Lack of effect of tumor necrosis factor-alpha -308 G/A polymorphism on severity of liver fibrosis in Tunisian hepatitis C virus (HCV)-infected patients.

OBJECTIVES: Tumor necrosis factor alpha (TNF-alpha) plays a key role in the immune response. An elevated plasma level of TNF-alpha was repeatedly observed in patients with active liver injury or cirrhosis regardless of the aetiology. The G/A transition at position -308 in the promoter region have been shown to influence TNF-alpha expression. In this study, we aimed to evaluate the impact of TNF-alpha -308 G/A functional polymorphism on fibrosis severity in Tunisian Hepatitis C Virus (HCV)-infected patients. METHODS: TNF-alpha -308 G/A polymorphism was evaluated by polymerase chain reaction (PCR) amplification followed by Restriction Fragment Length Polymorphism (RFLP) method in 53 chronic hepatitis C patients. Single-nucleotide polymorphism (SNP) frequencies were compared with regard to liver fibrosis severity as assessed by the METAVIR scoring system (F1-F2; n=22 versus F3-F4; n=31). RESULTS: The genotype distribution of the TNF-alpha -308 G/A polymorphism among the HCV-infected patients was as follows : GG : 67.9%, GA : 32.1%, AA : 0%. With regard to fibrosis score, no significant differences in TNF-alpha genotype distribution were observed between F1-F2 and F3-F4 patients (p=0.15). CONCLUSION: No significant association between TNF-alpha -308 polymorphism and and the severity of liver fibrosis was found in our Tunisian cohort.

NS5A(ISDR-V3) region genetic variability of Tunisian HCV-1b strains: Correlation with the response to the combined interferon/ribavirin therapy.

In the non-structural protein 5A (NS5A) of hepatitis C virus (HCV), mutations within the interferon sensitivity-determining region (ISDR), the PKR-binding domain (PKR-BD), the variable region 3 (V3), and the interferon/ribavirin resistance-determining region (IRRDR) have been correlated with the IFN-based therapy response. In Tunisia, where a high prevalence of HCV-1b has been found, no data regarding the implication of NS5A in treatment response were available. The current study examined the relationship between the pre-treatment mutation number within ISDR, PKR-BD, V3, IRRDR, as well as in the entire ISDR-V3 region of NS5A (aa 2209-2379) and the response to the 48-week course of combined IFN plus ribavirin therapy in 15 HCV-1b-infected Tunisian patients. Referring to HCV-J sequence, a significant high genetic variability was observed within PKR-BD in the sustained virological responder patients compared to non-responders (P = 0.040). More importantly, when considering the entire region from ISDR to V3, referred to as NS5A(ISDR-V3), a clear difference in the mutation number was observed between sustained virological responders (19.6 +/- 3.16) and non-responders (15.0 +/- 1.41) (P = 0.002). Additionally, a more detailed analysis of NS5A(ISDR-V3) region revealed an elevated degree of mutation rate within the region located between amino acids 2282 and 2308 (P = 0.0006). Interestingly, an analysis of specific amino acid variations defined proline and serine at position 2300 as signature patterns for sensitive and resistant strains, respectively. The genetic variability within the NS5A region of HCV-1b strains was associated with the response to the combined IFN plus ribavirin therapy in our Tunisian cohort.

Association of interleukin-18 polymorphisms and plasma level with the outcome of chronic HCV infection.

Hepatitis C virus (HCV) infection is the main cause of chronic liver disease throughout the world, and may progress to cirrhosis and hepatocellular carcinoma (HCC). Immunological factors, especially cytokines and some host genetic variations, rather than direct HCV action, seem to play an important role in the pathogenesis of HCV infection. Elevated levels of interleukin-18 (IL-18) were described previously for chronically (HCV)-infected patients. This study is aimed at investigating IL-18 promoter polymorphisms (-607C/A and -137G/C) in HCV-infected patients with different disease severities (chronic hepatitis C, liver cirrhosis and HCC) and establishing an association between these polymorphisms and IL-18 plasma concentration with the outcome of chronic HCV infection. The carriage of at least one C allele at position -607 (CC + CA) was associated with a higher risk of cirrhosis and HCC (P = 0.032). Compared with controls, HCV-infected patients had significantly higher levels of IL-18 (P = 0.0001) that correlate with disease severity (P = 0.01, P = 0.001, P = 0.0006, respectively). In conclusion, we supposed a possible implication of IL-18 promoter polymorphisms in the pathogenesis of chronic HCV infection.

Evaluation of a total core antigen assay for the diagnosis of hepatitis C virus infection in hemodialysis patients.

Hemodialysis patients are recognized as a group at high risk of infection with hepatitis C virus (HCV). Therefore, such a population should be screened routinely for the presence of HCV viremia. Since nucleic acid techniques remain expensive and largely unavailable in many laboratories in the developing world, the present study assesses the clinical usefulness of the HCV core antigen enzyme immunoassay for the diagnosis of HCV infection in dialysis patients. One hundred seventy-five dialysis patients were screened for the presence of anti-HCV antibodies and HCV RNA in the serum. One hundred twenty-eight serum samples were collected from the 76 patients who were anti-HCV antibody- and/or HCV RNA-positive. These were evaluated for total HCV core antigen. Of these samples, 55 had sufficient volume to be further tested to quantify HCV RNA by reverse transcription polymerase chain reaction (RT-PCR). Genotyping of the HCV strains showed that the majority belonged to genotype 1b (77%). The HCV core antigen assay showed a sensitivity and specificity of 84% and 89%, respectively. The use of core antigen assay has enabled the early detection of three patients who developed an acute hepatitis C infection during the period of study. A correlation study was undertaken between the quantitative values of viral load, expressed as pg/ml of HCV core antigen in serum, and viral RNA in UI/ml. A significant correlation was observed (Pearson's correlation coefficient: 0.552; P<0.001). In conclusion, detection of HCV core antigen in serum is an inexpensive, reliable, and highly specific assay that can be useful in most laboratory settings to diagnose HCV infection, and especially in laboratories where nucleic acid technologies are not yet available.

[High prevalence and incidence of hepatitis C virus infections among dialysis patients in the East-Centre of Tunisia]. / Prévalence et incidence élevées de l'infection par le virus de l'hépatite C chez les hémodialysés dans la région Centre-Est de la Tunisie.

Hemodialysed patients are recognised as a group at increased risk of infection with hepatitis C virus (HCV). The aim of this study was to determine the prevalence and incidence of HCV infection among dialysis patients of the east-centre part of Tunisia. Two hundred and seventy-six patients dialysed until 2001 were recruited within seven hemodialysis units located in the cities of Sousse, Monastir and Mahdia. The serum markers of HCV infection were tested over the period of March 2000-December 2002, by a 3rd generation ELISA test for antibodies and by qualitative RT-PCR technique for viral RNA. The prevalence of anti-HCV antibodies and of HCV RNA was 32.6% (90 patients) and 25.7% (71 patients), respectively. Between 1998 and 2002, 20 new infections were documented in five of the seven dialysis units corresponding to an incidence of 2.34% per year, with an average time of contamination after the beginning of dialysis of 4.6 years. If all the infections are assessed to have occurred during dialysis, the density of incidence of HCV contamination was 4.4% per year of dialysis. A high correlation was noticed between the presence of HCV markers in serum and the duration of dialysis (F = 34.15, P < 0.0001). In the absence of other risk factors (transfusion, drug-addiction), these results plead for the nosocomial transmission of the observed HCV infections. A phylogenetic analysis of the E2 hypervariable region of the viral genome is in progress to confirm this assumption.

Evaluation of a prototype HCV NS5b assay for typing strains of hepatitis C virus isolated from Tunisian haemodialysis patients.

Hepatitis C virus (HCV) strains isolated from 68 haemodialysis Tunisian patients exhibiting chronic infection were genotyped targeting the NS5b region of the HCV genome using a prototype assay developed by Bayer HealthCare-Diagnostics (TRUGENE NS5b HCV). The overall results were compared to those obtained with another assay of the same company based on sequencing of the 5' non-coding region (TRUGENE HCV 5'NC genotyping kit). All strains could be typed by the 5'NC typing kit, but only 62 (91; 2%) by the NS5b prototype assay. All the 62 strains typed by both methods exhibited the same pattern at the type level: 57 were type 1, 3 were type 2, and 2 were type 4. At the subtype level, eight strains that gave undetermined results by the 5'NC kit were successfully typed by the NS5b kit; eight additional strains exhibited discrepant results. The overall agreement between the two assays was 74.2% at the subtype level. In conclusion, the NS5b region appears to be much more accurate than the 5'NC region to subtype HCV strains, especially in those isolated from patients attending haemodialysis centres where the subtype distribution suggests frequent nosocomial transmissions.