Introduction
Hepatitis B virus (HBV) infection is the most common cause of acute and chronic liver
disease worldwide; it continues to be a serious global health problem [1]. The course of
chronic HBV infection progresses from light liver disease with remission to active liver
disease [2,3]. Approximately one-third of the world’s population (2 billion people) is
estimated to have been exposed to this virus. It is known that about 3–4 million people carry
HBV in our country [4].
With the development of highly sensitive molecular techniques, HBV DNA has been
detected more frequently in hepatitis B surface antigen-negative (HBsAg(−)) persons. The
first occurrences were observed in 1980 in patients with hepatocellular carcinoma (HCC) or
chronic hepatitis; these patients were HVC-negative [HCV(−)] and had no other underlying
causes. In subsequent years, occult HBV infection (OHBI) has been observed in patients
without liver disease and in persons with completely normal liver function.
OHBI is defined as undetectable serum HBsAg levels and the presence of HBV DNA in the
serum, lymphocytes, or liver. The presence of hepatitis B core antigen (HBcAg) or HBsAg
may be positive or negative in these individuals [5-7]. Today, PCR can be used as a
diagnostic tool for the detection of HBV DNA in patient sera. HBV DNA levels exist at low
titers in patients with OHBI. Today, the clinical and biological spectra of OHBI are not
precisely known. HIV infection, nonalcoholic liver disease, and other liver diseases (e.g., in
HIV-infected and hemodialysis patients) may be associated with OHBI [8-12]. OHBI can
also serve as an additional risk factor for HCC, alcoholic cirrhosis, nonalcoholic fatty liver
disease, and progression of HIV infection [13,14].
The gold standard method for the diagnosis of OHBI is the analysis of HBV DNA
extracts from the liver and blood. However, liver extracts are only available in a minority of
cases; therefore, the diagnosis of OHBI is most often based on the analysis of serum samples.
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the country.
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In general, OHBI is defined as the presence of HBV DNA in the livers of individuals who
test negative for HBsAg, regardless of the presence of HBV DNA in the serum [15].
The etiology of OHBI is unknown in 10% of patients with liver cirrhosis, termed
cryptogenic liver cirrhosis [16]. OHBI may be an important etiological cause of cryptogenic
cirrhosis.
OHBI frequency is not known in patients with cryptogenic cirrhosis in our region. We
aimed to determine the frequency of OHBI in patients with cryptogenic cirrhosis who have
been monitored in our hepatology clinic.
METHODS
This cross-sectional study was conducted at Adana Numune Training and Research Hospital.
Patients who were monitored for cryptogenic cirrhosis in our clinic between January 2010
and January 2017 were enrolled in the study. Cirrhosis was diagnosed using biopsy,
laboratory, or radiological findings of hepatic failure and the presence of portal hypertension.
Forty-four patients were diagnosed with cryptogenic cirrhosis, according to the following
criteria: absence of serological markers of HBV, HBC, and mitochondrial, nuclear, and
smooth muscle antigens; normal ceruloplasmin, iron, and alpha 1-antitrypsin levels; and
absence of ingestion of alcohol or other hepatic toxins.
Hepatic and viral markers
Quantification of serological markers was performed using the platform ARCHITECT
ci16200 (Abbott, USA). Serological markers of liver function, such as aspartate
aminotransferase (AST) and alanine aminotransferase (ALT), were quantified using the
activated alanine or aspartate aminotransferase assay (Abbott, USA). Serological markers of
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at their first mention in the main text.
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HBV infection were measured using the chemiluminescent microparticle immunoassay
(Abbott, USA).
HBV DNA detection
HBV DNA was extracted from 200 μL serum by the Cobas AmpliPrep method, according to
the manufacturer’s instructions. The Cobas TaqMan 48 analyzer was used for automated realtime
PCR amplification and detection of PCR products, according to the manufacturer’s
instructions. The data generated were analyzed by Amplilink software. HBV DNA levels are
expressed in international units per milliliter (IU/mL).
Ethical considerations
The study was approved by the Ethics Committee of Çukurova University (No:32-2016).
Statistical analysis
Data are presented as mean ± standard deviation. The data were analyzed using SPSS.
RESULTS
Forty-four patients were enrolled in this study. Table I shows the demographic and clinical
data of patients with cryptogenic cirrhosis in the study group. The mean follow-up time was
3.05 ± 1.55 years. Among the patients with cryptogenic cirrhosis, 29 (65.1%) were males and
15 (34.9%) were females (male to female ratio = 1.93). The mean age at the time of the study
was 62.93 ± 14.11 years.
The overall study group had a mild increase in serum transaminase levels, with median serum
ALT and AST levels of 45 IU/L (range: 10–40 IU/L) and 48 IU/L (range: 0–40 IU/L),
respectively.
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Of the 44 study participants, 22 patients (50%) were positive for HBsAg and 24 (54.5%)
were positive for HBcAg. In our study, the frequency of OHBI was 4.5% (2/44) in patients
with cryptogenic cirrhosis. All OHBI cases were older than 40 years of age.
Table I. General characteristics of the patients in this study
DISCUSSION
Chronic HBV infection is characterized by the presence of HBV DNA in serum and
persistent HBsAg. OHBI is the persistence of the viral genome in the livers of HbsAg(−)
individuals with low titers of HBV DNA. It has been reported that there is a high prevalence
of OHBI in patients with HIV, chronic HCV, or HCC, in those undergoing hemodialysis, in
those with criptogenic liver disease, in IV drug users, in those undergoing frequent blood
transfusions (hemophiliacs, etc.), and in blood donors [8-12]. OHBI can also be an additional
risk factor for HCC, alcoholic cirrhosis, nonalcoholic fatty liver disease, and progressive HIV
infection [13,14].
In our study, we investigated 44 patients with cryptogenic cirrhosis to evaluate the prevalence
of OHBI. OHBI was found in 4.5% in patients with cryptogenic cirrhosis. All OHBI cases
were older than 40 years of age. We reviewed several studies of the prevalence of OHBI were
reviewed. Honarkar et al. reported the prevelance of OHBI to be 22% in 35 cases of cirrhosis
using a tissue PCR method [17]. Kaviani et al. reported the prevalence of OHBI to be 1.9% in
104 cases of cryptogenic cirrhosis using a real-time PCR method [18]. Finally, in a study
conducted by Hashemi et al., OHBI frequency was identified to be 4% in patients with
cryptogenic cirrhosis [19]. OHBI prevalence has also been investigated by country.
Heringlake et al. did not detect any hidden viral infections in 162 German patients with
cryptogenic liver cirrhosis [20]. Fang et al. reported the prevalence of OHBI to be 22.8% in
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159 Chinese patients [21]. Song et al. examined 1,091 HbsAg(−) adults in the general Korean
population, finding an OHBI frequency of 0.7% [22].
Generally, the OHBI frequency is 0%–10% in people without liver disease, 11%–19% in
patients with chronic hepatitis, and 12%–61% in HCC patients [23-25]. In our study, all
patients were cirrhotic with an OHBI frequency of 4.5%. This finding is also consistent with
previously published results.
Serum HBV DNA levels are generally less than 200 IU/mL in patients with OHBI, which is
significantly lower than in HBsAg(+) individuals [26, 27]. In our study, HBV DNA levels of
two HBV DNA-positive cases were 22 and 2,345 IU/mL.
Depending on the HBV antibodies detected, OHBI may be seronegative [anti-HBc(−) and
anti-HBs(−)] or seropositive [anti-HBc(+) and/or anti-HBs(+)]. More than 80% of individuals
with OHBI are seropositive for these antigens. In this study, two patients who had detectable
HBV DNA levels were also positive for HBsAg. When all cases were considered, HBsAg
was found in 50% of patients. The presence of HBsAg is reported to be approximately 32%
in our country [28].
The presence of HBcAg can influence the prevalence of OHBI. In some studies, OHBI is
more common in patients who are positive for HBcAg than in patients who are negative for
HBcAg [11,26,29]. However, there is no significant correlation between OHBI and the
presence of either HBcAg or HBsAg [29]. In our study group, HBcAg positivity was 54%.
Two patients with detectable HBV DNA were also positive for HBcAg.
No other studies have investigated the frequency of OHBI in patients with cryptogenic
cirrhosis in our country. The prevalence of OHBI has been reported to be 0%–36.4% in
different patient groups and blood donors from Turkey. According to this study, OHBI was
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0%–36.4% in HCV(+) hemodialysis patients, 7% in HCV(−) hemodialysis patients, and
3.4%–19% in HBsAg(−) patients with detectable HBV DNA [28-33].
Despite its potential clinical importance, information on the effects of OHBI in patients with
chronic liver disease is limited, because the detection of HBV DNA may require a liver
biopsy, which is not routinely performed in clinical settings. In addition, many of the
previous studies addressing this issue only obtained small sample sizes, studied
heterogeneous populations, or used cross-sectional methods with variable detection
sensitivities for the analysis of liver or serum HBV DNA levels, which may explain
discrepancies in the effects of OHBI on patients with chronic liver disease.
Conclusion
Our study revealed the incidence of OHBI to be 4.5% in patients with cryptogenic cirrhosis.
OHBI can be a causative factor of cirrhosis and progressive liver decomposition in these
patients. We recommend that patients with cryptogenic cirrhosis be examined for OHBI. It is
also recommended that patients with OHBI be closely monitored and treated with highly
potent antiviral drugs.
STUDY LIMITATIONS
In all cases examined, liver biopsy specimens were unavailable; thus, we analyzed serum
samples using real-time PCR, which may underestimate the prevalence of OHBI. The number
of positive cases was thus insufficient to determine any