Low-pass whole genome sequencing as a cost-effective alternative to chromosomal microarray analysis for low- and middle-income countries.

Low-pass whole genome sequencing (LP-WGS) has been applied as alternative method to detect copy number variants (CNVs) in the clinical setting. Compared with chromosomal microarray analysis (CMA), the sequencing-based approach provides a similar resolution of CNV detection at a lower cost. In this study, we assessed the efficiency and reliability of LP-WGS as a more affordable alternative to CMA. A total of 1363 patients with unexplained neurodevelopmental delay/intellectual disability, autism spectrum disorders, and/or multiple congenital anomalies were enrolled. Those patients were referred from 15 nonprofit organizations and university centers located in different states in Brazil. The analysis of LP-WGS at 1x coverage (>50kb) revealed a positive testing result in 22% of the cases (304/1363), in which 219 and 85 correspond to pathogenic/likely pathogenic (P/LP) CNVs and variants of uncertain significance (VUS), respectively. The 16% (219/1363) diagnostic yield observed in our cohort is comparable to the 15%-20% reported for CMA in the literature. The use of commercial software, as demonstrated in this study, simplifies the implementation of the test in clinical settings. Particularly for countries like Brazil, where the cost of CMA presents a substantial barrier to most of the population, LP-WGS emerges as a cost-effective alternative for investigating copy number changes in cytogenetics.

Catalytic properties of amylases produced by Cunninghamella echinulata and Rhizopus microsporus.

The present work aimed to characterize and compare the catalytic properties of amylases from Cunninghamella echinulata and Rhizopus microsporus. The highest production of amylase by C. echinulata, 234.94 U g-1 of dry substrate (or 23.49 U mL-1), was obtained using wheat bran as a substrate, with 50-55% initial moisture and kept at 28 °C for 48 h. The highest production of amylases by R. microsporus, 224.85 U g-1 of dry substrate (or 22.48 U mL-1), was obtained cultivating wheat bran with 65% initial moisture at 45 °C for 24 h. The optimal activity of the amylases was observed at pH 5.0 at 60 °C for C. echinulata enzymes and at pH 4.5 at 65 °C for R. microsporus. The amylases produced by C. echinulata were stable at pH 4.0-8.0, while the R. microsporus enzymes were stable at pH 4.0-10.0. The amylases produced by C. echinulata remained stable for 1 h at 50 °C and the R. microsporus amylases maintained catalytic activity for 1 h at 55 °C. The enzymatic extracts of both fungi hydrolyzed starches from different plant sources and showed potential for liquefaction of starch, however the amylolytic complex of C. echinulata exhibited greater saccharifying potential.

ß-glucosidase from thermophilic fungus Thermoascus crustaceus: production and industrial potential.

Microbial ß-glucosidases can be used in several industrial processes, including production of biofuels, functional foods, juices, and beverages. In the present work, production of ß-glucosidase by solid state cultivation of the fungus Thermoascus crustaceus in a low-cost cultivation medium (comprising agroindustrial residues) was evaluated. The highest production of ß-glucosidase, about 415.1 U/g substrate (or 41.51 U/mL), was obtained by cultivating the fungus in wheat bran with 70% humidity, during 96 h at 40°C. The enzymatic activity was optimum at pH 4.5 and 65°C. ß-Glucosidase maintained its catalytic activity when incubated at a pH range of 4.0-8.0 and temperature of 30-55°C. The enzyme was strongly inhibited by glucose; even when the substrate and glucose concentrations were equal, the inhibition was not reversed, suggesting a non-competitive inhibition. In the presence of up to 10% ethanol, ß-glucosidase maintained its catalytic activity. In addition to ß-glucosidase, the enzymatic extract showed activity of 36 U/g for endoglucanase, 256.2 U/g for xylanase, and 18.2 U/g for ß-xylosidase. The results allow to conclude that the fungus T. crustaceus has considerable potential for production of ß-glucosidase and xylanase when cultivated in agroindustrial residues, thereby reducing the cost of these biocatalysts.

Low-fat frankfurters from protein concentrates of tilapia viscera and mechanically separated tilapia meat.

In order to develop a healthy low-fat frankfurter-type sausage, different formulations were developed with tilapia viscera surimi (T1) and two with mechanically separated tilapia meat (MSTM) surimi (T2 and T3), all without pig lard addition. Due to technological problems observed for T1 sausage during cooking, it was not further investigated. The functionality of the other two formulations was evaluated based on proximate composition, pH, water activity, and texture. Finally, microbiological and sensory analyses based on acceptance tests were performed. Listeria monocytogenes and Salmonella spp. were found to be absent. T2 showed higher frequencies for the attributes color (90.0%) and overall acceptability (86.7%), while T3 showed higher frequencies for taste (86.7%) and texture (96.7%). The surimi concentration was reflected in the physical properties of the sausages. It was found that the addition of MSTM surimi to sausage favored greater cutting strength (3.9 N for T2 and 4.9 N for T3). Beyond the surimi utilization, the total replacement of pig lard by cassava starch and soybean protein had also contributed with the texture properties.