In vitro Selection and Characterization of a Light-up DNA Aptamer for Thiazole Orange.

A new DNA aptamer that binds to the target Thiazole Orange-biotin (TO1-biotin) was isolated after nine rounds of in vitro selection. The selection was performed on streptavidin-coated beads with the target bound to the surface and with free dye in solution in higher selection rounds to select for slower off-rate binding. Using next-generation sequencing (NGS), the libraries after the 4th and 9th rounds of selection were sequenced to identify enriched sequences. Several sequence families emerged, showing superior fluorescence enhancement and high affinity for the target compared to the other families obtained by NGS analysis. These sequence families were further studied to understand the binding interactions better. Primary sequence and secondary structure analysis tools were used to identify a hypothetical three-tiered G-quadruplex motif for these families. This indicates that the TO1-biotin DNA aptamer identified here uses a similar ligand-binding topology to the original Mango RNA aptamer.

Biophysical characterization and design of a minimal version of the Hoechst RNA aptamer.

RNA aptamers are oligonucleotides, selected through Systematic Evolution of Ligands by EXponential Enrichment (SELEX), that can bind to specific target molecules with high affinity. One such molecule is the RNA aptamer that binds to a blue-fluorescent Hoechst dye that was modified with bulky t-Bu groups to prevent non-specific binding to DNA. This aptamer has potential for biosensor applications; however, limited information is available regarding its conformation, molecular interactions with the ligand, and binding mechanism. The study presented here aims to biophysically characterize the Hoechst RNA aptamer when complexed with the t-Bu Hoechst dye and to further optimize the RNA sequence by designing and synthesizing new sequence variants. Each variant aptamer-t-Bu Hoechst complex was evaluated through a combination of fluorescence emission, native polyacrylamide gel electrophoresis, fluorescence titration, and isothermal titration calorimetry experiments. The results were used to design a minimal version of the aptamer consisting of only 21 nucleotides. The performed study also describes a more efficient method for synthesizing the t-Bu Hoechst dye derivative. Understanding the biophysical properties of the t-Bu Hoechst dye-RNA complex lays the foundation for nuclear magnetic resonance spectroscopy studies and its potential development as a building block for an aptamer-based biosensor that can be used in medical, environmental or laboratory settings.

Cross-Binding of Adenosine by Aptamers Selected Using Theophylline.

We recently reported that some adenosine binding aptamers can also bind caffeine and theophylline with around 20-fold lower affinities. This discovery led to the current work to examine the cross-binding of adenosine to theophylline aptamers. For the DNA aptamer for theophylline, cross-binding to adenosine was observed, and the affinity was 18 to 38-fold lower for adenosine based on assays using isothermal titration calorimetry and ThT fluorescence spectroscopy. The binding complexes were characterized using NMR spectroscopy, and both adenosine and theophylline showed an overall similar binding structure to the DNA theophylline aptamer, although small differences were also observed. In contrast, the RNA aptamer did not show binding to adenosine, although both aptamers have very similar relative selectivity for various methylxanthines including caffeine. After a negative selection, a few new aptamers with completely different primary sequences for theophylline were obtained and they did not show binding to adenosine. Thus, there are many ways for aptamers to bind theophylline and some can have cross-binding to adenosine. In biology, theophylline, caffeine, and adenosine can bind to the same protein receptors to regulate sleep, and their binding to the same DNA motifs may suggest an early role of nucleic acids in similar regulatory functions.

Perceptions of Resettled Refugee Congolese Women: Maintaining Cultural Traditions during Resettlement.

Conflict-displaced refugees have increased significantly globally. The Democratic Republic of Congo is the leading country with refugees in the United States, where many resettle in Ohio. Women refugees are highly vulnerable, yet little literature has focused on them. Furthermore, maintaining cultural traditions can provide comfort during the tumultuous resettlement process. Therefore, this study used mixed methods to understand the perceptions of Congolese refugee women on maintaining cultural traditions during resettlement in Ohio. Translator-assisted, orally administered demographic survey and face-to-face interviews were conducted among resettled Congolese refugee women (n = 20) 18 and older, who arrived in the United States from 2011 to 2018, and were currently receiving Ohio resettlement agency assistance. Researchers applied descriptive coding and thematic analysis to identify themes and subthemes. Three themes were identified among the resettled Congolese refugee women regarding maintaining cultural traditions in the United States. The three themes comprised (1) clothing and dressing, (2) food, and (3) parenting style. Our work examined resettled refugee Congolese women's perceptions of maintaining their culture after resettling in Ohio. These study findings could assist community engagers with insights and practical recommendations on supportive services for resettled Congolese women and a deeper understanding of complex acculturative situations facing them during resettlement.

Structure-function characterization of an aldo-keto reductase involved in detoxification of the mycotoxin, deoxynivalenol.

Deoxynivalenol (DON) is a mycotoxin, produced by filamentous fungi such as Fusarium graminearum, that causes significant yield losses of cereal grain crops worldwide. One of the most promising methods to detoxify this mycotoxin involves its enzymatic epimerization to 3-epi-DON. DepB plays a critical role in this process by reducing 3-keto-DON, an intermediate in the epimerization process, to 3-epi-DON. DepBRleg from Rhizobium leguminosarum is a member of the new aldo-keto reductase family, AKR18, and it has the unusual ability to utilize both NADH and NADPH as coenzymes, albeit with a 40-fold higher catalytic efficiency with NADPH compared to NADH. Structural analysis of DepBRleg revealed the putative roles of Lys-217, Arg-290, and Gln-294 in NADPH specificity. Replacement of these residues by site-specific mutagenesis to negatively charged amino acids compromised NADPH binding with minimal effects on NADH binding. The substrate-binding site of DepBRleg is larger than its closest structural homolog, AKR6A2, likely contributing to its ability to utilize a wide range of aldehydes and ketones, including the mycotoxin, patulin, as substrates. The structure of DepBRleg also suggests that 3-keto-DON can adopt two binding modes to facilitate 4-pro-R hydride transfer to either the re- or si-face of the C3 ketone providing a possible explanation for the enzyme's ability to convert 3-keto-DON to 3-epi-DON and DON in diastereomeric ratios of 67.2% and 32.8% respectively.

Mycotoxin Metabolism by Edible Insects.

Mycotoxins are a group of toxic secondary metabolites produced in the food chain by fungi through the infection of crops both before and after harvest. Mycotoxins are one of the most important food safety concerns due to their severe poisonous and carcinogenic effects on humans and animals upon ingestion. In the last decade, insects have received wide attention as a highly nutritious, efficient and sustainable source of animal-derived protein and caloric energy for feed and food purposes. Many insects have been used to convert food waste into animal feed. As food waste might contain mycotoxins, research has been conducted on the metabolism and detoxification of mycotoxins by edible insects. The mycotoxins that have been studied include aflatoxins, fumonisins, zearalenone (ZEN), vomitoxin or deoxynivalenol (DON), and ochratoxins (OTAs). Aflatoxin metabolism is proved through the production of hydroxylated metabolites by NADPH-dependent reductases and hydroxylases by different insects. ZEN can be metabolized into α- and ß-zearalenol. Three DON metabolites, 3-, 15-acetyl-DON, and DON-3-glucoside, have been identified in the insect DON metabolites. Unfortunately, the resulting metabolites, involved enzymes, and detoxification mechanisms of OTAs and fumonisins within insects have yet to be identified. Previous studies have been focused on the insect tolerance to mycotoxins and the produced metabolites; further research needs to be conducted to understand the exact enzymes and pathways that are involved.

UV treatment for degradation of chemical contaminants in food: A review.

Application of ultraviolet (UV) irradiation for the degradation of chemical contaminants in food products has gained more and more interest in the past two decades. The majority of the research in this field was on mycotoxins, especially aflatoxins and patulin, with limited studies on pesticide residues and other chemical contaminants in food. These studies have been focused on identifying the structure and toxicity of degradation products, investigating the influence of UV treatment factors on the degradation efficiency, determining the impact of UV treatment on the quality of food products, and developing updated UV treatment methods such as TiO2 induced photocatalytic degradation. The summary of published literatures provided insights into future research opportunities in this area, which include determining a standard for the UV treatment description, working with naturally contaminated samples rather than artificially spiked samples, conducting pilot plant or industrial scale studies, examining more targets and conducting multi-targets studies, and developing more innovative methods for UV treatment.

A review of research on plant-based meat alternatives: Driving forces, history, manufacturing, and consumer attitudes.

The topic of plant-based meat alternatives (PBMAs) has been discussed for several decades, but it has only recently become one of the hottest topics in the food and research communities. With the purpose of investigating the current situation of scientific research on PBMA and determining future research opportunities, the driving forces for PBMA development, a brief history of its progression, key technologies required for production, and the resulting consumer attitudes are summarized. Environmental, human health, and animal welfare concerns are the main factors that have driven the development of PBMA. Although its history can trace back to ancient Asian civilizations, the first generation of PBMA originated in 1960s and a new generation of PBMA designed for carnivore was developed in recently years. Structuring methods such as extrusion and shear cell techniques have been widely studied, but improvements toward the overall appearance and flavor, biological and chemical safety control, as well as the selection of protein sources are also very important for PBMA production. The consumer acceptance of PBMA remains unsatisfactory but is continually improving. Based on those knowledge, future research opportunities include developing more effective strategies for consumer education, providing more scientific evidence for the health properties of PBMA, finding more suitable protein sources to improve the quality of the final products, improving the appearance and flavor, further examining and securing the chemical safety, exploring the structure formation mechanism during the extraction or shearing processes, and developing methods and standards for a quality evaluation of PBMA.

Validating a path model of adherence to prenatal care recommendations among pregnant women.

OBJECTIVE: To date, no study has reported a diagrammatic path model that involves patient-provider communication on pregnant women's adherence to prenatal care recommendations. To bridge this gap, this study aimed to validate a path model to display the direct and indirect influences on adherence to prenatal care recommendations. METHODS: A cross-sectional study founded on an evidence-based proposed theoretical framework was conducted among pregnant women (18-45 years) in their second or third trimester (n = 401) in the Midwestern United States. The proposed theoretical framework examined multiple levels of influences. Previously validated instruments were pilot tested and modified. Path analysis was conducted to validate the best-fit path model. RESULTS: The path model showed shared decision-making and motivation significantly predicted adherence to prenatal care recommendations and accounted for 10% (R2) of the variance. Cultural competency, interaction, perceived discrimination, and satisfaction accounted for 30% (R2) of the variance of shared decision-making. Patient's trust, distrust, and self-efficacy accounted for 5% (R2) of motivation's variance. CONCLUSION: Our work identified the best-fit path model for adherence to prenatal care recommendations. PRACTICE IMPLICATIONS: Incorporating findings from this study could assist prenatal care providers in understanding many complex variables affecting prenatal care, ultimately reducing infant mortality.

The Relationships Between Perceived Discrimination and Utilization of Mental Health Services Among African Americans and Caribbean Blacks.

Although all ethnic/racial groups underutilize mental health services, only about one-third of African Americans and Caribbean Blacks in need utilize mental health services. An extensive literature review led us to examine if an association between perceived discrimination and mental health utilization exists. The National Survey of American Life 2001-2003 nationally representative dataset was analyzed. Variables including utilization of mental health services, perceived discrimination, and socio-demographic variables were assessed for their relationships. The study found that each of the following predictors statistically significant: being male, being employed, having household income higher than 100,000, perceiving no discrimination, being African American, or being Caribbean Black are less likely to utilize mental health services than their counterparts. This study contributes to the emerging body of evidence demonstrating that perceived discrimination has a strong connection with the utilization of mental health services.

Influences of Socio-Demographic Factors and Health Utilization Factors on Patient-Centered Provider Communication.

Patient-centered care has been documented as a measure of quality of health care and has been associated with positive health outcomes. However, the effect of health utilization on improving patient-centered communication has not been investigated. This study examined the effect of three important kinds of health utilization: routine check-up, frequency of provider visits in the last year, and quality of health care to patient-centered provider communication. Cross-sectional data from 3,608 respondents to Health Information National Trends Survey-Cycle 4 2014 were analyzed. Multiple regressions were used to examine the association of sociodemographic factors and health utilization to patient-centered provider communication. Results showed that adults above 50 years and women reported higher patient-centered provider communication. Hispanic and Asian versus White respondents reported poorer patient-centered provider communication. Respondents with routine checkups between 1 and 2 years, 2 and 5 years, 5 or more years and none were all negatively associated with patient-centered provider communication in comparison with routine checkup within 1 year. Respondents who didn't visit health provider within past year had poorer patient-centered provider communication when compared to those who visited once. Finally, higher quality of healthcare experience was associated with higher patient-centered provider communication. Thus, this study highlights that race and ethnicity, age, and gender are significant factors that influence patient-centered provider communication; and specifically higher quality of healthcare experience, one provider visit within past year, and annual routine checkup as measures of health utilization predicts improved patient-centered provider communication.

Synthesis, physiochemical characterization, and biocompatibility of a chitosan/dextran-based hydrogel for postsurgical adhesion prevention.

An amine-functionalized succinyl chitosan and an oxidized dextran were synthesized and mixed in aqueous solution to form an in situ chitosan/dextran injectable, surgical hydrogel for adhesion prevention. Rheological characterization showed that the rate of gelation and moduli were tunable based on amine and aldehyde levels, as well as polymer concentrations. The CD hydrogels have been shown to be effective post-operative aids in prevention of adhesions in ear, nose, and throat surgeries and abdominal surgeries in vivo. In vitro biocompatibility testing was performed on CD hydrogels containing one of two oxidized dextrans, an 80 % oxidized (CD-100) or 25 % (CD-25) oxidized dextran. However, the CD-100 hydrogel showed moderate cytotoxicity in vitro to Vero cells. SC component of the CD hydrogel, however, showed no cytotoxic effect. In order to increase the biocompatibility of the hydrogel, a lower aldehyde level hydrogel was developed. CD-25 was found to be non-cytotoxic to L929 fibroblasts. The in vivo pro-inflammatory response of the CD-25 hydrogel, after intraperitoneal injection in BALB/c mice, was also determined by measuring serum TNF-α levels and by histological analysis of tissues. TNF-α levels were similar in mice injected with CD-25 hydrogel as compared to the negative saline injected control; and were significantly different (P < 0.05) as compared to the positive, lipopolysaccharide, injected control. Histological examination revealed no inflammation seen in CD hydrogel injected mice. The results of these in vitro and in vivo studies demonstrate the biocompatibility of the CD hydrogel as a post-operative aid for adhesion prevention.