CFTR Mutations in Three LatinAmerican Countries

C.M. Restrepo,1 L. Pineda,2 A. Rojas-Martınez,3 C.A. Gutierrez,1 A. Morales,2 Y. Gomez,1M.C. Villalobos,3 L. Borjas,2 W. Delgado,2 A. Myers,1 and H.A. Barrera-Saldana3*1Unidad de Genetica, Instituto de Ciencias Basicas, Facultad de Medicina, Universidad Colegio Mayor de NuestraSenora del Rosario, Bogota, Colombia

2Unidad de Genetica Medica, Facultad de Medicina, Universidad del Zulia, Maracaibo, Zulia, Venezuela3Departamento de Bioquımica, Facultad de Medicina, Universidad Autonoma de Nuevo Leon, Monterrey,Nuevo Leon, Mexico

We analyzed 192 cystic fibrosis (CF) allelesin three Latin American countries: Mexico,Colombia, and Venezuela. Mutation screen-ing was performed by polymerase chain re-action (PCR) and a reverse dot blot detec-tion kit that enables determination of 16 ofthe most common CF mutations worldwide.Mutations were detected in 47.9% of thescreened CF alleles. The most prevalent CFallele was DF508 (39.6%). The remaining 16non-DF508 detectable mutations repre-sented 8.3% of the CF alleles. Among them,the G542X, N1303K, and 3849+10kb C>Twere the most common. Although the fre-quency of DF508 described here is lowerthan that reported for Caucasian popula-tions, including in Spain, it is remarkablethat mutation prevalences found in thisstudy resemble those observed in Spain.Two of these mutations, G542X and3849+10kb C>T, that were relevant in thisanalysis, have a particularly high incidencein Spanish communities. The low frequencyof DF508 described here may be explainedby the Amerindian, Caucasian, and Blackadmixture that occurred in Latin Americaafter the discovery of the New World, andalso by the probable occurrence of muta-tions contributed by the original natives,which were undetectable in this analysis.Am. J. Med. Genet. 91:277–279, 2000.© 2000 Wiley-Liss, Inc.

KEY WORDS: cystic fibrosis; allele/muta-tion frequencies; ethnicgroups; Latin America

INTRODUCTION

Cystic fibrosis (CF) is the most common autosomalrecessive disease in Caucasian populations [Boat et al.,1989]. More than 800 mutations/polymorphisms havebeen described since the positional cloning, identifica-tion, and sequencing of the cystic fibrosis transmem-brane regulator gene (CFTR) 10 years ago [CFGACweb site]. The DF508 mutation, a 3 basepair (3 bp)deletion, results in the loss of the phenylalanine resi-due at position 508 of the protein CFTR, and is themost frequent CF-causing mutation [CFGAC, 1990].This mutation accounts for approximately 70% of allCF mutations and it is found particularly in Cauca-sians [CFGAC, 1990]. The remaining 30% are com-prised of more than 800 other mutations, genericallynamed non-DF508, affecting Caucasians and otherethnic groups [CFGAC web site]. The DF508 allelefrequencies show a northwest-southeast gradientdistribution across Europe ranging from 80–40%, re-spectively [CFGAC, 1990; Romeo and Devoto, 1990].Published data for CF mutations in Latin Americanpopulations are scarce. They include some reports fromArgentina [CFGAC, 1990; Aulehla-Scholz et al., 1990],Brazil [Raskin et al., 1999], Cuba [Collazo et al., 1995],Mexico [Rojas-Martınez et al., 1992; Orozco et al., 1993;Villalobos-Torres et al., 1997], some studies of Hispanicpatients in the USA [Grebe et al., 1994; Arzimanoglouet al., 1995], and other isolated reports. The LatinAmerican populations are mixtures from immigrantscoming originally from Asia and the Pacific Islands(Amerindians) and, more recently, from Europe andAfrica, after Columbus discovered America. Spaniardsbrought the major Caucasian background present to-day in the Spanish-speaking countries of LatinAmerica. It is possible that the recent immigrants mayhave introduced new CF alleles, changing the allelefrequencies in the region. The present work determines

Grant sponsor: “Catedras Patrimoniales de Excelencia II” fromthe National Council for Science and Technology (CONACYT).

*Correspondence to: Dr. Hugo A. Barrera-Saldana, Departa-mento de Bioquımica, Facultad de Medicina, Universidad Au-tonoma de Nuevo Leon. Av. Francisco I. Madero y Calle Dr. Edu-ardo Aguirre Pequeno, Col. Mitras Centro, CP 64460. Monterrey,Nuevo Leon, Mexico. E-mail: hbarrera@uanl.mx

Received 7 June 1999; Accepted 4 January 2000

American Journal of Medical Genetics 91:277–279 (2000)

© 2000 Wiley-Liss, Inc.

the frequencies of 16 CFTR mutations in three Span-ish-speaking Latin American countries, and comparesthese frequencies with the CF allelic frequencies re-ported for Spain.

MATERIALS AND METHODS

The present study analyzed 96 unrelated LatinAmerican patients with CF diagnosis, distributed asfollows: 45 patients from Monterrey, Mexico (90 chro-mosomes), 24 patients from Bogota, Colombia (48 chro-mosomes), and 27 patients from Maracaibo, Venezuela(54 chromosomes). All patients were ascertained in ge-netic services in each city. The CF diagnosis was es-tablished by clinical findings of chronic pulmonary and/or gastrointestinal disease, with at least two abnormalsweat chloride tests. Blood samples were drawn frompatients in whom clinical results were highly sugges-tive of CF.

Genomic DNA from whole blood was isolated fromleukocytes as previously described [Villalobos-Torres etal., 1997]. The isolated DNA from each patient wasamplified by polymerase chain reaction (PCR) using akit for reverse dot blot detection of 16 common CF mu-tations: DF508, R553X, G542X, G551D, N1303K,W1282X, R117H, R334W, R347P, A455E, DI507,1717-1 G>A, R560T, 3849+10kb C>T, 621+1 G>T,S549N [Villalobos-Torres et al., 1997]. This kit in-cluded the respective pairs of primers to analyze thedifferent mutated segments along the CF gene. Thebiotin-labeled amplicons were denatured and hybrid-ized with immobilized allele-specific oligonucleotidesprobes (ASOs) as described previously [Villalobos-Torres et al., 1997].

RESULTS

The present study characterized the molecular defectin 192 CF alleles from Colombia, Mexico, and Venezu-ela using a panel for ascertainment of the 16 most com-mon CFTR mutations worldwide. Results of this studyare shown in Table I. The overall diagnostic efficacy ofthe PCR-ASO probe approach for the detection of CFmutations in the analyzed patients was 47.9% (33.3,45.8, and 57.8% for Venezuela, Colombia, and Mexico,respectively). The most common mutation was theDF508 deletion, with an overall frequency of 39.6%,

ranging from 47.8% in Mexico to 29.6% in Venezuela.Detectable non-DF508 mutations accounted for 8.3% ofthe CF alleles in the sample. These detectable non-DF508 alleles were distributed as follows: 3.7% in Ven-ezuela, 10.0% in Mexico, and 10.4% in Colombia. Thenext three most common alleles determined in the pre-sent study were G542X (4.7% present in the threecountries studied), N1303K (1% present in Mexico andColombia), and 3849+10kb C>T (1% present only intwo Mexican CF chromosomes). Besides DF508 andG542X, no other CF alleles were detected in the Ven-ezuelan sample. In addition to these alleles, only threeother CF mutations were detected: 621+1 G>T, S549N,and W1282X. The G551D mutation, the third mostcommon mutation worldwide, was not found in thisanalysis. Unknown mutations (undetectable with thekit used) represent 52.1% of the CF alleles in thisstudy. They ranged from 42.2–66.7% in the analyzedcountries.

DISCUSSION

The distribution of CF mutations reported here fol-lows the trend of Caucasian populations, in which theDF508 frequency is overrepresented and the other mul-tiple CF mutations account only for very small percent-ages. But the average frequency of DF508 mutation inthe studied countries is lower than the frequency es-tablished for North America and Europe (about 66%)[CFGAC, 1994]. Differences in the prevalence of DF508among the analyzed countries are notorious. Theprevalence found in Mexico is very close to those fre-quencies of about 50% reported in Spain, the USA, andBrazil (Table II) [Casals et al., 1997; Grebe et al., 1994;Arzimanoglou et al., 1995; Raskin et al., 1999]. Preva-lences of DF508 found in Colombia and Venezuela areclose to those of 39% and 34% found in a previous Mexi-can study and in Cuba, respectively [Orozco et al.,1993; Collazo et al., 1995].

G542X mutation, the second most common alleleworldwide, has a higher prevalence in Colombia (6.3%).In Mexico, the prevalence found is about half that of8.4% found in Spain [Casals et al., 1997], but a previ-ous study of this mutation in Mexican CF patients re-vealed a frequency of 7.2% [Villarreal et al., 1996].N1303K and 3849+10kb C>T alleles, the third andtwelfth most common CF mutations in the world [CF-GAC, 1994], represent the third most common allelesin our study. The three additional mutations found(W1282X, 621+1G>T, and S549N) represent only 1.5%of the CF allele occurrence in this analysis.

This study suggests an important Spanish contribu-tion for the 47.8% of CF mutations ascertained in thesethree countries, but it also demonstrates regional dif-ferences in allele prevalence. Besides the differences inDF508 occurrence, the allele frequencies of G542X andN1303K mutations are similar for Mexico and Colom-bia, but differ from those found in Venezuela. Thesedifferences in ethnic background have been outlinedbefore in studies in the USA when countries of origin ofCF chromosomes were analyzed for the southwesternand eastern US-Latin patients [Grebe et al., 1994;Arzimanoglou et al., 1995]. Here, we suggest a more

TABLE I. Frequencies and (Chromosome Number) of CFMutations Detected in Three Latin American Countries

CF mutationMexico% (90)

Colombia% (48)

Venezuela% (54)

Total% (192)

DF508 47.8 (43) 35.4 (17) 29.6 (16) 39.6 (76)G542X 4.4 (4) 6.3 (3) 3.7 (2) 4.7 (9)N1303K 1.1 (1) 2.1 (1) 0.0 (0) 1.0 (2)3849 + 10kb

C > T 2.2 (2) 0.0 (0) 0.0 (0) 1.0 (2)W1282X 0.0 (0) 2.1 (1) 0.0 (0) 0.5 (1)621 + 1 G > T 1.1 (1) 0.0 (0) 0.0 (0) 0.5 (1)S549N 1.1 (1) 0.0 (0) 0.0 (0) 0.5 (1)Non-DF508a 10.0 (9) 10.4 (5) 3.7 (2) 8.3 (16)Unknown 42.2 (38) 54.2 (26) 66.7 (36) 52.1 (100)

aDetected with the 16 CF mutation panel in this study.

278 Restrepo et al.

complicated picture, resulting from subregional differ-ences in ethnic background that are common inside thethree studied countries. Although there are no confi-dent demographic studies to make these determina-tions, Monterrey, Mexico, is a city with a predomi-nantly Spanish-Native background (Mestizo), perhapssimilar to the background prevalent in Bogota, Colom-bia. Maracaibo, Venezuela, has a different background,due to an important Black contribution (Mulatto andZambo). A recent published report from Brazil also re-marks these subregional differences [Raskin et al.,1999].

The detection efficacy of the ASO panel used (47.9%)was low. Studies of Latin American patients carriedout in the USA determined 58% and 67% of CF alleles[Grebe et al., 1994; Arzimanoglou et al., 1995]. Thestudy performed in Brazil detected 56% of CF alleleswith a 5-CF mutation panel [Raskin et al., 1999]. Thehigh frequency of undetectable CF mutations (52.1%)in the present study requires additional analyses todetermine their nature and their contribution to thegenetic pool in each country or subregion. The efficacyof the CF-ASO probe methodology used may be in-creased if African mutations, as 3120+1 G>A are in-cluded. This will be of particular relevance for Colom-bia and Venezuela, which have an important Blackinfluence. Additionally, it can be expected that some“CF Amerindian alleles” will be found that currentlyremain undiscovered, which will be important to de-sign new ASO panels for the ascertainment of CF mu-tations and to develop genetic counseling in LatinAmerica.

ACKNOWLEDGMENTS

We thank Dr. Randall Saiki from Roche MolecularSystems for providing the 16 CFTR mutation detectionkits, Dr. Fernando Rivas and Dr. Jose Marıa Cantu atthe Division de Genetica, CIBO, IMSS in Guadalajara,Mexico, for valuable suggestions, and R.M. Chandler-Burns for critical reading of the manuscript. A.R.-M. issupported by the “Catedras Patrimoniales de Exce-lencia II” program from the National Council forScience and Technology (CONACYT). H.A.B.-S. thanksCONACYT and the Global Network for Cell and Mo-lecular Biology (GNMCB) for their support.

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TABLE II. Percentage of CF Mutations Detected in Studies of Latin American Populations*

CF mutation High prevalenceThis report(n 4 192)

Brazil[Raskin et al.,

1999](n 4 172)

US Latins[Grebe et al.,

1994](n 4 129)

US Latins[Arzimanoglou

et al., 1995](n 4 48)

Spain[Casals et al.,

1997](n 4 1280)

Worldwide[CFGAC,

1994](n 4 43,849)

DF508 Caucasian 39.6 47.0 45.7 47.9 53.2 66.0G542X Spain 4.7 5.5 5.4 2.1 8.4 2.4G551D England 0.0 0.2 0.0 2.1 0.3 1.6N1303K Italy 1.0 2.3 0.0 6.3 2.7 1.3W1282X Jew. Ashkenazi 0.5 na 0.8 0.0 0.8 1.2R553X Germany 0.0 0.8 0.8 2.1 0.3 0.7621 + 1 G > T French-Canadian 0.5 na 0.0 0.0 0.5 0.7R1162X Italy na na 1.6 0.0 1.6 0.33849 + 10kb C > T J. Sephardic/Spain 1.0 na 2.3 0.0 0.4 0.2R334W Not determined 0.0 na 1.6 6.3 1.6 0.1S549N Not determined 0.5 na 0.0 na na <0.1

*na, not ascertained.

CF Mutations in Latin America 279