448

with normal than in those with abnormal Alat, these results being obtained with an experimental recombinant immunoblot assay for anti-HCV." The best documented recent prospective study on posttransfusion hepatitis, done in Amsterdam in open heart surgery patients, showed the same diagnostic sensitivity for Alat screening and for anti-HCV (065 and 063, respectively).9 Six other centres contributed results to the Rome meeting.6 Pooling these unpublished results (from the Ortho anti-HCV ELISA evaluators meeting, in Paris on June 30, 1989) decreased the sensitivity of anti-HCV to 046. Furthermore, of 46 recipients with posttransfusion NANBH in whom anti-HCV developed 18 (40%) had received anti-HCV-negative blood only. Among the 55 patients without anti-HCV 44 (80%) had received only anti-HCV-negative blood. No information was provided on the Alat status of donors. Nevertheless, it does seem that today’s anti-HCV antibody test (anti-C-100) misses some infectious donations. Alat screening seems to eliminate units containing a high virus load irrespective of whether or not they are anti-HCV positive. Blood with normal Alat that is anti-HCV positive seems to have a low virus load or none.1o There is compelling evidence supporting Alat screening of blood used for transfusion and for the preparation of plasma derivatives. Additional anti-HCV screening will only increase the safety of individual units. For large plasma pools, however, only Alat screening significantly decreases the virus concentration of the pool; the effect of anti-HCV screening is negligible and may even increase the virus concentration of the pool. Until the impact of eliminating HCV antibodies from the plasma is known with respect to the fractionation behaviour of HCV or HCV/anti-HCV complexes, we see no need to exclude anti-HCVpositive plasma for fractionation purposes in settings where Alat screening is routine. Central Laboratory, Swiss Red Cross Blood Transfusion Service, CH-3000 Bern 22, Switzerland

JOHANN J. BURCKHARDT HANS FRIEDLI ANDREAS GARDI HANS-JÖRG HEINIGER

1. Aach RD, Szmuness W, Mosley JW, et al. Serum alanine aminotransferase of donors in relation to the risk of non-A, non-B hepatitis in recipients: the TransfusionTransmitted Viruses Study. N Engl J Med 1981; 304: 989-94. 2. Alter HJ, Purcell RH, Holland PV, et al. Donor transaminase and recipient hepatitis: impact on blood transfusion services. JAMA 1981; 246: 630-34. 3. Silverstein MD, MUlley AG, Dienstag JL. Should donor blood be screened for elevated alanine aminotransferase levels? A cost-effectiveness analysis. JAMA

1984; 252: 2839-45. Screening for hepatitis infectivity among blood donors: a model for blood safety? Arch Pathol Lab Med 1989; 113: 227-31. 5. Kuo G, Choo QL, Alter HJ, et al. An assay for circulating antibodies to a major etiologic virus of human non-A, non-B hepatitis. Science 1989; 244: 362-64. 6. First International Meeting on Hepatitis C Virus (Sept 14-15, 1989, Rome) (abstr). 7. 1990 International Symposium on Viral Hepatitis and Liver Disease (April 5-8, 1990, Houston); abstr 444, 452, 472, 481. 8. Morgan C, Hyland C, Young IF. Hepatitis C antibody and transaminase activities in blood donors. Lancet 1990; 335: 921. 9. Van der Poel CL, Reesink HW, Schaasberg W, et al. Infectivity of blood seropositive for hepatitis C antibodies. Lancet 1990; 335: 558-60. 10. Garson JA, Tedder RS, Briggs M, et al. Detection of hepatitis C viral sequences in blood donations by "nested" polymerase chain reaction and prediction of infectivity. Lancet 1990; 335: 1419-22. 4. Dodd RY.

Hepatitis C virus RNA in symptomless donors implicated in post-transfusion non-A, non-B hepatitis SIR,-Dr Garson and colleagues (June 16, p 1419) demonstrate the use of the polymerase chain reaction (PCR) to distinguish between infectious and non-infectious blood donors who have antibody to hepatitis C virus (HCV). Here we report, as a part of a prospective study initiated in January, 1988, the detection of HCV-RNA sequences by PCR in the sera of seven regular blood donors associated with three cases of post-transfusion non-A, non-B

hepatitis (P-NANBH). The implicated donors had no history of exposure to HCV risk factors and transaminase activities (ALT) were persistently normal, even after their blood came under suspicion. NANBH in a transaminase recipient was defined as an increase in ALT greater than 2-5 times the upper limit of normal at least twice during

Detection of HCV RNA in donors implicated in post-transfusion NANBH. PCR products hybridised with 32P-labelled oligonucleotide internal to amplified sequence. + =positive control (acute hepatitis C); - =negative control (distilled water): a donor 3; b donor 2; c donor 1. (Lower molecular weight of cDNA amplified from donor 1 is probably due to incomplete denaturation of RNA template.) =

=

=

and by exclusion of other hepatotropic viruses. AntiHCV was sought in sera stored at -20°C by ELISA and recombinant immunoblot assay (RIBA, Ortho Diagnostics). RNA was extracted from 200 ul of serum, reverse-transcribed into cDNA and amplified by PCR. The sequences of the outer primers were 5’-GCATGTCATGATGTAT-3’ (anti-sense) and 5’ACAATACGTGTGTCAC-3’ (sense).’1 Forty cycles of denaturation for 1 min at 94°C, annealing for 1 min at 45°C, and extension for 2 min at 72°C were done. Sera from donors 1 (M, 62) and 2 (M, 37) gave borderline anti-HCV reactivities by ELISA (absorbance/cut-off ratios 106 and 1-18, respectively). The reactivity of serum from donor 1 was not confirmed on the repeat testing. Serum from donor 3 (M, 40) was repeatedly negative. On RIBA, sera from donor 1 scored as weak reactive (1 + to both C-100 and 5-1-1 antigen) while those from donors 2 and 3 were indeterminate. Reverse transcribed HCV-RNA sequences could be amplified from all three donors (figure). The specificity of the PCR products was confirmed by independent hybridisation with two different oligonucleotides internal to the amplified sequence. The presence of HCV in the blood of these three donors was confirmed by their involvement in NANBH transmission. Recipients 1 (F, 36 y) and 3 (F, 43 y) had acute NANBH 6 and 1111 weeks, respectively, after transfusion, and anti-HCV antibodies appeared 2 and 9 weeks after onset of the disease. These two patients had received blood from four other donors, all anti-HCV andPCR negative. Recipient 2 (M, 36) had a mixed ALT (153 IU/1) 14 weeks after transfusion. The ALT returned to normal one month later and no seroconversion to anti-HCV was observed during 8 months of

follow-up

follow-up. Garson

et

al

note

that HCV antibodies

can

be detected in

non-infectious, HCV-RNA negative donors. Our results, on the other hand, provide evidence for the existence of symptomless infectious HCV carriers even in the absence of detectable antiHCV. These data confirm that hepatitis C can be transmitted from donors negative for anti-HCV2 and emphasise the need of methods to detect HCV viraemia directly. We thank Dr M.

Houghton (Chiron Corporation) for supplying the

oligonucleotides. Institution of Virology, University of Milan, 20133 Milan, Italy

A. R. ZANETTI E. TANZI G. ZEHENDER

Blood Transfusion Centre, Hospital of Tradate

E. MAGNI C. INCARBONE

CONBIOTEC,

A. ZONARO D PRIMI E. CARIANI

Hospital of Brescia 1. Weiner

AJ, Kuo G, Bradley DW, et al. Detection of hepatitis C viral sequences in non-A, non-B hepatitis. Lancet 1990; 335:1-3. 2 van der Poel CL, Reesink HW, Lelie PN, et al. Anti-hepatitis C antibodies and non-A, non-B post-transfusion hepatitis in the Netherlands. Lancet 1989; ii: 297-98.

Hepatitis C virus RNA in symptomless donors implicated in post-transfusion non-A, non-B hepatitis.

448 with normal than in those with abnormal Alat, these results being obtained with an experimental recombinant immunoblot assay for anti-HCV." The b...
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