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BACKGROUND: Membrane proteins constitute up to 30% of the human proteome. These proteins have special properties because the transmembrane segments are embedded into lipid bilayer while extramembranous parts are in different environments. Membrane proteins have several functions and are involved in numerous diseases. A large number of prediction methods have been introduced to predict protein subcellular localization as well as the tolerance or pathogenicity of amino acid substitutions. RESULTS: We tested the performance of 22 tolerance predictors by collecting information on membrane proteins and variants in them. The analysis indicated that the best tools had similar prediction performance on transmembrane, inside and outside regions of transmembrane proteins and comparable to overall prediction performances for all types of proteins. PON-P2 had the highest performance followed by REVEL, MetaSVM and VEST3. Further, we tested with the high quality dataset also the performance of seven subcellular localization predictors on membrane proteins. We assessed separately the performance for single pass and multi pass membrane proteins. Predictions for multi pass proteins were more reliable than those for single pass proteins. CONCLUSIONS: The predictors for variant effects had better performance than subcellular localization tools. The best tolerance predictors are highly reliable. As there are large differences in the performances of tools, end-users have to be cautious in method selection.
Assuntos
Biologia Computacional/métodos , Variação Genética , Espaço Intracelular/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Substituição de Aminoácidos , Benchmarking , Transporte ProteicoRESUMO
Background: The upper airways of cystic fibrosis (CF) persons are an evolutionary niche where genetically adapted bacterial strains are selected for lung infection. The microbiological studies conducted up to now on the upper airways are not easily comparable. Methods: Using classical culture methods, we simultaneously studied the microbiological status of upper and lower airways in persons not chronically infected with P. aeruginosa. Each person had a single upper airways sampling and a concomitant lower airways sampling. Lower airways sampling was performed by oropharyngeal swab or sputum collection. Using a quasi-experimental design of study, we evaluated the performance of 2 different upper airways' sampling methods, nasal lavage according to method described by Mainz or nasal lavage with a rhino-set. Pain was measured with appropriate scales. Results: A total of 194 persons were enrolled in this study. Pathogenic flora was found in 128 (6.6%) of 194 upper airways samples and in 164 (84.6%) lower airways samples. A statistically significant difference between the upper airways and the lower airways was found in the isolation of S. aureus and non-fermenter gram negatives. Nasal lavage according to Mainz resulted in the isolation of more non-fermenter gramnegatives than the rhino-set (p < 0.05). No differences were found in the pain caused bythe two methods. Conclusions: In our study population, cultures of the upper airway and lower airway differ in CF persons. In people sampled with nasal lavage according to Mainz more non-fermenter gram negatives were detected than with rhino-set. The two sampling methods were comparable with regard to the caused pain, nasal lavage according to Mainz method being quicker to perform.
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The sweat test (ST) is the current diagnostic gold standard for cystic fibrosis (CF). Many CF centres have switched from the Gibson-Cooke method to the Macroduct system-based method. We used these methods simultaneously to compare CF screening outcomes. STs using both methods were performed simultaneously between March and December 2022 at CF Centre in Florence. We included newborns who underwent newborn bloodspot screening (NBS), newborns undergoing transfusion immediately after birth, and children with CF screen-positive, inconclusive diagnosis (CFSPID). We assessed 72 subjects (median age 4.4 months; range 0-76.7): 30 (41.7%) NBS-positive, 18 (25.0%) newborns who underwent transfusion, and 24 (33.3%) children with CFSPID. No significant differences were found between valid sample numbers, by patient ages and groups (p = 0.10) and between chloride concentrations (p = 0.13), except for sweat chloride (SC) measured by the Gibson-Cooke and Macroduct methods in CFSPID group (29.0, IQR: 20.0-48.0 and 22.5, IQR: 15.5-30.8, respectively; p = 0.01). The Macroduct and Gibson-Cooke methods showed substantial agreement with the SC values, except for CFSPID, whose result may depend on the method of sweat collection. In case of invalid values with Macroduct, the test should be repeated with Gibson-Cooke method.
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BACKGROUND: Improved therapy in CF has led to an overall improvement in nutritional status. The objectives of our study are: to cross-sectionally assess nutritional status and serum levels of fat-soluble vitamins; to retrospectively evaluate the efficacy of modulators on nutritional status and fat-soluble vitamin levels. METHODS: In patients younger than 2 years of age, we evaluated growth, in patients aged 2-18 years, we assessed BMI z-scores, and in adults, we assessed absolute BMI values. Levels of 25(OH)D, vitamins A, and E were measured. RESULTS: A cross-sectional analysis was conducted on 318 patients, 109 (34.3%) with pancreatic sufficiency. Only three patients were under 2 years old. In 135 patients aged 2-18 years, the median BMI z-score was 0.11, and 5 (3.7%) patients had malnutrition (z-score ≤ 2SD). In 180 adults, the median BMI was 21.8 kg/m2. Overall, 15 (13.7%) males (M) and 18 (25.3%) females (F) were underweight (18 < BMI > 20); 3 (2.7%) M and 5 (7.0%) F had a BMI < 18. Suboptimal 25(OH)D levels were found in patients with pancreatic insufficiency. The prevalence of deficiency of vitamins A and E is low. After one year of treatment with modulators, the increase in BMI was more consistent (M: 1.58 ± 1.25 kg/m2 F: 1.77 ± 1.21 kg/m2) in elexacaftor/tezacaftor/ivacaftor (ETI)-treated patients compared with other modulators, with a significant increase in levels of all fat-soluble vitamins. CONCLUSIONS: Malnutrition is present in a limited number of subjects. The prevalence of subjects with suboptimal 25(OH)D levels is high. ETI showed a beneficial effect on nutritional status and circulating levels of fat-soluble vitamins.
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Pancreatitis-Associated Protein (PAP)-based Cystic Fibrosis (CF) newborn bloodspot screening (NBS) protocols detect less CFTR-Related Metabolic Syndrome (CRMS)/CF Screen Positive, Inconclusive Diagnosis (CFSPID). We prospectively evaluated the impact of PAP as the second step of the CF NBS protocol, before the CFTR genetic analysis, on NBS outcomes and CRMS/CFSPID detection in the Tuscany region, Italy. In parallel to the usual protocol (IRT/DNA, protocol 1), PAP was analyzed in IRT-positive infants (IRT/PAP/DNA, protocol 2) from 1 June 2020 until 31 May 2022. We defined an infant as NBS positive if PAP was >1.8 µg/L for IRT value 99th percentile-100 µg/L or >0.6 µg/L for IRT value >100 µg/L. To increase the positive predictive value (PPV) of protocol 2, we retrospectively lowered the upper IRT range value from 100 to 90 µg/L (modified protocol 2). We identified 8 CF and 13 CRMS/CFSPID with protocol 1, 5 CF and 5 CRMS/CFSPID with protocol 2 and 8 CF and 5 CRMS/CFSPID with modified protocol 2. With the PAP-based protocols, we observed a reduction of sweat tests, healthy carrier detection and a significant increase in PPV to 15.38%. Further data are needed in order to evaluate the outcomes of CRMS/CFSPID after a long follow-up.