Bird, A.

Galloway, M

Gibbs, B.

Kannemeyer, J.

Kirsch, A.E.

Robson, S.

Smuts, H.

Tucker, T.J.

Voigt, M.

English

AJOL - African Journals Online

SAMJARTICLES,MRC/UCT liver Research Centre, Departments of Medicine andVirology, University of Cape TownT J Tucker. MS 018M Voigl MS QIB. FCS'lSAJ. MMoo (Med!S Robson. MS OlS. Pt-.o. FCP lSA)J Kannemeyer. O!p Meo TI!C1IR E Kirsch. MS 018. MO. DSc. FCFH Smuts. PhDHepatitis C virus infectionrate in volunteer blooddonors from the WesternCape - comparison ofscreening tests and peRIntroduction. Hepatitis G virus (HGV) antibodyseroprevalence studies overestimate the true infectionrate. No data exist on the incidence of HCV or its clinicalfeatures in blood donors of sub-Saharan Africa.Aims. To establish the true incidence of HCV infection involunteer blood donors in the Western Gape, and comparerisk factors and clinical and biochemical features ofviraemic and non-viraemic subjects.Methods. All donors attending the Western ProvinceBlood Transfusion Service between December 1992 andAugust 1994 were screened prospectively for anti-HCVusing the Abbott second-generation assay. Positivedonors were evaluated clinically and biochemically. Theirsera were examined for HCV-RNA by the polymerasechain reaction (PGR).Results. Of 66314 donors screened,~275(0.41 %) wereanti-HCV-positive. Of these 13.6% were PGR-positive(0.056% ot all donors). PGR-positive patients had morerisk factors for HCV acquisition (P < 0.01), symptoms ofhepatitis (P = 0.02) and clinical sigf)s of liver dis~ase(P = 0.05) and higher alanine (P < 0.0001) and aspartateaminotransferase levels (P < 0.0001) than PGR-negativedonors. However, clinical and biochemical features did notdiscriminate adequately between PCR-positive andnegative donors. Uver biopsies performed in 9 of 13PGR-positive cases showed mild inflammation, but nocirrhosis.significant number of patients were not managed accordingto the protocol; the importance of adequate analgesia andfluid management needs to be re-emphasised.A recent local study has highlighted the limited time manymothers have for child care and how this is often left toolder siblings or grandparents. '3 Children are thus bothvictims and carers. School health education represents anopportunity to emphasise both burn prevention strategiesand first aid. We have started a 'Child-ta-Child' programmewhich recognises the role that many children play in childcare. ,.. In this programme the children are themselves askedto identify the important causes of burns and to designposters and health promotion messages that may reducethe incidence.The series reported here is hospital-based and thereforeunderestimates the true burden of disease in the community.Forjuoh et al. 's have described a rapid epidemiologicalmethod for determining the prevalence and incidence ofburns in the community by using burn scars as a proxymeasure. Such a community survey would be useful toclarify this issue.If health education and prevention interventions are to bea success there is a need for further research to assesspresent knowledge and practice and to determine the extentto which the latter needs to be modified. Interventionsshould target not just mothers but schoolchildren andgrandparents. Our study has highlighted burns as animportant cause of paediatric morbidity and a major sourceof hospital expenditure. This has led us to examine ourmanagement critically and to initiate interventions aimed atreducing the incidence and severity of paediatric burns.REFERENCES1 Manc'awl M. Rome- CJ. AccIdents In chIldren. adolescents and young aDults: Amajor public health problem. World Healfll Srar 0 1986; 39: 227-231.2. Onuba O. Pattern 01 burns InJuf) In NIgerian children. Trap Doer 1988: 18: 106-108.3. UNICEF- ChIldren ana women in SoUlh Afnca: a slluallonal analysIs. Preloria:UNICEF. 1993.-1. Courtnght p. Haile O. Koh,s E. The epIdemIology 01 burns to rural Ethiopia. JEpldemrol Comm Health 1993: 47: 19·22_5. 810ch C. The Bums Handbook. Cape Town: Monlca FaJrall CommunIcatIons.1987.6. King M. Primary Surgery. Vol. 2. O_lord: Oxford Medical PublicatIons. 1992.7. Onuba 0. Udoidlok E. The problems and prevention of burns In developingcountries. Bums 1987: 13: 382-3858. Oarko OF. Wachtel TL. Ward HW. er al. AetIological data on hOSpllalised burnspatIents. Bums 1986: 12: 391,9. Lyngdorf P. Epidemiology 01 severe burns. Bums 1986; 12: .:191.10. Lad,lan AAO. AccidentaJ scalds and burns in infancy and childhOOd. J TropPedratr 1987: 33: 199·202.11. Cheng JCY. Leung KS. Lam lC-L. Leung PC. An analysis of 1 70.:1 burn Injuries inHong Kong children. Burns 1990; 16: 182-184_12 Lmdblad BE. Terl<:elsen CJ. Domest,c burns among children. Bums 1990; 16:254-256_13 Chopra M. Ross F. QualirafJve $rucy of file Causes of Malnutrition ITI a Rural Areaof Sourh Afnca. Report for the World Bank Part,Clpatory PoverTy Assessment_Pretona: World Sank. 1996.14. Chopra M. Herhen C. Mak,ng the hnks oetween health and eaUCatiOn ,n ruralKwalulu~NataJ.Pa~r dehverea at SChool Health Conference. Umversr of lt1eWestern Cape. 20 - 22 Jan 199615 Forjuoh SN. Keyl PM. D,enet-Wesl M, Sm,th GS. Guyer 6. Prevalence and age-spec,fic incidence of ourns In Ghanaian child,en_ J Trop PealalT 1995; 41: 273-277.Accepted 14 Nov 1996.T J Tucker, M Voigt, A Bird, S Robson,J Kannemeyer, M Galloway. A E Kirsch,B Gibbs,H SmutsWestern PrO',ince Blood Transfusion Service, Pinelands, W CapeA Bird. MS ChS. FFPalh (SA)B Gibbs. flMLSM Galloway. DIp Med TeenSAMj Volume 87 No. j .\fay 1997Conclusion. Hepatitis C viraemia was present in 0.056%of blood donors. Only 13.6% of sUbjects with positiveserological tests had evidence of viraemia. Risk factorswere more common in viraemic donors. Viraemic subjectshad clinical and biochemical features of liver disease moreoften than non-viraemic donors. Mild disease activity waspresent in all 9 subjects biopsied.S Afr Med J 1997; 87: 603-605.Hepatitis C virus (HCV) transmission by whole blood (and itscomponents) has been significantly reduced since theintroduction of routine blood donor screening with enzyme-linked immunosorbent assays (ELlSAs) for anti-HCVantibodies.12 The predictive value for viraemia of a positiveELlSA result is affected by both the populationseroprevalence and the method used to confirm viraemia(Bayes' theorem). Although second~ and third-generationimmunoassays have improved sensitivity and specificitycompared with earlier ELlSAs,3.4 these continue to identifypeople without detectable viraemia.1.2 This may in part bedue to the retention of antibodies after clearance of HGV ormay result from nonspecific antibody reactivity. The goldstandard for confirming HCV viraemia is the reverse-transcription polymerase chain reaction (AT-PCR) usingprimers specific for the conserved 5' untranslated region ofHCV. Although recombinant immunoblot assays have beenused to confirm positive ELlSAs, these lack both thesensitivity and specificity of PCR.1.25Accurate estimates of HGV prevalence are essential, aspublic health planning depends on these results.s False-positive anti-HCV results reduce the size of the already smallblood donor pool and, in addition, SUbjects informed of aninitial positive ELlSA result, their families and sexual partnersexperience unnecessary anxiety.Although several seroprevalence studies have beenperformed in the general South African population,7-Q no datacurrently exist on the true incidence of hepatitis C viraemiain the South African blood donor population. Weprospectively studied all blood donors who presented at theWestern Province Blood Transfusion Service (WPBTS)between December 1992 and August 1994 in order todetermine the prevalence of hepatitis G viraemia in thosewith a positive ELlSA. Where possible clinical, biochemicaland additional serological studies were performed and,where indicated, histological evaluation of the liver wasundertaken.Subjects and methodsAll donors attending WPBTS between December 1992 andAugust 1994 were tested for the presence of HCVantibodies using the Abbott second-generation ELlSA(ELlSA-2) (Abbott Laboratories, North Chicago, USA). Anti-HCV-positive donors were referred to the liver clinic, GrooteSchuur Hospital for full clinical and serological assessmentand, where clinically indicated, a liver biopsy. Clinicalevaluation included a detailed history of possible risk factorsfor HCV transmission, such as intravenous drug abuse, pastblood transfusions and tattoos, and a history of priorsymptoms of hepatitis. The Ortho anti-HCV ElISA (OrthoDiagnostic Systems Inc., Aaritan, NJ, USA) was used atGroote Schuur Hospital as a supplementary assay in thereferred donors. Anti-HCV-positive donors who failed topresent to the clinic also had supplementary serologicaltests and PGA performed on their stored sera (stored at-70°C), where sufficient serum was available. In addition, theAbbott ELlSA was compared to the Ortho second- (until04/94) or third-generation ELlSA on all initially reactiveserum samples.HCVamplificationRNA was extracted from 200 IJI serum using Total ANAIsolation Reagent (Advanced Biotechnologies Ltd, London,UK) and tRNA on ice; chloroform/iso-amyl alcohol wasadded in a ratio of 24: 1 and the mixture Vigorously blendedand left on ice for 5 minutes; thereafter the homogenate wascentrifuged at 12 000 rpm for 15 minutes. The supernatantcontaining the ANA was removed and precipitated inisopropanol overnight at -20oG, pelleted, washed twice with75% ethanol, air-dried and resuspended in 20 IJI diethylpyrocarbonate (DepC)-treated ultra-pure water with 20 URNasin (Promega Corp., Madison, WI, USA) per sample. Asingle tube AT-PGR was used to form and amplify HCVcDNA as described previously.10 Both outer and nestedprimers used were those described by Chan et al.,11 whichgave a PGR product of 251 bp; this was visualised on a 2%agarose gel with ethidium bromide staining. The productwas transferred to a nylon membrane and probed with adigoxigenin-Iabelled HCV probe (Boehringer Mannheim,GmbH Biochemica, Mannheim, Germany).StatisticsAesults are presented as the mean and standard deviation(SO), unless otherwise specified. The groups were comparedusing the unpaired t-test for continuous data and the chi-square test for categorical data; a P-value < 0.05 wasconsidered significant.ResultsSerology and PCROf the 66 314 donors screened with Abbott ELlSA-2, 275(0.41 %) were antibody-positive. Of these, 100 (36%)presented to the liver clinic at Groote Schuur Hospital wherethey underwent full clinical evaluation. Stored serum from anadditional 84 anti-HCV-positive subjects who did not presentfor examination was analysed by PCR. Insufficient serumfrom the remaining 91 subjects was available for testing.Twenty-five of the 184 (13.6%) serum samples subjected toPGR analysis were found to be positive. The incidence ofviraemia in the group as a whole was therefore 0.056%.All PCR-positive donors were anti-HCV positive by boththe Ortho and Abbott ELlSA. One hundred and fifty-ninedonors were found to be Abbott ELlSA-positive but PCA-negative. Of these, 50 were positive, 107 negative and 2indeterminate when the Ortho ELlSA was used. It was notpossible to comment on the comparative value of the OrthoELlSA, since it was only used in Abbott ELlSA-2-positivedonors.Valuml8i Na. 5 }tay 199; SAJ\'1)Clinical analysisOne hundred donors were assessed at Groate SchuurHospital, of whom 16 were PGR-positive and 84 PCR-negative. The proportion of PGR-positive subjectspresenting for examination was similar to the PGR-positiveproportion of the group that did not present for examination(16% v. 11%; p; 0.30).Although the number of PGR-positive subjects was small,they had significantly more identifiable risk factors for HCVacquisition and more past symptoms compatible withhepatitis, while a significantly greater number hadhepatomegaly and abnormal clinical findings on examinationthan those who were PGR-negative. Serum alanineaminotransferase (ALl) and aspartate aminotransferase(AST) levels showed significant differences between thePGR-negative and PGR-positive groups (Table I).Table I. Donors with a positive anti·HCV ELI$A with and withoutviraemia showed significant differences in respect of medicalhistory, physical examination and biochemical analysisPGR-positive PGR-negative P-valueRisk factors 37.5% 11.9% 0.01Symptoms 12.5% 1.2% 0.02Hepatomegaly 25% 8.3% 0.05ALT (U/I) 34.9 ± 18.8 16.8 ± 10.3 P < 0.0001AST (U/I) 24.3 ± 10.25 14.4±5.1 P < 0.0001Combinations of EUSA with clinical and biochemicalfindings failed to enhance the ability to predict viraemicdonors. Combining EUSA and transaminase results did notimprove upon ELlSA results alone, as only 62.5% of thePGR-positive donors had ALT levels above the upper limit ofnormal. In addition, 9 donors who were anti-HCV-positivebut PGR-negative had a raised ALT level (4 positive for bothAbbott and Ortho ELlSAs and 5 with Abbott ELlSA posilivityonly).Nine PGA-positive donors were considered by theirphysicians to require a liver biopsy. All biopsies showed mildinflammatory activity and features compatible with HGVinfection, but none showed evidence of cirrhosis. Portaltracts were enlarged in 77%. and bile duct damage wasnoted in 55%. periportal fibrosis in 45% and piecemealnecrosis in 45%. Thirty-three per cent had steatosis and22% poorly formed lymphoid follicles.DiscussionThis stUdy shows that only a small number of Western Capeblood donors with serology suggestive of HCV infectionhave detectable viraemia. The prevalence of antibodies inCape Town donors (0.41 %) is similar to that described involuntary donors in northern Europe and the USA.'.2·12 In ourstudy virus was only detected in 13.6% of seropositivesubjects or 0.056% of all donors. This is similar to a study of287 332 British blood donors by Mulimer et al. where 0.35%of donors had HGV antibodies but only 5% of the latter hadviraemia detected by PCR.2 The poor predictive value ofserology for viraemia may be at least partially due to the lowHGV prevalence in the population we studied (Bayes'SAMJARTICLEStheorem). Other possible reasons for the discrepancy mustinclude donors who retain antibodies to HCV after clearingthe virus. as well as nonspecific binding of antibodies in theELlSA.A small group of the PGR-negative donors (4.8%) hadelevated transaminase levels. Although it is possible that asmall number of these had true HGV infection, this isunlikely given that the sensitivity of our PGA assay isextremely high. Quality control studies at our laboratory.performed as part of the Eurohep2 stud!, showed that ourPGA assay consistently detected as few as 20 genomeequivalents/ml serum. While undetectable hepatitis Cviraemia is possible in these donors, other commonconditions inclUding alcoholic liver disease, non-alcoholicsteatohepatitis or hepatitis G infection'3 are more likely to bethe cause of the elevated transaminase levels.The combination of EUSA results and transaminaseelevations and clinical variables did not assist in thedistinction between patients with and without viraemiabetter than ELlSA alone. The ELlSA- and PGR-positivegroup had significantly higher AST and ALT levels than theELlSA-positive, PGR-negative group, but in both groupsthere was too wide a variation for this finding to be of anysignificant assistance in individual cases. Similarly, althoughclinical symptoms and signs and a history of risk factorswere more common in the viraemic than the non-viraemicdonors, these did not assist the prediction of viraemia.In conclusion, screening for HGV infection by ELlSAsignificantly over-estimates the incidence of HGV infection.All anti-HGV-positive SUbjects should be referred forconfirmation of viraemia prior to appropriate managementand counselling.REFERENCES1. Oow BC. Cootl! I. Manor H. er al. Confirmation of hepatitis C virus antibody inblood donors. J Med Viro11993; 41: 215-220.2. Mutimer DJ. Harrison RF. O·Oonnel KB. El al. Hepat,tis C virus Infection i... theasymptomat,c 8nt,$I1 blOGd donor. J Viral HepaulJ:S 1995; 2: 47-53.3. CrIDll A. Vatensa M. Fab'ano C. e/ al. Third generalion hepat,ns C VirUS tests Inasymptomauc anu·HCV posit,ve blood donOl'$. J Heparo/l994; 21: 730~734.4. McDonald KL M,lIs WA. Wood RC. et aL Evaluation of climcal and·laboraloryaspects of antibooy lesls for detecllon of hepat'!Is C v,rus infectIon trl blooddonors and reclPlen!s fTom a low fisk populatIon. TransfuSion 1994: 34: 202-208.5. Garso", JA. Clewley JP. SImmonds P. er al. Hepatltls C viraemia In Unite<!Kingdom blood donors. Vox Sang 1992; 62: 218-2236. Vo,gt MD. 8ird A. Kirsch RE. er al. National strategy for the prevent,on andmanagemenl of transrusion-associated hepatrllS. S Afr Med J 1996; 86: 245-250.7. Abdool Karim SS. Ta'l OR. Hepat,tis C virus infection in urban and ruralNatallKwaZulu. S Afr Med J 1993: 83: 191-193.8. Son, PN. TaJt DR. Kenoyer DG, et al. HepautlS C Virus antibodies among riskgroups in a South African area endemic for hepat,tis B virus. J Med Viral 1993;40: 65-68.g. Tucker T. Kirsch RE. Louw SJ. lsaacs S, Kannemeyer J. Robson SC. Hepatitis EIn South Alnca: eVidence Ior sporadiC spread anCl Increased seroprevalence Inrural areas_ J Med Viral 1996~ 50: 117·119_10 Smuts H. Kannemeyer J. Genoryplng of hepaulIs C virus In South Atrica_ J ClinMlcrobio/1995. 33: 1679-1681.11. Chan SW. ,",cOmlsh F. Holmes EC, er al. AnalySIS of a new hepatitiS C virus typeanCllts phylogenic relationship to eXlstmg vanants. J Gen Virol 1992; 73: t 131.1141.12 AnClerson se. Halhaway T. Kuramoto 1K. et al. Comparison of two second·generation anti·hepatitis C virus EL/SA on 21431 US blood donor samples. J ViralHepatitiS 1995; 2: 55-6113. Lmnen J, Wages J jun. Zhang-Keck Z-V. er al. Molecular clomng and diseaseaSSOCiatIon of hepaulls G virus: a transfusion-transmIssible agent. SCience 1996;271: 505-508.Accepted 3 Feb 1997.SAM/ "al,m,87 Na. 5 May 1997~

Hepatitis C virus infection rate in volunteer blood donors from the Western Cape - comparison of screening tests and peR

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Hepatitis C virus infection rate in volunteer blood donors from the Western Cape - comparison of screening tests and peR

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