The frequency of Familial Mediterranean fever gene mutationsand genotypes at Kirikkale and comparison with the meanof regional MEFV mutation frequency of Turkey

Derya Beyza Sayın Kocakap • Aysen Gunel-Ozcan •

Feryal Cabuk • Cuneyt Ensari

Received: 12 December 2012 / Accepted: 24 December 2013 / Published online: 1 January 2014

� Springer Science+Business Media Dordrecht 2013

Abstract In this study we have retrospectively analysed

the mutation spectrum of the 351 Familial Mediterranean

fever patients referred to Kırıkkale University Faculty of

Medicine, Department of Medical Genetics Laboratory

over a period of 5 years and compared them with Turkey’s

mean. We have found 11 different mutations, including

rare mutations such as F479L, K695R, M680I(G/A) and 45

different genotypes showing the heterogeneity of MEFV

mutations in Central Anatolia. The most three prevalent

mutations were M694V (14.8 %), E148Q (7.1 %) and

M680I(G/C) (4.1 %) in accordance with the literature. We

have also investigated R202Q in our routine molecular

diagnosis. Mutation causing R202Q (c.605G [ A) change

was described as a frequent polymorphism and G allele

was found in linkage disequilibrium (LD) with M694V.

There are limited number of studies investigating R202Q,

some of them implicate that its homozygote state is disease

causing. We showed the high frequency of R202Q

(23.7 %) with and without M694V in all the groups ana-

lysed and its high LD rate with M694V in the diagnosed

group. Our study is reflecting the mutational heterogeneity

of MEFV and summarize mutational spectrum of Turkey’s

geographical regions and overall Turkey.

Keywords MEFV � Mutation � Turkey �Allelic frequencies

Introduction

Familial Mediterranean fever (FMF) (OMIM #249100) is a

hereditary autosomal recessive disorder, which is caused

by mutations in MEFV (FMF) gene resulting with sterile

inflammation in the peritoneum, synovium, or pleura. The

main clinical features of the disease is short, self resolving

recurrent attacks of fever and, acute abdominal, joint or

chest pain; myalgia and erythema. The most severe com-

plication of FMF is renal failure secondary to renal amy-

loidosis. In some cases renal amyloidosis may develop as

the first clinical manifestation [1].

FMF is mainly observed in eastern Mediterranean peo-

ple including Turks, Armenians, non-Ashkenazi Jews and

Arabs however it can be seen throughout the world due to

population movements. In Turkey, the estimated preva-

lence of FMF is 1/1,000, and the carrier rate is as high as

1/5 and most of the FMF patients originate mainly from the

non-Mediterranean regions of Turkey [2].

The gene responsible for FMF, MEFV is located on the

short arm of chromosome 16, has 10 exons and encodes a

781 amino acid pyrin/marenostrin protein. Although the

exact role of pyrin is not yet determined, it is thought that it

D. B. Sayın Kocakap (&) � A. Gunel-Ozcan � F. Cabuk

Department of Medical Genetics, School of Medicine, Kırıkkale

University, 71450 Kirikkale, Turkey

e-mail: dsayin@yahoo.com

Present Address:

A. Gunel-Ozcan

Center for Stem Cell Research and Development, Stem Cell

Sciences Department, Institute of Health Sciences, Hacettepe

University, Ankara, Turkey

Present Address:

F. Cabuk

Eli Lilly and Company, Istanbul, Turkey

C. Ensari

Division of Pediatric Nephrology, Department of Pediatrics,

School of Medicine, Kırıkkale University, Kirikkale, Turkey

123

Mol Biol Rep (2014) 41:1419–1426

DOI 10.1007/s11033-013-2986-4

plays a role in caspase-1 and interleukin-1b pathways,

leading to apoptosis-associated protein expression and anti-

inflammatory activity [3, 4]. To date, around 270 sequence

variants have been identified most of which are clustered in

exon 10 [5]. Albeit this mutation multiplicity, four com-

mon mutations (M680I(G/C), M694V, M694I, V726A)

account for 85 % FMF chromosomes in geographical areas

where FMF is frequent [6]. Mutations of the MEFV gene

diminish pyrin expression, which abolish its inhibitory

effect on caspase-1-mediated activation of interleukin-1band leading to increased inflammation [7].

The aim of our study is to evaluate MEFV mutation

frequency of Kırıkkale region over a period of 5 years and

to compare it, with MEFV mutation frequency according to

the geographical regions and overall Turkey.

Materials and methods

Patients

In this retrospective study a total of 351 suspected FMF

patients of which 30 definitely diagnosed, who referred to

Kirikkale University Faculty of Medicine, Department of

Medical Genetics Laboratory, over a period of 5 years,

between December 2007 and December 2012 were inves-

tigated. All patients have one or more FMF related

symptoms; mainly abdominal pain, fever and arthralgia.

Only 30 patients could be definitely diagnosed according to

Tel-Hashomer Critera and classified as ‘‘diagnosed’’ group,

for the rest of our patients it could not be possible to obtain

detailed clinical course and they remained as prediagnosed

or suspected FMF patients consisting the ‘‘suspected’’

group. Our genotyping results between August 2005 and

November 2007 were published previously [8]. All the

patients were from Central Anatolia, mainly from Kirikkale

and were referred from various inpatient and outpatient

clinics of Kirikkale University Suleyman Demirel Educa-

tion and Research Hospital, primarily Pediatrics (152),

Internal Medicine (113) and Physical Medicine and

Rehabilitation (53) Departments. All patients or their par-

ents signed informed consent prior to genetic testing and

the study was approved by the local Ethics Committee.

Detection of MEFV mutation

Genomic DNA was extracted from peripheral blood sam-

ples with EDTA using EZ-10 Spin Column Genomic DNA

KIT for blood samples (BIOBASIC, CANADA) according

to manufacturer’s instructions. MEFV mutation analysis

was performed with two different reverse hybridisation

based commercial kits, which can detect 12 and 15 com-

mon mutations; FMF Strip Assay, ViennaLab

Labordiagnostika GmbH and AID Diagnostica GMBH

respectively. According to manufacturer’s instructions

target DNA regions were amplified with multiplex poly-

merase chain reaction (PCR) using biotinylated primers.

PCR products were selectively hybridized to a test strip

containing allele-specific oligonucleotide probes, immobi-

lized as an array of parallel lines for each mutation. Bound

biotinylated sequences are detected using streptavidin–

alkaline phosphatase and color substrates. Both tests

include E148Q (exon2), P369S (exon3), F479L (exon5);

M680I(G/C), M680I(G/A), M694V, M694I, K695R,

V726A, A744S and R761H (exon10) mutations, while

I692del (exon10) is included in the first mentioned; S108R,

R202Q (exon2); V487M (exon5) and Y688X (exon10) are

included in the second mentioned kit. AID Diagnostica’s

kit has been used since November 2010.

Statistical analysis

Patients are studied in three groups as suspected, diagnosed

and overall. Mutation frequencies and genotypes, genders

are analysed with Microsoft Office Excel 2007 programme.

Results

The total number of analysed FMF patients is 351, of these

321 are suspected patients, and 30 are definitely diagnosed

as FMF by Pediatry Clinic [9]. Female/male ratio of the

patients is 1.44 (207/144); 1.41 (188/133) and 1.72 (19/11)

as overall, suspected and diagnosed patients respectively.

The mean age is 21 ± 14.3 (range 2–75); 22 ± 14.8 (range

2–75) and 8.6 ± 3.8 (range 2–16) as overall, suspected and

diagnosed patients respectively. We found at least one

mutation in 202 of 351 (57.55 %) patients; 13 (6 diag-

nosed) had 4 mutations, homozygous R202Q and M694V;

18 (3 diagnosed) had 3 mutations; 17 (2 diagnosed) had 2

homozygous mutations, 53 (7 diagnosed) had 2 compound

heterozygous mutations; 101 (9 diagnosed) had 1 mutation

and 149 (3 diagnosed) had none of the mutations covered

in the tests. A total of 45 different genotypes were found

among our patients. The distribution of patients’ genotypes

is given in Table 1. The most frequent genotypes are

R202Q/Wt and E148Q/Wt found in 24 (6.8 %) and 20

(5.9 %) patients respectively. Interestingly, among the

genotypes carrying two or more mutations most common

ones are R202Q/M694V (17 patient, 4.8 %) and the

genotype of all 13 patients with four mutations is R202Q/

R202Q/M694 V/M694 V (3.7 %), reflecting high linkage

disequilibrium (LD) rate of the two mutations as described

below.

1420 Mol Biol Rep (2014) 41:1419–1426

123

Table 1 The distribution of

patients’ genotypesSuspected Diagnosed Overall

Four mutations

R202Q/R202Q/M694V/M694V 7 6 13

Total 7 (2.2 %) 6 (20 %) 13 (3.7 %)

Three mutations

E148Q/E148Q/M694V 1 0 1

E148Q/R202Q/M694V 2 1 3

E148Q/P369S/M694V 0 1 1

E148Q/K695R/V726A 1 0 1

R202Q/R202Q/M694V 4 1 5

R202Q/M680I(G/C)/M694V 2 0 2

R202Q/M680I(G/A)/M694V 1 0 1

R202Q/M694V/M694V 1 0 1

R202Q/M694V/V726A 2 0 2

R202Q/M694V/R761H 1 0 1

Total 15 (4.7 %) 3 (10 %) 18 (5.1 %)

Two mutations

Homozygote

E148Q/E148Q 2 1 3

R202Q/R202Q 3 0 3

M680I(G/C)/M680I(G/C) 1 1 2

M694V/M694V 6 0 6

V726A/V726A 3 0 3

Total 15 (4.7 %) 2 (6.7 %) 17 (4.8 %)

Compound heterozygote

E148Q/R202Q 2 0 2

E148Q/P369S 3 0 3

E148Q/M680I(G/C) 4 1 5

E148Q/M680I(G/A) 1 0 1

E148Q/M694V 5 0 5

E148Q/V726A 1 0 1

R202Q/P369S 1 0 1

R202Q/M680I(G/C) 1 0 1

R202Q/M694V 13 4 17

R202Q/K695R 2 1 3

R202Q/A744S 1 0 1

F479L/M694V 1 0 1

F479L/V726A 1 0 1

M680I(G/C)/M680I(G/A) 1 0 1

M680I(G/C)/M694V 5 0 5

M680I(G/C)/V726A 1 0 1

M694V/V726A 1 1 2

M694V/R761H 1 0 1

M694V/A744S 1 0 1

Total 46 (14.3 %) 7 (23.3 %) 53 (15.1 %)

One mutation

E148Q 19 1 20

R202Q 22 2 24

P369S 3 0 3

F479L 3 0 3

Mol Biol Rep (2014) 41:1419–1426 1421

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Mutation causing R202Q (c.605G [ A) change was

described as a frequent polymorphism and G allele was

found in linkage disequilibrium with M694V [5]. We

analysed R202Q mutation in 213 persons (30 diagnosed),

and found that in 58 alleles (57.4 % of R202Q alleles)

R202Q and M694V were in LD, while in 43 alleles R202Q

and in 46 alleles M694V were found solely. The frequency

of the R202Q allele was found to be 23.7 % (101 alleles).

When we have excluded R202Q mutation which was

described as a frequent polymorphism, we found that 176

of 351 (50.1 %) patients had at least one mutation; 151/321

(47 %) in suspected and 25/30 (83.3 %) patients in diag-

nosed group. Table 2 shows the allelic frequency of the

tested mutations except R202Q. While M694V (14.8 %),

E148Q (7.1 %), M680I-(G/C) (4.1 %) and V726A (3.9 %)

mutations were the most frequent; we have never detected

S108R,V487M, Y688X, M694I and I692del mutations.

In a total of 351 people investigated, 47 had family

history (13.4 %), 29 (female/male:20/9) of 321 (9.03 %)

suspected, 18 (female/male: 12/6) of 30 (60 %) diagnosed.

Discussion

In this study MEFV mutation analysis results of 351

patients who were under the research of priliminary diag-

nosis of FMF is given. We have evaluated 12 mutations of

the MEFV gene among 138 and 15 mutations among 213

patients at the Molecular Diagnostic Laboratory of Kir-

ikkale University, Department of Medical Genetics. Kir-

ikkale, a small city in Central Anatolia where FMF is

prevalent, is located 80 km east of Ankara and does not

take too much immigration. Our patients are studied in

three different groups as suspected, diagnosed and overall

as described in the materials and methods part, patients

section. When we have analysed the literature reporting

Turkish MEFV mutation frequencies, we have noticed that

some of the publications were made from molecular

diagnostic laboratories and some of them were made from

different clinics. While the formers report mainly MEFV

mutation frequencies of FMF suspected patients, without

definite diagnosis of FMF and clinical course; the laters

report mainly MEFV mutation frequencies, usually with the

clinical course of the diagnosed (according to diagnostic

criteria for FMF) FMF patients. We have assembled vari-

ous studies from Turkey and grouped them according to

their patient profiles in order to compare with our mutation

profile (suspected, with symptoms of FMF, without definite

diagnosis; and definitely diagnosed, with definite diagnosis

according to Tel-Hashomer or Livneh Diagnostic Criteria).

Table 3 shows the most common mutation frequencies of

the studies in suspected, definitely diagnosed and overall

groups in Turkey, according to the region where the study

was made. Although some reports are made from referral

centers of all Turkey, we can generalize them as being

local referral centers (because people prefer to go to the

Table 1 continuedSuspected Diagnosed Overall

M680I(G/C) 10 0 10

M694V 15 1 16

K695R 4 1 5

V726A 11 2 13

A744S 3 0 3

R761H 2 2 4

Total 92 (28.7 %) 9 (30 %) 101(28.8 %)

Zero mutation 146 (45.5 %) 3 (10 %) 149 (42.5 %)

Total 321 30 351

Table 2 The allelic frequency of the tested mutations except R202Q

Suspected alleles

n (%)

Diagnosed alleles

n (%)

Overall alleles

n (%)

M694V 83 (12.9) 21(35) 104 (14.8)

E148Q 44 (6.9) 6 (10) 50 (7.1)

M680I(G/C) 24 (4.1) 3 (5) 27 (4.1)

V726A 22 (3.7) 3 (5) 25 (3.9)

P369S 8 (1.3) 1 (1.7) 9 (1.3)

K695R 7 (1.1) 2 (3.3) 9 (1.3)

R761H 4 (0.6) 2 (3.3) 6 (0.9)

F479L 5 (0.8) 0 (0) 5 (0.7)

A744S 5 (0.8) 0 (0) 5 (0.7)

M680I(G/

A)

3 (0.5) 0 (0) 3 (0.4)

Total allele 642 60 702

We didn’t include R202Q mutation frequency in the table in order to

prevent frequency error which can be caused by the LD of R202Q/

M694V mutations

1422 Mol Biol Rep (2014) 41:1419–1426

123

nearest center) and as reflecting the mutation rate of their

own geographical region in spite of the whole country.

The aim of our study is to compare MEFV mutation

frequency of Kirikkale region to the MEFV mutation

frequency of geographical regions and overall Turkey in

suspected, diagnosed and overall groups. We have evalu-

ated 12/15 mutations of the MEFV gene among 351 FMF

suspected patients, only 30 of them have definite diagnosis

Table 3 MEFV Allele counts and mutation frequencies of different studies from Turkey

References M694V

(%)

E148Q

(%)

M680I(G/C)

(%)

V726A

(%)

P369S

(%)

K695R

(%)

Patient

(n)

Region

Gunel-Ozcan et al.* [8] 17 (6.8) 21 (8.3) 11 (4.4) 11 (4.4) 5 (2.0) 0 (0) 126 Central Anatolia

Erden et al.* [14] 62 (31.6) 22 (11.2) 8 (4.1) 18 (9.2) 2 (1.0) 0 (0) 98 Central Anatolia

Ceylan et al.* [21] 201 (18.9) 53 (5.0) 61 (5.7) 56 (5.3) 9 (0.9) 10 (0.9) 532 Central Anatolia

Dundar et al.* [22] 607 (14.7) 228 (5.5) 315 (7.6) 197 (4.8) 41 (1.0) 8 (0.2) 2067 Central Anatolia

Ozdemir et al.* [23] 1017 (15.2) 476 (7.1) 354 (5.3) 267 (4) 78 (1.2) 8 (0.1) 3340 Central Anatolia

Ceylan et al.* [17] 228 (14.2) 71 (4.4) 62 (3.9) 87 (5.4) 17 (1.1) 9 (0.6) 802 Central Anatolia

Dogan et al.* [24] 312 (13.5) 143 (6.2) 124 (5.4) 69 (3.0) 28 (1.2) 11 (0.5) 1152 Central Anatolia

Akar et al.** [28] 200 (43.5) 3 (0.7) 55 (12.0) 51 (12.0) N/A 4 (0.9) 230 Central Anatolia

Yalcınkaya et al.** [16] 173 (51.8) N/A 53 (15.9) 48 (14.4) N/A N/A 167 Central Anatolia

Yılmaz et al.** [7] 464 (51.6) 29 (3.2) 84 (9.3) 28 (3.1) N/A N/A 450 Central Anatolia

Olgun et al.** [29] 31 (51.7) 0 (0) 15 (25) 5 (8.3) N/A N/A 30 Central Anatolia

Demirkaya et al.** [30] 330 (50) 9 (1.4) 93 (14.9) 64 (9.7) N/A N/A 330 Central Anatolia

Dusunsel et al.** [31] 126 (61.8) 7 (3.4) 25 (12.3) 10 (4.9) 1 (0.5) 1 (0.5) 102 Central Anatolia

Ureten et al.** [18] 205 (39.4) 32 (6.2) 49 (9.4) 42 (8.1) 2 (0.4) 0 (0) 260 Central Anatolia

Total* 2444 (15.0) 1014 (6.2) 935 (5.8) 705 (4.3) 180 (1.1) 46 (0.3) 8117 Central Anatolia

Total** 1529 (51.5) 80 (2.9) 374 (11.9) 248 (7.9) 3 (0.4) 5 (0.4) 1569 Central Anatolia

Total for overall 3973 (20.5) 1094 (5.7) 1309 (6.8) 953 (4.9) 183 (1.1) 51 (0.3) 9686 Central Anatolia

Akın et al.* [25] 375 (15.6) 132 (5.5) 94 (3.9) 102 (4.3) 20 (0.8) 9 (0.4) 1201 Aegean

Ozalkaya et al.* [11] 150 (24.4) 41 (6.7) 50 (8.1) 31 (5.0) 9 (1.5) 9 (1.5) 308 Aegean

Coker et al.* [15] 599 (33.9) 201 (11.4) 165 (9.3) 120 (6.8) 32 (1.8) 18 (1.0) 883 Aegean

Solak et al.** [32] 146 (36.1) 56 (13.9) 48 (11.9) 37 (9.2) N/A N/A 202 Aegean

Ozturk et al.** [33] 301(34.3) 98 (11.2) 63 (7.2) 65 (7.4) 31 (3.5) 9 (1.0) 438 Aegean

Total* 1124 (23.4) 374 (7.8) 309 (6.4) 253 (5.3) 61 (1.3) 36 (0.8) 2392 Aegean

Total** 447 (34.7) 154 (11.0) 111 (8.6) 102 (7.9) 31 (3.5) 9 (1.0) 640 Aegean

Total for overall 1571 (25.8) 528 (8.7) 420 (6.9) 355 (5.8) 92 (1.8) 45 (0.9) 3032 Aegean

Evliyaoglu et al.* [26] 21 (3.2) 64 (9.6) 9 (1.4) 13 (1.9) 16 (2.4) 2 (0.3) 332 Southeastern

Anatolia

Ertekin et al.** [34] 42 (51.2) 3 (3.7) 10 (12.2) 5 (6.1) 0 (0) 1 (1.2) 41 Eastern Anatolia

Etem et al.** [35] 180 (21.7) 159 (19.2) 79 (9.5) 81 (9.8) 31 (3.7) 8 (1.0) 415 Eastern Anatolia

Total** 222 (24.3) 162 (17.8) 89 (9.8) 86 (9.4) 31 (3.4) 9 (1.0) 456 Eastern Anatolia

Total for overall 243 (15.4) 226 (14.3) 98 (6.2) 99 (6.3) 47 (3.0) 11 (0.7) 788 Southeastern/

Eastern

Anatolia

Sahin et al.* [27] 278 (15.0) 91 (4.9) 95 (5.1) 50 (2.7) 20 (1.1) 7 (0.4) 929 Black Sea

Yılmaz et al.** [4] 84 (53.9) 15 (9.6) 25 (16.0) 4 (2.6) 6 (3.9) 0 (0) 78 Black Sea

Total for overall 362 (18.0) 106 (5.3) 120 (6.0) 54 (2.7) 26 (1.3) 7 (0.3) 1007 Black Sea

Inal et al.** [36] 114 (46.0) 9 (3.6) 17 (6.9) 16 (6.5) 2 (0.8) 0 (0) 124 Mediterranean

Turkish FMF Study Group** [2] 1121 (51.4) N/A 313 (14.4) 188 (8.6) N/A N/A 1090 Turkey

Total* 3867 (16.4) 1543 (6.6) 1348 (5.7) 1021 (4.3) 277 (1.2) 91 (0.4) 11770 Turkey

Total** 3517 (44.4) 420 (7.8) 929 (11.8) 644 (8.1) 73 (2.9) 23 (0.8) 3957 Turkey

Total for overall 7384 (23.5) 1963 (6.8) 2277 (7.2) 1665 (5.3) 350 (1.3) 114 (0.4) 16693 Turkey

* Suspected

** Diagnosed

Mol Biol Rep (2014) 41:1419–1426 1423

123

of FMF and all of them are pediatric. The major limitation

of our study is the small number of the diagnosed group,

even so our genotyping results will contribute to the

Turkish FMF patients’ mutation profile. We have found 11

different mutations and 45 different genotypes. In all three

groups (suspected, diagnosed, overall) analysed, M694V

(14,8 % overall) is the most prominent mutation, E148Q

(7.1 % overall), M680I(G/C) (4.1 % overall) and V726A

(3.9 % overall) are second, third and fourth prominent

mutations respectively. When we compare our frequencies

with those given in Table 3 we can see that M694V is the

most prominent mutation in overall Turkey group, and in

all the regions separately consistent with results presented

here. But, our M694V frequency is lower, when compared

with most of the studies given in Table 3, and is closest to

the frequencies of Eastern and Southeastern Anatolia. In

our previous study E148Q was the most prominent muta-

tion, while M694V was the second. We also detected rare

mutations such as K695R, R761H, F479L and M680I(G/A)

and patients with three mutations, which were not detected

in our previous study, showing the mutational heteroge-

neity in Turkish FMF patients [10]. The increase of the

M694V frequency and detection of rare mutations may be

attributed to the increase of the patients studied, reflecting a

bigger part of the population. Among the rare mutations,

the frequency of M680I(G/A) is the highest in our study in

the suspected group. Interestingly none of the studies

investigated so far has observed this mutation among def-

initely diagnosed patients (data not shown). K695R (1.3 %)

is found to be relatively high when compared to Turkey’s

overall and regional frequencies especially in the diag-

nosed group (3.3 %). Only Ozalkaya et al. [11] have found

higher frequency in the suspected group than presented

here.

Overall Turkey, in Central Anatolia and in Black Sea

Regions the second most common mutation is M680I(G/C)

while in Aegean and Eastern-Southeastern Anatolia it is

E148Q. In this study also, E148Q is the second most

common mutation, for the suspected and overall groups, its

frequency is near to the mean of Turkey. There are con-

flicting views about the role of E148Q mutation on FMF

pathogenesis. Although it was described as a disease

causing mutation with low penetrance and mild symptoms,

it has also been suggested to be a normal sequence variant

[12, 13]. Our E148Q mutation frequency is in the second

place in all three groups, supporting the idea that it is a

disease causing mutation.

M680I(G/C) mutation is one of the most prevalent

mutations in Turkey [2], but although it is the third most

common mutation in our study, its frequency is quite low:

for the definitely diagnosed group its frequency is the

lowest of all studies investigated and for the suspected

group its frequency is similar with our previous study and

Erden et al.’s study [8, 14].

As we have summarized Turkish MEFV mutation fre-

quencies in Table 3, we can clearly see that in both diag-

nosed and suspected groups mutation frequencies show

regional differences which can be attributed to the cultural

mosaicism of Turkey. We can also notice that mutation

frequencies of diagnosed group is approximately twice of

the suspected group for M680I(G/C), V726A, P369S and

K695R mutations and three times higher for M694V, we

can speculate that it can be attributable to the awareness of

the high prevalence of FMF in Turkish population and

increased usage of molecular tests in the diagnosis of

patients with FMF-like symptoms. This result also implies

that it is important to pay attention on patients’ profile to

give accurate results and make effective comparison when

reporting allelic frequencies.

In 53 patients (7 diagnosed), we have identified 2 het-

erozygous mutations, more likely to be a—a compound

heterozygous state, rather than the two mutations in the

same chromosome, which is a rarer situation. If mutations

are located in the same chromosome the patient would have

a wild type allele and be a FMF carrier, otherwise he/she

will be compound heterozygote and FMF patient. Most of

the diagnostic tests used, do not discriminate if two

mutations are in cis or trans position, and so if the patient is

carrier or ‘‘FMF patient’’. In order distinguish the two

situations, genotyping of the parents or siblings will be

usefull.

When excluded R202Q mutation, three patients had

three mutations which are E148Q/E148Q/M694V, E148Q/

P369S/M694V and E148Q/K695R/V726A, all of them

bear E148Q mutation. There are a lot of studies reporting

more than two mutations in MEFV gene, which are

M694V/P369S/E148Q [15, 25], M680I(G/C)/R761H/

E148Q [25, 33], E148Q/M694V/K695R, E148Q/E148Q/

P369S [25]; E148Q/P369S/M680I(G/C) [15, 33]; M694V/

M680I/V726A [4]; M694V/E148Q/V726A, M680I/M680I/

M694I [32]; E148Q/E148Q/M694V [33]. As seen above

most genotypes carry E148Q mutation in accordance with

our findings. To best of our knowledge this is the first

report of E148Q/K695R/V726A genotype from Turkey.

Although most studies indicate that FMF has no sex

preponderance [15, 16] we have found a female predomi-

nance in all the groups analysed. There are some reports

which are in accordance with female dominance in FMF

patients [8, 17, 18, 31]. In our previous study 63 % of

patients were female, and this ratio has given us the

opportunity to suggest that there may be an epigenetic

effect. In this present study 59 % of the overall group and

63 % of the diagnosed group are female in accordance with

our previous study.

1424 Mol Biol Rep (2014) 41:1419–1426

123

Even though FMF is a clinically diagnosed disease,

molecular diagnostic tests are helpful to detect disease-

causing mutations, confirmation of the diagnosis and

diagnosis of the presymptomatic patients’ relatives.

Inheritance pattern of FMF is autosomal recessive imply-

ing that only patients carrying two mutant alleles are

symptomatic. There are many studies indicating that there

are clinically diagnosed FMF patients who carry hetero-

zygous MEFV mutations or zero mutation [19, 20]. In our

diagnosed group only 40 % of patients carry two mutant

alleles, in agreement with the hypothesis that one mutant

allele may be associated with mild FMF symptoms [17].

We must also consider that there might be rare mutations,

which are not included in the strip assay or other unknown/

modifier loci implied in FMF.

In this study we retrospectively analysed the mutation

spectrum of the suspected and diagnosed FMF patients

referred to Kirikkale University Faculty of Medicine,

Department of Medical Genetics’ Laboratory over a period

of 5 years and compared them with Turkey’s mean. We

have found 11 different mutations, including rare mutations

such as F479L, K695R, M680I(G/A) and 45 different

genotypes showing the heterogeneity of MEFV mutations

in Central Anatolia. The most prevalent mutation was

M694V and 55.8 % of M694V alleles were in linkage

disequilibrium with R202Q. There are limited number of

studies investigating R202Q which is known as a sequence

variant, some of them implicate that its homozygote state is

disease causing. We showed the high frequency of R202Q

with and without M694V in all the groups analysed and its

high linkage disequilibrium rate with M694V in the diag-

nosed group. In conclusion our study is reflecting the

mutational heterogeneity of MEFV and summarize muta-

tional spectrum of Turkey’s geographical regions and

overall Turkey.

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