Neuroradiological features of patients with bilateral macronodular adrenocortical disease and meningiomas associated or not with genetic variants of ARMC5- a case series.

INTRODUCTION: Meningiomas are the most common primary brain and central nervous system tumors, accounting for approximately 40% of these tumors. The most important exams for the radiological study of meningiomas are computed tomography (CT) and magnetic resonance imaging (MRI). We aimed to analyze the radiological features of patients with meningioma related to the simultaneous presence of bilateral macronodular adrenocortical disease (BMAD), with or without pathogenic variants of ARMC5. METHODS: This study included 10 patients who were diagnosed with BMAD. All of them had a radiological diagnosis of expansive brain lesions suggestive of meningioma. All patients underwent brain MRI and a neuroradiolgist analyzed the following parameters: number, site and size of lesions; presence of calcification, edema and bone involvement. RESULTS AND DISCUSSION: Eight patients presented with germline variants of ARMC5; the other 2, did not. The most significant result was the incidence of multiple meningiomas, which was 50% in BMAD patients, whereas the average incidence described thus far is lower than 10%. Considering location, the 22 tumors in the BMAD patients were 5 convexity tumors (22.7%), and 17 skull base tumors (77.2%), the opposite proportion of patients without BMAD. A total of 40.9% of the tumors had calcification, 9% had cerebral edema and 40.9% had bone invasion due to hyperostosis. The literature describes meningioma calcification in 25% of patients, bone invasion by tumor hyperostosis in 20%, and cerebral edema in approximately 60%. CONCLUSION: Relevant results were found considering the rate of multiple meningiomas and tumor location. This finding reinforces the need for further research into the neurological effects caused by genetic variants of ARMC5 in patients with BMAD.

Emergence of bedaquiline-resistant tuberculosis and of multidrug-resistant and extensively drug-resistant Mycobacterium tuberculosis strains with rpoB Ile491Phe mutation not detected by Xpert MTB/RIF in Mozambique: a retrospective observational study.

BACKGROUND: In 2021, an estimated 4800 people developed rifampicin-resistant tuberculosis in Mozambique, 75% of which went undiagnosed. Detailed molecular data on rifampicin-resistant and multidrug-resistant (MDR) tuberculosis are not available. Here, we aimed at gaining precise data on the determinants of rifampicin-resistant and MDR tuberculosis in Mozambique. METHODS: In this retrospective observational study, we performed whole-genome sequencing of 704 rifampicin-resistant Mycobacterium tuberculosis complex (Mtbc) strains submitted to the National Tuberculosis Reference Laboratory (NTRL) in Maputo, Mozambique, between 2015 and 2021. Phylogenetic strain classification, genomic resistance prediction, and cluster analysis were performed. FINDINGS: Between Jan 1, 2015, and July 31, 2021, 2606 Mtbc isolates with an isoniazid or rifampicin resistance were identified in the NTRL biobank, of which, 1483 (56·9%) were from men, 1114 (42·7%) from women, and nine (0·4%) were unknown. Genome-based drug-resistant prediction classified 704 Mtbc strains as rifampicin resistant. 628 (89%) of the 704 Mtbc strains were classified MDR; of those, 146 (23%) were pre-extensively drug resistant (pre-XDR; additional fluoroquinolone resistance), and 24 (4%) extensively drug resistant (XDR; combined fluoroquinolone and bedaquiline resistance). Overall, 61 (9%) of 704 strains revealed resistance to bedaquiline: five (7%) of 76 rifampicin resistant plus bedaquiline resistant, 32 (7%) of 458 MDR plus bedaquiline resistant, and 24 (100%) of 24 XDR. Prevalence of bedaquiline resistance increased from 3% in 2016 to 14% in 2021. The cluster rate (12 single-nucleotide polymorphism threshold) was 42% for rifampicin-resistant strains, 78% for MDR strains, 94% for pre-XDR strains, and 96% for XDR Mtbc strains. 31 (4%) of 704 Mtbc strains, belonging to a diagnostic escape outbreak strain previously described in Eswatini (group_56), had an rpoB Ile491Phe mutation which is not detected by Xpert MTB/RIF (no other rpoB mutation). Of these, 23 (74%) showed additional resistance to bedaquiline, 13 (42%) had bedaquiline and fluoroquinolone resistance, and two (6%) were bedaquiline, fluoroquinolone, and delamanid resistant. INTERPRETATION: Pre-XDR resistance is highly prevalent among MDR Mtbc strains in Mozambique and so is bedaquiline resistance; and the frequency of bedaquiline resistance quadrupled over time and was found even in Mtbc strains without fluoroquinolone resistance. Importantly, strains with Ile491Phe mutation were frequent, accounting for 31% (n=10) of MDR plus bedaquiline-resistant strains and 54% (n=13) of XDR Mtbc strains. Given the current diagnostic algorithms and treatment regimens, both the emergence of rifampicin resistance due to Ile491Phe and bedaquiline resistance might jeopardise MDR tuberculosis prevention and care unless sequencing-based technology is rolled out. The potential cross border spread of diagnostic escape strains needs further investigation. FUNDING: The German Ministry of Health through the Seq_MDRTB-Net project, the Deutsche Forschungsgemeinschaft under Germany's Excellence Strategy Precision Medicine in Inflammation and the Research Training Group 2501 TransEvo, the Leibniz Science Campus Evolutionary Medicine of the Lung, and the German Ministry of Education and Research via the German Center for Infection Research.

Immunoinformatic Approach for Rational Identification of Immunogenic Peptides Against Host Entry and/or Exit Mpox Proteins and Potential Multiepitope Vaccine Construction.

COVID-19 has intensified humanity's concern about the emergence of new pandemics. Since 2018, epidemic outbreaks of the mpox virus have become worrisome. In June 2022, the World Health Organization declared the disease a global health emergency, with 14 500 cases reported by the Centers for Disease Control and Prevention in 60 countries. Therefore, the development of a vaccine based on the current virus genome is paramount in combating new cases. In view of this, we hypothesized the obtainment of rational immunogenic peptides predicted from proteins responsible for entry of the mpox virus into the host (A17L, A26L/A30L, A33R, H2R, L1R), exit (A27L, A35R, A36R, C19L), and both (B5R). To achieve this, we aligned the genome sequencing data of mpox virus isolated from an infected individual in the United States in June 2022 (ON674051.1) with the reference genome dated 2001 (NC_003310.1) for conservation analysis. The Immune Epitope Database server was used for the identification and characterization of the epitopes of each protein related to major histocompatibility complex I or II interaction and recognition by B-cell receptors, resulting in 138 epitopes for A17L, 233 for A28L, 48 for A33R, 77 for H2R, 77 for L1R, 270 for A27L, 72 for A35R, A36R, 148 for C19L, and 276 for B5R. These epitopes were tested in silico for antigenicity, physicochemical properties, and allergenicity, resulting in 51, 40, 10, 34, 38, 57, 25, 7, 47, and 53 epitopes, respectively. Additionally, to select an epitope with the highest promiscuity of binding to major histocompatibility complexes and B-cell receptor simultaneously, all epitopes of each protein were aligned, and the most repetitive and antigenic regions were identified. By classifying the results, we obtained 23 epitopes from the entry proteins, 16 from the exit proteins, and 7 from both. Subsequently, 1 epitope from each protein was selected, and all 3 were fused to construct a chimeric protein that has potential as a multiepitope vaccine. The constructed vaccine was then analyzed for its physicochemical, antigenic, and allergenic properties. Protein modeling, molecular dynamics, and molecular docking were performed on Toll-like receptors 2, 4, and 8, followed by in silico immune simulation of the vaccine. Finally, the results indicate an effective, stable, and safe vaccine that can be further tested, especially in vitro and in vivo, to validate the findings demonstrated in silico.

Applying the bioisosterism strategy to obtain lead compounds against SARS-CoV-2 cysteine proteases: An in-silico approach.

SARS-CoV-2 cysteine proteases are essential nonstructural proteins due to their role in the formation of the virus multiple enzyme replication-transcription complex. As a result, those functional proteins are extremely relevant targets in the development of a new drug candidate to fight COVID-19. Based on this fact and guided by the bioisosterism strategy, the present work has selected 126 out of 1050 ligands from DrugBank website. Subsequently, 831 chemical analogs containing bioisosteres, some of which became structurally simplified, were created using the MB-Isoster software, and molecular docking simulations were performed using AutoDock Vina. Finally, a study of physicochemical properties, along with pharmacokinetic profiles, was carried out through SwissADME and ADMETlab 2.0 platforms. The promising results obtained with the molecules encoded as DB00549_BI_005, DB04868_BI_003, DB11984_BI_002, DB12364_BI_006 and DB12805_BI_004 must be confirmed by molecular dynamics studies, followed by in vitro and in vivo empirical tests that ratify the advocated in-silico results.

Antisynthetase Syndrome With Predominant Pulmonary Involvement: A Case Report.

Antisynthetase syndrome (ASyS) is an autoimmune disease characterized by the presence of aminoacyl-transfer RNA synthetase antibodies. Its clinical presentation is variable and may include interstitial lung disease (ILD), myositis, arthritis, fever, Raynaud's phenomenon, and "mechanic's hands." ILD is more prevalent in this entity when compared to other idiopathic inflammatory myopathies and imparts greater severity to the condition. Here, we report the case of a 42-year-old female patient who sought care for severe ILD and persistent fever. Her diagnosis was made only after the detection of anti-Jo1 autoantibodies. Treatment was refractory to both prednisone monotherapy and cyclophosphamide pulse therapy, requiring the introduction of rituximab. A high degree of clinical suspicion is required to allow early diagnosis of ASyS in patients with pulmonary involvement in the absence of accompanying muscle weakness or other clinical symptoms.

Implementation of targeted next-generation sequencing for the diagnosis of drug-resistant tuberculosis in low-resource settings: a programmatic model, challenges, and initial outcomes.

Targeted next-generation sequencing (tNGS) from clinical specimens has the potential to become a comprehensive tool for routine drug-resistance (DR) prediction of Mycobacterium tuberculosis complex strains (MTBC), the causative agent of tuberculosis (TB). However, TB mainly affects low- and middle-income countries, in which the implementation of new technologies have specific needs and challenges. We propose a model for programmatic implementation of tNGS in settings with no or low previous sequencing capacity/experience. We highlight the major challenges and considerations for a successful implementation. This model has been applied to build NGS capacity in Namibia, an upper middle-income country located in Southern Africa and suffering from a high-burden of TB and TB-HIV, and we describe herein the outcomes of this process.

Detection of herpesviruses in neotropical primates from São Paulo, Brazil.

Transmission of herpesvirus between humans and non-human primates represents a serious potential threat to human health and endangered species conservation. This study aimed to identify herpesvirus genomes in samples of neotropical primates (NTPs) in the state of São Paulo, Brazil. A total of 242 NTPs, including Callithrix sp., Alouatta sp., Sapajus sp., and Callicebus sp., were evaluated by pan-herpesvirus polymerase chain reaction (PCR) and sequencing. Sixty-two (25.6%) samples containing genome segments representative of members of the family Herpesviridae, including 16.1% for Callitrichine gammaherpesvirus 3, 6.1% for Human alphaherpesvirus 1, 2.1% for Alouatta macconnelli cytomegalovirus, and 0.83% for Cebus albifrons lymphocryptovirus 1. No co-infections were detected. The detection of herpesvirus genomes was significantly higher among adult animals (p = 0.033) and those kept under human care (p = 0.008671). These findings confirm the importance of monitoring the occurrence of herpesviruses in NTP populations in epizootic events.

Technical comparison of MinIon and Illumina technologies for genotyping Chikungunya virus in clinical samples.

New-generation sequencing (NGS) techniques have brought the opportunity for genomic monitoring of several microorganisms potentially relevant to public health. The establishment of different methods with different mechanisms provides a wide choice, taking into account several aspects. With that in mind, the present aim of the study was to compare basic genomic sequencing metrics that could potentially impact genotyping by nanopores from Oxford Nanopore Technologies and by synthesis from Illumina in clinical samples positive for Chikungunya (CHIKV). Among the metrics studied, running time, read production, and Q score were better represented in Illumina sequencing, while the MinIOn platform showed better response time and greater diversity of generated files. That said, it was possible to establish differences between the studied metrics in addition to verifying that the distinctions in the methods did not impact the identification of the CHIKV virus genotype.

Progress and promise of alternative animal and non-animal methods in biomedical research.

Cell culture and invertebrate animal models reflect a significant evolution in scientific research by providing reliable evidence on the physiopathology of diseases, screening for new drugs, and toxicological tests while reducing the need for mammals. In this review, we discuss the progress and promise of alternative animal and non-animal methods in biomedical research, with a special focus on drug toxicity.

A comparative analysis of clinical, functional and radiographic outcomes of patients with and without fusion between the first and second metatarsals after the lapidus procedure.

BACKGROUND: The aim of this study was to compare the clinical, functional and radiographic outcomes in patients who underwent the original Lapidus procedure (OLP) with fusion and undesired non-fusion between the first (M1) and second (M2) metatarsals. METHODS: A retrospective and comparative analysis of 29 patients (38 feet) who underwent the OLP was performed. They were divided in two groups: (1) 23 feet in which fusion occurred and (2) 15 with undesired non-fusion. Clinical and functional data were assessed with the VAS for pain, AOFAS, LEFS and SF-12. SF-12 comprises physical and mental health scales (PCS-12 and MCS-12). Radiographic parameters assessed were bony and soft tissue forefoot widths (BSFW), intermetatarsal-angle (IMA) and HV-angle (HVA). RESULTS: Separately, the groups presented significant improvements in all questionnaires (p < 0.001), except on MCS-12 (fusion p = 0.08 and non-fusion p = 0.27). When comparing both groups, patients with fusion had higher AOFAS scores (p < 0.05). Both groups presented the same improvements on BSFW, IMA and HVA (p = 0.09, p = 0.16, p = 0.52 and p = 0.63). CONCLUSION: Excellent results were observed, even when without fusion between M1 and M2. Patients who evolved with fusion between the M1 and M2 showed greater improvements in the AOFAS score. LEVEL OF EVIDENCE: III, retrospective comparative study.

Histopathologic and Immunohistochemical Assessment of Acute Respiratory Distress Syndrome (ARDS): Challenges and Complexities of Postmortem Diagnosis.

Acute respiratory distress syndrome (ARDS) is a life-threatening condition due to acute lung injury (ALI), characterized by rapid-onset respiratory failure, leading to the clinical manifestations of poor lung compliance, severe hypoxemia, and dyspnea. ARDS/ALI has many causes, most commonly related to infections (sepsis, pneumonia), traumas, and multiple transfusions. The objective of this study is to assess the performance of postmortem anatomopathological examination in identifying etiological agents associated with ARDS or ALI in deceased patients from the State of São Paulo from 2017 to 2018. A retrospective cross-sectional study was performed based on the final outcome obtained by histopathology, histochemical, and immunohistochemical examination for ARDS/ALI differential diagnosis at the Pathology Center of the Adolfo Lutz Institute in São Paulo, Brazil. Of the 154 patients clinically diagnosed with ARDS or ALI, 57% tested positive for infectious agents, and the most frequent outcome was influenza A/H1N1 virus infection. In 43% of cases, no etiologic agent was identified. The opportunity to establish a diagnosis, identify particular infections, confirm a microbiological diagnosis, and uncover unanticipated etiologies is provided by postmortem pathologic analysis of ARDS. A molecular assessment could improve the diagnosis accuracy and lead to research into host responses and public health measures.

Use of High-Resolution Geospatial and Genomic Data to Characterize Recent Tuberculosis Transmission, Botswana.

Combining genomic and geospatial data can be useful for understanding Mycobacterium tuberculosis transmission in high-burden tuberculosis (TB) settings. We performed whole-genome sequencing on M. tuberculosis DNA extracted from sputum cultures from a population-based TB study conducted in Gaborone, Botswana, during 2012-2016. We determined spatial distribution of cases on the basis of shared genotypes among isolates. We considered clusters of isolates with ≤5 single-nucleotide polymorphisms identified by whole-genome sequencing to indicate recent transmission and clusters of ≥10 persons to be outbreaks. We obtained both molecular and geospatial data for 946/1,449 (65%) participants with culture-confirmed TB; 62 persons belonged to 5 outbreaks of 10-19 persons each. We detected geospatial clustering in just 2 of those 5 outbreaks, suggesting heterogeneous spatial patterns. Our findings indicate that targeted interventions applied in smaller geographic areas of high-burden TB identified using integrated genomic and geospatial data might help interrupt TB transmission during outbreaks.

Genomic epidemiology of SARS-CoV-2 during the first four waves in Mozambique.

Mozambique reported the first case of coronavirus disease 2019 (COVID-19) in March 2020 and it has since spread to all provinces in the country. To investigate the introductions and spread of SARS-CoV-2 in Mozambique, 1 142 whole genome sequences sampled within Mozambique were phylogenetically analyzed against a globally representative set, reflecting the first 25 months of the epidemic. The epidemic in the country was marked by four waves of infection, the first associated with B.1 ancestral lineages, while the Beta, Delta, and Omicron Variants of Concern (VOCs) were responsible for most infections and deaths during the second, third, and fourth waves. Large-scale viral exchanges occurred during the latter three waves and were largely attributed to southern African origins. Not only did the country remain vulnerable to the introductions of new variants but these variants continued to evolve within the borders of the country. Due to the Mozambican health system already under constraint, and paucity of data in Mozambique, there is a need to continue to strengthen and support genomic surveillance in the country as VOCs and Variants of interests (VOIs) are often reported from the southern African region.

Disseminated Mucor indicus infection in a marmoset (Callithrix sp.).

Mucormycosis is rarely described in Platyrrhines. Herein, we describe the pathologic and molecular features of a gastric and hepatic infection by Mucor indicus in a marmoset (Callithrix sp.).

Infections by entomopathogenic fungi in common green iguanas (Iguana iguana) in captivity in Brazil.

Entomopathogenic fungi, widely available biological agents used to control agricultural pests, are sporadically reported to cause focal or disseminated infection in reptiles and mammals, including humans. This study summarizes the clinical presentation, histopathological and molecular findings by panfungal polymerase chain reaction and sequencing of four cases of hypocrealean fungal infections in captive common green iguanas (Iguana, iguana). One case of granulomatous pneumonia, hepatitis and serositis was related to Metarhizium flavoviride complex infection. Two disseminated fungal infection cases, with scarce inflammatory cell infiltration, were caused by Beauveria bassiana while there was one case of multifocal granulomatous and necrotizing pneumonia by Purpureocillium spp. To the best of our knowledge, this is the first report of fatal mycosis infection due to entomopathogenic fungi in captive common green iguanas.

Co-production of Classes A and B Carbapenemases BKC-1 and VIM-2 in a Clinical Pseudomonas Putida Group Isolate from Brazil.

Emergence of resistance to classical antimicrobial agents is a public health issue, especially in countries with high antimicrobial consumption rates. Carbapenems have been employed as first-choice option for empirical treatment complicated infections. However, in the last decades, frequency of carbapenemase-producing Gram-negative bacteria has rising, demanding the use of alternative antimicrobial agents. By sequencing the entire genomes with short and long reads technologies, we report the isolation and genomic characterization of a carbapenem-resistant Pseudomonas clinical isolate. The identification based on average nucleotide identity indicates a putative new species into the Pseudomonas putida Group, which carries both the blaBKC-1 and blaVIM-2 carbapenemase genes. The blaBKC-1 was found to be on a transferable IncQ plasmid backbone, whereas blaVIM-2 was found in a new integron, In2126 (intl1∆-blaVIM-2-aacA7-blaVIM-2∆-aacA27-3'CS), described in this study. Our findings indicate that co-occurrence of classes A and B carbapenemase enzymes underscores the evolving emergence of more complex antimicrobial resistance in opportunistic pathogens.

A Study on the Anti-NF-κB, Anti-Candida, and Antioxidant Activities of Two Natural Plant Hormones: Gibberellin A4 and A7.

INTRODUCTION: Gibberellins (GA) are terpenoids that serve as important plant hormones by acting as growth and response modulators against injuries and parasitism. In this study, we investigated the in vitro anti-NF-κB, anti-Candida, and antioxidant activity of gibberellin A4 (GA4) and A7 (GA7) compounds, and further determined their toxicity in vivo. METHODS: GA4 and GA7 in vitro toxicity was determined by MTT method, and nontoxic concentrations were then tested to evaluate the GA4 and GA7 anti-NF-κB activity in LPS-activated RAW-luc macrophage cell culture (luminescence assay). GA4 in silico anti-NF-κB activity was evaluated by molecular docking with the software "AutoDock Vina", "MGLTools", "Pymol", and "LigPlot+", based on data obtained from "The Uniprot database", "Protein Data Bank", and "PubChem database". The GA4 and GA7 in vitro anti-Candida effects against Candida albicans (MYA 2876) were determined (MIC and MFC). GA7 was also evaluated regarding the viability of C. albicans preformed biofilm (microplate assay). In vitro antioxidant activity of GA4 and GA7 was evaluated against peroxyl radicals, superoxide anions, hypochlorous acid, and reactive nitrogen species. GA4 and GA7 in vivo toxicity was determined on the invertebrate Galleria mellonella larvae model. RESULTS: Our data show that GA4 at 30 µM is nontoxic and capable of reducing 32% of the NF-κB activation on RAW-luc macrophages in vitro. In vitro results were confirmed via molecular docking assay (in silico), since GA4 presented binding affinity to NF-κB p65 and p50 subunits. GA7 did not present anti-NF-κB effects, but exhibited anti-Candida activity with low MIC (94 mM) and MFC (188 mM) values. GA7 also presented antibiofilm properties at 940 mM concentration. GA4 did not present anti-Candida effects. Moreover, GA4 and GA7 showed antioxidant activity against peroxyl radicals, but did not show scavenging activity against the other tested radicals. Both compounds did not affect the survival of G. mellonella larvae, even at extremely high doses (10 g/Kg). CONCLUSION: Our study provides preclinical evidence indicating that GA4 and GA7 have a favorable low toxicity profile. The study also points to GA4 and GA7 interference with the NF-κB via, anti-Candida activity, and a peroxyl radical scavenger, which we argue are relevant biological effects.