Ephedra foeminea as a Novel Source of Antimicrobial and Anti-Biofilm Compounds to Fight Multidrug Resistance Phenotype.

Plants are considered a wealthy resource of novel natural drugs effective in the treatment of multidrug-resistant infections. Here, a bioguided purification of Ephedra foeminea extracts was performed to identify bioactive compounds. The determination of antimicrobial properties was achieved by broth microdilution assays to evaluate minimal inhibitory concentration (MIC) values and by crystal violet staining and confocal laser scanning microscopy analyses (CLSM) to investigate the antibiofilm capacity of the isolated compounds. Assays were performed on a panel of three gram-positive and three gram-negative bacterial strains. Six compounds were isolated from E. foeminea extracts for the first time. They were identified by nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) analyses as the well-known monoterpenoid phenols carvacrol and thymol and as four acylated kaempferol glycosides. Among them, the compound kaempferol-3-O-α-L-(2″,4″-di-E-p-coumaroyl)-rhamnopyranoside was found to be endowed with strong antibacterial properties and significant antibiofilm activity against S. aureus bacterial strains. Moreover, molecular docking studies on this compound suggested that the antibacterial activity of the tested ligand against S. aureus strains might be correlated to the inhibition of Sortase A and/or of tyrosyl tRNA synthase. Collectively, the results achieved open interesting perspectives to kaempferol-3-O-α-L-(2″,4″-di-E-p-coumaroyl)-rhamnopyranoside applicability in different fields, such as biomedical applications and biotechnological purposes such as food preservation and active packaging.

Transcriptomic Profiling of Human Limbus-Derived Stromal/Mesenchymal Stem Cells-Novel Mechanistic Insights into the Pathways Involved in Corneal Wound Healing.

Limbus-derived stromal/mesenchymal stem cells (LMSCs) are vital for corneal homeostasis and wound healing. However, despite multiple pre-clinical and clinical studies reporting the potency of LMSCs in avoiding inflammation and scarring during corneal wound healing, the molecular basis for the ability of LMSCs remains unknown. This study aimed to uncover the factors and pathways involved in LMSC-mediated corneal wound healing by employing RNA-Sequencing (RNA-Seq) in human LMSCs for the first time. We characterized the cultured LMSCs at the stages of initiation (LMSC-P0) and pure population (LMSC-P3) and subjected them to RNA-Seq to identify the differentially expressed genes (DEGs) in comparison to native limbus and cornea, and scleral tissues. Of the 28,000 genes detected, 7800 DEGs were subjected to pathway-specific enrichment Gene Ontology (GO) analysis. These DEGs were involved in Wnt, TGF-ß signaling pathways, and 16 other biological processes, including apoptosis, cell motility, tissue remodeling, and stem cell maintenance, etc. Two hundred fifty-four genes were related to wound healing pathways. COL5A1 (11.81 ± 0.48) and TIMP1 (20.44 ± 0.94) genes were exclusively up-regulated in LMSC-P3. Our findings provide new insights involved in LMSC-mediated corneal wound healing.

Molecular dissemination of emerging antibiotic, biocide, and metal co-resistomes in the Himalayan hot springs.

Growing resistance among microbial communities against antimicrobial compounds, especially antibiotics, is a significant threat to living beings. With increasing antibiotic resistance in human pathogens, it is necessary to examine the habitats having community interests. In the present study, a metagenomic approach has been employed to understand the causes, dissemination, and effects of antibiotic, metal, and biocide resistomes on the microbial ecology of three hot springs, Borong, Lingdem, and Yumthang, located at different altitudes of the Sikkim Himalaya. The taxonomic assessment of these hot springs depicted the predominance of mesophilic organisms, mainly belonging to the phylum Proteobacteria. The enriched microbial metabolism assosiated with energy, cellular processes, adaptation to diverse environments, and defence were deciphered in the metagenomes. The genes representing resistance to semisynthetic antibiotics, e.g., aminoglycosides, fluoroquinolones, fosfomycin, vancomycin, trimethoprim, tetracycline, streptomycin, beta-lactams, multidrug resistance, and biocides such as triclosan, hydrogen peroxide, acriflavin, were abundantly present. Various genes attributing resistance to copper, arsenic, iron, and mercury in metal resistome were detected. Relative abundance, correlation, and genome mapping of metagenome-assembled genomes indicated the co-evolution of antibiotic and metal resistance in predicted novel species belonging to Vogesella, Thiobacillus, and Tepidimona genera. The metagenomic findings were further validated with isolation of microbial cultures, exhibiting resistance against antibiotics and heavy metals, from the hot spring water samples. The study furthers our understanding about the molecular basis of co-resistomes in the ceological niches and their possible impact on the environment.

Metagenomics revealing molecular profiling of community structure and metabolic pathways in natural hot springs of the Sikkim Himalaya.

BACKGROUND: Himalaya is an ecologically pristine environment. The geo-tectonic activities have shaped various environmental niches with diverse microbial populations throughout the Himalayan biosphere region. Albeit, limited information is available in terms of molecular insights into the microbiome, including the uncultured microbes, of the Himalayan habitat. Hence, a vast majority of genomic resources are still under-explored from this region. Metagenome analysis has simplified the extensive in-depth exploration of diverse habitats. In the present study, the culture-independent whole metagenome sequencing methodology was employed for microbial diversity exploration and identification of genes involved in various metabolic pathways in two geothermal springs located at different altitudes in the Sikkim Himalaya. RESULTS: The two hot springs, Polok and Reshi, have distinct abiotic conditions. The average temperature of Polok and Reshi was recorded to be 62 °C and 43 °C, respectively. Both the aquatic habitats have alkaline geochemistry with pH in the range of 7-8. Community profile analysis revealed genomic evidence of plentiful bacteria, with a minute fraction of the archaeal population in hot water reservoirs of Polok and Reshi hot spring. Mesophilic microbes belonging to Proteobacteria and Firmicutes phyla were predominant at both the sites. Polok exhibited an extravagant representation of Chloroflexi, Deinococcus-Thermus, Aquificae, and Thermotogae. Metabolic potential analysis depicted orthologous genes associated with sulfur, nitrogen, and methane metabolism, contributed by the microflora in the hydrothermal system. The genomic information of many novel carbohydrate-transforming enzymes was deciphered in the metagenomic description. Further, the genomic capacity of antimicrobial biomolecules and antibiotic resistance were discerned. CONCLUSION: The study provided comprehensive molecular information about the microbial treasury as well as the metabolic features of the two geothermal sites. The thermal aquatic niches were found a potential bioresource of biocatalyst systems for biomass-processing. Overall, this study provides the whole metagenome based insights into the taxonomic and functional profiles of Polok and Reshi hot springs of the Sikkim Himalaya. The study generated a wealth of genomic data that can be explored for the discovery and characterization of novel genes encoding proteins of industrial importance.

Structural Characteristics, Physicochemical Properties, and Digestibility Analysis of Resistant Starch Type-V Prepared from Debranched Corn Starch and Fatty Acid Complexation.

Corn starch was gelatinized and treated with a metagenomic type 1 pullulanase (PulM), increasing the proportion of linear glucan chains. The debranched corn starch (DCS), containing amylose helices, was subjected to complexation with fatty acid molecules at moderate temperatures (50-60 °C). The amylose-lipid complexes prepared using saturated fatty acids, e.g., capric acid (CA) and lauric acid (LA), displayed higher CI values as compared to that of unsaturated fatty acid compounds, e.g., undecylenic acids (UAs) and oleic acid (OA). The DCS-fatty acid complex was estimated to contain about 14% of rapidly digested starch (RDS), 26% of slowly digested starch (SDS), and 60% of resistant starch V (RS-5). RS-5 samples exhibited high resistance toward digestive enzymatic hydrolysis. The surface microdetails of RS-5 were examined by scanning electron microscopy (SEM), depicting small spherulite-like structural aggregates. X-ray diffraction pattern analysis estimated about 46% of the crystallinity of RS-5. Thermal attributes of RS-5 were examined by thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) analysis, depicting the increase in melting enthalpies after the complexation of fatty acid molecules with debranched corn starch. Comparative DSC thermograms divulged a relatively higher stability of RS-5 as compared to that of RS-3. The findings advocated the potentiality of RS-5 (nondigestible DCS-LA complex) as a functional, valuable ingredient in the food industry.

Corneal stem cells niche and homeostasis impacts in regenerative medicine; concise review.

The limbal stem cells niche (LSCN) is an optimal microenvironment that provides the limbal epithelial stem cells (LESCs) and strictly regulates their proliferation and differentiation. Disturbing the LSCN homeostasis can lead to limbal stem cell dysfunction (LSCD) and subsequent ocular surface aberrations, such as corneal stromal inflammation, persistent epithelial defects, corneal neovascularisation, lymphangiogenesis, corneal opacification, and conjunctivalization. As ocular surface disorders are considered the second main cause of blindness, it becomes crucial to explore different therapeutic strategies for restoring the functions of the LSCN. A major limitation of corneal transplantation is the current shortage of donor tissue to meet the requirements worldwide. In this context, it becomes mandatory to find an alternative regenerative medicine, such as using cultured limbal epithelial/stromal stem cells, inducing the production of corneal like cells by using other sources of stem cells, and using tissue engineering methods aiming to produce the three-dimensional (3D) printed cornea. Limbal epithelial stem cells have been considered the magic potion for eye treatment. Epithelial and stromal stem cells in the limbal niche hold the responsibility of replenishing the corneal epithelium. These stem cells are being used for transplantation to maintain corneal epithelial integrity and ultimately sustain optimal vision. In this review, we summarised the characteristics of the LSCN and their current and future roles in restoring corneal homeostasis in eyes with LSCD.

Moringa oleifera impedes protein glycation and exerts reno-protective effects in streptozotocin-induced diabetic rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Moringa oleifera is a valued plant with wide distribution in tropical and subtropical regions of the world. It is traditionally used for the treatment of fever, infections, rheumatism, cancer, improving cardiac, renal and hepatic functions, and regulating blood glucose level. The plant has been scientifically reported for the anti-inflammatory, antioxidant, renoprotective, and anti-diabetic properties. Diabetic patients are prone to develop end-stage renal diseases due to incidence of diabetes-induced renal dysfunctions. Given that, increased production and accumulation of advanced glycation end-products (AGEs) play a conspicuous role in the development of diabetes-linked renal dysfunctions, nature-based interventions with AGEs inhibitory activity can prevent renal dysfunctions leading to renoprotection. AIM OF THE STUDY: The study aimed to demonstrate the preventive effects of the ethanolic extract of the leaves of Moringa oleifera (EEMO) on protein glycation and its further assessment for the renoprotective effect in diabetic rats. MATERIALS AND METHODS: Antiglycation activity of EEMO was assessed in vitro using bovine serum albumin. For reno-protective activity assessment, streptozotocin (STZ)-induced diabetic rats were orally treated with EEMO (100 mg/kg) or standard antiglycation agent aminoguanidine (100 mg/kg) for consecutive 8 weeks. The effects on glucose homeostasis, renal functions, and renal morphology were assessed by clinical biochemistry, molecular and histological examination. RESULTS: Presence of EEMO efficiently prevented glucose-, fructose- or methylglyoxal-mediated glycation of protein. Under in vivo set-up, compared to diabetic control rats, EEMO treatment effectively improved the glucose tolerance and body weight, and reduced the serum levels of triglycerides and total cholesterol. Additionally, EEMO administration significantly ameliorated renal dysfunctions in diabetic rats characterized by improved levels of creatinine, urea nitrogen, and uric acid in serum, and total protein level in urine, accompanied by improved kidney morphology. The diabetes-associated pro-inflammatory response characterized by upregulated expression of the inducible nitric oxide synthase (iNos), activation of nuclear factor kappa B (NF-κB) and the raised levels of inflammatory factors, interleukin-1 beta (IL-1ß) and interleukin-6 (IL-6) in renal tissue was significantly attenuated in EEMO-treated rats. Moreover, EEMO treatment diminished renal reactive oxygen species (ROS) levels in diabetic animals. CONCLUSIONS: Our study demonstrated that EEMO prevented AGEs formation and ameliorated renal dysfunctions in diabetic rats by blocking inflammatory/oxidative pathways. Our observations justify M. oleifera as a potential source of therapeutic interventions for diabetic nephropathy management.

Ashwagandha (Withania somnifera) and Shunthi (Zingiber officinale) in mild and moderate COVID-19: An open-label randomized controlled exploratory trial.

BACKGROUND: Ayurveda interventions have been used for prophylaxis and care during the COVID-19 pandemic in India and have shown promising results in promoting early clinical recovery from COVID-19. OBJECTIVE: To assess the efficacy and safety of Ashwagandha [Withania somnifera (L.) Dunal] tablet and Shunthi (Zingiber officinale Roscoe) capsule in mild and moderate COVID-19 compared to conventional standard care. METHODS: A randomized controlled exploratory trial was conducted at a designated COVID-19 care center in India with 60 participants having mild or moderate COVID-19. Ashwagandha, two tablets (250 mg each), and Shunthi, two capsules (500 mg each) twice daily for 15 days, were given orally to the participants in the Ayurveda group (AG) and the control group (CG) received conventional standard care. The outcome measures included clinical recovery rate, the proportion of participants with negative RT-PCR assay for COVID-19 on day 7 and day 15, mean time to attain clinical recovery, change in pro-inflammatory markers, serum IgG for COVID-19, HRCT chest findings, disease progression and incidence of adverse events (AE). RESULTS: A total of 60 participants were enrolled, and the data of 48 participants (AG = 25 and CG = 23) were considered for the statistical analysis. The mean time for clinical recovery was reduced by almost 50 % in the AG (6.9 days) compared to CG (13.0 days) (p < 0.001). The proportion of participants who attained viral clearance in AG was 76.0 % compared to 60.8 % in the CG (RR= 1.24, 95 % CI: 0.841, 1.851, p-value = 0.270). Changes in the pro-inflammatory markers, serum IgG for COVID-19, and HRCT chest findings were comparable in both groups, and no AE or disease progression was reported. CONCLUSIONS: The Ayurveda interventions, Ashwagandha and Shunthi, can effectively reduce the duration of clinical recovery and improve time for viral clearance in mild and moderate COVID-19. These interventions were observed to be safe and well-tolerated during the duration of the trial. TRIAL REGISTRATION: Clinical Trial Registry of India - CTRI/2020/08/027224.

Biodegradation of plastics for sustainable environment.

Plastics are a kind of utility product that has become part and parcel of one's life. Their continuous usage, accumulation, and contamination of soil and water pose a severe threat to the biotic and abiotic components of the environment. It not only increases the carbon footprints but also contributes to global warming. This calls for an urgent need to develop novel strategies for the efficient degradation of plastics. The microbial strains equipped with the potential of degrading plastic materials, which can further be converted into usable products, are blessings for the ecosystem. This review comprehensively summarizes the microbial technologies to degrade different plastic types, such as polyethylene (PE), polyethylene terephthalate (PET), polystyrene (PS), polyvinyl chloride (PVC), polypropylene (PP), and polyurethane (PU). The study also describes the utilization of degraded plastic material as feedstock for its conversion into high-value chemicals.

Insulin resistance corresponds with a progressive increase in NOD1 in high fat diet-fed mice.

PURPOSE: Innate immune components participate in obesity-induced inflammation, which can contribute to endocrine dysfunction during metabolic diseases. However, the chronological activation of specific immune proteins such as Nucleotide-binding oligomerization domain-containing protein 1 (NOD1) and relevance to cellular crosstalk during the progression of obesity-associated insulin resistance (IR) is not known. METHODS: The NOD1 signaling in various insulin-sensitive metabolic tissues during the progression of diet-insulin resistance was assessed in C57BL/6J mice fed with 60% high-fat diet (HFD) for 4, 8, 12, and 16 weeks. Intestinal permeability was measured using FITC-dextran. NOD1 activating potential was analyzed using HEK-Blue mNOD1 cells. RESULTS: HFD-fed mice showed progressive induction of glucose intolerance and impairment of insulin signaling in key metabolic tissues. We found a time-dependent increase in intestinal permeability coupled with transport and accumulation of NOD1 activating ligand in the serum of HFD-fed mice. We also observed a progressive accumulation of γ-D-glutamyl-meso-diaminopimelic acid (DAP), a microbial peptidoglycan ligand known to activate NOD1, in serum samples of the HFD-fed mice. There was also a progressive increase in transcripts levels of NOD1 in bone marrow-derived macrophages during HFD-feeding. In addition, skeletal muscle, adipose and liver, the key insulin sensitive metabolic tissues also had a time-dependent increase in transcripts of NOD1 and Rip2 and a corresponding activation of pro-inflammatory responses in these tissues. CONCLUSION: These data highlight the correlation of inflammation and insulin resistance to NOD1 activation in the bone marrow derived macrophages and insulin responsive metabolic tissues during high fat diet feeding in mice.

AYUSH-64 as an adjunct to standard care in mild to moderate COVID-19: An open-label randomized controlled trial in Chandigarh, India.

OBJECTIVE: To determine the therapeutic efficacy and safety of AYUSH-64 as an add-on to standard care in mild to moderate COVID-19. DESIGN SETTING, AND INTERVENTIONS: This open-label randomized controlled parallel-group trial was conducted at a designated COVID care centre in India in 80 patients diagnosed with mild to moderate COVID-19 and randomized into two groups. Participants in the AYUSH-64 add-on group (AG) received AYUSH-64 two tablets (500 mg each) three times a day for 30 days along with standard conventional care. The control group (CG) received standard care alone. MAIN OUTCOME MEASURES: Proportion of participants who attained clinical recovery on day 7, 15, 23 and 30, proportion of participants with negative RT-PCR assay for COVID-19 at each weekly time point, change in pro-inflammatory markers, metabolic functions, HRCT chest (CO-RADS category) and incidence of Adverse Drug Reaction (ADR)/Adverse Event (AE). RESULTS: Out of 80 participants, 74 (37 in each group) contributed to the final analysis. Significant difference was observed in clinical recovery in the AG (p < 0.001 ) compared to CG. Mean duration for clinical recovery in AG (5.8 ± 2.67 days) was significantly less compared to CG (10.0 ± 4.06 days). Significant improvement in HRCT chest was observed in AG (p = 0.031) unlike in CG (p = 0.210). No ADR/SAE was observed or reported in AG. CONCLUSIONS: AYUSH-64 as adjunct to standard care is safe and effective in hastening clinical recovery in mild to moderate COVID-19. The efficacy may be further validated by larger multi-center double-blind trials.

A novel ß-glucosidase from a hot-spring metagenome shows elevated thermal stability and tolerance to glucose and ethanol.

ß-glucosidase causes hydrolysis of ß-1,4-glycosidic bond in glycosides and oligosaccharides. It is an industrially important enzyme owing to its potential in biomass processing applications. In this study, computational screening of an extreme temperature aquatic habitat metagenomic resource was done, leading to the identification of a novel gene, bglM, encoding a ß-glucosidase. The comparative protein sequence and homology structure analyses designated it as a GH1 family ß-glucosidase. The bglM gene was expressed in a heterologous host, Escherichia coli. The purified protein, BglM, was biochemically characterized for ß-glucosidase activity. BglM exhibited noteworthy hydrolytic potential towards cellobiose and lactose. BglM, showed substantial catalytic activity in the pH range of 5.0-7.0 and at the temperature 40 °C-70 °C. The enzyme was found quite stable at 50 °C with a loss of hardly 20% after 40 h of heat exposure. Furthermore, any drastically negative effect was not observed on the enzyme's activity in the presence of metal ions, non-ionic surfactants, metal chelating, and denaturing agents. A significantly high glucose tolerance, retaining 80% relative activity at 1 M, and 40% at 5 M glucose, and ethanol tolerance, exhibiting 80% relative activity in 10% ethanol, enrolled BglM as a promising enzyme for cellulose saccharification. Furthermore, its ability to catalyze the hydrolysis of daidzin and polydatin ascertained it as an admirably suited biocatalyst for enhancement of nutritional values in soya and wine industries.

Biotransformation of hydroquinone into α-arbutin by transglucosylation activity of a metagenomic amylosucrase.

Arbutin is a naturally occurring glycosylated product of hydroquinone. With the ability to interrupt melanin biosynthesis in epidermal cells, it is a promising cosmetic ingredient. In this study, a novel amylosucrase, Asmet, identified from a thermal spring metagenome, has been characterized for arbutin biosynthesis. Asmet was able to catalyze transglucosylation of hydroquinone to arbutin, taking sucrose as glycosyl donor, in the temperature range of 20 °C to 40 °C and pH 5.0 to 6.0, with the relative activity of 80% or more. The presence of chloride salts of Li, K, and Na at 1 mM concentration did not exhibit any notable effect on the enzyme's activity, unlike Cu, Ni, and Mn, which were observed to be detrimental. The hydroquinone (20 mM) to sucrose ratio of 1:1 to 1:10 was appropriate for the catalytic biosynthesis of arbutin. The maximum hydroquinone to arbutin conversion of 70% was obtained in 24 h of Asmet led catalysis, at 30 °C and pH 6.0. Arbutin production was also demonstrated using low-cost feedstock, table sugar, muscovado, and sweet sorghum stalk extract, as a replacement for sucrose. Whole-cell catalysis of hydroquinone to arbutin transglucosylation was also established.

A novel cold-active type I pullulanase from a hot-spring metagenome for effective debranching and production of resistant starch.

Pullulanase is a potent enzyme for starch debranching. In this study, a novel type I pullulanase (PulM) was identified from the metagenome of a thermal aquatic habitat that exhibits optimal activity of debranching at 40 °C temperature and pH 6.0 to 7.0. More than 50% enzymatic activity was detected at the low temperature of 4 °C, determining it a cold-active type I pullulanase. It was able to efficiently catalyze the hydrolysis of α-1,6-glycosidic linkages in pullulan, with a specific activity of 177 U mg-1. The results determined PulM to be a potential starch debranching biocatalyst, causing a significant increase of about 80% in the apparent amylose content of potato starch. Retrogradation of the debranched starch resulted in the formation of resistant starch 3. The yield of resistant starch was estimated to be about 45%. The resistant starch exhibited higher crystallinity, enhanced heat-stability, and resistance to α-amylase digestion, as compared to native starch.

A Multifunctional Peptide From Bacillus Fermented Soybean for Effective Inhibition of SARS-CoV-2 S1 Receptor Binding Domain and Modulation of Toll Like Receptor 4: A Molecular Docking Study.

Fermented soybean products are traditionally consumed and popular in many Asian countries and the northeastern part of India. To search for potential agents for the interruption of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Spike glycoprotein 1 (S1) and human angiotensin-converting enzyme 2 (ACE2) receptor interactions, the in silico antiviral prospective of peptides identified from the proteome of kinema was investigated. Soybean was fermented using Bacillus licheniformis KN1G, Bacillus amyloliquefaciens KN2G and two different strains of Bacillus subtilis (KN2B and KN2M). The peptides were screened in silico for possible antiviral activity using two different web servers (AVPpred and meta-iAVP), and binding interactions of selected 44 peptides were further explored against the receptor-binding domain (RBD) of the S1 protein (PDB ID: 6M0J) by molecular docking using ZDOCK. The results showed that a peptide ALPEEVIQHTFNLKSQ (P13) belonging to B. licheniformis KN1G fermented kinema was able to make contacts with the binding motif of RBD by blocking specific residues designated as critical (GLN493, ASN501) in the binding of human angiotensin-converting enzyme 2 (ACE2) cell receptor. The selected peptide was also observed to have a significant affinity towards human toll like receptor 4 (TLR4)/Myeloid Differentiation factor 2 (MD2) (PDB ID: 3FXI) complex known for its essential role in cytokine storm. The energy properties of the docked complexes were analyzed through the Generalized Born model and Solvent Accessibility method (MM/GBSA) using HawkDock server. The results showed peptidyl amino acids GLU5, GLN8, PHE11, and LEU13 contributed most to P13-RBD binding. Similarly, ARG90, PHE121, LEU61, PHE126, and ILE94 were appeared to be significant in P13-TLR4/MD2 complex. The findings of the study suggest that the peptides from fermented soy prepared using B. licheniformis KN1G have better potential to be used as antiviral agents. The specific peptide ALPEEVIQHTFNLKSQ could be synthesized and used in combination with experimental studies to validate its effect on SARS-CoV-2-hACE2 interaction and modulation of TLR4 activity. Subsequently, the protein hydrolysate comprising these peptides could be used as prophylaxis against viral diseases, including COVID-19.

Isolation and Characterization of an Endophytic Fungus Colletotrichum coccodes Producing Tyrosol From Houttuynia cordata Thunb. Using ITS2 RNA Secondary Structure and Molecular Docking Study.

An endophytic fungus isolated from healthy leaf tissues of Houttuynia cordata Thunb., an ethnomedicinal plant of North East India, showed a considerable amount of antimicrobial activity. The ethyl acetate extract of the fungal culture filtrates displayed promising antimicrobial activity against a panel of clinically significant pathogens including Candida albicans, Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. Bioassay guided purification of the organic extract using column and thin layer chromatography yielded a pure homogenous compound which was identified using spectroscopic methods (essentially by 1H NMR and MS) as tyrosol, a well-known phenylethanoid present in several natural sources. Besides, molecular docking studies against tyrosyl tRNA synthetases (TyrRS) of S. aureus (PDB ID: 1JIL) and E. coli (PDB ID: 1VBM), and CYP45014α-lanosterol demethylase (CYP51) of C. albicans (PDB ID: 5FSA) revealed tyrosol has a strong binding affinity with the enzyme active site residues. The fungus was identified as Colletotrichum sp. and characterized by its genomic ITS rDNA and ITS2 sequences. Phylogenetic analyses showed clustering of our isolate with Colletotrichum coccodes. Species of Colletotrichum are also reported to be plant pathogens. Therefore, to confirm the endophytic lifestyle of the isolate, ITS2 RNA secondary structure study was undertaken. The result indicated our isolate exhibited differences in the folding pattern as well as in motif structures when compared to those of pathogenic C. coccodes. The findings indicated that endophytic fungi harboring H. cordata could be explored as a potent source of antimicrobial agents.

Flavonoids from Polyalthia longifolia prevents advanced glycation end products formation and protein oxidation aligned with fructose-induced protein glycation.

Advanced glycation end products (AGEs) are reactive chemical entities formed by non-enzymatic reaction between reducing sugars and amino group of proteins. Enhanced accumulation of AGEs and associated protein oxidation contribute to pathogenesis of diabetes-associated complications. Here, we evaluated the inhibitory activity of flavonoid compounds isolated from the leaves of Polyalthia longifolia on formation of AGEs and protein oxidation. Antiglycation activity was determined by measuring the formation of AGE fluorescence intensity, Nε-(carboxymethyl) lysine, and level of fructosamine. Protein oxidation was examined using levels of protein carbonyls and thiol group. Compounds significantly (p < 0.001) restricted the formation of fluorescent AGEs in fructose- BSA and methylglyoxal-BSA systems. Furthermore, there was a decrease in levels of fructosamine and protein carbonyls, and elevation in level of thiol group in fructose-BSA in presence of flavonoids. In summary, flavonoids from Polyalthia longifolia inhibit fructose-mediated protein glycation and oxidation, and can be potential agent for preventing AGE-mediated diabetic complications.

Current Status and Potential Therapeutic Strategies for Using Non-coding RNA to Treat Diabetic Cardiomyopathy.

Diabetic cardiomyopathy (DMCM) is the leading cause of mortality and morbidity among diabetic patients. DMCM is characterized by an increase in oxidative stress with systemic inflammation that leads to cardiac fibrosis, ultimately causing diastolic and systolic dysfunction. Even though DMCM pathophysiology is well studied, the approach to limit this condition is not met with success. This highlights the need for more knowledge of underlying mechanisms and innovative therapies. In this regard, emerging evidence suggests a potential role of non-coding RNAs (ncRNAs), including micro-RNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs) as novel diagnostics, mechanisms, and therapeutics in the context of DMCM. However, our understanding of ncRNAs' role in diabetic heart disease is still in its infancy. This review provides a comprehensive update on pre-clinical and clinical studies that might develop therapeutic strategies to limit/prevent DMCM.

Nod1-mediated lipolysis promotes diacylglycerol accumulation and successive inflammation via PKCδ-IRAK axis in adipocytes.

Chronic inflammation contributes to obesity mediated metabolic disturbances, including insulin resistance. Obesity is associated with altered microbial load in metabolic tissues that can contribute to metabolic inflammation. Different bacterial components such as, LPS, peptidoglycans have been shown to underpin metabolic disturbances through interaction with host innate immune receptors. Activation of Nucleotide-binding oligomerization domain-containing protein 1 (Nod1) with specific peptidoglycan moieties promotes insulin resistance, inflammation and lipolysis in adipocytes. However, it was not clear how Nod1-mediated lipolysis and inflammation is linked. Here, we tested if Nod1-mediated lipolysis caused accumulation of lipid intermediates and promoted cell autonomous inflammation in adipocytes. We showed that Nod1-mediated lipolysis caused accumulation of diacylglycerol (DAG) and activation of PKCδ in 3T3-L1 adipocytes, which was prevented with a Nod1 inhibitor. Nod1-activated PKCδ caused downstream stimulation of IRAK1/4 and was associated with increased expression of proinflammatory cytokines such as, IL-1ß, IL-18, IL-6, TNFα and MCP-1. Pharmacological inhibition or siRNA mediated knockdown of IRAK1/4 attenuated Nod1-mediated activation of NF-κB, JNK, and the expression of proinflammatory cytokines. These results reveal that Nod1-mediated lipolysis promoted accumulation of DAG, which engaged PKCδ and IRAK1/4 to augment inflammation in 3T3-L1 adipocytes.

Metagenomic Insights Into the Taxonomic and Functional Features of Kinema, a Traditional Fermented Soybean Product of Sikkim Himalaya.

Kinema is an ethnic, naturally fermented soybean product consumed in the Sikkim Himalayan region of India. In the present study, the whole metagenome sequencing approach was adopted to examine the microbial diversity and related functional potential of Kinema, consumed in different seasons. Firmicutes was the abundant phylum in Kinema, ranging from 82.31 to 93.99% in different seasons, followed by Actinobacteria and Proteobacteria. At the species level, the prevalent microorganisms were Bacillus subtilis, Bacillus amyloliquefaciens, Bacillus licheniformis, Corynebacterium glutamicum, Bacillus pumilus, and Lactococcus lactis. The abundance of microbial species varied significantly in different seasons. Further, the genomic presence of some undesirable microbes like Bacillus cereus, Proteus mirabilis, Staphylococcus aureus, Proteus penneri, Enterococcus faecalis, and Staphylococcus saprophyticus, were also detected in the specific season. The metagenomic analysis also revealed the existence of bacteriophages belonging to the family Siphoviridae, Myoviridae, and Podoviridae. Examination of the metabolic potential of the Kinema metagenome depicted information about the biocatalysts, presumably involved in the transformation of protein and carbohydrate polymers into bioactive molecules of health-beneficial effects. The genomic resource of several desirable enzymes was identified, such as ß-galactosidase, ß-glucosidase, ß-xylosidase, and glutamate decarboxylase, etc. The catalytic function of a novel glutamate decarboxylase gene was validated for the biosynthesis of γ-aminobutyric acid (GABA). The results of the present study highlight the microbial and genomic resources associated with Kinema, and its importance in functional food industry.