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The natural fermentation of antibiotics, along with semi-synthetic and synthetic approaches, is one of the most important methods for their production. The majority of the antibiotic market comes from the fermentation of high-yielding (HY) fungal strains. These strains have been obtained since the 1950s from wild-type (WT) isolates as a result of classical strain improvement (CSI) programs primarily involving multi-round random mutagenesis and screening. However, the molecular basis leading to high-yield production was unknown. In recent years, due to the application of multiomic approaches, key changes that occur in CSI programs, with WT strains that become HY industrial producers of a particular antibiotic, have begun to be understood. It becomes obvious that, during CSI, certain universal events are selected, which lead both to a direct increase in the production of the target metabolite and affect other vital processes of the cell (side mutations). These key events include: the upregulation of the target biosynthetic gene cluster (BGC), changes in the system of global regulation, disruption of alternative BGCs, the rearrangement of energy fluxes in favor of the target SM (secondary metabolite), changes in the regulation of the response to stress, and the redirection of primary metabolic pathways to obtain more precursors for target production. This knowledge opens up the possibility of both introducing targeted changes using genetic engineering methods when creating new producers and increasing the production of CSI strains as a result of fermentation with low-molecular compounds, targeted to compensate for the effects of side mutations.

The high-yielding production of pharmaceutically significant secondary metabolites in filamentous fungi is obtained by random mutagenesis; such changes may be associated with shifts in the metabolism of polyamines. We have previously shown that, in the Acremonium chrysogenum cephalosporin C high-yielding strain (HY), the content of endogenous polyamines increased by four- to five-fold. Other studies have shown that the addition of exogenous polyamines can increase the production of target secondary metabolites in highly active fungal producers, in particular, increase the biosynthesis of β-lactams in the Penicillium chrysogenum Wis 54–1255 strain, an improved producer of penicillin G. In the current study, we demonstrate that the introduction of exogenous polyamines, such as spermidine or 1,3-diaminopropane, to A. chrysogenum wild-type (WT) and HY strains, leads to an increase in colony germination and morphological changes in a complete agar medium. The addition of 5 mM polyamines during fermentation increases the production of cephalosporin C in the A. chrysogenum HY strain by 15–20% and upregulates genes belonging to the beta-lactam biosynthetic cluster. The data obtained indicate the intersection of the metabolisms of polyamines and beta-lactams in A. chrysogenum and are important for the construction of improved producers of secondary metabolites in filamentous fungi.

The addition of exogenous polyamines increases the production of antibiotic cephalosporin C (CPC) in Acremonium chrysogenum high-yielding (HY) strain during fermentation on a complex medium. However, the molecular basis of this phenomenon is still unknown. In the current study, we developed a special synthetic medium on which we revealed the opposite effect of polyamines. The addition of 1,3-diaminopropane resulted in an increase in the yield of CPC by 12–15%. However, the addition of spermidine resulted in a decrease in the yield of CPC by 14–15% and accumulation of its metabolic pathway precursor, deacetylcephalosporin C (DAC); the total amount of cephems (DAC and CPC) was the same as after the addition of DAP. This indicates that spermidine, but not 1,3-diaminopropane, affects the final stage of CPC biosynthesis, associated with the acetylation of its precursor. In both cases, upregulation of biosynthetic genes from beta-lactam BGCs occurred at the same level as compared to the control; expression of transport genes was at the control level. The opposite effect may be due to the fact that N1-acetylation is much more efficient during spermidine catabolism than for 1,3-diaminopropane. The addition of spermidine, but not 1,3-diaminopropane, depleted the pool of acetyl coenzyme A by more than two-fold compared to control, which could lead to the accumulation of DAC.

The widespread introduction of antibiotics into medical practice, starting in the late 1940s and early 1950s, radically transformed healthcare, raised it to a qualitatively new level, allowed saving human lives in previously hopeless situations, and became one of the most important factors leading to an increase in the life expectancy of the population of Earth by more than 20 years [...]

Filamentous fungi are one of the most important producers of secondary metabolites. Some of them can havse a toxic effect on the human body, leading to diseases. On the other hand, they are widely used as pharmaceutically significant drugs, such as antibiotics, statins, and immunosuppressants. A single fungus species in response to various signals can produce 100 or more secondary metabolites. Such signaling is possible due to the coordinated regulation of several dozen biosynthetic gene clusters (BGCs), which are mosaically localized in different regions of fungal chromosomes. Their regulation includes several levels, from pathway-specific regulators, whose genes are localized inside BGCs, to global regulators of the cell (taking into account changes in pH, carbon consumption, etc.) and global regulators of secondary metabolism (affecting epigenetic changes driven by velvet family proteins, LaeA, etc.). In addition, various low-molecular-weight substances can have a mediating effect on such regulatory processes. This review is devoted to a critical analysis of the available data on the “turning on” and “off” of the biosynthesis of secondary metabolites in response to signals in filamentous fungi. To describe the ongoing processes, the model of “piano regulation” is proposed, whereby pressing a certain key (signal) leads to the extraction of a certain sound from the “musical instrument of the fungus cell”, which is expressed in the production of a specific secondary metabolite.

The vast majority of cultural heritage objects consist of materials that can be subjected to biodeterioration. Currently, there is significant number of publications showing which materials are capable of destroying certain organisms, and what conservation and restoration procedures are required. However, there is neither a clear classification of these diverse events nor their visual representation. In our review, for the first time, an attempt is made to compare the type of biodeterioration, based on the destruction of a particular material, with a specific color of the rainbow. In this regard, a cultural heritage objects made of a single material are designated as one color icon; and those made of composite materials are designated as pictogram consisting of several icons of corresponding colors. For example, a stone sculpture, in accordance with the rainbow code, is assigned a gray color, which was introduced to visualize stone materials. The drum corresponds to a pictogram consisting of violet (corresponds to leather) and brown (corresponds to wood). A work of easel painting on canvas corresponds to a pictogram consisting of a red color icon (corresponds to canvas) and a gold color icon (corresponds to painting materials). We used cold color shades to denote basic inorganic materials, and cold color shades to denote organic materials. The proposed rainbow code for biodetermination is an open platform that can be expanded by adding new colors for new materials introduced, and allows to translate potentially any cultural heritage object into a pictogram with colors that correspond to the materials used in its manufacture. Such a graphical interpretation can help both systematize the storage conditions of museum exhibits and facilitate understanding of the processes of biodeterioration of composite materials.

From the 1950s to the present, the main tool for obtaining fungal industrial producers of secondary metabolites remains the so-called classical strain improvement (CSI) methods associated with multi-round random mutagenesis and screening for the level of target products. As a result of the application of such techniques, the yield of target secondary metabolites in high-yielding (HY) strains was increased hundreds of times compared to the wild-type (WT) parental strains. However, the events that occur at the molecular level during CSI programs are still unknown. In this paper, an attempt was made to identify characteristic changes at the genome level that occurred during CSI of the Acremonium chrysogenum WT strain (ATCC 11550) and led to the creation of the A. chrysogenum HY strain (RNCM F-4081D), which produces 200–300 times more cephalosporin C, the starting substance for obtaining cephalosporin antibiotics of the 1st–5th generations. We identified 3730 mutational changes, 56 of which led to significant disturbances in protein synthesis and concern: (i) enzymes of primary and secondary metabolism; (ii) transporters, including MDR; (iii) regulators, including cell cycle and chromatin remodeling; (iv) other processes. There was also a focus on mutations occurring in the biosynthetic gene clusters (BGCs) of the HY strain; polyketide synthases were found to be hot spots for mutagenesis. The obtained data open up the possibility not only for understanding the molecular basis for the increase in cephalosporin C production in A. chrysogenum HY, but also show the universal events that occur when improving mold strains for the production of secondary metabolites by classical methods.

Acremonium chrysogenum is the only industrial producer of the antibiotic cephalosporin C (CPC), the starting substance for manufacturing cephalosporins of the first to fifth generations. Strains produced for industrial use are significantly improved by multiple rounds of random mutagenesis; however, the molecular basis for such changes is not fully understood. In this study, we attempt to elucidate key changes that occurred at the proteome level in the CSI program of A. chrysogenum HY (RNCM F-4081D), with CPC production 300-fold higher than that in the parental A. chrysogenum WT strain (ATCC 11550). Our work reveals that more than 30% of proteins are differentially expressed at different stages of fermentation. Among the identified changes, the most critical appears to be upregulation of beta-lactam biosynthetic enzymes. The data also suggest shifts in the primary metabolic pathways, providing building blocks for beta-lactam synthesis reactions, including the amino acid precursors cysteine and valine and the substrate for the expandase reaction, α-ketoglutarate. Changes in energy flows in favor of targeted metabolic pathways are also revealed. High-yielding CPC production appears to be accompanied by oxidative stress, as key oxidative stress enzymes are upregulated. Our findings are consistent with previous investigations describing changes that occurred in other fungal strains improved by classical methods. This points to general key changes leading to high-yield production. A deeper understanding of these features is important for predicting the target effects of improved industrial producers of secondary metabolites.

The transformation of steroids by microorganisms is widely used in medical biotechnology. A huge group of filamentous fungi is one of the most promising taxa for screening new biocatalytic reactions in order to obtain pharmaceutically significant steroids. In this work, we screened 10 filamentous fungi-destructors of egg tempera for the ability to biotransform androst-4-en-3,17-dione (AD) during cultivation in a liquid nutrient medium or in a buffer solution. These taxonomically unrelated strains, belonging to the classes Eurotiomycetes, Dothideomycetes and Sordariomycetes, are dominant representatives of the microbiome from halls where works of tempera painting are stored in the State Tretyakov Gallery (STG, Moscow, Russia). Since the binder of tempera paints, egg yolk, contains about 2% cholesterol, these degrading fungi appear to be a promising group for screening for steroid converting activity. It turned out that all the studied fungi-destructors are able to transform AD. Some strains showed transformation efficiency close to the industrial strain Curvularia lunata RNCIM F-981. In total, 33 steroids formed during the transformation of AD were characterized, for 19 of them the structure was established by gas chromatography/mass spectrometry analysis. In this work, we have shown for the first time that fungi-destructors of tempera paintings can efficiently transform steroids.

Assaying cellular responses to antimicrobial molecules is one way to understand modes of action of potential drugs. This is often achieved via transcriptomics and proteomics, but simple, inexpensive methods for rapid characterization are lacking. To bridge this gap, we assayed changes in the abundance of a panel of 64 “sentinel” proteins fused to GFP in the yeast Saccharomyces cerevisiae using flow cytometry. This method produced expected patterns for classical antifungals and allowed inference of common mechanisms between known and novel compounds. Single-cell data also revealed diverging responses in mitochondrial protein abundance in response to thiazolidine antifungals, and perturbations of the cell cycle caused by various compounds. Finally, the method provided insight into the unknown mode of action of alkylated nucleosides, which can be used against fungi residing on works of art. These substances elevate levels of proteins involved in the biosynthesis of aromatic amino acids (AAA), as well as in oxidative stress. Furthermore, deficiencies of Trp and Tyr biosynthesis increased the efficacy of these compounds, while antioxidants reduced it. Most surprisingly, antioxidant effectiveness relied on AAA biosynthesis. Thus, our approach and its possible modifications for other microbes provide an easy and reliable platform for revealing modes of action of novel compounds. This paper reports a valuable tool to characterize cellular responses at the single cell level, providing a powerful means to screen compounds for bioactivity and study the mechanism of these compounds.