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Saudi Journal of Kidney Diseases and Transplantation
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Year : 2007  |  Volume : 18  |  Issue : 1  |  Page : 107-113
The Impact of Polymerase Chain Reaction Assays for the Detection of Hepatitis C Virus Infection in a Hemodialysis Unit

1 Department of Nephrology and Dialysis, The Al Hada Armed Forces Hospital, Taif, Saudi Arabia
2 Department of Laboratory Medicine and Molecular Pathology, The Al Hada Armed Forces Hospital, Taif, Saudi Arabia

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Hepatitis C virus (HCV) infection is most often diagnosed by detection of antibodies against the virus (HCV Ab). However, it has been reported that some HCV Ab negative patients test positive for HCV-RNA. Over a study period of 30 months, all patients on hemodialysis at the Al Hada Armed Forces Hospital in Taif, Saudi Arabia were tested monthly for HCV Ab and twice per year for HCV-RNA. HCV Ab was tested by a third generation microparticle enzyme immunoassay (MEIA), and HCV-RNA by a qualitative hepatitis-RNA assay, second version (COBAS Amplicor ® PCR), which was recently introduced in the Molecular Pathology Laboratory of our hospital. Of the 180 patients studied, 34 (18.9%) had positive HCV Ab, and of the 146 HCV Ab negative patients, five patients tested positive for HCV-RNA (3.42 %). Our study further finds that, when applying HCV Ab testing only, some patients with HCV viremia may be undetected. For better HCV infection control, routine HCV­RNA testing of dialysis patients should be considered, particularly in areas where the infection is common and in units applying isolation policies.

Keywords: Hepatitis C, Hepatitis C virus antibodies, Hepatitis C-PCR, Hemodialysis, Infection control.

How to cite this article:
Hussein MM, Mooij JM, Hegazy MS, Bamaga MS. The Impact of Polymerase Chain Reaction Assays for the Detection of Hepatitis C Virus Infection in a Hemodialysis Unit. Saudi J Kidney Dis Transpl 2007;18:107-13

How to cite this URL:
Hussein MM, Mooij JM, Hegazy MS, Bamaga MS. The Impact of Polymerase Chain Reaction Assays for the Detection of Hepatitis C Virus Infection in a Hemodialysis Unit. Saudi J Kidney Dis Transpl [serial online] 2007 [cited 2021 Sep 28];18:107-13. Available from: https://www.sjkdt.org/text.asp?2007/18/1/107/31857

   Introduction Top

Hepatitis C virus (HCV) infection is associated with a poor prognosis for survival among dialysis patients. [1],[2] Commonly, HCV is diagnosed by detection of antibodies against the virus (HCV Ab). However, it has been reported that 2.5 to 12% of patients with active HCV infection, as manifested by detectable serum levels of hepatitis C-RNA (HCV-RNA), test negative for antibodies against the virus. [3],[4],[5] In some patient groups, such as immunocompromised patients, in which the antibody response is suppressed, this number might be even higher. In one study from Saudi Arabia, 28% of the dialysis patients who were anti-HCV negative by third generation ELISA tested positive for HCV-RNA. [6]

These findings indicate that significant numbers of patients with active HCV infection will remain undetected when only anti-HCV Ab assays are used, with unforeseen consequences. In our hospital, we recently introduced an HCV-RNA assay to test all patients routinely for HCV infection, in addition to standard hepatitis C serology screening. This study was undertaken to detect the prevalence of HCV-RNA in HCV Ab negative patients.

   Patients and Methods Top

Since mid 2003, blood samples from all patients on hemodialysis (HD) at the Al Hada Armed Forces Hospital in Taif, Saudi Arabia were taken on a monthly basis for detection of anti-HCV Ab, and in addition twice per year for detection of the presence of HCV-RNA. Anti-HCV antibodies were analyzed using a third generation micro­particle enzyme immunoassay (MEIA) (Abbott Axsym ®, Abbott Laboratories, Abbott Park, IL, USA). The sensitivity and specificity of this test, as mentioned by the manufacturer, are 100 % and 99.6 %, respectively. [7] HCV-RNA was analyzed through a qualitative hepatitis-RNA assay (COBAS Amplicor ® PCR), version 2 (Roche Diagnostics, Branchburg, NJ, USA). [8] This assay has a detection limit of approximately 50 copies of HCV-RNA per ml; the number of copies ranges from 60 to 100 per ml for plasma and serum, respectively. [9]

Most of the patients who tested positive with the HCV qualitative assay underwent further quantitative testing with the COBAS Amplicor HCV Monitor TM Test, version 2 (Roche Diagnostics, Branchburg, NJ, USA). [10] The values of this test results are given as IU/ml. All HCV-PCR tests were carried out at the Molecular Pathology Laboratory of our hospital by medical technologists who were trained by Roche to run the reverse-transcriptase PCR assays. The tests are performed in a specially designated area for molecular protocols to prevent cross contaminations.

For the purpose of this study, the duration on dialysis was defined as the time on dialysis both prior to the study and during the study period. The end of the study period was set at December 31, 2005, or earlier if the patient was transplanted, expired, or lost to follow-up (e.g. transferred to another hospital). Patient data, including age (at the end of the study) and duration on dialysis are presented as mean ± standard deviation, are shown in [Table - 1]. The correlations between two parameters (duration on dialysis and number of HCV Ab positive patients) were calculated using the two-tailed Pearson bivariate correlation coefficient. The level of significance was set at p < 0.05.

   Results Top

The number of patients involved during the study period (30 months) was 180 (86 males and 94 females). The ages of the patients ranged from 8 - 103 years, with a mean of 54.63 ± 18.68 years. The duration on dialysis ranged from 1 - 396 months with a mean of 41.26 ± 50.97 months. During the study period, a total of 34 patients (18.9 %) tested positive for HCV Ab. There was a strongly significant correlation between the duration on dialysis and the number of patients with positive HCV Ab (p = < 0.0001).

Of the 146 patients who tested negative for HCV Ab, five had detectable levels of HCV­RNA (3.4 %). The total duration on dialysis of three of these five patients (patient # 1, 2 and 5) was less than six months each. Two of them were kidney transplant patients who had returned to dialysis following chronic rejection; one of them was known to be anti­HCV Ab negative and HCV-RNA positive prior to the start of dialysis. The third patient had rapidly progressive glomerulonephritis due to ANCA-positive vasculitis, and had undergone intensive immunosuppressive treatment, including plasmapheresis, prior to the start of dialysis. The remaining two patients were on dialysis for 50 and 10 months, respectively, each before the start of the study. In one of them (patient # 3), the HCV-RNA test, was negative when performed initially, but six months later, it had become positive. In the second patient (patient # 4), the HCV-RNA test result was positive at inclusion and remained so throughout the observation period.

The HCV Ab results (done monthly) remained negative during the study period in four of the five patients who were initially HCV Ab negative and HCV-RNA positive. Overall, the study period lasted three months in one patient and four months each in two patients, while in the fifth patient it lasted for 10 months. In one patient (patient # 4) the HCV-RNA became "equivocal" at six months. Before the next assay could be done, this patient expired due to a hepatitis-unrelated event (post-operative cardiac arrest). None of the two patients who were already on dialysis at the start of the study had received blood transfusions during the preceding year.

Liver function tests were mildly abnormal in the two patients who had a failed kidney transplant. In the other three patients, the liver function tests remained within normal limits.

   Discussion Top

The prevalence and incidence of HCV infection among patients on dialysis is consistently higher than in the general population, with the prevalence in dialysis patients ranging from 8% in North America to 90% in some countries in the Middle East.[11] There is a considerable amount of evidence to suggest a nosocomial transmission of HCV within the hemodialysis units, although the exact modes of transmission are not fully clarified. A nosocomial transmission of HCV infection in dialysis units has recently been documented using molecular techniques. [12] Known risk factors include duration on dialysis, the number of blood transfusions, breakdown in standard infection control practices, physical proximity to an infected patient, and inadequate sterilization of dialysis machines between patient sessions. [11],[13],[14],[15]

The presence of patients who are HCV Ab negative and HCV-RNA positive might be another risk factor; however, routine testing for HCV-RNA is not available in most dialysis centers and is not included in the most recent guidelines of the Centers for Disease Control (CDC) for preventing transmission of infection among dialysis patients. [15] A possible explanation for the absence of antibodies in a patient with active viremia might be the known suppressed cellular immunity in the dialysis population, causing a reduced ability to mount a detectable antibody response to the virus. In this context, it is known that hepatitis B vaccinations in dialysis patients do not always result in an antibody response (despite doubling of the vaccine dose). [16],[17],[18] A similar lack of immune response to vaccination has also been associated with the malnutrition-inflammation-cachexia syndrome (MICS), which leads to a poor prognosis in dialysis patients. [19]

Failure to detect HCV Ab in an HCV­RNA positive patient might also occur in the so-called "window" period between infection and anti-HCV seroconversion, which might be prolonged in dialysis patients. [20] In some other cases, it may be due to disappearance of anti-HCV antibody despite persistence of viremia.[11] The prevalence of positive HCV Ab patients in our dialysis center was 18.9%. Five patients in the HCV Ab negative group were positive for HCV-RNA. One of them was known to be HCV Ab negative and HCV-RNA positive before dialysis, while two patients were shown to be HCV-RNA positive at the start of the dialysis period; thus, the infection might have been acquired prior to commence­ment of dialysis treatment. However, one patient, who had already been on dialysis for 50 months, became positive for HCV­RNA during the study period, while for another patient, who had been on dialysis for 10 months and HCV-RNA positive and Ab negative at the start of the study, the HCV serology became "equivocal" after six months during the study period, again suggesting a recent infection.

The prevalence of HCV positive serology in our unit was found to be well below the 47.8% reported for dialysis patients in the Kingdom of Saudi Arabia as a whole in 2002.[21] The reason for this difference might be that in our center, in addition to adherence to universal infection control precautions [11],[13],[14],[15] from an early stage, specific policies were developed for dialysis of HCV Ab positive patients to prevent the spread of infection. They include isolation from other patients and the use of designated dialysis machines in these patients. [22] These isolation policies are not recommended as routine strategies to control nosocomial transmission of HCV in dialysis units and are not included in the CDC guidelines. [15] Nevertheless, they were applied in our unit in view of the high prevalence of HCV infection in the area.

Despite these isolation policies, a potential risk for spread of the infection always remains, if some of the HCV-serology negative patients happen to be active carriers of the virus. This might explain why, during the study period, despite the adherence to universal infection control precautions, and the designation of special machines to HCV Ab positive patients, fresh cases of HCV infection, possibly dialysis-related, were seen in our unit. This observation might also be supported by the finding in our study of a strong positive correlation between the duration on dialysis and HCV Ab positivity. Some investigators recommend routine testing for HCV-RNA in all dialysis patients [5], [23], [24] while others recommend it only for the HCV Ab negative patients. [25] The CDC guidelines recommend performing the test only for patients with abnormal liver function tests and negative HCV-serology. [15]

In areas with a high prevalence and incidence of HCV infection, and in view of the signi­ficant morbidity and mortality associated with HCV infection, testing for HCV-viremia might be a valuable tool, despite the high costs, to improve the diagnosis of HCV infection and, by doing so, to reduce the risk of spreading. [5], [20], [23], [24], [26]

   References Top

1.Stehman-Breen CO, Emerson S, Gretch D, Johnson RJ. Risk of death among chronic dialysis patients infected with hepatitis C virus. Am J Kidney Dis 1998; 32(4): 629-34.  Back to cited text no. 1    
2.Nakayama E, Akiba T, Marumo F, Sato C. Prognosis of anti-hepatitis C virus antibody-positive patients on regular hemodialysis therapy. J Am Soc Nephrol 2000; 11(10): 1896-902.  Back to cited text no. 2    
3.Alter MJ, Margolis HS, Krawczynski K, et al. The natural history of community­acquired hepatitis C in the United States. The Sentinel Counties Chronic non-A, non-B Hepatitis Study Team. N Engl J Med 1992; 327(27): 1899-905.  Back to cited text no. 3    
4.Pereira BJ, Levey AS. Hepatitis C virus infection in dialysis and renal trans­plantation. Kidney Int 1997; 51(4): 981-99.  Back to cited text no. 4    
5.Schroter M, Feucht HH, Schafer L, Zollner B, Laufs R. High percentage of seronegative HCV infections in haemodialysis patients: the need for PCR. Intervirology 1997; 40 (4): 277-8.  Back to cited text no. 5    
6.Al Meshari K, Al Ahdal M, Alfurayh O, Ali A, De Vol E, Kessie G. New insights into hepatitis C virus infection of hemodialysis patients: the implications. Am J Kidney Dis 1995; 25(4): 572-8.  Back to cited text no. 6    
7.Abbott Axsym System®, HCV version 3, 2002, Package insert. Abbott Diagnostics Division, Abbott Park, Il, USA.  Back to cited text no. 7    
8.Cobas Amplicor, Hepatitis C Virus test, version 2.0., 2001. Product Insert. Roche Molecular Systems Inc., Branchburg, NJ, USA.  Back to cited text no. 8    
9.Lee SC, Antony A, Lee N, et al. Improved version 2.0 qualitative and quantitative AMPLICOR reverse transcription-PCR tests for hepatitis C virus RNA: calibration to international units, enhanced genotype reactivity, and performance characteristics. Clin Microbiol 2000; 38(11):4171-9.  Back to cited text no. 9    
10.Cobas Amplicor HCV Monitorm Test, version 2.0, 2000. Product Insert. Roche Molecular Systems Inc., Branchburg, NJ, USA.  Back to cited text no. 10    
11.Natov SN, Murthy BV, Pereira BJ. Hepatitis and Human Immunodeficiency virus infections in end-stage renal disease patients. In: Henrich WL, ed. Principles and practice of dialysis, 3 rd ed. Philadelphia, PA, USA: Lippincott Williams & Wilkins, 2004; 323-351.  Back to cited text no. 11    
12.Hmaied F, Ben Mamou M, Saune­Sandres K, et al. Hepatitis C virus infection among dialysis patients in Tunisia: incidence and molecular evidence for nosocomial transmission. J Med Virol 2006; 78(2):185-91.  Back to cited text no. 12    
13.Tokars JI, Arduino MJ, Alter MJ. Infection control in haemodialysis units. In: Berman SJ, Pien FD, guest Eds. Infections in patients with chronic renal failure. Moellering RC, Consulting Ed. Infectious Disease Clinics of North America. Philadelphia, PA, USA: Saunders, 2001; 15 (3): 797-812.  Back to cited text no. 13    
14.Huraib SO. Hepatitis C in Dialysis Patients. Saudi J Kidney Dis Transplant 2003; 14 (4): 442-50.  Back to cited text no. 14    
15.Recommendations for Preventing Transmission of Infections Among Chronic Hemodialysis patients. Centers for Disease Control (CDC), MMWR, April 27, 2001/ 50 (RR05); 1-43.  Back to cited text no. 15    
16.Stevens CE, Alter HJ, Taylor PE, Zang EA, Harley EJ, Szmuness W. Hepatitis B vaccine in patients receiving hemodialysis. Immunogenicity and efficacy. N Engl J Med 1984; 311(8): 496-501.  Back to cited text no. 16    
17.Descamps-Latscha B, Witko-Sarsat V, Jungers P. Infection and immunity in end­stage renal disease. In: Henrich WL ed. Principles and practice of dialysis, 3rd ed. Philadelphia, PA, USA: Lippincott Williams & Wilkins, 2004; 307-322.  Back to cited text no. 17    
18.Mitwalli A. Responsiveness to hepatitis B vaccine in immunocompromised patients by doubling the dose scheduling. Nephron 1996;73(3):417-20.  Back to cited text no. 18    
19.Kalantar-Zadeh K, Miller LG, Daar ES. Diagnostic discordance for hepatitis C virus infection in hemodialysis patients. Am J Kidney Dis 2005; 46(2): 290-300.  Back to cited text no. 19    
20.Schroeter M, Zoellner B, Polywka S, Laufs R, Feucht HH. Prolonged time until seroconversion among hemo-dialysis patients: the need for HCV PCR. Intervirology 2005; 48(4): 213-5.  Back to cited text no. 20    
21.Shobokshi OA, Serebour FE, Al Drees AZ, et al. Hepatitis C virus sero­prevalence rate among Saudis. Saudi Med J 2003; 24 (Suppl. 2): S81-6.  Back to cited text no. 21    
22.Shamshirsaz AA, Kamgar M, Bekheirnia MR, et al. The role of hemodialysis machines dedication in reducing Hepatitis C transmission in the dialysis setting in Iran: a multicenter prospective intervene­tional study. BMC Nephrol 2004; 5(1):13.  Back to cited text no. 22    
23.Fabrizi F, Poordad FF, Martin P. Diagnostic workup of hepatitis C and the patient on maintenance dialysis. Int J Artif Organs 2001; 24(12): 843-852.  Back to cited text no. 23    
24.Kocabas E, Seyrek N, Paydas S, et al. Detection of hepatitis B and C infection by polymerase chain reaction among hemo­dialysis patients. Nephron 2002; 91(1):178-80.  Back to cited text no. 24    
25.Beccari M, Rizzolo L, Ottolenghi A, Sorgato G. Hepatitis C virus screening strategies in haemodialysis units. Nephrol Dial Transplant 2002; 17 (8): 1536-7.  Back to cited text no. 25    
26.Rigopoulou EI, Stefanidis I, Liaskos C, et al. HCV-RNA qualitative assay based on transcription mediated amplification improves the detection of hepatitis C virus infection in patients on hemodialysis: results from five hemodialysis units in central Greece. J Clin Virol 2005; 34(1):81-5.  Back to cited text no. 26    

Correspondence Address:
Magdi M Hussein
Department of Nephrology and Dialysis, Al Hada Armed Forces Hospital, P.O.Box 1347, TAIF
Saudi Arabia
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