Secondary bacterial infection is a major cause of severe progression and death in viral pneumonia. The excessive recruitment of neutrophils accompanied by the formation of neutrophil extracellular traps (NETs) plays an important role in excessive inflammation caused by viral-bacterial co-infection, but effective pharmacological interventions remain lacking. Our study revealed that quercitrin (QTN), a flavonoid commonly found in traditional Chinese medicine and daily diets, significantly reduced mortality, recruitment of neutrophils, the production of inflammatory cytokines, and viral and bacterial loads in H1N1 influenza and methicillin-resistant Staphylococcus aureus (MRSA) coinfection-induced pneumonia in mice. Mechanistically, QTN down-regulates the interaction between Histone H3 and myeloperoxidase (MPO), thereby mitigating excessive inflammation induced by NETs. Our study demonstrates that inhibiting NETosis with QTN is one of the effective strategies for co-infection treatment.

Spinosad, a secondary metabolite produced by Saccharopolyspora spinosa, is a polyketide macrolide insecticide with low toxicity and environmental friendliness. Owing to the high level of DNA methylation and unclear regulatory mechanisms, gene engineering to increase spinosad production is challenging. Limited improvements in yield have been observed with heterologous expression or partial overexpression of the 74-kb spinosyn gene cluster (spn), and research on the overexpression of the complete spinosyn gene cluster is lacking. The plasmid pCM265-spn was constructed using CRISPR/Cas9-mediated Transformation-Associated Recombination cloning to enable the overexpression of the complete spn gene cluster in Sa. spinosa. The engineered strain Sa. spinosa-spn achieved a 124

The coinfection of respiratory viruses and bacteria is a major cause of morbidity and mortality worldwide, despite the development of vaccines and powerful antibiotics. As a macromolecule that is difficult to absorb in the gastrointestinal tract, a homogeneous polysaccharide from Houttuynia cordata (HCPM) has been reported to exhibit anti-complement properties and alleviate influenza A virus (H1N1)-induced lung injury; however, the effects of HCPM without in vitro antiviral and antibacterial activities on more complicated pulmonary diseases resulting from viral-bacterial coinfection remains unclear. This study established a representative coinfection murine pneumonia model infected with H1N1 (0.2 LD50) and methicillin-resistant Staphylococcus aureus (MRSA, 107 CFU). HCPM significantly improved survival rate and weight loss, and ameliorated gut-lung damage and inflammatory cytokine production. Interestingly, the therapeutic effect of HCPM on intestinal damage preceded that in the lungs. Mechanistically, HCPM inhibited the overactivation of the intestinal complement (C3a and C5a) and suppressed the activation of the NLR family pyrin domain-containing 3 (NLRP3) pathway, which contributes to the regulation of the Treg/Th17 cell balance in the gut-lung axis. The results indicate the beneficial effects of an anti-complement polysaccharide against viral-bacterial coinfection pneumonia by modulating crosstalk between multiple immune regulatory networks.

This study investigated the phytochemical profiles and bioactivities of two edible boletes from Southwestern China, Phlebopus portentosus and Butyriboletus roseoflavus. A total of 33 secondary metabolites, comprising 15 alkaloids, 4 pulvinic acid derivative pigments, and 14 ergosterols, were isolated and identified. To our best knowledge, boletesine A (1), boletesine B (2), and cis-xerocomic acid (16) were previously undescribed compounds. The new structures were established by extensive spectroscopic methods and chemical calculations. Compound 1 features a hitherto unknown hybrid skeleton formed between a 2-formylpyrrole-alkaloid and a dopacetic acid (DOPAC) via a Michael addition reaction. Bioactivity assays revealed the neuroprotective effects of compounds 18 and 19 against Aβ25–35- or H2O2-induced toxicity. In a cytotoxic assay against a small panel of cancer cell lines, compound 9 exhibited significant activity against HeLa cells (IC50 = 10.76 µM), while 33 demonstrated broad-spectrum cytotoxicity against Hela229, SGC7901, PC-3, and BEL7402 cells (IC50s in the range of 20~30 µM). Of particular note is the anti-influenza virus activities against A/H3N2 and B/Victoria strains of compounds 22 and 26 (EC50 values ranging from 3.6 to 9.6 µM). Along with these, compound 29 showed a moderate antiviral effect against coxsackievirus B3. These findings underscore the therapeutic potential of the two edible boletes in addressing neurodegenerative diseases, cancer, and viral infections, paving the way for their prospective applications in the development of functional foods and pharmaceuticals.

A new 10-membered lactone derivative (diaporolide, 1) and three known polyketides (2-4) were obtained from the endophytic fungus Diaporthe sp. RYF-Br-5, which was derived from the fresh branches of Keteleeria pubescens, a rare endemic conifer native to China. Through a detailed spectroscopic analysis and theoretical ECD calculations, the structure and absolute stereochemistry of compound 1 were confirmed. In antimicrobial assays, compound 1 exhibited an inhibitory effect against Staphylococcus aureus with an MIC value of 64 mu g/mL.

Eight previously undescribed solanapyrone analogues, sphasolanapyrones A-H (1-8), together with four structurally related known compounds (9-12) were obtained from the solid fermentation of Nigrospora sphaerica MZW-A, an endophytic fungus isolated from the fresh branches of the endangered conifer Pinus wangii. This study represents the first investigation on the secondary metabolites of endophytic fungus associated with this precious plant. The structures and absolute configurations of compounds 1-8 were elucidated by extensive spectroscopic analysis and electronic circular dichroism (ECD) calculations. 1 and 2 are first examples of 3,4-vic diol solanapyrone analogues, whereas 7 and 8 are rare 17-nor-solanapyrones. Among these isolates, 3, 4, 5, and 11 showed weak inhibitory against Staphylococcus aureus. In addition, compound 5 displayed weak cytotoxic effects against several tumor cell lines.

Four undescribed polyketides, beshanzones A (1) and B (2) as well as beshanhexanols A (3) and B (4), along with three known ones (5-7) were isolated from the rice fermentation of two endophytic fungi associated with the critically endangered Chinese endemic conifer Abies beshanzuensis. gamma-Butyrolactone derivatives 1, 2, and 5 were isolated from Phomopsis sp. BSZ-AZ-2, an interesting strain that drawn our attention this time. The cyclohexanol derivatives 3, 4, 6, and 7 were obtained during a follow-up investigation on Penicillium commune BSZ-P-4-1. The chemical structures including absolute configurations of compounds 1-4 were determined by spectroscopic methods, Mo2(OAc)4 induced electronic circular dichroism (IECD), GIAO NMR calculations and DP4+ probability analyses. In particular, compound 2 contains a novel 5/5 bicyclic ring system, which might be biogenetically derived from the known compound 5 through hydrolysis followed by an Aldol reaction. All isolates were evaluated for their antimicrobial activities against a small panel of bacterial and fungal pathogens. Compounds 6 and 7 showed moderate inhibitory activities against Candida albicans, with MIC values of 16 and 32 mu g/mL, respectively.

New Delhi metallo-beta-lactamase-1 (NDM-1) has rapidly disseminated worldwide, leading to multidrug resistance and worse clinical prognosis. Designing and developing effective NDM-1 inhibitors is a critical and urgent challenge. In this study, we constructed a library of long-lasting nitroxoline derivatives and identified ASN-1733 as a promising dual-functional antibiotic. ASN-1733 can effectively compete for Ca2+ on the bacterial surface, causing the detachment of lipopolysaccharides (LPS), thereby compromising the outer membrane integrity and permeability and exhibiting broad-spectrum bactericidal activity. Moreover, ASN-1733 demonstrated wider therapeutic applications than nitroxoline in mouse sepsis, thigh and mild abdominal infections. Furthermore, ASN-1733 can effectively inhibit the hydrolytic capability of NDM-1 and exhibits synergistic killing effects in combination with meropenem against NDM-1 positive bacteria. Mechanistic studies using enzymatic experiments and computer simulations revealed that ASN-1733 can bind to key residues on Loop10 of NDM-1, hindering substrate entry into the enzyme's active site and achieving potent inhibitory activity (K-i = 0.22 mu M), even in the presence of excessive Zn2+. These findings elucidate the antibacterial mechanism of nitroxoline and its derivatives, expand their potential application in the field of antibacterial agents and provide new insights into the development of novel NDM-1 inhibitors. [GRAPHICS] .

An investigation on the secondary metabolites from a rice culture broth of the endophytic fungus Neurospora terricola HDF-Br-2 derived from the vulnerable conifer Pseudotsuga gaussenii led to the isolation and characterization of 34 structurally diverse polyketides (1-34). Seven of them are previously undescribed, including five unprecedented dihydropyran-containing (terricoxanthones A-E, 1-5, resp.) and one rare tetrahydrofurancontaining (terricoxanthone F, 6) dimeric xanthones. The structures were elucidated by spectroscopic methods and single-crystal X-ray diffraction analyses. Terricoxanthones each were obtained as a racemic mixture. Their plausible biosynthetic relationships were briefly proposed. Compounds 6, aspergillusone A (8), and alatinone (27) displayed considerable inhibition against Candida albicans with MIC values of 8-16 mu g/mL. 4-Hydroxyvertixanthone (12) and 27 exhibited significant inhibitory activities against Staphylococcus aureus, with MIC values of 4-8 mu g/mL. Furthermore, compounds 8 and 27 could disrupt biofilm of S. aureus and C. albicans at 128 mu g/mL. The findings not only extend the skeletons of xanthone dimers and contribute to the diversity of metabolites of endophytes associated with the endangered Chinese conifer P. gaussenii, but could further reveal the important role of protecting plant species diversity in support of chemical diversity and potential sources of new therapeutics.

Two new tetraketide-derived phenol rhamnosides [botryrhamnosides A (1) and B (2)] and a new rhamnosylated tryptophol alkaloid (botryrhamnoside C, 3), along with seven related known compounds (4-10) were isolated from the solid culture of Botryosphaeria dothidea LE-07, an endophytic fungus residing in the leaves of the rare medicinal plant Chinese tulip tree (Liriodendron chinense). Their structures with the absolute configurations were determined by a combination of spectroscopy methods, comparing specific rotations, electronic circular dichroism (ECD) calculations, and single-crystal X-ray diffraction analysis. Compounds 1 and 2 are rare tetraketide-derived resorcinols incorporating a L-rhamnose moiety, while 3 represents the first example of rhamnose-bound tryptophol derivatives produced by microorganisms. These metabolites were evaluated in vitro for their antimicrobial and anti-neuroinflammation activities. The rhamnosylated derivatives 1-5 displayed potent antibacterial activity against Escherichia coli, with MIC values in the range of 8-16 mu g/mL. Compound 2 attenuated neuroinflammation in lipopolysaccharide (LPS)-induced BV-2 microglial cells, by decreasing the level of pro-inflammatory mediators [nitric oxide (NO), tumor necrosis factor-alpha (TNF-alpha), and interleukin 6 (IL-6)] and down-regulating the mRNA expression of inducible nitric oxide synthase (iNOS). In addition, compound 8 exhibited remarkable inhibitory effect against the ATP-citrate lyase (ACL), an emerging drug target for hyperlipidemia and related glycolipid metabolic disorders, with an IC50 value of 5.32 mu M.

Fungichromin is a polyene macrolide antibiotic with potent killing activity against a broad range of agricultural pathogens and filamentous fungi and a wide range of potential applications. The production of fungichromin is still hampered by poor fermentation yield and high cost. In this study, the whole genome sequencing of fungichromin-producing Streptomyces sp. WP-1 was conducted, and the fungichromin biosynthetic gene cluster was identified. Comparative analysis revealed that the fungichromin biosynthetic gene cluster contains two regulatory genes, ptnF, and ptnR. The roles of ptnF and ptnR were determined through knockout and complementation. The yield of fungichromin was increased by overexpressing these two regulatory genes, as well as the crotonyl CoA reductase/carboxylase gene ptnB in Streptomyces sp. WP-1. The yield of fungichromin was increased to 8.5 g/L using a combination of genetic engineering and a medium optimization strategy, which is the highest fermentation titer recorded. • Confirmation of the positive regulation of ptnF and ptnR on fungichromin. • Improvement of fungichromin production by the construction of ptnF, ptnR, and ptnB overexpression strains. • Improvement of fungichromin production by the addition of soybean oil and copper ions at optimal concentration.

Photodynamic inactivation (PDI) has received increasing attention as a promising approach to combat Candida albicans infections. This study aimed to evaluate the synergistic effect of a new BODIPY (4,4-difluoro-boradiazaindacene) derivative and hydrogen peroxide on C. albicans. BDP-4L in combination with H2O2 demonstrated enhanced photokilling efficacy. In suspended cultures of C. albicans, the maximum decrease was 6.20 log and 2.56 log for PDI using BDP-4L (2.5 μM) with or without H2O2, respectively. For mature C. albicans biofilms, 20 μM BDP-4L plus H2O2 eradicated C. albicans, causing an over 6.7 log count reduction in biofilm-associated cells, while only a reduction of ~ 1 log count was observed when H2O2 was omitted. Scanning electron microscopy analysis and LIVE/DEAD assays suggested that PDI using BDP-4L plus H2O2 induced more damage to the cell membrane. Correspondingly, amplification of nucleic acids release was observed in biofilms treated with the combined PDI. Additionally, we also discovered that the addition of hydrogen peroxide potentiated the generation of 1O2 in PDI using the singlet oxygen sensor green probe. Collectively, BDP-4L combined with H2O2 presents a promising approach in the treatment of C. albicans infections.

Four undescribed palmarumycin-type spirodioxynaphthalenes (phyligustricins A-D) and a known biogenetic precursor (palmarumycin BG1) were isolated from a solid fermentation of Phyllosticta ligustricola HDF-L-2, an endophyte associated with the endangered Chinese conifer Pseudotsuga gaussenii. The structures were elucidated by spectroscopic methods, single-crystal X-ray diffraction analyses, and electronic circular dichroism calculations. Both phyligustricins A and B have an unprecedented spirodioxynaphthalene-derived skeleton containing an extra 4H-furo [3,2-c]pyran-4-one moiety, while phyligustricins C and D are p-hydroxy-phenethyl substituted spirodioxynaphthalenes. The plausible biosynthetic relationships of the isolates were briefly proposed. Phyligustricins C and D and palmarumycin BG1 showed considerable antibacterial activity against Staphylococcus aureus, each with an MIC value of 16 μg/mL. Palmarumycin BG1 displayed significant inhibitory effects against ACL and ACC1, with IC50 values of 1.60 and 8.00 μM, respectively.

Phosphate concentration above 10 mM reduces the production of many secondary metabolites; however, the phenomenon is not mechanistically understood yet. Specifically, the problem of phosphorus limitation in antibiotic production remains unresolved. This study investigates the phosphorus inhibition effect on spinosad production and alleviates it by calcium and phosphate supplementation to fermentation media. Furthermore, we examined the mechanism of fatty acids–induced increase in polyketides production. Four phosphates that were supplemented into the fermentation media include NaH2PO4, Na2HPO4, KH2PO4, and K2HPO4 and NaH2PO4 was found to be the most effective phosphate. Under the optimal phosphate condition of supplementing 20 mM NaH2PO4 on the fourth day and 5 g/L CaCO3, the maximal spinosad production reached 520 mg/L, showing a 1.65-fold increase over the control treatment. In the NaH2PO4-CaCO3 system, the de novo fatty acid biosynthesis was significantly downregulated while spinosad biosynthesis and β-oxidation were upregulated. The coordination of de novo fatty acid biosynthesis and β-oxidation promoted intracellular acetyl-CoA concentration. The results demonstrate that NaH2PO4-CaCO3 combined addition is a simple and effective strategy to alleviate phosphorus inhibition effect through the regulation of fatty acid metabolism and accumulation of immediate precursors. This information improves our understanding of phosphates’ influence on the large-scale production of polyketides.

BACKGROUND:Photodynamic therapy (PDT) may be an alternative treatment of Candida albicans (C. albicans) infections. The aim of this study was to investigate the antifungal effect of PDT mediated by a new photosensitizer (PS) derived from BODIPY (BDP-4L) on C. albicans biofilms. METHODS:C. albicans biofilms were incubated with BDP-4L of different concentrations and then irradiated at the light doses of 1.8, 3.6, 5.4, 7.2 and 9.0 J/cm2. XTT reduction assay was conducted to determine the PS concentration and PDT parameters. Confocal light scanning microscopy (CLSM) and scanning electron microscope (SEM) were used to visualize and quantify the effect of BDP-4L on C. albicans biofilms after PDT. RESULTS:C. albicans biofilms were inactivated in light dose-dependent and PS concentration-dependent manners using BDP-4L as PS. Without irradiation, no inactivation effect was observed when PS concentrations varied from 5 μM to 80 μM. 40 μM PS with 3.6 J/cm2 irradiation resulted in a significant reduction of 83.8% in biofilm metabolic activities. CLSM assay demonstrated that cell viability was obviously inhibited by 82.6%. SEM images revealed ruptured and rough cell surface, indicating increased cell membrane permeability after PDT. CONCLUSIONS:Our results suggested that BDP-4L mediated PDT exhibited a favorable antifungal effect on C. albicans biofilms.

A new N-methoxy-1-pyridone alkaloid [chromenopyridin A (1)] and four known compounds (2-5) were isolated and identified from the endophytic fungus Penicillium nothofagi P-6, which was derived from the bark of the critically endangered conifer Abies beshanzuensis. Their structures were elucidated by extensive spectroscopic analyses and single-crystal X-ray diffraction. Among the isolates, compound 1 showed considerable cytotoxicities against the A549 and Hela human cancer cell lines, with IC50 values of 14.7 and 11.3 mu M. In addition, compounds 1 and 4 exhibited potent antibacterial activity against Staphylococcus aureus with MIC values of 62.5 and 15.6 mu g/mL, respectively.

A structurally unprecedented prenylated indole alkaloid, together with five known analogues, was isolated from an endophytic fungus associated with the endangered conifer Abies beshanzuensis .

Three new (psganpenes A-C), one known eremophilane sesquiterpene and one newrelated derivative (psganpene D) were isolated from the culture of endophytic fungus Nemania sp. HDF-Br-5, which derived from Pseudotsuga gaussenii Flous, anendangered conifer endemic to China. Their structures were deduced by extensive spectroscopic methods. All isolates were screened for cytotoxicity and antibacterial activity. Psganpene D showed promising cytotoxicity against KB, HCT116 and Hela cell strains, with IC50 values in the range of 15.1-32.3 mu M.