Blastocystis occurrence and subtype diversity in European wild boar (Sus scrofa) from the Iberian Peninsula.

The ongoing increase in wild boar populations across Europe has fostered human-wildlife conflicts, including the transmission of emerging pathogens with zoonotic importance. Blastocystis is a ubiquitous, faecal-oral transmitted protist that can cause gastrointestinal illnesses and is observed in humans and animals worldwide. The role of wildlife in the epidemiology of Blastocystis is insufficiently understood. Thus, we investigated the occurrence and subtype diversity of Blastocystis in free-ranging wild boars from the Iberian Peninsula using conventional PCR and next-generation amplicon sequencing of a fragment of the ssu RNA gene. A total of 459 wild boar faecal samples were collected across Spain (n = 360) and Portugal (n = 99) between 2014 and 2021. Blastocystis was present in 15.3% (70/459; 95% CI 12.1-18.9) of the wild boars analysed, and its occurrence was significantly higher in Portugal (34.3%, 34/99; 95% CI 25.1-44.6) than in Spain (10.0%, 36/360; 95% CI 7.1-13.6). Seven Blastocystis subtypes (ST5, ST10b, ST13-ST15, ST24b, and ST43) were detected among the surveyed wild boar populations, with greater variability detected in Portuguese samples. ST5 was identified in all the Blastocystis-positive animals, whereas 14.3% of them harboured ST mixed colonisations. Our results demonstrate that Blastocystis ST5 is particularly adapted to infect wild boars. The additional identification of zoonotic STs reinforces the role of wild boars as spreaders of zoonotic infections with public health significance.

DNA methylation governs the sensitivity of repeats to restriction by the HUSH-MORC2 corepressor.

The human silencing hub (HUSH) complex binds to transcripts of LINE-1 retrotransposons (L1s) and other genomic repeats, recruiting MORC2 and other effectors to remodel chromatin. How HUSH and MORC2 operate alongside DNA methylation, a central epigenetic regulator of repeat transcription, remains largely unknown. Here we interrogate this relationship in human neural progenitor cells (hNPCs), a somatic model of brain development that tolerates removal of DNA methyltransferase DNMT1. Upon loss of MORC2 or HUSH subunit TASOR in hNPCs, L1s remain silenced by robust promoter methylation. However, genome demethylation and activation of evolutionarily-young L1s attracts MORC2 binding, and simultaneous depletion of DNMT1 and MORC2 causes massive accumulation of L1 transcripts. We identify the same mechanistic hierarchy at pericentromeric α-satellites and clustered protocadherin genes, repetitive elements important for chromosome structure and neurodevelopment respectively. Our data delineate the epigenetic control of repeats in somatic cells, with implications for understanding the vital functions of HUSH-MORC2 in hypomethylated contexts throughout human development.

Establishing the Performance of Next-Generation Amplicon Sequencing for Detection of Giardia duodenalis in Ready-to-Eat Packaged Leafy Greens.

Giardia duodenalis is a globally distributed intestinal parasite that commonly infects both humans and animals. G. duodenalis is a species complex, which includes eight assemblages that vary both in genetic structure and host specificity. The prevalence of mixed-assemblage G. duodenalis cysts on food, an understudied infection route for G. duodenalis, remains unknown. In the present study, a method able to detect G. duodenalis mixed-assemblage infections using next-generation amplicon sequencing (NGS) of the beta-giardin gene was applied in combination with the US-FDA's BAM Chapter 19b protocol for the detection of G. duodenalis from fresh produce to ascertain the limit of detection of G. duodenalis on leafy greens. Ready-to-eat baby Romaine lettuce was inoculated with 5 (n = 5), 20 (n = 10), 100 (n = 10), 200 (n = 10), or 1,000 (n = 10) G. duodenalis cysts of the assemblage B strain H3. Detection of G. duodenalis was successful in 100% of the samples seeded with 1,000, 200, and 100 cysts, in 50% of the samples seeded with 20 cysts, and in none of the samples seeded with 5 cysts. We thus demonstrate robust detection of G. duodenalis on packaged leafy greens using the BAM Chapter 19B method coupled with assemblage-sensitive NGS. This protocol provides a new diagnostic tool useful for both prevalence studies and outbreak investigations involving fresh produce that may assist in better describing the role of G. duodenalis in foodborne illness and in protecting consumers from contaminated fresh produce.

A cross-sectional survey of Blastocystis sp. and Dientamoeba fragilis in non-human primates and their caregivers in Czech zoos.

Intestinal protists in the gut microbiome are increasingly studied, but their basic epidemiology is not well understood. We explored the prevalence, genetic diversity, and potential zoonotic transmission of two protists colonizing the large intestine - Blastocystis sp. and Dientamoeba fragilis - in 37 species of non-human primates (NHPs) and their caregivers in six zoos in the Czech Republic. We analyzed 179 fecal samples (159 from NHPs, 20 from humans) by qPCR. Blastocystis sp. was detected in 54.7% (98/179) of samples, in 24 NHP species and in 57.2% of NHP samples (prevalence ranged between 36 and 80%), and in 35% of human samples (prevalence ranged between 0 and 67%). Using next generation amplicon sequencing, nine Blastocystis subtypes (ST1-ST5, ST7, ST8, and two novel subtypes) were identified. The two new Blastocystis subtypes (named ST47 and ST48) were described using Nanopore sequencing to produce full-length reference sequences of the small subunit ribosomal RNA gene. Some subtypes were shared between NHPs and their caregivers, suggesting potential zoonotic transmission. Mixed subtype colonization was frequently observed, with 52% of sequenced samples containing two or more subtypes. Dientamoeba was found only in NHPs with a prevalence of 6%. This study emphasizes the critical role of molecular diagnostics in epidemiological and transmission studies of these protists and calls for further research to better understand their impact on public health.

Sarcopenia as a Predictive Factor for Carboplatin Toxicity in Patients with Advanced Non-Small Cell Lung Cancer.

Sarcopenia in cancer patients often negatively impacts various outcomes. Carboplatin, a first-line chemotherapy for non-small cell lung cancer (NSCLC), is dosed based on body weight, which doesn't account for sarcopenia. This study evaluated the association between sarcopenia and carboplatin-related toxicity in NSCLC patients. Patients with locally advanced or metastatic NSCLC treated with carboplatin were included. Toxicity events during the first two cycles of treatment were recorded. Sarcopenia was assessed using pretreatment computed tomography scans analyzed with Slice-O-Matic V4.2 software, defining sarcopenia as a skeletal muscle index (SMI) of <52.4 cm2/m2 for men and <38.5 cm2/m2 for women. Among 146 patients, 52% had sarcopenia. Hematological toxicity occurred in 71.2% of all patients and 77.6% of those with sarcopenia. The fat-free mass index (FFMI) was independently associated with hematological toxicity and dose-limiting toxicity (DLT), which was observed in 55.5% of patients. Sarcopenia significantly correlates with hematological toxicity and DLT during carboplatin treatment in NSCLC patients. Given its prevalence and noninvasive detection, further research is needed to understand its impact on treatment outcomes.

IgG, IgM, and Nonstructural Protein 1 Response Profiles after Receipt of Tetravalent Dengue Vaccine TAK-003 in a Phase 2 Randomized Controlled Trial.

The profiles of vaccine-induced dengue antibodies may differ from those produced following natural infection and could potentially interfere with the interpretation of diagnostic tests. We assessed anti-dengue IgG and IgM antibodies, and nonstructural protein 1 antigen profiles in the serum of adults who received a single dose of the tetravalent dengue vaccine TAK-003 as either an initially developed high-dose formulation or the standard approved formulation in a phase 2 study in Singapore (#NCT02425098). Immunoglobulin G and IgM profiles during the first 30 days postvaccination varied by baseline serostatus (microneutralization assay). Nonstructural protein 1 antigen was not detected in the serum of any participants. Vaccine-induced IgG and IgM antibodies can affect serological confirmation of subsequent dengue infection in vaccinees. These results highlight the limitations of using serological tests for dengue diagnosis, particularly in a postvaccination setting, and emphasize the need for more sensitive antigen- and molecular-based testing for accurate dengue diagnosis.

Electron transport chain capacity expands yellow fever vaccine immunogenicity.

Vaccination has successfully controlled several infectious diseases although better vaccines remain desirable. Host response to vaccination studies have identified correlates of vaccine immunogenicity that could be useful to guide development and selection of future vaccines. However, it remains unclear whether these findings represent mere statistical correlations or reflect functional associations with vaccine immunogenicity. Functional associations, rather than statistical correlates, would offer mechanistic insights into vaccine-induced adaptive immunity. Through a human experimental study to test the immunomodulatory properties of metformin, an anti-diabetic drug, we chanced upon a functional determinant of neutralizing antibodies. Although vaccine viremia is a known correlate of antibody response, we found that in healthy volunteers with no detectable or low yellow fever 17D viremia, metformin-treated volunteers elicited higher neutralizing antibody titers than placebo-treated volunteers. Transcriptional and metabolomic analyses collectively showed that a brief course of metformin, started 3 days prior to YF17D vaccination and stopped at 3 days after vaccination, expanded oxidative phosphorylation and protein translation capacities. These increased capacities directly correlated with YF17D neutralizing antibody titers, with reduced reactive oxygen species response compared to placebo-treated volunteers. Our findings thus demonstrate a functional association between cellular respiration and vaccine-induced humoral immunity and suggest potential approaches to enhancing vaccine immunogenicity.

Correlates of protection against symptomatic SARS-CoV-2 in vaccinated children.

The paucity of information on longevity of vaccine-induced immune responses and uncertainty of the correlates of protection hinder the development of evidence-based COVID-19 vaccination policies for new birth cohorts. Here, to address these knowledge gaps, we conducted a cohort study of healthy 5-12-year-olds vaccinated with BNT162b2. We serially measured binding and neutralizing antibody titers (nAbs), spike-specific memory B cell (MBC) and spike-reactive T cell responses over 1 year. We found that children mounted antibody, MBC and T cell responses after two doses of BNT162b2, with higher antibody and T cell responses than adults 6 months after vaccination. A booster (third) dose only improved antibody titers without impacting MBC and T cell responses. Among children with hybrid immunity, nAbs and T cell responses were highest in those infected after two vaccine doses. Binding IgG titers, MBC and T cell responses were predictive, with T cells being the most important predictor of protection against symptomatic infection before hybrid immunity; nAbs only correlated with protection after hybrid immunity. The stable MBC and T cell responses over time suggest sustained protection against symptomatic SARS-CoV-2 infection, even when nAbs wane. Booster vaccinations do not confer additional immunological protection to healthy children.

Catalytic, Spectroscopic, and Theoretical Studies of Fe4S4-Based Coordination Polymers as Heterogenous Coupled Proton-Electron Transfer Mediators for Electrocatalysis.

Iron-sulfur clusters play essential roles in biological systems, and thus synthetic [Fe4S4] clusters have been an area of active research. Recent studies have demonstrated that soluble [Fe4S4] clusters can serve as net H atom transfer mediators, improving the activity and selectivity of a homogeneous Mn CO2 reduction catalyst. Here, we demonstrate that incorporating these [Fe4S4] clusters into a coordination polymer enables heterogeneous H atom transfer from an electrode surface to a Mn complex dissolved in solution. A previously reported solution-processable Fe4S4-based coordination polymer was successfully deposited on the surfaces of different electrodes. The coated electrodes serve as H atom transfer mediators to a soluble Mn CO2 reduction catalyst displaying good product selectivity for formic acid. Furthermore, these electrodes are recyclable with a minimal decrease in activity after multiple catalytic cycles. The heterogenization of the mediator also enables the characterization of solution-phase and electrode surface species separately. Surface enhanced infrared absorption spectroscopy (SEIRAS) reveals spectroscopic signatures for an in situ generated active Mn-H species, providing a more complete mechanistic picture for this system. The active species, reaction mechanism, and the protonation sites on the [Fe4S4] clusters were further confirmed by density functional theory calculations. The observed H atom transfer reactivity of these coordination polymer-coated electrodes motivates additional applications of this composite material in reductive H atom transfer electrocatalysis.

Early Incorporation to Palliative Care (EPC) in Patients With Advanced Non-Small Cell Lung Cancer: The PACO Randomized Clinical Trial.

BACKGROUND: Patients with non-small cell lung cancer (NSCLC) experience a considerable disease burden, evident in symptomatic and psychological spheres. Advanced cancer represents a complex scenario for patients and the healthcare team. Early palliative care (EPC) has been proven as a clinically meaningful strategy in this context by several randomized trials but not in a resource-limited setting. This study aimed to evaluate the effect of EPC compared with standard oncological care (SOC) in patients with metastatic NSCLC in Mexico. MATERIALS AND METHODS: A prospective, randomized clinical trial was conducted at Instituto Nacional de Cancerologia in Mexico. All patients had histologically confirmed metastatic NSCLC without previous treatment. Patients were randomly assigned (1:1) to receive SOC or SOC + EPC. The EPC group was introduced to the palliative care team at baseline after randomization, which was integrated by psychologists, bachelor's in nutrition, specialized nurses, and physicians. Patients randomized to this arm had programmed visits to meet with the team at baseline and through the 2nd, 4th-, and 6th cycles thereafter. The primary endpoint was overall survival (OS); secondary outcomes included quality of life (QoL), anxiety and depression, and symptom intensity. They were assessed using the instruments EORTC QLQ-C30 questionnaire, Edmonton Symptom Assessment Scale (ESAS), and the Hospital Anxiety and Depression Scale (HADS) (clinicaltrials.gov [NCT01631565]). Questionnaires were completed at baseline, at 2nd, 4th, and 6th cycles of treatment. RESULTS: Between March 2012 and June 2015, 201 patients were assessed for eligibility and 146 were enrolled and allocated to receive EPC (73) or SOC (73). Median OS for patients in the EPC vs SOC arm was 18.1 months (95% CI, 7.9-28.4) and 10.5 months (95% CI, 4.7-16.2) (P = .029). Having a poor performance status (HR 1.7 [1.2-2.5]; P = .004) and allocation to the control group (HR 1.5 [1.03-2.3]; P = .034) were independently associated with a worse OS. Those patients with a global QoL > 70 at baseline had a better OS if they were In the EPC arm (38.7 months (95% CI, 9.9-67.6) vs SOC 21.4 months (95% CI, 12.4-30.3)). Mean QoL had a numerical improvement in patients allocated to EPC after 6 cycles of follow-up, nonetheless this difference was not statistically significant (55.1 ±â€…23.7 vs 56.9 ±â€…25.3; P = .753). There were no significant differences in anxiety and depression at all study points. CONCLUSIONS: EPC is associated with a significant improvement in OS, although, we observed that the greatest benefit of providing EPC was observed in those with a global QoL > 70 at baseline. This study did not identify significant changes in terms of QoL or symptom burden between the study groups after follow-up. Evidence robustly suggests that EPC should be considered part of the multidisciplinary treatment of metastatic NSCLC patients since diagnosis. According to our study, EPC can be implemented in low- or middle-income countries (LMIC).

Molecular detection and characterization of Blastocystis in herbivore livestock species in Portugal.

Blastocystis is a ubiquitous intestinal protist in humans and animals worldwide. The traditional livestock free-roaming raising system in rural communities increases the risk of infection with contact with a wider range of pathogens transmitted via the faecal-oral route associated with that wildlife-livestock-human interface. However, no studies have been conducted to determine the occurrence and subtype distribution of Blastocystis in livestock in Portugal. Here, we collected 180 faecal samples from herbivore livestock (cattle, goats, horses, and sheep) in different regions of the country to investigate Blastocystis prevalence and subtype diversity using PCR and next-generation amplicon sequencing. Blastocystis was present in 40.6% (73/180; 95% CI: 33.31-48.11) of the samples (goats, 81.0%; sheep, 60.9%; cattle, 32.2%). None of the horse samples were Blastocystis-positive. Eighteen subtypes were detected (ST1-ST3, ST5-ST7, ST10, ST13, ST14, ST21, ST23-ST26, ST30, ST42-ST44). Mixed infections were detected in 97.3% of the Blastocystis-positive samples. Potentially zoonotic subtypes were identified in 75.0%, 96.4%, and 100% of the Blastocystis-positive specimens collected from cattle, sheep, and goats, respectively. These results demonstrate that cattle, sheep, and goats harbour a high diversity of Blastocystis subtypes in the study regions. Importantly, our data provide novel molecular evidence strongly suggesting that some Blastocystis STs/ST subgroups may have differential host specificity.

Mirtazapine as Appetite Stimulant in Patients With Non-Small Cell Lung Cancer and Anorexia: A Randomized Clinical Trial.

Importance: Currently there is no standard therapy to improve cancer-related anorexia, hampering survival. Mirtazapine has been suggested as a feasible option in this context. Objectives: To assess the effect of mirtazapine on appetite and energy consumption in patients with advanced non-small cell lung cancer (NSCLC). Design, Setting, and Participants: This randomized, double-blind, placebo-controlled clinical trial including adults was performed in a tertiary cancer care center from August 2018 to May 2022 with a follow-up of 8 weeks. Overall, 134 patients were screened; 114 were assessed for eligibility and 28 were excluded. Interventions: Patients were randomized in a 1:1 ratio to receive mirtazapine, 15 mg, or placebo for 2 weeks followed by a dose escalation to 30 mg until week 8 or placebo. Both groups received nutritional assessment and dietary advice. Main outcomes and measures: Appetite was assessed by the Anorexia Cachexia Scale and energy intake. Dietary parameters were evaluated at baseline, 4 weeks, and 8 weeks, with a 24-hour dietary recall, and energy quantification based on the Mexican system of nutritional equivalents. Results: A total of 86 patients met the inclusion criteria and were randomized to the placebo (n = 43) or the mirtazapine group (n = 43). The mean (SD) age was 63.5 (11.2) years, 41 were women (57.7%) and had adenocarcinoma, Eastern Cooperative Oncology Group performance status scale score of 1, stage IV NSCLC, and were receiving first-line treatment. Baseline characteristics were similar between groups. There was no difference in appetite scores in patients who received mirtazapine or placebo after 4 and 8 weeks. After 4 weeks, mirtazapine significantly increased energy intake (379.3 kcal; 95% CI, 1382.6-576.1; P < .001) including proteins (22.5 g; 95% CI, 11.5-33.4; P = .001), carbohydrates (43.4 g; 95% CI, 13.1-73.8; P = .006), and fats (13.2 g; 95% CI, 6.0-20.4; P = .006). Fats intake was significantly higher in patients in the mirtazapine group (14.5 g vs 0.7 g; P = .02) after 8 weeks. The mirtazapine group significantly decreased the proportion of patients with sarcopenia (82.8% vs 57.1%, P = .03) at 8 weeks. Patients on mirtazapine tolerated the treatment well, but reported a higher perception of nightmares at 2 weeks based on a 10 cm VAS score (0 [25th-75th percentile, 0-1] vs 0 [25th-75th percentile, 0-0] in the control group; P = .009) but this finding was nonsignificant after 4 and 8 weeks. Conclusion and Relevance: In this randomized clinical trial of patients with advanced NSCLC, there was no difference in appetite scores in all patients who received mirtazapine or placebo, but the mirtazapine group had a significant increase in energy intake through the 4- and 8-week follow-up, mainly in fat intake, which is a better and crucial source of energy. The addition of mirtazapine in the treatment of patients with advanced NSCLC and anorexia may help these patients achieve their energy requirements and improve health-related quality of life, specifically emotional and cognitive functioning. Trial Registration: ClinicalTrials.gov Identifier: NCT04748523.

PROTOCOL FOR HUMAN EXPOSURE TO OPIOIDS AND CONCENTRATED MEDETOMIDINE USED IN FIELD APPLICATIONS.

Wildlife professionals routinely use potent sedatives and anesthetics when chemically immobilizing wildlife and zoo species in remote environments. Accidental exposure to these prescription veterinary drugs is rare but could be rapidly fatal. Commonly used agents include opioids and α2 adrenoreceptor agonists. These drugs can be reversed with specific antagonists; however, they are often not approved for human use. The protocol created here can be used by wildlife health professionals in a field setting with basic human emergency medical response training in coordination with local Emergency Medical Services (EMS). Key components include, building local relationships between EMS and wildlife professionals, focused EMS training, administering opioid and α2 adrenergic antagonists off label, and local evacuation procedures. This framework could allow wildlife management agencies or zoos to mitigate the risk of human exposures to these commonly used drugs, significantly improving occupational safety in an otherwise high-risk environment.

Division of Blastocystis ST10 into three new subtypes: ST42-ST44.

The Blastocystis subtype ST10 has been recognized to contain a great deal of diversity at the sequence level, potentially indicating the presence of multiple new STs within the clade. However, the data needed to validate these new STs were not available. To help resolve this diversity, full-length small subunit (SSU) rRNA gene reference sequences were generated using Oxford Nanopore MinION long-read sequencing from 21 samples representing multiple domestic and wild hosts and geographic regions and covering the sequence diversity previously described using fragments of the SSU rRNA gene. Phylogenetic and pairwise distance analyses were used to compare full-length sequences of the SSU rRNA gene generated in this study with all other valid STs of Blastocystis. We present data supporting the division of ST10/ST23 cluster into five subtypes, ST10, ST23, and three new subtypes with the proposed ST designations of ST42, ST43, and ST44. As the host range of Blastocystis continues to expand with new subtypes and new hosts being frequently identified, the reference sequences provided in this study will assist in accurate sequence classification and help to clarify the epidemiology of this common intestinal microeukaryote.

Mechanism of Benzene Hydroxylation on Tri-Iron Oxo-Centered Cluster-Based Metal-Organic Frameworks.

High-valent Fe(IV)-oxo species derived upon reactions of N2O with Fe(II) centers-embedded in the framework of tri-iron oxo-centered-based metal-organic frameworks (MOFs)- selectively affect the conversion of benzene-to-phenol via electrophilic addition to arene C-H bonds akin to oxygen transfer mechanisms in the P450 enzyme. The Fe(II) species identified by Mössbauer spectroscopy can be titrated in situ by the addition of NO to completely suppress benzene oxidation, verifying the relevance of Fe(II) centers. Observed inverse kinetic isotope effects in benzene hydroxylation preclude the involvement of H atom transfer steps from benzene to the Fe(IV)-oxo species and instead suggest that the electrophilic iron-oxo group adds to an sp2 carbon of benzene, resulting in a change in the hybridization from sp2-to-sp3. These mechanistic postulates are affirmed in Kohn-Sham density functional calculations, which predict lower barriers for additive mechanisms for arene oxidation than H atom abstraction steps. The calculations show that the reaction proceeds on the pentadectet spin surface and that a non-innocent ligand participates in the transfer of the H atom. Following precedent literature which demonstrates that these Fe(IV)-oxo species react with C-H bonds in alkanes via hydrogen atom abstraction to form alcohols, it appears that iron(IV)-oxo species in MOFs exhibit duality in their reactions with inert hydrocarbon substrates akin to enzymes-if the C-H bonds are in saturated aliphatic hydrocarbons, then activation occurs via hydrogen abstraction, while if the C-H bonds are aromatic, then activation occurs by addition rearrangement.

Perilesional edema diameter associated with brain metastases as a predictive factor of response to radiotherapy in non-small cell lung cancer.

Background: Different prognostic scales exist in patients with brain metastasis, particularly in lung cancer. The Graded Prognostic Assessment for lung cancer using molecular markers (Lung-molGPA index) for brain metastases is a powerful prognostic tool that effectively identifies patients at different risks. However, these scales do not include perilesional edema diameter (PED) associated with brain metastasis. Current evidence suggests that PED might compromise the delivery and efficacy of radiotherapy to treat BM. This study explored the association between radiotherapy efficacy, PED extent, and gross tumor diameter (GTD). Aim: The aim of this study was to evaluate the intracranial response (iORR), intracranial progression-free survival (iPFS), and overall survival (OS) according to the extent of PED and GT. Methods: Out of 114 patients with BM at baseline or throughout the disease, 65 were eligible for the response assessment. The GTD and PED sum were measured at BM diagnosis and after radiotherapy treatment. According to a receiver operating characteristic (ROC) curve analysis, cutoff values were set at 27 mm and 17 mm for PED and GT, respectively. Results: Minor PED was independently associated with a better iORR [78.8% vs. 50%, OR 3.71 (95% CI 1.26-10.99); p = 0.018] to brain radiotherapy. Median iPFS was significantly shorter in patients with major PED [6.9 vs. 11.8 months, HR 2.9 (95% CI 1.7-4.4); p < 0.001] independently of other prognostic variables like the Lung-molGPA and GTD. A major PED also negatively impacted the median OS [18.4 vs. 7.9 months, HR 2.1 (95% CI 1.4-3.3); p = 0.001]. Conclusion: Higher PED was associated with an increased risk of intracranial progression and a lesser probability of responding to brain radiotherapy in patients with metastatic lung cancer. We encourage prospective studies to confirm our findings.