Lessons learnt over two decades of vaccination against hepatitis B in Italy.

This article reviews topics covered and discussed at the Meeting: "Vaccini e vaccinazioni. Migliorare l'oggi e preparare il domani", held in Genoa, Italy, on 12 September 2014. Data presented at the meeting, clearly showed that: 1) hepatitis B vaccination can confer long-term protection and there is no need for booster in immunocompetent vaccinees; 2) vaccination is highly effective in protecting population from clinical acute or chronic HBV infections, including hepatocellular carcinoma; 3) children vaccinated as infants with hexavalent vaccines maintain immunological memory 5 years after priming, but further studies are needed to assess whether immunity persists during the adolescence and adulthood when risk of exposure to HBV becomes higher; 4) the emergence of vaccine-escape mutants and Pol-gene mutants during antiviral therapy - which can result in changes in the S-gene - is of some concern, but at present there is no evidence that such mutants may pose a threat to the established programs of vaccination.

Hepatitis E: a puzzling double-faced disease.

Viral hepatitis type E is highly endemic in many developing countries, where large water-borne epidemics caused by viral genotype 1 and - to a lesser degree - by genotype 2 cyclically occur, resulting in high morbidity and mortality, especially among pregnant women. In developed countries, the disease is usually diagnosed in travelers coming back from endemic countries, but an increasing number of sporadic locally acquired hepatitis cases caused by genotype 3 and 4 have recently been reported. The wide-spread distribution of HEV3 and HEV4 in domestic pigs, wild boars, deer, as well as in other mammals, suggests that infections caused by these genotypes may have a zoonotic source. HEV3 infection can evolve to chronic infection in immunosuppressed patients; in addition, it may be associated with neurological disorders and extrahepatic manifestations. Two recently developed recombinant vaccines have proved to be safe and effective. One of such vaccines has recently been licensed for use in China.

The state of hepatitis B and C in Europe: report from the hepatitis B and C summit conference*.

Worldwide, the hepatitis B virus (HBV) and the hepatitis C virus (HCV) cause, respectively, 600,000 and 350,000 deaths each year. Viral hepatitis is the leading cause of cirrhosis and liver cancer, which in turn ranks as the third cause of cancer death worldwide. Within the WHO European region, approximately 14 million people are chronically infected with HBV, and nine million people are chronically infected with HCV. Lack of reliable epidemiological data on HBV and HCV is one of the biggest hurdles to advancing policy. Risk groups such as migrants and injecting drug users (IDU) tend to be under-represented in existing prevalence studies; thus, targeted surveillance is urgently needed to correctly estimate the burden of HBV and HCV. The most effective means of prevention against HBV is vaccination, and most European Union (EU) countries have universal vaccination programmes. For both HBV and HCV, screening of individuals who present a high risk of contracting the virus is critical given the asymptomatic, and thereby silent, nature of disease. Screening of migrants and IDUs has been shown to be effective and potentially cost-effective. There have been significant advances in the treatment of HCV and HBV in recent years, but health care professionals remain poorly aware of treatment options. Greater professional training is needed on the management of hepatitis including the treatment of liver cancer to encourage adherence to guidelines and offer patients the best possible outcomes. Viral hepatitis knows no borders. EU Member States, guided by the EU, need to work in a concerted manner to implement lasting, effective policies and programmes and make tackling viral hepatitis a public health priority.

Early co-circulation of different clades of influenza A/H1N1v pandemic virus in northern Italy.

INTRODUCTION: The spatial diffusion over time of pandemic influenza A/HINI virus (A/HIN1v) was surveyed in Northern Italy (nearly 10 million inhabitants)from April to December 2009, and the molecular characteristics of circulating viruses were analyzed to identify the appearance of drift variants. About 45% of analyzed samples were laboratory-confirmed cases of A/HINlv. Sporadic cases occurred until the middle of June 2009, then, case numbers began to increase delineating distinct epidemiological phases of viral circulation. METHODS: RNA was extracted using RNeasy Mini kit (QIAGEN GmbH, Germany). Virological diagnosis of A/HINlv infection was carried out by real-time RT-PCR assay. Sequence analysis of hemagglutinin (HA) gene was performed through a RT-PCR assay specific for a 995 bp fragment (nt. 64-1,058) in the HAl domain. The nucleotide sequences were obtained by automated DNA sequencing. The HAl sequences were aligned with other sequences collected from GenBank database by ClustalX software. The multiple sequence alignment was used to perform a basic phylogenetic analysis and a phylogenetic tree from HA sequences was constructed. RESULTS: The HA gene sequences ofA/HINlv analyzed segregated into three genetically distinct clades and were characterized by the appearance of amino acid variations that were progressively fixed in the field viral population under scrutiny. CONCLUSIONS: These data suggest an early co-circulation of genetically distinct A/HNINv variants and emphasize the importance of a close molecular surveillance to detect rapidly the spread of new viral variants and to define their epidemiological impact.

Emerging and re-emerging infections at the turn of the millennium.

After World War II, mankind believed that infectious diseases were on the way to being defeated. Unfortunately, they still are the second worldwide cause of death. Globalization changes promote the emergence of new infections and pandemics; international deliveries and travelling facilitate the dissemination of infectious agents; man-induced environmental changes create new opportunities for contacts between species, leading to infections in aberrant hosts, including man; global warming enables insects, a major vector of pathogens, to thrive in more countries. The main pandemics have been caused by viruses, such as HIV and novel subtypes of influenza viruses. In addition, prion proteins are a threat. The transmission of the Creutzfeld Jakob disease variant through blood transfusion and the recent discovery of prion protein in the spleen of a haemophilia patient are a matter of further concern. The end of the war against infectious diseases is not in sight. Mankind's battle with pathogens has lasted millennia and is destined to continue.

A 2010 update on occult hepatitis B infection.

Occult hepatitis B virus infection is a challenging issue whose virological and clinical relevance has been a source of long-lasting debate. By definition, OBI is characterized by the persistence of HBV-DNA in the liver tissue (and in some cases also in the serum) in absence of HBsAg. According to the HBV serological profile, OBI may be antibody (anti-HBc alone or together with anti-HBs) positive (seropositive OBI) or antibody negative (seronegative OBI). OBI is a complex biological entity with possible relevant clinical implications, mainly related to the intrahepatic persistence of viral cccDNA and to a strong suppression of viral replication and gene expression. Clinical observations suggest that OBI carriers may be a source of HBV transmission through blood transfusion or orthotopic liver transplantation (OLT). The state of suppression of viral replication and gene expression may be discontinued when an immunosuppressive status occurs, leading to typical hepatitis B with severe - and some times - fulminant course. The long-lasting persistence of the virus in the liver may provoke a very mild but continuing necro-inflammation that (if other causes of liver damage cohexist) may contribute over time to the progression of the chronic liver damage towards cirrhosis. In addition, OBI is supposed to be an important risk factor to HCC development since it maintains the pro-oncogenic properties typical of the overt infection.

Hepatitis C virus outbreak in a haemodialysis unit: learning from failures.

The investigation of an outbreak of hepatitis C virus in an Italian haemodialysis (HD) centre showed that three patients acquired infection with the same strain, affecting a chronically hepatitis C virus (HCV)-infected patient receiving HD in the same room and during the same shifts. Through our observational analysis many possible modes of transmission were identified, but none could be definitively identified as the route of HCV spread in this small cluster. This outbreak confirms that repeated opportunities for nosocomial HCV transmission may occur among HD patients due to several breaches in the standard precautions for bloodborne infections by healthcare staff.

Long-term immunogenicity of a plasma-derived hepatitis B vaccine in HIV seropositive and HIV seronegative hemophiliacs.

Short-term studies indicate that hepatitis B vaccines are safe and satisfactorily immunogenic in hemophiliacs. The duration of immunity in these immunocompromised patients, however, is not known. To determine this, we studied 78 hemophiliacs prospectively 2, 3, and 4 years after the initial vaccination with a plasma-derived vaccine given as three monthly injections followed by a fourth booster injection at month 14. The duration of immunity clearly depended on whether the patients were infected with the human immunodeficiency virus (HIV). In HIV seronegative hemophiliacs (n = 67), there was a progressive decline in titers of antibody to the hepatitis B surface antigen (anti-HBs), but antibody was still detectable 4 years later in all of them. From the curves of decline of antibody titers, it appears that there is no need to revaccinate patients for at least 5 to 6 years. The HIV seropositive hemophiliacs (n = 11) not only started from much lower anti-HBs titers, but 5 of 11 lost anti-HBs. None of the 45 patients treated with concentrates during the postvaccination period developed serologic signs of hepatitis B, even though 6 of them had come into contact with live or inactivated hepatitis B virus as shown by the occurrence of spontaneous anamnestic antibody responses. This vaccine and schedule of vaccination afford a prolonged duration of immunity in HIV seronegative hemophiliacs, but HIV seropositive hemophiliacs have a risk of losing immunity early.

A solid-phase enzyme immunoassay (EIA) for detection of HBeAg and anti-HBe.

A sensitive, reproducible and easily performed enzyme immunoassay (EIA) based on a sandwich technique is described for serological detection of HBeAg and anti-HBe. EIA appears to be 600 times more sensitive than immunodiffusion and counterimmunoelectrophoresis and its performance compares well with available radioimmunoassays.

Antibody to hepatitis C virus after a vapour-heated factor VIII concentrate. The Study Group of the Fondazione dell'Emofilia.

To evaluate whether or not clotting factor concentrates exposed to virucidal procedures transmitted hepatitis C, sera obtained in 1984-1986 from 27 previously untreated hemophiliacs infused with a vapour-heated factor VIII concentrate were tested retrospectively for the antibody to the hepatitis C virus (anti-HCV). A 2-year-old hemophiliac, negative for anti-HCV before administration of concentrate, seroconverted at week 12 and remained anti-HCV positive thereafter. Both his parents were anti-HCV negative and he had no other household contact. The patient had also become HBsAg positive at week 8 and had at the same time a marked elevation of alanine aminotransferase. His double infection with the hepatitis B and C viruses indicates that hot vapour was not completely effective in inactivating these viruses.

Effect of ursodeoxycholic acid administration in patients with acute viral hepatitis: a pilot study.

BACKGROUND: Ursodeoxycholic acid (UDCA) is able to improve biochemical markers of cholestasis, with a parallel decrease in transaminases, in various cholestatic liver diseases. AIM: To evaluate the effects of UDCA administration on acute viral hepatitis-related cholestasis and the course of acute viral hepatitis. METHODS: Seventy-nine consecutive patients with acute viral hepatitis (HBV: 43, HCV: 11, HAV: 15, HEV: 3, Non A-E: 7) were randomized to receive either UDCA for 3 weeks or no treatment. Liver biochemistry and serum bile acid determinations were run at weekly intervals. RESULTS: No significant differences were observed in mean percentage decreases in transaminases between treated and untreated patients. By contrast, cholestatic indexes decreased significantly more quickly in patients treated with UDCA than in controls, and this effect was more evident in patients with increasing alanine transaminase levels at admission. After a peak at the end of the first week of therapy, serum levels of conjugated ursodeoxycholic acid (CUDCA) showed a gradual decrease. Conjugated cholic acid (CCA) and chenodeoxycholic acid (CCDCA) showed a progressive decrease with the resolution of viral hepatitis, but no influence of UDCA administration was observed. CONCLUSIONS: Our study demonstrates that UDCA significantly improves cholestatic indices in patients with acute viral hepatitis, but this effect does not seem to affect the course of the illness.

Viral antigens and antibodies in hepatitis B infection.

Hepatitis B virus antigens and antibodies were detected in the sera of acute and persistently infected patients. Evidence of active virus replication was confined to immediately before or during the initial detection of hepatitis B surface antigen during acute hepatitis B. Hepatitis B core antibody appeared during the period of antigenaemia and preceded recovery. Hepatitis B e antigen was dound in a proportion of sera which contained significant levels of virus particles. In contrast, all sera containing hepatitis B virus particles from persistently infected patients treated by maintenance haemodialysis also contained the e antigen. Among a group of 50 persistent carriers of hepatitis B virus, significant levels of virus production occurred in the presence of antibody to e antigen. In addition, evidence of exposure to hepatitis B virus was found among 3% of blood donors in whose sera the surface antigen was not detected by radioimmunoassay. The significance of these findings is discussed in relation to the aetiology of hepatitis type B.

Is HCV transmitted by the vertical/perinatal route?

Hepatitis type C is the major aetiological cause of both parenterally transmitted and cryptogenic, sporadic or community acquired nonAnonB hepatitis. The lack of known parenteral risk factors in a consistent number of cases with nonAnonB hepatitis has stimulated the search of other possible modes of viral transmission. The aim of this report is to review the evidence both for and against vertical/perinatal transmission of HCV from anti-HCV positive mothers to infants.

Localization of hepatitis C virus antigen(s) by immunohistochemistry on fixed-embedded liver tissue.

Using two sources of primary antibodies, we immunohistochemically stained hepatitis C virus-related antigen(s) on fixed-embedded liver specimens. These antigens were localized in the cytoplasm of hepatocytes. The results obtained serologically correlated well with immunohistochemistry.

Screening of pregnant women and hepatitis B prophylaxis in newborns.

Prevention of perinatal hepatitis B includes: (1) screening of pregnant women for hepatitis B surface antigen, and (2) immunoprophylaxis of babies at risk. HBIG treatment seems to be of some efficacy in preventing the HBsAg carrier state while it permits passive active immunization to occur. The disadvantage of HBIG is that it confers only temporary immunity. Therefore, if infection does not occur, babies will still be susceptible to the virus when passively administered anti-HBs will no longer be circulating. On the other hand, vaccine provides a long term but not immediate protection. Therefore the ideal approach in post-exposure prophylaxis is a combination of passive plus active immunization. The aim is to provide an immediate protection, with the HBIG, and a long term immunity, with the vaccine, to babies born to HBsAg carrier mothers.

Status and significance of testing for hepatitis B surface antigen and surface antibody.

Following Blumberg's discovery of hepatitis B surface antigen (HBsAg), many attempts have been made to develop several in vitro diagnostic techniques for the detection of this antigen and its homologous antibody. The two-dimensional micro-Ouchterlony immunodiffusion has been the first technique used, rapidly replaced by procedures of increasing sensitivity characterized as second-generation and the currently available third-phase tests which include radioimmunoassay (RIA), reverse passive haemagglutination (RPHA), reverse passive latex agglutination (RPLA) and enzyme immunoassay (EIA). Among these, RIA appears to be the most sensitive and specific, whereas EIA, RPHA and RPLA have the advantage of long shelf-life of stable reagents, no need for sophisticated and expensive equipment and no hazard associated with the handling of radioactive isotopes. Moreover, the sensitivity of EIA should increase by objective reading with a colorimeter. The most sensitive method for the detection of surface antibody (anti-HBs) is again RIA, whereas passive haemagglutination (PHA) had the advantage of providing titres. Finally EIA, based on inhibition of a known amount of HBsAg, has at least the same sensitivity as PHA, but has the advantage that reagents are more stable and that it permits screening for both HBsAg and anti-HBs with the same reagents at the same time. The application of these highly sensitive techniques for the detection of HBsAg and anti-HBs has resulted in a consistent reduction in the incidence of post-transfusion hepatitis type B and in a better understanding of the aetiology, epidemiology and natural history of this infection.

Hepatitis type C: modes of transmission and preventive measures.

There is general agreement that the hepatitis C virus is efficiently transmitted parenterally, while data on viral transmission from mothers to babies or by sexual or non-sexual household contact are conflicting. In Italy, hepatitis C accounts for approximately 20% of all cases of acute viral hepatitis. It is more frequent among young persons (15-24 years old), among those living in the southern part of the country or on the islands, and in males than in females (3:1). In recent years the incidence rate has dropped and significant changes in the HCV transmission patterns have been observed. As a consequence of the implementation of anti-HCV screening and the introduction of increasing restrictions on donor eligibility, the proportion of cases arising from blood and blood component transfusions has significantly declined. A candidate vaccine to protect against hepatitis C virus is now under development.

Immune response to hepatitis B vaccine in staff and patients in renal dialysis units.

Anti-HBs response was detected in 96 per cent of staff members in three haemodialysis units after three 20 microgram doses of hepatitis B vaccine and in 82 per cent of adult patients treated with three 40 microgram doses. The percentage of responders and levels of antibody remained unchanged at 12 months from the beginning of the trial. Three out of six children injected with three 20 microgram doses in a paediatric haemodialysis unit remained free from markers of HBV infection and had high levels of anti-HBs after the second dose of vaccine. The other three children who developed serological markers of HBV infection seroconverted to anti-HBc within six months from the first dose and, in one of them, antigenaemia at three and four months was detected.

Outcome of mother to infant acquired GBV-C/HGV infection.

Twelve children born to hepatitis C virus antibody GBV-C/HGV RNA positive mothers who acquired GBV-C/HGV infection by the vertical or perinatal route were studied. Most (91%) were persistently GBV-C/HGV RNA positive up to 12 months of age. Four out of six cases who acquired GBV-C/HGV alone had normal alanine amino transferase activities. Long lasting evidence of hepatocellular injury was detected only in children with GBV-C/HGV and hepatitis C virus and HIV coinfection.