Historical and geographical distribution of the founder mutation c.610G>A; p.Ala204Thr in the CLCNKB gene linked to Bartter syndrome type III in Spain.

Bartter syndrome (BS) type III caused by variation within CLCNKB has a heterogeneous presentation, extending from severe to very mild, and is the most common in Spain [1-4]. The p. Ala204Thr variation is the most recurrent in our country and is only observed in Spanish descendants, suggesting a founder effect [1-4].

A cohort of 21 Spanish patients with a clinical history consistent with BS was analysed and 15 patients from different unrelated families were characterized as BS type III (Supplementary data, Table S1). Thirteen out of 15 (87%) are carrying the variant c.610G>A; p.Ala204Thr (Supplementary data, Figure S1) in homozygosis or as a part of a compound heterozygous. Characterization of the CLCNKB locus and its neighbourhood by SNP arrays showed a shared haplotype of seven single nucleotide polymorphisms (SNPs) (Figure 1) that co-segregates with this variant in the affected individuals, a fact that strongly reflects the presence of a founder mutation and excludes the alternative hypothesis of a hot spot. We noted that all the subjects' ancestry originated from an area of the centre of Spain, where all the localities were founded between 10th and 14th centuries (afterward repopulated as a result of The Reconquest), which are crossed by historical-pecuary routes (Cañadas Reales; Table 1 and Figure 2). For hundreds of years, a seasonal movement (North–South and vice versa) of people along these livestock roads took place (the transhumance), suggesting a regional founder effect of this variant. Remarkably, our cohort’s grandparents were likely to have been born in the decades either side of 1920–30, so the spatial distribution of the genetic structure described in this study would reflect that the genetic structure seen within Spain around the beginning of the 20th century is a result of regional genetic isolation.

FIGURE 1:

Haplotypes reconstruction (1 to 5) relative to seven markers (SNPs) regarding NM_000085.4: NM_000085.4:c.610G>A(p.Ala204Thr) mutation in the CLCNKB of three families type (as example). A) SNP markers selected and position into CLCNKA/B locus. From the top: rs2268341, rs1739822, rs1739828, rs1763613, rs1763612, rs1739839, rs1544131, and their poblational frequencies (source: gnomAD). B) Homozygous consanguineous family, C) Homozoygous non-consanguineous familiy, D) Compound heterozygous case, E) Compound heterozygous case without including c.610G>A.

Table 1.

Genetic characteristics, geographical and historical ancestry origin of 13 patients with p.Ala204Thr

Case	Age/sex	Paternal mutation	Maternal mutation	Father ancestry origin/century	Mother ancestry origin/century
				Cañada	Cañada
1	49/Male	p.Ala204Thr	p.Ala204Thr	San Pablo de los Montes (Toledo)/13th	San Pablo de los Montes (Toledo)/ 13th
				Cañada Real Segoviana (4)	Cañada Real Segoviana (4)
2	26/Male	p.Ala204Thr	p.Ala204Thr	Cantalejo (Segovia)/10th	Cuellar (Segovia)/11th
				Cañada Real Soriana Occidental (6)	Cañada Real Leonesa Oriental (3)
3	39/Male	p.Ala204Thr	p.Ala204Thr	El Casar de Escalona (Toledo)/12th	Pozuelo del Rey (Madrid)/12th
				Cañada Real Leonesa Oriental (3)	Cañada Real Galiana (7)
4	40/Female	p.Ala204Thr	p.Ala204Thr	Urda (Toledo)/11th	Urda (Toledo)/11th
				Cañada Real Leonesa Oriental (3)	Cañada Real Leonesa Oriental (3)
5	62/Female	p.Ala204Thr	p.Ala204Thr	Fuencarral (Madrid)/12th	Medina del Campo (Valladolid)/11th
				Cañada Real Galiana (7)	Cañada Real Leonesa Occidental (2)
6	21/Female	p.Ala204Thr	p.Ala204Thr	Caleruega (Burgos)/10th	Caleruega (Burgos)/10th
				Cañada Real Segoviana (4)	Cañada Real Segoviana (4)
7	39/Female	p.Ala204Thr	p.Met1_His654del	San Pedro de Gaillos (Segovia)/10th	Ferrol (Galicia)
				Cañada Real Soriana Occidental (6)
8	12/Male	p.Ala204Thr	p.Ala204Thr	Valdemanco (Madrid)/14th	Valdemanco (Madrid)/14th
				Cañada Real Segoviana (4)	Cañada Real Segoviana (4)
9	11/Female	p.Glu442Gly	p.Ala204Thr	Madrid-Jaen	Madrid, Fuensalida (Toledo)/11th, 13th
					Cañada Real Segoviana (4)
					Cañada Real Galiana (7)
10	52/Female	p.Ala204Thr	p.Ala204Thr	Corral de Almaguer (Toledo)/13th	Corral de Almaguer (Toledo)/13th
				Cañada Real Soriana Oriental (5)	Cañada Real Soriana Oriental (5)
11	37/Male	p.Ala204Thr	p.Ala204Thr	Almagro (Ciudad Real)/13th	Bolaños de Calatrava (Ciudad Real)/13th
				Cañada Real Soriana Oriental (5)	Cañada Real Soriana Oriental (5)
				Cañada Real de Cuenca (8)	Cañada Real de Cuenca (8)
12	26/Female	p.Ala204Thr	p.Ala204Thr	Guzmán (Burgos)/11th	Madrigalejo (Cáceres)/11th
				Cañada Real Segoviana (4)	Cañada Real de La Plata (1)
					Cañada Real Leonesa Occidental (2)
13	57/Female	p.Leu439Pro	p.Ala204Thr	Mundaca (Bilbao)	Alcalá de Henares (Madrid)/11th
					Cañada Real Galiana (7)

FIGURE 2:

Historical-geographical relationships between Spanish type III Bartter syndrome patients studied (dot blue), who carries the NM_000085.4:c.610G>A (p.Ala204Thr) mutation in the CLCNKB gene and historical pathways (Cañadas Reales). Each individual is represented by a dot placed the centroid of their grandparents’ birthplaces. The mutation can be related to a transhumance population established into or in the vicinities of pathways of historical importance (Cañadas Reales).

The cultural and linguistic impact of Muslim rule in the Iberian Peninsula is well-known but limited to the extent, timing and geographic spread of genetic mixing between immigrants and Iberians over several centuries after the initial Muslim conquest (711 CE) [5, 6]. During The Reconquest, Christian-controlled territory in the North moved gradually Southwards from the mid-8th century. The Castilian kings moved in with their armies but in what amounted to a veritable frontier movement, struggled to repopulate their new territories. Transhumance could thrive in this largely unpopulated landscape. The population movements were facilitated by the subsequent establishment of ‘The Honourable Concejo de la Mesta’ (year 1273 CE) by King Alfonso X (The Wise), regulating the migration of people through historical pathways (Cañadas Reales). Interestingly, the distribution of p.Ala204Thr in the BS population was strikingly very similar to that established for different populations and dialects presented around 1300 CE (Old Kingdom of Castile) [5, 6]. Since p.Ala204Thr is not found in populations from North Africa, Arab origin or Sephardic-Jewish ancestry, this mutation seems to have originated in the primitive population of Spain, who either remained unmixed with the Muslim invaders and Jewish people or originated after the onset of The Reconquest (which finished in 1492 CE) [5, 6], spreading with the re-population of the reconquered lands and establishment of the transhumance. In addition, the p.Ala204Thr variation is also present in Latin American individuals with Spanish ancestry, but exclusively in a heterozygous way (source: gnomAD). This fact, along with finding several heterozygous carriers (for p. Ala204Thr) in individuals with ancestry originated from Mestanza (a town founded in the 11th century, localized at the South of Ciudad Real) and a dot of confluence of Cañada Real Segoviana (4) and Cañada Real Soriana Oriental (5) (Figure 2), seemed to support our hypothesis. Our data are also consistent with studies establishing a strong association between specific-unique mutations and population founder effects on BS [7-9], such as in Korea and Japan [8, 9] or Costa Rican populations [10] (Supplementary data, Table S2).

Our findings support the p.Ala204Thr variant as a founder mutation originating from the central part of Spain, and it is conceivable that the greater concentration of individuals within this area was the consequence of an offspring from a common ancestor. Thus, the knowledge of the spectrum of mutations and their geographical distribution of this variant will allow us a more effective detection strategy in individuals with BS type III, or extend it to other countries with large Spanish populations.

SUPPLEMENTARY DATA

Supplementary data are available at ckj online.

Click here for additional data file.