Collaborative model for diagnosis and treatment of very rare diseases: experience in Spain with thymidine kinase 2 deficiency.

BACKGROUND: Mitochondrial diseases are difficult to diagnose and treat. Recent advances in genetic diagnostics and more effective treatment options can improve patient diagnosis and prognosis, but patients with mitochondrial disease typically experience delays in diagnosis and treatment. Here, we describe a unique collaborative practice model among physicians and scientists in Spain focused on identifying TK2 deficiency (TK2d), an ultra-rare mitochondrial DNA depletion and deletions syndrome. MAIN BODY: This collaboration spans research and clinical care, including laboratory scientists, adult and pediatric neuromuscular clinicians, geneticists, and pathologists, and has resulted in diagnosis and consolidation of care for patients with TK2d. The incidence of TK2d is not known; however, the first clinical cases of TK2d were reported in 2001, and only ~ 107 unique cases had been reported as of 2018. This unique collaboration in Spain has led to the diagnosis of more than 30 patients with genetically confirmed TK2d across different regions of the country. Research affiliate centers have led investigative treatment with nucleosides based on understanding of TK2d clinical manifestations and disease mechanisms, which resulted in successful treatment of a TK2d mouse model with nucleotide therapy in 2010. Only 1 year later, this collaboration enabled rapid adoption of treatment with pyrimidine nucleotides (and later, nucleosides) under compassionate use. Success in TK2d diagnosis and treatment in Spain is attributable to two important factors: Spain's fully public national healthcare system, and the designation in 2015 of major National Reference Centers for Neuromuscular Disorders (CSURs). CSUR networking and dissemination facilitated development of a collaborative care network for TK2d disease, wherein participants share information and protocols to request approval from the Ministry of Health to initiate nucleoside therapy. Data have recently been collected in a retrospective study conducted under a Good Clinical Practice-compliant protocol to support development of a new therapeutic approach for TK2d, a progressive disease with no approved therapies. CONCLUSIONS: The Spanish experience in diagnosis and treatment of TK2d is a model for the diagnosis and development of new treatments for very rare diseases within an existing healthcare system.

Genome-wide assessment of diversity and differentiation between original and modern Brown cattle populations.

Identifying genomic regions involved in the differences between breeds can provide information on genes that are under the influence of both artificial and natural selection. The aim of this study was to assess the genetic diversity and differentiation among four different Brown cattle populations (two original vs. two modern populations) and to characterize the distribution of runs of homozygosity (ROH) islands using the Illumina Bovine SNP50 BeadChip genotyping data. After quality control, 34 735 SNPs and 106 animals were retained for the analyses. Larger heterogeneity was highlighted for the original populations. Patterns of genetic differentiation, multidimensional scaling, and the neighboring joining tree distinguished the modern from the original populations. The FST -outlier identified several genes putatively involved in the genetic differentiation between the two groups, such as stature and growth, behavior, and adaptability to local environments. The ROH islands within both the original and the modern populations overlapped with QTL associated with relevant traits. In modern Brown (Brown Swiss and Italian Brown), ROH islands harbored candidate genes associated with milk production traits, in evident agreement with the artificial selection conducted to improve this trait in these populations. In original Brown (Original Braunvieh and Braunvieh), we identified candidate genes related with fat deposition, confirming that breeding strategies for the original Brown populations aimed to produce dual-purpose animals. Our study highlighted the presence of several genomic regions that vary between Brown populations, in line with their different breeding histories.

Comparative selection signature analyses identify genomic footprints in Reggiana cattle, the traditional breed of the Parmigiano-Reggiano cheese production system.

Reggiana is an autochthonous cattle breed reared mainly in the province of Reggio Emilia, located in the North of Italy. Reggiana cattle (originally a triple-purpose population largely diffused in the North of Italy) are characterised by a typical solid red coat colour. About 2500 cows of this breed are currently registered to its herd book. Reggiana is now considered a dual-purpose breed even if it is almost completely dedicated to the production of a mono-breed branded Protected Designation of Origin Parmigiano-Reggiano cheese, which is the main driver of the sustainable conservation of this local genetic resource. In this study, we provided the first overview of genomic footprints that characterise Reggiana and define the diversity of this local cattle breed. A total of 168 Reggiana sires (all bulls born over 35 years for which semen was available) and other 3321 sires from 3 cosmopolitan breeds (Brown, Holstein and Simmental) were genotyped with the Illumina BovineSNP50 panel. ADMIXTURE analysis suggested that Reggiana breed might have been influenced, at least in part, by the other three breeds included in this study. Selection signatures in the Reggiana genome were identified using three statistical approaches based on allele frequency differences among populations or on properties of haplotypes segregating in the populations (fixation index (FST); integrated haplotype score; cross-population extended haplotype homozygosity). We identified several regions under peculiar selection in the Reggiana breed, particularly on bovine chromosome (BTA) 6 in the KIT gene region, that is known to be involved in coat colour pattern distribution, and within the region of the LAP3, NCAPG and LCORL genes, that are associated with stature, conformation and carcass traits. Another already known region that includes the PLAG1 gene (BTA14), associated with conformation traits, showed a selection signature in the Reggiana cattle. On BTA18, a signal of selection included the MC1R gene that causes the red coat colour in cattle. Other selection sweeps were in regions, with high density of quantitative trait loci for milk production traits (on BTA20) and in several other large regions that might have contributed to shape and define the Reggiana genome (on BTA17 and BTA29). All these results, overall, indicate that the Reggiana genome might still contain several signs of its multipurpose and non-specialised utilisation, as already described for other local cattle populations, in addition to footprints derived by its ancestral origin and by its adaptation to the specialised Parmigiano-Reggiano cheese production system.

Genome-wide analyses reveal the regions involved in the phenotypic diversity in Sicilian pigs.

Nero Siciliano (Sicilian Black, SB) is a local pig breed generally of uniform black color. In addition to this officially recognized breed, there are animals showing morphological characteristics resembling the SB but with gray hair (Sicilian Grey, SG). The SG, compared with the SB, also shows a more compact structure with greater transverse diameters, higher average daily gains and lower thickness of the back fat. In this study, using the Illumina PorcineSNP60 BeadChip, we run genome-wide analyses to identify regions that may explain the phenotypic differences between SB (n = 21) and SG (n = 27) individuals. Combining the results of the two case-control approaches (GWAS and FST ), we identified two significant regions, one on SSC5 (95 401 083 bp) and one on SSC15 (55 051 435 bp), which contains several candidate genes related to growth traits in pig. The results of the Bayesian population differentiation approach identified a marker near the MGAT4C, a gene associated with average daily gain in pigs. Finally, scanning the genome for runs of homozygosity islands, we found that the two groups have different runs of homozygosity islands, with several candidate genes involved in coat color (in SG) or related to different pig performance traits (in SB). In summary, the two analyzed groups differed for several phenotypic traits, and genes involved in these traits (growth, meat traits and coat color) were detected. This study provided another contribution to the identification of genomic regions involved in phenotypic variability in local pig populations.

Deoxynucleoside Therapy for Thymidine Kinase 2-Deficient Myopathy.

OBJECTIVE: Thymidine kinase 2, encoded by the nuclear gene TK2, is required for mitochondrial DNA maintenance. Autosomal recessive TK2 mutations cause depletion and multiple deletions of mtDNA that manifest predominantly as a myopathy usually beginning in childhood and progressing relentlessly. We investigated the safety and efficacy of deoxynucleoside monophosphate and deoxynucleoside therapies. METHODS: We administered deoxynucleoside monophosphates and deoxynucleoside to 16 TK2-deficient patients under a compassionate use program. RESULTS: In 5 patients with early onset and severe disease, survival and motor functions were better than historically untreated patients. In 11 childhood and adult onset patients, clinical measures stabilized or improved. Three of 8 patients who were nonambulatory at baseline gained the ability to walk on therapy; 4 of 5 patients who required enteric nutrition were able to discontinue feeding tube use; and 1 of 9 patients who required mechanical ventilation became able to breathe independently. In motor functional scales, improvements were observed in the 6-minute walk test performance in 7 of 8 subjects, Egen Klassifikation in 2 of 3, and North Star Ambulatory Assessment in all 5 tested. Baseline elevated serum growth differentiation factor 15 levels decreased with treatment in all 7 patients tested. A side effect observed in 8 of the 16 patients was dose-dependent diarrhea, which did not require withdrawal of treatment. Among 12 other TK2 patients treated with deoxynucleoside, 2 adults developed elevated liver enzymes that normalized following discontinuation of therapy. INTERPRETATION: This open-label study indicates favorable side effect profiles and clinical efficacy of deoxynucleoside monophosphate and deoxynucleoside therapies for TK2 deficiency. ANN NEUROL 2019;86:293-303.

Genome-wide association studies for milk production traits in Valle del Belice sheep using repeated measures.

Genome-wide association studies (GWASes) have become a powerful tool for identifying genomic regions associated with important traits in livestock. Milk production traits in dairy sheep are measured at different time points during their life span. Using phenotypic data generated from longitudinal traits could improve the power of association studies but until now have received less attention in GWASes as a methodology and has not been implemented. The aim of this study was to carry out a GWAS for milk production traits in Valle del Belice sheep using repeated measures. After quality control, 469 ewes and 37 228 SNPs were retained for the analysis, and phenotypic data included 5586 test-day records for five milk production traits (milk yield, MY; fat yield and percentage, FY and F%; protein yield and percentage, PY and P%). Nine SNPs located within or close to known genes were found to be associated with milk production traits. In particular, rs398340969, associated with both milk yield and protein yield, is located within the DCPS gene. In addition, rs425417915 and rs417079368, both associated with both fat percentage and protein percentage, are located within the TTC7B gene and at 0.37 Mb within the SUCNR1 gene respectively. In summary, the use of repeated records was beneficial for mapping genomic regions affecting milk production traits in the Valle del Belice sheep.

Genome-wide association study reveals the locus responsible for microtia in Valle del Belice sheep breed.

Microtia is a congenital deformity of the outer ear with phenotypes varying from a small auricle to total absence (anotia). The genetic basis is still poorly understood, and very few studies have been performed in sheep. Valle del Belice sheep is a breed showing microtia. The aim of this study was to identify the potential genomic regions involved in microtia in sheep. A total of 40 individuals, 20 with microtia and 20 normal, were genotyped with the Illumina OvineSNP50 BeadChip. The comparison among the results from a genome-wide association study, Fisher's exact test and FST analysis revealed a single strong association signal: rs419889303 on chromosome 1, located within intron 3 of the CLRN1 gene. Our study suggests for the first time that this novel candidate gene is responsible for microtia in sheep. Additional analysis based on the sequencing would help confirm our findings and allow for the proposal of a precise genetic basis for microtia in sheep.

Genome-wide identification of runs of homozygosity islands and associated genes in local dairy cattle breeds.

Runs of homozygosity (ROH) are widely used as predictors of whole-genome inbreeding levels in cattle. They identify regions that have an unfavorable effect on a phenotype when homozygous, but also identify the genes associated with traits of economic interest present in these regions. Here, the distribution of ROH islands and enriched genes within these regions in four dairy cattle breeds were investigated. Cinisara (71), Modicana (72), Reggiana (168) and Italian Holstein (96) individuals were genotyped using the 50K v2 Illumina BeadChip. The genomic regions most commonly associated with ROHs were identified by selecting the top 1% of the single nucleotide polymorphisms (SNPs) most commonly observed in the ROH of each breed. In total, 11 genomic regions were identified in Cinisara and Italian Holstein, and eight in Modicana and Reggiana, indicating an increased ROH frequency level. Generally, ROH islands differed between breeds. The most homozygous region (>45% of individuals with ROH) was found in Modicana on chromosome 6 within a quantitative trail locus affecting milk fat and protein concentrations. We identified between 126 and 347 genes within ROH islands, which are involved in multiple signaling and signal transduction pathways in a wide variety of biological processes. The gene ontology enrichment provided information on possible molecular functions, biological processes and cellular components under selection related to milk production, reproduction, immune response and resistance/susceptibility to infection and diseases. Thus, scanning the genome for ROH could be an alternative strategy to detect genomic regions and genes related to important economic traits.

Runs of homozygosity reveal genome-wide autozygosity in Italian sheep breeds.

The availability of dense single nucleotide polymorphism (SNP) assays allows for the determination of autozygous segments based on runs of consecutive homozygous genotypes (ROH). The aim of the present study was to investigate the occurrence and distribution of ROH in 21 Italian sheep breeds using medium-density SNP genotypes in order to characterize autozygosity and identify genomic regions that frequently appeared in ROH within individuals, namely ROH islands. After filtering, the final number of animals and SNPs retained for analyses were 502 and 46 277 respectively. A total of 12 302 ROH were identified. The mean number of ROH per breed ranged from 10.58 (Comisana) to 44.54 (Valle del Belice). The average length of ROH across breeds was 4.55 Mb and ranged from 3.85 Mb (Biellese) to 5.51 Mb (Leccese). Valle del Belice showed the highest value of inbreeding on the basis of ROH (FROH  = 0.099), whereas Comisana showed the lowest (FROH  = 0.016), and high standard deviation values revealed high variability in autozygosity levels within each breed. Differences also existed in the length of ROH. Analysis of the distribution of ROH according to their size showed that, for all breeds, the majority of the detected ROH were <10 Mb in length, with a few long ROH >25 Mb. The levels of ROH that we estimated here reflect the inbreeding history of the investigated sheep breeds. These results also highlight that ancient and recent inbreeding have had an impact on the genome of the Italian sheep breeds and suggest that several animals have experienced recent autozygosity events. Comisana and Bergamasca appeared as the less consanguineous breeds, whereas Barbaresca, Leccese and Valle del Belice showed ROH patterns typically produced by recent inbreeding. Moreover, within the genomic regions most commonly associated with ROH, several candidate genes were detected.

Penalized classification for optimal statistical selection of markers from high-throughput genotyping: application in sheep breeds.

The identification of individuals' breed of origin has several practical applications in livestock and is useful in different biological contexts such as conservation genetics, breeding and authentication of animal products. In this paper, penalized multinomial regression was applied to identify the minimum number of single nucleotide polymorphisms (SNPs) from high-throughput genotyping data for individual assignment to dairy sheep breeds reared in Sicily. The combined use of penalized multinomial regression and stability selection reduced the number of SNPs required to 48. A final validation step on an independent population was carried out obtaining 100% correctly classified individuals. The results using independent analysis, such as admixture, F st, principal component analysis and random forest, confirmed the ability of these methods in selecting distinctive markers. The identified SNPs may constitute a starting point for the development of a SNP based identification test as a tool for breed assignment and traceability of animal products.

Monurelle Biogel® vaginal gel in the treatment of vaginal dryness in postmenopausal women.

OBJECTIVES: To evaluate efficacy, tolerability and safety of Monurelle Biogel® vaginal gel for treatment of vaginal dryness. METHODS: Multicenter, national, randomized, controlled vs. no-treatment, open-label study. Ninety-five postmenopausal women were randomized (48 to Monurelle Biogel® and 47 to no treatment). Primary endpoint was the change of Verbal Rating Scale (VRS) total score of vaginal atrophy (VA) symptoms after 8-week treatment. The main secondary endpoints were VRS single-item score, Vaginal Health Index (VHI) score, Maturation Index (MI), Female Sexual Function Index (FSFI), and Female Sexual Distress Scale-Revised (FSDS-R). RESULTS: The VRS total score was statistically significant in favor of the treatment group on day 28 (p = 0.001) but not on day 56 (p = 0.064). By excluding women who were not sexually active, the total VRS scores reached the criteria for clinical success in 27/43 subjects (62.8%) in the control arm and in 38/46 subjects (82.6%) in the treatment arm (p = 0.035) on day 56. The VHI score significantly changed in the active arm (4.71 ± 4.85 vs. 0.28 ± 1.71) (p < 0.001) on day 56. Even the MI significantly improved, with an increase in the percentage of superficial cells (p = 0.01). The improvements in both VHI and MI were still present at the follow-up visit after the discontinuation of the treatment (day 84). Sexual function and distress showed a statistical significant difference on day 56. CONCLUSIONS: Monurelle Biogel® vaginal gel applied twice daily for 8 weeks is effective in relieving vaginal dryness and other VA symptoms. Such a clinical meaningful effect persists at least 4 weeks and is supported by an improvement in the vaginal environment. Trial Registration clinicaltrials.gov Identifier: NCT02994342.

Genome-wide analysis in endangered populations: a case study in Barbaresca sheep.

Analysis of genomic data is becoming increasingly common in the livestock industry and the findings have been an invaluable resource for effective management of breeding programs in small and endangered populations. In this paper, with the goal of highlighting the potential of genomic analysis for small and endangered populations, genome-wide levels of linkage disequilibrium, measured as the squared correlation coefficient of allele frequencies at a pair of loci, effective population size, runs of homozygosity (ROH) and genetic diversity parameters, were estimated in Barbaresca sheep using Illumina OvineSNP50K array data. Moreover, the breed's genetic structure and its relationship with other breeds were investigated. Levels of pairwise linkage disequilibrium decreased with increasing distance between single nucleotide polymorphisms. An average correlation coefficient <0.25 was found for markers located up to 50 kb apart. Therefore, these results support the need to use denser single nucleotide polymorphism panels for high power association mapping and genomic selection efficiency in future breeding programs. The estimate of past effective population size ranged from 747 animals 250 generations ago to 28 animals five generations ago, whereas the contemporary effective population size was 25 animals. A total of 637 ROH were identified, most of which were short (67%) and ranged from 1 to 10 Mb. The genetic analyses revealed that the Barbaresca breed tended to display lower variability than other Sicilian breeds. Recent inbreeding was evident, according to the ROH analysis. All the investigated parameters showed a comparatively narrow genetic base and indicated an endangered status for Barbaresca. Multidimensional scaling, model-based clustering, measurement of population differentiation, neighbor networks and haplotype sharing distinguished Barbaresca from other breeds, showed a low level of admixture with the other breeds considered in this study, and indicated clear genetic differences compared with other breeds. Attention should be given to the conservation of Barbaresca due to its critical conservation status. In this context, genomic information may have a crucial role in management of small and endangered populations.

Genomic inbreeding estimation in small populations: evaluation of runs of homozygosity in three local dairy cattle breeds.

In the local breeds with small population size, one of the most important problems is the increase of inbreeding coefficient (F). High levels of inbreeding lead to reduced genetic diversity and inbreeding depression. The availability of high-density single nucleotide polymorphism (SNP) arrays has facilitated the quantification of F by genomic markers in farm animals. Runs of homozygosity (ROH) are contiguous lengths of homozygous genotypes and represent an estimate of the degree of autozygosity at genome-wide level. The current study aims to quantify the genomic F derived from ROH (F ROH) in three local dairy cattle breeds. F ROH values were compared with F estimated from the genomic relationship matrix (F GRM), based on the difference between observed v. expected number of homozygous genotypes (F HOM) and the genomic homozygosity of individual i (F MOL i ). The molecular coancestry coefficient (f MOL ij ) between individuals i and j was also estimated. Individuals of Cinisara (71), Modicana (72) and Reggiana (168) were genotyped with the 50K v2 Illumina BeadChip. Genotypes from 96 animals of Italian Holstein cattle breed were also included in the analysis. We used a definition of ROH as tracts of homozygous genotypes that were >4 Mb. Among breeds, 3661 ROH were identified. Modicana showed the highest mean number of ROH per individual and the highest value of F ROH, whereas Reggiana showed the lowest ones. Differences among breeds existed for the ROH lengths. The individuals of Italian Holstein showed high number of short ROH segments, related to ancient consanguinity. Similar results showed the Reggiana with some extreme animals with segments covering 400 Mb and more of genome. Modicana and Cinisara showed similar results between them with the total length of ROH characterized by the presence of large segments. High correlation was found between F HOM and F ROH ranged from 0.83 in Reggiana to 0.95 in Cinisara and Modicana. The correlations among F ROH and other estimated F coefficients were generally lower ranged from 0.45 (F MOL i -F ROH) in Cinisara to 0.17 (F GRM-F ROH) in Modicana. On the basis of our results, recent inbreeding was observed in local breeds, considering that 16 Mb segments are expected to present inbreeding up to three generations ago. Our results showed the necessity of implementing conservation programs to control the rise of inbreeding and coancestry in the three Italian local dairy cattle breeds.

[Knowledge level of hypertensive patients about hypertension. Relationship between knowledge level and hypertension control]. / Estudio Conocimiento: grado de conocimiento sobre hipertensión arterial de nuestros pacientes. Relación con el nivel de control de la misma.

OBJECTIVE: To assess the knowledge of the hypertensive patients about their hypertension and their relation to its control. MATERIAL AND METHODS: Cross-sectional study among 400 hypertensive patients, all over 18 years, selected from 50 primary-care centres, who responded to an hypertension-related survey. Included variables were survey items, age, gender, educational level, professional occupation, blood pressure data and antihypertensive treatment. The obtained differences were analyzed using the chi-square test, Kruskal-Wallis, Wilcoxon, Anova and Bonferroni methods. RESULTS: There were 323 valid surveys. 52.9% of respondents were women, the average age: 65.4 years (SD: 11.2), 54.8% of them had primary education. 39.6% were aware of the objectives of systolic BP control. Only 19.6% having knowledge of those for diastolic BP control, with no differences between controlled and uncontrolled (systolic BP: 39% vs 38.1%, P=.887; diastolic BP: 19.2% vs 21%, P=.721). Over 70% knew about lifestyle changes, without significant differences between controlled and uncontrolled respondents. 82% of controlled respondents, and 79% of those uncontrolled, recognized the chronical nature of the treatment (P=.548), but 15.1% of the controlled respondents and 12.4% of uncontrolled respondents did not see the relation between the treatment and hypertension control (P=.525). 31.1% believed to be well-controlled, but in fact was not. CONCLUSIONS: Our patients doesn't know blood pressure targets of control. There isn't relationship between this knowledge and control of hypertension.

Estudio Conocimiento: grado de conocimiento sobre hipertensión arterial de nuestros pacientes. Relación con el nivel de control de la misma / Knowledge level of hypertensive patients about hypertension. Relationship between knowledge level and hypertension control

Objetivo: Evaluar el conocimiento de nuestros pacientes hipertensos sobre su HTA y su relación con el control de la misma. Material y métodos: Estudio descriptivo transversal con 400 hipertensos, mayores de edad, seleccionados de forma sistemática consecutiva de 50 consultas de atención primaria, que respondieron una encuesta sobre HTA. Las variables recogidas fueron los ítems de la encuesta, edad, sexo, nivel educacional, ocupación, cifras de presión arterial y tratamiento antihipertensivo. Las diferencias se analizaron con los test ji-cuadrado, Kruskal-Wallis, Wilcoxon, Anova y Bonferroni según distribución normal. Resultados: Fueron válidas 323 encuestas. El 52,9% fueron mujeres, edad media de 65,4años (DE: 11,2). El 54,8% contaban con estudios primarios. El 39,6% conocían los objetivos de control de PA sistólica, y solo el 19,6% los de PA diastólica, sin diferencias entre controlados y no controlados (PA sistólica: 39% vs 38,1%, p = 0,887; PA diastólica: 19,2% vs 21%, p = 0,721). Más del 70% conocían las modificaciones del estilo de vida, sin diferencias entre controlados y no controlados. El 82% de los controlados y el 79% de los no controlados reconocieron la cronicidad del tratamiento (p = 0,548), pero el 15,1% de los controlados y el 12,4% de los no controlados no lo relacionaban con el control de la HTA (p = 0,525). El 31,1% creían estar bien controlados aunque no lo estaban. Conclusiones: Los pacientes hipertensos conocen en baja frecuencia cuáles son los objetivos de control, sin encontrar relación entre el conocimiento del problema y el control del mismo

Objective: To assess the knowledge of the hypertensive patients about their hypertension and their relation to its control. Material and methods: Cross-sectional study among 400 hypertensive patients, all over 18 years, selected from 50 primary-care centres, who responded to an hypertension-related survey. Included variables were survey items, age, gender, educational level, professional occupation, blood pressure data and antihypertensive treatment. The obtained differences were analyzed using the chi-square test, Kruskal-Wallis, Wilcoxon, Anova and Bonferroni methods. Results: There were 323 valid surveys. 52.9% of respondents were women, the average age: 65.4 years (SD: 11.2), 54.8% of them had primary education. 39.6% were aware of the objectives of systolic BP control. Only 19.6% having knowledge of those for diastolic BP control, with no differences between controlled and uncontrolled (systolic BP: 39% vs 38.1%, P = .887; diastolic BP: 19.2% vs 21%, P = .721). Over 70% knew about lifestyle changes, without significant differences between controlled and uncontrolled respondents. 82% of controlled respondents, and 79% of those uncontrolled, recognized the chronical nature of the treatment (P = .548), but 15.1% of the controlled respondents and 12.4% of uncontrolled respondents did not see the relation between the treatment and hypertension control (P = .525). 31.1% believed to be well-controlled, but in fact was not. Conclusions: Our patients doesn’t know blood pressure targets of control. There isn’t relationship between this knowledge and control of hypertension

The genome-wide structure of two economically important indigenous Sicilian cattle breeds.

Genomic technologies, such as high-throughput genotyping based on SNP arrays, provided background information concerning genome structure in domestic animals. The aim of this work was to investigate the genetic structure, the genome-wide estimates of inbreeding, coancestry, effective population size (Ne), and the patterns of linkage disequilibrium (LD) in 2 economically important Sicilian local cattle breeds, Cinisara (CIN) and Modicana (MOD), using the Illumina Bovine SNP50K v2 BeadChip. To understand the genetic relationship and to place both Sicilian breeds in a global context, genotypes from 134 other domesticated bovid breeds were used. Principal component analysis showed that the Sicilian cattle breeds were closer to individuals of Bos taurus taurus from Eurasia and formed nonoverlapping clusters with other breeds. Between the Sicilian cattle breeds, MOD was the most differentiated, whereas the animals belonging to the CIN breed showed a lower value of assignment, the presence of substructure, and genetic links with the MOD breed. The average molecular inbreeding and coancestry coefficients were moderately high, and the current estimates of Ne were low in both breeds. These values indicated a low genetic variability. Considering levels of LD between adjacent markers, the average r(2) in the MOD breed was comparable to those reported for others cattle breeds, whereas CIN showed a lower value. Therefore, these results support the need of more dense SNP arrays for a high-power association mapping and genomic selection efficiency, particularly for the CIN cattle breed. Controlling molecular inbreeding and coancestry would restrict inbreeding depression, the probability of losing beneficial rare alleles, and therefore the risk of extinction. The results generated from this study have important implications for the development of conservation and/or selection breeding programs in these 2 local cattle breeds.

Study of polymorphisms in the promoter region of ovine ß-lactoglobulin gene and phylogenetic analysis among the Valle del Belice breed and other sheep breeds considered as ancestors.

The aim of this work was to sequence the promoter region of ß-lactoglobulin (BLG) gene in four sheep breeds, in order to identify polymorphisms, infer and analyze haplotypes, and phylogenetic relationship among the Valle del Belice breed and the other three breeds considered as ancestors. Sequencing analysis and alignment of the obtained sequences showed the presence of 36 single nucleotide polymorphisms (SNPs) and one deletion. A total of 22 haplotypes found in "best" reconstruction were inferred considering the 37 polymorphic sites identified. Haplotypes were used for the reconstruction of a phylogenetic tree using the Neighbor-Joining algorithm. The number of polymorphisms identified showed high variability within breeds. Analysis of genetic diversity indexes showed that the Sarda breed presented the lowest nucleotide diversity, whereas the Comisana breed presented the highest one. Comparing the nucleotide diversity among breeds, the highest value was obtained between Valle del Belice and Pinzirita breeds, whereas the lowest one was between Valle del Belice and Sarda breeds. Considering that polymorphisms in the promoter region of BLG gene could have a functional role associated with milk composition, the lowest value of nucleotide diversity between Valle del Belice and Sarda breeds may be related to a higher similarity of milk composition of these two breeds compared to the others. Further analyses will be conducted in order to evaluate the possible correlation between the genetic diversity indexes and the BLG content in milk of our breeds.

Polymorphisms of beta-lactoglobulin promoter region in three Sicilian goat breeds.

Several beta-lactoglobulin (BLG) polymorphisms have been described within the proximal promoter region and coding region of the caprine gene, although no genetic variants affecting the protein amino acid composition and/or expression level have been characterized so far. Binding sites for several transcription factors (TFs) are present in the BLG promoter region. The aims of this work were to sequence the full-length promoter region of three Sicilian goat breeds in order to identify polymorphisms, analyze the identified haplotypes, search for differences between breeds for the presence of polymorphisms in this gene region, search for putative TFs binding sites, and check if polymorphisms lay within the identified TFs binding sites. The promoter region of BLG gene in Sicilian goat breeds showed high level of polymorphism due to the presence of 36 single nucleotide polymorphisms (SNPs). Association between polymorphic sites was computed within the whole sample analyzed and 18 haplotypes were inferred. Binding sites for three milk protein binding factors (MPBFs) and four nuclear factor-I (NF-I) were found within BLG promoter region based on the ovine sequence. The identification of some SNPs within TFs binding sites allowed hypothesizing the loss of TFs. Further studies are in progress to evaluate the effect of these mutations on binding affinity of TFs, the functional interaction of the TFs with the goat BLG promoter, and the relationship of the polymorphisms with BLG gene expression and milk production and composition.