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).
Virological Patterns of Hepatitis Virus Coinfection
The virological data are summarized in Table 2.
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.
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).
HDV RNA DETECTION IN GROUPS 1BC AND 2B
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.
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.
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.
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.
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.
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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).© 2009 Lippincott Williams & Wilkins, Inc.