A gene cluster encoding a nonribosomal peptide synthetase-like enzyme catalyzes γ-aromatic butenolides.

Sea cucumber-derived fungi have attracted much attention due to their capacity to produce an incredible variety of secondary metabolites. Genome-wide information on Aspergillus micronesiensis H39 obtained using third-generation sequencing technology (PacBio-SMRT) showed that the strain contains nonribosomal peptide synthetase (NRPS)-like gene clusters, which aroused our interest in mining its secondary metabolites. 11 known compounds (1-11), including two γ-aromatic butenolides (γ-AB) and five cytochalasans, were isolated from A. micronesiensis H39. The structures of the compounds were determined by NMR and ESIMS, and comparison with those reported in the literature. From the perspective of biogenetic origins, the γ-butyrolactone core of compounds 1 and 2 was assembled by NRPS-like enzyme. All of the obtained compounds showed no inhibitory activity against drug-resistant bacteria and fungi, as well as compounds 1 and 2 had no anti-angiogenic activity against zebrafish.

Novel Sulfonylurea Derivatives as Potential Antimicrobial Agents: Chemical Synthesis, Biological Evaluation, and Computational Study.

Methicillin-resistant Staphylococcus aureus (MRSA) is a worldwide health threat and has already tormented humanity during its long history, creating an urgent need for the development of new classes of antibacterial agents. In this study, twenty-one novel sulfonylurea derivatives containing phenyl-5-vinyl and pyrimidinyl-4-aryl moieties were designed and synthesized, among which, nine compounds exhibited inhibitory potencies against Gram-positive bacterial strains: MRSA (Chaoyang clinical isolates), S. aureus ATCC6538, vancomycin-resistant Enterococci-309 (VRE-309), and Bacillus subtilis ATCC 6633. Especially, 9i and 9q demonstrated inhibitory activities against the four bacterial strains with minimum inhibitory concentrations (MICs) of 0.78-1.56 µg/mL, and quite a few of other MRSA clinical strains with MICs of 0.78 µg/mL, superior to those of the positive controls vancomycin (MIC of 1 µg/mL) and methicillin (MIC of >200 µg/mL). This is the very first time that sulfonylurea derivatives have been identified as promising inhibitors against different MRSA clinical isolates. In addition, all the MIC values of the synthesized compounds against Candida albicans were greater than 100 µg/mL. Since the reported anti-Candida activities of sulfonylureas were due to acetohydroxyacid synthase (AHAS) inhibition, the molecular target against MRSA for the target sulfonylureas was thought to be a different mode of action. Density functional theory (DFT) calculations were finally performed to understand the structure-activity relationships, based on which, significant differences were observed between their HOMO maps for compounds with strong antibacterial activities and weak anti-MRSA effects. The present results hence provide valuable guidance for the discovery of novel agents to treat bacterial infections, especially against MRSA.

Sesquiterpenes and polyphenols with glucose-uptake stimulatory and antioxidant activities from the medicinal mushroom Sanghuangporus sanghuang.

A chemical investigation on the fermentation products of Sanghuangporus sanghuang led to the isolation and identification of fourteen secondary metabolites (1-14) including eight sesquiterpenoids (1-8) and six polyphenols (9-14). Compounds 1-3 were sesquiterpenes with new structures which were elucidated based on NMR spectroscopy, high resolution mass spectrometry (HRMS) and electronic circular dichroism (ECD) data. All the isolates were tested for their stimulation effects on glucose uptake in insulin-resistant HepG2 cells, and cellular antioxidant activity. Compounds 9-12 were subjected to molecular docking experiment to primarily evaluate their anti-coronavirus (SARS-CoV-2) activity. As a result, compounds 9-12 were found to increase the glucose uptake of insulin-resistant HepG2 cells by 18.1%, 62.7%, 33.7% and 21.4% at the dose of 50 µmol·L-1, respectively. Compounds 9-12 also showed good cellular antioxidant activities with CAA50 values of 12.23, 23.11, 5.31 and 16.04 µmol·L-1, respectively. Molecular docking between COVID-19 Mpro and compounds 9-12 indicated potential SARS-CoV-2 inhibitory activity of these four compounds. This work provides new insights for the potential role of the medicinal mushroom S. sanghuang as drugs and functional foods.

New meroterpenoid compounds from the culture of mushroom Panus lecomtei.

Two new meroterpenoid compounds (1 and 2) together with five known meroterpenoid derivatives (3-7) were isolated from solid culture of mushroom Panus lecomtei. The structures of new compounds were confirmed by the analysis of NMR and HR-ESI-MS spectroscopic data. The biosynthetic pathway of 1-7 was postulated. All isolated compounds were evaluated for antibacterial activities against Staphylococcus aureus, Escherichia coli, Bacillus subtilis, Pseudomonas aeruginosa and Bacillus Calmette-Guérin. Compound 3 exhibited weak antibacterial activity against Bacillus Calmette-Guérin with the inhibition rate of 83.6% at 100 µmol·L-1. Other compounds showed no antibacterial activities against all tested pathogens at 100 µmol·L-1.

Antifungal activity, kinetics and molecular mechanism of action of garlic oil against Candida albicans.

The antifungal activity, kinetics, and molecular mechanism of action of garlic oil against Candida albicans were investigated in this study using multiple methods. Using the poisoned food technique, we determined that the minimum inhibitory concentration of garlic oil was 0.35 µg/mL. Observation by transmission electron microscopy indicated that garlic oil could penetrate the cellular membrane of C. albicans as well as the membranes of organelles such as the mitochondria, resulting in organelle destruction and ultimately cell death. RNA sequencing analysis showed that garlic oil induced differential expression of critical genes including those involved in oxidation-reduction processes, pathogenesis, and cellular response to drugs and starvation. Moreover, the differentially expressed genes were mainly clustered in 19 KEGG pathways, representing vital cellular processes such as oxidative phosphorylation, the spliceosome, the cell cycle, and protein processing in the endoplasmic reticulum. In addition, four upregulated proteins selected after two-dimensional fluorescence difference in gel electrophoresis (2D-DIGE) analysis were identified with high probability by mass spectrometry as putative cytoplasmic adenylate kinase, pyruvate decarboxylase, hexokinase, and heat shock proteins. This is suggestive of a C. albicans stress responses to garlic oil treatment. On the other hand, a large number of proteins were downregulated, leading to significant disruption of the normal metabolism and physical functions of C. albicans.

Synthesis and evaluation of isatin-ß-thiosemicarbazones as novel agents against antibiotic-resistant Gram-positive bacterial species.

Methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE) have caused an increasing mortality rate, which means that antibiotic resistance is becoming an important health issue. In the course to screen new agents for resistant bacteria, we identified that a series of isatin-ß-thiosemicarbazones (IBTs) could inhibit the growth of MRSA and VRE. This was the first time that the "familiar" IBT compounds exhibited significant anti Gram-positive pathogen activity. Against a clinical isolated MRSA strain, 20 of the 51 synthesized compounds showed minimum inhibitory concentration (MIC) data of 0.78 mg/L and another 12 novel compounds had MICs of 0.39 mg/L. Moreover, these compounds also inhibited Enterococcus faecalis and VRE at similar levels, indicating that IBTs might have different mode of action compared with vancomycin. For these IBTs, comparative field analysis (CoMFA) models were further established to understand the structure-activity relationships in order to design new compounds from steric and electrostatic contributions. This work has suggested that IBTs can be considered as potential lead compounds to discover antibacterial inhibitors to combat drug resistance.

ABC transporters coupled with the elevated ergosterol contents contribute to the azole resistance and amphotericin B susceptibility.

Most screening approaches produce compounds that target survival genes and are likely to generate resistance over time. Simply having more drugs does not address the potential emergence of resistance caused by target mutation, drug efflux pumps over-expression, and so on. There is a great need to explore new strategies to treat fungal infections caused by drug-resistant pathogens. In this study, we found that azole-resistant Candida albicans with CaCDR1 and CaCDR2 over-expression is hypersensitive against amphotericin B (AmB) by our high throughput synergy screening (HTSS). In contrast, Δcdr1 and Δcdr2 knockout strains were resistant to AmB. Moreover, clinical isolates with increased expression of CaCDR1 and CaCDR2 demonstrated susceptibility to AmB, which can also synergize with the efflux pumps inducer fluphenazine (FPZ). Finally, the increased drug susceptibility to AmB in azole-resistant C. albicans with drug efflux pumps over-expression was consistent with the elevated expression of CaERG11 and its associated ergosterols in clinical isolates. Our data implies that the level of ergosterol contents determines the susceptibility to azoles and AmB in C. albicans. Deep understanding of the above mechanisms would offer new hope to treat drug-resistant C. albicans.

Salinibacillus xinjiangensis sp. nov., a halophilic bacterium from a hypersaline lake.

A Gram-positive, endospore-forming, rod-shaped bacterium, designated isolate J4(T), was isolated from a neutral saline lake sample from Xinjiang Uyghur Autonomous Region, China, and subjected to a polyphasic taxonomic investigation. Phylogenetic analysis based on 16S rRNA gene sequence revealed that strain J4(T) is most closely related to Salinibacillus aidingensis 25-7(T) (with 96.7 % similarity), Salinibacillus kushneri 8-2(T) (96.5 %), Ornithinibacillus scapharcae TW25(T) (96.4 %), Salirhabdus euzebyi CVS-14(T) (96.4 %) and Ornithinibacillus californiensis MB-9(T) (96.2 %). Chemotaxonomic analysis showed menaquinone-7 (MK-7) to be the major isoprenoid quinone of strain J4(T); diphosphatidylglycerol and phosphatidylglycerol were the major cellular polar lipids and the cell wall contained meso-diaminopimelic acid as the diagnostic diamino acid. The major cellular fatty acids were iso-C15 : 0 and anteiso-C15 : 0. The genomic DNA G+C content of strain J4(T) was determined to be 36.2 mol%. Strain J4(T) was positive for catalase activity and negative for oxidase activity. Strain J4(T) was observed to grow at 25-50 °C (optimal 35-42 °C), pH 6.5-8.0 (optimal 7.0-7.5) and in media containing 1-21 % (w/v) NaCl (optimal 9-12 %). Based on these data, strain J4(T) represents a novel species of the genus Salinibacillus and the name Salinibacillus xinjiangensis sp. nov. is proposed. The type strain is J4(T) ( = CGMCC 1.12331(T) = JCM 18732(T)).

Ophiobolins P-T, five new cytotoxic and antibacterial sesterterpenes from the endolichenic fungus Ulocladium sp.

Five ophiobolane sesterterpenes, ophiobolins P-T, and three known compounds, 6-epi-21,21-O-dihydroophiobolin G, 6-epi-ophiobolin G and 6-epi-ophiobolin K, were isolated from the acetone extract of the endolichenic fungus Ulocladium sp. by using OSMAC method. Their structures were elucidated on the basis of spectroscopic analysis. The absolute configuration of the 18,19-diol moieties in ophiobolin Q was assigned using the Frelek's method. The cytotoxic effects on KB and HepG2 cell lines, antibacterial activity against Bacillus subtilis, methicillin-resistant Staphylococcus aureus, and Bacille Calmette-Guerin were evaluated for all isolated compounds. Ophiobolin T and 6-epi-ophiobolin G exhibited the most potent cytotoxic activity against HepG2 with IC50 of 0.24 and 0.37 µM, respectively. In antibacterial assay, ophiobolins P and T showed moderate antibacterial activity against B. subtilis and meticillin-resistant S. aureus. Ophiobolin T also displayed moderate antibacterial activity against the Bacille Calmette-Guerin strain.

Synthesis and evaluation of novel sulfenamides as novel anti Methicillin-resistant Staphylococcus aureus agents.

A total of 29 novel sulfenamide compounds were synthesized, spectroscopically characterized and evaluated in vitro for antimicrobial activity against various infectious pathogens. Compounds 1b and 2c exhibited potent inhibition against clinical Methicillin-resistant Staphylococcus aureus (MRSA) strains with minimum inhibitory concentration (MIC) values of 1.56 µg/mL.

Identification of blapsins A and B as potent small-molecule 14-3-3 inhibitors from the insect Blaps japanensis.

In this study, we report three novel naturally occurring compounds, blapsins A (1) and B (2), and blapsamide (3) from the ethanol extract of the stink beetle, Blaps japanensis. The structures of these compounds were determined using spectroscopic methods. Compound 3 is a phenolic compound bearing a formamido group in the structure. Functional studies revealed that compounds 1 and 2 potently inhibited 14-3-3 protein-protein interactions (PPIs) with IC(50) values of 9.2 and 10.0 µM as determined by an ELISA assay, and 2.0 and 2.5 µM in an FP assay, respectively. These compounds represent the first example of natural small-molecule 14-3-3 inhibitors.

Polyketides with antimicrobial activity from the solid culture of an endolichenic fungus Ulocladium sp.

Two new polyketides, 7-hydroxy-3, 5-dimethyl-isochromen-1-one (1) and 6-hydroxy-8-methoxy-3a-methyl-3a,9b-dihydro-3H-furo[3,2-c]isochromene-2,5-dione (2), along with eleven known compounds, 5'-methoxy-6-methyl-biphenyl-3,4,3'-triol (3), 7-hydroxy-3-(2-hydroxy-propyl)-5-methyl-isochromen-1-one (4), rubralactone (5), isoaltenuene (6), altenuene (7), dihydroaltenuenes A (8), altenusin (9), alterlactone (10), 6-O-methylnorlichexanthone (11), norlichexanthone (12), and griseoxanthone C (13) were isolated from the culture of the endolichenic fungus Ulocladium sp. Compound 2 was obtained as a racemate with an unprecedented chemical skeleton. The NMR data assignments for 3 and 4 were achieved for the first time. Compounds 1-13 were screened for their antimicrobial and radical scavenging activities. Compound 1 showed some antifungal activity against Candida albicans SC 5314 with IC(50) of 97.93 ± 1.12 µM. Compounds 11-13 showed strong activity against Bacillus subtilis with IC(50) in the range of 1-5 µM. Compound 12 significantly inhibited the growth of methicillin-resistant Staphylococcus aureus with IC(50) of 20.95 ± 1.56 µM. Compounds 9 and 10 showed strong radical scavenging activity in comparison with vitamin C. The plausible biosynthetic pathways for compounds 1, 2, and 4-8 were discussed.

Chemical constituents from endophytic fungus Fusarium oxysporum.

A new oxysporidinone analogue (1) and a new 3-hydroxyl-2-piperidinone derivative (2), along with the known compounds (-)-4,6'-anhydrooxysporidinone (3), (+)-fusarinolic acid (4), gibepyrone D (5), beauvercin (6),cerevisterol (7), fusaruside (8), and (2S,2'R,3R,3'E,4E,8E)-1-O-D-glucopyranosyl-2-N-(2'-hydroxy-3'-octadecenoyl)-3-hydroxy-9-methyl-4,8-sphingadienine (9) were isolated from Fusarium oxysporum. Compounds 1-9 were evaluated for cytotoxicity using the MTT method against cancer cell lines, PC-3, PANC-1, and A549. Beauvericin showed cytotoxicity against PC-3, PANC-1, and A549 with IC(50) value of 49.5 ± 3.8, 47.2 ± 2.9, and 10.4 ± 1.6µM, respectively. Beauvericin also exhibited anti-bacterial activity towards methicillin-resistant Staphylococcus aureus (MIC=3.125 µg/mL) and Bacillus subtilis (MIC=3.125 µg/mL).

Yuhushiella deserti gen. nov., sp. nov., a new member of the suborder Pseudonocardineae.

A thermotolerant, Gram-stain-positive, aerobic, sporangium-forming actinomycete, strain RA45(T), was isolated from a desert region in Xinjiang Uigur Autonomous Region, north-western China. Comparative analysis of the 16S rRNA gene sequence and phenotypic characterization revealed that strain RA45(T) belonged phylogenetically to the family Pseudonocardiaceae of the suborder Pseudonocardineae. Strain RA45(T) showed more than 5  % 16S rRNA gene sequence divergence from recognized species of genera in the family Pseudonocardiaceae, forming a distinct lineage within the evolutionary radiation occupied by the genera Amycolatopsis, Prauserella, Thermocrispum, Saccharomonospora, Saccharopolyspora and Sciscionella, but distinct from each of them. The affiliation to the family was supported by the presence of suborder- and family-specific 16S rRNA signature nucleotides, a DNA G+C content of 69.9 mol%, the presence of meso-diaminopimelic acid, ribose, arabinose, glucose and galactose, which are characteristic components of cell-wall chemotype IV of actinomycetes, the presence of menaquinone MK-9(H4) as the major respiratory lipoquinone, a lack of mycolic acids and the presence of an N-acetylated type of muramic acid. However, strain RA45(T) differed from known genera of the family in its polar lipid composition: the major phospholipids were phosphatidylethanolamine, phosphatidylinositol mannosides, phosphatidylmethylethanolamine, diphosphatidylglycerol, phospholipids of unknown structure and phospholipids of unknown structure containing glucosamine (phospholipid type IV). Based on its morphological, chemotaxonomic and phylogenetic characteristics, strain RA45(T) is considered to represent a novel species of a new genus in the family Pseudonocardiaceae, for which the name Yuhushiella deserti gen. nov., sp. nov. is proposed. The type strain of Yuhushiella deserti is RA45(T) (=CGMCC 4.5579(T) =JCM 16584(T)).

Antituberculosis agents and an inhibitor of the para-aminobenzoic acid biosynthetic pathway from Hydnocarpus anthelminthica seeds.

Investigation on the extracts of Hydnocarpus anthelminthica seeds led to the isolation of three new compounds, anthelminthicins A-C (1-3, resp.), and two known ones, namely chaulmoogric acid (4) and ethyl chaulmoograte (5). Their structures were determined mainly by using spectroscopic techniques. The absolute configuration at the cyclopentenyl moiety of compound 2 was rationalized by quantum calculations. Base hydrolysis, followed by optical-rotation comparison, allowed assignment of the configuration of chaulmoogric-acid moiety of compounds 3 and 5. Biological assays revealed that compounds 1-5 significantly inhibit Mycobacterium tuberculosis (MTB) growth with MIC values of 5.54, 16.70, 4.38, 9.82, and 16.80 microM, respectively. Compound 3 was found to inhibit the pathway between chorismate and para-aminobenzoic acid (pAba) with a MIC value of 11.3 microM, representing a new example of pAba inhibitor isolated from a natural source. All compounds were not toxic to Candida albicans SC5314 at a concentration up to 100 microM.

Verrucosispora sediminis sp. nov., a cyclodipeptide-producing actinomycete from deep-sea sediment.

An actinomycete, designated MS426T, the culture broth of which showed potent antimicrobial activity, was isolated from a deep-sea sediment sample of the South China Sea. An almost-complete sequence of the 16S rRNA gene of strain MS426T was determined and aligned with those of representatives of the family Micromonosporaceae available in public databases. Phylogenetic trees were inferred by using three algorithms. Strain MS426T formed a branch adjacent to Verrucosispora lutea YIM 013T in a distinct cluster occupied only by strains of the genus Verrucosispora. Strain MS426T was distinguishable from the type strains of the two described Verrucosispora species by using a combination of chemical and morphological markers and by DNA-DNA relatedness. On the basis of these genotypic and phenotypic differences, the novel antimicrobial strain with pharmaceutical potential represents a novel species, for which the name Verrucosispora sediminis sp. nov. is proposed. The type strain is MS426T (=CGMCC 4.3550T =JCM 15670T).

Amycolatopsis marina sp. nov., an actinomycete isolated from an ocean sediment.

A Gram-positive, aerobic, non-motile actinobacterium, designated strain Ms392A(T), was isolated from an ocean-sediment sample collected from the South China Sea. The isolate contained chemical markers that supported chemotaxonomic assignment to the genus Amycolatopsis. On the basis of an analysis of 16S rRNA gene sequence similarities, strain Ms392A(T) represents a novel subclade within the genus Amycolatopsis, with Amycolatopsis palatopharyngis 1BDZ(T) as its closest phylogenetic neighbour (99.4 % similarity). However, DNA-DNA hybridization demonstrated that strain Ms392A(T) was distinct from A. palatopharyngis AS 4.1729(T) (48.6 % relatedness). The polyphasic analysis demonstrated that the ocean isolate can be clearly distinguished from recognized species of the genus Amycolatopsis. Therefore, strain Ms392A(T) represents a novel species of the genus Amycolatopsis, for which the name Amycolatopsis marina sp. nov. is proposed. The type strain is Ms392A(T) (=CGMCC 4.3568(T) =NBRC 104263(T)).