RSS was first described in 1953 [1]. It is a clinically and genetically heterogeneous condition. Clinical diagnosis is based on various characteristics including slow growth before and after birth, relative macrocephaly, a prominent forehead at a young age, hemihypotrophy, and fifth finger clinodactyly. Five different diagnostic scoring systems of RSS have been published. These systems take into account most major clinical features and sometimes minor clinical features, and are on the whole quite similar. These five scoring systems were recently compared in the Journal of Medical Genetics in terms of sensitivity, specificity, and positive or negative predictive value [2]. In this evaluation, the scoring system described by Netchine et al. [3] showed a high degree of specificity and sensitivity. This scoring system has also been used by other teams from various countries in the diagnosis of RSS (see for instance [4]). Furthermore, the article describing this scoring system [3] was reviewed by the editorial board of The Journal of Clinical Endocrinology & Metabolism (JCEM) and received an award in 2008 from the Endocrine Society. This article has been cited in PUBMED 128 times. The first described molecular abnormality in RSS was the maternal uniparental disomy of chromosome 7 (mUPD7). This defect is present in 5- 10% of individuals affected by RSS [5, 6]. This observation suggests the aberrant dosage of imprinted genes may be involved in the pathology of RSS, although the gene(s) involved remain unknown. Recently, abnormalities of 11p15, a locus also harboring imprinted genes, have been described. This region contains two separate imprinting clusters: the telomeric domain (ICR1) that regulates H19 and IGF2, and the centromeric domain (ICR2) that regulates the long non-coding RNA KCNQ1OT1 and CDKN1C genes. Of note, cytogenetic, genetic, and epigenetic disturbances of the 11p15 region encompassing ICR1 and ICR2 are associated with the overgrowth disorder Beckwith Wiedemann syndrome (BWS). BWS is a clinical and molecular mirror of RSS, and around 8% of cases involve loss of function mutations of CDKN1C [7]. In RSS, loss of methylation (LOM) of the paternal ICR1 is the most prevalent defect. This epigenetic change is found in up to 60% of RSS patients [3, 8], although it has been reported to occur less frequently in some populations (e.g. with a frequency of ~31% in Japanese patients) [4]. Approximately 30% of RSS patients are ? idiopathic ? with no molecular anomaly yet identified.

General comments on the items of the scoring system described in [3]: Young children with RSS have a very different clinical profile than older children or adults with RSS. Feeding difficulties are much more pronounced in early life and therefore have to be assessed at a young age. Older children and adults may become overweight. Feeding difficulties can be difficult to assess; therefore, one of the conclusions of our paper in 2007 was to replace the item ? feeding difficulties ? with ? feeding difficulties and /or a BMI two SDs below the mean at around two years of age ?. The prominent forehead is distinct from relative macrocephaly. Indeed, relative macrocephaly is characterized by a growth restriction affecting mostly the body, whereas circumference of the head is relatively less unaffected. A prominent forehead however, is a dysmorphic feature that describes the characteristic shape of these patients' forehead protruding over the plane of the face. This feature tends to disappear in late childhood and therefore has to be assessed around two years of age. Body asymmetry is a feature of RSS, but is not present in all RSS patients. This feature is more frequent in RSS patients with epigenetic changes to 11p15 than in patients with mUPD7. For example, in our sample of 145 RSS patients with 11p15 LOM, 79% of patients had hemihypotrophy, whereas in a sample of 16 RSS patients with mUPD7, none of the patients were affected by hemihypotrophy (p<0.0001). This is probably due to the fact that the 11p15 LOM occurs in a mosaic manner, such that the proportion of cells that possess this epigenetic change may differ between the left and right side of the body. The item post natal growth retardation also has to be assessed around two years of age. This is because most RSS patients are treated with growth hormone generally starting between two and four years of age, to improve their growth rate. Bone age and pubertal status also has to be taken into account when analyzing the growth of a child affected by RSS. Indeed, these children frequently have an advanced bone age due to aggressive adrenarche and early puberty. Thus, even though their height may be normal during childhood, their final height may be lower than average due to premature bone fusion.

Specific comments in response to this letter The diagnosis of RSS in the patients reported by Brioude et al. is not questionable because all the patients carrying the mutation fulfilled the clinical criteria of RSS as described by Netchine et al. in 2007. Furthermore, these patients also fulfill the RSS clinical criteria according to other validated RSS clinical scoring systems such as the Birmingham scoring system [2]. However, as mentioned above, the clinical diagnosis of RSS is often difficult. RSS scoring systems are helpful to orient the diagnosis; nonetheless, it is also very important that an experienced clinical geneticist or pediatrician make this diagnosis. The pediatrician (IO) who referred this family to our laboratory has a lot of clinical experience in RSS and has followed two generations of this family (second and third). The pediatrician (IO) became convinced of the diagnosis of a very rare case of familial RSS when the third generation of this family came to the clinic for consultation. In the clinical scoring system described by Netchine et al. [3], body asymmetry is one of the six criteria for the clinical diagnosis of RSS. However, in a family carrying a CDKN1C mutation, body asymmetry is not expected to occur because the molecular defect is present in all the cells of the body (unlike the mosaic epigenetic change at the 11p15 locus). This hypothesis is in accordance with the observation that patients with Beckwith Wiedemann syndrome who carry CDKN1C loss of function mutations are not usually affected by body asymmetry [7]. Similarly, patients with IMAGe syndrome with a mutation in CDKN1C do not have body asymmetry [9]. Thus, the absence of body asymmetry does not exclude the diagnosis of RSS in this particular case. Furthermore, we found that RSS patients with mUPD7 do not show body asymmetry, and some RSS patients with 11p15 LOM are also unaffected (20% in our sample of patients). As mentioned above, all the patients described here had feeding difficulties and/or a BMI two SDs below the mean at around two years of age. We can consider a low BMI to be equivalent to feeding difficulties, as mentioned in the legend of table 2 and proposed in our paper in 2007 [3]. Relative macrocephaly was not present at birth for all the patients presented here (notably in the propositus IV.1). However, this trait was present at two years of age in all patients carrying the mutation. Therefore, this characteristic may help to distinguish RSS patients with a mutation in CDKN1C from patients with 11p15 LOM. However, the absence of relative macrocephaly is not enough to preclude the clinical diagnosis of RSS in these patients. Although the triangular face does not appear to be obvious in the pictures of the patients, this feature has been confirmed by the patients' physician (IO). Furthermore, this trait (triangular face and prominent forehead) is usually less obvious in adult patients. Besides, a triangular face is not a clinical criterion for RSS diagnosis and is not part of our scoring system. All the patients described here presented growth retardation: Length at birth and then height and/or weight was two SDs below the mean according to French reference charts both at birth and after birth. All the patients with the CDKN1C mutation did not attain a normal height at the age of 2 years. Patient III.7 received a long-term therapy consisting of growth hormones and Triptorelin acetate and thus reached a final height of 157 cm. However this patient's sister, (patient III.6) received growth hormone therapy for a shorter duration than her sister resulting in a lower final height. Today, most RSS patients are treated with growth hormones and the best improvement in final height is attained when the rapid advancement in bone age is also treated. Individuals II.4 and III.10 do not carry the CDKN1C mutation, and cannot be considered as having RSS because they do not present the clinical criteria as described above. These two individuals did not show relative macrocephaly at birth or at the age of two years. Their size at birth was either normal (II.4), or, in the case of III.10, small, but nonetheless larger than new-born babies corresponding to family members who carry the mutations. Individuals II.4 and III.10 had a normal BMI, and II.4 had a normal adult height without any therapy. We agree with the point that individual III.10 showed a short final height despite growth hormone therapy. This patient was followed like the other patients with short stature of this family by the same pediatrician (IO), who did not clinically assess her as having RSS. To date, the causal mechanism of her short adult height has not been elucidated.

CONCLUSION Defining the clinical criteria of a clinically and molecularly heterogeneous syndrome like RSS is not straightforward. Nonetheless, we are convinced that the family described here, represents a rare case of familial RSS. The diagnosis of RSS was made by a physician experienced in RSS who had the opportunity to follow and treat two generations of this exceptional family. The scoring system described by Netchine et al. [3], but also other RSS scoring systems such as those evaluated by Dias et al. [2] also favor this diagnosis. The absence of body asymmetry does not rule out the diagnosis of RSS, as in mUPD7 RSS cases or in 20% of 11p15 LOM cases. In this case, the absence of body asymmetry is consistent with the genetic cause identified. This CDKN1C mutation is therefore a new genetic cause of familial RSS. The CDKN1C mutation identified in this RSS familial case affects the same amino acid as a mutation described by Arboleda et al. [9] in patients with IMAGe syndrome. This observation will be of great interest for the readers of The Journal of Medical Genetics. IMAGe syndrome has some similarities with RSS, including fetal and postnatal growth retardation and a prominent forehead, but is also typically characterized by bone anomalies and a severe adrenal insufficiency that were not present in this family. It is striking to note that a mutation on the exact same amino acid, but with a different substitution, can lead to severe adrenal insufficiency in one case and a normal adrenal function in the other case, and therefore to two distinct syndromes. As of 2013, 60 years after the first description of RSS, several molecular causes of RSS have been identified (mUPD7 and various types of 11p15 anomalies) accounting for about 70% of etiologies. We suggest that CDKN1C mutations are a new molecular cause of familial RSS. Given the clinical and molecular heterogeneity of RSS we suggest that RSS should no longer be considered as a syndrome but as a spectrum of disorders with a variety of causes. This would make it easier to understand why RSS is clinically and molecularly heterogeneous. However, it is important for RSS patients to be identified as belonging to a same spectrum and not to multiple categories of very rare conditions, because these patients share a lot of clinical manifestations and have similar treatment needs. This will allow the development of common clinical guidelines for these patients, and well help in the implementation of clinical trials that are always difficult to establish for rare diseases. The identification of exact genetic defect is also a cornerstone for an accurate genetic counseling.

1 Silver HK, Kiyasu W, George J, Deamer WC. Syndrome of congenital hemihypertrophy, shortness of stature, and elevated urinary gonadotropins. Pediatrics 1953;12(4):368-76. 2 Dias RP, Nightingale P, Hardy C, Kirby G, Tee L, Price S, Macdonald F, Barrett TG, Maher ER. Comparison of the clinical scoring systems in Silver -Russell syndrome and development of modified diagnostic criteria to guide molecular genetic testing. J Med Genet 2013;50(9):635-9. 3 Netchine I, Rossignol S, Dufourg MN, Azzi S, Rousseau A, Perin L, Houang M, Steunou V, Esteva B, Thibaud N, Demay MC, Danton F, Petriczko E, Bertrand AM, Heinrichs C, Carel JC, Loeuille GA, Pinto G, Jacquemont ML, Gicquel C, Cabrol S, Le Bouc Y. 11p15 imprinting center region 1 loss of methylation is a common and specific cause of typical Russell-Silver syndrome: clinical scoring system and epigenetic-phenotypic correlations. J Clin Endocrinol Metab 2007;92(8):3148-54. 4 Fuke T, Mizuno S, Nagai T, Hasegawa T, Horikawa R, Miyoshi Y, Muroya K, Kondoh T, Numakura C, Sato S, Nakabayashi K, Tayama C, Hata K, Sano S, Matsubara K, Kagami M, Yamazawa K, Ogata T. Molecular and clinical studies in 138 Japanese patients with Silver-Russell syndrome. PLoS One 2013;8(3):e60105. 5 Kotzot D, Schmitt S, Bernasconi F, Robinson WP, Lurie IW, Ilyina H, Mehes K, Hamel BC, Otten BJ, Hergersberg M, et al. Uniparental disomy 7 in Silver-Russell syndrome and primordial growth retardation. Hum Mol Genet 1995;4(4):583-7. 6 Preece MA, Price SM, Davies V, Clough L, Stanier P, Trembath RC, Moore GE. Maternal uniparental disomy 7 in Silver-Russell syndrome. J Med Genet 1997;34(1):6-9. 7 Brioude F, Lacoste A, Netchine I, Vazquez MP, Auber F, Audry G, Gauthier -Villars M, Brugieres L, Gicquel C, Le Bouc Y, Rossignol S. Beckwith- Wiedemann Syndrome: Growth Pattern and Tumor Risk according to Molecular Mechanism, and Guidelines for Tumor Surveillance. Horm Res Paediatr 2013;80:457-65. 8 Gicquel C, Rossignol S, Cabrol S, Houang M, Steunou V, Barbu V, Danton F, Thibaud N, Le Merrer M, Burglen L, Bertrand AM, Netchine I, Le Bouc Y. Epimutation of the telomeric imprinting center region on chromosome 11p15 in Silver-Russell syndrome. Nat Genet 2005;37(9):1003-7. 9 Arboleda VA, Lee H, Parnaik R, Fleming A, Banerjee A, Ferraz-de-Souza B, Delot EC, Rodriguez-Fernandez IA, Braslavsky D, Bergada I, Dell'angelica EC, Nelson SF, Martinez-Agosto JA, Achermann JC, Vilain E. Mutations in the PCNA-binding domain of CDKN1C cause IMAGe syndrome. Nat Genet 2012;44(7):788-92.

Conflict of Interest:

None declared

Re: Disruption of RAB40AL function leads to Martin-Probst syndrome, a rare X-linked multisystem neurodevelopmental human disorder. Bedoyan JK, Schaibley VM, Peng W, Bai Y, Mondal K, Shetty AC, Durham M, Micucci JA, Dhiraaj A, Skidmore JM, Kaplan JB, Skinner C, Schwartz CE, Antonellis A, Zwick ME, Cavalcoli JD, Li JZ, Martin DM. J Med Genet. 2012 May;49(5):332- 40.

With great interest we have read the article by Bedoyan and colleagues. The authors state that disruption of RAB40AL function by a dinucleotide missense change (chrX:102,079,078-102,079,079AC?GA p.D59G; hg18) causes Martin-Probst syndrome, a rare X-linked disorder characterised by deafness, cognitive impairment, short stature and distinct craniofacial dysmorphisms, among other clinical features. However, we have reasons to question that conclusion.

We have identified the same dinucleotide missense change in four out of 446 index patients from families with X-linked intellectual disability (XLID) by performing high-throughput sequencing of all X chromosome- specific exons. However, one patient in whom we had identified the p.D59G substitution also carried a deletion on chromosome 14 that unequivocally explained the clinical phenotype, which led us to speculate that in this family the RAB40AL change may not be causative of the phenotype. Therefore, we carried out segregation analysis in the remaining three families. One family was not informative for the RAB40L change. In this family both the clinical phenotype and segregation analyses of additionally identified variants suggested that this family more likely carries a pathogenic mutation in another XLID gene. In the remaining two families, Sanger sequencing confirmed the RAB40AL p.D59G substitution in the index patients. However, in each of these families, one of the affected male sibs did not carry the p.D59G substitution. Conversely, the exome variant server (ESP6500) and dbSNP database report two normal males who are hemizygous for the p.D59G substitution (dbSNP134 rs145606134). In yet another XLID family, we have observed a frameshift mutation (chrX:102192671-102192672 [hg19], ins1bp, p.A143GfsX12) in RAB40AL. In this family, in a healthy hemizygous male as well as several healthy homozygous females carried the frameshift variant.

Taken together, these data seem to indicate that the RAB40AL variants observed are not related to the XLID in our families. Moreover, they suggest that the RAB40AL p.D59G substitution may not be the cause of Martin-Probst syndrome but may represent a rare benign variant. While it is not trivial to prove that a given variant is NOT involved in XLID, the aggregate evidence presented here suggests that the role of RAB40AL in XLID and in particular, that of the p.D59G substitution in Martin-Probst syndrome should be revisited.

Conflict of Interest:

None declared

We would like to comment on Dixon et al., 2012 who questioned the pathogenicity of private NLRP7 variants found in a heterozygous state in singleton cases with one to three hydatidiform moles. The interesting part of this work is in the analysis of nine patients with recurrent androgenetic moles despite that only reports demonstrating that three of these patients had had recurrent androgenetic moles are provided either in the supplementary materials (for case 6)[1] or referenced in the Materials and Methods and had been previously reported by the same group (for case 2 in Buyukurt et al.[2] and case 1 in van der Smagt et al.[3]). Among patients with recurrent androgenetic moles, the authors found only one patient with a non-synonymous variant, R413Q that is not reported in the 1000 Genomes database. Curiously, we previously reported another novel DNA substitution affecting the same amino acid, R413W, in a patient with one mole and one live birth[5].

We agree with the authors that finding a rare or private variant in a patient but not in controls does not imply its pathogenicity. However, when several patients, each with a single rare variant, are found, and if these variants are not found in controls [4-6], such variants cannot be ignored, and the question of their pathogenicity should be raised and experimental methods addressing this question should be performed. Our group has reported ten different variants in heterozygous state in patients with one to three moles, but not in our controls. Determining the pathogenicity of private missense variants is very challenging in human genetics. Below, we discuss the arguments that are in favour of, or against, the pathogenicity of the private missense we reported in heterozygous state.

First, a good and simple genetic argument to determine if a variant is pathogenic is its absence in controls, which was done for every variant. The fact that some of these variants were later reported in some populations in the 1000 Genomes database is not against their involvement in the disease. Because in these populations, these variants were found at low frequencies that are very close to the incidences of sporadic moles in these populations. Basically, the definition of a susceptibility variant implies its presence in the general population and only a subset of the subjects carrying it will develop the disease.

Second, another good genetic argument is to show segregation of the variant in question in families with other mutations but on different haplotypes. This was done for one missense, A719V, that we reported in two patients in a heterozygous state[4, 5,6]. This variant was recently reported in a fourth patient and again in a heterozygous state and not in controls [7]. Definitely, more data are needed for the other variants.

Third, as mentioned by Dixon et al., the predicted pathogenicity scores by Polyphen-2 for seven of the ten variants we reported as single defective alleles are equal or higher than the predicted pathogenicity scores for missenses seen in patients with two defective alleles.

Fourth, to date, the only known functional role of NLRP7 is its requirement for normal IL1? secretion by peripheral blood mononuclear cells that we demonstrated in cells carrying different NLRP7 mutations [8] and was confirmed using independent in vitro studies [9]. To investigate whether the identified single rare variants in our patients affect IL1? secretion, we had performed the cytokine assay on patients cells carrying each of the four variants, C399Y, G380R, A719V (Fig. 1 in8), and R413W (unpublished data), in a heterozygous state, and again these cells displayed reduced IL1? and TNF secretion.

Fifth, the reproductive outcomes of patients with one defective allele are milder (approximately 81.6% reproductive wastage, 18.4% live birth) than those of patients with two defective alleles (97.5% reproductive wastage, 2.5% live births) [5] and more severe than those of patients with sporadic moles (10-22% reproductive wastage, 90-78% live births) [10].

One of the arguments used by Dixon et al., [1] is that androgenetic moles are not caused by inherited variants is the fact that androgenetic moles have not been seen in families. While we agree with this good argument, we believe that this may not be the only explanation. Another possibility is that a genetic defect leading to an androgenetic mole implies having an oocyte whose pronucleus, by as yet unknown mechanisms, is not able to meet and fuse with the male pronucleus and form a diploid biparental zygote. Such defect may be very severe, lethal, and not compatible with the life of the patient when caused only by a genetic defect in a homozygous state. However, in a heterozygous state, such defect would be expected to be milder, compatible with the life of the patient, and will confer genetic susceptibility to androgenetic moles only in few of her pregnancies when combined with other environmental and/or genetic factors.

Taking into consideration the emerging views about the mechanisms leading to androgenetic moles and the presence of complex postzygotic aneuploidies at their origin, it is therefore unlikely to see the same resulting parental contribution to products of conception occurring in other family members and consequently such conceptions will have other histopathological features and lead to other forms of reproductive wastage (such as spontaneous abortions, blighted ovum, ectopic pregnancies, etc.). This would make it difficult to health care professionals to connect these different clinical entities under the same unifying common defect segregating in the family. We did notice familial aggregation of reproductive wastage in the relatives of some patients with moles but these are difficult to detect and require well-prepared questionnaire and pedigrees. In few instances, we saw patients with sporadic moles who reported the occurrence of moles in their mothers. Although, it is impossible to review the histopathology of the mother's moles to validate the patients' statements, which is a real limitation in this research field, however, based on family history, there are rare cases with dominant transmission of a mild genetic susceptibility. Because males do not manifest moles, theoretically, only 50% of families of patients who carry predisposing genetic variants are at risk to manifest reproductive wastage and of these families only those with females who have tried to conceive may manifest the disease. For all these reasons and the genetic complexity of the mechanisms leading to androgenetic moles, we believe that a very small fraction of patients with a single missense "mutation" would be expected to have another member who had had androgenetic moles.

In conclusion, we agree with Dixon et al., that recurrent androgenetic moles do not seem to be caused by a strong causal genetic defect. However, we believe that there are several arguments, at least for the time being, indicating that some of the single rare variants in NLRP7 predispose the patients to moles and reproductive wastage. With the advances in this field, the accumulation of more data from various groups, and the development of additional functional tests to determine easily the pathogenicity of rare variants of unknown significance, patients with one mutation or one private variant can be assured that their chances of having live births are much higher than patients with two defective alleles5.

References

1 Dixon PH, Trongwongsa P, Abu-Hayyah S, Ng SH, Akbar SA, Khawaja NP, Seckl MJ, Savage PM, Fisher RA. Mutations in NLRP7 are associated with diploid biparental hydatidiform moles, but not androgenetic complete moles. J Med Genet 2012;49(3):206-11.

2 Buyukkurt S, Fisher RA, Vardar MA, Evruke C. Heterogeneity in recurrent complete hydatidiform mole: presentation of two new Turkish families with different genetic characteristics. Placenta 2010;31(11):1023 -5.

3 van der Smagt JJ, Scheenjes E, Kremer JA, Hennekam FA, Fisher RA. Heterogeneity in the origin of recurrent complete hydatidiform moles: not all women with multiple molar pregnancies have biparental moles. Bjog 2006;113(6):725-8.

4 Deveault C, Qian JH, Chebaro W, Ao A, Gilbert L, Mehio A, Khan R, Tan SL, Wischmeijer A, Coullin P, Xie X, Slim R. NLRP7 mutations in women with diploid androgenetic and triploid moles: a proposed mechanism for mole formation. Hum Mol Genet 2009;18(5):888-97.

5 Messaed C, Chebaro W, Roberto RB, Rittore C, Cheung A, Arseneau J, Schneider A, Chen MF, Bernishke K, Surti U, Hoffner L, Sauthier P, Buckett W, Qian J, Lau NM, Bagga R, Engert JC, Coullin P, Touitou I, Slim R. NLRP7 in the spectrum of reproductive wastage: rare non-synonymous variants confer genetic susceptibility to recurrent reproductive wastage. J Med Genet 2011;48(8):540-8.

6 Qian J, Cheng Q, Murdoch S, Xu C, Jin F, Chebaro W, Zhang X, Gao H, Zhu Y, Slim R, Xie X. The Genetics of Recurrent Hydatidiform Moles in China: Correlations between NLRP7 Mutations, Molar Genotypes, and Reproductive Outcomes. Mol Hum Reprod 2011;17(10):612-9.

7 Landolsi H, Rittore C, Philibert L, Hmissa S, Gribaa M, Touitou I, Yacoubi MT. NLRP7 mutation analysis in sporadic hydatidiform moles in Tunisian patients: NLRP7 and sporadic mole. Arch Pathol Lab Med 2012;136(6):646-51.

8 Messaed C, Akoury E, Djuric U, Zeng J, Saleh M, Gilbert L, Seoud M, Qureshi S, Slim R. NLRP7, a NOD-like receptor protein, is required for normal cytokine secretion and co-localizes with the Golgi and the microtubule organizing center. J Biol Chem 2011;286(50):43313-433123.

9 Khare S, Dorfleutner A, Bryan NB, Yun C, Radian AD, de Almeida L, Rojanasakul Y, Stehlik C. An NLRP7-containing inflammasome mediates recognition of microbial lipopeptides in human macrophages. Immunity 2012;36(3):464-76.

10 Lan Z, Hongzhao S, Xiuyu Y, Yang X. Pregnancy outcomes of patients who conceived within 1 year after chemotherapy for gestational trophoblastic tumor: a clinical report of 22 patients. Gynecol Oncol 2001;83(1):146-8.

Conflict of Interest:

None declared

We have read the entire paper with great care. We totally agree with the findings of the respected authors in this respective article. The article seems to be used by many researchers as a reference article so we need to update the status of SMA diagnosis for non deleted SMA patients. Many new approaches have been employed to diagnose non deleted SMA. One of such methods is long range PCR method (LR-PCR) (Clemont et al., 2004). Secondly, we can not just focus on the mutations described in this article. We need to focus on all known mutations in the SMN1 gene in non deleted SMA patients. Although Clermont and colleagues claimed that 90% of the mutations in the SMN1 gene can be found by their method (Clermont et al.,2004), yet many other groups have reported individual mutations. We would like to enumerate the list of all these mutations. All the mutations are divided into four main groups; missence mutation, Nonsense mutation, Frame-shift mutation and Splice site mutations.

The missence mutations (total 25) have been reported in Ex.1 in type II and III SMA patients (Parsons et al., 1998), Ex. 2a in type II and III SMA patients (Sun et al., 2005), Ex.3 in type I (Cusco et al., 2004, Clermont et al., 2004, Prior et al., 2007, Sun et al., 2005, Koatni et al., 2007), Ex.3 in type II (Clermont et al., 2004, Sun et al., 2005), Ex.3 in type III SMA patient (Sun et al., 2005, Ex.4 in type I SMA patient (Zapletalova et al., 2007), Ex. 6 in type III SMA patients (Rochette et al., 1997, Clermont et al., 2004, Sun et al., 2005, Hahnen et al., 1997, Parson et al., 1998, Wirth et al., 1999), Ex.6 in type I SMA patient (Clermont et al., 2004), Ex.6 in type II SMA patient (Alias et al., 2009, Prior et al., 2007, Lefebvre et al., 1995, Wirth et al., 1999, Rochette et al., 1997, Clermont et al., 2004, Zapletalova et al., 2007, Hahnen et al., 1997, Parson et al., 1998, Sun et al., 2005, Ex.6 in type III SMA patients (Hahnen et al., 1997, Parsons et al., 1998, Wirth et al., 1999, Sun et al., 2005, Zapletalova et al., 2007, Alias et al., 2009, Burglen et al., 1996, Skordiset et al., 2001), Ex.7 in type I SMA patient (Talbot et al., 1997) and Ex.7 in type II and III SMA patients (Wang et al., 1998). The nonsense mutations (total 5) were curated to be included in Ex.1 in type I SMA patient (Wirth et al., 1999), Ex.1 in type III SMA patient (Sun et al., 2005), Ex.3 in type II SMA patient (Sossi et al., 2001, Prior et al., 2007), Ex.3 in type III SMA patient (Wirth et al., 1999, Sun et al., 2005, Sossi et al., 2001, Prior et al., 2007), Ex.3 in type I SMA patient (Bricchta et al., 2008), Ex.4 in type I SMA patient (Alias et al., 2009), Ex.5 in type I SMA patient (Tsai et al., 2001). Considering the fram-shift mutations (total 18), it included mutation in Ex.1 of type I SMA patient (Wirth et al., 1999), Ex.1 in type III SMA patient (Wirth et al., 1999), Ex.2a in type I SMA patient (Prior et al., 2007, Zapletalova et al., 2007), Ex.2a in type I SMA patient (Wirth et al., 1999, Alias et al., 2008), Ex.2b in type I SMA patient (Clermont et al., 2004), Ex. 2b in type III SMA patient (Wirth et al., 1999), Ex.3 in type I SMA patient (Brahe et al., 1996, Sossi et al., 2001, Alias et al., 2008), Ex.3 in type II SMA patient (Bussaglia et al., 1995, Martin et al., 2002, Cusco et al., 2003, Alias et al., 2008), Ex.3 in type III SMA patient (Alias et al., 2008, Clermont et al., 2004, Bussaglia et al., 1995, Martin et al., 2002, Cusco et al., 2003), Ex.4 in type I SMA patient (Clermont et al., 1997, Clermont et al., 2004), Ex.4 in type II SMA patient (Parons et al., 1998, Wirth et al., 1999), Ex.4 in type III SMA patient (Parson et al., 1998), Ex.5 in type I, II and III SMA patients (Wirth et al., 1999, Clermont et al., 2004), Ex.6 in type I, II and III SMA patients (Wirth et al., 1999, Clermont et al., 2004, Parsons et al., 1996, Parsons et al., 1998, Martin et al 2002, Clermont et al., 2004, Alias et al., 2008, Martin et al., 2002). The splice site mutations in the SMN1 gene in non deleted SMA patients include 40Int.4 in type I SMA patient (Brichta et al., 2008), 41Int.6 in type I SMA patient (Martin et al., 2002), 42Int.6 in type I SMA patient (Eggermann et al., 2008), 43Int.6 in type I SMA patient (Lefebvre et al., 1995), 44Int.7 in type I SMA patient (Wirth et al., 1999) and 45Int.7 in type II SMA patient (Lefebvre et al., 1995). The copy number analysis of the SMN1 gene is not enough as the SMA modified genes; SMN2 and NAIP are reported to modify the disease severity (Watihayati et al., 2009). We will emphasize that the title of the article is making the readers confusing as considering the article to be the diagnostic protocol for the spinal muscular atrophy in non deleted SMA patients. The severity of the disease could be estimated by many parameters even in the non deleted SMA patients and recently published report has prooven that (Watihayati et al., 2009) beside the report of Teguh and colleagues which states that only PCR detection of exon7 of the SMN1 is enough to diagnose SMA (Teguh et al., 2011) but yet the authors did not explain the diagnosis of non deleted SMA patients. Very humbly we would like to add this response with a request to the respected authors to consider the findings of SMN2 copy number and the NAIP gene deletion with the available data of this patient.

Conflict of Interest:

None declared