Naz OĞUZOĞLU, Pinar U. ONGANER, Ayşe ÖZER

Mutation and genotype analysis of hepatitis B virus on acute and chronic infected selected patients in Turkey

Kronik hepatit B virus infeksiyonu Türkiye'de % 8 oranında bulunmaktadır. Bu çalışmada, akut ve kronik hepatit B infeksiyonlu hastalar seçilerek X ve kor genlerine ait baz değişikliklerinin saptanması amaçlanmıştır. Restriksiyon fragment uzunluk polimorfizm yöntemi ile 50 hastanın S gen bölgesine göre genotipleri belirlenmiştir. 42 olguda genotip D (% 84), genotip A 6 olguda (% 12), ve genotip F 2 hastada (% 4) olmak üzere üç farklı genotip saptanmıştır. X ve bu bölge ile üstüste gelen prekor-kor bölgeleri özgün primerler kullanılarak çoğaltılmıştır. Ardından yapılan dizi analizi reaksiyonları ile hastaların baz dizileri belirlenmiştir. HBV X gen bölgesinde 12 farklı nokta mutasyon ve 4 farklı insersiyon tanımlanmıştır. Tüm hastalarda C_T and T_C baz değişimleri gözlenmiş olup söz konusu diğer mutasyonlar sırasıyla, (G-»A), (G-*C), (A->T), (A-»G), (C-*A), (C-»G), (A-»C), (G-*T), (T-s»G), (T-*A)'dır. AntiHBe pozitif 5 olguda viral replikasyonu arttırarak prognozun kötüleşmesine neden olan spesifik mutasyonlar tesbit edilmiştir. HBV infeksiyonları ve mekanizmasının hastalığın seyrindeki önemin anlaşılması için farklı bölgelerden seçilmiş akut ve kronik HBV ile infekte hastalar üzerinde daha fazla çalışma yapılması gerekmektedir.

Türkiye' de akut ve kronik infeksiyonlu hastalardaki hepatit B virüs mutasyon ve genotip analizi

Chronic hepatitis B virus infection is important in Turkey because of the intermediate prevalence (8 %) of persistent HBV infection. The purpose of this study is to demonstrate X and core gene mutations in chronic and acute HBV infected patients in the Turkish population. HBV genotypes were analysed using restriction fragment length polymorphism (RFLP) of the S gene region. The genotypes of fifty HBV infected patients were determined. Three different genotypes were found, genotype D; 42 (84 %), genotype A; 6 (12 %), and genotype F; 2 (4 %). The X and overlapping core regions have been amplified by PCR and sequenced. Seven different mutations were detected in the core region. In the X region, 12 different point mutations and 4 different insertions were detected. All samples had C^T and T^C alterations and the others as follows; (G-»A), (G-»C), (A-*T), (A-»G), (C-»A), (C-»G), (A-*C), (G—>T), (T—>G), (T-*A). Isolates from five AntiHBe positive five patients contained specific mutations, which are reported to increase viral replication and result in a poor prognosis. Further investigations should be carried out using more diverse Turkish patients in order to understand the clinical outcome of these alterations.

PDF

___

Akahane Y, Yamanaka T, Suzuki H, Sugai Y, Tsuda F, Yotsumoto S, Omi S, Okamoto H, Miyakawa Y, and Mayumi M. Chronic active hepatitis with hepatitis B virus DNA and antibody against e antigen in the serum. Disturbed synthesis and secretion of e antigen from hepatocytes due to a point mutation in the precore region. Gastroenterology. 99:1113-1119, 1990
Akarca US, and Lok AS. Naturally occurring hepatitis B virus core gene mutations. Hepatology. 22: 50-60, 1994.
Alexopoulou A, Karayiannis P, Hadziyannis SJ, Aiba N, and Thomas HC. Emergence and selection of HBV variants in an anti-HBe positive patient persistently infected with quasi-species. J Hepatology. 26: 748-753, 1997.
Arii M, Takada S, and Koike K. Identification of three essential regions of hepatitis B virus X protein for trans-activation function. Oncogene. 7:397-403,1992.
Aye TT, Bartholomeusz A, Shaw T, Bowden S, Breschkin A, McMillan J, Angus P, and Locarnini S. Hepatitis B virus polymerase mutations during antiviral therapy in a patient following liver transplantation. J Hepatology. 26:1148-1153, 1997.
Baumert TF, Marrone A, Vergalla J, and Liang TJ. (1998). Naturally occurring mutations definea novel function of the hepatitis B virus core promoter in core protein expression. / Virol. 72: 6785-6795,1998.
Blâckberg J, and Kidd-Ljunggren K. Genotypic differences in the hepatitis B virus core promoter and precore sequences during seroconversion from HBeAg to anti-HBe. JMed Virol. 60:107-112, 2000.
Bozdayi AM, Asian N, Bozdayi G, Turkyilmaz AR, Sengezer T, Wend U, Erkan O, Aydemir F,Zakirhodjaev S, Orucov S, Bozkaya H, Gerlich W, Karayalcin S, Yurdaydin C, and Uzunalimoglu O. Molecular epidemiology of hepatitis B, C and D viruses in Turkish patients. Arch Virol. 149: 2115-2129, 2004.
Brunetto MR, Stemler M, Bonino F, Schodel F, Oliveri F, Rizzetto M, Verme G, and Will H. A new hepatitis B virus strain in patients with severe anti-HBe positive chronic hepatitis. B J Hepatol. 10:258-261, 1990.
Brunetto MR, Giarin MM, Oliveri F, Chiaberge E, Baldi M, Alfarano A, Serra A, Saracco G, Verme G, Will H, et al. Wild-type and e antigen-minus hepatitis B viruses and course of chronic hepatitis. Proc NatlAcad Sci USA. 88: 4186-4190, 1991.
Brunetto MR, Giarin M, Saracco G, Oliveri F, Calvo P, Capra G, Randone A, Abate ML, Manzini P, Capalbo M., et al. Hepatitis B virus unable to secrete e antigen and response to interferon in chronic hepatitis B. Gastroenterology. 105: 845-850, 1993.
Carman WF, Jacyna MR, Hadziyannis S, Karayiannis P, McGarvey MJ, Makris A, and Thomas HC. Mutation preventing formation of hepatitis B e antigen in patients with chronic hepatitis B infection. Lancet. 8663: 588-591, 1989.
Chan HL, Hussain M, and Lok AS. Different hepatitis B virus genotypes are associated with different mutations in the core promoter and precore regions during hepatitis B e antigen seroconversion. Hepatology. 29: 976-84, 1999.
Chen IH, Huang CJ, and Ting LP. Overlapping initiator and TATA box functions in the basal core promoter of ' hepatitis B virus. / Virol. 69:3647-3657, 1995.
De Castro L, Niel C, and Gomes SA. Low frequency of mutations in the core promoter and precore regions of hepatitis B virus in anti-HBe positive Brazilian carriers. BMC Microbiol. 1: 1-10, 2001.
Hsu HY, Chang MH, Lee CY, Hsieh KH, Ni YH, Chen PJ, and Chen DS. Precore mutant of hepatitis B virus in childhood fulminant hepatitis B: an infrequent association. J Infect Dis. 171:776781, 1995.
Jardi R, Rodriguez F, Buti M, Costa X, Valdes A, Ailende H, Schaper M, Galimany R, Esteban R, and Guardia J. Mutations in the basic core promoter region of hepatitis B virus. Relationship with precore variants and HBV genotypes in a Spanish population of HBV carriers. / HepatoL 40: 507-514, 2004.
Jazayeri M, Basuni A A, Sran N, Gish R, Cooksley G, Locarnini S, and Carman WF. HBV core sequence: definition of genotype-specific variability and correlation with geographical origin. J Viral Hepat. 11: 488-501, 2004.
Kao JH, Chen PJ, Lai MY, and Chen DS. Acute exacerbations of chronic hepatitis B are rarely associated with superinfection of hepatitis B virus. Hepatology. 34: 817-23,2001.
Lavine.J, and Hirsch R. A system for studying the selective encapsidation of hepadna virus RNA. J. Virol. 63: 4257-4263, 1989.
Lee Yi, Hur GM, Suh DJ, and Kim SH. Novel pre-C/C gene mutants of hepatitis B virus in chronic active hepatitis: naturally occurring escape mutants. / Gen Virol. 77:1129-1138, 1996.
Li JS, Tong SP, Wen YM, Vitvitski L, Zhang Q, and Trepo C. Hepatitis B virus genotype A rarely circulates as an HBe-minus mutant: possible contribution of a single nucleotide in the precore region. / Virol. 67: 5402-5410, 1993.
Liang TJ, Hasegawa K, Munoz SJ, Shapiro CN, Yoffe B, McMahon BJ, Feng C, Bei H, Alter MJ, and Dienstag JL. Hepatitis B virus precore mutation and fulminant hepatitis in the United States. A polymerase chain reaction-based assay for the detection of specific mutation. / din Invest. 93: 550-555, 1994.
Lindh M, Horal P, Dhillon AP, Furuta Y, and Norkrans G. Hepatitis B virus carriers without precore mutations in hepatitis B e antigen-negative stage show more severe liver damage. Hepatology. 24: 494-501, 1996.
Locarnini S. Molecular virology of hepatitis B virus. Semin Liver Dis. 24:3-10, 2004.
Lok AS, Akarca U, and Greene S. Mutations in the pre-core region of hepatitis B virus serve to enhance the stability of the secondary structure of the pre-genome encapsidation signal. Proc Natl Acad Sci USA. 91: 4077-4081,1994.
Loriot MA, Marcellin P, Talbodec N, Guigonis V, Gigou M, Boyer N, Bezeaud A, Erlinger S, and Benhamou JP. Low frequency of precore hepatitis B virus mutants in anti-hepatitis B e-positive reactivation after loss of hepatitis B e antigen in patients with chronic hepatitis B. Hepatology. 21: 627-631, 1995.
Marcellin P, Castelnau C, Martinot-Peignoux M, and Boyer N. Natural history of hepatitis B. Minerva Gastroenterol Dietol. 51:63-75, 2005.
McMillan JS, Bowden DS, Angus PW, McCaughan GW, and Locarnini SA. Mutations in the hepatitis B virus precore/core gene and core promoter in patients with severe recurrent disease following liver transplantation. Hepatology. 24: 1371-1378, 1996. Mizokami M, Nakano T, Orito E, Tanaka Y, Sakugawa H, Mukaide M, and Robertson BH. Hepatitis B virus genotype assignment using restriction fragment length polymorphism patterns. FEBS Letters. 450: 66-71,1999.
Nowak MA, Bonhoeffer S, Hill AM, Boehme R, Thomas HC, and McDade H. Viral dynamics in hepatitis B virus infection. Proc Natl Acad Sci USA. 93: 4398-4402, 1996.
Onganer PU. Genotypes of the hepatitis B virus: basic and clinical aspects. Ann Microbiol. 54:325-334, 2004.
Okamoto H, Yotsumoto S, Akahane Y, Yamanaka T, Miyazaki Y, Sugai Y, Tsuda F, Tanaka T, Miyakawa Y, and Mayumi M. Hepatitis B viruses with precore region defects prevail in persistently infected hosts along with seroconversion to the antibody against e antigen. J Virol. 64:1298-1303, 1990.
Pollicino T, Campo S, and Raimondo G. PreS and core gene heterogeneity in hepatitis B virus (HBV) genomes isolated from patients long-lasting HBV chronic infection. Virology. 208: 672-677, 1995.
Robinson WS. Molecular events in the pathogenesis of hepadnavirus-associated hepatocellular carcinoma. Ann. Rev. Med. 45:297-323, 1994.
Rodriguez-Frias F, Buti M, Jardi R, Cotrina M, Viladomiu L, Esteban R, and Guardia J. Hepatitis B virus infection: precore mutants and its relation to viral genotypes and core mutations. Hepatology. 22:1641-1647, 1995.
Ruiz PA, Norder H, Robertson BH, and Magnius LO. Genotype H: a new Amerindian genotype of hepatitis B virus revealed in Central America. /. Gen Virol. 83: 2059-2073, 2002.
Santantonio T, Jung MC, Miska S, Pastore G, Pape GR, and Will H. High prevalence and heterogeneity of HBV preC mutants in anti-HBe-positive carriers with chronic liver disease in southern Italy. J Hepatol. 13: 78-81, 1991.
Schaefer S. Hepatitis B virus: significance of genotypes. / Viral Hepat. 12:111-124,2005.
Seeger C, and Mason WS. Hepatitis B biology. Microbiol j and Mol Biol Rew. 64: 51-68, 2000.
Su Q, Schroder H, Hofmann WJ, Otto G, Pichlmay R, and Bannasch P. Expression of hepatitis B virus X protein in HBV-infected human livers and hepatocellular carcinomas. Hepatology. 27:1109-1120, 1998.
Takada S, and Koike K. Three sites of the hepatitis B virus X protein cooperatively interact with cellular proteins. " Virology. 205: 503-510, 1994.
Tassopoulos NC, Volpes R, Pastore G, Heathcote J, Buti M, Goldin RD, Hawley S, Barber J, Condreay L, and Gray DF. Efficacy of lamivudine in patients with hepatitis B e antigeri-negative/hepatitis B virus DNA-positive (precore mutant) chronic hepatitis B. Lamivudine Precore Mutant Study Group. Hepatology. 29: 889-896, 1999.
Tillmann H, Trautwein C, Walker D, Michitaka K, Kubicka S, Boker K, and Manns M. Clinical relevance of mutations in the precpre genome of the hepatitis B virus. Gut. 37: 568-573, 1995.
Uysal P, Oguzoglu N, and Ozer A. HBV Genotyping Using Restriction Fragment Length Polymorphism (RFLP) on the Turkish Population. Marmara Medical Journal. 14: 213-216,2001.
Wai CT, and Fontana RJ. Clinical significance of hepatitis B virus genotypes, variants, and mutants. Clin Liver Dis. 8:321-352, 2004.