Case Report: A Series of Three Meningoencephalitis Cases Caused by Acanthamoeba spp. from Eastern India.

Acanthamoeba spp. are rare etiological agents of meningoencephalitis with high mortality. We present three cases of Acanthamoeba meningoencephalitis in immunocompetent individuals from Eastern India. The first patient presented with fever and headache; the second with headache, visual disturbance, and squint; and the third presented in a drowsy state. The cases presented on March 3, 18, and 21, 2023 respectively. The first two patients had concomitant tubercular meningitis for which they received antitubercular therapy and steroid. Their cerebrospinal fluid showed slight lymphocytic pleocytosis and increased protein. The diagnosis was done by microscopy, culture, and polymerase chain reaction. They received a combination therapy comprising rifampicin, fluconazole, and trimethoprim-sulfamethoxazole. The first patient additionally received miltefosine. She responded well to therapy and survived, but the other two patients died despite intensive care. Detection of three cases within a period of 1 month from Eastern India is unusual. It is imperative to sensitize healthcare providers about Acanthamoeba meningoencephalitis to facilitate timely diagnosis and treatment of the disease.

Plasma Gradient of Soluble Urokinase-Type Plasminogen Activator Receptor Is Linked to Pathogenic Plasma Proteome and Immune Transcriptome and Stratifies Outcomes in Severe COVID-19.

Disease caused by SARS-CoV-2 coronavirus (COVID-19) led to significant morbidity and mortality worldwide. A systemic hyper-inflammation characterizes severe COVID-19 disease, often associated with acute respiratory distress syndrome (ARDS). Blood biomarkers capable of risk stratification are of great importance in effective triage and critical care of severe COVID-19 patients. Flow cytometry and next-generation sequencing were done on peripheral blood cells and urokinase-type plasminogen activator receptor (suPAR), and cytokines were measured from and mass spectrometry-based proteomics was done on plasma samples from an Indian cohort of COVID-19 patients. Publicly available single-cell RNA sequencing data were analyzed for validation of primary data. Statistical analyses were performed to validate risk stratification. We report here higher plasma abundance of suPAR, expressed by an abnormally expanded myeloid cell population, in severe COVID-19 patients with ARDS. The plasma suPAR level was found to be linked to a characteristic plasma proteome, associated with coagulation disorders and complement activation. Receiver operator characteristic curve analysis to predict mortality identified a cutoff value of suPAR at 1,996.809 pg/ml (odds ratio: 2.9286, 95% confidence interval 1.0427-8.2257). Lower-than-cutoff suPAR levels were associated with a differential expression of the immune transcriptome as well as favorable clinical outcomes, in terms of both survival benefit (hazard ratio: 0.3615, 95% confidence interval 0.1433-0.912) and faster disease remission in our patient cohort. Thus, we identified suPAR as a key pathogenic circulating molecule linking systemic hyperinflammation to the hypercoagulable state and stratifying clinical outcomes in severe COVID-19 patients with ARDS.

Analysis of Radiation-Induced Changes in Cell Cycle and DNA Damage of Murine Hematopoietic Stem Cells by Multi-Color Flow Cytometry.

Exposure of bone marrow to genotoxic stress such as ionizing radiation (IR) results in a rapid decline of peripheral blood cells and stimulates entry of the normally quiescent hematopoietic stem cells (HSCs) into the cell cycle to reconstitute the hematopoietic system. While several protocols have employed flow cytometry analysis of bone marrow cells to study changes in specific cell populations with respect to cell cycle proliferation and/or expression of γ-H2AX, a marker of DNA damage, these parameters were examined in separate panels. Here, we describe a flow cytometry-based method specifically designed to examine cell cycle distribution using Ki-67 and FXCycle violet in combination with γ-H2AX in HSCs and hematopoietic progenitor cells (HPCs) within the same sample. This method is very useful, particularly in studies involving genotoxic stresses such as IR, which substantially reduce the absolute numbers of HSCs and HPCs available for staining. Additionally, we describe several important considerations for the analysis of markers of HSCs in irradiated versus unirradiated samples. Examples include the use of fluorescence minus one (FMO) controls, the gating strategy for markers whose expression is typically impacted by IR such as Sca1, tips for staining of intracellular antigens like Ki67, and ensuring the detection of signal from at least 500 events in each gate to ensure robustness of the results. © 2021 Wiley Periodicals LLC. Basic Protocol: Immunostaining protocol for bone marrow mononuclear cells using a multi-fluorophore panel.

Simultaneous exposure to chronic irradiation and simulated microgravity differentially alters immune cell phenotype in mouse thymus and spleen.

Deep-space missions may alter immune cell phenotype in the primary (e.g., thymus) and secondary (e.g., spleen) lymphoid organs contributing to the progression of a variety of diseases. In deep space missions, astronauts will be exposed to chronic low doses of HZE radiation while being in microgravity. Ground-based models of long-term uninterrupted exposures to HZE radiation are not yet available. To obtain insight in the effects of concurrent exposure to microgravity and chronic irradiation (CIR), mice received a cumulative dose of chronic 0.5 Gy gamma rays over one month ± simulated microgravity (SMG). To obtain insight in a dose rate effect, additional mice were exposed to single acute irradiation (AIR) at 0.5 Gy gamma rays. We measured proportions of immune cells relative to total number of live cells in the thymus and spleen, stress level markers in plasma, and change in body weight, food consumption, and water intake. CIR affected thymic CD3+/CD335+ natural killer T (NK-T) cells, CD25+ regulatory T (Treg) cells, CD27+/CD335- natural killer (NK1) cells and CD11c+/CD11b- dendritic cells (DCs) differently in mice subjected to SMG than in mice with normal loading. No such effects of CIR on SMG as compared to normal loading were observed in cell types from the spleen. Differences between CIR and AIR groups (both under normal loading) were found in thymic Treg and DCs. Food consumption, water intake, and body weight were less after coexposure than singular or no exposure. Compared to sham, all treatment groups exhibited elevated plasma levels of the stress marker catecholamines. These data suggest that microgravity and chronic irradiation may interact with each other to alter immune cell phenotypes in an organ-specific manner and appropriate strategies are required to reduce the health risk of crewmembers.

Loss of C/EBPδ Exacerbates Radiation-Induced Cognitive Decline in Aged Mice due to Impaired Oxidative Stress Response.

Aging is characterized by increased inflammation and deterioration of the cellular stress responses such as the oxidant/antioxidant equilibrium, DNA damage repair fidelity, and telomeric attrition. All these factors contribute to the increased radiation sensitivity in the elderly as shown by epidemiological studies of the Japanese atomic bomb survivors. There is a global increase in the aging population, who may be at increased risk of exposure to ionizing radiation (IR) as part of cancer therapy or accidental exposure. Therefore, it is critical to delineate the factors that exacerbate age-related radiation sensitivity and neurocognitive decline. The transcription factor CCAAT enhancer binding protein delta (C/EBPδ) is implicated with regulatory roles in neuroinflammation, learning, and memory, however its role in IR-induced neurocognitive decline and aging is not known. The purpose of this study was to delineate the role of C/EBPδ in IR-induced neurocognitive decline in aged mice. We report that aged Cebpd-/- mice exposed to acute IR exposure display impairment in short-term memory and spatial memory that correlated with significant alterations in the morphology of neurons in the dentate gyrus (DG) and CA1 apical and basal regions. There were no significant changes in the expression of inflammatory markers. However, the expression of superoxide dismutase 2 (SOD2) and catalase (CAT) were altered post-IR in the hippocampus of aged Cebpd-/- mice. These results suggest that Cebpd may protect from IR-induced neurocognitive dysfunction by suppressing oxidative stress in aged mice.

Socio-demographic and Clinico-Epidemiological Study of Scrub Typhus in Two Tertiary Care Hospitals of Kolkata.

Background and Aims: Scrub typhus is the commonest of the rickettsial diseases in India and is difficult to diagnose. Untreated cases have fatality rates of 30-45%. Eschar is present in 7-97% cases. Pneumonia and acute respiratory distress syndrome (ARDS) are frequent complications. Serum immunoglobulin M capture ELISA is the most sensitive test. Doxycycline is the drug of choice. Our objectives were to study the socio-demographic and clinic-epidemiological profiles of scrub typhus cases in two tertiary care hospitals in Kolkata, India. This was the first study of scrub typhus in Southern West Bengal and its neighboring areas. . Methods: Study was conducted over 16 months and all fever cases of Tropical Medicine / Medicine outpatients' clinics were evaluated. Results: Fourteen cases were diagnosed. 78.6% were from rural areas and 35.7% were farmers. Headache and fever were the commonest presenting complaints while eschar was found in only 21.4%. Serum IgM scrub typhus antibody was positive in all cases . Conclusion: Scrub typhus should be a differential diagnosis in acute febrile illness cases, as early diagnosis and therapy prevents complications.