KASP: a genotyping method to rapid identification of resistance in Plasmodium falciparum.

BACKGROUND: The emergence and spread of anti-malarial resistance continues to hinder malaria control. Plasmodium falciparum, the species that causes most human malaria cases and most deaths, has shown resistance to almost all known anti-malarials. This anti-malarial resistance arises from the development and subsequent expansion of Single Nucleotide Polymorphisms (SNPs) in specific parasite genes. A quick and cheap tool for the detection of drug resistance can be crucial and very useful for use in hospitals and in malaria control programmes. It has been demonstrated in different contexts that genotyping by Kompetitive Allele Specific PCR (KASP), is a simple, fast and economical method that allows a high-precision biallelic characterization of SNPs, hence its possible utility in the study of resistance in P. falciparum. METHODS: Three SNPs involved in most cases of resistance to the most widespread anti-malarial treatments have been analysed by PCR plus sequencing and by KASP (C580Y of the Kelch13 gene, Y86N of the Pfmdr1 gene and M133I of the Pfcytb gene). A total of 113 P. falciparum positive samples and 24 negative samples, previously analysed by PCR and sequencing, were selected for this assay. Likewise, the samples were genotyped for the MSP-1 and MSP-2 genes, and the Multiplicity of Infection (MOI) and parasitaemia were measured to observe their possible influence on the KASP method. RESULTS: The KASP results showed the same expected mutations and wild type genotypes as the reference method, with few exceptions that correlated with very low parasitaemia samples. In addition, two cases of heterozygotes that had not been detected by sequencing were found. No correlation was found between the MOI or parasitaemia and the KASP values of the sample. The reproducibility of the technique shows no oscillations between repetitions in any of the three SNPs analysed. CONCLUSIONS: The KASP assays developed in this study were efficient and versatile for the determination of the Plasmodium genotypes related to resistance. The method is simple, fast, reproducible with low cost in personnel, material and equipment and scalable, being able to core KASP arrays, including numerous SNPs, to complete the main pattern of mutations associated to P. falciparum resistance.

A Data-Driven Approach to Physical Fatigue Management Using Wearable Sensors to Classify Four Diagnostic Fatigue States.

Physical exercise contributes to the success of rehabilitation programs and rehabilitation processes assisted through social robots. However, the amount and intensity of exercise needed to obtain positive results are unknown. Several considerations must be kept in mind for its implementation in rehabilitation, as monitoring of patients' intensity, which is essential to avoid extreme fatigue conditions, may cause physical and physiological complications. The use of machine learning models has been implemented in fatigue management, but is limited in practice due to the lack of understanding of how an individual's performance deteriorates with fatigue; this can vary based on physical exercise, environment, and the individual's characteristics. As a first step, this paper lays the foundation for a data analytic approach to managing fatigue in walking tasks. The proposed framework establishes the criteria for a feature and machine learning algorithm selection for fatigue management, classifying four fatigue diagnoses states. Based on the proposed framework and the classifier implemented, the random forest model presented the best performance with an average accuracy of ≥98% and F-score of ≥93%. This model was comprised of ≤16 features. In addition, the prediction performance was analyzed by limiting the sensors used from four IMUs to two or even one IMU with an overall performance of ≥88%.

Gait Phase Detection for Lower-Limb Exoskeletons using Foot Motion Data from a Single Inertial Measurement Unit in Hemiparetic Individuals.

Due to the recent rise in the use of lower-limb exoskeletons as an alternative for gait rehabilitation, gait phase detection has become an increasingly important feature in the control of these devices. In addition, highly functional, low-cost recovery devices are needed in developing countries, since limited budgets are allocated specifically for biomedical advances. To achieve this goal, this paper presents two gait phase partitioning algorithms that use motion data from a single inertial measurement unit (IMU) placed on the foot instep. For these data, sagittal angular velocity and linear acceleration signals were extracted from nine healthy subjects and nine pathological subjects. Pressure patterns from force sensitive resistors (FSR) instrumented on a custom insole were used as reference values. The performance of a threshold-based (TB) algorithm and a hidden Markov model (HMM) based algorithm, trained by means of subject-specific and standardized parameters approaches, were compared during treadmill walking tasks in terms of timing errors and the goodness index. The findings indicate that HMM outperforms TB for this hardware configuration. In addition, the HMM-based classifier trained by an intra-subject approach showed excellent reliability for the evaluation of mean time, i.e., its intra-class correlation coefficient (ICC) was greater than 0 . 75 . In conclusion, the HMM-based method proposed here can be implemented for gait phase recognition, such as to evaluate gait variability in patients and to control robotic orthoses for lower-limb rehabilitation.

The retinal locus of fixation in simulations of progressing central scotomas.

Patients with central scotoma use a preferred retinal locus (PRL) of fixation to perform visual tasks. Some of the conditions that cause central scotoma are progressive, and as a consequence, the PRL needs to be adjusted throughout the progression. The present study investigates the peripheral locus of fixation in subjects under a simulation of progressive central scotoma. Five normally sighted subjects participated in the study. A foveally centered mask of varying size was presented to simulate the scotoma. Initially, subjects developed a peripheral locus of fixation under simulation of a 6° scotoma, which was used as a baseline. The progression was simulated in two separate conditions: a gradual progression and an abrupt progression. In the gradual progression, the diameter of the scotoma increased by a fixed amount of either 1° or 2° of visual angle, thus scotomas of 8°, 10°, and 11° of visual angle were simulated. In the abrupt progression, the diameter was adjusted individually to span the area of the visual field used by the current peripheral locus of fixation. Subjects located the peripheral locus of fixation along the same meridian under simulation of scotoma progression. Furthermore, no differences between the fixation stability of the baseline locus of fixation and the incremental progression locus of fixation were found whereas, in abrupt progression, the fixation stability decreased significantly. These results provide first insight into fixation behavior in a progressive scotoma and may contribute to the development of training tools for patients with progressive central maculopathies.

A preferred retinal location of fixation can be induced when systematic stimulus relocations are applied.

Patients with central vision loss obtain visual information by fixating on an object eccentrically with a preferred retinal locus of fixation (PRL). Patients do not always choose the most efficient PRL position, and as a consequence, visual performance is not always fully exploited. This study investigates whether PRLs can be induced by applying systematic stimulus relocations. The PRL was trained using a central scotoma simulation in 15 healthy subjects. They performed different visual tasks during four sessions, after which their reading performance was evaluated. In five subjects the stimulus was relocated to the left hemifield whenever a saccade would place the stimulus on the opposite hemifield. In five different subjects the relocation was inversed: The stimulus was located in the right hemifield. The relocation was 7.5° of visual angle and it was applied horizontally. Five additional subjects naturally chose the PRL location. They were used as the control group to evaluate the development of a PRL. After training, subjects performed visual search tasks on static stimuli. Evaluation after training showed that systematic stimulus relocations can be used to influence the development of the PRL. These results might be significant for the development of training strategies for the visually impaired.

Transfer of an induced preferred retinal locus of fixation to everyday life visual tasks.

Subjects develop a preferred retinal locus of fixation (PRL) under simulation of central scotoma. If systematic relocations are applied to the stimulus position, PRLs manifest at a location in favor of the stimulus relocation. The present study investigates whether the induced PRL is transferred to important visual tasks in daily life, namely pursuit eye movements, signage reading, and text reading. Fifteen subjects with normal sight participated in the study. To develop a PRL, all subjects underwent a scotoma simulation in a prior study, where five subjects were trained to develop the PRL in the left hemifield, five different subjects on the right hemifield, and the remaining five subjects could naturally chose the PRL location. The position of this PRL was used as baseline. Under central scotoma simulation, subjects performed a pursuit task, a signage reading task, and a reading-text task. In addition, retention of the behavior was also studied. Results showed that the PRL position was transferred to the pursuit task and that the vertical location of the PRL was maintained on the text reading task. However, when reading signage, a function-driven change in PRL location was observed. In addition, retention of the PRL position was observed over weeks and months. These results indicate that PRL positions can be induced and may further transferred to everyday life visual tasks, without hindering function-driven changes in PRL position.

Resembling breast milk: influence of polyamine-supplemented formula on neonatal BALB/cOlaHsd mouse microbiota.

Infant microbiota is influenced by numerous factors, such as delivery mode, environment, prematurity and diet (breast milk or formula). In addition to its nutritional value, breast milk contains bioactive substances that drive microbial colonisation and support immune system development, which are usually not present in infant formulas. Among these substances, polyamines have been described to be essential for intestinal and immune functions in newborns. However, their effect on the establishment of microbiota remains unclear. Therefore, the aim of the present study was to ascertain whether an infant formula supplemented with polyamines has an impact on microbial colonisation by modifying it to resemble that in breast-fed neonatal BALB/c mice. In a 4 d intervention, a total of sixty pups (14 d old) were randomly assigned to the following groups: (1) breast-fed group; (2) non-enriched infant formula-fed group; (3) three different groups fed an infant formula enriched with increasing concentrations of polyamines (mixture of putrescine, spermidine and spermine), following the proportions found in human milk. Microbial composition in the contents of the oral cavity, stomach and small and large intestines was analysed by quantitative PCR targeted at fourteen bacterial genera and species. Significantly different (P< 0·05) microbial colonisation patterns were observed in the entire gastrointestinal tract of the breast-fed and formula-fed mice. In addition, our findings demonstrate that supplementation of polyamines regulates the amounts of total bacteria, Akkermansia muciniphila, Lactobacillus, Bifidobacterium, Bacteroides-Prevotella and Clostridium groups to levels found in the breast-fed group. Such an effect requires further investigation in human infants, as supplementation of an infant formula with polyamines might contribute to healthy gastrointestinal tract development.

Reversible insertion of carbenes into ruthenium-silicon bonds.

The five-coordinate carbene complexes [Ru{κP,P,Si-Si(Me)(C6H4-2-PiPr2)2}Cl(═CHR)] (2, R = Ph; 3, R = SiMe3), analogues of the Grubbs catalyst, were prepared from the dimer [Ru(µ-Cl){κP,P,Si-Si(Me)(C6H4-2-PiPr2)2}]2 (1) and the corresponding diazoalkane N2CHR. The particular structural features that result from the presence of a strongly trans directing silyl group at the pincer ligand of these complexes are discussed on the basis of NMR information and the crystal structure of the vinylidene analogue [Ru{κP,P,Si-Si(Me)(C6H4-2-PiPr2)2}Cl(═C═CHPh)] (4), which was also obtained from 1 and phenylacetylene. The reactions of 3 with reagents such as P(OMe)3, CO, NCMe, and K(acac) illustrate that the first response of these carbene complexes to an increase of the coordination number around ruthenium is the insertion of the carbene ligand into the Ru-Si bond. These reactions also indicate that the insertion process is reversible and allows typical transformations of carbene ligands such as C-H functionalizations via carbene insertion (in the acac ligand) or the formation of ketene from CO. In addition, the reactions of 3 with terminal alkynes such as phenylacetylene or 3,3-dimethyl-1-butyne show that the inserted carbenes can also undergo reactions typical of metal-bound alkyls such as alkyne insertion and C-H reductive elimination.

Stability of phenolic compounds in apple and strawberry: Effect of different processing techniques in industrial set up.

Different technologies commonly employed by the fruit processing industry affect the phenolic content. This study aimed to establish the extent to which different processing techniques and conditions, tested at an industrial scale, affect phenolics, color, and sensory attributes of fresh strawberries and apples. The effects of freezing, thermal treatments, and high-pressure processing were investigated. In strawberries, mild and standard thermal treatments showed similar patterns for most phenolic groups; an increase in proanthocyanidins, no change in ellagic acid conjugates, and a major decrease in flavonols and anthocyanins. In apples, mild treatments and high-pressure processing had similar effects in all phenolic groups, with increases in dihydrochalcones, hydroxycinnamics, and proanthocyanidins and decreases in flavonols. However, the standard thermal treatment increased flavonols and dihydrochalcones concentrations. This study shows that each fruit behaves differently, and both technology and processing conditions should be customized accordingly to preserve or even increase the phenolic content.

Effect of Storage Conditions on the Stability of Polyphenols of Apple and Strawberry Purees Produced at Industrial Scale by Different Processing Techniques.

During a food product's life, storage conditions affect its composition of nutrients, bioactive compounds, and sensory attributes. In this research, strawberry and apple purees were selected as a model to examine how the storage of various purees industrially produced with different technologies affect the bioactive phenolic compounds, color, and sensory attributes. Specifically, fruit products processed on an industrial scale by different technologies including freezing, thermal treatment (mild and standard), and high-pressure processing were studied, as well as storage for up to 12 months at -20, 4, and 24 °C. In strawberry puree, storage conditions had a stronger impact on phenolic compound levels, particularly on anthocyanins, whereas in apple puree, the initial processing techniques exerted a greater influence than storage conditions, mainly caused by the hot or cold crushing processes. In general, proanthocyanidins were the major phenolic group and the most stable during storage, while anthocyanins were the group most affected by both processing and storage. Apple flavonols and dihydrochalcones were quite stable, while strawberry ellagitannins suffered higher degradations during storage. Through our analysis, it is found that during storage, the stability of polyphenols in each fruit is different, and processing and storage can be either detrimental or even beneficial. The selection of the ideal storage conditions (time and temperature) is a key factor to maintaining the polyphenol content in sensitive fruits such as strawberries. However, storage conditions are in some cases more important to minimizing the polyphenol losses than how the product is processed.

Prediction of refractive error and its progression: a machine learning-based algorithm.

OBJECTIVE: Myopia is the refractive error that shows the highest prevalence for younger ages in Southeast Asia and its projection over the next decades indicates that this situation will worsen. Nowadays, several management solutions are being applied to help fight its onset and development, nonetheless, the applications of these techniques depend on a clear and reliable assessment of risk to develop myopia. METHODS AND ANALYSIS: In this study, population-based data of Chinese children were used to develop a machine learning-based algorithm that enables the risk assessment of myopia's onset and development. Cross-sectional data of 12 780 kids together with longitudinal data of 226 kids containing age, gender, biometry and refractive parameters were used for the development of the models. RESULTS: A combination of support vector regression and Gaussian process regression resulted in the best performing algorithm. The Pearson correlation coefficient between prediction and measured data was 0.77, whereas the bias was -0.05 D and the limits of agreement was 0.85 D (95% CI: -0.91 to 0.80D). DISCUSSION: The developed algorithm uses accessible inputs to provide an estimate of refractive development and may serve as guide for the eye care professional to help determine the individual best strategy for management of myopia.

Do different robot appearances change emotion recognition in children with ASD?

Introduction: Socially Assistive Robotics has emerged as a potential tool for rehabilitating cognitive and developmental disorders in children with autism. Social robots found in the literature are often able to teach critical social skills, such as emotion recognition and physical interaction. Even though there are promising results in clinical studies, there is a lack of guidelines on selecting the appropriate robot and how to design and implement the child-robot interaction. Methods: This work aims to evaluate the impacts of a social robot designed with three different appearances according to the results of a participatory design (PD) process with the community. A validation study in the emotion recognition task was carried out with 21 children with autism. Results: Spectrum disorder results showed that robot-like appearances reached a higher percentage of children's attention and that participants performed better when recognizing simple emotions, such as happiness and sadness. Discussion: This study offers empirical support for continuing research on using SAR to promote social interaction with children with ASD. Further long-term research will help to identify the differences between high and low-functioning children.

Impact of fungal and plant metabolites application on early development stages of pea powdery mildew.

BACKGROUND: Pea powdery mildew incited by Erysiphe pisi represents a major constraint for pea crops worldwide. Crop protection is largely based on chemical control, although recently a renewed interest in the discovery of natural products as alternatives to synthetic fungicides application has emerged. Thus, 12 bioactive plant and fungal metabolites belonging to different class of natural compounds were evaluated, together with a commercial fungicide, at different concentrations on detached pea leaves for their potential to inhibit spore germination and subsequent stages of fungal growth. The most effective metabolites were tested at different concentrations in planta under controlled conditions to evaluate the level of control achieved by treatments before, concurrently and after pathogen inoculation. Pathogen development was macroscopically scored on whole plants as percentage of disease severity and area under the disease progress curve. RESULTS: Cavoxin, inuloxin C and sphaeropsidin A strongly inhibited E. pisi germination and haustoria formation and reduced colony size. This effect was dose dependent. These results were further confirmed in whole plants by spraying the metabolites on plant leaves for preventive or curative control, which reduced fungal developmental of E. pisi at levels comparable with those obtained by application of the fungicide. CONCLUSIONS: Cavoxin, inuloxin C and sphaeropsidin A have potential as alternatives to synthetic fungicides for the control of crop pathogens of economic importance such as powdery mildew. © 2019 Society of Chemical Industry.

Can positions in the visual field with high attentional capabilities be good candidates for a new preferred retinal locus?

The sustained component of visual attention lowers the perceptual threshold of stimuli located at the attended region. Attentional performance is not equal for all eccentric positions, leading to variations in perception. The location of the preferred retinal locus (PRL) for fixation might be influenced by these attentional variations. This study investigated the relation between the placement of sustained attention and the location of a developed PRL using simulations of central scotoma. Thirteen normally sighted subjects participated in the study. Monocular sustained attention was measured in discrete eccentric locations of the visual field using the dominant eye. Subsequently, a six degrees macular scotoma was simulated and PRL training was performed during eight ten-minutes blocks of trials. After training, every subject developed a PRL. Subjects with high attentional capabilities in the lower hemifield generally developed PRLs in the lower hemifield (n=10), subjects with high attentional capabilities in the upper hemifield developed PRLs in the upper hemifield (n=2) and one subject with similar attentional capabilities in the upper and lower hemifield developed the PRL on the upper hemifield. Analyzed individually, the results showed that 70% of the subjects had a PRL location in the hemifield where high attentional performance was achieved. These results suggest that attentional capabilities can be used as a predictor for the development of the PRL and are of significance for low vision rehabilitation and for the development of new PRL training procedures, with the option for a preventive attentional training in early macular disease to develop a favorable PRL.

Cell-free culture supernatant of Bifidobacterium breve CNCM I-4035 decreases pro-inflammatory cytokines in human dendritic cells challenged with Salmonella typhi through TLR activation.

Dendritic cells (DCs) constitute the first point of contact between gut commensals and our immune system. Despite growing evidence of the immunomodulatory effects of probiotics, the interactions between the cells of the intestinal immune system and bacteria remain largely unknown. Indeed,, the aim of this work was to determine whether the probiotic Bifidobacterium breve CNCM I-4035 and its cell-free culture supernatant (CFS) have immunomodulatory effects in human intestinal-like dendritic cells (DCs) and how they respond to the pathogenic bacterium Salmonella enterica serovar Typhi, and also to elucidate the molecular mechanisms involved in these interactions. Human DCs were directly challenged with B. breve/CFS, S. typhi or a combination of these stimuli for 4 h. The expression pattern of genes involved in Toll-like receptor (TLR) signaling pathway and cytokine secretion was analyzed. CFS decreased pro-inflammatory cytokines and chemokines in human intestinal DCs challenged with S. typhi. In contrast, the B. breve CNCM I-4035 probiotic strain was a potent inducer of the pro-inflammatory cytokines and chemokines tested, i.e., TNF-α, IL-8 and RANTES, as well as anti-inflammatory cytokines including IL-10. CFS restored TGF-ß levels in the presence of Salmonella. Live B.breve and its supernatant enhanced innate immune responses by the activation of TLR signaling pathway. These treatments upregulated TLR9 gene transcription. In addition, CFS was a more potent inducer of TLR9 expression than the probiotic bacteria in the presence of S. typhi. Expression levels of CASP8 and IRAK4 were also increased by CFS, and both treatments induced TOLLIP gene expression. Our results indicate that the probiotic strain B. breve CNCM I-4035 affects the intestinal immune response, whereas its supernatant exerts anti-inflammatory effects mediated by DCs. This supernatant may protect immune system from highly infectious agents such as Salmonella typhi and can down-regulate pro-inflammatory pathways.

Human intestinal dendritic cells decrease cytokine release against Salmonella infection in the presence of Lactobacillus paracasei upon TLR activation.

Probiotic bacteria have been shown to modulate immune responses and could have therapeutic effects in allergic and inflammatory disorders. However, little is known about the signalling pathways that are engaged by probiotics. Dendritic cells (DCs) are antigen-presenting cells that are involved in immunity and tolerance. Monocyte-derived dendritic cells (MoDCs) and murine DCs are different from human gut DCs; therefore, in this study, we used human DCs generated from CD34+ progenitor cells (hematopoietic stem cells) harvested from umbilical cord blood; those DCs exhibited surface antigens of dendritic Langerhans cells, similar to the lamina propria DCs in the gut. We report that both a novel probiotic strain isolated from faeces of exclusively breast-fed newborn infants, Lactobacillus paracasei CNCM I-4034, and its cell-free culture supernatant (CFS) decreased pro-inflammatory cytokines and chemokines in human intestinal DCs challenged with Salmonella. Interestingly, the supernatant was as effective as the bacteria in reducing pro-inflammatory cytokine expression. In contrast, the bacterium was a potent inducer of TGF-ß2 secretion, whereas the supernatant increased the secretion of TGF-ß1 in response to Salmonella. We also showed that both the bacteria and its supernatant enhanced innate immunity through the activation of Toll-like receptor (TLR) signalling. These treatments strongly induced the transcription of the TLR9 gene. In addition, upregulation of the CASP8 and TOLLIP genes was observed. This work demonstrates that L. paracasei CNCM I-4034 enhanced innate immune responses, as evidenced by the activation of TLR signalling and the downregulation of a broad array of pro-inflammatory cytokines. The use of supernatants like the one described in this paper could be an effective and safe alternative to using live bacteria in functional foods.

Infant formula supplemented with polyamines alters the intestinal microbiota in neonatal BALB/cOlaHsd mice.

Polyamines play a critical role in the development of intestinal and immune systems during the infant breastfeeding period, but the effect of polyamines on the microbiota has not been reported. The aim of our study was to characterize the impact on the colonization pattern in neonatal BALB/cOlaHsd mice after supplementing an infant formula (IF) with a mixture of putrescine (PUT), spermidine (SPD) and spermine (SPM). A total of 48 pups (14 days old) were randomly assigned to 4-day intervention groups as follows: breast-fed (unweaned) pups (n=12); weaned pups (n=12) fed an infant formula (IF); weaned pups (n=12) fed an IF enriched with a low concentration of PUT, SPD and SPM (2.10, 22.05 and 38.00 µg/day, respectively); and weaned pups (n=12) fed with IF enriched with a high concentration of PUT, SPD and SPM (8.40, 88.20 and 152.00 µg/day, respectively) of polyamines in accordance with normal proportions found in human milk. Microbiota composition was analyzed by fluorescent in situ hybridization (FISH) with flow cytometry detection. Microbiota changes in formula-fed mice were significantly greater following supplementation with polyamines (P<.01). Bifidobacterium group bacteria, Akkermansia-like bacteria and Lactobacillus-Enterococcus group levels were higher in the groups fed infant formula supplemented with polyamines, resulting in even higher numbers of bacteria than in the breastfed pups. Our findings indicate that infant formulas enriched with polyamines may interact with gut microbiota, suggesting that further studies in human infants are required to assess the impact of polyamines on both growth and microbiota levels.