Abstract Double strand break (DSB) repair primarily occurs through 3 pathways: non-homologous end-joining (NHEJ), alternative end-joining (Alt-EJ), and homologous recombination (HR). Typical methods to measure pathway usage include integrated cassette reporter assays or visualization of DNA damage induced nuclear foci. It is now well understood that repair of Cas9-induced breaks also involves NHEJ, Alt-EJ, and HR pathways, providing a new format to measure pathway usage. Here, we have developed a simple Cas9-based system with validated repair outcomes that accurately represent each pathway and then converted it to a droplet digital PCR (ddPCR) readout, thus obviating the need for Next Generation Sequencing and bioinformatic analysis with the goal to make Cas9-based system accessible to more laboratories. The assay system has reproduced several important insights. First, absence of the key Alt-EJ factor Pol θ only abrogates ∼50% of total Alt-EJ. Second, single-strand templated repair (SSTR) requires BRCA1 and MRE11 activity, but not BRCA2, establishing that SSTR commonly used in genome editing is not conventional HR. Third, BRCA1 promotes Alt-EJ usage at two-ended DSBs in contrast to BRCA2. This assay can be used in any system, which permits Cas9 delivery and, importantly, allows rapid genotype-to-phenotype correlation in isogenic cell line pairs.

ABSTRACT Based on available literature, practices, and agricultural movements, this study analyzed the conceptualization of agroecology in the West Balkans (Slovenia, Croatia, Bosnia and Herzegovina, Montenegro, Serbia, and North Macedonia). For decades, like Yugoslavia, they shared common agricultural development, and agroecology was viewed on a theoretical basis alone. Currently, cooperation exists between agroecology (scientific discipline) and organic agriculture (practical platform). All countries have committed to transition toward sustainable agriculture; however, lower productivity and economical/structural impediments have resulted in variable acceptance of agroecology. Nevertheless, agroecology is recognized as an alternative approach to conventional agriculture, with the potential to meet future food demands.

Executive functions (EF) play a crucial role in overall human functioning. Children with Autism Spectrum Disorder (ASD) often have EF deficits. The goal of this study was to examine EF and developmental domains in preschool children with ASD. The sample for this study comprised 32 children (27 boys, mean age 65.3 months, SD- 4.0 months) with ASD. The control group consisted of 32 typically developing children (16 boys, mean age 64.3 months, SD- 5.1 months). EF were assessed with Behavior Rating Inventory of Executive Function and developmental domains were assessed with the Developmental Assessment of Young Children. The results of this study indicated that children with ASD do not have uneven EF and developmental profiles, although the EF and developmental domains scores were more heterogeneous than in typically developing children. Children with ASD had substantially lower EF and developmental scores than typically developing children. Implications of these results are discussed.

Background: A precise three-point shot (3S) is considered a key parameter of success in a basketball game, and therefore the factors that affect its success have always attracted the attention of researchers. Aim: The aim of this research was a biomechanical-mathematical analysis of 3S in basketball, in order to determine the key parameters for performing a 3S. Results:  The research shows a model of shooting a basketball player from the central position of the shot with 6.75 m. The modeling led to the conclusion that the height of the throw, the speed and the angle of the throw of the ball have a positive and direct relationship with the angle at which the ball falls into the basket when it comes to a shot for three points. Conclusion:  The height of the throw, the speed and the angle of the ball have a positive and direct relationship with the angle at which the ball hits the basket when it comes to a shot for three points. Anthropometric characteristics of the player, such as the length of the arm, and the height of the player, directly lead to a positive relationship with the throwing angle.

One of the most important public- health tasks is to provide hygienic and healthy food and water, which will not have harmful consequences for the health of consumers, and in this way a safe food trade is ensured, therefore protecting everyone in the chain from the producer to the consumer. This paper presents the concentrations of heavy metals Fe, Mn, Zn and Cu, obtained by atomic absorption spectrophotometry (AAS), from 5 samples of thyme infusion (lat. Thymus serpyllum), which can be found on the market of Travnik. Sample infusions were prepared by holding the filter bags in boiling deionized water for 10 minutes. Mean values of concentrations of these heavy metals were for Fe 0.0508 ± 0.0139 mg/kg, Mn 0.06 ± 0.0069 mg/kg, Zn 0.0436 ± 0.0147 mg/kg, while the Cu content was not detected in any infusion of the said tea. The risk assessment of the intake of tested heavy metals for the health of consumers, expressed through the Hazard Index (HI), which was <1, shows that the samples of analyzed teas are safe for consumption.

This paper examines possibilities for improving the existing strategies of consistency management for highly-distributed transactional database in a hybrid cloud environment. With a detailed analysis of the existing consistency models for distributed database and standard strategies including Classic, Quorum and Tree Based Consistency (TBC), it is concluded that an improved advanced model of so-called visible adaptive consistency needs to be applied in a highly-distributed cloud environment, as necessary and sufficient degree of synchronization of all replicas. Along with the proposed model, research and development of an advanced novel strategy for consistency management Rose TBC (R-TBC) approach has been conducted, by improving standard TBC approach. Regarding implementation, a specific agglomerative Rose Tree Algorithm (RTA) has been developed, based on Bayesian hierarchical clustering and Graph Partitioning Algorithm - Multidimensional Data Clustering (GPA-MDC) intelligent partitioning of transactional Cloud Database Management System (CDBMS). The final result is constructed R-TBC model that changes in accordance with dynamic changes of entire heterogeneous CDBMS environment.

Microneedles (MNs) have been manufactured using a variety of methods from a range of materials, but most of them are expensive and time-consuming for screening new designs and making any modifications. Therefore, stereolithography (SLA) has emerged as a promising approach for MN fabrication due to its numerous advantages, including simplicity, low cost, and the ability to manufacture complex geometrical products at any time, including modifications to the original designs. This work aimed to print MNs using SLA technology and investigate the effects of post-printing curing conditions on the mechanical properties of 3D-printed MNs. Solid MNs were designed using CAD software and printed with grey resin (Formlabs, UK) using Form 3 printer (Formlabs, UK). MNs dimensions were 1.2 × 0.4 × 0.05 mm, arranged in 6 rows and 6 columns on a 10 × 10 mm baseplate. MNs were then immersed in an isopropyl alcohol bath to remove unpolymerized resin residues and cured in a UV-A heated chamber (Formlabs, UK). In total, nine samples were taken for each combination of curing temperature (35°C, 50°C, and 70°C) and curing time (5 min, 20 min, and 60 min). Fracture tests were conducted using a hardness apparatus TB24 (Erweka, Germany). MNs were placed on the moving probe of the machine and compressed until fracture. The optimization of the SLA process parameters for improving the strength of MNs was performed using the Taguchi method. The design of experiments was carried out based on the Taguchi L9 orthogonal array. Experimental results showed that the curing temperature has a significant influence on MN strength improvements. Improvement of the MN strength can be achieved by increasing the curing temperature and curing time.

Three fractions of tobacco waste (scrap, dust and midrib) were subjected to a high voltage electric discharge (HVED) assisted extraction procedure under different experimental conditions: solvent:solid ratio (300, 500, 700 mL/g), frequency (40, 70, 100 Hz) and treatment time (15, 30, 45 min), in order to study the influence of these conditions on the content of chlorogenic acid. The content of chlorogenic acid ranged from 1.54 to 3.66 mg/100 g for scrap, from 1.90 to 2.97 mg/100 g for dust, and from 2.30 to 3.38 mg/100 g for midrib extract, showing a strong dependence on the applied process parameters. The temperature change and the change in pH and electrical conductivity of the extracts after high voltage discharge treatment were also observed. The studied process parameters showed a statistically significant effect on the chemical and physical properties of the extracts from tobacco waste as well as on the content of chlorogenic acid, indicating the potential of HVED assisted processes in the separation of chlorogenic acid from tobacco industry waste. Multiple regression analysis was used to fit the results for the chlorogenic acid to a second order polynomial equation and the optimum conditions were determined.

Background: Diesel engine exhaust causes adverse health effects. Meanwhile, the impact of renewable diesel exhaust on human health is less known. In this study, nasal patency, pulmonary function, and self-rated symptoms were assessed in 19 healthy volunteers after two separate 3-hour exposures to renewable diesel (hydrotreated vegetable oil [HVO]) exhaust, and exposure to filtered air (FA) for comparison. The HVO exposures were generated with two modern non-road vehicles (2019) having either: 1) no aftertreatment system (HVOPM+NOx), or 2) an aftertreatment system containing a diesel oxidation catalyst and a diesel particulate filter (HVONOx). The exposure concentrations complied with current EU occupational exposure limits (OELs) of NO, NO2, formaldehyde, polycyclic aromatic hydrocarbons (PAHs), and future OELs of elemental carbon (EC) from 2023. Results: Exposure to HVOPM+NOx consisted of PM1 (≈90 µg m-3, 54 µg m-3 EC) and NOx (NO 3.4 ppm, NO2 0.6 ppm). The average total respiratory tract deposition of PM1 was 27 µg h-1. The deposition fraction of HVO PM1 was 40-50% higher compared to diesel exhaust PM1 from an older vehicle, due to smaller particle sizes of the HVOPM+NOx exhaust. Exposure to HVONOx consisted mainly of NOx (NO 2.0 ppm, NO2 0.7 ppm) with low level of PM1 (~1 µg m-3). Compared to filtered air, exposure to HVOPM+NOx and HVONOx caused higher incidence of self-reported symptoms (78%, 63%, respectively, vs. 28% for FA, p<0.03). Especially, exposure to HVOPM+NOx showed 40-50% higher eye and throat irritation symptoms. Compared to filtered air, a decrement in nasal patency was found for the HVONOx exposures (-18.1, 95%CI: -27.3 to -8.8 L min-1), and for the HVOPM+NOx (-7.4 (-15.6 to 0.8) L min-1). Overall, no change was indicated in the pulmonary function tests (spirometry, peak expiratory flow, forced oscillation technique), except a slight increase in FEV1/FVC after exposure to HVONOx.Conclusion: Short-term exposure to HVO exhaust below the EU OELs did not cause severe pulmonary function changes in healthy subjects. However, an increase in self-rated mild irritation symptoms, and mild decrease in nasal patency after two HVO exposures may indicate irritative effects from exposure to HVO exhaust from modern non-road vehicles below future OELs.