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Visceral Leishmaniasis (VL) is an endemic disease in Latin America, and the clinical outcome worsens when a patient has HIV co-infection. In these patients, the immune response is complex and cytokines profile variable. We evaluate Th1/Th2/Th17 cytokine profile in VL/HIV patients from Brazilian high endemic area. In this cross-sectional study, the serological cytokines production was compared with clinical and epidemiological traits of the VL/HIV, VL and HIV patients. VL/HIV patients are predominantly male (89.2 %) with relapses in 35.5 % of the cases. There is higher serum levels of IL-6 (p = 0.006) and IL-10 (p < 0.001) in VL/HIV patients. Furthermore, there is a moderate or strong positive correlation in the serum levels of IL-6 and TNF-α (Rho = 0.635, p < 0.001), IL-10 and IFN-γ (Rho = 0.523, p < 0.001), IL-6 and IL-10 (Rho = 0.506, p < 0.001), IL-2 and IL-4 (Rho = 0.506, p < 0.001). Then, VL/HIV patients who died during VL treatment (p = 0.033), patients with oedema (p = 0.011), and patients with jaundice (p = 0.019) had statistically high levels of IL-6. In conclusion, VL/HIV patients have production or reduction of specific cytokines, highlights IL-6 and IL-10., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Tyrosinases (TYRs) are type-3 copper proteins that are widely distributed in nature. They can hydroxylate and oxidize phenolic molecules and are mostly known for producing melanins that confer protection against photo induced damage. TYRs are also thought to play an important role in the 'latch mechanism', where high concentrations of phenolic compounds inhibit oxidative decomposition of organic biomass and subsequent CO 2 release, especially relevant in wetland environments. In the present study, we describe two TYRs, HcTyr1 and HcTyr2, from halophilic bacterium Hahella sp. CCB MM4 previously isolated at Matang mangrove forest in Perak, Malaysia. The structure of HcTyr1 was determined by X-ray crystallography at a resolution of 1.9 Å and represents an uncharacterized group of prokaryotic TYRs as demonstrated by a sequence similarity network analysis. The genes encoding the enzymes were cloned, expressed, purified and thoroughly characterized by biochemical methods. HcTyr1 was able to self-cleave its lid-domain (LID) in a protease independent manner, whereas the LID of HcTyr2 was essential for activity and stability. Both enzymes showed variable activity in the presence of different metals, surfactants and NaCl, and were able to oxidize lignin constituents. The high salinity tolerance of HcTyr1 indicates that the enzyme can be an efficient catalyst in the habitat of the host., (© 2024 The Author(s). FEBS Open Bio published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

The long-term physiological consequences of respiratory viral infections, particularly in the aftermath of the COVID-19 pandemic-termed post-acute sequelae of SARS-CoV-2 (PASC)-are rapidly evolving into a major public health concern 1-3 . While the cellular and molecular aetiologies of these sequelae are poorly defined, increasing evidence implicates abnormal immune responses 3-6 and/or impaired organ recovery 7-9 after infection. However, the precise mechanisms that link these processes in the context of PASC remain unclear. Here, with insights from three cohorts of patients with respiratory PASC, we established a mouse model of post-viral lung disease and identified an aberrant immune-epithelial progenitor niche unique to fibroproliferation in respiratory PASC. Using spatial transcriptomics and imaging, we found a central role for lung-resident CD8 + T cell-macrophage interactions in impairing alveolar regeneration and driving fibrotic sequelae after acute viral pneumonia. Specifically, IFNγ and TNF derived from CD8 + T cells stimulated local macrophages to chronically release IL-1β, resulting in the long-term maintenance of dysplastic epithelial progenitors and lung fibrosis. Notably, therapeutic neutralization of IFNγ + TNF or IL-1β markedly improved alveolar regeneration and pulmonary function. In contrast to other approaches, which require early intervention 10 , we highlight therapeutic strategies to rescue fibrotic disease after the resolution of acute disease, addressing a current unmet need in the clinical management of PASC and post-viral disease., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Postacute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (PASC) represent an urgent public health challenge and are estimated to affect more than 60 million individuals globally. Although a growing body of evidence suggests that dysregulated immune reactions may be linked with PASC symptoms, most investigations have primarily centered around blood-based studies, with few focusing on samples derived from affected tissues. Furthermore, clinical studies alone often provide correlative insights rather than causal mechanisms. Thus, it is essential to compare clinical samples with relevant animal models and conduct functional experiments to understand the etiology of PASC. In this study, we comprehensively compared bronchoalveolar lavage fluid single-cell RNA sequencing data derived from clinical PASC samples and a mouse model of PASC. This revealed a pro-fibrotic monocyte-derived macrophage response in respiratory PASC, as well as abnormal interactions between pulmonary macrophages and respiratory resident T cells, in both humans and mice. Interferon-γ (IFN-γ) emerged as a key node mediating the immune anomalies in respiratory PASC. Neutralizing IFN-γ after the resolution of acute SARS-CoV-2 infection reduced lung inflammation and tissue fibrosis in mice. Together, our study underscores the importance of performing comparative analysis to understand the cause of PASC and suggests that the IFN-γ signaling axis might represent a therapeutic target.

Capillary electrophoresis (CE) is an analytical method in which charged species are separated by attraction or repulsion performed in submillimeter diameter capillaries or micro- and nanofluidic channels through the application of a high voltage electric field. When capillary electrophoresis is assembled in a multicapillary instrument such as 96-well format (multiplexed), it becomes a powerful high-throughput system with the ability to simultaneously screen several types of samples like genetic mutations, metabolomes, kinase inhibitors, or enzymatic activities to name a few. The usage of a 96-multiplexed capillary electrophoresis system (96-MP-CE) represents a new platform for product-specific high-throughput screening of enzyme mutant libraries from directed evolution campaigns providing a comprehensive view on enzyme activity through the detection of all products formed. We describe the application of 96-MP-CE to screen mutant libraries of P450 BM3. MP-CE was used in directed evolution campaigns toward benzo-1,4-dioxane and α-isophorone., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

The risk of developing severe forms of tuberculosis has increased by the acquired immunodeficiency syndrome (AIDS) epidemic, lack of effective drugs to eliminate latent infection and the emergence of drug-resistant mycobacterial strains. Excessive inflammatory response and tissue damage associated with severe tuberculosis contribute to poor outcome of the disease. Our previous studies using mice deficient in the ATP-gated ionotropic P2X7 receptor suggested this molecule as a promising target for host-directed therapy in severe pulmonary tuberculosis. In this study, we assessed the effects of P2X7 pharmacological blockade on disease severity. First, we observed an increase in P2RX7 gene expression in the peripheral blood of tuberculosis patients compared to healthy donors. Lung leukocytes of mice infected with hypervirulent mycobacteria also showed increased expression of the P2X7 receptor. P2X7 blockade in mice with advanced tuberculosis recapitulated in many aspects the disease in P2X7-deficient mice. P2X7-directed therapy reduced body weight loss and the development of inflammatory and necrotic lung lesions, as well as delayed mycobacterial growth. Lower TNF-α production by lung cells and a substantial reduction in the lung GR-1 + myeloid cell population were observed after P2X7 inhibition. The effector CD4 + T cell population also decreased, but IFN-γ production by lung cells increased. The presence of a large population with characteristics of myeloid dendritic cells, as well as the increase in IL-6 production by lung cells, also indicate a qualitative improvement in the pulmonary immune response due to P2X7 inhibition. These findings support the use of drugs that target the P2X7 receptor as a therapeutic strategy to improve the outcome of pulmonary tuberculosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Santiago-Carvalho, Almeida-Santos, Bomfim, Souza, Silva, Melo, Amaral, Cione, Lasunskaia, Hirata, Alves-Filho, Nakaya, Alvarez and D’Império Lima.)

Hydroxy- or ketone- functionalized fatty acid methyl esters (FAMEs) are important compounds for production of pharmaceuticals, vitamins, cosmetics or dietary supplements. Biocatalysis through enzymatic cascades has drawn attention to the efficient, sustainable, and greener synthetic processes. Furthermore, whole cell catalysts offer important advantages such as cofactor regeneration by cell metabolism, omission of protein purification steps and increased enzyme stability. Here, we report the first whole cell catalysis employing an engineered P450 BM3 variant and cpADH5 coupled cascade reaction for the biosynthesis of hydroxy- and keto-FAMEs. Firstly, P450 BM3 was engineered through the KnowVolution approach yielding P450 BM3 variant YE&#95;M1&#95;2, (R47S/Y51W/T235S/N239R/I401 M) which exhibited boosted performance toward methyl hexanoate. The initial oxidation rate of YE&#95;M1&#95;2 toward methyl hexanoate was determined to be 23-fold higher than the wild type enzyme and a 1.5-fold increase in methyl 3-hydroxyhexanoate production was obtained (YE&#95;M1&#95;2; 2.75 mM and WT; 1.8 mM). Subsequently, the whole cell catalyst for the synthesis of methyl 3-hydroxyhexanoate and methyl 3-oxohexanoate was constructed by combining the engineered P450 BM3 and cpADH5 variants in an artificial operon. A 2.06 mM total product formation was achieved by the whole cell catalyst including co-expressed channel protein, FhuA and co-solvent addition. Moreover, the generated whole cell biocatalyst also accepted methyl valerate, methyl heptanoate as well as methyl octanoate as substrates and yielded ω-1 ketones as the main product., (Copyright © 2020. Published by Elsevier Inc.)