV strains and analyzed them with the sequences of other HEV strains by phylogenetic and coalescent analyses. Japan-indigenous genotype 3 HEV strains were divided into two major subtypes, namely, 3a and 3b; one minor subtype, 3e; and a few other unassigned lineages. The Japan-indigenous subtype 3e strains were closely related to European subtype 3e HEV strains and were comparatively rare in Japan; however, eight strains of the 12 cases we examined belonged to subtype 3e, indicating a close phylogenetic relationship, despite the lack of common risk factors. Coalescent analyses indicated that the Mie 3e strains seemed to have intruded into Mie prefecture about 10 years ago. Sporadic acute hepatitis E cases caused by the 3e strains occurred consistently from 2004 to 2011 in Mie prefecture. This is the first report of unexpected persistent occurrence of hepatitis by the European-type genotype 3 HEV, subtype 3e, in a country outside of Europe. Phylogenetic and coalescent analyses traced the history of the indigenization of the Mie 3e strains from Europe. Because hepatitis E cases caused by 3e strains are relatively rare in Japan, molecular evolutionary analyses of HEV infection in Mie prefecture is important for preventing a future hepatitis endemic or epidemic by 3e strains in Japan. (c) 2012 Elsevier B.V. All rights reserved.

Background: Since previous studies have investigated the population dynamics of Japan-indigenous genotype 3 hepatitis E virus (HEV) using virus sequences, more nucleotide sequences have been determined, and new techniques have been developed for such analysis. Aims: To prevent future hepatitis E epidemic in Japan, this study aimed to elucidate the cause of past HEV expansion. Methods: The epidemic history of Japan-indigenous genotype 3 HEV was determined using the coalescent analysis framework. Bayesian skyline plot (BSP) and Bayesian estimate of phylogeny with relaxed molecular clock models were calculated using Markov chain Monte Carlo sampling. Results: Japan-indigenous strains consist of New World strains (subtype 3a), Japanese strains (3b) and European strains (3e). The oldest lineage, 3b, appeared around 1929. Lineages 3a and 3e appeared around 1960. BSPs indicated similar radical population growth of the 3a and 3b lineages from 1960 to 1980. Conclusions: Population dynamics of the three lineages shared some common characteristics, but had distinguishing features. The appearance of 3a and 3e lineages coincides with the increase of large-race pig importation from Europe and the USA after 1960. The epidemic phase of 3a and 3b strains from 1960 to 1980 could be related to increased opportunity for HEV infection arising from large-scale pig breeding since 1960. Our observations revealed new findings concerning the close relationship between the epidemic history of Japan-indigenous genotype 3 HEV and the improvement of the Japanese pig industry. Infection control in pig farms should be an effective method of preventing HEV infection in humans.

From 2004 to 2010, we have seen four cases of sporadic acute hepatitis E in nothern or central district of Mie Prefecture, Japan, who contracted infection of rare genotype 3 hepatitis E virus (HEV) strains. The HEV strains recovered from the four patients were 99.3-99.5% identical to each other in the 412-nucleotide sequence within ORF2 and homologous to those circulating in European countries such as Spain, United Kingdom, and the Netherlands, suggesting the long-term maintenance of the European type genotype 3 HEV strains in a restricted area in Japan. None of these four patients had histories of consuming uncooked or undercooked meat and/or viscera from infected animals and of travel to European countries, and the source of infectious origin remains unknown. These results warrant careful observation of clinical and subclinical HEV infection in this area and implementation of infection prevention measures in the future. © 2011 The Japan Society of Hepatology.

症例は54歳の男性。経皮内視鏡的胃瘻造設術(percutaneous endoscopic gastrostomy:PEG)が施行された後、脳血管障害後遺症のため他院に入院中であった。X線検査で上部食道に鋭利な人工歯根を有する局部床義歯を認め、胃穹窿部に1本の鋭利な義歯を認めたため紹介となった。内視鏡的静脈瘤結紮術用オーバーチューブとネット鉗子を用いることにより、重篤な合併症を発症することなく、内視鏡的に異物除去を行うことが可能であった。この方法は合併症の予防に有用で、かつ、安全であると考えられた。義歯を有するPEG後栄養管理中の症例においては、口腔ケアの際に義歯が不安定になっていないかどうかを定期的に確認し、義歯の誤飲を未然に防ぐ注意が必要である。(著者抄録)

20歳以下の若年でほぼ同時期に発症したHLA-B27陽性の脊椎関節炎(SpA)の姉妹例を報告した。症例1(二人姉妹の姉)、症例2(症例1の妹)、母方伯母に強直性脊椎炎(AS)の既往があり、2例とも20歳以下で臀部痛などの脊椎関節炎症状が出現し、HLA-B27は陽性であった。SpAのAmor診断基準はそれぞれ11点、10点であったことからSpAと診断され、非ステロイド系抗炎症薬、サラゾスルファジン投与により臨床症状とCRPは正常化した。本症例は女系家族集積性の極めて高いSpAの姉妹例で、ASを含むSpAがまれなリウマチ病態であるなかで、見落とされやすい女性症例に留意する必要性を示すものであり、女性SpA例の特徴を理解するうえで貴重な症例であった。

BSCガストロストミーシステム:BSC gastrostomy system(以下BSCGSと略す)は、ガイドワイヤーを用いずに胃瘻造設後のカテーテル交換を行うことができるバンパータイプ・ボタン型カテーテルキットである。今回BSCGSに改良が加えられたため、交換に要する時間や、バイタルサインの変化について、ガイドワイヤーを用いる同タイプのキットとの比較検討を行った。BSCGSでは、有意に交換時間が短縮され(p<0.01)、バイタルサインの変化に有意差は認めず、有用かつ安全と考えられた。(著者抄録)

難消化性デキストリン含有食品「健糖楽茶」のショ糖負荷試験による血糖値抑制効果について二重盲検クロスオーバー法で検討した。血糖値に異常がない健康被験者20例を対象とした。対照食品あるいは試験食品を速やかに摂取させ、6分後に150mLのショ糖液を速やかに摂取させた。血糖値のピーク値は試験食品群、対照食品群ともショ糖液摂取直後の30分に現れ、血糖値は45分で有意に試験食品群で抑制された。摂取後60分、90分、120分では対照食品群との有意差はなかった。AUCは有意に試験食品群で抑制された。インスリン値の経時変化は、対照食品群と試験食品群に有意差はなく、二元配置分散分析では摂取後時間にのみに有意差を認めた。対照食品群に比べて試験食品群でインスリン値が低値になることはなかった。対照食品群で、血糖値とインスリン値には比較的強い相関を認めた。試験食品及び対照食品を摂取することによる有害事象は認めなかった。

難消化性デキストリン含有食品「健糖楽茶」の反復摂取における安全性について検討した。1回1包、1日3回の試験食品を1ヵ月間摂取する過剰摂取試験を実施した。試験開始前、試験開始2週間後、4週間後、終了後2週間目に理学検査、血液及び尿検査を実施した。29例の、試験食品の平均摂取回数は80.2回、平均摂取率は95.4%と良好なコンプライアンスであった。摂取期間中の有害事象(愁訴)は摂取後2週間に多く発生したが、いずれも軽症で数日以内に解消した。摂取前の愁訴人数と近い人数であった。尿潜血陽性は一過性で、摂取前の陽性反応が継続的に発生しなかった。試験中の脱落例はなく、1ヵ月間過剰摂取試験における血液検査結果及び尿検査の結果では、特別な有意差を示す変動は観察しなかった。いずれかの検査において血糖値が110mg/dLを超えた境界型被験者の臨床検査値にも異状はなかった。安全性はヒト臨床試験で確認した。

我々は、経皮内視鏡的胃瘻造設術(以下PEGと略す)の術後早期の感染症を予防する目的で、オーバーチューブ(以下OTと略す)と胃壁固定具(以下GDと略す)を用い、その有用性と安全性について、検討を行った。OTとGDの併用は肺炎の発症率を有意に低下させた(p<0.05)。PEG施行時に、OTとGDを併用することは、術後早期の肺炎の発症を予防するうえで有用であり、胃壁固定時やOT挿入時に重篤な合併症も認めず、安全であると思われた。(著者抄録)

Aim: To study hepatitis C virus (HCV) selection and hypervariable region-1 (HVR1) evolution in a chimpanzee chronically infected with HCV-1 over 12 years after inoculation with a human factor VIII concentrate contaminated with HCV. Methods: From the inoculum, the earliest chimpanzee plasma and 12 annual plasma samples, HCV fragments including HVR1 were amplified followed by cloning and sequencing. Results: Five HCV subtypes - 1a, 1b, 2a, 2b, 3a - and multiple 1a strains were identified in the inoculum. Two 1a strains were found in the earliest chimpanzee sample, while a single HCV-1 strain was detected in the 12 annual samples. None of the chimpanzee sequences were identical to those found in the inoculum. Over 12 years, HVR1 patterns changed irregularly, but a few patterns showed identical nucleotide or amino acid sequences. In the last three years, the variety of HVR1 patterns decreased, while the proportion of major patterns increased. These corresponded to a higher virus load and a lower number of amino acid substitutions. Simultaneously, the HVR1 sequences became more similar to the consensus sequence of the 1a subtype. Conclusion: HCV selection was observed from the inoculum to the inoculated chimpanzee and from the early acute hepatitis to the persistent chronic infection. The selection occurred at three levels: among subtypes after transmission, among isolates during acute hepatitis and among quasispecies in chronic infection.

62歳女。特発性肺線維症(IPF)の増悪でメチルプレドニゾロンによる入院治療が行われていたが黒色嘔吐、黒色便も出現した。意識清明、眼瞼結膜軽度貧血を認めた。胸部聴診で軽度ラ音を聴取し、心窩部に軽度圧痛を認めたが、腹部は平坦、弾性軟で肝脾は触知せず、下肢に軽度浮腫を認めた。また神経学的異常所見は認めなかった。上部消化管内視鏡で胃上体大彎後壁より毛細血管拡張を認めた。病変は血管集簇に接して長径10mm、短径5mmの類楕円形の血管集簇が連続して存在し出血がみられたため出血性胃血管拡張と診断した。

Here, the complete genome sequences for three hepatitis C virus (HCV) variants identified from China and belonging to genotype 6 are reported: km41, km42 and gz52557. Their entire genome lengths were 9430, 9441 and 9448 nt, respectively; the 5' untranslated regions (UTRs) contained 341, 342 and 339 nt, followed by single open reading frames of 9045, 9045 and 9057 nt, respectively; the 3' UTRs, up to the poly(U) tracts, were 41, 51 and 52 nt, respectively. Phylogenetic analyses showed that km41 is classified into subtype 6k and km42 into subtype 6n. Although gz52557 clustered distantly with subtype 6g, it appeared to belong to a distinct subtype. Analysis with 53 and 105 partial core and NS5B region sequences, respectively, representing 17 subtypes from 6a to 6q and three unassigned isolates of genotype 6 in co-analyses demonstrated that gz52557 was equidistant from all of these isolates, indicating that it belongs to a novel subtype. However, based on a recent consensus that three or more examples are required for a new HCV subtype designation, it is suggested that gz52557 remains unassigned to any subtype.

Subtype 1b is the most common strain of Hepatitis C virus (HCV) in China. Here, the molecular epidemiology and epidemic history of this strain were investigated by conducting phylogenetic and population genetic analyses of El and NS5B gene sequences sampled from nine Chinese cities. The phylogenetic analysis indicated the presence of two clusters of Chinese strains that did not include reference strains from other countries, suggesting that these clusters represent two independent chains of HCV transmission within China. The remaining Chinese isolates were more closely related to reference strains from other countries. The date of origin and past population dynamics of the two groups were investigated using a new population genetic method, the Bayesian skyline plot. The estimated dates of origin of both groups coincide with the period of the Chinese 'Cultural Revolution' during the years 1966-1976. Both groups grew at a rapid exponential rate between similar to 1970 and similar to 1990, after which transmission slowed considerably. Possible explanations for the groups' fast spread and subsequent slowdown are discussed, including parenteral transmission by unsafe injection, iatrogenic transmission by infected blood or blood products and improvements in blood safety since 1990. These results shed light on HCV transmission in China and may help to predict the future burden of HCV-related disease in the country.

During January-April, 2 000, 12 cases of acute hepatitis B were reported in Pierce County, Washington, compared with seven in all of 1999. Seven (58.3%) case patients were injection drug users (IDUs), three of whom were coinfected with hepatitis D virus (HDV) and died of fulminant hepatitis. Vaccination clinics were implemented at the local health department and needle exchange program to control the outbreak. We investigated this outbreak to determine risk factors for hepatitis B virus (HBV) transmission among IDUs. Hepatitis B cases were ascertained through routine surveillance and prevaccination testing at vaccination clinics. We conducted a case-control study comparing IDU case patients with HBV-susceptible IDUs identified at the vaccination clinics. Fifty-eight case patients were identified during January-December, 2000, 20 (34.5%) of whom were coinfected with HDV. Thirty-eight case patients (6S.S%) reported current IDU. In the case-control study, the 17 case patients were more likely than the 141 controls to report having more than one sex partner [odds ratio (OR) = 4.8, 95% confidence interval (CI) = 1.5-15.0], injecting more than four times a day (OR = 4.5, 95% CI= 1.2-15.6) and sharing drug cookers with more than two people (58.8% vs. 14.0%, OR = 14.0, 95% CI = 2.4-81.5). Results were similar after controlling for syringe sharing in multivariable analysis. IDUs should be vaccinated against hepatitis B and should be advised against sharing drug injection equipment.

Background: The response rates and duration of peginterferon alpha ( PEG- IFN- a) and ribavirin combination therapy in chronic hepatitis C genotype 4, the prevalent genotype in the Middle East and Africa, are poorly documented. Aims: To compare the efficacy and safety of 24, 36, or 48 weeks of PEG- IFN- α- 2b and ribavirin therapy in chronic hepatitis C genotype 4. Methods: In this prospective, randomised, double blind study, 287 patients with chronic hepatitis C genotype 4 were randomly assigned to PEG- IFN-α- 2b ( 1.5 mg/ kg) once weekly plus daily ribavirin ( 1000 - 1200 mg) for 24 weeks ( group A, n = 95), 36 weeks ( group B, n = 96), or 48 weeks ( group C, n = 96) and followed for 48 weeks after completion of treatment. Early viral kinetics and histopathological evaluation of pre- and post treatment liver biopsies were performed. The primary end point was viral clearance 48 weeks after completion of treatment. Results: Sustained virological response was achieved in 29%, 66%, and 69% of patients treated with PEGIFN-α- 2b and ribavirin for 24, 36, and 48 weeks, respectively, by intention to treat analysis. No statistically significant difference in sustained virological response rates was detected between 36 and 48 weeks of therapy ( p = 0.3). Subjects with sustained virological response showed greater antiviral efficacy ( e) and rapid viral load decline from baseline to treatment week 4 compared with non- responders and improvement in liver histology. The incidence of adverse events was higher in the group treated for 48 weeks. Conclusion: PEG- IFN- a- 2b and ribavirin for 36 or 48 weeks was more effective in the treatment of chronic hepatitis C genotype 4 than treatment for 24 weeks. Thirty six week therapy was well tolerated and produced sustained virological and histological response rates similar to the 48 week regimen.

The goal of this study was to adapt a long RT-PCR technique to amplify large PCR fragments from the genome of hepatitis C virus (HCV) isolates using clinical samples. This was done by using a reverse transcriptase devoid of RNase H activity and a mixture of two antibody-bound thermostable polymerases to combine the high processivity of Taq and the high fidelity of Pwo with its 3' → 5' exonuclease activity. Other modifications included gentle handling during RNA extraction, the absence of tRNA and random primers, a two-step reverse transcription procedure to optimize cDNA synthesis, and increasing the annealing temperature for primers. With this approach, the HCV-1 genome (nucleotides 35-9282) was amplified consistently as two overlapping fragments of 5344 and 4675 bp from a pooled chimpanzee plasma sample containing approximately 10(6) genome copies of HCV RNA/ml. Using the conditions that we identified, 96% of the complete genomic sequence of a distinct HCV genotype 6 variant (km45) was determined from less than 300 μ l of serum. This method should prove useful for molecular, epidemiological and clinical studies of hepatitis C where samples are limited but complete virus sequence is required, for example, identifying mutational hot spots of HCV under specific clinical conditions. © 2005 Elsevier B.V. All rights reserved.

To determine hepatitis C virus (HCV) genotype distribution in China, a total of 148 HCV RNA positive serum samples were collected from nine geographic areas and subjected to RT-PCR followed by direct DNA sequencing and phylogenetic analysis of the core, E1, and NS5B regions. HCV was genotyped in 139 (93.9%) samples. Among them subtype 1b was the most predominant [66% (92/139)] followed by 2a [14% (19/139)]. Of 92 subtype 1b isolates, 35 (38%) and 30 (33%) formed two clusters, designated groups A and B. Group A was prevalent throughout China, while group B was predominant in the central and southern regions. In three cities in the Pearl River Delta, subtype 6a replaced 2a as the second most predominant subtype, and in Kunming (southwest) multiple HCV genotypes/subtypes were present. New variants of HCV genotype 6 were discovered in three samples from Kunming and one in Guangzhou in the Pearl River Delta.

Acute hepatitis C virus (HCV) is typically defined as new viremia and antibody seroconversion. Rates and immunologic correlates of hepatitis C clearance have therefore been based on clearance of viremia only in individuals who initially had an antibody response. We sought to characterize the immunological correlates of clearance in patients with acute hepatitis C and their sexual contacts. We prospectively determined CD4(+) and CD8(+) cytotoxic T-lymphocyte responses in index patients with acute HCV and their sexual contacts who developed acute infection, either with or without spontaneous clearance, as well as those contacts who never developed viremia. Responses were measured using proliferation and ELISpot assays for CD4(+) and CD8(+) responses. We demonstrate in this prospective study that cellular immune responses can develop in exposed but persistently aviremic and antibody-negative individuals as well as those individuals with spontaneous clearance of acute HCV. These findings lend further credence to the importance of cellular immune responses in recovery from HCV and suggest that low exposure to HCV may lead to development of HCV-specific immune responses without ongoing HCV replication. This finding has important implications for HCV vaccine and therapeutic development.