In medicine and other academic settings, (doctoral) students often work in interdisciplinary teams together with researchers of pharmaceutical sciences, natural sciences in general, or biostatistics. They should be fundamentally taught good research practices, especially in terms of statistical analysis. This includes reproducibility as a central aspect. Acknowledging that even experienced researchers and supervisors might be unfamiliar with necessary aspects of a perfectly reproducible workflow, a lecture series on reproducible research (RR) was developed for young scientists in clinical pharmacology. The pilot series highlighted definitions of RR, reasons for RR, potential merits of RR, and ways to work accordingly. In trying to actually reproduce a published analysis, several practical obstacles arose. In this article, reproduction of a working example is commented to emphasize the manifold facets of RR, to provide possible explanations for difficulties and solutions, and to argue that harmonized curricula for (quantitative) clinical researchers should include RR principles. These experiences should raise awareness among educators and students, supervisors and young scientists. RR working habits are not only beneficial for ourselves or our students, but also for other researchers within an institution, for scientific partners, for the scientific community, and eventually for the public profiting from research findings.

OBJECTIVE: The spread of bovine spongiform encephalopathy (BSE) agent to small ruminants is still a major issue in the surveillance of transmissible spongiform encephalopathies (TSEs). L-type bovine spongiform encephalopathy (L-BSE) is an atypical form of BSE with an unknown zoonotic potential that is transmissible to cattle and small ruminants. Our current knowledge of bovine atypical prion strains in sheep and goat relies only on experimental transmission studies by intracranial inoculation. To assess oral susceptibility of goats to L-BSE, we orally inoculated five goats with cattle L-BSE brain homogenates and investigated pathogenic prion protein (PrPsc) distribution by an ultrasensitive in vitro conversion assay known as Real-Time Quaking Induced Conversion (RT-QuIC). RESULTS: Despite a prolonged observation period of 80 months, all these animals and the uninfected controls did not develop clinical signs referable to TSEs and tested negative by standard diagnostics. Otherwise, RT-QuIC analysis showed seeding activity in five out of five examined brain samples. PrPsc accumulation was also detected in spinal cord and lymphoreticular system. These results indicate that caprine species are susceptible to L-BSE by oral transmission and that ultrasensitive prion tests deserve consideration to improve the potential of current surveillance systems against otherwise undetectable forms of animal prion infections.

OBJECTIVE: Evidence show that cocaine and amphetamine regulated transcript-prepropeptide (CART-PT) gene variants may affect obesity related traits, but little is known about its end points. In the current study, we aimed to evaluate the interaction of CARTPT gene polymorphism with diet quality indices including dietary approaches to stop hypertension (DASH) and Mediterranean diet score (MDS) on cardio-metabolic risk factors. This cross sectional study recruited 288 apparently healthy obese individuals. Diet quality indices including DASH and MDS were evaluated using semi quantitative food frequency questionnaire (FFQ). Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used for CARTPT genotypes. RESULTS: No significant differences was reported for general characteristics and biochemical parameters across genotypes except for QUICKI among females (P = 0.01) and it was higher in heterozygous genotype. There was significant CARTPT-DASH interactions affecting serum fasting glucose level (P = 0.049). However, in relation to CERTPT-MDS interactions, the highest level of insulin (P = 0.003) and HOMA-IR (P = 0.003) values were shown among AA carriers in high adherence to MDS, while AA carriers in high compliance to MDS experienced decreased level of QUICKI (P = 0.001).

OBJECTIVE: The advancement of molecular techniques in an era in which high-throughput sequencing has revolutionized biology renders old-fashioned alternatives to high-throughput methods obsolete. Such advanced molecular techniques, however, are not yet accessible to economically disadvantaged region-based laboratories that still obtain DNA profiles using gel-based techniques. To explore whether cost-efficient techniques can produce results that are as robust as those obtained using high-throughput methods, we compared the performance of polyacrylamide gel electrophoresis (PAGE)- and capillary electrophoresis (CE)-derived genomic data in estimating genetic diversity and inferring relatedness using 70 individuals of fine flounder (Paralichthys adspersus) selected from a hatchery population and genotyped for five microsatellite loci. RESULTS: Here, we show that PAGE- and CE-derived genomic datasets yield comparable genetic diversity levels regarding allelic diversity measures and heterozygosity. However, relatedness inferred from each dataset showed that the categorization of dyads in the different relationship categories strongly differed. This suggests that while scientists can reliably use PAGE-derived genomic data to estimate genetic diversity, they cannot use the same for parentage testing. The findings could help laboratories committed to population research not be discouraged from using the PAGE system if high-throughput technologies are unavailable and the method is adequate to address the biological question.

OBJECTIVE: Convalescent plasma has been tried as therapy for various viral infections. Early observational studies of convalescent plasma treatment for hospitalized COVID-19 patients were promising, but randomized controlled studies were lacking at the time. The objective of this study was to investigate if convalescent plasma is beneficial to hospitalized patients with COVID-19. RESULTS: Hospitalized patients with confirmed COVID-19 and an oxygen saturation below 94% were randomized 1:1 to receive convalescent plasma in addition to standard of care or standard of care only. The primary outcome was number of days of oxygen treatment to keep saturation above 93% within 28 days from inclusion. The study was prematurely terminated when thirty-one of 100 intended patients had been included. The median time of oxygen treatment among survivors was 11 days (IQR 6-15) for the convalescent plasma group and 7 days (IQR 5-9) for the standard of care group (p = 0.4, median difference -4). Two patients in the convalescent plasma group and three patients in the standard of care group died (p = 0.64, OR 0.49, 95% CI 0.08-2.79). Thus no significant differences were observed between the groups. Trial registration ClinicalTrials NCT04600440, retrospectively registered Oct 23, 2020.

OBJECTIVE: Music-related physical and mental health conditions are common among post-secondary music students, with many studies reporting a prevalence greater than 70%. However, there is currently no consensus on appropriate, validated assessments for this population. The aim of this pilot study was to test the feasibility of an assessment protocol developed for a German longitudinal study with Canadian post-secondary music students, and to compare the health of music students to non-music students. Using a cross-sectional design, first-semester music and non-music control students were recruited at two campuses at the same university. Both groups completed questionnaires and physical testing, including range of motion, core strength, and pressure pain threshold. Nineteen music students and 50 non-music student controls participated in this study. RESULTS: The German protocol is feasible in a Canadian post-secondary setting. Canadian music students demonstrated similar health outcomes to those in the parent study. All participants demonstrated poorer mental and physical quality of life than the Canadian norms, though this was not statistically significant. The results of this study should be confirmed in a larger study. Future studies with larger sample sizes can provide further insight into the health of Canadian music students, providing a basis for prevention and intervention.

OBJECTIVE: Accurate determination of the mixing time in bioreactors is essential for the optimization of the productivity of bioprocesses. The aim of this work was to develop a simple optical method to determine the mixing time in a photobioreactor. The image processing method should be based on freeware tools, should not require programming skills, and thus could be used in education within high schools and in early stages of undergraduate programs. RESULTS: An optical method has been established to analyze images from recorded videos of mixing experiments. The steps are: 1. Extraction of a sequence of images from the video file; 2. Cropping of the pictures; 3. Background removal; and 4. Image analysis and mixing time evaluation based on quantification of pixel-to-pixel heterogeneity within a given area of interest. The novel method was generally able to track the dependency between aeration rate and mixing time within the investigated photobioreactor. In direct comparison, a PEARSON correlation coefficient of rho = 0.99 was obtained. Gas flow rates between 10 L h-1, and 300 L h-1 resulted from mixing times of between 48 and 14 s, respectively. This technique is applicable without programming skills and can be used in education with inexperienced user groups.

OBJECTIVE: The purpose of this study was to examine the ways that encouraged people to develop positive attitudes and perceptions toward inclusive education. The Japanese special needs education system for students with disabilities has been shifting from a segregated model to a more inclusive form which is the major challenge facing educational systems around the world. While support for inclusive practices has grown rapidly in Japan, their implementation requires more attention. Considering these situations, in the current study, we experimentally manipulated future-oriented thinking and examined whether positive perceptions about inclusive education was enhanced if people acknowledged and realized that an inclusive society may improve the long-term welfare of not only people with disabilities but also people without disabilities or functional limitations. RESULTS: Our results partially confirmed that future-oriented thinking encouraged positive perceptions of inclusive education. It increased only when participants thought about the future employment of people with/without disabilities. No significant effects were found for the present orientation or control conditions.

OBJECTIVE: The use of benchtop metabolic profiling technology based on nuclear magnetic resonance (NMR) was evaluated in a small cohort of cats with a view to applying this as a viable and rapid metabolic tool to support clinical decision making. RESULTS: Urinary metabolites were analysed from four subjects consisting of two healthy controls and two chronic kidney disease (CKD) IRIS stage 2 cases. The study identified 15 metabolites in cats with CKD that were different from the controls. Among them were acetate, creatinine, citrate, taurine, glycine, serine and threonine. Benchtop NMR technology is capable of distinguishing between chronic kidney disease case and control samples in a pilot feline cohort based on metabolic profile. We offer perspectives on the further development of this pilot work and the potential of the technology, when combined with sample databases and computational intelligence techniques to offer a clinical decision support tool not only for cases of renal disease but other metabolic conditions in the future.

OBJECTIVE: The proliferation of false information on COVID-19 mostly through social media is adversely affecting control efforts. The objective of this study was to identify areas where targeted effective messaging can be useful in demystifying misinformation against COVID-19. RESULTS: The study showed high levels of misinformation on COVID-19 in the study area [mean score 2.71; standard deviation (SD) 1.5]. The highest levels of misinformation were observed in Dr. Ruth Segomotsi Mompati district, North West province (mean score: 3.84; SD: 2.1) and Sedibeng district, Gauteng province (mean score: 3.56; SD 1.7). Higher levels of misinformation were reported by those aged 18-24 years (mean score: 3.48; SD: 1.8), and men (mean score: 2.73; SD: 1.8). Across the two provinces, we identified geospatial hot and coldspots of misinformation highlighting the need to implement point of care strategies such as targeted messaging. Findings showed the need for targeted interventions to young people, students, those with low levels of education and the self-employed in the two districts more importantly, as South Africa expands its nationwide vaccination roll-out.

OBJECTIVE: Most epidemiological studies on suicidal behavior have been focused on high-income country settings. This study examine factors associated with suicidal behaviors among school-attending adolescents in three island nations. In this secondary analysis of the publicly available 2015 nationally representative GSHS data, we tested demographic, social, and behavioral attributes using multivariable logistic regression to association with suicide attempts. RESULTS: Within the recall period, 13.6% of participants reported having attempted suicide one or more times in the Cook Islands, 10.8% in Curaçao, and 9.8% in East Timor. In the Cook Islands, suicide ideation (AOR = 19.42, 95% CI = 9.11-41.41), anxiety (AOR = 2.51, 95% CI = 1.08-5.82), physical bullying (AOR = 3.3, 95% CI = 1.10-9.91), and cigarette smoking (AOR = 3.82, 95% CI = 1.38-10.54) were associated with suicide attempts. For Curaçaoo, suicide ideation (AOR = 7.88, 95% CI = 5.20-11.95) and suicide planning (AOR = 7.01, 95% CI = 4.24-11.60) were statistically significant. While for East Timor, suicide ideation (AOR = 4.59, 95% CI = 2.14-9.88), suicide planning (AOR = 3.36, 95% CI = 1.76-6.29), bullying victimization (AOR = 2.69, 95% CI = 1.02-7.12), and serious injuries (AOR = 2.22, 95% CI = 1.31-3.74) were statistically significant. Suicide attempt is relatively common in each of the three island nations. The socioeconomic context of adolescents might play a significant role in moderating suicidal behavior. Therefore, prevention efforts should be grounded in view of geographic, demographic, and socioeconomic contexts of the populations at risk.

OBJECTIVES: SARS-CoV-2, severe respiratory syndrome coronavirus-2, is an RNA virus that emerged from China sweeping the globe in the form of a pandemic that became an international public health concern. This pilot study aimed to describe the genetic variation and molecular epidemiology of SARS-CoV-2 in Palestine in fall 2020. RESULTS: To achieve these aims, whole genome sequencing of SARS-CoV-2, phylogenetic analysis, haplotype networking and genetic diversity analysis were performed. These analyses revealed a unique spike mutation H245N that has never been reported before. The phylogenetic analysis depicted that three clusters existed in Palestinian SARS-CoV-2 genome sequences, in which cluster-I comprised the majority of clusters by 90%. Congruently, the haplotype network analysis depicted the same three clusters with a total of 39 haplotypes. The genetic diversity analysis showed that Cluster-I is highly diverse as confirmed by statistically significant mutation rate indices, Tajima's D and Fu-Li's-F tests (- 2.11 and 2.74, respectively), highest number of mutations (Eta = 120), highest number of haplotypes (h = 17), and highest average number of nucleotide differences between any two sequences (S = 118). The study confirmed the high genetic diversity among the Palestinian of SARS-CoV-2 which possessed high number of mutations including one which was reported for the first time.

OBJECTIVE: Improvements in balance performance through balance training programs in children have been reported in several studies. However, the influence of balance training modalities (e.g., training period, frequency, volume) on the training effectiveness have not yet been studied. To address this shortfall, the present study investigated the effects of balance training duration and volume (i.e., 240 min during 4 weeks versus 360 min during 6 weeks) on measures of static and dynamic balance performance in healthy children (N = 29) aged 10 years. RESULTS: Irrespective of balance training duration and volume, significant pre- to post-test improvements were found for variables of static (i.e., one-legged stance on foam ground, reduced number of floor contacts: p = .041, ηp2 = .15) and dynamic (i.e., Lower Quarter Y Balance test, increased anterior reach distance: p = .038, ηp2 = .15) balance performance but no group × test interactions were detected. These findings indicate that balance training is effective to improve static and dynamic balance performance in healthy children, but the effectiveness seems unaffected by the applied training duration and volume. Trial Registration Current Controlled Trials ISRCTN75170753 (retrospectively registered at 12th April, 2021).

Many research participants are willing to be contacted about future research opportunities, however this question is not always asked. Furthermore, if participants do consent for contact about future research, this information is not always accessible to other research teams. We discuss our experience of recruiting individuals who have previously taken part in healthcare research and suggest potential strategies to support this process and enable greater research participation.

OBJECTIVE: Coronaviruses (CoVs) are natural commensals of bats. Two subgenera, namely Sarbecoviruses and Merbecoviruses have a high zoonotic potential and have been associated with three separate spillover events in the past 2 decades, making surveillance of bat-CoVs crucial for the prevention of the next epidemic. The study was aimed to elucidate the presence of coronavirus in fresh bat guano sampled from Wind Cave Nature Reserve (WCNR) in Sarawak, Malaysian Borneo. Samples collected were placed into viral transport medium, transported on ice within the collection day, and preserved at - 80 °C. Nucleic acid was extracted using the column method and screened using consensus PCR primers targeting the RNA-dependent RNA polymerase (RdRp) gene. Amplicons were sequenced bidirectionally using the Sanger method. Phylogenetic tree with maximum-likelihood bootstrap and Bayesian posterior probability were constructed. RESULTS: CoV-RNA was detected in ten specimens (47.6%, n  = 21). Six alphacoronavirus and four betacoronaviruses were identified. The bat-CoVs can be phylogenetically grouped into four novel clades which are closely related to Decacovirus-1 and Decacovirus-2, Sarbecovirus, and an unclassified CoV. CoVs lineages unique to the Island of Borneo were discovered in Sarawak, Malaysia, with one of them closely related to Sarbecovirus. All of them are distant from currently known human coronaviruses.

OBJECTIVE: The aims of this research were to determine the genomic properties of BI-EHEC to control Enterohemorrhagic Escherichia coli (EHEC), which was isolated from previous study. Genomic analysis of this phage is essential for the assessment of this bacteriophage for further application as food preservatives. RESULTS: Genome of BI-EHEC was successfully annotated using multiPhATE2. Structural and lytic cycle-related proteins such as head, tail, capsid, and lysozyme (lysin) were annotated. The phylogenetic tree of tail fiber protein and BRIG results showed that BI-EHEC was similar to phages of the same host in the bacteriophage genome database. There were no indications of virulence properties, antibiotic resistance genes and lysogenic protein among annotated genes which implied BI-EHEC followed a lytic life cycle. PHACTS analysis was done to confirm this notion further and yielded a lytic cycle result. Further analysis using CARD found that BI-EHEC does not contain residual ARGs per recommended parameter. Furthermore, BI-EHEC confirmed as lytic bacteriophage, making it a good candidate for biocontrol agent.

OBJECTIVE: NGFR/p75NTR and NRADD/NRH proteins are closely related structurally and are encoded by genes that arose from a duplication event early in vertebrate evolution. The transmembrane domain (TMD) of NGFR is cleaved by γ-secretase but there is conflicting data around the susceptibility to γ-secretase cleavage of NRADD proteins. If NGFR and NRADD show differential susceptibility to γ-secretase, then they can be used to dissect the structural constraints determining substrate susceptibility. We sought to test this differential susceptibility. RESULTS: We developed labelled, lumenally-truncated forms of zebrafish Ngfrb and Nradd and a chimeric protein in which the TMD of Nradd was replaced with the TMD of Ngfrb. We expressed these in zebrafish embryos to test their susceptibility to γ-secretase cleavage by monitoring their stability using western immunoblotting. Inhibition of γ-secretase activity using DAPT increased the stability of only the Ngfrb construct. Our results support that only NGFR is cleaved by γ-secretase. Either NGFR evolved γ-secretase-susceptibility since its creation by gene duplication, or NRADD evolved to be refractory to γ-secretase. Protein structure outside of the TMD of NGFR is likely required for susceptibility to γ-secretase.

The adoption and incentivisation of open and transparent research practices is critical in addressing issues around research reproducibility and research integrity. These practices will require training and funding. Individuals need to be incentivised to adopt open and transparent research practices (e.g., added as desirable criteria in hiring, probation, and promotion decisions, recognition that funded research should be conducted openly and transparently, the importance of publishers mandating the publication of research workflows and appropriately curated data associated with each research output). Similarly, institutions need to be incentivised to encourage the adoption of open and transparent practices by researchers. Research quality should be prioritised over research quantity. As research transparency will look different for different disciplines, there can be no one-size-fits-all approach. An outward looking and joined up UK research strategy is needed that places openness and transparency at the heart of research activity. This should involve key stakeholders (institutions, research organisations, funders, publishers, and Government) and crucially should be focused on action. Failure to do this will have negative consequences not just for UK research, but also for our ability to innovate and subsequently commercialise UK-led discovery.

OBJECTIVE: The Angiotensin 1 converting enzyme (ACE1) gene plays a critical role in regulating blood pressure and thus, it has become a major therapeutic target of antihypertensives. Single nucleotide polymorphisms (SNPs) occurring within a gene most especially at the functional segment of the genes alter the structure-function relationship of that gene. RESULTS: Our study revealed that five nsSNPs of the ACE1 gene were found to be potentially deleterious and damaging and they include rs2229839, rs14507892, rs12709442, and rs4977 at point mutations P351R, R953Q, I1018T, F1051V, and T1187M. The protein stability predictive tools revealed that all the nsSNPs decreased stability of the protein and the Consurf server which estimates the evolutionary conservation profile of a protein showed that three mutants were in the highly conserved region. In conclusion, this study predicted potential druggable deleterious mutants that can be further explored to understand the pathological basis of cardiovascular disease.

OBJECTIVES: During the COVID-19 pandemic, most countries implemented lockdowns that motivated changes in the dietary patterns, physical activity, and body mass index (BMI) of consumers worldwide, as well as the emergence of new food marketing strategies in social media. We sought to design and validate a methodology for monitoring and evaluating the Facebook marketing strategies of multinational fast-food chains in response to the COVID-19 pandemic. DATA DESCRIPTION: We developed three datasets. First, a dataset with the Uniform Resource Locators (URLs) of 1015 Facebook posts of five fast-food chains present in Argentina, Bolivia, Guatemala, and Peru. Second, a dataset of 106 content-analyzed posts we used in a pilot to determine intercoder reliability using statistical tests. Third, a dataset of a final sample of the 1015 content-analyzed posts that we used to determine the variables most frequently used. Following a mixed-methods approach, we developed 29 variables that recorded general information, as well as the marketing strategies we identified in the posts, including 14 COVID-19 specific variables. These data should help to monitor the social media marketing strategies that fast-food chains have introduced during the COVID-19 lockdowns, thus providing initial evidence about how they could be contributing to an unhealthy food environment.

Erratum in BMC Res Notes. 2022 Feb 14;15(1):53.

OBJECTIVES: Assessing the likelihood of success of induction of labour using cervical volume is an important research question. DATA DESCRIPTION: We provide data generated in a prospective observational study which was carried out at North Colombo Teaching Hospital, Ragama, Sri Lanka. Study conducted to compare pre-induction digital cervical assessment, sonographic cervical length, and sonographic cervical volume with vaginal delivery rate within 24 h. Inductions with 100 singleton pregnancies at term were included.

Erratum in BMC Res Notes. 2022 Jul 14;15(1):246.

OBJECTIVE: Cisplatin is a conventional anticancer drug that generates reactive oxygen species and causes apoptosis. However, many cancer cells develop alterations in the ATP binding cassette transporter responsible for the uptake and efflux process, which leads to resistance. Many natural products have shown potential to compete with ATP binding cassette transporter and may sensitize resistant cells to cisplatin. Studies have shown pro-oxidant effect of carotenoids that promote apoptosis of cancer cells. Bixin and fucoxanthin are well-known carotenoids with known antioxidant properties, however their bioactivity in lung cancer cells, clinically known to develop resistance due to ATP binding cassette transporter, has been minimally studied. This study is the first to investigate the potential of bixin and fucoxanthin to sensitize human lung cancer cell line, A549 and cervical cancer cell line, HeLa, to cisplatin. Drug combination method developed by Chou and Talalay theorem was employed. RESULT: Employing the best combination ratio, this study shows selective sensitization of cancer cells to cisplatin after bixin and fucoxanthin treatment. Further study on the mechanism of action in specific types of cancer cells is warranted. It may improve cisplatin sensitivity in tumors and rational use of cancer drugs.

OBJECTIVES: Bombyx mori is the key contributor to industrial silk production. The maximum production of silk occurs during 5th instar. The posterior silk glands in the larvae are responsible for the production of the main component of silk fibre-fibroin. The expression of genes and their regulation are dependent on the chromatin architecture. The nuclear matrix supports its structure and function by anchoring specific regions to regulate gene expression. The major constituent of the nuclear matrix, crucial to its structural and temporal maintenance, is its RNA. Therefore, the study of nuclear matrix RNA of the posterior silk glands on different days of 5th instar larval development is essential to understand its association to differential expression of genes. DATA DESCRIPTION: The tissue-specific developmental association of nuclear matrix RNA (NuMat RNA) at the genome level has not been done so far for any organism. Bombyx mori, CSR2 X CSR4 is the most popular dihybrid strain in India. The nuclear matrix RNA was isolated from day 1, day 5 and day 7 of 5th instar posterior silk glands of Bombyx mori. The NuMat RNA was sequenced using Illumina platform. The reads obtained were processed and the datasets were deposited in NCBI.

OBJECTIVE: We recently developed the self-management system using the HF points and instructions to visit hospitals or clinics when the points exceed the pre-specified levels. We found that the self-management system decreased the hospitalization for HF with an increase in unplanned visits and early intervention in the outpatient department. However, it is unclear whether we managed severe HF outpatients who should have been hospitalized. In this study, we aimed to compare HF severity in rehospitalized patients with regard to self-management system use. RESULTS: We retrospectively enrolled 306 patients (153 patients each in the system user and non-user groups) using propensity scores (PS). We compared HF severity and length of readmission in rehospitalized patients in both groups. During the 1-year follow-up period, 24 system users and 43 non-system users in the PS-matched cohort were hospitalized. There were no significant differences between the groups in terms of brain natriuretic peptide levels at readmission, maximum daily intravenous furosemide dose, percentage of patients requiring intravenous inotropes, duration of hospital stay and in-hospital mortality. These results suggest that the HF severity in rehospitalized patients was not different between the two groups.

Reproducibility and research integrity are essential tenets of every scientific study and discovery. They serve as proof that an established and documented work can be verified, repeated, and reproduced. New knowledge in the biomedical science is built on the shoulders of established and proven principles. Thus, scientists must be able to trust and build on the knowledge of their colleagues. Scientific innovation and research discoveries especially in the field of medicine has contributed to improving the lives of patients and increasing life expectancies. However, the growing concerns of failure to comply with good scientific principles has resulted in issues with research integrity and reproducibility. Poor reproducibility and integrity, therefore, may lead to ineffective interventions and applications. Here we comment on research reproducibility in basic medical and life sciences with regards to issues arising and outline the role of stakeholders such as research institutions and their employees in addressing this crisis.

OBJECTIVE: We observed that the addition of glucose enhanced the expression of sigX and sigM, encoding extra-cytoplasmic function sigma factors in Bacillus subtilis. Several regulatory factors were identified for this phenomenon, including YqfO, CshA (RNA helicase), and YlxR (nucleoid-associated protein). Subsequently, the relationships among these regulators were analyzed. Among them, YqfO is conserved in many bacterial genomes and may function as a metal ion insertase or metal chaperone, but has been poorly characterized. Thus, to further characterize YqfO, we performed RNA sequencing (RNA-seq) analysis of YqfO in addition to CshA and YlxR. RESULTS: We first performed comparative RNA-seq to detect the glucose-responsive genes. Next, to determine the regulatory effects of YqfO in addition to CshA and YlxR, three pairs of comparative RNA-seq analyses were performed (yqfO/wt, cshA/wt, and ylxR/wt). We observed relatively large regulons (approximately 420, 780, and 180 for YqfO, CshA, and YlxR, respectively) and significant overlaps, indicating close relationships among the three regulators. This study is the first to reveal that YqfO functions as a global regulator in B. subtilis.

OBJECTIVES: This study aimed to explore mechanism of colistin resistance amongst Klebsiella pneumoniae isolates through plasmid mediated mcr-1 gene in Pakistan. Carbapenem and Colistin resistant K. pneumoniae isolates (n  = 34) stored at - 80 °C as part of the Aga Khan University Clinical Laboratory strain bank were randomly selected and subjected to mcr-1 gene PCR. To investigate mechanisms of resistance, other than plasmid mediated mcr-1 gene, whole genome sequencing was performed on 8 clinical isolates, including 6 with colistin resistance (MIC  >  4 μg/ml) and 2 with intermediate resistance to colistin (MIC  >  2 μg/ml). RESULTS: RT-PCR conducted revealed absence of mcr-1 gene in all isolates tested. Whole genome sequencing results revealed modifications in Lipid A-Ara4N pathway. Modifications in Lipid A-Ara4N pathway were detected in ArnA_ DH/FT, UgdH, ArnC and ArnT genes. Mutation in ArnA_ DH/FT gene were detected in S3, S5, S6 and S7 isolates. UgdH gene modifications were found in all isolates except S3, mutations in ArnC were present in all except S1, S2 and S8 and ArnT were detected in all except S4 and S7. In the absence of known mutations linked with colistin resistance, lipid pathway modifications may possibly explain the phenotype resistance to colistin, but this needs further exploration.

OBJECTIVE: The mango tree Mangifera indica is known as one of the botanical sources of propolis in Tropical regions. There are two different materials which bees can collect from a mango tree to produce propolis: the resin of the tree bark, and the latex found on the fruits. We performed the study of the chemical profile of mango resin in comparison with propolis in order to clarify its importance as propolis source. RESULTS: We compared the chemical profiles (by GC-MS analysis of ethanol extracts after silylation) of the resin and samples of propolis: of stingless bees (3 Vietnames, 2 Indonesian), and one of Apis mellifera from Thailand. In the resin and all propolis samples, 25 compounds were identified: fatty acids, cardanols (alk(en)yl phenols), cardols, anacardic acids, triterpene alcohols and ketones, cycloartane type triterpenic acids. All samples have the same qualitative composition but there are important quantitative differences. Considering literature data on mango latex, we conclude that bees of different species, make use of the two propolis sources offered by mango: bark resin and fruit latex, in different proportions. We also confirmed for the first time the presence of alk(en)yl phenols and anacardic acids in the tree bark resin of mango.

OBJECTIVE: Interferon-gamma (IFN-γ) is overexpressed in rheumatoid synovium and thought to be involved in the pathogenesis of rheumatoid arthritis (RA). In this study, we examined our hypothesis that IFN-γ activates innate immune cells and upregulates inflammatory cytokines. Peripheral blood neutrophils were stimulated with IFN-γ in the presence or absence of Janus kinase (JAK) inhibitors. Interleukin-6 (IL-6) mRNA and protein expression were analyzed using real-time polymerase chain reaction (PCR) method and enzyme-linked immunosorbent assay. Protein phosphorylation of JAKs or STAT1 was assessed by Western blot using phospho-specific antibodies. RESULTS: IFN-γ stimulation induces IL-6 expression in protein and mRNA levels in human neutrophils. Furthermore, IFN-γ stimulation induces JAK1/JAK2 phosphorylation and downstream signal transducer and activator of transcription (STAT) 1 phosphorylation in human neutrophils. Although all JAKi, blocked IFN-γ-induced JAK1.2/STAT1 phosphorylation at higher concentrations (100 nM), baricitinib most efficiently inhibited IFN-γ-induced JAK1.2/STAT1 phosphorylation at lower concentrations (≤ 25 nM). Among these JAKi, baricitinib was the most potent regulator for IFN-γ-induced IL-6 production in human neutrophils. Our data indicate that IFN-γ upregulates IL-6 production via the JAK1/2-STAT1 pathway in human innate immune cells. Furthermore, this IFN-γ-mediated IL-6 induction via JAK/STAT was downregulated by JAKi.

OBJECTIVE: The main purpose of this paper is to present an accelerated nonstandard finite difference method for solving the singularly perturbed Burger-Huxley equation in order to produce more accurate solutions. RESULTS: The quasilinearization technique is used to linearize the nonlinear term. A nonstandard methodology of Mickens procedure is used in the spatial direction and also within the first order temporal direction that construct the first-order finite difference approximation to solve the considered problem numerically. To accelerate the rate of convergence from first to second-order, the Richardson extrapolation technique is applied. Numerical experiments were conducted to support the theoretical results.