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Saudi Journal of Kidney Diseases and Transplantation
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RENAL DATA FROM THE ARAB WORLD Table of Contents   
Year : 2000  |  Volume : 11  |  Issue : 2  |  Page : 218-222
Hepatitis C Virus Antibodies in Dialysis Patients in Tunisia: A Single Center Study


1 Laboratoire d’immunologie, Hôpital Charles Nicolle, Tunisia
2 Service de Nephrologie et de Médecine Interne, Hôpital Charles Nicolle, Tunisia
3 Centre d’hémodialyse, Boukornine, Tunisia

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   Abstract 

Fifty-eight patients on maintenance hemodialysis in a dialysis unit at Tunis, Tunisia were tested for anti-hepatitis C virus (anti-HCV) antibodies by second generation ELISA test, and for HCV-RNA by nested reverse transcriptase polymerase chain reaction (RT-PCR) of 5' non-coding region. Specificity of the antibodies was confirmed by immunoblot test. HCV genotype was defined using INNO-LIPA test. Twenty-seven out of 58 patients (46.5%) were reactive by ELISA. Transaminase levels were assessed over a six-month period and showed normal average values. Fourteen of the 27 anti-HCV positive patients (51%) were positive by RT-PCR. Type 1b HCV genotype was the most prevalent, seen in all the dialysis patients and one patient in addition, was co­infected with genotype 4. There was a significant correlation between the duration on dialysis (over five years) and the prevalence of anti-HCV-positive patients (P<0.005) while no correlation existed between the number of blood transfusions and the presence of anti­HCV antibodies. The present study illustrates the high prevalence of HCV infection among Tunisian dialysis patients (51%) and indicates that the spread may be nosocomial rather than transfusion-related.

Keywords: Hemodialysis, Hepatitis C, Tunisia.

How to cite this article:
Sassi F, Gorgi Y, Ayed K, Abdallah T B, Lamouchi A, Maiz H B. Hepatitis C Virus Antibodies in Dialysis Patients in Tunisia: A Single Center Study. Saudi J Kidney Dis Transpl 2000;11:218-22

How to cite this URL:
Sassi F, Gorgi Y, Ayed K, Abdallah T B, Lamouchi A, Maiz H B. Hepatitis C Virus Antibodies in Dialysis Patients in Tunisia: A Single Center Study. Saudi J Kidney Dis Transpl [serial online] 2000 [cited 2019 Oct 14];11:218-22. Available from: http://www.sjkdt.org/text.asp?2000/11/2/218/36683

   Introduction Top


Hepatitis viruses have become the major infectious problems in patients on main­tenance hemodialysis (HD). Introduction of a preventive strategy by hepatitis B vaccination has considerably reduced the trans­mission of hepatitis B virus among these patients. Of late, hepatitis C virus (HCV) has become the major cause of acute and chronic hepatitis in this patient group. A significantly increased prevalence (12%) of antibodies to HCV (anti-HCV) was initially noted in HD patients by Jeffers et al. [1] This finding was subsequently confirmed by reports from different countries with prevalence rates ranging between 2 and 68%. [2],[3],[4]

The prevalence of anti-HCV in Tunisian blood donors and dialysis patients has been previously reported. [5],[6] However, there are no published data on the prevalence of HCV­RNA positivity or the clinical features of hepatitis C in Tunisian dialysis patients. This study was performed to find out the prevalence of anti-HCV-positive and HCV­RNA-positive patients in a Tunisian dialysis center and as well as the influence of viremia and host factors on serum amino-transferase values. Also, since there is growing evidence in HCV affected patients for a major influence of HCV genotype on the therapeutic efficacy of interferon alpha, we studied the HCV genotype in our HCV-RNA positive patients.


   Patients and Methods Top


Our study was conducted on 58 patients on maintenance HD in a dialysis unit at Tunis. There were 35 males and 23 females with age ranging from 16 to 77 years. Thirty-seven (63%) had history of blood transfusion(s). Forty-five patients were vaccinated against hepatitis B virus and two were HBsAg positive. The following data were collected on all the study patients: age and sex, dialysis details, hepatitis B status, blood transfusion and liver function tests over a six-month period.

All the patients were screened for anti­HCV using second generation ELISA assay (MONOLISA anti-HCV PLUS, Pasteur) which detects antibodies against core antigens and non-structural antigens encoded by the NS3 and NS4 regions. All ELISA anti­HCV-reactive samples were tested for anti­HCV specificity by a recombinant immunoblot assay (DESISCAN HCV PLUS, Pasteur). HCV-RNA was searched for by a 5' non-coding region nested­polymerase chain reaction (PCR) and subsequent genotyping was performed using the INNO-LIPA probe.

Extraction of the HCV-RNA and Synthesis of cDNA

50 ml of the serum was mixed with 400 µl of RNAZOL (Bioprobe System, RNA-BTM, ref. RNA B 03) and incubated for 10 minutes in ice. After lysis and denaturation, 45 µl of chloroform was added. The mixture was incubated for 20 minutes in ice and centrifuged following which 250 µl of cold isopropanol was added to the supernatant. This solution was incubated all night long at -20°C and then centrifuged. The ensuing precipitate was washed in 500 µl of cold ethanol followed by centrifugation. The precipitate was then dried in a speed vac and dissolved in 10 µl of Diethyl pyro­carbonate 0.1% DEPC/H 2 O and 2 µl of random primers (pDN 6 ), which constitute the non-specific universal primers used for cDNA synthesis. cDNA was synthesized using the reverse transcriptase, Avian Myeloblastosis Virus-Reverse Transcriptase (AMV-RT). The amplification of the 5' non-coding region of cDNA was achieved according to the manufacturer (INNO-LIPA II AMPLIFICATION, Inno-genetics).

The serum alanine amino transferase (ALT) levels were estimated on a monthly basis for a total period of six months. To minimize laboratory errors, measurement was made in one setting for all the six collected samples of each patient.


   Results Top


Anti-HCV Antibodies

Anti-HCV, assessed by second generation ELISA tests, was positive in 27 patients (46.5%). No correlation was noted between the presence of these antibodies and the age and sex of the patients. The sero-positive and negative patients' average age was 48 years and 49 years respectively. Among the anti-HCV-positive patients, 16 had received varying number of blood transfusions, nine of whom were transfused before 1995, when mandatory anti-HCV screening of blood donors was first introduced in Tunisia. The other 11 patients had not been transfused at any time.

The association between the number of transfusions, duration on dialysis and the presence of anti-HCV antibodies was analysed. There was significant association only between the period on dialysis and prevalence of anti-HCV-positivity (P<0.05). Of the 27 anti-HCV positive patients, 20 (74.1%) had been on HD for more than five years, as against only eight of the 31 anti-HCV negative patients (25.8%).

The serum ALT levels, serially measured were normal in both HCV-positive and negative patients.

Search for the Viral RNA in Serum

Simple PCR for HCV RNA from sera was negative in all patients. The application of the nested-RT-PCR gave 14 positive results among the 27 anti-HCV-positive patients (51%), 10 of whom had been transfused, and four others who had not been transfused at any time. None of the anti­HCV-negative patients had positive PCR.


   Discussion Top


We studied sera of patients obtained from one center of HD in Tunis for anti-HCV using a combination of immunoassays, immunoblot and nested-PCR. The prevalence of anti-HCV in the study patients was 46.5% as compared to the prevalence in the general population of 0.5-1%. [5] Prevalence of anti-HCV in European countries is about 20%, with marked differences between different dialysis units (5-47%). [7],[8] The high prevalence in our center may be related to various risk factors including blood transfusion, partial immunodeficiency, frequent parenteral manipulations and the high concentration of infected patients in dialysis units.

In HD as well as transplanted anti-HCV­positive subjects, the increase of transa­minases, particularly ALT, is unpredictable. [9] Persistence of elevation of transaminases has been observed in only 30% of the anti­-HCV-positive patients. [10],[11] A lower preva­lence has been observed in certain other studies. In the study of Olmer et al on 114 hemodialysed patients, only one patient presented with elevated ALT level. [12] In our study, monthly measurement over a period of six months did not show increased ALT levels in any patient.

The mode of transmission of HCV is still not clear and represents one of the major management problems faced by dialysis units. We did not find any correlation between blood transfusions and the frequency of infection by HCV. However, there was close correlation between the number of anti-HCV-positive patients and the duration on dialysis. This makes a case for nosocomial transmission of HCV in dialysis units where the number of infected patients is high and where the material management does not take into account the patient's viral status.

The presence of anti-HCV was associated with detectable viremia in 51% of our study patients testifying a viral replication, and this was associated with normal levels of transaminases. It is important to note that HCV viremia can fluctuate in hemodialysed patients, regardless of an inherent technical problem to the PCR. [13] Also, the serological results for HCV do not allow specifying the infectiousness of the patient. Even after recovery, antibodies can persist for several years. Only a liver biopsy would allow one to assess histological score and the develop­ment of chronic viral hepatitis.[14] In the dialysed patient who is likely to benefit from interferon treatment, the serological result is not a predictive factor of response to treatment.

The determination of the HCV virus genotype in patients on HD appears useful in assessing the epidemiology of a dialysis center by the evidence of "cluster". Also, studies have shown that patients infected by genotypes other than 1b respond better to interferon alpha treatment than patients

with 1b HCV infection. [15],[16] In our study, the most frequently encountered genotype was type 1b, although we had a patient co­infected with 1b and 4. It is important to note that genotype 4 is reported to be absent in blood donors in Tunisia. It is generally present in the Middle East and in Central Africa. The existence of a second genotype in our patient would suggest a reinfection or co-infection by HCV.

In conclusion, our study indicates that dialysis patients may be grouped into three categories with respect to HCV: (i) patients with no viremia and with an immune response, (ii) patients with viremia and immune response, (iii) and patients with no viremia and no immune response. All these patients are usually asymptomatic with regard to liver disease. The first two categories may reflect the different stages of HCV infection and imply that detection of both anti-HCV antibodies and HCV­ RNA are needed for an adequate diagnosis.

 
   References Top

1.Jeffers LJ, Perez GO, De Medina MD, et al. Hepatitis C infection in two urban hemo­dialysis units. Kidney Int 1990;38:320-2.  Back to cited text no. 1  [PUBMED]  
2.Machida J, Yamaguchi K, Ueda S, et al. High incidence of hepatitis C virus anti­bodies in hemodialysis patients. Nephron 1992;60:117-8.  Back to cited text no. 2  [PUBMED]  
3.Vitale C, Tricerri A, Marangella M, et al. Epidemiology of hepatitis C virus infection in dialysis units: first-versus second generation assays. Nephron 1993; 64:315-6.  Back to cited text no. 3  [PUBMED]  
4.Simon N, Courouce AM, Lemarrec N, Trepo C, Ducamp S. A twelve-year natural history of hepatitis C virus infection in hemodialyzed patients. Kidney Int 1994; 46:504-11.  Back to cited text no. 4    
5.Gorgi Y, Yalaoui S, Ben Nejma HL, et al. Detection of hepatitis C virus in the general population of Tunisia. Bull Soc Path Exot 1998;91:177.  Back to cited text no. 5    
6.Hachicha J, Hammami A, Masmoudi H, et al. Viral hepatitis C in chronic hemodialyzed patients in Southern Tunisia. Prevalence and risk factors. Ann Med Interne Paris 1995;146(5):295-8.  Back to cited text no. 6    
7.Zeldi JB, Depner TA, Kuramoto IK, Gish RG, Holland PV. The prevalence of hepatitis C virus antibodies among hemo-dialysis patients. Ann Intern Med 1990; 112:958-60.  Back to cited text no. 7    
8.Percentage of anti-hepatitis C positive hemodialysis patients in all European and Mediterranean countries, Vienna 1994, Reports of EDTA (Abst).  Back to cited text no. 8    
9.Courouce AM, Bouchardeau F, Chauveau P, et al. Hepatitis C virus (HCV) infection in hemodialysed patients: HCV-RNA and anti­HCV antibodies (third generation assays). Nephrol Dial Transplant 1995; 10:234-9.  Back to cited text no. 9    
10.Pol S, Romeo R, Zins B, et al. Hepatitis C virus RNA in anti-HCV positive hemo­dialyzed patients; significance and thera­peutic implications. Kidney Int 1993; 44(5):1097-100.  Back to cited text no. 10    
11.Chan TM, Lok AS, Cheng IK, Chan RT. Prevalence of hepatitis C virus infection in hemodialysis patients: a longitudinal study comparing the results of RNA and anti-body assays. Hepatology 1993;17:5-8.  Back to cited text no. 11  [PUBMED]  
12.Olmer M, Bouchouareb D, Zandotti C, de micco P, de Lamballerie X. Transmission of the hepatitis C virus in an hemodialysis unit: evidence for nosocomial infection. Clin Nephrol 1997;47:263-70.  Back to cited text no. 12  [PUBMED]  
13.Dussol B, de Lamballerie X, Brunet P, et al. Is hepatitis C virus-RNA detection by nested polymerase chain reaction clinically relevant in hemodialysis patients? Clin Nephrol 1996;45:257-60.  Back to cited text no. 13  [PUBMED]  
14.Al-Wakeel J, Malik GH, Al-Mohaya S, et al. Liver disease in dialysis patients with antibodies to hepatitis C virus. Nephrol Dial Transplant 1996;11:2265-8.  Back to cited text no. 14    
15.Nousbaum JB, Pol S, Nalpas B, et al. Hepatitis C virus type 1b (II) infection in France and Italy. Ann Intern Med 1995; 122:161-8.  Back to cited text no. 15  [PUBMED]  [FULLTEXT]
16.Martinot-Peignoux M, Marcellin P, Pouteau M, et al. Pretreatment serum hepatitis C virus RNA levels and hepatitis C virus genotype are the main and inde-pendent prognostic factors of sustained response to interferon alpha therapy in chronic hepatitis C. Hepatology 1995;22: 1050-6.  Back to cited text no. 16  [PUBMED]  

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Correspondence Address:
K Ayed
Laboratoire d’immunologie, Hôpital Charles Nicolle, Boulevard du 9 Avril (1006)
Tunisia
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PMID: 18209319

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    Abstract
    Introduction
    Patients and Methods
    Results
    Discussion
    References
 

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