Elevated serum β2-microglobulin in individuals coinfected with hepatitis B and hepatitis D virus in a rural settings in Southwest Nigeria
Okoror et al. BMC Res Notes
Elevated serum β2-microglobulin in individuals coinfected with hepatitis B and hepatitis D virus in a rural settings in Southwest Nigeria
Lawrence Ehis Okoror 0
Ayodele Oluwaseun Ajayi 0
Oluwaseun Benjamin Ijalana 1
0 Department of Microbiology, Federal University , PMB 373, Oye-Ekiti, Ekiti State , Nigeria
1 Biosolution Technologies , Olusegun Obasanjo Way, Akure , Nigeria
Objective: Coinfection of hepatitis B virus (HBV) with hepatitis D virus (HDV) has being reported to increase severity of progression to hepatocellular carcinoma (HCC) and liver cirrhosis (LC). Beta microglobulin (2βM) which is present on the surfaces of blood cells in acceptable levels is a tumor marker which may become elevated in disease conditions. This study hence observed the prevalence of HBV and HDV coinfection in a rural population and their 2βM concentration. Results: Of the 368 samples, 66 (17.9%) were positive to hepatitis B surface antigen (HBsAg) and 33 (50%) were coinfected with HDV, 8 (2.1%) were monoinfected with HDV. 2βM concentration increased beyond the normal level in individuals coinfected with HBV and HDV as compared with the monoinfected individuals. Coinfection resulted in the increased concentration of 2βM in HBV and HDV coinfection and the likelihood of progression to HCC and LC may not be ruled out. Monoinfection with HDV also had high 2βM concentration but this is due to having being infected with a non-detected HBV or chronic infection in which HBV is clearing.
2βmicroglobulin; Hepatitis B; Hepatitis D; ELISA; Coinfection; Monoinfection
Hepatitis B virus (HBV) is known to infect approximately
350 million people [
] who are chronic carriers. Over 2
billion people are infected [
] which makes the virus
a major public health problem with over 1 million death
worldwide. HBV causes diseases of varying severity from
acute hepatitis  to chronic liver diseases, cirrhosis,
hepatocellular carcinoma to fulminant hepatitis [
HBV is a DNA virus in the family Hepadna viridae, it
is highly transmissible so that even a very small quantity
of blood is enough to transmit the virus. Major routes
of entry of the virus include wounds, abrasions, mucous
membranes, bites and scratching [
]. The virus
replicates quickly and more rapidly with very high titre in the
blood making it more infectious than HIV and
hepatitis C, because of this any contact of the blood with the
mucosal membranes has a high risk of contracting the
virus. The high titre of the virus in the blood makes it
easy for transmission via sharp objects like needle stick
injuries with a risk of 1–6% if the source of infection is
positive for HBV and negative for hepatitis e antigen and
22–40% if positive for both [
]. Risk of mother to child
has also been reported [
]. In Nigeria, HBV is endemic
with over 12% [
] seropositivity, though Nigerians are
said to be chronic carriers, reports vary from population
to population. Umolu et al.  reported a 5.8% rate in
blood donors in Benin City. In a study among university
students, Uneke et al. [
] reported 9% seropositivity.
Hepatitis D virus (HDV) is a defective RNA virus
which occurs only in HBsAg positive individuals [
as it could only multiply in the presence of HBV infected
cells. It has being reported to account for more severe
complications in HBV infected individuals leading to
rapid progression to cirrhosis, hepatocellular carcinoma
and death, unlike those infected with HBV alone [
]. The occurrence of HDV could be as a result of
super infection of chronic HBV infection or they could
co-occur in the acute infection [
In Nigeria information regarding HDV is scanty but
a few have reported HDV in chronic liver diseases [
Due to the complications likely to be produced by HBV
and HDV confection, more awareness needs to be
created in Nigeria and sub-Saharan Africa.
Beta-2-microglobulin is one of the major
histocompatibility complex class molecules on the cell surfaces of
all nucleated cells. It interacts and stabilizes the tertiary
structure of the major histocompatibility complex class 1
]. It is a member of the human leucocyte
antigen (HLA) where it is expressed in certain diseases
conditions such as tumor in elevated concentration in
the blood and detected in serum. Hepatitis D virus
infection in HBV infected individuals increases the chances of
development to hepatocellular carcinoma which may be
seen in the elevated level or concentration of 2βM.
We report the co-occurrence of HBV and HDV in
relation to their 2βM concentration in a rural population.
To our knowledge this is the first study in Nigeria to use
2-β-microglobulin to evaluate the state of HBV/HDV
Materials and methods
A total of 368 blood samples were collected from
apparently healthy individuals in a rural suburb in South West
Nigeria. Participants included in this study were either
HBV positive or HDV positive or both and must have
lived in the study population for the past 1 year. All
participant confirmed participation, parental consent was
obtained for children below the age of 18. HBV, HDV
mono-infected individuals served as controls.
Sera were separated from whole blood by centrifuging
at 3000 rpm and tested for hepatitis B surface antigen
(HBsAg), HDV and 2βM using the quantitative ELISA
technique for all the 3 parameters and test performed as
directed by the manufacturer of the ELISA kit (WKEA
Medical Supplies, China). Absorbance of all the samples
was read on a microplate reader (Thermomax, Molecular
Devices, USA) at 450 nm optical density. HBsAg, HDV
and 2βM concentration was determined using curves
from myassays ELISA analysis on line software for all
samples plotted against the standards.
Data analysis was done using the openepi which is an
online epidemiological software and SAS University
Edition determining seroprevalence of HDV in HBsAg
positive patients and expressed as percentage of the study
group/Pearson Chi square was used to determine the
association between the concentration of 2βM and HBV/
HDV infection. Linear correlation analysis was used to
determine the association between age and the
concentration of 2βM and HBV/HDV infection.
Of the 368 samples tested for HBsAg, 240 (65.2%) were
females while 128 (34.8%) were males, 66 (17.9%) were
positive to HBsAg of which 39 (59.1%) were females and
27 (40.9%) were males. A total of 33 (50%) individuals
were coinfected with HBV and HDV of which 8 (24.2%)
were males while 25 (75.8) were females. Eight
individuals (2.1%) were monoinfected with HDV from which
there were 4 (50%) males and 4 (50%) females (Table 1).
Age group 21–30 had the highest number of individuals
29 (12.9%), they also have the highest number of
individuals testing positive to HBV/HDV coinfection. Chi
square (χ2 = 271.9; p ≤ 0.0000001) shows that there is
significant difference between all results across the table.
The major finding in this study is the significant increase
in 2βM concentration in HBV/HDV coinfection (Fig. 1)
compared with HBsAg monoinfection and HDV
monoinfection. Yegane et al. [
] reported the role of 2βM in
monitoring chronic HBV infection where they estimated
the serum 2βM level in individuals with chronic HBV
infection and found that serum 2βM was higher in cases
of chronic HBV as compared to controls without HBV.
Though they did not work on HBV/HDV coinfection,
their observation was similar to the result obtained in
this study as 23.7% of patients monoinfected with HBV,
29% monoinfected with HDV and 39% coinfection have
2βM level well above the acceptable serum concentration
and high incidence of HDV in chronic HBV has being
reported. Yeagane et al. [
] then concluded that 2βM
is an indicator for monitoring chronic hepatitis which
is in agreement with this study as the trend of increase
suggests that coinfection of HBV/HDV have influenced
the increment in 2βM concentration in coinfected
individuals. This result is similar to a study by Casey et al.
] where they reported 64% of individuals were
coinfected with HBV and HDV and concluded that
coinfection of HBV with HDV was responsible for the frequent
hepatitis outbreak in a military setting. Wanch et al. [
reported the influence of age in elevated 2βM
concentration in patients infected with tuberculosis with the
older ages having higher concentration. Garcia-Garcia
et al. [
] also confirmed this report which was also seen
in this study as the older age groups had higher
concentration of 2βM which may be due to longer exposure to
HBV and becoming chronic as against the younger
population who has just recently been exposed. Figure 2 shows
individual 2βM concentration from the 101 positive
individuals for all the 3 parameters tested and age,
individuals from the older age groups had higher concentration of
2βM though prevalence was higher in mid ages had more
individuals showing elevated 2βM levels. There was more
elevated concentration of 2βM in individuals with
coinfection of HBV and HDV than those with monoinfection
in the mid ages. Though there were cases in this study
with high 2βM concentration in monoinfected
individuals, this was actually noticed in people with older ages
and could have been due to other cofounders as earlier
reported by Wanch et al. [
] and chronic HBV infection.
Figure 2 also reveals a positive correlation between age
and HBV and HDV monoinfection (r = 0.8912; CI 95%).
This also confirmed that as age increases 2βM
concentration increases in individuals coinfected with HBV/
HDV in significant proportion. This is also supported by
reports from Wanch et al. [
] where they reported that
age has an influence in the increase in 2βM
concentration in patients infected with tuberculosis with the older
ages having more prevalence. Garcia-Garcia et al. [
also confirmed this report. This study further reports that
HBV is still a major public health concern which case is
worsened by HDV infection especially in the rural areas
with limited health facilities and public health awareness.
Findings in this study when compared to earlier reports
shows that HBV is actually on the rise especially in the
rural areas. Umolu et al. [
] reported 5.8% in blood
donors while Uneke et al. [
] reported 9% seropositivity
among university students. These two reports dealt with
enlightened population where there are health facilities
and public health awareness. The chances of progression
to HCC and LC is more in the rural areas as evident in
this results from this study with the involvement of HDV
and the elevated 2βM. Though this is the first report
using 2βM to monitor HBV/HDV coinfection in Nigeria.
This study also reveals a rise in HBV infection (17.6%)
despite available vaccines against the virus in Nigeria
a rise from Umolu et al. [
] and Alao et al. [
separately reported over 12% HBsAg which was similar
to what was obtained in this study; however the trend is
increasing. Udeze et al. [
] reported a 9.0% HBsAg
positivity among university students in Nigeria a report which
is lower than the 17.9% from this study this can be said to
be due to different population studied. An Egyptian study
reported 21% prevalence [
]. This rise in prevalence in
HBV infection despite the vaccination campaign may
also be as a result of increasing population and also the
likely involvement of other immune suppressing diseases
not captured in this study. There may be a possibility of
reduction of HBV in the population in the near future
due to the vaccination campaign but there could be high
prevalence of HBV-associated LC and HCC due to long
latency of chronic hepatitis which eventually involve the
HDV coinfection which was earlier reported to be due
to chronic HBV infection leading to rapid progression to
HCC and LC.
Eight individuals (2.1%) were monoinfected with HDV
from which there were 4 (50%) males and 4 (50%) females
(Table 1) this we suggest have resulted from HBV chronic
infection. Though there is paucity of reports on
monoinfection with HDV, a few reports have it that
monoinfection with HDV may occur in individuals who had HBV
infection earlier and may lead to serious sequelae where
it becomes persistence and develops to hepatocellular
carcinoma more severe and quicker than a coinfection
with HBV since it may have resulted as a result of chronic
HBsAg. Katja et al. [
] inoculated mice with HDV and
concluded that HDV could persist even in humans for
over 8 weeks before being rescued by HBV and thereby
develop more virulence. However, mechanism for HDV
monoinfection is still being investigated.
This study confirms elevated 2βM concentration in HBV/
HDV coinfected individuals. It is also suggested that 2βM
concentration could also be used as a marker to
complications that could arise due to HBV/HDV coinfection.
We also conclude that there is little awareness campaign
in the rural areas as to danger posed by HBV and or
HBV/HDV coinfection. Therefore awareness campaign
in the rural communities must be stepped up in order to
reduce the rising HBV/HDV infection.
• We were unable to assay for other debilitating
diseases that could lead to immunocompromise and
then enhance HBV infection.
• We did not confirm other diseases that could cause
increase in 2βM concentration but this did not
undermine our results as we have HBV and HDV
monoinfection served as controls.
• Confounding factors like chemicals not covered in
this study are likely to cause elevated 2βM though
there influence were taking care of by controls.
2βM: 2-beta microglobulin; HBV: hepatitis B virus; HDV: hepatitis D virus;
HCC: hepato cellular carcinoma; LC: liver cirrhosis; HBsAg: hepatitis B surface
antigen; DNA: deoxyribonucleic acid; RNA: ribonucleic acid; HLA: human
leucocyte antigen; ELISA: enzyme linked immunosorbent assay.
LEO conceived the study was supervised sample collection and all through
the laboratory work till writing the manuscript. AO jointly carried laboratory
work as well as sample collection. He was also involved in the peer review
process. OBI did a lot of laboratory work and was also involved in the final
build up of the manuscript he also contributed in the peer review process, he
contributed immensely during the design of the project. All authors read and
approved the final manuscript.
All our undergraduate students who participated in this study are well
appreciated as well as management of Joseph Ayo Babalola University for providing
the foundation for the study.
The authors declare that they have no competing interests.
Availability of data and materials
Consent to publish
This study was approved by the ethical committee/research committee of
Joseph Ayo Babalola University, whose medical centre the study was started.
All participants agreed to participate in the study on the condition of
anonymity and verbal consent was obtained, parental consent was obtained for
children below the age of 18.
Authors funded the research on their own.
Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.
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