Elisabetta Madonna

Hepatitis C virus infection in an endemic area of Southern Italy 14 years later: Evidence for a vanishing infection

BACKGROUND In a 1996 survey, prevalence of hepatitis C virus antibodies (anti-HCV) in a southern Italian town was 12.6%. AIMS To identify changes in the epidemiology of hepatitis C virus (HCV) infection. METHODS Anti-HCV, HCV-RNA (PCR, detection limit 15 IU/mL), HCV genotype (Innolipa). Were performed in a random 1:4 systematic sample of the general population. Multiple logistic regression analysis was used to estimate factors independently associated with the likelihood of anti-HCV positivity. RESULTS Of 1012 subjects, 58 (5.7%) were anti-HCV-positive, compared to 12.6% 14 years earlier. Prevalence was 0.4% in individuals <30 years old and 31.8% in those ≥ 70 years old. Among 139 HCV-negative in 1996 re-sampled in 2010, only one had seroconverted (incidence: 0.05 × 100 persons/year). Alanine transaminase levels were elevated in 8 (13.8%). HCV-RNA was detected by PCR in 46.5% anti-HCV-positive subjects. In 2010 59% were genotype 2-infected, in 1996 50.7% genotype 1-infected. Previous use of non-disposable glass syringes was a strong independent predictor (OR 3.2; CI 95%=1.4-7.3). CONCLUSION Epidemiology of HCV infection in an endemic area of south Italy has changed over 14 years, now largely confined to the oldest age group; this seems to be due to the disappearance of its past main mode of transmission, namely the use of glass syringes.

Seroprevalence and anti‐HEV persistence in the general population of the Republic of San Marino

The prevalence of anti‐HEV was assessed in 2,233 subjects aged 20–79 years in the Republic of San Marino in the years 1990–1991. The sera were tested by ELISA and further confirmed by Western blot (WB) analysis. The overall anti‐HEV prevalence was 1.5%. A significant trend by age was observed. Anti‐HEV prevalence was 0.6% in subjects <30 years and 3.3% in those older than 70 years of age. Family size larger than four persons (OR = 3.8; 95% CI = 1.8–13.2) was the sole independent predictor of anti‐HEV positivity in the multivariate analysis. Anti‐HAV and anti‐HEV prevalences did not show a parallel trend by age. No association was found either between hepatitis E virus (HEV) or hepatitis C virus (HCV) infections. Follow‐up samples 5 years apart were available for 38 out of 54 (70%) anti‐HEV ELISA‐positive subjects. Eight out of 22 (37%) WB‐confirmed anti‐HEV–positive subjects were still anti‐HEV–positive after 5 years. However, anti‐HEV remained positive in all but two (75%) of the subjects with WB‐confirmed ELISA positivity value of S/CO ≥ 2 (cutoff 1.2), but in only 2 out of the 14 subjects (14%) with a WB‐con‐ firmed ELISA positivity value of S/CO < 2 (P < 0.005). None of the 16 subjects ELISA‐positive but not WB‐confirmed was anti‐HEV–positive 5 years apart. Therefore, only a relative proportion of subjects once infected with HEV maintain for at least 5 years anti‐HEV antibodies. J. Med. Virol. 58:49–53, 1999.

A cohort study to evaluate persistence of hepatitis B immunogenicity after administration of hexavalent vaccines

BackgroundIn 2001, two hexavalent vaccines were licensed in Italy (Hexavac®, Infanrix Hexa®), and since 2002 were extensively used for primary immunization in the first year of life (at 3, 5, 11/12 months of age). In 2005, the market authorization of Hexavac® was precautionary suspended by EMEA, because of doubts on long-term protection against hepatitis B virus. The objectives of this study were to evaluate the persistence of antibodies to anti-HBs, in children in the third year of life, and to investigate the response to a booster dose of hepatitis B vaccine.MethodsParticipant children were enrolled concomitantly with the offering of anti-polio booster dose, in the third year of life. Anti-HBs titers were determined on capillary blood samples. A booster dose of hepatitis B vaccine was administered to children with anti-HBs titers < 10 mIU/ml, with the monovalent precursor product of the previously received hexavalent vaccine. HBsAb titers were tested again one month after the booster.ResultsSera from 113 children previously vaccinated with Hexavac®, and from 124 vaccinated with Infanrix Hexa® were tested for anti-HBs. Titers were ≥ 10 mIU/ml in 69% and 96% (p < 0,0001) respectively. The proportion of children with titers ≥ 100 mIU/ml did also significantly differ among groups (27% and 78%; p < 0,0001).Post-booster, 93% of children achieved titers ≥ 10 mIU/ml, with no significant difference by vaccine group.DiscussionFifteen months after third dose administration, a significant difference in anti-HBs titers was noted in the two vaccine groups considered. Monovalent hepatitis B vaccine administration in 3-year old children induced a proper booster response, confirming that immunologic memory persists in children with anti-HBs titers < 10 mIU/ml. However, long-term persistence of HBV protection after hexavalent vaccines administration should be further evaluated over time.

Hepatitis B Virus Infection in Health Care Workers in Albania: a Country still Highly Endemic for HBV Infection

Background:Health care workers (HCW) have an elevated risk of acquiring and transmitting parenteral infections. The aim of this study was to evaluate the prevalence of hepatitis B virus (HBV) and hepatitis C virus (HCV) markers with the final goal to encourage HBV vaccination of the non-immune Albanian HCW.Methods:Among 480 HCW enrolled, 92 were physicians, 246 were nurses/techniques, 120 were auxiliary workers and 22 were office workers.Results:The HBsAg, anti-HBc and anti-HCV prevalence were 8.1%, 70% and 0.6%, respectively. The highest (11.4%) HBsAg prevalence was observed in the youngest age group (20–30 years of age). High HBsAg prevalence (7.2–7.5%) was detected also in age groups above 30 years. The highest HBsAg prevalence (12.6%) was found in the auxiliaries. The anti-HBc prevalence increased significantly with age from 59% in HCWs younger than 39 years to 87% among those older than 50 years. After adjustments for different job categories, age older than 40 years remained independently associated with anti-HBc positivity (OR = 2.9; 95% CI 1.9–4.6) and inversely associated with the lack of HBV immunity or infection markers (OR = 0.4; 95% CI 0.2–7). Of 142 HBsAg negative and/or anti-HBc Ab negative sera, 28 (20%) tested positive for anti-HBs. The 114 remaining individuals with no HBV infection or immunity markers were vaccinated against HBV infection.Conclusions:A high HBV infection rate and low HBV vaccination coverage were found in Albanian HCW. Albania is a Mediterranean country still highly endemic for HBV infection and new strategies to promote HBV vaccination are to be adopted.

Seroprevalence of hepatitis E virus (HEV) antibody and the possible association with chronic liver disease: a case-control study in Albania

A case-control study involving 109 in-patients with chronic liver disease and 190 in-patients with no apparent liver disease was conducted to evaluate the seroprevalence of anti-HEV antibodies and the possible association with chronic liver disease. Among cases, the anti-HEV prevalence was 36.6% which increased significantly by age; among controls, the prevalence was 12.1% (P<0.05) and was similar among age groups <60 years. Among cases, aged >50 years (OR 4.0, 95% CI 1.4-11) and the presence of end stage liver disease (ESLD) (OR 4.3, 95% CI 1.4-12.8) were associated independently with anti-HEV positivity. The mean optical density, determined by anti-HEV immunoenzymatic test, was significantly higher among patients with ESLD, compared to the other patients. These results indicate that there is a high seroprevalence of anti-HEV in patients with chronic liver disease and a possible association between HEV infection and/or anti-HEV production and advanced stage chronic liver disease.

High prevalence of anti-hepatitis E virus antibodies among blood donors in central Italy, February to March 2014

Prevalence of anti-hepatitis E virus (HEV) antibodies is highly variable in developed countries, which seems partly due to differences in assay sensitivity. Using validated sensitive assays, we tested 313 blood donors attending a hospital transfusion unit in central Italy in January and February 2014 for anti-HEV IgG and IgM and HEV RNA. Data on HEV exposure were collected from all donors. Overall anti-HEV IgG prevalence was 49% (153/313). Eating raw dried pig-liver sausage was the only independent predictor of HEV infection (adjusted prevalence rate ratio = 2.14; 95% confidence interval: 1.23-3.74). Three donors were positive for either anti-HEV IgM (n = 2; 0.6%) or HEV RNA (n = 2; 0.6%); they were completely asymptomatic, without alanine aminotransferase (ALT) abnormalities. Of the two HEV RNA-positive donors (both harbouring genotype 3), one was anti-HEV IgG- and IgM-positive, the other was anti-HEV IgG- and IgM-negative. The third donor was positive for anti-HEV IgG and IgM but HEV RNA-negative. HEV infection is therefore hyperendemic among blood donors (80% men 18-64 years-old) from central Italy and associated with local dietary habits. Nearly 1% of donors have acute or recent infection, implying potential transmission to blood recipients. Neither ALT nor anti-HEV IgM testing seems useful to prevent transfusion-transmitted HEV infection.

Molecular characterisation of human hepatitis E virus from Italy: comparative analysis of five reverse transcription-PCR assays

BackgroundHepatitis E (HEV) is an important public-health concern as a major cause of enterically transmitted hepatitis worldwide. In industrialised countries it is considered rare, and largely confined to travellers returning from endemic areas. However, autochthonous (locally acquired) HEV infection is also emerging in these regions. The infection is caused by different genotypes, depending on whether it is travel-related or autochthonous. Conventional RT-PCR followed by sequencing of PCR products can identify HEV genotype and, depending on the region, the subtype, thus helping in defining the origin of infection and tracing the source of contamination.MethodsWe re-analysed a collection of serum samples previously confirmed as hepatitis E positive by anti-HEV IgM and IgG assays as well as by Real-Time PCR, with the aim to compare the performances of five different broad range RT-PCR assays that could be provided for molecular characterisation of HEV. This approach is certainly valuable to investigate the molecular epidemiology of acute hepatitis E in countries where co-circulation of different genotypes occurs, like Italy.ResultsSamples were analyzed by five assays targeting the ORF1, ORF2, and ORF2/3 regions. The sensitivity of these assays varied significantly, depending on the target region. Only 46% of samples tested positive by nested PCR; moreover, no single method was able to detect all positive samples. Most sequences originated from patients who had travelled to endemic areas (genotype 1), while the minority originated from Italian patients with no travel history (genotype 3).ConclusionBroad range methods for molecular characterization of HEV still need to be improved to detect all circulating strains.

Diagnosis of HEV infection by serological and real-time PCR assays: a study on acute non-A-C hepatitis collected from 2004 to 2010 in Italy.

BackgroundThe impact of hepatitis E in developed countries, like Italy, still requires a clear definition. In the present study, we evaluated HEV infection in patients with acute non-A-C hepatitis by an approach comparing data from Real-time PCR and serological assays.MethodsIn a first analysis, sera from 52 patients hospitalized with a diagnosis of acute viral non-A-C hepatitis in Italy were tested by in-house Real-Time PCR assay for identification of Hepatitis E Virus (HEV) RNA and by anti-HEV IgM and IgG assays. In a subsequent analysis, selected samples were evaluated by additional IgM tests to confirm diagnosis.ResultsAmong the 52 samples, 21 showed positive results for all three markers (IgM, IgG and HEV RNA). One patient showed HEV RNA as single marker. Uncertain results were found in 8 samples while the remaining 22 were negative for all markers. Further analysis of the 8 undefined samples by additional IgM tests confirmed HEV infection in 1 patient. Overall, acute HEV infections were reliably identified in 23 (44.2%) out of 52 patients.ConclusionsIn the present paper, we performed a study evaluating HEV infection in 52 sporadic non-A-C acute hepatitis cases. All samples were collected from 2004 to 2010 in Italy. By a diagnostic strategy based on genomic and serological assays we identified HEV infections in 23 out of 52 patients (44.2%), a percentage higher than previous estimates. Thus, the actual impact of HEV infections in Italy needs to be further evaluated on a national scale by a diagnostic strategy based on multiple and last generation assays.

Key Role of Sequencing to Trace Hepatitis A Viruses Circulating in Italy During a Large Multi-Country European Foodborne Outbreak in 2013

Background Foodborne Hepatitis A Virus (HAV) outbreaks are being recognized as an emerging public health problem in industrialized countries. In 2013 three foodborne HAV outbreaks occurred in Europe and one in USA. During the largest of the three European outbreaks, most cases occurred in Italy (>1,200 cases as of March 31, 2014). A national Task Force was established at the beginning of the outbreak by the Ministry of Health. Mixed frozen berries were early demonstrated to be the source of infection by the identity of viral sequences in patients and in food. In the present study the molecular characterization of HAV isolates from 355 Italian cases is reported. Methods Molecular characterization was carried out by PCR/sequencing (VP1/2A region), comparison with reference strains and phylogenetic analysis. Results A unique strain was responsible for most characterized cases (235/355, 66.1%). Molecular data had a key role in tracing this outbreak, allowing 110 out of the 235 outbreak cases (46.8%) to be recognized in absence of any other link. The data also showed background circulation of further unrelated strains, both autochthonous and travel related, whose sequence comparison highlighted minor outbreaks and small clusters, most of them unrecognized on the basis of epidemiological data. Phylogenetic analysis showed most isolates from travel related cases clustering with reference strains originating from the same geographical area of travel. Conclusions In conclusion, the study documents, in a real outbreak context, the crucial role of molecular analysis in investigating an old but re-emerging pathogen. Improving the molecular knowledge of HAV strains, both autochthonous and circulating in countries from which potentially contaminated foods are imported, will become increasingly important to control outbreaks by supporting trace back activities, aiming to identify the geographical source(s) of contaminated food, as well as public health interventions.

Evaluation of rapid tests for diagnosis of acute hepatitis E

BACKGROUND Hepatitis E virus diagnosis still presents difficulties due to discordant results among diagnostic tests. OBJECTIVES The aim of this study was to evaluate the performance of two rapid tests for detection of anti-HEV IgM antibodies. STUDY DESIGN The rapid tests were compared with three commercial anti-HEV ELISA assays and one Real-Time PCR assay on 59 sera from patients with acute viral non-AC hepatitis. RESULTS The presence of anti-HEV IgM antibodies was evaluated by two rapid tests (Wantai and Assure) on 25 HEV RNA positive samples. Anti-HEV IgM antibodies were detected in 24/25 and 23/25 samples respectively. The sensitivity and specificity of Wantai and Assure Rapid tests were evaluated using the 25 HEV RNA positive samples and 50 HEV RNA negative samples (including sera from acute-phase HAV and HBV infections and blood donors). Overall, the sensitivity of Wantai Rapid and Assure Rapid tests was 96.1% and 92.6% respectively; the specificity of the 2 tests was 100%. CONCLUSION Our data suggest the potential use of anti-HEV IgM rapid assays as a first line test in primary health care settings, particularly useful for patients with chronic liver disease or pregnant women who urgently need an antiviral treatment.