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Frequency of hepatitis B virus DNA in anti-HBc positive, HBsAg negative blood donors in Rasht, northern Iran
Transfusion and Apheresis Science 45 (2011) 195–197
Contents lists available at SciVerse ScienceDirect
Transfusion and Apheresis Science journal homepage: www.elsevier.com/locate/transci
Frequency of hepatitis B virus DNA in anti-HBc positive, HBsAg negative blood donors in Rasht, northern Iran Ali Khamesipour a, Zahra Mohtasham Amiri b, Sedigheh Amini Kafiabad c, Farshid Saadat d, Fariborz Mansour-ghanaei e, Abdoul-Reza Esteghamati f, Reza Jafari Shakib d,⇑ a
Center for Research and Training in Skin Diseases and Leprosy, Tehran University of Medical Sciences, Tehran, Iran Department of Community Medicine, Road Trauma Research Center of Guilan, Medical Faculty, Guilan University of Medical Sciences, Rasht, Iran c Department of Immunohematology, Research Center of Iranian Blood Transfusion Organization, Hemmat Highway, Tehran, Iran d Department of Microbiology and Immunology, Medical Faculty, Guilan University of Medical Sciences, Rasht, Iran e Gastrointestinal and Liver Disease Research Center, Medical Faculty, Guilan University of Medical Sciences, Rasht, Iran f Department of Pediatrics, Iran Medical University, Tehran, Iran b
a r t i c l e Keywords: Occult HBV Blood donors Anti-HBc
i n f o
a b s t r a c t Background: One of the important factors in the ensuing safety of blood transfusion is to use a sensitive screening assay for detection of blood-born infective agents such as HBV which transmits through transfusion. To improve the detection rate of HBV infection in blood donors, a cross-sectional study was conducted in Rasht, which is the largest city in the north of Iran to explore the possibility of using anti-HBc as a screening test. Study design and methods: A total of 2041 blood samples negative for HBsAg, Anti-HCV, Anti-HIV I, II and RPR were tested to detect anti-HBc and then the positive anti-HBc samples were further checked for the presence of HBV DNA. Results: The prevalence of anti-HBc positive samples was 3.8% and HBV DNA was detected in only one sample. Conclusions: This study showed that anti-HBc positive blood donors may be a source of HBV transmission and further study for evaluation of HBV DNA in anti-HBc positive blood units is needed. Ó 2011 Elsevier Ltd. All rights reserved.
1. Introduction Blood transfusion saves millions of lives and supports modern surgery and chemotherapy but might cause unwanted reactions such as hepatitis B virus (HBV) infection. The risk of HBV transmission by transfusion is higher than HCV and HIV [1,2]. Although HBsAg screening reduces the Abbreviations: HBV, hepatitis B Virus; HCV, hepatitis C Virus; HIV, human Immunodeficiency Virus; RPR, rapid Plasma Reagin; NAT, nucleic Acid Amplification Technology; Ag, antigen. ⇑ Corresponding author. Address: Department of Microbiology and Immunology, Medical Faculty, Guilan University of Medical Sciences, P.O. Box 41635-3363, Rasht, Iran. Tel.: +98 131 6690099; fax: +98 131 6690036. E-mail address: [email protected] (R.J. Shakib). 1473-0502/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.transci.2011.08.005
rate of HBV transmission, HBsAg negative blood donors may still transmit HBV related to the seronegative window period or occurs during occult HBV infection [3]. Occult HBV infection is defined as the presence of HBV DNA in individuals with undetectable HBsAg outside of the window period [3,4]. The frequency of occult HBV infection depends on the sensitivity of the method of HBsAg and HBV DNA detection, and the HBV prevalence in the population [5]. Occult HBV is most frequently seen in anti-HBc positive individuals as the only serological marker but it might be seen in individuals with no HBV serological marker [4,6]. The clinical significance of occult HBV infection remains largely undefined but impacts on clinical situations such as; (1) transmission of HBV infection through blood trans-
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A. Khamesipour et al. / Transfusion and Apheresis Science 45 (2011) 195–197
fusion, haemodialysis, organ transplantation – especially orthotropic liver transplantation (2) reactivation of occult HBV infection which leads to acute hepatitis B and sometimes even fulminant hepatitis ususally is seen in immuncompromised individuals (3) chronic liver disease: in chronic HCV patients, co-infection with occult HBV infection often associated with progression of the disease and failure of IFN-a therapy (4) hepatocellular carcinoma [7,8]. The real prevalence of occult HBV infection is unknown. The data generated by various studies are not comparable mainly due to differences in the methodology used, however, HBV DNA detection using a PCR method is the most sensitive test currently used, but there is no valid standard assay to detect occult HBV infection [7,9]. It seems that a complementary test is needed to precisely diagnose HBV infection in HBsAg negative blood donors. However, each country needs to develop its own strategy to screen blood samples based on HBV prevalence, detection of infection by different serologic/NAT (nucleic acid amplification technology) screening methods, and cost effectiveness of the screening method to assure that safe blood is used [10–12]. In some countries, both HBsAg and anti-HBc screening tests are routinely used to reduce the rate of HBV transmision. Such screening is not suitable for countries with a high HBV infection rate, because using such criteria may cause most of the donated blood to be discarded [13,14]. An important tool for HBV detection is to use NAT which is expensive and not available everywhere. The aim of this study is to explore the possibility of using anti-HBc as a screening test to improve the HBV detection rate in the donated blood in the city of Rasht which is the largest city northern Iran. 2. Materials and methods 2.1. Subjects In this cross sectional study, blood samples which were donated by volunteers to the Iranian Blood Transfusion Organization in Rasht were randomly selected for detection of anti-HBc antibody. The blood samples were collected in 6 ml vaccum tubes containing gel and clot activator. The tubes were centrifuged at 3500 rpm for 10 min, and sera were separated and stored at 20° C until use. A first-time blood donor was defined as a donor who donated blood for the first time. A regular donor was defined as a donor who donated blood more than once during one year, and a repeat donor was defined as donor who had a history of previous donation but the interval between two donations was more than one year. 2.2. Methods 2041 blood samples which were negative for HBsAg, anti-HCV, anti-HIV I, II, and RPR were tested for anti-HBc (Diasorin, Italy). Then the anti-HBc positive sera were evaluated to detect HBV DNA. DNA extractions were done using a commercial DNA extraction kit (QIAamp DNA Mini Kit) and extracted DNA was analyzed using commercial real time PCR kit (artus HBV LC PCR Kit). A positive sample
was rechecked with a sensitive homemade PCR kit (locally produced and used by the Iranian Blood Transfusion Organization; IBTO) and the amplified PCR products were analyzed using agarose gel electrophoresis followed by ethidium bromide staining and ultraviolet visualization. The experiments were performed under strict conditions to avoid any contamination. 3. Results Characteristics of the volunteers regarding frequency of blood donotion, age and sex are summarized in Table 1. The mean age is 36.23 ± 10.81. A total of 78 out of 2041 of the HBsAg negative blood samples (3.8%) were found to be positive for anti-HBc. One out of the 78 anti-HBc positive samples was positive for HBV DNA. The positive sample was further confirmed using another PCR (home made). 4. Discussion The prevalence of HBsAg in Iran varies from 1.2% to 9.7% [15]. Currently in Iran, blood samples are checked only for HBsAg as a marker of HBV infection. HBV DNA was detectable in 0–4.6% of anti-HBc positive blood donors studied in different countries of Asia [12,16]. There are reports from Iran which show that using anti-HBc screening in blood donors is useful [17–20,22,23]. In a study performed in Rafsanjan, in 2006, it was shown that out of 270 healthy blood donors, 14 (5.2%) samples were positive for anti-HBc then, when the samples were further tested using PCR, HBV-DNA was detected in 4 out of 14 (28.57%) [17]. In another study carried out in the city of Isfahan, the anti-HBc was seen in 43 (8%) out of 545 HBsAg negative blood donors and when the anti-HBc positive, HBsAg negative blood donors were tested by conventional PCR, HBV DNA was positive in 5 (11.6%) blood samples [18]. In a study completed in Shiraz in 2006 in which 2000 serum samples negative for HBsAg from healthy blood donors were tested for anti-HBc. The results showed that 131 (6.55%) of the blood samples were positive. Among the anti-HBc positive samples, HBV DNA was detected in 16 (12.2%) samples using conventional PCR [19]. The results of the above studies showed that the presence of HBV DNA in anti-HBc positive individuals in three Table 1 Characteristic of study group. Characteristic Donor status Regular Repeat First-time Sex Male Female Age 17–24 25–34 35–44 45–54 55–64 >65
No. (%) (n = 2041)
Anti-HBc+ (%)
1089 (53.4%) 421 (20.6%) 531 (26%)
45 (4.13%) 15 (3.56%) 18 (3.39%)
1919 (94%) 122 (6%)
74 (3.86%) 4 (3.28%)
336 (16.5%) 643 (31.5%) 561 (27.5%) 392 (19.2%) 104 (5.1%) 5 (0.25%)
4 (1.19%) 13 (2.02%) 23 (4.1%) 27 (6.89%) 10 (9.61%) 1 (20%)
A. Khamesipour et al. / Transfusion and Apheresis Science 45 (2011) 195–197
cities in the center of Iran were considerably high and using anti-HBc to screen blood samples reduces the rate of HBV transmission in blood transfusion. In the current study, anti-HBc was used to screen the samples and a PCR method was used to detect HBV DNA in anti-HBc positive samples in Rasht in the north of Iran, the results showed that HBV DNA is present in one sample out of 78 anti-HBc positive HBsAg negative blood donors. The discrepancy which was seen in the three studies mentioned might be due to the sampling; in the present study the blood samples were taken from the accessory bag instead of in cord blood bags which diminishes the rate of contamination as shown by Amini et al. [20]. Another explanation is that the frequency of occult HBV infection is dependent on the HBV prevalence in the population [5] and the prevalence of HBV in blood donors in Rasht and Guilan provinces is lower than in many of the other parts of Iran [21]. In a recent study it was shown that the prevalence of HBV-DNA in anti-HBc positive blood donors in Tehran is 199 (9.95%) in 2000 blood donors, but when 50 samples negative for anti HBs and positive for anti HBc were tested for HBV DNA using the Arthus kit, the same kit used in the current study, only 2 samples were positive for HBV DNA which is lower than in the previous studies [22]. In 2007, Amini and her colleagues showed that 230 (11.5%) out of 2000 samples were positive for anti-HBc and 3 out of 230 samples were positive for HBV DNA [20]. The results of a recent study completed in Arak, showed that antiHBc was positive in 11 (2.1%) of 531 blood donors but no positive HBV-DNA was found in 11 anti-HBc positive samples which might be due to not enough sample size [23]. In conclusion, the results showed that HBV DNA exists in individuals with a positive anti-HBc in Rasht. The rate of anti-HBc positivity is less than in many other parts of the country. As anti-HBc positive blood donors may be a potential source of HBV transmission further study for evaluation of HBV DNA in anti-HBc positive blood units is strongly recommended. Type of contribution Ali Khamesipour: design of the study, writing the manuscript and critical revising. Zahra Mohtasham Amiri: acquisition of data, analysis and interpretation of data. Sedigheh Amini Kafiabad: performing the experiments. Farshid Saadat: performing the experiments. Fariborz Mansour-ghanaei: writing the manuscript and critical revising. Abdoul-Reza Esteghamati: design of the study, final approval of the version to be submitted. Reza Jafari Shakib: design of the study, acquisition of data, performing the experiments, analysis and interpretation of data, writing the manuscript and critical revising. Acknowledgments The authors are grateful to Mr. Mohammadreza Heidarzadeh from the Guilan blood bank, Rasht, Iran, Dr Zohreh Sharifi and Fahimeh Ranjbar Kermani from the Iranian
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Blood Transfusion Organization-Research Center for technical assistance and Dr Hooshang Lahooti from the university of Sydney. This study was supported by the Regional Office for the Eastern Mediterranean EMRO/WHO, small Grant ID No. SGS07/41. References [1] Schreiber GB, Busch MP, Kleinman SH, Korelitz JJ. The risk of transfusion-transmitted viral infections. The Retrovirus Epidemiology Donor Study. N Engl J Med 1996;334:1685–90. [2] Comanor L, Holland P. Hepatitis B virus blood screening: unfinished agendas. Vox Sang 2006;91:1–12. [3] Candotti D, Allain JP. Transfusion-transmitted hepatitis B virus infection. J Hepatol 2009;51:798–809. [4] Torbenson M, Thomas DL. Occult hepatitis B. Lancet Infect Dis 2002;2:479–86. [5] Allain JP. Occult hepatitis B virus infection. Transfus Clin Biol 2004;11:18–25. [6] Chemin I, Zoulim F, Merle P, Arkhis A, Chevallier M, Kay A. High incidence of hepatitis B infections among chronic hepatitis cases of unknown aetiology. J Hepatol 2001;34:447–54. [7] Raimondo G, Pollicino T, Cacciola I, Squadrito G. Occult hepatitis B virus infection. J Hepatol 2007;46:160–70. [8] Hu KQ. Occult hepatitis B virus infection and its clinical implications. J Viral Hepat 2002;l(9):243–57. [9] Raimondo G, Pollicino T, Romanò L, Zanetti AR. A 2010 update on occult hepatitis B infection. Pathol Biol (Paris) 2010;58:254–7. [10] Kuhns MC, Busch MP. New strategies for blood donor screening for hepatitis B virus: nucleic acid testing versus immunoassay methods. Mol Diagn Ther 2006;10:77–91. [11] Altunay H, Kosan E, Birinci I, Aksoy A, Kirali K, Saribas S, et al. Are isolated anti-HBc blood donors in high risk group? The detection of HBV DNA in isolated anti-HBc cases with nucleic acid amplification test (NAT) based on transcription-mediated amplification (TMA) and HBV discrimination. Transfus Apher Sci 2010;43:265–8. [12] Shang G, Yan Y, Yang B, Shao C, Wang F, Li Q, et al. DNA+/HBsAgblood donors identified by HBV NAT in Shenzhen, China. Transfus Apher Sci 2009;41:3–7. [13] Kleinman SH, Busch MP. HBV: amplified and back in the blood safety spotlight. Transfusion 2001;41:1081–5. [14] Wang JT, Lee CZ, Chen PJ, Wang TH, Chen DS. Transfusiontransmitted HBV infection in an endemic area: the necessity of more sensitive screening for HBV carriers. Transfusion 2002;42:1592–7. [15] Alavian SM, Hajarizadeh B, Ahmadzad-Asl M, Kabir A, BagheriLankarani K. Hepatitis B Virus Infection in Iran: A Systematic Review. Hep Mon 2008;8:281–94. [16] Liu CJ, Chen DS, Chen PJ. Epidemiology of HBV infection in Asian blood donors: Emphasis on occult HBV infection and the role of NAT. J Clin Virol 2006;36(Suppl. 1). [17] Jafarzadeh A, Kazemi Arababadi M, Mirzaee M, Pourazar A. Occult hepatitis B virus infection among blood donors with antibodies to hepatitis B core antigen. Acta Medica Iranica 2008;46:27–32. [18] Pourazar A, Salehi M, Jafarzadeh A, Kazemi M, Oreizi F, Shariatnezhad K. Detection of HBV DNA in HBsAg normal blood donors. Iranian J Immunol 2005;2:172–6. [19] Behzad-Behbahani A, Mafi-Nejad A, Tabei SZ, Lankarani KB, Torab A, Moaddeb A. Anti-HBc & HBV-DNA detection in blood donors negative for hepatitis B virus surface antigen in reducing risk of transfusion associated HBV infection. Indian J Med Res 2006;123:37–42. [20] Amini Kafi-abad S, Talebian A, Moghtadaie M, Ranjbar Kermani F, Ferdowsian F, Samie S, et al. Detection of hepatitis B virus DNA (PCR) in HBsAg negative, anti-HBc positive blood donors in Tehran province. SJIBTO 2007;3:379–87 [in Persian]. [21] Mansour Ghanaei F, Fallah MS, Jafarshad R, Joukar F, Salari A, Tavafzadeh R. Prevalence of hepatitis B surface antigen and hepatitis C virus antibody and their risk factors among Guilan’s volunteer blood donors (1998–2003). Hep Mon 2007;7:239–41. [22] Alizadeh Z, Sharifi Z, Samiei S, Talebian A. Survey of frequency and load of HBV-DNA in HBsAg negative blood donors using real-time PCR. Vox Sanguinis 2008; 95(Suppl. 1): 285, P625. [23] Sofian M, Aghakhani A, Izadi N, Banifazl M, Kalantar E, Eslamifar A, et al. Lack of occult hepatitis B virus infection among blood donors with isolated hepatitis B core antibody living in an HBV low prevalence region of Iran. Int J Infect Dis 2010;14:e308–10.
Contents lists available at SciVerse ScienceDirect
Transfusion and Apheresis Science journal homepage: www.elsevier.com/locate/transci
Frequency of hepatitis B virus DNA in anti-HBc positive, HBsAg negative blood donors in Rasht, northern Iran Ali Khamesipour a, Zahra Mohtasham Amiri b, Sedigheh Amini Kafiabad c, Farshid Saadat d, Fariborz Mansour-ghanaei e, Abdoul-Reza Esteghamati f, Reza Jafari Shakib d,⇑ a
Center for Research and Training in Skin Diseases and Leprosy, Tehran University of Medical Sciences, Tehran, Iran Department of Community Medicine, Road Trauma Research Center of Guilan, Medical Faculty, Guilan University of Medical Sciences, Rasht, Iran c Department of Immunohematology, Research Center of Iranian Blood Transfusion Organization, Hemmat Highway, Tehran, Iran d Department of Microbiology and Immunology, Medical Faculty, Guilan University of Medical Sciences, Rasht, Iran e Gastrointestinal and Liver Disease Research Center, Medical Faculty, Guilan University of Medical Sciences, Rasht, Iran f Department of Pediatrics, Iran Medical University, Tehran, Iran b
a r t i c l e Keywords: Occult HBV Blood donors Anti-HBc
i n f o
a b s t r a c t Background: One of the important factors in the ensuing safety of blood transfusion is to use a sensitive screening assay for detection of blood-born infective agents such as HBV which transmits through transfusion. To improve the detection rate of HBV infection in blood donors, a cross-sectional study was conducted in Rasht, which is the largest city in the north of Iran to explore the possibility of using anti-HBc as a screening test. Study design and methods: A total of 2041 blood samples negative for HBsAg, Anti-HCV, Anti-HIV I, II and RPR were tested to detect anti-HBc and then the positive anti-HBc samples were further checked for the presence of HBV DNA. Results: The prevalence of anti-HBc positive samples was 3.8% and HBV DNA was detected in only one sample. Conclusions: This study showed that anti-HBc positive blood donors may be a source of HBV transmission and further study for evaluation of HBV DNA in anti-HBc positive blood units is needed. Ó 2011 Elsevier Ltd. All rights reserved.
1. Introduction Blood transfusion saves millions of lives and supports modern surgery and chemotherapy but might cause unwanted reactions such as hepatitis B virus (HBV) infection. The risk of HBV transmission by transfusion is higher than HCV and HIV [1,2]. Although HBsAg screening reduces the Abbreviations: HBV, hepatitis B Virus; HCV, hepatitis C Virus; HIV, human Immunodeficiency Virus; RPR, rapid Plasma Reagin; NAT, nucleic Acid Amplification Technology; Ag, antigen. ⇑ Corresponding author. Address: Department of Microbiology and Immunology, Medical Faculty, Guilan University of Medical Sciences, P.O. Box 41635-3363, Rasht, Iran. Tel.: +98 131 6690099; fax: +98 131 6690036. E-mail address: [email protected] (R.J. Shakib). 1473-0502/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.transci.2011.08.005
rate of HBV transmission, HBsAg negative blood donors may still transmit HBV related to the seronegative window period or occurs during occult HBV infection [3]. Occult HBV infection is defined as the presence of HBV DNA in individuals with undetectable HBsAg outside of the window period [3,4]. The frequency of occult HBV infection depends on the sensitivity of the method of HBsAg and HBV DNA detection, and the HBV prevalence in the population [5]. Occult HBV is most frequently seen in anti-HBc positive individuals as the only serological marker but it might be seen in individuals with no HBV serological marker [4,6]. The clinical significance of occult HBV infection remains largely undefined but impacts on clinical situations such as; (1) transmission of HBV infection through blood trans-
196
A. Khamesipour et al. / Transfusion and Apheresis Science 45 (2011) 195–197
fusion, haemodialysis, organ transplantation – especially orthotropic liver transplantation (2) reactivation of occult HBV infection which leads to acute hepatitis B and sometimes even fulminant hepatitis ususally is seen in immuncompromised individuals (3) chronic liver disease: in chronic HCV patients, co-infection with occult HBV infection often associated with progression of the disease and failure of IFN-a therapy (4) hepatocellular carcinoma [7,8]. The real prevalence of occult HBV infection is unknown. The data generated by various studies are not comparable mainly due to differences in the methodology used, however, HBV DNA detection using a PCR method is the most sensitive test currently used, but there is no valid standard assay to detect occult HBV infection [7,9]. It seems that a complementary test is needed to precisely diagnose HBV infection in HBsAg negative blood donors. However, each country needs to develop its own strategy to screen blood samples based on HBV prevalence, detection of infection by different serologic/NAT (nucleic acid amplification technology) screening methods, and cost effectiveness of the screening method to assure that safe blood is used [10–12]. In some countries, both HBsAg and anti-HBc screening tests are routinely used to reduce the rate of HBV transmision. Such screening is not suitable for countries with a high HBV infection rate, because using such criteria may cause most of the donated blood to be discarded [13,14]. An important tool for HBV detection is to use NAT which is expensive and not available everywhere. The aim of this study is to explore the possibility of using anti-HBc as a screening test to improve the HBV detection rate in the donated blood in the city of Rasht which is the largest city northern Iran. 2. Materials and methods 2.1. Subjects In this cross sectional study, blood samples which were donated by volunteers to the Iranian Blood Transfusion Organization in Rasht were randomly selected for detection of anti-HBc antibody. The blood samples were collected in 6 ml vaccum tubes containing gel and clot activator. The tubes were centrifuged at 3500 rpm for 10 min, and sera were separated and stored at 20° C until use. A first-time blood donor was defined as a donor who donated blood for the first time. A regular donor was defined as a donor who donated blood more than once during one year, and a repeat donor was defined as donor who had a history of previous donation but the interval between two donations was more than one year. 2.2. Methods 2041 blood samples which were negative for HBsAg, anti-HCV, anti-HIV I, II, and RPR were tested for anti-HBc (Diasorin, Italy). Then the anti-HBc positive sera were evaluated to detect HBV DNA. DNA extractions were done using a commercial DNA extraction kit (QIAamp DNA Mini Kit) and extracted DNA was analyzed using commercial real time PCR kit (artus HBV LC PCR Kit). A positive sample
was rechecked with a sensitive homemade PCR kit (locally produced and used by the Iranian Blood Transfusion Organization; IBTO) and the amplified PCR products were analyzed using agarose gel electrophoresis followed by ethidium bromide staining and ultraviolet visualization. The experiments were performed under strict conditions to avoid any contamination. 3. Results Characteristics of the volunteers regarding frequency of blood donotion, age and sex are summarized in Table 1. The mean age is 36.23 ± 10.81. A total of 78 out of 2041 of the HBsAg negative blood samples (3.8%) were found to be positive for anti-HBc. One out of the 78 anti-HBc positive samples was positive for HBV DNA. The positive sample was further confirmed using another PCR (home made). 4. Discussion The prevalence of HBsAg in Iran varies from 1.2% to 9.7% [15]. Currently in Iran, blood samples are checked only for HBsAg as a marker of HBV infection. HBV DNA was detectable in 0–4.6% of anti-HBc positive blood donors studied in different countries of Asia [12,16]. There are reports from Iran which show that using anti-HBc screening in blood donors is useful [17–20,22,23]. In a study performed in Rafsanjan, in 2006, it was shown that out of 270 healthy blood donors, 14 (5.2%) samples were positive for anti-HBc then, when the samples were further tested using PCR, HBV-DNA was detected in 4 out of 14 (28.57%) [17]. In another study carried out in the city of Isfahan, the anti-HBc was seen in 43 (8%) out of 545 HBsAg negative blood donors and when the anti-HBc positive, HBsAg negative blood donors were tested by conventional PCR, HBV DNA was positive in 5 (11.6%) blood samples [18]. In a study completed in Shiraz in 2006 in which 2000 serum samples negative for HBsAg from healthy blood donors were tested for anti-HBc. The results showed that 131 (6.55%) of the blood samples were positive. Among the anti-HBc positive samples, HBV DNA was detected in 16 (12.2%) samples using conventional PCR [19]. The results of the above studies showed that the presence of HBV DNA in anti-HBc positive individuals in three Table 1 Characteristic of study group. Characteristic Donor status Regular Repeat First-time Sex Male Female Age 17–24 25–34 35–44 45–54 55–64 >65
No. (%) (n = 2041)
Anti-HBc+ (%)
1089 (53.4%) 421 (20.6%) 531 (26%)
45 (4.13%) 15 (3.56%) 18 (3.39%)
1919 (94%) 122 (6%)
74 (3.86%) 4 (3.28%)
336 (16.5%) 643 (31.5%) 561 (27.5%) 392 (19.2%) 104 (5.1%) 5 (0.25%)
4 (1.19%) 13 (2.02%) 23 (4.1%) 27 (6.89%) 10 (9.61%) 1 (20%)
A. Khamesipour et al. / Transfusion and Apheresis Science 45 (2011) 195–197
cities in the center of Iran were considerably high and using anti-HBc to screen blood samples reduces the rate of HBV transmission in blood transfusion. In the current study, anti-HBc was used to screen the samples and a PCR method was used to detect HBV DNA in anti-HBc positive samples in Rasht in the north of Iran, the results showed that HBV DNA is present in one sample out of 78 anti-HBc positive HBsAg negative blood donors. The discrepancy which was seen in the three studies mentioned might be due to the sampling; in the present study the blood samples were taken from the accessory bag instead of in cord blood bags which diminishes the rate of contamination as shown by Amini et al. [20]. Another explanation is that the frequency of occult HBV infection is dependent on the HBV prevalence in the population [5] and the prevalence of HBV in blood donors in Rasht and Guilan provinces is lower than in many of the other parts of Iran [21]. In a recent study it was shown that the prevalence of HBV-DNA in anti-HBc positive blood donors in Tehran is 199 (9.95%) in 2000 blood donors, but when 50 samples negative for anti HBs and positive for anti HBc were tested for HBV DNA using the Arthus kit, the same kit used in the current study, only 2 samples were positive for HBV DNA which is lower than in the previous studies [22]. In 2007, Amini and her colleagues showed that 230 (11.5%) out of 2000 samples were positive for anti-HBc and 3 out of 230 samples were positive for HBV DNA [20]. The results of a recent study completed in Arak, showed that antiHBc was positive in 11 (2.1%) of 531 blood donors but no positive HBV-DNA was found in 11 anti-HBc positive samples which might be due to not enough sample size [23]. In conclusion, the results showed that HBV DNA exists in individuals with a positive anti-HBc in Rasht. The rate of anti-HBc positivity is less than in many other parts of the country. As anti-HBc positive blood donors may be a potential source of HBV transmission further study for evaluation of HBV DNA in anti-HBc positive blood units is strongly recommended. Type of contribution Ali Khamesipour: design of the study, writing the manuscript and critical revising. Zahra Mohtasham Amiri: acquisition of data, analysis and interpretation of data. Sedigheh Amini Kafiabad: performing the experiments. Farshid Saadat: performing the experiments. Fariborz Mansour-ghanaei: writing the manuscript and critical revising. Abdoul-Reza Esteghamati: design of the study, final approval of the version to be submitted. Reza Jafari Shakib: design of the study, acquisition of data, performing the experiments, analysis and interpretation of data, writing the manuscript and critical revising. Acknowledgments The authors are grateful to Mr. Mohammadreza Heidarzadeh from the Guilan blood bank, Rasht, Iran, Dr Zohreh Sharifi and Fahimeh Ranjbar Kermani from the Iranian
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Blood Transfusion Organization-Research Center for technical assistance and Dr Hooshang Lahooti from the university of Sydney. This study was supported by the Regional Office for the Eastern Mediterranean EMRO/WHO, small Grant ID No. SGS07/41. References [1] Schreiber GB, Busch MP, Kleinman SH, Korelitz JJ. The risk of transfusion-transmitted viral infections. The Retrovirus Epidemiology Donor Study. N Engl J Med 1996;334:1685–90. [2] Comanor L, Holland P. Hepatitis B virus blood screening: unfinished agendas. Vox Sang 2006;91:1–12. [3] Candotti D, Allain JP. Transfusion-transmitted hepatitis B virus infection. J Hepatol 2009;51:798–809. [4] Torbenson M, Thomas DL. Occult hepatitis B. Lancet Infect Dis 2002;2:479–86. [5] Allain JP. Occult hepatitis B virus infection. Transfus Clin Biol 2004;11:18–25. [6] Chemin I, Zoulim F, Merle P, Arkhis A, Chevallier M, Kay A. High incidence of hepatitis B infections among chronic hepatitis cases of unknown aetiology. J Hepatol 2001;34:447–54. [7] Raimondo G, Pollicino T, Cacciola I, Squadrito G. Occult hepatitis B virus infection. J Hepatol 2007;46:160–70. [8] Hu KQ. Occult hepatitis B virus infection and its clinical implications. J Viral Hepat 2002;l(9):243–57. [9] Raimondo G, Pollicino T, Romanò L, Zanetti AR. A 2010 update on occult hepatitis B infection. Pathol Biol (Paris) 2010;58:254–7. [10] Kuhns MC, Busch MP. New strategies for blood donor screening for hepatitis B virus: nucleic acid testing versus immunoassay methods. Mol Diagn Ther 2006;10:77–91. [11] Altunay H, Kosan E, Birinci I, Aksoy A, Kirali K, Saribas S, et al. Are isolated anti-HBc blood donors in high risk group? The detection of HBV DNA in isolated anti-HBc cases with nucleic acid amplification test (NAT) based on transcription-mediated amplification (TMA) and HBV discrimination. Transfus Apher Sci 2010;43:265–8. [12] Shang G, Yan Y, Yang B, Shao C, Wang F, Li Q, et al. DNA+/HBsAgblood donors identified by HBV NAT in Shenzhen, China. Transfus Apher Sci 2009;41:3–7. [13] Kleinman SH, Busch MP. HBV: amplified and back in the blood safety spotlight. Transfusion 2001;41:1081–5. [14] Wang JT, Lee CZ, Chen PJ, Wang TH, Chen DS. Transfusiontransmitted HBV infection in an endemic area: the necessity of more sensitive screening for HBV carriers. Transfusion 2002;42:1592–7. [15] Alavian SM, Hajarizadeh B, Ahmadzad-Asl M, Kabir A, BagheriLankarani K. Hepatitis B Virus Infection in Iran: A Systematic Review. Hep Mon 2008;8:281–94. [16] Liu CJ, Chen DS, Chen PJ. Epidemiology of HBV infection in Asian blood donors: Emphasis on occult HBV infection and the role of NAT. J Clin Virol 2006;36(Suppl. 1). [17] Jafarzadeh A, Kazemi Arababadi M, Mirzaee M, Pourazar A. Occult hepatitis B virus infection among blood donors with antibodies to hepatitis B core antigen. Acta Medica Iranica 2008;46:27–32. [18] Pourazar A, Salehi M, Jafarzadeh A, Kazemi M, Oreizi F, Shariatnezhad K. Detection of HBV DNA in HBsAg normal blood donors. Iranian J Immunol 2005;2:172–6. [19] Behzad-Behbahani A, Mafi-Nejad A, Tabei SZ, Lankarani KB, Torab A, Moaddeb A. Anti-HBc & HBV-DNA detection in blood donors negative for hepatitis B virus surface antigen in reducing risk of transfusion associated HBV infection. Indian J Med Res 2006;123:37–42. [20] Amini Kafi-abad S, Talebian A, Moghtadaie M, Ranjbar Kermani F, Ferdowsian F, Samie S, et al. Detection of hepatitis B virus DNA (PCR) in HBsAg negative, anti-HBc positive blood donors in Tehran province. SJIBTO 2007;3:379–87 [in Persian]. [21] Mansour Ghanaei F, Fallah MS, Jafarshad R, Joukar F, Salari A, Tavafzadeh R. Prevalence of hepatitis B surface antigen and hepatitis C virus antibody and their risk factors among Guilan’s volunteer blood donors (1998–2003). Hep Mon 2007;7:239–41. [22] Alizadeh Z, Sharifi Z, Samiei S, Talebian A. Survey of frequency and load of HBV-DNA in HBsAg negative blood donors using real-time PCR. Vox Sanguinis 2008; 95(Suppl. 1): 285, P625. [23] Sofian M, Aghakhani A, Izadi N, Banifazl M, Kalantar E, Eslamifar A, et al. Lack of occult hepatitis B virus infection among blood donors with isolated hepatitis B core antibody living in an HBV low prevalence region of Iran. Int J Infect Dis 2010;14:e308–10.