Manoj Kushwaha

The time-consuming nature of conventional bacterial identification methods, coupled with rising resistance to antibiotics, presents a serious challenge in the rapid diagnosis and treatment of patients suffering from severe infection and sepsis. In this study, we demonstrate that the MasSpec Pen (MSPen) technology enables rapid identification of the most frequently occurring bacterial species in human infections. A total of 755 clinical bacterial isolates prospectively collected from pediatric and adult patients were analyzed using the MSPen to create a library of molecular profiles. The mass spectra obtained presented distinct molecular patterns for bacterial species, with high relative abundances of bacterial metabolites and lipids, including unique quinolone signaling molecules in Pseudomonas aeruginosa , phosphoethanolamine (PE) lipids in Enterobacteriaceae , and phosphatidylglycerol (PG) lipids in Staphylococcus species. Logistic and multinomial lasso models using relative abundances or logarithmic-ratio abundance calculations achieved >95% accuracy for Gram stain typing and >92% for species-level identification in isolates. Random forest models yielded comparable performance, reinforcing the robustness of these findings across approaches. Many of the metabolite and lipid predictive features from each isolate classifier were detected when directly analyzing highly infected synovial fluid, tissue, pleural fluid, and breast aspirate specimens from patients. Notably, significant variations in the natural isotopic abundance of specific lipids were observed in infected patient samples, indicating presence of bacterial infection. This study demonstrates the potential of MSPen for the direct identification of bacterial-specific metabolites and lipids in clinical isolates and specimens, offering promise for future advancements in the MSPen technology for rapid bacterial identification. Teaser.: MSPen shows potential for rapid, direct identification of bacterial metabolites and lipids, enabling accurate bacterial classification.

This manuscript reports a chemical investigation involving reverse-phase semipreparative HPLC-based isolation, followed by mass spectrometric and NMR-based characterization of three endophytic fungi recovered from Albizia lebbeck (L.) Benth. The study yielded two compounds (1-2) from ALE-75 (Sacharicola bicolor), three (3-5) from ALE-85 (Aspergillus pseudoglaucus), and one (6) from ALE-121 (Aspergillus eurotium). The extract from Aspergillus pseudoglaucus exhibited antioxidant potential (IC50 = 11.92 µg/mL in DPPH assay) and moderate cytotoxicity against lung (A549, IC50 = 35 µg/ml), colon (HCT-116, IC50 value of 45.4 µg/ml), and breast (T47D, IC50 value of 2.8 µg/ml) cancer cells, with growth inhibition of 89%, 77%, and 74% at 100 µg/ml. ALE-121 extract showed strong cytotoxicity against lung (A549, IC50 = 11.2 µg/ml), colon (HCT-116, IC50 = 12.5 µg/ml) and breast (T47D, IC50 = 15 µg/ml) cancer cells, with inhibition rates of 85%, 84%, and 75% at 100 µg/ml. Compound 6 (Austrocortinin) showed strong cytotoxicity against these cancer lines, with inhibition rates of 89%, 87%, and 67% at 10 µg/ml. Compound 1 was initially identified through the fermentation process of the endophytic fungus Coriolopsis sp. J5, which was isolated from the mangrove Ceriops tagal and compound 2 in Penicillium chrysogenum LF066 from the marine sponge Tethya aurantium. Compounds 3-5, originally found in Chaetomium globosum and Aspergillus flavus, whereas Compound 6 was initially discovered in the Australian Cortinarius toadstool. Although these compounds have been identified previously, this is the first instance of their reporting in the aforementioned endophytic fungi isolated from Albizia lebbeck.

In this study, we employed liquid chromatography-tandem mass spectrometry (LC-MS/MS) to perform chemo-profiling on four fungal species: three Trichoderma species─two strains of Trichoderma asperellum (S3-3 and S3-1) and one Trichoderma longibrachiatum (FS-46)─and one species from the Tolypocladium genus, Tolypocladium inflatum (MTCC-3538). Our analysis identified 78 peptaibols featuring α-aminoisobutyric acid (Aib). Among these, we discovered both cyclic and linear Peptaibols and lipopeptaibols with Aib, ranging in putative molecular weights from 1033.4 to 1953.6 g/mol, corresponding to peptide lengths of 10-20 amino acids. In total, 78 peptaibols, including lipopeptaibols, were identified through LC-MS/MS. Of these, 43 were recognized as new peptaibols based on mass fragmentation patterns and sequence data from the MS/MS analysis. Our findings suggest that LC-MS/MS-based large-scale studies of the peptaibiome will enable the discovery of novel peptaibols and provide a valuable reference for understanding fragmentation patterns of peptaibols, cyclic peptaibols, and lipopeptaibols. Moreover, LC-MS/MS allows rapid identification of peptaibiotics at low concentrations directly in extracts, offering a potential quality control method for selecting beneficial Trichoderma strains for crop protection, while their fragmentation patterns and plausible sequences can guide synthetic design.

In this study, we employed liquid chromatography–tandem mass spectrometry (LC–MS/MS) to perform chemo-profiling on four fungal species: three Trichoderma species—two strains of Trichoderma asperellum (S3-3 and S3-1) and one Trichoderma longibrachiatum (FS-46)—and one species from the Tolypocladium genus, Tolypocladium inflatum (MTCC-3538). Our analysis identified 78 peptaibols featuring α-aminoisobutyric acid (Aib). Among these, we discovered both cyclic and linear Peptaibols and lipopeptaibols with Aib, ranging in putative molecular weights from 1033.4 to 1953.6 g/mol, corresponding to peptide lengths of 10-20 amino acids. In total, 78 peptaibols, including lipopeptaibols, were identified through LC-MS/MS. Of these, 43 were recognized as new peptaibols based on mass fragmentation patterns and sequence data from the MS/MS analysis. Our findings suggest that LC-MS/MS-based large-scale studies of the peptaibiome will facilitate the discovery of novel peptaibols and provide a valuable reference for understanding the fragmentation patterns of peptaibols, cyclic peptaibols, and lipopeptaibols.

The bioassay-guided fractionation of the extract of aerial parts of Enicostemma littorale resulted in two fractions 3 and 4 with moderate and potent antioxidant activity, respectively. The purification of fraction 3 gave swertiamarin (1), while the LCMS profile of fraction 4 unveiled the presence of another constituent along with swertiamarin. The extensive purification of fraction 4 led to the unusual isolation of mangiferin (2) from E. littorale. The uncommon isolation of mangiferin from E. littorale motivated us to conduct its in silico and in vitro screening as an anti-inflammatory agent. Both studies have proved mangiferin to be a promising anti-inflammatory molecule with a binding energy of -9.17 kcal/mol against Cyclooxygenase-2 protein and IC₅₀ of 146.07 nanomolar. This study is the first report of the isolation of mangiferin, a xanthone glycoside from E. littorale.Communicated by Ramaswamy H. Sarma.

[This corrects the article DOI: 10.1021/acsomega.2c03037.].

The Rosellinia sanctae-cruciana extract was subjected to detailed liquid chromatography tandem mass spectrometry studies. A total of 38 peaks were annotated to m/z 508.26, m/z 510.28, m/z 524.26, m/z 526.28, m/z 540.26, m/z 542.27, and m/z 584.28 [M + H]⁺. The accurate mass, mutually supported UV/vis spectra, and database search identified these compounds as cytochalasins. Systematic dereplication helped identify a peak at m/z 540.26 [M + H]⁺ as the new compound. Further, the identified compound was purified by high-performance liquid chromatography and characterized by 2D NMR to be 19,20-epoxycytochalasin N1, a new optical isomer of 19,20-epoxycytochalasin-N. It exhibited substantial cytotoxicity with IC₅₀ values ranging from 1.34 to 19.02 μM. This study shows a fast approach for dereplicating and identifying novel cytochalasin metabolites in crude extracts.

Velutibol A (1), a new 14-residue peptaibol was isolated from the Himalayan cold habitat fungus Trichoderma velutinum. The structural characterization was carried out by 1D and 2D NMR studies, and tandem mass studies, and Marfey's method aided in determining the stereochemistry of the amino acids. The CD analysis revealed folding of the peptide in a 3₁₀-helical conformation. The intramolecular H-bonding was determined by an NMR-VT experiment. Cytotoxic evaluation was carried out against a panel of cancer cell lines. The cell cycle assay was carried out on human myeloid leukaemia (HL-60) cells and revealed the formation of apoptotic bodies and DNA damage in a dose-dependent manner. Three other peptaibols namely velutibol B (2), velutibol C (3), and velutibol D (4) were also isolated in trace amounts from the psychotropic fungus and characterized through tandem mass spectroscopy and Marfey's analysis.

Present study relates to the effect of valproic acid, an epigenetic modifier on the metabolic profile of Aspergillus fumigatus (GA-L7), an endophytic fungus isolated from Grewia asiatica L. Seven secondary metabolites were isolated from A. fumigatus (GA-L7) which were identified as: pseurotin A, pseurotin D, pseurotin F2, fumagillin, tryprostatin C, gliotoxin and bis(methylthio)gliotoxin. Addition of valproic acid in the growth medium resulted in the alteration of secondary metabolic profile with an enhanced production of a metabolite, fumiquinazoline C by tenfolds. In order to assess the effect of valproic acid on the biosynthetic pathway of fumiquinazoline C, we studied the expression of the genes involved in its biosynthesis, both in the valproic acid treated and untreated control culture. The results revealed that all the genes i.e. Afua_6g 12040, Afua_6g 12050, Afua_6g 12060, Afua_6g 12070 and Afua_6g 12080, involved in the biosynthesis of fumiquinazoline C were overexpressed significantly by 7.5, 8.8, 3.4, 5.6 and 2.1 folds respectively, resulting in overall enhancement of fumiquinazoline C production by about tenfolds.