Vaccine 25 (2007) 3133–3136

Impact of hepatitis B vaccination in a highly endemic area of

south Italy and long-term duration of anti-HBs antibody
in two cohorts of vaccinated individuals
Giuliano Da Villa a , Luisa Romanò b , Alessio Sepe a , Raffaele Iorio c ,
Nunzio Paribello c , Alessandra Zappa b , Alessandro R. Zanetti b,∗
a Italian Institute for Prevention of Liver Diseases, Naples, Italy
b Department of Public Health-Microbiology-Virology, University of Milan,
Via C. Pascal 36, 20133 Milan, Italy
c ASL Napoli 3, Naples, Italy

Available online 22 January 2007

Abstract

The aims of this study were to evaluate the impact of hepatitis B vaccination on the changing pattern of HBV infection in a former
hyperendemic area (Afragola, South Italy), and to assess the long-term persistence of anti-HBs in two cohorts of individuals vaccinated as
infants 18 and 23 years ago. Our data shows a significant decline in the prevalence of hepatitis B virus (HBV) markers in the general population
from 1978 to 2006 (HBsAg: 13.4% versus 0.91%; anti-HBc: 66.9% versus 7.6%; p < 0.001). Data from two cohorts of vaccinees provides
further evidence regarding the long-term persistence of vaccine-induced anti-HBs. Data here reported indicates that the implementation of
vaccination had a great impact in the control and prevention of hepatitis B in Italy.
© 2007 Elsevier Ltd. All rights reserved.

Keywords: Hepatitis B vaccination; HBV prevalence; Long-term protection

1. Introduction To prevent and control hepatitis B in Italy, vaccination tar-

Between the 1960s and 1970s, Italy was considered a
country at intermediate to high level of hepatitis B endemic-
ity. According to the nationwide surveillance system (Istituto
Nazionale di Statistica, ISTAT), the reported incidence of
acute viral hepatitis at the beginning of the 1970s was 98
per 100,000 inhabitants and approximately 60% of the noti-
fied cases were estimated to be caused by the hepatitis B
virus (HBV) [1–2]. The incidence and prevalence of hep-
atitis B surface antigen (HBsAg) carriers were higher in
the southern regions of the country compared to the north
[3]. In particular, studies performed at the time showed
that the greater area of Naples was hyperendemic for HBV
[1–5].
∗ Corresponding author. Tel.: +39 02 50315126; fax: +39 02 50315120.
E-mail address: alessandro.zanetti@unimi.it (A.R. Zanetti).
geted to high risk individuals was first introduced on regional
scale in 1983 and became mandatory for all newborns and
adolescents nationwide in 1991 [6–8].
We evaluated the impact of hepatitis B vaccination in
changing the pattern of HBV infection in the general pop-
ulation of Afragola, a town with approximately 70,000
inhabitants, located in the greater area of Naples, in which the
general prevalence of HBsAg was higher than 13% in the late
1970s [4]. The aim of this study was to compare the preva-
lence of HBsAg and HBV core antigen antibody (anti-HBc)
in healthy individuals in 1978 to that observed in 2006, 23
years following the introduction of vaccination. The second
aim of this investigation was to study the long-term persis-
tence of anti-hepatitis B surface antigen antibody (anti-HBs)
in two cohorts of children vaccinated with anti-hepatitis B
plasma-derived vaccine in 1983 and with DNA recombinant
vaccine in 1988.

0264-410X/$ – see front matter © 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.vaccine.2007.01.044
3134 G. Da Villa et al. / Vaccine 25 (2007) 3133–3136

Table 1
Demographic characteristics of healthy individuals enrolled in 1978 and
2006 in Afragola, Naples
1978 2006
Total population 2690 1540
Male 1651 (61.4) 929 (60.3)
Female 1039 (38.6) 611 (39.7)
6–14 Years
Total 511 360
Male 316 (61.8) 237 (65.8)
Female 195 (38.2) 123 (34.2)
15–20 Years
Total 360 300
Fig. 1. Age-specific prevalence of HBsAg in healthy individuals tested in
Male 190 (52.8) 172 (57.3)
1978 and 2006.
Female 170 (47.2) 128 (42.7)
>25–58 Years
Total 1819 880 3. Statistical analysis
Male 1145 (62.9) 527 (59.9)
Female 674 (37.1) 353 (40.1) Differences in frequency were detected by the χ2 test, and
Values in parenthesis are in percent. p < 0.05 was considered significant.

4. Results
2. Materials and method
4.1. Declining prevalence of HBV markers
Between January and April 2006, we collected blood sam-
ples from 1540 subjects living in Afragola, and tested them Taken together, 361 out of 2690 (13.4%) individuals tested
for HBsAg and anti-HBc antibody. Individuals enrolled in in 1978 were positive for HBsAg and 66.9% (1800/2690) for
this study were comparable for age and gender distribution anti-HBc. In 2006, the overall prevalence of HBsAg and anti-
to those tested for the same markers of HBV infection in HBc dropped to 0.91% (14/1540) and to 7.6% (117/1540),
1978 (Table 1). All individuals under 25 years of age in 2006 respectively (p < 0.001). As shown in Figs. 1 and 2, in
had been vaccinated against hepatitis B as infants or ado- 1978 the age-specific prevalence of HBsAg and anti-HBc
lescents according to the Italian law, while vaccination was antibody, were 6.8% (35/511) and 56.9% (291/511) in indi-
administered to approximately 15% of those older than 25 viduals 6–14 years old; 10.3% (37/360) and 60% (216/360)
years. in those aged 15–20 years; 15.9% (289/1819) and 71.1%
In addition, we collected blood samples from two cohorts (1293/1819) in individuals 25–58 years old, respectively.
of young people vaccinated against hepatitis B as infants. In In 2006, the corresponding figures decreased dramati-
particular, cohort A included 88 individuals (48 males, 40 cally to 0.28% (1/360) and 0.56% (2/360); 0.33% (1/300)
females) born in 1983 to HBsAg positive mothers, who had and 1.7% (5/300); 1.4% (12/880) and 12.5% (110/880),
been given HBV hyper-immune gamma globulins (HBIG, respectively.
Biagini, Italy) at birth followed by three paediatric doses
of plasma-derived HB vaccine (Hevac-B Pasteur, France) at
months 1, 2 and 3, according to the schedule in use at that
time.
Cohort B included 112 individuals (83 males, 29 females)
born to HBsAg negative mothers who were administered
three paediatric doses of recombinant HB vaccine (Engerix
B, SKB, Belgium) in 1988, at 3, 5 and 11 months of age.
HBsAg and anti-HBc antibody were assayed by radio
immunoassays (Ausria and Corab, Abbott Labs, IL, USA)
in samples collected in 1978; the same markers and anti-HBs
antibody were assayed by IMx (Abbott Labs, IL, USA) in
samples collected in 2006.
Hepatitis B viral DNA was detected by the COBAS Taq-
Man HBV test (Roche Molecular Systems, NJ, USA) with a Fig. 2. Age-specific prevalence of anti-HBc in healthy individuals tested in
95% detection limit of 6.7 IU/ml. 1978 and 2006.
 G. Da Villa et al. / Vaccine 25 (2007) 3133–3136
Table 2
Prevalence of anti-HBs and anti-HBc antibodies in two cohorts of vaccinees
immunised as infants in 1983 (group A) and in 1988 (group B), as detected
in 2006
Group A (N = 88) Group B (N = 112)
Anti-HBs positive 83 (94.3) 84 (75)
Anti-HBc positive 0 3a (2.7)
Values in parenthesis are in percent.
a All were HBsAg negative, anti-HBs positive, and with undetectable HBV
DNA.
4.2. Duration of vaccine-induced anti-HBs
As shown in Table 2, 83 out of 88 (94.3%) individuals
born to HBsAg positive mothers (group A), who were given
HBIG and hepatitis B plasma-derived vaccine in the first few
months of life maintained protective concentrations of anti-
HBs (>10 mIU/ml) 23 years after immunisation. All these
vaccinees were HBsAg and anti-HBc antibody negative.
Concerning group B, 84 out of 112 (75%) subjects vac-
cinated with recombinant DNA vaccine retained protective
levels of anti-HBs over 18 years after the primary course of
vaccination. Three (2.7%) vaccinees had anti-HBc antibody
in absence of detectable HBsAg and HBV-DNA and with
negative history of clinically overt hepatitis B.
5. Discussion
Before the implementation of vaccination, Italy was con-
sidered a highly endemic country for hepatitis B, with higher
incidence and prevalence rates detected in the southern
regions compared to the north [1–5]. A large survey carried
out in 1978 in Afragola, a town of approximately 70,000
inhabitants located in the greater area of Naples, showed that
over 80% of the population had been infected with HBV, and
13.4% of these were HBsAg carriers [4]. Due to this high
endemicity, the Italian health authorities decided to imple-
ment a vaccination policy as soon as anti-hepatitis B vaccine
became available.
Thus, selective vaccination of individuals from high-risk
groups (e.g., health care workers, household contacts with
chronically infected people, intravenous drug users, some
categories of patients, babies born to HBsAg carrier moth-
ers, etc.) was initiated on regional scale in 1983, and universal
vaccination of all infants and 12 years old adolescents became
mandatory on national scale in 1991 [6–8].
Due to the high circulation of HBV in Afragola in
the late 1970s, a pilot study providing universal immuni-
sation to all newborns has been implemented since 1983
[9]. With the combination of such strategies of vaccination,
both at regional and national levels, the Italian population
aged between 0 and 25 years is now immunised against
hepatitis B.
As reported by the national surveillance system for acute
viral hepatitis (SEIEVA, Rome), the morbidity rate of acute
3135
viral hepatitis B per 100,000 inhabitants has dramatically
declined in the post-vaccination era, plummeting from 12 in
1985 to 1.3 in 2005 [10]. The decline was even more striking
in individuals aged between 15 and 24 years, in whom the
morbidity rate per 100,000 people dropped from 41 to 0.5 in
the same period of time. In addition, the prevalence of HBV
markers of infection has dropped to near zero among chil-
dren and young people during the last decades. For example,
the prevalence of anti-HBc antibody among Italian recruits,
which was 16.8% in 1981, decreased to less than 1% in 2001.
In accordance with this tendency, our survey carried out in
the former hyperendemic area of Afragola clearly shows a
highly significant decline in the prevalence of HBV mark-
ers of infection in the general population between 1978 and
2006 (HBsAg: 13.4 vs. 0.91%, p < 0.001; anti-HBc: 66.9
vs. 7.6%, p < 0.001). In particular, the prevalence of HBsAg
dropped from 8.2% (72/871) in 1978 to 0.3% (2/660) in
2006 in individuals under 20 years of age, confirming that
new generations of children and teenagers are growing up
with almost no markers of HBV infection. Of utmost impor-
tance, a significant decline in the prevalence of HBV markers
was also observed in adults who, in the majority of cases,
were not vaccinated. This fact may be partly attributed to the
herd immunity induced by the high coverage rate of children
immunisation resulting from our mandatory policy of vacci-
nation and in part to the general improvement in the standard
of living and hygiene, and the introduction of public health
measures, such as the use of universal precautions in medi-
cal settings, refining of blood screening together with social,
behavioural and demographic changes which took place in
the last few decades [11].
These findings are in agreement with those reported in
other high endemic countries where a substantial decrease
in the disease burden has followed the implementation of
long-standing policies of vaccination [12–15].
Finally, data from the two cohorts of individuals vacci-
nated as infants provides further evidence on the long-term
persistence of vaccine-induced anti-HBs as previously
reported [16]. It is hard to say whether the higher percent-
age of vaccinees found with protective levels of antibody
(≥10 mIU/ml) among those born to HBsAg positive mothers
compared to those born to HBsAg negative mothers (94.3%
versus 75%, p < 0.001) was due to a different immuno-
genicity of the two types of vaccines used (plasma-derived
versus recombinant DNA), to the different schedules of
immunisation or to a different degree of natural booster sec-
ondary to HBV exposure, conceivably higher in individuals
whose mothers were HBsAg carriers. Three (2.7%) vacci-
nees had markers of HBV infection (anti-HBc). Since they
all belonged to the group of individuals born to HBsAg neg-
ative mothers, we can infer that they acquired HBV infection
horizontally after immunisation. At enrolment, all these vac-
cinees were in good health and none had a history or presented
signs or symptoms of hepatitis B. Since the acquisition of
HBV in young age is often associated with the development
of a chronic carrier state, we can speculate that vaccina-
3136 G. Da Villa et al. / Vaccine 25 (2007) 3133–3136
tion might have partially protected these vaccinees whose
exposure to HBV resulted in subclinical infections.
This data further confirms that both plasma-derived and
DNA recombinant vaccines are highly immunogenic and can
confer long-term protection.
In conclusion, a body of evidence, including our finding,
indicates that the implementation of hepatitis B vaccination
had played a central role in the control and prevention of
hepatitis B in Italy.
Acknowledgement
We are extremely grateful to Mrs. C. Tagliacarne for skil-
ful technical assistance.
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