Viral Interference Between Hepatitis B, C, and D Viruses in Dual and Triple Infections in HIV-Positive Patients

Morsica, Giulia*; Bagaglio, Sabrina*; Cicconi, Paola†; Capobianchi, Maria R‡; Pellizzer, Giampietro§; Caramello, Pietro‖; Orani, Anna¶; Moioli, Cristina#; Rizzardini, Giuliano**; Uberti-Foppa, Caterina*; Puoti, Massimo††; Monforte, Antonella d'Arminio†; for the Hepa I.C.o.N.A the Icona Foundation Study Groups

JAIDS Journal of Acquired Immune Deficiency Syndromes:
doi: 10.1097/QAI.0b013e3181add592
Clinical Science

Objective: To investigate the reciprocal inhibitory effects of hepatitis B virus (HBV)/hepatitis C virus (HCV)/hepatitis D virus (HDV) infections in naive and previously antiretroviral-experienced HIV-positive patients.

Design: This retrospective study involved 72 consecutive patients of the Italian Cohort Naive Antiretroviral cohort: 21 coinfected with HBV/HCV (group 1BC), 18 infected with HBV (group 2B), and 33 infected with HCV (group 3C).

Methods: Viral interference between HBV and HCV was assessed by means of the qualitative detection, quantification, and genotyping of each virus; HDV infection was assessed by means of genomic amplification.

Results: Univariate analysis showed that HBV DNA was less frequently detected in group 1BC than in group 2B (16 of 21 vs 18 of 18; P = 0.02), their HBV load was significantly lower (median 3.9 vs 5.4 log10 HBV DNA copies/mL; P = 0.002), and they more frequently carried HBV genotype D (12 of 13 vs 4 of 11; P = 0.0071). HCV RNA was less frequently detected in group 1BC than in group 3C (12 of 21 vs 33 of 33; P < 0.0001), and HDV RNA was more frequently detected in group 1BC than in group 2B (9 of 21 vs 2 of 18; P = 0.028). Multivariate analysis of the HBV-infected subjects showed that the risk of HCV coinfection was associated with older age [relative risk 0.28, 95% confidence interval (CI): 0.09 to 0.90; P = 0.033 for every 10 years older] and intravenous drug use (relative risk 73, 95% CI: 2.4 to >999.999; P = 0.013). The only predictor of HBV coinfection in HCV-infected individuals was a lower HCV load (relative risk 0.30, 95% CI: 0.11 to 0.79 for every additional log10 HCV RNA; P = 0.015).

Conclusion: HBV and HCV showed alternative dominant replication in the I.Co.N.A. cohort, with HBV having a more unfavorable effect on HCV replication.

Author Information

From the *Department of Infectious Diseases, San Raffaele, Scientific Institute, Milan, Italy; †Clinic of Infectious and Tropical Diseases, University of Milan, San Paolo Hospital, Milan Italy; ‡Laboratory of Virology, INMI L. Spallanzani, Rome, Italy; §Clinic of Infectious Diseases, Vicenza Hospital, Vicenza, Italy; ‖Department of Infectious Diseases, Amedeo di Savoia Hospital, Turin, Italy; ¶Department of Infectious Diseases, Lecco Hospital, Lecco, Italy; #Department of Infectious Diseases, Niguarda Hospital, Milan, Italy; **Department of Infectious Diseases, L Sacco Hospital, Milan, Italy; and ††Department of Infectious and Tropical Diseases, University of Brescia, Spedali Civili Hospital, Brescia, Italy.

Received for publication September 11, 2008; accepted March 30, 2009.

‡‡§§See Appendix.

The Icona Study Foundation is sponsored by unrestricted grants from Abbott, Boehringer-Ingelheim, Bristol Myers Squibb, Gilead, GSK, Janssen-Cilag, and Pfizer.

Correspondence to: Giulia Morsica, MD, Department of Infectious Diseases, Via Stamira d'Ancona 20, 20127 Milan, Italy (e-mail:

Article Outline
Back to Top | Article Outline


HIV, hepatitis B (HBV), hepatitis C (HCV), and hepatitis D (HDV) viruses are biologically different but, despite their different life cycles and modes of gene expression, share common transmission routes1 and so double or triple infection with hepatotropic viruses can occur in some HIV-infected patients.

The results of molecular studies of HIV-negative patients concerning the dominant role of either virus in HBV/HCV coinfection are contradictory: Some suggest that the dominant virus is HCV2-4 and others that it is HBV5,6; furthermore, in the case of triple hepatitis, HDV inhibits both HBV and HCV replication.7 However, the reciprocal effects of the viruses in HIV-infected patients with multiple viral infections are still unclear.8-10

Experimental evidence strongly supports the view that the replicative dominance of a virus is affected by the chronological sequence of infection, with the more recent playing an inhibitory role.11-13 Pontisso et al14 suggested that HBV dominance over HCV replication seems to lead to a state of latent HCV infection characterized by the absence of circulating HCV particles and the presence of HCV genomic sequences in the liver. However, more recent studies of HIV-infected subjects indicate that the absence of serum HCV RNA in the case of HBV coinfection is probably associated with the elimination of HCV.15,16

HBV infection is associated with the emergence of genomic mutations occurring randomly along the HBV genome that lead to the generation of different viral populations.17 The most prevalent mutation involving the pre-core (pre-C) region is the transition of TGG to TAG at nucleotide 1896, which creates a translational stop codon (codon 28) that causes the failure of HBeAg serum secretion.18 The pre-C codon stop mutation leading to a more stable epsilon structure is crucial for the pre-genomic encapsidation signal and the start of HBV DNA synthesis.

The reciprocal inhibition of dual HBV/HCV or triple HBV/HCV/HDV infection still needs further clarification, particularly in HIV-infected subjects in whom the immunological and virological factors related to HIV infection may interact with and modify the replication status of hepatitis B, C, and D.

In an attempt to clarify the factors or conditions that may affect viral interplay in such patients, we analyzed the replicative status of HIV, HBV, HCV, and HDV and virological factors including HBV/HCV genotypes and the presence of a pre-C stop codon, in a well-characterized HIV-positive cohort.

Back to Top | Article Outline



The Italian Cohort Naive Antiretroviral (I.Co.N.A) is a multicenter, prospective, observational study that started on April 1, 1997 and includes HIV-positive subjects who were antiretroviral drug naive at the time of enrolment. The patients come from 69 Infectious Disease Centers in Italy and give their written informed consent before enrolment. Their demographic, clinical, and laboratory data and information concerning specific therapies are recorded in the Internet cohort database at the time of enrolment and at least every 6 months during follow-up. The routine 6-monthly screening procedures for all the patients include immunovirological parameters and serological tests for HCV (anti-HCV antibodies, HCV-Ab) and HBV [hepatitis B surface antigen (HBsAg), hepatitis B surface antibodies (HBsAb)], whereas HCV RNA, hepatitis B core antibodies (HBcAb), and HBeAg/HBeAb are available for a subset of patients but not systematically recorded. The plasma samples are collected and stored on a voluntary basis by the participating centers at the same times.

From the 6200 HIV-positive subjects in the I.Co.N.A cohort as of April 2005, we selected those judged to be infected with HBV and/or HCV on the basis of their HBsAg and HCV-Ab positivity: 212 (3.4%) were infected by HBV, 2425 (39.1%) by HCV, and 194 (3.1%) by both. We then retrospectively studied those for whom plasma samples for the evaluation of virological data were available. The HCV-Ab-positive patients were also selected on the basis of their anti-HBcIgG negativity. The patients' demographic, clinical, and virological data were representative of the I.Co.N.A cohort as a whole, except in the case of the HCV-infected patients; although selected consecutively, the patients included in the analysis had lower CD4+ cell counts at the time the plasma samples were examined than those that were not included: 261 cells per microliter (interquartile ratio, IQR: 162-573) vs 420 cells per microliter (IQR: 226-615). HDV infection was assessed by searching for HDV RNA as anti-HDV immunoglobulin M and immunoglobulin G antibody data were not recorded in the database for all the patients.

Back to Top | Article Outline
Nucleic Acid Extraction

To detect HCV and HDV RNA, total RNA was extracted from 200 μL of plasma stored at −80°C using a commercially available kit (TRIzol LS; Gibco, BRL Grand Island, NY) in accordance with the manufacturer's instructions. To detect and quantify HBV DNA, total DNA was extracted from 200 μL of plasma using a QIAamp DNA Mini Kit (QIAGEN S.p.A., Milan, Italy) in accordance with the manufacturer's instructions.

Back to Top | Article Outline
Qualitative HBV DNA Detection, Quantification, Genotyping, and Pre-Core Mutant Characterization
Qualitative Detection

HBV DNA was amplified by means of a heminested polymerase chain reaction (PCR) using primers spanning the partial preS1/preS2 region (outer: sense 2817-2839, antisense 685-704; inner: antisense 3081-3098) and pre-core/core (preC/C) regions of the HBV genome (outer: sense 1746-1763; outer antisense 2286-2270; inner: antisense 1960-1978). The sensitivity limit was 10−5 of cloned HBV DNA (2.6 copies/mL).

Back to Top | Article Outline
Quantification by Means of Real-Time PCR

HBV DNA was quantified by means of an in-house real-time PCR using primers located within the pre-C/C regions of the HBV genome (GenBank Accession No. AF121241): sense primer hbv1857 (5′-CAC TGT TCA AGC CTC CAA GCT-3′; nt 1857-1877) and antisense primer hbv1928 (5′-CAA ATT CTT TAT AAG GGT CAA TGT CAA T-3′; nt 1928-1901). A probe of 15 bp (5′-TGC CTT GGG TGG CTT-3′; nt 1880-1894), which recognizes a region downstream of primer hbv1857, was synthesized (PE Biosystems, Warrington, United Kingdom) with the FAM reporter dye covalently linked to its 5′-end and a downstream nonfluorescent quencher (minor groove binder [MGB]).

The targeted amplicon of 71 bp was selected within a pre-C/C region of HBV that is conserved among different isolates. An extensive search of 2 databases (EMBL and GenBank) indicated that neither the primers nor the probe shared significant homology with other known nucleotide sequences.

Back to Top | Article Outline
Plasmid Preparation

HBV DNA was extracted from the 200 μL of plasma as described above, and the PCR products were cloned into the pCRII plasmid using the TOPO-TA cloning kit (Invitrogen, Corp, San Diego, CA) in accordance with the manufacturer's instructions.

Back to Top | Article Outline
Real-Time PCR Conditions

All the reactions were optimized to obtain the best amplification kinetics under the same cycling conditions (2 minutes at 50°C, 15 minutes at 95°C, and 40 cycles of 15 seconds at 95°C, and 1 minute per cycle at 60°C) and with the same reaction mixture composition. All the reactions were performed in a final volume of 25 μL containing 100 mM each of dATP, dCTP, and dGTP; 200 mM dUTP; 4 mM MgCl2; 1TaqMan buffer A; 0.625 U of AmpliTaq Gold, 0.25 U of uracil-N-glycosylase; and 10 μL of DNA template. Target-specific primers and a probe were used at final concentrations of 300 and 200 nM, respectively. Each sample was tested in triplicate, and the mean value was recorded. The sensitivity limit of the real-time PCR was 103-108 copies per milliliter.

Back to Top | Article Outline
HBV Genotyping and Pre-Core HBV Mutant Characterization

The HBV genotype and pre-core mutants were, respectively, determined by directly sequencing the PCR products of the partial pre-S1/pre-S2 region and pre-C/C domain.19,20 The PCR products were sequenced by means of an ABI 373 automated sequencer using PRISM Dye terminator cycle sequencing (Perkin Elmer; Foster City, CA) in accordance with the manufacturer's instructions, and HBV was genotyped by making a phylogenetic comparison with database sequences of pre-S domains representing genotypes A-H. The pre-core mutants were determined by comparing the HBV sequences obtained from our patients with that of the ayw HBV prototype.

Back to Top | Article Outline
Qualitative and Quantitative HCV RNA Analysis and Genotyping

After reverse transcription, 5 μL cDNA was amplified in 50 μL of PCR mixture containing10 pmol of an outer primer set encompassing the 5′-untranslated region of the HCV genome. For the second PCR, 2 μL of the first amplification products was amplified with 10 pmol of an inner primer set.21 The sensitivity limit of the in-house-nested PCR assays was 10 copies per milliliter for the amplification of the HCV genome. HCV RNA levels were determined using an Amplicor HCV Kit (HCV Monitor Test; Roche Diagnostic System, Basel, Switzerland; detection threshold <600 copies/mL). HCV was genotyped by means of direct sequencing of the PCR products derived from the 5′-untranslated region and alignment with the respective prototype.22

Back to Top | Article Outline
HDV RNA Detection

HDV infection was evaluated by detecting HDV RNA in stored frozen plasma samples. After reverse transcription with outer antisense primer D-3, 5 μL cDNA was amplified with 10 pmoles of outer primers encompassing the HDVAg of the HDV genome: outer sense primer D-0 5′-AGTGGCTCTCCCTTAGCCAT-3′ (nt 813-832); outer antisense D-3 5′-TGAACCCCCTCGAAGGTGGA-3′ (nt 1147-1128); inner sense primer D1 5′-GTCCTCCTTCGGATGCCCAG-3′ (nt 847-866); and inner antisense D2 5′-GAGTCCCGGAGTCCCCCTT-3′ (nt 1084-1066).

Nested PCR was performed on 2 μL of the first amplification products as a template with 10 pmol of the inner primer set. The first PCR was conducted for 38 cycles of 30 seconds at 94°C, 30 seconds at 55°C, and 30 seconds at 72°C. The second PCR was carried out under the same cycling conditions except for the annealing temperature, which was 60°C instead of 55°C. The last cycle was extended for an extra 5 minutes at 72°C.

Back to Top | Article Outline
Statistical Analysis

The patients were divided into 3 groups: group 1BC with HBV/HCV coinfection, group 2B with HBV infection, and group 3C with HCV infection. Group 1BC was compared with groups 2B and 3C in terms of demographics and immunovirological characteristics using the χ2 test for discrete variables and Wilcoxon test for continuous variables. Logistic regression models were used for the multivariable analyses. One model included groups 1BC and 2B and considered HCV infection as the outcome; the analyzed variables were HIV risk factor [intravenous drug users (IVDU) vs non-IVDU], age (per 10 years), HDV RNA positivity, and HBV DNA quantification (per additional log10 copies/mL). To study the role of the HBV genotype in HCV coinfection, a logistic regression subanalysis considered only the patients for whom the HBV genotype was available, with the variables HBV genotype (D vs non-D), HDV infection (yes/no), and HBV DNA (per additional log10).

A separate logistic regression analysis model included groups 1BC and 3C and considered HBV infection as the outcome; the analyzed variables were HIV risk factor (IVDU vs non-IVDU), age (per 10 years), sex, CD4+ cells (per additional 100 cells/μL), HIV RNA (per additional log10 copies/mL), HCV genotype (genotype 1 vs non-1), and HCV RNA (per additional log10 copies/mL). All the parameters analyzed were considered at the time of plasma sample collection. The significance tests were all 2 sided, and a P value of <0.05 was considered statistically significant.

Back to Top | Article Outline


Demographic and Clinical Features

Among 72 consecutive HIV-positive subjects with serological markers for HBV and/or HCV infection and available biological specimens, 21 (group 1BC) were HBV/HCV coinfected, 18 (group 2B) had HBV infection alone, and 33 (group 3C) had HCV infection. The last 2 groups were chosen as internal controls for each infection (HBV or HCV) with reference to the study group of HBV/HCV coinfected individuals.

Ten of the 39 subjects with serological markers of HBV infection were on antiretroviral therapy (ART), including 7 on lamivudine-including ART; the median duration of lamivudine treatment was 9.5 months (IQR: 0.4-18.6). None of the 3 patients receiving ART without lamivudine had been previously exposed to any drug active against HBV; the newer drugs for treating HBV infection, such as adefovir, entecavir, and tenofovir, did not form part of their ART. None of the patients (including those with HCV infection alone) had received interferon alpha for active chronic hepatitis.

The subjects in group 1BC were younger than those in group 2B but had the same median age as those in group 3C (Table 1). Analysis of the risk factors for HIV transmission showed that transmission routes were similar in groups 1BC and 3C, whereas the prevalence of sexual transmission was higher in group 2B than group 1BC (Table 1). The 3 groups were comparable in terms of the immunovirological parameters related to HIV infection (Tables 1, 2).

Group 1BC had higher median alanine aminotransferase (ALT) levels at the time of specimen examination: 83 IU/L (IQR: 50-150) as against 41 IU/L (IQR: 32-97) in group 2B and 52 IU/L (IQR: 35-65) in group 3C. However, only the difference between group 1BC and group 2B was statistically significant (P = 0.03; Table 1).

Back to Top | Article Outline
Virological Patterns of Hepatitis Virus Coinfection

The virological data are summarized in Table 2.

Back to Top | Article Outline
Virological Characteristics of HBV in Groups 1BC and 2B

Active HBV replication was found in 16 of the 21 subjects with serological evidence of dual HBV/HCV infection (the remaining 5 had undetectable HBV DNA levels), and HBV DNA was detected in all 18 patients with single HBV infection: The difference between the groups was statistically significant (P = 0.02). The median HBV load was significantly higher in group 2B than 1BC: 5.4 (IQR: 4.6-8.9) log10 copies per milliliter vs 3.9 (IQR: 3.0-7.0) HBV DNA log10 copies per milliliter. Table 3 shows the ART of the 10 patients receiving it and their HBV DNA status.

The HBV genotype was successfully determined in 13 of the 16 subjects in group 1BC and 11 of the 18 in group 2B. HBV genotyping was not possible in 5 subjects in group 1BC because of their undetectable HBV DNA levels; in the case of 7 subjects in group B, the amplification products were insufficient to obtain genomic sequences.

Interestingly, HBV genotyping in group 2B showed the presence of genotype D in 4 subjects, genotype G in 6, and genotype A in 1, whereas genotype D was found in 12 of the 13 group 1BC patients (the other carried genotype A), and there were no cases of genotype G. The difference in the distribution of HBV genotypes in these 2 groups was statistically significant (P = 0.0071).

The A pre-core mutation (G1896 → A, codon 28) that leads to the defective synthesis of HBeAg was detected in 3 of 14 patients with dual HBV/HCV infection and 1 of 15 patients with single HBV infection. Its prevalence was similar in the 2 groups.

Multivariate analysis showed that age [relative risk (RR) 0.098, 95% confidence interval (CI): 0.011 to 0.853; P = 0.033] and transmission route (IVDU vs non-IVDU; RR 73, 95% CI: 2.5 to >999.999; P = 0.013) were independent variables associated with HBV/HCV coinfection.

Back to Top | Article Outline
Virological Characteristics of HCV in Groups 1BC and 3C

Twelve of the 21 patients in group 1BC and all 33 in group 3C were positive for HCV RNA, a difference in frequency that was statistically significant (P < 0.0001); the amounts of HCV RNA detected were similar in the 2 groups. Multivariate analysis showed that the risk of HBV coinfection in HCV-infected individuals was related to a lower HCV load (RR 0.30, 95% CI: 0.11 to 0.79; P = 0.015). HCV genotypes were equally distributed in group 1BC and group 3C (Table 2).

Back to Top | Article Outline


HDV RNA was more frequently detected in group 1BC (9 of 21 patients, 43%) than in group 2B (2 of 18; 11%; P = 0.028). The frequency of HBV DNA detection was similar in the patients in groups 1BC and 2B with HDV coinfection (8 of 9 and 2 of 2; P = 0.62), as was the median HBV load: 4.05 (IQR: 3.76-4.66) log10 copies per milliliter vs 6.84 (IQR: 4.15-9.52) log10 copies per milliliter (P = 0.19). All 9 HDV RNA-positive patients in group 1BC were IVDUs, and 8 of 8 carried HBV genotype D; one of the 2 HDV RNA-positive individuals in group 2B carried HBV genotype D and the other genotype G.

Table 4 summarizes the risk factors and virological characteristics in group 1BC on the basis of the presence of HDV RNA.

Back to Top | Article Outline


Demographic Characteristics

The subjects in group 1BC were younger than those in group 2B but had the same median age as those in group 3C. As most of the subjects in groups 1BC and 3C were ex-IVDUs, whereas sexual exposure was prevalent in group 2B, their younger age may have been a consequence of the different transmission route.

Mean ALT level in group 1BC was higher than in group 2B but similar to that observed in group 3C, thus suggesting that chronic carriers of HBsAg were prevalent in group 2B despite their higher HBV viral load. However, as alcohol intake was not investigated, we cannot exclude the possibility that it was higher in groups 1BC and 3C because of the similar frequency of IVDUs.

Furthermore, a single determination of ALT levels is not sufficient to draw any definite conclusion concerning the activity of liver disease, and most of the subjects had not undergone a liver biopsy or their histological results were not available.

Our finding is partially in line with those obtained in HIV-negative patients with HBV/HCV coinfection,23,24 although, in these cases, ALT activity was related to the synergistic effect of the dual infection (assessed on the basis of anti-HCV Ab and HBsAg positivity) because it was associated with higher ALT levels than either HBV or HCV infection alone.

Back to Top | Article Outline
HBV Virological Profiles in Groups 1BC and 2B

The difference in the detectability of HBV DNA between groups 1BC and 2B was statistically significant, and so it is likely that in some cases with serological evidence of dual HBV/HCV infection, HBV was inhibited by HCV; however, only a small number of patients in group 1BC had undetectable HBV DNA levels. Similarly, HBV DNA levels were higher in group 2B than in group 1BC, although both groups had comparable CD4+ cell counts and HIV viremia levels. These findings suggest that HCV is dominant in some cases of dual HBV/HCV infection and that HIV infection does not affect HBV replication under conditions of relatively preserved immune competence.

HBV genotype D was prevalent in the patients with HBV/HCV coinfection, whereas genotype G was also detected in those with single HBV infection. The frequency of the HBV genotypes was significantly different in the 2 groups, but it was not possible to analyze the independent predictive role of the risk factor for HIV and the infecting genotype in the multivariable model because most of the IVDUs were genotype D carriers and the 2 variables were colinear.

Fewer than 20% of the patients with HBV/HCV coinfection became infected as a result of sexual intercourse, and none of these carried genotype G. On the other hand, sexual exposure was a risk factor for HIV transmission in all 6 subjects with genotype G in group 2B.

We hypothesize that genotype G was more frequently sexually transmitted in our cohort of HIV-infected subjects, as has been reported in other small series of HIV-negative and HIV-positive patients.25,26 Interestingly, despite Mediterranean basin prevalence of HBV genotype D harboring the G to A mutation at position 1896 of the pre-core region, we found the G1896 wild-type strain in the subjects with dual HBV/HCV infection and in those with single HBV infection. Finally, the comparable rate of the pre-core stop codon mutant in groups 1BC and 2B argues against the possibility of increased replication efficiency (known to be associated with the emergence of pre-core mutants)27,28 in 1 of the 2 groups.

Back to Top | Article Outline
HCV Characteristics in Groups 1BC and 3C

A significant number of the HBV/HCV coinfected subjects did not show active HCV replication possibly because of viral clearance after a previous HCV infection. Interestingly, a recent study29 has found markedly increased HCV clearance in chronic HBsAg carriers with HIV coinfection, although the authors did not exclude mutual interference in viral genome replication as a probable cause. Furthermore, they did not investigate the chronic HBsAg carriers with HCV RNA clearance by looking for HBV DNA and HDV RNA genomic sequences, which may modify HCV replication.

The HCV genotype had no effect on dual HBV/HCV in comparison with single HCV infection. In particular, the distribution of HCV genotype 1, which may be associated with a higher HCV load in HIV-infected subjects,30 was similar in the 2 groups.

Back to Top | Article Outline
HDV in HBV/HCV Coinfection and Single HBV Infection

It is well known that HDV infection is more prevalent in the Mediterranean area than in Western countries and may depend on risk practices.1 The prevalence of HDV markers in European cohorts of chronic HBsAg carriers with HIV coinfection has been estimated as varying from 4% to 44%,8,31 and the presence of HDV RNA was found in 62% of HBV/HCV coinfected subjects in a US cohort of HIV-infected hemophiliacs.32

Eleven percent of our HIV-infected patients with HBV coinfection were positive for HDV RNA and 43% of those with HBV/HCV infection, which is in line with the findings of previous studies carried out in Europe.1,33 The HDV RNA-positive subjects with dual HBV/HCV infection were prevalently IVDUs but, as HDV can be transmitted parenterally or sexually,9 the higher prevalence of HDV RNA in group 1BC cannot be fully explained by the different risk practices, and it is possible that the alternating dominance of HBV and HCV may interfere with HDV RNA detectability.

It has been shown that HDV suppresses both HBV and HCV replication,8,34,35 but only 1 of these studies involved HIV-infected patients with multiple infection. The discrepancy between our finding and that of this study of HIV-infected subjects may have been due to the more sensitive PCR assays we used to detect HBV/HCV genomic sequences or to the clinical characteristics of the patients because most of the HIV-positive subjects with multiple infections did not receive any antiretroviral treatment.

A more recent study9 of HIV-infected patients found that 36.8% of HBV-infected subjects had detectable HDV RNA and that HBV load was not significantly less in those with dual HBV/HDV infection than in those with single HBV infection. Similarly, we found HBV and HCV replication in both HDV-negative and HDV-positive patients, thus suggesting that HDV does not suppress or dramatically decrease HBV and HCV replication in plasma samples.

It is well known that lamivudine is effective against HBV and HIV, but only 7 of our HBV-infected subjects were on a lamivudine-including ART regimen (3 in group 1BC and 4 in group 2B) and their ART had no effect on HBV load in comparison with the untreated patients. We hypothesize that the possible activity of lamivudine on HBV and HDV replication was ineffective, but the small sample size and differences in the duration of lamivudine treatment do not allow any definite conclusions.

A cross-sectional study provides snapshot of viral interactions at replicative levels. Our and others' previous cross-sectional studies of HIV-negative and HIV-positive subjects have revealed similar interference concerning multiple viral infection. It is unlikely that the inherent genomic factors analyzed changed over time and so the results provide important information concerning the distribution of HBV/HCV genotypes and pre-core stop codon mutants in our patients, especially those with HBV/HCV coinfection. However, differences in risk practices may limit their applicability.

In conclusion, reciprocal inhibition may occur in HIV-infected subjects with dual HBV/HCV infection, with HBV having a stronger inhibitory effect on the replication of HCV. CD4+ cell counts and HIV viral load in or relatively immune competent patients did not affect the dominance of one virus over the other, and the detection of HDV RNA did not seem to modify HBV and HCV replication in the patients with dual HBV/HCV infection or single HBV infection.

Our findings of different HBV genotypes and HDV replication patterns in groups 1BC and 2B encourages the use of such analyses in larger cohorts of subjects characterized by the same risk factor for viral acquisition. Careful monitoring of virus dominance and HBV genotypes will add important information for treatment strategies because the current standards of treatment for chronic HBV and HCV infection differ and recent data show an association between HBV genotype and the response of HBV infection to treatment.

Back to Top | Article Outline


1. Gaeta GB, Precone DF, Cozzi-Lepri A, et al. Multiple viral infections. J Hepatol. 2006;44:S108-S113.
2. Crespo J, Lozano JL, Carte B, et al. Viral replication in patients with concomitant hepatitis B and C virus infection. Eur J Clin Microbiol Infect Dis. 1997;16:445-451.
3. Fong T-L, Di Bisceglie A, Waggoner JG, et al. The significance of antibody to hepatitis C virus in patients with chronic hepatitis B. Hepatology. 1991;14:64-67.
4. Liaw Y-F. Role of hepatitis C virus in dual and triple hepatitis virus infection. Hepatology. 1995;22:1101-1108.
5. Zarski J-P, Bohn B, Bastie A, et al. Characteristics of patients with dual infection by hepatitis B and C viruses. J Hepatol. 1998;28:27-33.
6. Koike K, Yasuda K, Yotsuyanagi H, et al. Dominant replication of either virus in dual infection with hepatitis viruses B and C. J Med Virol. 1995;45:236-239.
7. Mathurin P, Thibault V, Kadidja K, et al. Replication status and histological features of patients with triple (B, C, D) and dual (B, C) hepatic infections. J Viral Hepat. 2000;7:15-22.
8. Arribas JR, Gonzáles-García JJ, Lorenzo A, et al. Single (B or C), dual (BC or BD) and triple (BCD) viral hepatitis in HIV-infected patients in Madrid, Spain. AIDS. 2005;19:1361-1365.
9. Sheng WH, Hung CC, Kao JH, et al. Impact of hepatitis D virus infection on the long-term outcomes of patients with hepatitis B virus and HIV coinfection in the era of highly active antiretroviral therapy: a matched cohort study. Clin Infect Dis. 2007;44:988-995.
10. Maida I, Ríos MJ, Pérez-Saleme L, et al. Profile of patients triply infected with HIV and the hepatitis B and C viruses in the HAART era. AIDS Res Human Retroviruses. 2008;24:679-683.
11. Tsiquaye KN, Portmann B, Tovey G, et al. Non-A, non-B hepatitis in persistent carriers of hepatitis B virus. J Med Virol. 1983;11:179-189.
12. Liaw YF, Yeh CT, Tsai SL. Impact of acute hepatitis B virus superinfection on chronic hepatitis C virus infection. Am J Gastroenterol. 2000;95:2978-2980.
13. Sagnelli E, Coppola N, Messina V, et al. HBV superinfection in hepatitis C virus chronic carriers, viral interaction, and clinical course. Hepatology. 2002;36:1285-1291.
14. Pontisso P, Ruvoletto MG, Fattovich G, et al. Clinical and virological profiles in patients with multiple hepatitis virus infections. Gastroenterology. 1993;105:1529-1533.
15. Martín-Carbonero L, Barriero P, Jiménez-Galán G, et al. Clearance of hepatitis C virus in HIV-infected patients with multiple chronic viral hepatitis. J Viral Hepat. 2007;14:392-395.
16. Soriano V, Barreiro P, Martín-Carbonero L, et al. Treatment of chronic hepatitis B or C in HIV-infected patients with dual viral hepatitis. J Infect Dis. 2007;195:1181-1183.
17. Brunetto MR, Rodriguez UA, Bonino F. Hepatitis B virus mutants. Intervirology. 1999;42:69-80.
18. Carman WF, Jacyna MR, Hadziyannis S, et al. Mutation preventing formation of hepatitis B e antigen in patients with chronic hepatitis B infection. Lancet. 1989;2:588-591.
19. Bortolotti F, Guido M, Bartolacci S, et al. Chronic hepatitis B in children after e antigen seroclearance: final report of a 29-year longitudinal study. Hepatology. 2006;43:556-562.
20. De Mitri MS, Cassini R, Morsica G, et al. Virological analysis, genotypes and mutational patterns of the HBV precore/core gene in HBV/HCV-related hepatocellular carcinoma. J Viral Hepat. 2006;13:574-581.
21. Bagaglio S, De Mitri MS, Lodrini S, et al. Mutations in the E2-PePHD region of hepatitis C virus type 1b in patients with hepatocellular carcinoma. J Viral Hepat. 2005;12:243-250.
22. Morsica G, Bagaglio S, Ghezzi S, et al. Hepatitis C virus (HCV) coinfection in a cohort of HIV positive long-term non-progressors: possible protective effect of infecting HCV genotype on HIV disease progression. J Clin Virol. 2007;39:82-86.
23. Crespo J, Lozano JL, De la Cruz F, et al. Prevalence and significance of hepatitis C viremia in chronic active hepatitis B. Am J Gastroenterol. 1994;89:1147-1151.
24. Raimondo G, Brunetto MR, Pontisso P, et al. The Associazione Italiana Studio del Fegato (AISF) cooperative group. Longitudinal evaluation reveals a complex spectrum of virological profiles in hepatitis B virus/hepatitis C virus-coinfected patients. Hepatology. 2006;43:100-107.
25. Kato H, Gish RG, Bzowei N, et al. Eight genotypes (A-H) of hepatitis B virus infecting patients from San Francisco and their demographic, clinical, and virological characteristics. J Med Virol. 2004;73:516-521.
26. Alvarado-Esquivel C, Sablon E, Conde-González CJ, et al. Molecular analysis of hepatitis B virus isolates in Mexico: predominant circulation of hepatitis B virus genotype H. World J Gastroenterol. 2006;28:6540-6545.
27. Tong S-P, Li J-S, Vitvitski L, et al. Replication capacities of natural and artificial precore stop codon mutants of hepatitis B virus: Relevance of pregenome encapsidation signal. Virology. 1992;191:237-245.
28. De Mitri MS, Morsica G, Cassini R, et al. Low replication and variability of HBV pre-core in concomitant infection with hepatitis B and hepatitis C viruses. Arch Virol. 2007;152:395-404.
29. Melendez-Morales L, Konkle BA, Preiss L, et al. Chronic hepatitis B and other correlates of spontaneous clearance of hepatitis C virus among HIV-infected people with hemophilia. AIDS. 2007;31:1631-1636.
30. Soriano V, Mocroft A, Rockstroh J, et al. EuroSIDA Study Group. Spontaneous viral clearance, viral load, and genotype distribution of hepatitis C virus (HCV) in HIV-infected patients with anti-HCV antibodies in Europe. J Infect Dis. 2008;198:1337-1344.
31. Chlabicz S, Grzeszczuk A, Lapínski TW, et al. Search for hepatitis delta (HDV) infection in hepatitis C patients in north-eastern Poland. Comparison with anti-HDV prevalence in chronic hepatitis B. Eur J Epidemiol. 2003;18:559-561.
32. Eyster ME, Sanders JC, Battegay M, et al. Suppression of hepatitis C virus (HCV) replication by hepatitis D virus (HDV) in HIV-infected hemophiliacs with chronic hepatitis B and C. Dig Dis Sci. 1995;40:1583-1588.
33. Denis F, Adjide CC, Rogez JP, et al. Seroprevalence of HBV, HCV, and HDV hepatitis markers in 500 patients infected with human immunodeficiency virus. Pathol Biol. 1997;45:701-708.
34. Sagnelli E, Coppola N, Scolastico C, et al. Virological and clinical aspects of reciprocal inhibitory effect of hepatitis B, C, and Delta viruses in patients with chronic hepatitis. Hepatology. 2000;32:1106-1110.
35. Jardi R, Rodriguez F, Buti M, et al. Role of hepatitis B, C, and D viruses in dual and triple infection: influence of viral genotypes and hepatitis B precore and basal core promoter mutations on viral replicative interference. Hepatology. 2001;34:404-410.
Back to Top | Article Outline

F. Ancarani, A. Antinori, G. Antonucci, R. Bruno, M. R. Capobianchi, A. Cingolani, A. Cozzi-Lepri, A. d'Arminio Monforte, M Galli, E. Girardi, N. Marino, G. Morsica, P. Narciso, C. Pastecchia, P. Pizzaferri, M. Puoti, T. Santantonio, and G. Verucchi.

ICONA Foundation (Italy)-Central Coordinator: A. d'Arminio Monforte. Steering Committee: A. Ammassari, A. Antinori, C. Arici, C. Balotta, P. Bonfanti, M. R. Capobianchi, A. Castagna, F. Ceccherini-Silberstein. A. Cozzi-Lepri, A. d'Arminio Monforte, A. De Luca, C. Gervasoni, E. Girardi, S. Lo Caputo, R. Murri, C. Mussini, M. Puoti, and C. Torti. Governing Body: M. Moroni (Chair), G. Carosi, R. Cauda, F. Chiodo, A. d'Arminio Monforte, G. Di Perri, M. Galli, R. Iardino, G. Ippolito, A. Lazzarin, F. Mazzotta, R. Panebianco, G. Pastore, and CF. Perno. Participating Physicians and Centers-Italy: M. Montroni, G. Scalise, A. Costantini, and A. Riva (Ancona); U. Tirelli and F. Martellotta (Aviano); G. Pastore and N. Ladisa (Bari); F. Suter and F. Maggiolo (Bergamo); F. Chiodo, V. Colangeli, and C. Fiorini (Bologna); G. Carosi, G. Cristini, C. Torti, C. Minardi, and D. Bertelli (Brescia); T. Quirino (Busto Arsizio); P. E. Manconi and P. Piano (Cagliari); E. Pizzigallo and M. Dalessandro (Chieti); G. Carnevale and A. Zoncada (Cremona); F. Ghinelli and L. Sighinolfi (Ferrara); F. Leoncini, F. Mazzotta, M. Pozzi, and S. Lo Caputo (Florence); B. Grisorio and S. Ferrara (Foggia); G. Pagano, G. Cassola, A. Alessandrini, and R. Piscopo (Genoa); F. Soscia and L. Tacconi (Latina); A. Orani and P. Perini (Lecco); D. Tommasi and P. Congedo (Lecce); F. Chiodera and P. Castelli (Macerata); M. Moroni, A. Lazzarin, G. Rizzardini, L. Caggese, A. d'Arminio Monforte, A. Galli, S. Merli, C. Pastecchia, and M. C. Moioli (Milan); R. Esposito and C. Mussini (Modena); A. Gori and S. Cagni (Monza), N. Abrescia, A. Chirianni, CM. Izzo, M. De Marco, R. Viglietti, and E. Manzillo (Naples); C. Ferrari, P. Pizzaferri (Parma); G. Filice and R. Bruno (Pavia); G. Magnani and M. A. Ursitti (Reggio Emilia); M. Arlotti and P. Ortolani (Rimini); R. Cauda, M Andreoni, A. Antinori, G. Antonucci, P. Narciso, V. Tozzi, V. Vullo, A. De Luca, M. Zaccarelli, R. Acinapura, P. De Longis, M. P. Trotta, M. Lichtner, and F. Carletti (Rome); M. S. Mura and M. Mannazzu (Sassari); P. Caramello, G. Di Perri, G. C. Orofino, and M. Sciandra (Turin); E. Raise and F. Ebo (Venice); and G. Pellizzer and D. Buonfrate (Vicenza). Cited Here...

© 2009 Lippincott Williams & Wilkins, Inc.