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Prognostic markers are the biomarkers used to measure the disease progression and patient outcome regardless of treatment in coronavirus disease 2019 (COVID-19). This study aimed to analyze laboratory parameters as prognostic markers for the early identification of disease severity. In this study, 165 patients attending Sukraraj Tropical and Infectious Disease Hospital with COVID-19 were enrolled and divided into severe and non-severe groups. The demographic data, underlying co-morbidities, and laboratory findings were analyzed and compared between severe and non-severe cases. The correlation between the disease criticality and laboratory parameters was analyzed. Cut-off values of parameters for severe patients were speculated through the receiver operating characteristics (ROC) curve, and regression analysis was performed to determine the risk factors. Patients with severe COVID-19 infection had significantly higher absolute neutrophil count, neutrophil–lymphocyte ratio (NLR), platelet–lymphocyte ratio (PLR), ferritin, positive carbohydrate reactive protein (CRP), glucose, urea, creatinine, and aspartate aminotransferase, while lower absolute lymphocyte count, absolute eosinophil count (AEC), and red blood cell count in comparison to non-severe infection. ROC analysis gave a cut-off value (sensitivity, specificity) of age, AEC, NLR, PLR, and ferritin as 47.5 years (70.2, 64.7%), 335 cells/mm3 (74, 67%) 3.3 (68.4, 63.7%), 129 (77.2, 51%), and 241 ng/mL (74.0%, 65.0%) respectively. Risk factor analysis showed higher age, low AEC, high ferritin, and positive CRP as independent risk factors associated with severe COVID-19 infection. Hematological and inflammatory markers, including novel NLR and PLR, should be assessed to aid clinicians in the early identification of severe cases, prioritization of cases, and effective management to decrease the mortality of COVID-19 patients.

Keshav Lamichhane,1,* Nabaraj Adhikari,1,* Anup Bastola,2 Lina Devkota,2 Parmananda Bhandari,2 Binod Dhungel,1 Upendra Thapa Shrestha,1 Bipin Adhikari,3 Megha Raj Banjara,1 Komal Raj Rijal,1 Prakash Ghimire1 1Central Department of Microbiology, Tribhuvan University, Kathmandu, Nepal; 2Sukraraj Tropical and Infectious Disease Hospital, Teku, Kathmandu, Nepal; 3Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine, University of Oxford, Oxford, UK*These authors contributed equally to this workCorrespondence: Komal Raj RijalCentral Department of Microbiology, Tribhuvan University, Kirtipur, Kathmandu, NepalEmail rijalkomal@gmail.comIntroduction: Oropharyngeal candidiasis are the commonest fungal infections among HIV-positive patients. The main objective of this study was to explore biofilm-producing Candida species causing oropharyngeal infections among HIV patients attending Sukraraj Tropical and Infectious Diseases Hospital (STIDH) in Kathmandu, Nepal.Methods: Oropharyngeal swabs were collected from the HIV-positive patients between July and December 2019. A total of 174 oropharyngeal swabs were cultured on Sabouraud Dextrose Agar (SDA). All samples were inoculated on SDA slants supplemented with chloramphenicol and underwent incubation at 37°C for 24– 48 hours. Any visible growth reported was processed for the identification of the species. Candida species were differentiated based on the growth and colour of the isolates on CHROM agar candida. Biofilm production in Candida species was determined by the microtiter plate method (MPM). Antifungal susceptibility testing was performed using the disc diffusion method.Results: Among 174 oropharyngeal samples, 23.6% (n=41/174) of them had oropharyngeal infections and 36.6% of the oropharyngeal infections (15/41) had CD4 T-lymphocytes count below 200 cells/mm3 who were also active tobacco users (p< 0.05). Among Candidial growth, 61% (25/41) were Candida albicans and 39% (16/41) were non-albicans. Of 41 Candida spp., 65% (27/41) were biofilm producers. An equal proportion of Candida albicans (4 isolates) and non-albicans (4 isolates) were strong biofilm producers. C. albicans isolates were sensitive towards clotrimazole (96%; 24/25) and fluconazole (92%; 23/25), whereas sensitivity towards ketoconazole was only 48% (12/25). Non-albicans Candida was highly sensitive to amphotericin-B (62.5%; 10/16) followed by clotrimazole (56.2%; 9/16). The biofilm-producing Candida isolates showed the highest resistivity (51.9%; 14/27) to ketoconazole and lowest (22.2%; 6/27) to clotrimazole.Conclusion: Oropharyngeal candidiasis is a common opportunistic infection among HIV-infected individuals. The majority of cases of oropharyngeal candidiasis are caused by biofilm producers Candida albicans and non-albicans Candida. Biofilm producers Candida were more resistant towards commonly used antifungal drugs.Keywords: oral candidiasis, HIV, Candida albicans, biofilm, antifungal susceptibility test

Rooku KC,1,* Sadiksha Adhikari,1,* Anup Bastola,2 Lina Devkota,2 Parmananda Bhandari,2 Prabina Ghimire,3 Bipin Adhikari,4 Komal Raj Rijal,1 Megha Raj Banjara,1 Prakash Ghimire1 1Central Department of Microbiology, Tribhuvan University, Kathmandu, Nepal; 2Sukraraj Tropical and Infectious Diseases Hospital, Kathmandu, Nepal; 3Nepal Medical College, Jorpati, Kathmandu, Nepal; 4Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine, University of Oxford, Oxford, UK*These authors contributed equally to this workCorrespondence: Komal Raj RijalCentral Department of Microbiology, Tribhuvan University, Kirtipur, Kathmandu, NepalEmail rijalkomal@gmail.comIntroduction: Opportunistic bacterial and fungal infections are the major cause of morbidity and mortality among immune suppressed HIV-positive patients. The main objective of this study was to determine bacterial and fungal organisms causing respiratory infections and their susceptibility to commonly prescribed antimicrobials among HIV patients attending a tertiary infectious disease hospital in Kathmandu.Methods: Sputum samples were collected from the HIV-positive patients attending Sukraraj Tropical and Infectious Disease Hospital (STIDH) from August 2017 to March 2018. A total of 100 sputum samples were cultured on conventional bacterial and fungal culture media. Bacterial and fungal isolates were identified based on their colony characteristics, microscopic morphology and various biochemical tests. Antibiotic susceptibility test (AST) of bacterial isolates was performed by modified Kirby Bauer disc diffusion method.Results: Out of 100 sputum samples cultured, 24% (n=24) showed bacterial growth, 42% (n=42) showed fungal growth and 10% (n=10) had both bacterial and fungal growth. Among bacteria, 91.6% (n=22) were monomicrobial and 8.4% (n=2) were polymicrobial in growth, of which, Klebsiella pneumoniae (37.5%) were predominant isolates, followed by Pseudomonas aeruginosa (29.2%), and Escherichia coli (16.7%). The antibiotic susceptibility test (AST) showed 68% (17/25) of bacterial isolates were multi-drug resistant (MDR) and among them 41.2% (7/17) were found to be extended spectrum β lactamase (ESBL) producers. Fungal growth was observed in 42% of samples (42/100). A total of six different species of Candida and four different genera of molds were identified. On species differentiation, Candida albicans (20%) were followed by Candida parapsilosis (4%), and Candida dubliniensis (3%); and various molds were Aspergillus fumigatus (4%), Aspergillus flavus (2%), and Penicillium species (5%). CD4 count was inversely associated with bacterial and fungal infections. Fifty percent of the patients with the fungal infections had a CD4 count below 200. No fungal organisms were isolated from HIV-positive patients under antifungal drug treatment.Conclusion: HIV-positive patients with a CD4 count less than 200 cells/μL are more vulnerable to opportunistic infections of bacterial and fungal origin. Early isolation, identification and appropriate treatment can reduce mortality due to co-infections. Routine screening of opportunistic pathogens is critical to contain the disease progression.Keywords: HIV, CD4, bacterial and fungal infection, opportunistic infections

Glaucoma is a group of ocular diseases that cause irreversible blindness. It is characterized by multifactorial degeneration of the optic nerve axons and retinal ganglion cells (RGCs), resulting in the loss of vision. Major components of glaucoma pathogenesis include glia-driven neuroinflammation and impairment of mitochondrial dynamics and bioenergetics, leading to retinal neurodegeneration. In this review article, we summarize current evidence for the emerging role of apolipoprotein A-I binding protein (AIBP) as an important anti-inflammatory and neuroprotective factor in the retina. Due to its association with toll-like receptor 4 (TLR4), extracellular AIBP selectively removes excess cholesterol from the plasma membrane of inflammatory and activated cells. This results in the reduced expression of TLR4-associated, cholesterol-rich lipid rafts and the inhibition of downstream inflammatory signaling. Intracellular AIBP is localized to mitochondria and modulates mitophagy through the ubiquitination of mitofusins 1 and 2. Importantly, elevated intraocular pressure induces AIBP deficiency in mouse models and in human glaucomatous retina. AIBP deficiency leads to the activation of TLR4 in Müller glia, triggering mitochondrial dysfunction in both RGCs and Müller glia, and compromising visual function in a mouse model. Conversely, restoring AIBP expression in the retina reduces neuroinflammation, prevents RGCs death, and protects visual function. These results provide new insight into the mechanism of AIBP function in the retina and suggest a therapeutic potential for restoring retinal AIBP expression in the treatment of glaucoma.

The mechanical properties of solid tumors influence tumor cell phenotype and the ability to invade surrounding tissues. Using bioengineered scaffolds to provide a matrix microenvironment for patient-derived glioblastoma (GBM) spheroids, this study demonstrates that a soft, brain-like matrix induces GBM cells to shift to a glycolysis-weighted metabolic state, which supports invasive behavior. We first show that orthotopic murine GBM tumors are stiffer than peritumoral brain tissues, but tumor stiffness is heterogeneous where tumor edges are softer than the tumor core. We then developed 3D scaffolds with μ-compressive moduli resembling either stiffer tumor core or softer peritumoral brain tissue. We demonstrate that the softer matrix microenvironment induces a shift in GBM cell metabolism toward glycolysis, which manifests in lower proliferation rate and increased migration activities. Finally, we show that these mechanical cues are transduced from the matrix via CD44 and integrin receptors to induce metabolic and phenotypic changes in cancer cells.

Spinal tuberculosis usually involves intervertebral disc, pardiscal vertebra and has associated perivertebral collection. Involvement of vertebral body including the posterior element, noncontagious vertebral involvement and sparing of intervertebral discs is typical of metastasis. Index case, a 19 years old Nepali patient presented to our hospital with complaints of severe backache. Blood and urine investigations were normal except elevated erythrocyte sedimentation rate and C- reactive protein. A contrast enhanced computed tomography (CT) showed lytic lesions in clivus, C7, T10, and L3 vertebral bodies with involvement of right sided posterior elements of C7 and T10. Mild surrounding soft tissue was seen. Contrast enhanced MRI showed similar findings and also lesions in S1 and right sacral ala. Possibility of metastatic lesion was considered over tuberculosis. A CT guided biopsy from right pedicle of T10 vertebra confirmed tubercular etiology.

A-Kinase anchoring protein 1 (AKAP1) is a multifunctional mitochondrial scaffold protein that regulates mitochondrial dynamics, bioenergetics, and calcium homeostasis by anchoring several proteins, including protein kinase A, to the outer mitochondrial membrane. Glaucoma is a complex, multifactorial disease characterized by a slow and progressive degeneration of the optic nerve and retinal ganglion cells (RGCs), ultimately resulting in vision loss. Impairment of the mitochondrial network and function is linked to glaucomatous neurodegeneration. Loss of AKAP1 induces dynamin-related protein 1 dephosphorylation-mediated mitochondrial fragmentation and loss of RGCs. Elevated intraocular pressure triggers a significant reduction in AKAP1 protein expression in the glaucomatous retina. Amplification of AKAP1 expression protects RGCs from oxidative stress. Hence, modulation of AKAP1 could be considered a potential therapeutic target for neuroprotective intervention in glaucoma and other mitochondria-associated optic neuropathies. This review covers the current research on the role of AKAP1 in the maintenance of mitochondrial dynamics, bioenergetics, and mitophagy in RGCs and provides a scientific basis to identify and develop new therapeutic strategies that could protect RGCs and their axons in glaucoma.

Increased secretion of hyaluronic acid (HA), a glycosaminoglycan abundant in the brain extracellular matrix (ECM), correlates with worse clinical outcomes for glioblastoma (GBM) patients. GBM cells aggressively invade the brain parenchyma while encountering spatiotemporal changes in their local ECM, including HA concentration. To investigate how varying HA concentrations affect GBM invasion, patient-derived GBM cells are cultured within a soft, 3D matrix in which HA concentration is precisely varied and cell migration observed. Data demonstrate that HA concentration can determine the invasive activity of patient-derived GBM cells in a biphasic and highly sensitive manner, where the absolute concentration of HA at which cell migration peaked is specific to each patient-derived line. Furthermore, evidence that this response relies on phosphorylated ezrin, which interacts with the intracellular domain of HA-engaged CD44 to effectively link the actin cytoskeleton to the local ECM is provided. Overall, this study highlights CD44-HA binding as a major mediator of GBM cell migration that acts independently of integrins and focal adhesion complexes and suggests that targeting HA-CD44-ezrin interactions represents a promising therapeutic strategy to prevent tumor cell invasion in the brain.

The interplay between the gut microbiome and fermented foods has significant implications for human health and nutrition. Fermented foods, rich in beneficial microorganisms such as Lactic Acid Bacteria (LAB) including Lactobacillus spp., have attracted considerable research interest due to their potential probiotic effects. The six Nepalese fermented foods selected for this research—homemade yogurt, commercial yogurt, gundruk, tama, chhurpi, and achaar—are recognized for their diverse microbial profiles and purported health benefits, commonly consumed across various regions of Nepal. This study aims to elucidate their probiotic potential through tests such as acid and salt tolerance and their antimicrobial effects against five pathogens (Staphylococcus aureus, Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa, and Salmonella spp.). A total of 120 samples of these six different fermented foods were collected from households and local markets in Chitwan District. Among 120 fermented food samples, 103 samples (85.83%) were found to be LAB positive. Out of the total sample, 64 samples (53.30%) were found to be Lactobacillus spp. positive. These positive isolates were further tested for their pH tolerance, NaCl tolerance, and antibacterial potential. Upon research, 18 isolates were found to be tolerant at pH 2, and 21 isolates were found to be tolerant at 8% NaCl concentration. The highest antibacterial effect of Lactobacillus spp. was found against Staphylococcus aureus with ZOI 18 mm and lowest against Escherichia coli. The study here contributes to the importance of utilizing the microbial diversity present in fermented foods, along with providing valuable insights for developing novel probiotic interventions for enhancing gut and overall health. Moreover, the research highlights the potential of traditional fermented foods as a valuable source of probiotic strains, paving the way for innovative approaches to improve health through diet.

The COVID-19 pandemic has affected higher education institutions (HEIs) worldwide and reshaped the existing educational system. Due to travel constraints and physical separation, there has been a global shift toward distance learning, and Nepal is no exception. This research intends to assess the practicality of online education by evaluating learners' experiences amidst COVID-19. A cross-sectional study was directed among HEIs students in Nepal using self-structured questionnaires. Our study revealed that 64.6% of the respondents were unsatisfied with online classes. More than half of the respondents (53.4%) use cell phones for online studies. Online education was reported to be unappealing to 28.8% of respondents. Variables such as age group (p = 0.05), enjoying class (p < 0.001), hours spent for an online class in a day (p = 0.05), and period for educational work using an electronic device (p = 0.1) were found significant with satisfaction level using both bivariate test and inferential test of univariate binary logistics regression. The challenges and opportunities encountered among students and faculties are highlighted along with the recommendations for fortifying communication in online-based teaching/learning.

Alzheimer's disease (AD) is a worldwide problem. Currently, there are no effective drugs for AD treatment. Scrophularia buergeriana Miquel (SB) is a traditional herbal medicine used in Korea to treat various diseases. Our previous studies have shown that ethanol extract of SB roots (SBE, Brainon®) exhibits potent anti-amnesic effects in Aβ1-42- or scopolamine-treated memory impairment mice model and neuroprotective effects in a glutamate-induced SH-SY5Y cell model. In this study, we evaluated the therapeutic effects of Brainon® and its mechanism of action in senescence-accelerated mouse prone 8 (SAMP8) mice. Brainon® (30 or 100 mg/kg/day) was orally treated to six-month-old SAMP8 mice for 12 weeks. Results revealed that Brainon® administration effectually ameliorated cognitive deficits in Y-maze and passive avoidance tests. Following the completion of behavioral testing, western blotting was performed using the cerebral cortex. Results revealed that Brainon® suppressed Aβ1-42 accumulation, Tau hyperphosphorylation, oxidative stress, and inflammation and alleviated apoptosis in SAMP8 mice. Brainon® also promoted synaptic function by downregulating the expression of AChE and upregulating the expression of p-CREB/CREB and BDNF. Furthermore, Brainon® restored SAMP8-reduced expression of ChAT and -dephosphorylated of ERK and also decreased AChE expression in the hippocampus. Furthermore, Brainon® alleviated AD progression by promoting mitophagy/autophagy to maintain normal cellular function as a novel finding of this study. Our data suggest that Brainon® can remarkably improve cognitive deficiency with the potential to be utilized in functional food for improving brain health.

Glioblastoma (GBM) is characterized by exceptionally high intratumoral heterogeneity. However, the molecular mechanisms underlying the origin of different GBM cell populations remain unclear. Here, we found that the compositions of ribosomes of GBM cells in the tumour core and edge differ due to alternative RNA splicing. The acidic pH in the core switches before messenger RNA splicing of the ribosomal gene RPL22L1 towards the RPL22L1b isoform. This allows cells to survive acidosis, increases stemness and correlates with worse patient outcome. Mechanistically, RPL22L1b promotes RNA splicing by interacting with lncMALAT1 in the nucleus and inducing its degradation. Contrarily, in the tumour edge region, RPL22L1a interacts with ribosomes in the cytoplasm and upregulates the translation of multiple messenger RNAs including TP53. We found that the RPL22L1 isoform switch is regulated by SRSF4 and identified a compound that inhibits this process and decreases tumour growth. These findings demonstrate how distinct GBM cell populations arise during tumour growth. Targeting this mechanism may decrease GBM heterogeneity and facilitate therapy.

Glioblastoma (GBM) is characterized by extensive microvascular hyperproliferation. In addition to supplying blood to the tumor, GBM vessels also provide trophic support to glioma cells and serve as conduits for migration into the surrounding brain, promoting recurrence. Here, we enrich CD31-expressing glioma vascular cells (GVCs) and A2B5-expressing glioma tumor cells (GTCs) from primary GBM and use RNA sequencing to create a comprehensive molecular interaction map of the secreted and extracellular factors elaborated by GVCs that can interact with receptors and membrane molecules on GTCs. To validate our findings, we utilize functional assays, including a hydrogel-based migration assay and in vivo mouse models to demonstrate that one identified factor, the little-studied integrin binding sialoprotein (IBSP), enhances tumor growth and promotes the migration of GTCs along the vasculature. This perivascular niche interactome will serve as a resource to the research community in defining the potential functions of the GBM vasculature.

This study was aimed to determine the antibacterial activity of root bark, leaves, and pericarp extract of Diploknema butyracea and to evaluate the prospective antioxidant activity, total flavonoid, polyphenol, and carbohydrate content. The plant parts were collected and extracted by cold maceration, using hexane, ethyl acetate, methanol, and distilled water. Phytochemical screening of different samples of D. butyracea in different solvents revealed the presence of varied extent of alkaloid, saponin, terpenoid, anthraquinones, tannin, cardiac glycoside, flavonoid, carbohydrate, polyphenol, protein and amino acid, resin, and phytosterol. Our study showed that methanolic root bark extract exhibited the potent antimicrobial activity against Staphylococcus aureus, Staphylococcus epidermidis, and Klebsiella pneumonia with an average zone of inhibition of 17.33 mm, 14.33 mm, and 13.0 mm, respectively. Surprisingly, all of the extracts were insensitive to Escherichia coli. The lowest minimum bactericidal concentration (MBC), 4.6 mg/ml, was observed with the aqueous pericarp extract against S. epidermidis and the highest was of 50 mg/ml shown by ethyl acetate pericarp against K. pneumonia. Our results showed that both the polar and nonpolar components present in the different parts of D. butyracea exhibit prominent antibacterial activities against different bacterial strains. The in vitro 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity showed that the methanol extract of root barks displayed the most potent antioxidant activity (IC50 : 6.1 µg/ml). The total polyphenol content of the plant part extracts was observed between 19.48 ± 0.23 and 123.48 ± 1.84 µg gallic acid equivalent/mg of dry extract weight. Likewise, flavonoid content ranged from 40.63 ± 1.28 µg to 889.72 ± 3.40 μg quercetin equivalent/mg of dry extract weight and total carbohydrate content ranged from 11.92 ± 0.60 µg to 174.72 ± 0.60 µg glucose equivalent per/mg dry extract weight. Overall, our study showed that the root bark, pericarp, and leaves extract of D. butyracea evinced prominent antibacterial properties against various pathogenic bacterial strains.

Parkinson's disease (PD) is characterized by loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) of the midbrain. Restoration of nigrostriatal dopamine neurons has been proposed as a potential therapeutic strategy for PD. Because currently used PD therapeutics only help relieve motor symptoms and do not treat the cause of the disease, highly effective drugs are needed. Vildagliptin, a dipeptidyl peptidase 4 (DPP4) inhibitor, is an anti-diabetic drug with various pharmacological properties including neuroprotective effects. However, the detailed effects of vildagliptin against PD are not fully understood. We investigated the effects of vildagliptin on PD and its underlying molecular mechanisms using a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mouse model and a 1-methyl-4-phenylpyridium (MPP+)-induced cytotoxicity model. Vildagliptin (50 mg/kg) administration significantly attenuated MPTP-induced motor deficits as evidenced by rotarod, pole, and nest building tests. Immunohistochemistry and Western blot analysis revealed that vildagliptin increased tyrosine hydroxylase-positive cells in the SNpc and striatum, which was reduced by MPTP treatment. Furthermore, vildagliptin activated MPTP-decreased PI3k/Akt and mitigated MPTP-increased ERK and JNK signaling pathways in the striatum. Consistent with signaling transduction in the mouse striatum, vildagliptin reversed MPP+-induced dephosphorylation of PI3K/Akt and phosphorylation of ERK and JNK in SH-SY5Y cells. Moreover, vildagliptin attenuated MPP+-induced conversion of LC3B-II in SH-SY5Y cells, suggesting its role in autophagy inhibition. Taken together, these findings indicate that vildagliptin has protective effects against MPTP-induced motor dysfunction by inhibiting dopaminergic neuronal apoptosis, which is associated with regulation of PI3k/Akt, ERK, and JNK signaling transduction. Our findings suggest vildagliptin as a promising repurposing drug to treat PD.

Protein quality control mechanisms play an important role in cancer progression by providing adaptive responses and morphologic stability against genome-wide copy number alterations, aneuploidy, and conformation-altering somatic mutations. This dependency on protein quality control mechanisms creates a vulnerability that may be exploited for therapeutic benefits by targeting components of the protein quality control mechanism. Recently, valosin-containing protein (VCP), also known at p97 AAA-ATPase, has emerged as a druggable target in cancer cells to affect their dependency on protein quality control. Here, we show that VCP inhibitors induce cytotoxicity in several ovarian cancer cell lines and these compounds act synergistically with mifepristone, a drug previously shown to induce an atypical unfolded protein response. Although mifepristone at a clinically achievable dose induces a weak unfolded protein response, it enhances the cytotoxic effects of VCP inhibitor CB-5083. Mechanistically, mifepristone blocks the cytoprotective effect of ATF6 in response to endoplasmic reticulum (ER) stress while activating the cytotoxic effects of ATF4 and CHOP through the HRI (EIF2AK1)-mediated signal transduction pathway. In contrast, CB-5083 activates ATF4 and CHOP through the PERK (EIF2AK3)-mediated signaling pathway. This combination activates ATF4 and CHOP while blocking the adaptive response provided by ATF6, resulting in increased cytotoxic effects and synergistic drug interaction.