Refermentation with yeast and lactic acid bacteria isolates: a strategy to improve the flavor of green coffee beans.

BACKGROUND: Yeast and lactic acid bacteria (LAB) play an important part in the post-harvest fermentation of coffee. This study applied lab-scale fermentation to commercial green coffee beans using dry coffee pulp as the substrate, with the aim of modifying coffee-bean flavor. In addition to spontaneous fermentation, yeast and LAB isolated from coffee beans and dried coffee pulp were added during fermentation. RESULTS: Co-inoculation of yeast and LAB showed a significant effect on the chlorogenic acid content after between 24 and 72 h of fermentation. Acetic, citric, malic, lactic, and quinic acids were shown to be affected significantly (P < 0.05) by fermentation and inoculation. Gas chromatography detected that esters, alcohols, aldehydes, furans, and pyrazines were the primary compounds in the coffee beans. Certain volatile groups were present in greater concentrations and broader varieties within the inoculated beans. The highest cupping scores were given to beans that had been co-inoculated with yeast and LAB. CONCLUSION: Overall, the use of yeasts and LAB starters showed potential to create coffee beverages with desirable characteristics by standardized fermentation. © 2024 Society of Chemical Industry.

Efficient production of human interleukin-3 from Escherichia coli using protein disulfide isomerase b'a' domain.

Human interleukin-3 (IL3) is a multifunctional cytokine essential for both clinical and biomedical research endeavors. However, its production in Escherichia coli has historically been challenging due to its aggregation into inclusion bodies, requiring intricate solubilization and refolding procedures. This study introduces an innovative approach employing two chaperone proteins, maltose binding protein (MBP) and protein disulfide isomerase b'a' domain (PDIb'a'), as N-terminal fusion tags. Histidine tag (H) was added at the beginning of each chaperone protein gene for easy purification. This fusion of chaperone proteins significantly improved IL3 solubility across various E. coli strains and temperature conditions, eliminating the need for laborious refolding procedures. Following expression optimization, H-PDIb'a'-IL3 was purified using two chromatographic methods, and the subsequent removal of the H-PDIb'a' tag yielded high-purity IL3. The identity of the purified protein was confirmed through liquid chromatography coupled with tandem mass spectrometry analysis. Biological activity assays using human erythroleukemia TF-1 cells revealed a unique two-step stimulation pattern for both purified IL3 and the H-PDIb'a'-IL3 fusion protein, underscoring the protein's functional integrity and revealing novel insights into its cellular interactions. This study advances the understanding of IL3 expression and activity while introducing novel considerations for protein fusion strategies.

Novel Anti-Mesothelin Nanobodies and Recombinant Immunotoxins with Pseudomonas Exotoxin Catalytic Domain for Cancer Therapeutics.

Recombinant immunotoxins (RITs) are fusion proteins consisting of a targeting domain linked to a toxin, offering a highly specific therapeutic strategy for cancer treatment. In this study, we engineered and characterized RITs aimed at mesothelin, a cell surface glycoprotein overexpressed in various malignancies. Through an extensive screening of a large nanobody library, four mesothelin-specific nanobodies were selected and genetically fused to a truncated Pseudomonas exotoxin (PE24B). Various optimizations, including the incorporation of furin cleavage sites, maltose-binding protein tags, and tobacco etch virus protease cleavage sites, were implemented to improve protein expression, solubility, and purification. The RITs were successfully overexpressed in Escherichia coli, achieving high solubility and purity post-purification. In vitro cytotoxicity assays on gastric carcinoma cell lines NCI-N87 and AGS revealed that Meso(Nb2)-PE24B demonstrated the highest cytotoxic efficacy, warranting further characterization. This RIT also displayed selective binding to human and monkey mesothelins but not to mouse mesothelin. The competitive binding assays between different RIT constructs revealed significant alterations in IC50 values, emphasizing the importance of nanobody specificity. Finally, a modification in the endoplasmic reticulum retention signal at the C-terminus further augmented its cytotoxic activity. Our findings offer valuable insights into the design and optimization of RITs, showcasing the potential of Meso(Nb2)-PE24B as a promising therapeutic candidate for targeted cancer treatment.

Crotamine-based recombinant immunotoxin targeting HER2 for enhanced cancer cell specificity and cytotoxicity.

Crotamine, one of the major toxins present in the venom of the South American rattlesnake Crotalus durissus terrificus, exhibits potent cytotoxic properties and has been suggested for cancer therapy applications. However, its selectivity for cancer cells needs to be improved. This study designed and produced a novel recombinant immunotoxin, HER2(scFv)-CRT, composed of crotamine and single-chain Fv (scFv) derived from trastuzumab targeting human epidermal growth factor receptor 2 (HER2). The recombinant immunotoxin was expressed in Escherichia coli and purified using various chromatographic techniques. The cytotoxicity of HER2(scFv)-CRT was assessed in three breast cancer cell lines, demonstrating enhanced specificity and toxicity in HER2-expressing cells. These findings suggest that the crotamine-based recombinant immunotoxin has the potential to expand the repertoire of recombinant immunotoxin applications in cancer therapy.

Predictors of Intention to Get a COVID-19 Vaccine of Health Science Students: A Cross-Sectional Study.

BACKGROUND: Vaccination is one of the interventions that can be employed to control coronavirus disease (COVID-19). The aim of this study is to identify factors that affect vaccination intention among health science students, by using the health belief model (HBM) as a theoretical framework. METHODS: This study was undertaken in April 2021 using a convenience sample strategy among health science students in Tra Vinh University, which is a university in South Vietnam. Participants were required to complete the self-report questionnaire. All data were calculated by using STATA 14 software, to determine the factors associated with COVID-19 vaccine acceptance, a multivariable logistic regression analysis was performed, along with odds ratio (OR) and 95% confidence interval (95% CI). RESULTS: A total of 412 eligible participants were taken part in the study, with a questionnaire response rate of 48.2%, reporting that 77.7% of participants intended to receive a COVID-19 vaccination when available. An overall score of knowledge was 7.8 ± 1.5 (range 0-10), a positive attitude toward vaccination including perceived susceptibility and severity (3.1 ± 0.7), perceived benefits (3.1 ± 0.7) and cues to action (4.0 ± 0.6). Determinants of willingness to get a COVID-19 vaccination included their field of education, knowledge and factors in HBM model including perceived susceptibility and severity, perceived benefits and cues to action (all p < 0.05). In contrast, perceived barriers had a negative association with the probability of vaccine acceptance (p < 0.05). The study did indicate a number of participants not willing to receive a vaccination due to, mainly, safety (67.4%), development issues (58.7%) and side effects (79.3%). CONCLUSION: The theoretical framework recognized factors that influence vaccine acceptance. These are considered evidence for the development of planning strategies to optimize the vaccination uptake and offer success in vaccination campaigns for both health science students and the general population.

Soluble Cytoplasmic Expression and Purification of Immunotoxin HER2(scFv)-PE24B as a Maltose Binding Protein Fusion.

Human epidermal growth factor receptor 2 (HER-2) is overexpressed in many malignant tumors. The anti-HER2 antibody trastuzumab has been approved for treating HER2-positive early and metastatic breast cancers. Pseudomonas exotoxin A (PE), a bacterial toxin of Pseudomonas aeruginosa, consists of an A-domain with enzymatic activity and a B-domain with cell binding activity. Recombinant immunotoxins comprising the HER2(scFv) single-chain Fv from trastuzumab and the PE24B catalytic fragment of PE display promising cytotoxic effects, but immunotoxins are typically insoluble when expressed in the cytoplasm of Escherichia coli, and thus they require solubilization and refolding. Herein, a recombinant immunotoxin gene was fused with maltose binding protein (MBP) and overexpressed in a soluble form in E. coli. Removal of the MBP yielded stable HER2(scFv)-PE24B at 91% purity; 0.25 mg of pure HER2(scFv)-PE24B was obtained from a 500 mL flask culture. Purified HER2(scFv)-PE24B was tested against four breast cancer cell lines differing in their surface HER2 level. The immunotoxin showed stronger cytotoxicity than HER2(scFv) or PE24B alone. The IC50 values for HER2(scFv)-PE24B were 28.1 ± 2.5 pM (n = 9) and 19 ± 1.4 pM (n = 9) for high HER2-positive cell lines SKBR3 and BT-474, respectively, but its cytotoxicity was lower against MDA-MB-231 and MCF7. Thus, fusion with MBP can facilitate the soluble expression and purification of scFv immunotoxins.

Soluble Prokaryotic Overexpression and Purification of Human GM-CSF Using the Protein Disulfide Isomerase b'a' Domain.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a member of the colony-stimulating factor (CSF) family, which functions to enhance the proliferation and differentiation of hematopoietic stem cells and other hematopoietic lineages such as neutrophils, dendritic cells, or macrophages. These proteins have thus generated considerable interest in clinical therapy research. A current obstacle to the prokaryotic production of human GM-CSF (hGM-CSF) is its low solubility when overexpressed and subsequent complex refolding processes. In our present study, the solubility of hGM-CSF was examined when combined with three N-terminal fusion tags in five E. coli strains at three different expression temperatures. In the five E. coli strains BL21 (DE3), ClearColi BL21 (DE3), LOBSTR, SHuffle T7 and Origami2 (DE3), the hexahistidine-tagged hGM-CSF showed the best expression but was insoluble in all cases at each examined temperature. Tagging with the maltose-binding protein (MBP) and the b'a' domain of protein disulfide isomerase (PDIb'a') greatly improved the soluble overexpression of hGM-CSF at 30 °C and 18 °C. The solubility was not improved using the Origami2 (DE3) and SHuffle T7 strains that have been engineered for disulfide bond formation. Two conventional chromatographic steps were used to purify hGM-CSF from the overexpressed PDIb'a'-hGM-CSF produced in ClearColi BL21 (DE3). In the experiment, 0.65 mg of hGM-CSF was isolated from a 0.5 L flask culture of these E. coli and showed a 98% purity by SDS-PAGE analysis and silver staining. The bioactivity of this purified hGM-CSF was measured at an EC50 of 16.4 ± 2 pM by a CCK8 assay in TF-1 human erythroleukemia cells.

Anthropometric Indexes for Predicting High Blood Pressure in Vietnamese Adults: A Cross-Sectional Study.

BACKGROUND: The rate of hypertension and obesity is increasing in Vietnamese society. This study aimed to focus on assessing the relationship between anthropometric indexes (body mass index (BMI), waist-to-height ratio (WHtR), waist-to-hip ratio (WHR)) and high blood pressure to determine which is the best predictor for high blood pressure among adults over 18 years in Vietnam. METHODS: A cross-sectional study of 1636 people was conducted. People who were over 18 years old, healthy or had hyperlipidemia and on the treatment were recruited. Patients with cancer, heart disease, diabetes, kidney disease, and hypertension were excluded. Information on demographics, smoking and drinking habits, weight, height, waist circumference, and blood pressure was collected. Areas under receiver operating characteristic curves (AUCs) were examined to determine the predictability of anthropometric indicators for high blood pressure in men and women. Logistic regression analysis, stratified by gender, was performed to examine the association between anthropometric indexes and high blood pressure. RESULTS: In this study, the percentage of people with high blood pressure was 10.51%. The AUC for the WHtR was significantly greater than for the BMI for both genders. Logistic regression demonstrated that only WHtR had a significant positive association with high blood pressure among women. The optimal WHtR cut-off value for predicting high blood pressure in men and women were 0.47 and 0.50, respectively. CONCLUSION: Among the indicators analyzed in this study, WHtR was the best for the predicting of the presence of high blood pressure, in both men and women. However, WHtR could only explain the changes of high blood pressure in women.

Knowledge, Attitude, and Practices Regarding COVID-19 Among Chronic Illness Patients at Outpatient Departments in Ho Chi Minh City, Vietnam.

BACKGROUND: The novel coronavirus disease (COVID-19) has become a major threat to human life around the world. This study aims to assess the knowledge, attitude, and practices regarding COVID-19 among people with chronic diseases at the outpatient departments in Ho Chi Minh City. METHODS: A cross-sectional study was carried out between February and March 2020 using a convenience sampling strategy in three hospitals in Ho Chi Minh City (HCMC) via the use of a structured self-administered questionnaire. Factors relating to practices, prevalence ratio (PR), and 95% confidence interval were estimated by using the Poisson regression with robust options. P-value <0.05 was considered as statistically different. RESULTS: A total of 522 participants had a mean age of 51.5 ± 10.6 years. Most of them reported seeing information regarding the COVID-19 pandemic (93.7%) via television and social media (72.8% and 62.1%, respectively). Just over two-thirds of the participants (68.4%) answered with sufficient knowledge of COVID-19. Most respondents had a positive attitude toward COVID-19 (90.8%), although some misconceptions existed. Almost over three-fourths of them (77.2%) maintained good practices for prevention. The rate of good practices in those who had sufficient knowledge was 1.24 times greater than that among those who had insufficient knowledge (PR 1.24, 95% CI: 1.10-1.41, P<0.05). Also, the rate of good practices in males was lower than that of females (PR: 0.91, 95% CI: 0.83-0.99, P<0.05). CONCLUSION: There still exists an amount of insufficient knowledge and negative attitude regarding COVID-19, which may be barriers to good prevention practices among chronic illness patients. Education programs need to continue via television and social media and emphasize that people with chronic diseases are more likely to experience severe symptoms, including death from COVID-19. Additionally, management authorities should prolong specific policies to protect the more vulnerable in our community.

Evaluation of the leaching behavior of incineration bottom ash using seawater: A comparison with standard leaching tests.

Batch and column tests were conducted on untreated incineration bottom ash (IBA) samples from two incineration plants in Singapore, using seawater as the leachant. The main objective of this study was to investigate the change in the leaching behavior of certain elements (i.e. As, Cd, Cr, Cu, Ni, Pb, Sb, Se and Zn) when IBA comes into contact with seawater. Such an investigation using seawater as leachant was not commonly carried out when investigating leaching behavior in IBA. The leaching tests were then carried out on the same IBA samples using DI water, as a comparison. Lower level of leaching was observed for Pb and Zn when seawater was used as the leachant. Cr and Sb showed significant cumulative release at Liquid-to-Solids (L/S) ratio 5 in the seawater column leaching. The influence of Dissolved Organic Carbon (DOC) on Cu leaching seems to decrease after L/S 2 when using seawater in the column test. Although the leaching behavior of IBA was affected when seawater was used, for the column test, there was no significant difference during the initial release when compared to DI water. The initial L/S fractions collected were important as the low L/S ratios represent the pore water concentration and the maximum output in an actual application. The results from this study would be useful for the future study on using IBA in marine applications.

Synergistic interactions of vancomycin with different antibiotics against Escherichia coli: trimethoprim and nitrofurantoin display strong synergies with vancomycin against wild-type E. coli.

Gram-negative bacteria are normally resistant to the antibiotic vancomycin (VAN), which cannot significantly penetrate the outer membrane. We used Escherichia coli mutants that are partially sensitive to VAN to study synergies between VAN and 10 other antibiotics representing six different functional categories. We detected strong synergies with VAN and nitrofurantoin (NTR) and with VAN and trimethoprim (TMP) and moderate synergies with other drugs, such as aminoglycosides. These synergies are powerful enough to show the activity of VAN against wild-type E. coli at concentrations of VAN as low as 6.25 µg/ml. This suggests that a very small percentage of exogenous VAN does enter E. coli but normally has insignificant effects on growth inhibition or cell killing. We used the results of pairwise interactions with VAN and the other 10 antibiotics tested to place VAN into a functional category of its own, as previously defined by Yeh et al. (P. Yeh, A. I. Tschumi, and R. Kishony, Nat Genet 28:489-494, 2006, http://dx.doi.org/10.1038/ng1755).

Development of supportive services for frontotemporal dementias through community engagement.

The effect of early onset frontotemporal dementias (FTD) on spouses and children is profound, requiring different types of support services from pre-existing Alzheimer's disease interventions already in place. This article explores how the needs of families living with FTD resulted in three programme initiatives developed at Baycrest (an academic health sciences centre focused on ageing, in Toronto, Canada) to meet the needs of this population. These included an Internet-based videoconferencing support group for spouses, a website that provides support and counsel for children and their parents, and an adult day programme designed for FTD patients. The strength of these interventions is that services were developed with involvement of stakeholders in FTD care from the start, to deal with gaps in services in a sustainable way.

Neuroimaging of delusions in Alzheimer's disease.

MEDLINE, Embase and PsycINFO were searched using the keywords "imaging", "neuroimaging", "CT", "MRI", "PET", "SPECT", "Alzheimer's", "dementia", "delusions" and "psychosis" to find studies specifically assessing or reporting on neuroimaging of delusions in Alzheimer's Dementia (AD), separate from hallucinations or psychosis in general in AD. Twenty-five studies were found meeting criteria and are included in this review which reports on structural, regional perfusion, metabolic and receptor binding imaging modalities assessing delusions as a whole, as well as persecutory and misidentification delusional subtypes. The majority of studies implicate right-sided pathology, primarily frontal lobe. Left-frontal predominance and release, secondary to right-sided pathology, may create a hyperinferential state resulting in the formation of delusions. This perturbation and imbalance of normal networks is associated with delusional phenomenology. Temporal lobe structures are also important in misidentification syndromes, which have a different natural history than paranoid delusions. Consistent with the neuropathological and genetic literature, neuroimaging has shown that paranoid versus misidentification delusions are associated with different phenomenology and different neural substrates. Delusional subtype is an important factor in understanding the neurobiological underpinnings of delusions in dementia. We also discuss methodological issues related to neuroimaging of delusions in AD.

Neurobiology of delusions in Alzheimer's disease.

Alzheimer's disease (AD) is associated with cognitive and functional impairment as well as neuropsychiatric sequelae, including psychotic symptoms such as delusions and hallucinations. Strong evidence supports the need to study delusions separate from hallucinations. Integrating the epidemiology, clinical correlates, and neuropathological and genetic literature for delusions in AD allows us to speculate on etiology and mechanisms. Plaque and tangle deposition in individuals with susceptible alleles of serotonergic, muscarinic, nicotinic, or Apoε4 genes appears to result in disruption of cortical circuitry, culminating in delusions. While delusions in AD correspond to a phenotype distinct from AD without delusions, subtypes of delusions may also define further distinct clinical entities. Persecutory delusions may occur earlier in the illness and have a more significant genetic component than misidentification delusions, which are associated with increased cognitive impairment and advanced dementia. Clearly distinguishing between these two syndromes is essential to making progress in the area of delusions in AD.