In the present study, we demonstrate for the first time that WES can be seen as a relevant alternative for molecular diagnosis of NDM. Since an accurate molecular diagnosis for this condition can lead to very dramatic improvements in patient care, development of reliable and cost efficient methods for quick and accurate DNA analysis are of major interest.
Currently, patients with NDM are evaluated using Sanger sequencing, which is far more expensive per sequenced base-pairs than WES. Indeed, the standard sequencing of a single PCR fragment costs 67.50€ ($82) for the only French hospital laboratory specialized in NDM molecular diagnosis (Robert Debré hospital, Paris). This price includes consumables, equipment amortization, personnel salary and hospital overhead costs. Therefore, the total cost for the French National Insurance for the sequencing of KCNJ11, ABCC8 and INS alone, which requires 42 PCR fragments is 2,835€ ($3,440; ~0.45€ or ~$0.55 per bp). In comparison, the all inclusive cost of WES for NDM, which will detect mutations in KCNJ11, ABCC8 and INS as well as rarer genetic aetiologies of NDM, is currently 3,274€ ($4,146; <<0.001€ or $ per bp) per sample, by performing a sequencing on two channels and in 76 bp paired-end configuration (CNRS-UMR8199, Lille, France). Moreover, it is very likely that WES cost will fall in next months towards 2,000€ ($2,528) or even less.
We believe that the WES protocol is less labour intensive and time-consuming than the standard Sanger protocol for genetically heterogeneous disorders requiring several large genes to be screened. A WES run involving four DNA samples can be completed in two weeks, including the time required to analyse the data, which is comparable to the time required by current Sanger sequencing of ABCC8 only with its 39 exons. The p.Q485H mutation was missed six years ago by the research assistant in charge of the sequence reading. Although we can assume that mutation detection bio-informatics tools were less efficient a few years ago (the hospital laboratory used the PhredPhrap software in 2004) and that current methods are more accurate, the Sanger protocol and specially the semi-automated reading of sequence traces is always laborious and demanding (thus expensive), and a double-check of sequence readings by two different persons is performed in several French diagnostic laboratories in order to avoid any errors in the mutation identification process.
WES method is not only a cost-effective tool for molecular diagnosis; it should be also seen as an excellent tool for further genetic research and identification of novel causal mutations. Indeed, in the French NDM cohort, half of PNDM cases are still not elucidated 
. Classical linkage analyses are generally not successful as many NDM mutations occur de novo
or are not fully penetrant. Most NDM genes have been found via candidate gene analyses but this approach has now reached its limits 
. However, WES typically yields thousands of ‘novel’ genetic variants (i.e.
not yet present in human genome variants databases). Therefore, the identification of truly causal variants would be strongly facilitated by the development of a high quality WES database of novel mutations found in both elucidated cases or in cases of unknown aetiology as well as in controls coming from same ethnicity. WES would also permit the identification of putative NDM modifier genes, a very challenging task for targeted gene analysis.
We are quite confident that the p.Q485H mutation is likely to be functional given the non ambiguous prediction of its putative damaging effect. In addition, the clinical data from the patient fit well with the features of PNDM linked to a ABCC8
mutation (e.g. de novo
mutation associated with very early-onset of the disease and attention disorder) 
We believe that other NDM patients should be assessed with the same protocol as DNA quality may change the WES accuracy. Also, our DNA capture was not totally perfect as we could miss approximately 6% of exomic SNPs present in the high-resolution oligonucleotide genotyping array. Furthermore, the exomic coverage was not homogeneous between NDM genes (), thus we could suspect that the WES accuracy would not be the same for all NDM genes. Therefore, despite high WES mean coverage and elevated rates of both concordance and sensitivity in mutation detection, it is also necessary to assess and to verify the homogeneity of the target capture, specially in the genes of interest that have to be screened for molecular diagnosis ().
Details on exomic sequencing depth in NDM genes, obtained through the WES of the PNDM patient.
Knowing that the capture technology is improving day after day (by enriching exomic loci poorly captured with the previous kits), our present study suggests that it will be possible to soon update the protocols for molecular diagnosis of NDM 
. We propose that after discovery of severe hyperglycemia in a neonate who is negative for serological markers of type 1 diabetes, a preliminary assessment of abnormalities of chromosome 6q24 can be performed (as at this stage, it is too early to guess NDM will be permanent or transient) followed by the search of a mutation in both KCNJ11
using Sanger sequencing as these two genes can be easily and quickly sequenced. If negative, we propose a WES analysis of the patient's DNA which is the most comprehensive way to fully explore the molecular causes of this NDM case.