RESEARCH ARTICLE

Genetic Screening for Krabbe Disease: LearningFrom the Past and Looking to the FutureMichal Macarov,1,2 Joel Zlotogora,3 Vardiella Meiner,1 Zinab Khatib,2 Vivi Sury,1 Getu Mengistu,1

Ruth Bargal,1 Esther Shmueli,1 Bela Meidan,1 and Marsha Zeigler1*1Department of Human Genetics and Metabolic Diseases, Hadassah, Hebrew University Hospital, Jerusalem, Israel2The Monique and Jacques Roboh Department of Genetic Research, Hadassah, Hebrew University Hospital, Jerusalem, Israel3Department of Community Genetics, Public Health Services, Ministry of Health, Jerusalem, Israel

Received 4 January 2010; Accepted 23 October 2010

In Israel, Krabbe disease is frequent in two Moslem Arab villages

in the Jerusalem area. In this paper we present our experience

of almost four decades with diagnosis of Krabbe disease, carrier

screening and prenatal diagnosis. The screening program is

well accepted by the community, and there is a clear trend

towards premarital testing. The screening program and prenatal

diagnosis have led to a decrease in the incidence of Krabbe

disease from 1.6 per 1,000 live births to 0.82 per 1,000.

� 2011 Wiley-Liss, Inc.

Key words: Krabbe disease; carrier screening; prenatal

diagnosis

INTRODUCTION

Krabbe disease, also known as globoid cell leukodystrophy, is a

severe lethal autosomal recessive neurodegenerative disorder

caused by the deficiency of beta galactocerebrosidase activity. The

incidence of the disorder in the general population is about one

in 100,000 live births [for review, see Wenger et al., 2005]. In Israel,

Krabbe disease is frequent in two Moslem Arab villages in the

Jerusalem area and in a Druze community in northern Israel. In

the two Arab villages there is one founder mutation [D528N]. In

the Druze community the founder mutation is I583S [Zlotogora

et al., 1985, 1991]. In this report we present our experience of almost

four decades in screening for patients and carriers of Krabbe disease

in these two villages.

MATERIALS AND METHODS

The PopulationThe two villages A and B are within one and a half miles from each

other and are part of the Jerusalem municipality. In 2007 they

included 25,000 inhabitants, almost all being Moslem Arabs; in the

same year there were 592 live births in village A and 429 in village B

(State of Israel Ministry of Interior).

Diagnosis of the Disease

Beta galactocerebrosidase activity and mutation analysis are per-

formed as previously described [Wenger and Williams, 1991; Rafi

et al., 1996].

Until 1997 the diagnosis of the disease was based on the enzy-

matic assay; heterozygote screening being problematic due to the

overlap of the enzyme activity between normal subjects and het-

erozygotes. Starting in 1997, after the characterization of a unique

mutation responsible for the disease [Rafi et al., 1996], the molec-

ular test is used for diagnosis and heterozygote determination.

Carrier ScreeningBetween the years 1999 and 2002 a pilot screening program was

performed that was funded by the Israeli Ministry of Science and

Technology and later by the Jerusalem Foundation. Since 2003

the program has been financed by the Israeli Ministry of Health

as part of a nation-wide screening program [Zlotogora et al., 2009].

The screening program was originally aimed at married couples

in which both spouses originate from one of the two villages. The

screening is in general sequential, when one spouse is examined

*Correspondence to:

Marsha Zeigler, Department of Human Genetics, Hadassah, Hebrew

University Hospital, P.O. Box 12000, 91120 Jerusalem, Israel.

E-mail: zeigler@hadassah.org.il

Published online 22 February 2011 in Wiley Online Library

(wileyonlinelibrary.com).

DOI 10.1002/ajmg.a.33815

How to Cite this Article:Macarov M, Zlotogora J, Meiner V, Khatib Z,

Sury V, Mengistu G, Bargal R, Shmueli E,

Meidan B, Zeigler M. 2011. Genetic screening

for Krabbe disease: Learning from the past

and looking to the future.

Am J Med Genet Part A 155:574–576.

� 2011 Wiley-Liss, Inc. 574

and if found to be a carrier, the recommendation is that the other

spouse undergo testing. Carrier couples are referred for genetic

counseling, and are offered the possibility of prenatal diagnosis

either by chorionic villi sampling (CVS) or amniocentesis. Today,

with the advent of new technologies we can offer carrier couples pre

-implantation genetic diagnosis (PGD).

A nurse living in one of the villages is responsible for the

operation and administration of the screening program. She also

draws the blood and brings it to the Department of Human

Genetics, Hadassah Medical Center. Each individual in the carrier

screening is asked to fill out a form that includes age, identification

number and marital status. The turn around time is between 2 to

4 weeks, and the results are given to tested individuals by the nurse.

The genetic counseling for couples at risk is performed in one of the

genetic clinics in Jerusalem, and the prenatal diagnosis is performed

at the Department of Human Genetics, Hadassah Medical Center.

Educational ProgramThe first step in the educational program was to familiarize the

medical personnel and the religious leaders in the villages with the

aims of the screening program. This was done by arranging a number

of informal meetings with them. Lectures at high-schools and educa-

tional information for women were provided by the nurse in charge

of the community outreach. Also individuals visiting the medical

clinics in the villages received information about the program.

RESULTS

Family SurveyBetween 1977 and 1983 four children from village A were diagnosed

to have Krabbe disease by enzymatic assay of galactocerebrosidase

activity. This prompted us to conduct a survey in the village in 1986

[Levy-Lahad, 1987]. At that time the disease was found in seven

families, in whom 11 parents of affected children originated from a

large extended family in village A and the three other parents originated

from village B. As a result of this survey, the incidence of Krabbe disease

in villageA was calculated to be 1.6 affected children per 1,000 live births

between 1973 and 1987 [Zlotogora et al., 1991]. Later the number of

cases from villageB increased,andbetween1989and2007the incidence

of Krabbe disease was similar in the two villages.

Carrier ScreeningIn the 5 years in which the population carrier screening was performed

in the two villages (2003–2007), 1,772 individuals were tested, of whom

149 were found to be carriers. Thus, the calculated carrier frequency of

Krabbe disease in the two villages is 1:12. Only one married carrier

couple was discovered through the screening program. The most

probable explanation is that despite counseling the partner of the

carrier did not always come for examination (no exact data available),

and that approximately 30% of individuals were tested before marriage.

Prenatal DiagnosisBetween 1989 and 2007, 17 carrier couples underwent 58 prenatal

diagnostic procedures; 47 by CVS and 11 by amniocentesis. Sixteen

of these 17 couples had one or more previously affected children.

Thirty-five normal fetuses were diagnosed of which 33 ended in

live births, and two were spontaneously aborted after CVS. Twenty-

three affected fetuses were diagnosed, resulting in termination of

20 pregnancies, and one spontaneous abortion after CVS. Two

affected children were born to couples who decided not to termi-

nate pregnancy after the diagnosis of an affected fetus by amnio-

centesis. One couple had a previously affected child and this was

their last pregnancy. The other couple had no previously affected

children and the spouses were found to be carriers during pregnan-

cy through the program. In the following pregnancy the woman had

a CVS procedure and a healthy child was born.

Postnatal DiagnosisA total of 20 affected infants were born between 1989 and 2007. Ten

affected infants were born in 1989–1998, six were born during the

pilot study in 1999–2002 and four were born between 2003 and

2007 after the initiation of the population carrier screening. The

calculated incidence of Krabbe disease among live births decreased

from 1.66 per 1,000 in 1999–2002 (6/3,600 total live births) to

0.82 per 1,000 in 2003–2007 (4/4,876). Of the 20 affected infants,

two were diagnosed during pregnancy, and one was born to a couple

that had not been properly referred to genetic counseling. The man,

who was found to be a carrier was advised by his family physician

not to marry within the family. His wife, being from another family

in the same village was therefore not examined.

DISCUSSION

The Department of Human Genetics at Hadassah Medical Center

has been involved with the diagnosis and prevention of Krabbe

disease in the Jerusalem area for almost four decades. The large

number of affected children born during these years allowed

the medical personnel involved with the population to make early

diagnoses according to the first symptoms, place of birth and

galactocerebrosidase activity. Before the characterization of the

founder mutation in this population, the only way to prevent

the disease was for couples at increased risk to undergo an invasive

prenatal diagnosis and termination of affected pregnancies. Most of

the couples who had a previously affected child used this option and

33 normal children were born to couples in whom a prenatal

diagnosis test was normal. There was only one couple that opted

for prenatal diagnosis before the birth of an affected child. Although

this couple performed amniocentesis they opted not to terminate

the pregnancy of an affected fetus. This observation is in line with

previous findings among the Arab population in Israel where some

of the women with affected fetuses will not interrupt the pregnancy

when the diagnosis is performed late in pregnancy. The reason for

this being religious believes and/or when they have not had a

previously affected child [Zlotogora and Reshef, 1998]. Today,

pre-implantation genetic diagnosis may provide a more suitable

option to couples who wish to avoid termination of pregnancy, and

this option is offered to carrier couples.

The characterization of the disease-causing mutation enabled

us to design a pilot carrier screening program which confirmed

the suspected frequency of the mutation in the two villages. Our

MACAROV ET AL. 575

experience with carrier screening for Krabbe disease was the

basis for the implementation of a national program offered to

the Israeli population for the prevention of severe genetic disease

with a frequency higher than 1:1,000 live births [Zlotogora et al.,

2009].

It is interesting, that in the two villages examined, a clear trend

toward premarital testing is observed, whereas in other communi-

ties in which the national screening program is offered—either

Jewish, Druze, or Moslem—almost all individuals screened are

married. This is probably due to the efficient community outreach

in the two villages that has led to increased awareness of the

existence of Krabbe disease and the possibility of being examined.

In addition, the religious authority in the villages strongly recom-

mends that young people be tested before marriage. Premarital

screening offers a convenient option for prevention of genetic

diseases since carrier couples can decide not to proceed with

the marriage process and by this avoid the increased risk and the

possible necessity for a decision whether to terminate pregnancies

of affected fetuses or not. This option suits the norms of traditional

and religious societies in which termination of pregnancy is allowed

only under strict circumstances. For example, in the obligatory

premarital tests for beta-thalassemia in the Gaza strip, most of the

couples found to be at risk opted not to marry [Tarazi et al., 2007].

Whereas the advantage of premarital screening is obvious, one

major problem that should not be ignored is that it may lead to

stigmatization at the individual level, especially in the case of

women in these communities. This stigmatization can be reduced

if the male partner is examined first, or may be avoided by perform-

ing premarital carrier matching instead of individual testing. In

premarital carrier matching, the results are given to the couple

in terms of whether there is a risk, rather than to an individual

as to whether he or she is a carrier. In this way only when

both partners are found to be carriers will the information be

given, therefore reducing the number of individuals subjected to

stigmatization. Premarital carrier matching has been implemented

successfully in the ultra-orthodox Jewish community by the Dor

Yeshorim Organization [Ekstein and Katzenstein, 2001].

Prevention of the birth of affected infants has been achieved in

the villages either by premarital carrier testing or by prenatal

diagnosis. As mentioned, almost all carrier couples that opted for

prenatal diagnosis had a previously affected child or children, and

in most cases the pregnancies of affected fetuses were terminated.

Since the implementation of the prevention program in the villages

the incidence of Krabbe disease at birth declined from 1.6 per 1,000

live births to 0.82 per 1,000.

Looking back at our experience of almost four decades it would

appear that our intervention is only partially successful. One

important achievement of the program is that it is well accepted

by the community. We are not aware of stigmatization at the

individual or the community level. This program allowed many

of the families who had affected children to have healthy children.

However, much still remains to be done, since our purpose is to

offer a true choice to every member of the community. This has not

yet been achieved as several children were born to couples that did

not know about the possibilities of prevention.

One of the drawbacks is that we failed to know how many

partners of carriers did not come for testing and for what reason:

was it because they decided not to get married or whether the

individual tested had no partner.

REFERENCES

Ekstein J, Katzenstein H. 2001. The Dor Yeshorim story: Community-based carrier screening for Tay-Sachs disease. Adv Genet 44:297–310.

Levy-Lahad E. Krabbe disease in an inbred community: Screening for thedetection of affected children and families at risk. MD thesis, Hadassah-Hebrew University Medical School. 1987.

Rafi MA, Luzi P, Zlotogora J, Wenger DA. 1996. Two different mutationsare responsible for Krabbe disease in the Druze and Moslem Arabpopulations in Israel. Hum Genet 97:304–308.

Tarazi I, Al Najjar E, Lulu N, Sirdah M. 2007. Obligatory premarital tests forbeta-thalassaemia in the Gaza Strip: Evaluation and recommendations.Int J Lab Hematol 29:111–118.

Wenger DA, Williams C. 1991. Screening for lysosomal disorders. In:Hommes FA, editor. Techniques in diagnostic human biochemicalgenetics. New York: Wiley-Liss. pp 587-617.

Wenger DA, Suzuki K, Suzuki Y, Suzuki K. 2005. Galactosylceramide lipidosis:Globoidcell leukodystrophy(Krabbedisease). In:ScriverCR,BeaudetAL, SlyWS, Valle D, Vogelstein B, editors. The metabolic and molecular bases ofinherited disease (OMMBID). New York, NY: McGraw-Hill.

Zlotogora J, Reshef N. 1998. Prenatal testing for genetic disorders amongArabs. Prenat Diagn 18:219–224.

Zlotogora J, Regev R, Zeigler M, Iancu TC, Bach G. 1985. Krabbe disease:Increased incidence in a highly inbred community. Am J Med Genet21:765–770.

Zlotogora J, Levy-Lahad E, Legum C, Iancu TC, Zeigler M, Bach G. 1991.Krabbe disease in Israel. Isr J Med Sci 27:196–198.

Zlotogora J, Carmi R, Lev B, Shalev SA. 2009. A targeted population carrierscreening program for severe and frequent genetic diseases in Israel. Eur JHum Genet 17:591–597.

576 AMERICAN JOURNAL OF MEDICAL GENETICS PART A