Immune gene expression and epigenetic potential affect the consumption of risky food by female house sparrows.

When organisms move into new areas, they are likely to encounter novel food resources. Even if they are nutritious, these foods can also be risky, as they might be contaminated by parasites. The behavioural immune system of animals could help them avoid the negative effects of contaminated resources, but our understanding of behavioural immunity is limited, particularly whether and how behavioural immunity interacts with physiological immunity. Here, we asked about the potential for interplay between these two traits, specifically how the propensity of an individual house sparrow (Passer domesticus) to take foraging risks was related to its ability to regulate a key facet of its immune response to bacterial pathogens. Previously, we found that sparrows at expanding geographic range edges were more exploratory and less risk-averse to novel foods; in those same populations, birds tended to over-express Toll-like receptor 4 (TLR4), a pattern-recognition receptor that distinguishes cell-wall components of Gram-negative bacteria, making it the major sensor of potentially lethal gut microbial infections including salmonellosis. When we investigated how birds would respond to a typical diet (i.e., mixed seeds) spiked with domesticated chicken faeces, birds that expressed more TLR4 or had higher epigenetic potential for TLR4 (more CpG dinucleotides in the putative gene promoter) ate more food, spiked or not. Females expressing abundant TLR4 were also willing to take more foraging risks and ate more spiked food. In males, TLR4 expression was not associated with risk-taking. Altogether, our results indicate that behaviour and immunity covary among individual house sparrows, particularly in females where those birds that maintain more immune surveillance also are more disposed to take foraging risks.

Combined Gap-Polymerase Chain Reaction and Targeted Next-Generation Sequencing Improve α- and ß-Thalassemia Carrier Screening in Pregnant Women in Vietnam.

Vietnam has a high thalassemia burden. We collected blood samples from 5880 pregnant Vietnamese women during prenatal health checks to assess thalassemia carrier frequency using combined gap-polymerase chain reaction (gap-PCR) and targeted next-generation sequencing (NGS). Thalassemia carriers were identified with prevalence of 13.13% (772), including 7.82% (460) carriers of α-thalassemia (α-thal), 5.31% (312) carriers of ß-thalassemia (ß-thal), and 0.63% (37) concurrent α-/ß-thal carriers. Deletional mutations (368) accounted for 80.0% of α-thal carriers, of which, --SEA (Southeast Asian) (n = 254; 55.0%) was most prevalent, followed by the -α3.7 (rightward) (n = 66; 14.0%) and -α4.2 (leftward) (n = 45; 9.8%) deletions. Hb Westmead (HBA2: c.369C>G) (n = 53) and Hb Constant Spring (Hb CS or HBA2: c.427T>C) (in 28) are the two most common nondeletional α-globin variants, accounting for 11.5 and 6.0% of α-thal carriers. We detected 11 different ß-thal genotypes. Hb E (HBB: c.79G>A) (in 211) accounted for 67.6% of ß-thal carriers. The most common ß-thal genotypes were associated with mutations at codon 17 (A>T) (HBB: c.52A>T), codons 41/42 (-TTCT) (HBB: c.126_129delCTTT), and codon 71/72 (+A) (HBB: c.217_218insA) (prevalence 0.70%, 0.68%, and 0.2%, respectively). Based on mutation frequencies calculated in this study, estimates of 5021 babies in Vietnam are affected with clinically severe thalassemia annually. Our data suggest a higher thalassemia carrier frequency in Vietnam than previously reported. We established that combining NGS with gap-PCR creates an effective large-scale thalassemia screening method that can detect a broad range of mutations.

Determinants of aflatoxin exposures in Kenyan School-aged children.

Aflatoxins are naturally occurring food toxins known to contaminate cereals with a carry-over effect in milk and meat products from farm animals raised on contaminated feed. In children, continuous consumption of aflatoxin-contaminated food is linked to immune suppression, vaccine interference and growth faltering while in adult populations, carcinogenesis in the liver has been established. We evaluate the main determinants of aflatoxin exposures among children recruited from primary schools in Makueni and Siaya Counties. A five-part questionnaire was administered to collect information from randomly selected participants. AflatoxinB1-lysine adducts in children's sera and total aflatoxins in food samples were analyzed by High-Performance Liquid Chromatography with Fluorescence detection. Using Chi-squared tests and Kruskal-Wallis tests, children from low-income households had the highest aflatoxin exposure, p-value = 0.0029. Smaller family size, greater food diversity, and good farming practices were associated with low aflatoxin exposures p < 0.001. Individual households living under severe levels of poverty were evidently exposed to higher levels of aflatoxins.

Excess Risk of Tuberculosis Infection Among Extra-household Contacts of Tuberculosis Cases in an African City.

BACKGROUND: Although households of tuberculosis (TB) cases represent a setting for intense transmission of Mycobacterium tuberculosis, household exposure accounts for <20% of transmission within a community. The aim of this study was to estimate excess risk of M. tuberculosis infection among household and extra-household contacts of index cases. METHODS: We performed a cross-sectional study in Kampala, Uganda, to delineate social networks of TB cases and matched controls without TB. We estimated the age-stratified prevalence difference of TB infection between case and control networks, partitioned as household and extra-household contacts. RESULTS: We enrolled 123 index cases, 124 index controls, and 2415 first-degree network contacts. The prevalence of infection was highest among household contacts of cases (61.5%), lowest among household contacts of controls (25.2%), and intermediary among extra-household TB contacts (44.9%) and extra-household control contacts (41.2%). The age-adjusted prevalence difference between extra-household contacts of cases and their controls was 5.4%. The prevalence of infection was similar among the majority of extra-household case contacts and corresponding controls (47%). CONCLUSIONS: Most first-degree social network members of TB cases do not have adequate contact with the index case to experience additional risk for infection, but appear instead to acquire infection through unrecognized exposures with infectious cases in the community.

Identification of potent inhibitors of the sortilin-progranulin interaction.

High-throughput screening methods have been used to identify two novel series of inhibitors that disrupt progranulin binding to sortilin. Exploration of structure-activity relationships (SAR) resulted in compounds with sufficient potency and physicochemical properties to enable co-crystallization with sortilin. These co-crystal structures supported observed SAR trends and provided guidance for additional avenues for designing compounds with additional interactions within the binding site.

Influenza Vaccine Efficacy and Effectiveness in Pregnant Women: Systematic Review and Meta-analysis.

OBJECTIVE: Pregnant women have a high risk for complications from influenza infection, but vaccination rates within this group remain low in the US and other countries. The efficacy and effectiveness of the influenza vaccine are a key determinant of vaccine uptake. This review aimed to synthesize the available evidence on the protection of both seasonal and monovalent pandemic H1N1 (pdmH1N1) vaccine against laboratory-confirmed influenza (LCI), influenza-like illness (ILI), and respiratory illness (RI). METHODS: A search of the literature was undertaken from Pubmed, Embase, ClinicalTrials.gov, and Cochrane Central Register of Controlled Trials up to Aug 1, 2018. Both observational studies and clinical trials were considered. RESULTS: Nineteen studies were identified from 11 countries. Women with pdmH1N1 vaccination had a lower risk of getting LCI (Relative risk [RR] 0.3, 95% confidence interval [CI] 0.26-0.35) and ILI (RR 0.15, 95% CI 0.06-0.36)). The pooled estimate from three randomized clinical trials (RR 0.47, 95% CI 0.31-0.71) and two case control studies (OR 0.37, 95% CI 0.23-0.61) showed that the seasonal vaccine was protective against LCI. The seasonal vaccine was not protective against ILI (RR 0.95, 95% CI 0.88-1.03). This association was similar for the outcome of RI (RR 0.81, 95% CI 0.55-1.20). CONCLUSION: This analysis bolsters existing evidence that influenza vaccines are effective among pregnant women. Additional public health efforts are needed to promote physician recommendations of influenza vaccination in pregnancy.

Feasibility, Reliability, and Validity of the Vietnamese Version of the Clinical Dementia Rating.

INTRODUCTION: In Vietnam, Alzheimer's disease (AD) and other dementias have become an increasingly important public health problem among the elderly. Achieving a diagnosis tool with high reliability and validity is essential. The Clinical Dementia Rating (CDR) is a global clinical scale with established diagnostic and severity-ranking utility that has been widely employed in epidemiological studies in an international context. OBJECTIVE: The aims of this study were to establish the Vietnamese version of the CDR (V-CDR) and evaluate the feasibility, reliability, and validity of this version for diagnosing and classifying cognitive functions in the elderly. METHOD: One hundred and fifty-three elderly outpatients at a clinic of Cho Ray Hospital, Vietnam, were screened with the Mini Mental State Examination (MMSE) for potential cognitive impairment. All those who scored ≤26 points were included in the study and were subsequently remitted to the V-CDR and clinical assessment for diagnosis. Reliability was assessed through internal consistency (Cronbach α), intra- and interrater reliability (weighted κ). Concurrent and discriminative validity of the V-CDR were assessed. RESULTS: The V-CDR had an excellent internal consistency for each of the 2 raters (Cronbach α 0.90 and 0.96) and excellent agreement in both intra- and interrater reliability (weighted κ 0.84 [95% CI 0.74-0.94] and 0.82 [95% CI 0.72-0.93], respectively). The sensitivity and specificity for detecting dementia were 93.6 and 100%, respectively. The positive and negative predictive value were 100 and 96.4%, respectively. The agreement of V-CDR and clinical assessment was excellent (weighted κ 0.94 [95% CI 0.88-0.99]). V-CDR was substantially better than MMSE at distinguishing between mild cognitive impairment and normal cognitive function (AUC = 0.957, 95% CI 0.893-1.000 vs. AUC 0.594, 95% CI 0.441-0.746). CONCLUSIONS: The V-CDR is a feasible, reliable, and valid instrument which should be used in clinical practice for diagnosing and classifying the different dementia stages in the elderly.

Analysis of the evolution of the detection limits of electrochemical nucleic acid biosensors II.

This critical review of electrochemical biosensors allowing direct detection of nucleic acid targets reports on different transduction pathways and their latest breakthroughs. A classification of the various strategies based on the nature of the electrochemical transduction is established to emphasize the efficiency of each of them. It provides an overall picture of the detection limit of the various approaches developed during the last two decades. Graphical Abstract Detection limits evolutions of electrochemical DNA biosensors.

Electrochemical Impedance Spectroscopy in a Droplet of Solution for the Investigation of Liquid/Solid Interface.

The local electrochemical behavior of a solid-liquid interface can be studied by electrochemical impedance spectroscopy (EIS). The investigated surface area can be delimited by adding a drop of solution, which forms an interface between the liquid drop and the working electrode, and performing the measurements inside. The size of the drop must be sufficiently small for a simultaneous wettability characterization (from the contact angle measurement) and appropriately large so that wettability is not influenced by the presence of the working and the counter electrode inserted in the droplet. In this work, we showed that EIS measurements can be performed in a solution droplet of 2 to 4 µL, although the electrochemical cell lacks the usual geometry. For our measurements, we studied a model system consisting of a KCl aqueous solution of [Fe(CN)6]3-/4- redox couple at a Pt electrode. All the results were compared with those obtained for a bulk configuration. The sessile drop configuration and the EIS response were modeled using finite element method for different electrode sizes and configurations to account for electrochemical kinetics and both current and potential distributions.

The Electrochemical Behavior of TiN/316LSS Material in Simulated Body Fluid Solution.

We report on the fabrication and the electrochemical behavior of TiN film on the 316L stainless steel (316LSS) material in simulated body fluid (SBF) solution for implant application. The characterization results indicate that the coated TiN is completely crystalline with (111) crystal orientation. Electrochemical results of 316LSS and TiN/316LSS material after 21 days of immersion in SBF show that the durability of the TiN/316LSS is much higher than that of 316LSS, which registers a very low corrosion current density (about tens of nA cm(-2)). The formation of hydroxyapatite on the surface of the TiN/316LSS is also confirmed by SEM, EDX, X-ray and IR spectroscopy.

Electrodeposition and Characterization of Hydroxyapatite on TiN/316LSS.

The deposition of TiN on stainless steel substrates may improve the stability and compatibility of this material with bone, which may be advantageously exploited for the elaboration of advanced pros- thetic devices. In this work, TiN-coated 316LSS (by way of DC magnetron sputtering) was used as a starting material for investigating the electrochemical post-deposition of hydroxyapatite (HAp) which has a composition close to that of bone. Electrodeposition was carried out starting from an aqueous medium containing solubilized Ca(NO3)2 and NH4H2PO4 in the presence of H2O2. We report the influence of experimental conditions on the morphology of the obtained HAp coating on TiN/316LSS. The effect of applied potential, temperature, H2O2 concentration, pH and duration of reaction were thoroughly discussed on the basis of X-ray diffraction (XRD), scanning electron microscopy (SEM), Fourier Transform Infrared (FTIR) spectroscopy and Energy Dispersive X-ray Spectroscopy (EDX) results. This method appears advantageous for producing HAp-coated implant materials.

Sensitive and selective amplification of methylated DNA sequences using helper-dependent chain reaction in combination with a methylation-dependent restriction enzymes.

We have developed a novel technique for specific amplification of rare methylated DNA fragments in a high background of unmethylated sequences that avoids the need of bisulphite conversion. The methylation-dependent restriction enzyme GlaI is used to selectively cut methylated DNA. Then targeted fragments are tagged using specially designed 'helper' oligonucleotides that are also used to maintain selection in subsequent amplification cycles in a process called 'helper-dependent chain reaction'. The process uses disabled primers called 'drivers' that can only prime on each cycle if the helpers recognize specific sequences within the target amplicon. In this way, selection for the sequence of interest is maintained throughout the amplification, preventing amplification of unwanted sequences. Here we show how the method can be applied to methylated Septin 9, a promising biomarker for early diagnosis of colorectal cancer. The GlaI digestion and subsequent amplification can all be done in a single tube. A detection sensitivity of 0.1% methylated DNA in a background of unmethylated DNA was achieved, which was similar to the well-established Heavy Methyl method that requires bisulphite-treated DNA.

A panel of genes methylated with high frequency in colorectal cancer.

BACKGROUND: The development of colorectal cancer (CRC) is accompanied by extensive epigenetic changes, including frequent regional hypermethylation particularly of gene promoter regions. Specific genes, including SEPT9, VIM1 and TMEFF2 become methylated in a high fraction of cancers and diagnostic assays for detection of cancer-derived methylated DNA sequences in blood and/or fecal samples are being developed. There is considerable potential for the development of new DNA methylation biomarkers or panels to improve the sensitivity and specificity of current cancer detection tests. METHODS: Combined epigenomic methods - activation of gene expression in CRC cell lines following DNA demethylating treatment, and two novel methods of genome-wide methylation assessment - were used to identify candidate genes methylated in a high fraction of CRCs. Multiplexed amplicon sequencing of PCR products from bisulfite-treated DNA of matched CRC and non-neoplastic tissue as well as healthy donor peripheral blood was performed using Roche 454 sequencing. Levels of DNA methylation in colorectal tissues and blood were determined by quantitative methylation specific PCR (qMSP). RESULTS: Combined analyses identified 42 candidate genes for evaluation as DNA methylation biomarkers. DNA methylation profiles of 24 of these genes were characterised by multiplexed bisulfite-sequencing in ten matched tumor/normal tissue samples; differential methylation in CRC was confirmed for 23 of these genes. qMSP assays were developed for 32 genes, including 15 of the sequenced genes, and used to quantify methylation in tumor, adenoma and non-neoplastic colorectal tissue and from healthy donor peripheral blood. 24 of the 32 genes were methylated in >50% of neoplastic samples, including 11 genes that were methylated in 80% or more CRCs and a similar fraction of adenomas. CONCLUSIONS: This study has characterised a panel of 23 genes that show elevated DNA methylation in >50% of CRC tissue relative to non-neoplastic tissue. Six of these genes (SOX21, SLC6A15, NPY, GRASP, ST8SIA1 and ZSCAN18) show very low methylation in non-neoplastic colorectal tissue and are candidate biomarkers for stool-based assays, while 11 genes (BCAT1, COL4A2, DLX5, FGF5, FOXF1, FOXI2, GRASP, IKZF1, IRF4, SDC2 and SOX21) have very low methylation in peripheral blood DNA and are suitable for further evaluation as blood-based diagnostic markers.

M. tuberculosis Infection Attributable to Exposure in Social Networks of Tuberculosis Cases in an Urban African Community.

Background: The persistence of tuberculosis today and its global disparity send a powerful message that effective tuberculosis control must respond to its regional epidemiology. Active case finding through contact investigation is a standard protocol used for tuberculosis control, but its effectiveness has not been established, especially in endemic areas. Methods: To quantify the potential effectiveness of contact investigation in Kampala, Uganda, we used a cross-sectional design to evaluate the social networks of 123 tuberculosis index cases and 124 controls without tuberculosis. Results: Tuberculous infection was present in 515 of 989 tuberculosis case contacts (52.1%) and 396 of 1026 control contacts (38.6%; adjusted prevalence ratio, 1.4; 95% CI, 1.3-1.6). The proportion of infected participants with known exposure within the social network of the tuberculosis case was 35%. The population-attributable fraction was 11.1% for any known exposure, with 7.3% attributable to household exposure and 3.4% attributable to extrahousehold exposure. Conclusions: This low population-attributable fraction indicates that contact tracing in the social networks of index cases will have only a modest effect in reducing tuberculous infection in a community. New approaches to community-level active case finding are needed.

Reactogenicity and immunogenicity of heterologous prime-boost immunization with COVID-19 vaccine.

BACKGROUND: The objective of the present work was to assess the reactogenicity and immunogenicity of heterologous COVID-19 vaccination regimens in clinical trials and observational studies. METHODS: PubMed, Cochrane Library, Embase, MedRxiv, BioRxiv databases were searched in September 29, 2021. The PRISMA instruction for systemic review was followed. Two reviewers independently selected the studies, extracted the data and assessed risk of bias. The quality of studies was evaluated using the New Castle-Ottawa and Cochrane risk of instrument. The characteristics and study outcome (e.g., adverse events, immune response, and variant of concern) were extracted. RESULTS: Nineteen studies were included in the final data synthesis with 5 clinical trials and 14 observational studies. Heterologous vaccine administration showed a trend toward more frequent systemic reactions. However, the total reactogenicity was tolerable and manageable. Importantly, the heterologous prime-boost vaccination regimens provided higher immunogenic effect either vector/ mRNA-based vaccine or vector/ inactivated vaccine in both humoral and cellular immune response. Notably, the heterologous regimens induced the potential protection against the variant of concern, even to the Delta variant. CONCLUSIONS: The current findings provided evidence about the higher induction of robust immunogenicity and tolerated reactogenicity of heterologous vaccination regimens (vector-based/mRNA vaccine or vector-based/inactivated vaccine). Also, this study supports the application of heterologous regimens against COVID-19 which may provide more opportunities to speed up the global vaccination campaign and maximize the capacity to control the pandemic.

Discovery of MK-1454: A Potent Cyclic Dinucleotide Stimulator of Interferon Genes Agonist for the Treatment of Cancer.

Stereochemically and structurally complex cyclic dinucleotide-based stimulator of interferon genes (STING) agonists were designed and synthesized to access a previously unexplored chemical space. The assessment of biochemical affinity and cellular potency, along with computational, structural, and biophysical characterization, was applied to influence the design and optimization of novel STING agonists, resulting in the discovery of MK-1454 as a molecule with appropriate properties for clinical development. When administered intratumorally to immune-competent mice-bearing syngeneic tumors, MK-1454 exhibited robust tumor cytokine upregulation and effective antitumor activity. Tumor shrinkage in mouse models that are intrinsically resistant to single-agent therapy was further enhanced when treating the animals with MK-1454 in combination with a fully murinized antimouse PD-1 antibody, mDX400. These data support the development of STING agonists in combination with pembrolizumab (humanized anti-PD-1 antibody) for patients with tumors that are partially responsive or nonresponsive to single-agent anti-PD-1 therapy.

Nutrition and growth outcomes are affected by aflatoxin exposures in Kenyan children.

Aflatoxin exposure, malnutrition and growth impairment in children present significant public health problems in low- and middle-income countries. Recent epidemiology studies show that exposure to aflatoxins through dietary sources in early life contributes to growth retardation among children. However, the findings remain inconclusive due to limited comparative studies in high versus low aflatoxin exposure regions. This cross-sectional study presents aflatoxin exposure levels among children aged 6 to 12 years, and further evaluates the association between aflatoxin exposure levels, malnutrition and growth impairment in Kenya, East Africa. AFB1-lysine adducts are validated biomarkers of exposure and were quantified using HPLC with fluorescence detection. All children (n = 746) had detectable levels of AFB1-lysine adducts in serum, range 0.65-518.9 pg/mg albumin with a geometric mean (GM) of 10.5 (95%CI 9.4-11.7) pg/mg albumin. The Geometric Means (GM) of AFB1-lysine adducts were 14.0 (95%CI 12.5, 15.7) pg/mg albumin and 8.2 (95%CI 7.6, 8.8) pg/mg albumin (p-value < 0.001), among children recruited from Makueni and Siaya Counties, respectively. While the study confirms higher human exposure levels in Makueni county, it provides an initial data set for aflatoxin exposure levels among children recruited from Siaya County. In multivariate analysis, after adjusting for socio-economic indicators, farming practices, and household dietary patterns, increasing one unit of log AFB1-lysine was associated with decreasing Weight-for-age z-score (WAZ) by -0.13, p-value = 0.019 among all children aged 6-12 years. Among children 6 to 9 years, WAZ decreases by -0.11 (-0.54, -0.01), p-value = 0.049. Additional growth parameters Height-for-age z-score (HAZ) and Weight-for-height z-score (WHZ) do not reach statistical significance. HAZ decreases by -0.08, p-value = 0.337 and WHZ decreases by -0.17, p-value = 0.437 with every increase in log AFB1-lysine. These data suggest that efforts must be put in place to control for aflatoxin exposure in order to achieve better growth outcomes.

Discovery of a Novel cGAMP Competitive Ligand of the Inactive Form of STING.

Drugging large protein pockets is a challenge due to the need for higher molecular weight ligands, which generally possess undesirable physicochemical properties. In this communication, we highlight a strategy leveraging small molecule active site dimers to inhibit the large symmetric binding pocket in the STING protein. By taking advantage of the 2:1 binding stoichiometry, maximal buried interaction with STING protein can be achieved while maintaining the ligand physicochemical properties necessary for oral exposure. This mode of binding requires unique considerations for potency optimization including simultaneous optimization of protein-ligand as well as ligand-ligand interactions. Successful implementation of this strategy led to the identification of 18, which exhibits good oral exposure, slow binding kinetics, and functional inhibition of STING-mediated cytokine release.

Helper-Dependent Chain Reaction (HDCR) for Selective Amplification of Methylated DNA Sequences.

Over the last few years a number of restriction enzymes that cut DNA only if cytosines within their recognition sequences are methylated have been characterized and become commercially available. Cleavage with these enzymes to release DNA fragments in a methylation-dependent manner can be combined with a novel method of amplification, Helper Dependent Chain Reaction (HDCR), to selectively amplify these fragments. HDCR uses "Helper" oligonucleotides as templates for extension of the free 3' end of target fragments to incorporate tag sequences at the ends of fragments. These tag sequences are then used for priming of amplification of target fragments. Modifications to the amplification primers (Drivers) and the Helpers ensure that there is selection for the sequences within target fragments with each cycle of amplification. The combination of methylation-dependent enzymes and HDCR allows the sensitive and selective amplification of methylated DNA sequences without the need for bisulfite treatment.

PIM3 proto-oncogene kinase is a common transcriptional target of divergent EWS/ETS oncoproteins.

Despite significant structural diversity, present evidence suggests that EWS/ETS fusion proteins promote oncogenesis by transcriptionally modulating a common set of target genes. In order to identify these genes, microarray expression analyses were performed on NIH 3T3 polyclonal populations expressing one of three EWS/ETS fusion genes. The majority of these genes can be grouped into seven functional categories, including cellular metabolism and signal transduction. The biologic significance of these target genes was pursued. The effects of modulating genes involved in metabolism were assessed by flux studies and demonstrated shifts in glucose utilization and lactate production as a result of EWS/FLI1 expression. The proto-oncogene coding for serine/threonine kinase PIM3 was found to one of several genes encoding signal transduction proteins that were up-regulated by EWS/ETS fusions. PIM3 was found to be expressed in a panel of human Ewing's family tumor cell lines. Forced expression of PIM3 promoted anchorage-independent growth. Coexpression of a kinase-deficient PIM3 mutant attenuated EWS/FLI1-mediated NIH 3T3 tumorigenesis in immunodeficent mice.