Methicillin-resistant Staphylococcus aureus (MRSA), a WHO high-priority pathogen that can cause great harm to living beings, is a primary cause of death from antibiotic-resistant infections. In the present study, six new compounds, including fumindoline A–C (1–3), 12β, 13β-hydroxy-asperfumigatin (4), 2-epi-tryptoquivaline F (17) and penibenzophenone E (37), and thirty-nine known ones were isolated from the marine-derived fungus Aspergillus fumigatus H22. The structures and the absolute configurations of the new compounds were unambiguously assigned by spectroscopic data, mass spectrometry (MS), electronic circular dichroism (ECD) spectroscopic analyses, quantum NMR and ECD calculations, and chemical derivatizations. Bioactivity screening indicated that nearly half of the compounds exhibit antibacterial activity, especially compounds 8 and 1 1, and 33–38 showed excellent antimicrobial activities against MRSA, with minimum inhibitory concentration (MIC) values ranging from 1.25 to 2.5 μM. In addition, compound 8 showed moderate inhibitory activity against Mycobacterium bovis (MIC: 25 μM), compound 10 showed moderate inhibitory activity against Candida albicans (MIC: 50 μM), and compound 13 showed strong inhibitory activity against the hatching of a Caenorhabditis elegans egg (IC₅₀: 2.5 μM).

Fungal natural products play a prominent role in the development of pharmaceuticalagents. Two new cyclic tetrapeptides (CTPs), westertide A (1) and B (2), with eight known compounds (3–10) were isolated from the fungus Aspergillus westerdijkiae guided by OSMAC (one strain-many compounds) and molecular networking strategies. The structures of new compounds were unambiguously determined by a combination of NMR and mass data analysis, and chemical methods. All of the isolates were evaluated for antimicrobial effects, synergistic antifungal activity, cytotoxic activity, and HDAC inhibitory activity. Compounds 1–2 showed synergistic antifungal activity against Candida albicans SC5314 with the presence of rapamycin and weak HDAC (histone deacetylase) inhibitory activity. These results indicate that molecular networking is an efficient approach for dereplication and identification of new CTPs. CTPs might be a good starting point for the development of synergistic antifungal agents.

Under the guidance of LC-MS/MS-based molecular networking, seven new verrucosidin derivatives, penicicellarusins A-G (3–9), were isolated together with three known analogues from the fungus Penicillium cellarum. The structures of the new compounds were determined by a combination of NMR, mass and electronic circular dichroism spectral data analysis. The absolute configuration of penicyrone A (10) was corrected based on X-ray diffraction analyses. Bioactivity screening indicated that compounds 1, 2, and 4 showed much stronger promising hypoglycemic activity than the positive drug (rosiglitazone) in the range of 25–100 μM, which represents a potential new class of hypoglycemic agents. Preliminary structure-activity relationship analysis indicates that the formation of epoxy ring on C₆-C₇ in the structures is important for the glucose uptake-stimulating activity. The gene cluster for the biosynthesis of 1–12 is identified by sequencing the genome of P. cellarum and similarity analysis with the gene cluster of verrucosidins in P. polonicum.

Two new cyclohexadepsipeptides japonamides A (1) and B (2) were isolated from the ethyl acetate extract of a marine-sponge-derived fungus Aspergillus japonicus based on molecular networking. Their structures were elucidated by comprehensive spectral analysis and their absolute configurations were confirmed by Marfey’s method. Compounds 1 and 2 showed no antifungal activities against Candida albicans SC5314 measured by the broth microdilution method but exhibited prominent synergistic antifungal activities in combination with fluconazole, ketoconazole, or rapamycin. The Minimum inhibitory concentrations (MICs) of rapamycin, fluconazole, and ketoconazole were significantly decreased from 0.5 to 0.002 μM, from 0.25 to 0.063 μM, and from 0.016 to 0.002 μM, in the presence of compounds 1 or 2 at 3.125 μM, 12.5 μM, and 6.25 μM, respectively. Surprisingly, the combination of compounds 1 or 2 with rapamycin showed a strong synergistic effect, with fractional inhibitory concentration index (FICI) values of 0.03.

Exogenous genes are generally expressed by integration into the chromosomes of Pichia pastoris. However, systematic studies on the chromosomal position effect are lacking, and locations that are conducive to the high expression of foreign genes are rarely reported. In this study, a genomic random insertion mutagenesis library for P. pastoris was successfully constructed using the piggyBac (PB) transposon system. Through sequencing, the sequence TTAA was identified as the major recognition site of the PB transposon, which exhibited relatively high coverage on P. pastoris chromosomes, making it a valuable tool for studying position effect variegation in P. pastoris. Using the enhanced green fluorescent protein gene (eGFP) as a reporter, two libraries including low-expression positions and high-expression positions were obtained by flow cytometry. The low-expression sites were mainly located upstream of ORFs around the promoter region and downstream near the terminator region, while the high-expression sites were predominantly located at the gene interior. KEGG and GO analyses showed that genes in high-expression positions were significantly enriched in the ATP-dependent chromatin remodeling and histone binding pathways, and genes in low-expression positions were significantly enriched in the MAPK signaling pathway, autophagy, mitochondrial autophagy, ABC transporters, and the arginine synthesis pathway. This study has clarified the genome-wide landscape of position effect variegation in P. pastoris. Additionally, it has provided novel insights into high-throughput screening strategies for strains with high exogenous gene expression.