Molecular epidemiological study of variegate porphyria (VP) to determine the frequency of the founder gene mutation in South Africa

Koegelenberg, Adriaan Jacobus (2003-03)

Thesis (MSc) -- Stellenbosch University, 2003.

Thesis

ENGLISH ABSTRACT: Variegate porphyria (VP; OMIM 176200) is caused by mutations in the protoporphyrinogen oxidase gene (PPOX) and is inherited as an autosomal dominant trait, displaying incomplete penetrance. Manifestation of VP includes photosensitivity and potentially fatal acute porphyric attacks. The incidence of VP in South Africa is the highest in the world due to a founder effect. In 1960 Geoffrey Dean estimated the frequency of VP to be 0.3% in the Caucasian South African population. However, this estimate is questionable on the grounds of genealogical and biochemical methods used, and as indicated by more recent records from diagnostic laboratories reported here. The aim of this study was to determine the frequency of the VP founder mutation (R59W) within two South African populations (Caucasian and mixed ancestry) by means of a highly specific DNA test. Various methods of blood sample collection, DNA isolation and R59W mutation detection, suitable for a large-scale population-screening study, were evaluated. Blood samples were obtained from 4 072 participants at blood transfusion clinics, pathology clinics and maternity wards from three provinces within South Africa. Blood sample collection on FTA cards (Whatman BioScience) with subsequent DNA isolation and SSCP analysis were found to be the most appropriate methods. Four of the participants tested were positive for the R59W mutation. All four were from the 3 233 Caucasian individuals tested while three of these were Afrikaansspeaking, confirming the high prevalence of the founder mutation in the South African Afrikaans-speaking Caucasian population (Afrikaner), as previously reported. One of the two adult R59W positive participants was unaware of her carrier status, in accordance with incomplete penetrance of the trait recently estimated as 60%. The mother of one of the newborn babies found to be R59W positive was also not aware of VP in the family, indicating that ignorance regarding VP status in South Africa is a matter of some concern. The estimated frequency of 0.12% (4 / 3 233) for the R59W mutation is significantly lower than the frequency of 0.3% estimated previously, and analysis of the different populations sampled yielded interesting results. Only one of 2 093 (0.05%) participants in the Caucasian blood transfusion sample was mutationpositive. It is possible that the sample from this group was not unbiased, as porphyries were discouraged to donate blood in the past. One of 761 (0.13%) Caucasian participants from pathology clinics was mutation-positive. This sample was probably not unbiased either, since suspected porphyries are referred to these clinics by physicians. Two of 379 (0.53%) Caucasian newborn individuals tested positive, yielding a much higher frequency than the previous estimate by Dean. Although the sample size was very small, it was probably unbiased and therefore provides the best estimate. It is suggested that the sample size of this group be increased in order to improve the accuracy of the founder mutation frequency estimation in South Africa and to determine whether the issue of underdiagnosis and the resulting risk of potentially fatal acute attacks should be addressed by appropriate genetic testing in the future.

AFRIKAANSE OPSOMMING: Variegate porfirie (VP; OMIM 176200) word veroorsaak deur mutasies in die protoporforinogeen oksidase geen (PPOX) en vertoon 'n outosomaal dominante oorerwingspatroon met onvolledige penetransie. Die manifestasie van VP sluit fotosensitiwiteit en potensieël noodlottige akute aanvalle in. Die voorkoms van VP in Suid-Afrika is die hoogste in die wêreld as gevolg van 'n stigtereffek. In 1960 het Geoffrey Dean die frekwensie van VP in die Suid-Afrikaanse Kaukasiese populasie beraam as 0.3%. Hierdie beraming word egter bevraagteken op grond van die genealogiese en biochemiese metodes wat gebruik is, soos aangedui deur meer onlangse bevindinge van diagnostiese laboratoria. Die doel van hierdie studie was om die frekwensie van die VP stigtermutasie (R59W) in twee Suid-Afrikaanse populasies (Kaukasiese en van gemengde herkoms) deur middel van 'n hoogs spesifieke DNS toets te beraam. Verskeie metodes van bloedmonsterinsameling, DNS isolasie en R59W mutasie deteksie, geskik vir 'n grootskaalse populasiesifting studie, is ondersoek. Bloedmonsters van 4 072 vrywilligers by bloedoortappingsklinieke, patologieklinieke en kraamsale van drie provinsies binne Suid-Afrika is verkry. Daar is gevind dat bloedmonsterversameling op FTA kaartjies (Whatman BioScience) met gepaardgaande DNS isolasie en SSCP analise die mees effektiewe metodes is. Vier van die individue wat getoets is, was positief vir die R59W mutasie. AI vier was afkomstig van die 3 233 Kaukasiese monsters, terwyl drie Afrikaanssprekend was, en dus die hoë voorkoms van die stigtermutasie in die Suid-Afrikaanse Afrikaansspekende Kaukasiese bevolking (Afrikaner) bevestig. Een van die twee volwasse R59W positiewe individue was onbewus van haar draer status, wat ooreenstem met die onlangse beraming van 'n 60% penetransie vir VP. Die ma van een van die R59W positiewe pasgebore babas was ook onbewus van VP in die familie. Hierdie uitslae impliseer dat onkunde aangaande VP status "n rede tot kommer in Suid-Afrika mag wees. Die beraamde frekwensie van 0.12% (4/3233) vir die R59W mutasie was baie laer as die vorige beraamde frekwensie van 0.3%. Analise van die verskillende monsterpopulasies het egter interessante resultate gelewer. Slegs een van 2 093 (0.05%) deelnemers in die Kaukasiese bloedoortappingsgroep was mutasie-positief. Dit is moontlik dat die bloedmonsters verkry vanaf hierdie bron nie ongeselekteerd was nie, aangesien porfirielyers in die verlede afgeraai is om bloed te skenk. Een van 761 (0.13%) Kaukasiese individue verkry van patologieklinieke was mutasie-positief. Hierdie groep was ook nie ongeselekteerd nie, aangesien geneeshere moontlike porfirielyers na hierdie klinieke verwys. Twee van 379 (0.53%) Kaukasiese pasgebore babas het positief getoets, wat "n baie hoër frekwensie is as wat deur Dean beraam is. Alhoewel hierdie monsterpopulasie baie klein was, was dit ongeselekteerd en het dit waarskynlik die mees akkurate beraming gelewer. Daar word dus voorgestel dat die aantal monsters in hierdie groep vermeerder word om "n meer akkurate beraming van die stigtermutasie in Suid-Afrika te verkry en sodoende te bepaal of onderdiagnose en die gepaardgaande risiko van noodlottige akute aanvalle in die toekoms deur gepaste genetiese toetsing aangespreek behoort te word.

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