Hepialiamides A-C: Aminated Fusaric Acid Derivatives and Related Metabolites with Anti-Inflammatory Activity from the Deep-Sea-Derived Fungus Samsoniella hepiali W7.

A systematic investigation combined with a Global Natural Products Social (GNPS) molecular networking approach, was conducted on the metabolites of the deep-sea-derived fungus Samsoniella hepiali W7, leading to the isolation of three new fusaric acid derivatives, hepialiamides A-C (1-3) and one novel hybrid polyketide hepialide (4), together with 18 known miscellaneous compounds (5-22). The structures of the new compounds were elucidated through detailed spectroscopic analysis. as well as TD-DFT-based ECD calculation. All isolates were tested for anti-inflammatory activity in vitro. Under a concentration of 1 µM, compounds 8, 11, 13, 21, and 22 showed potent inhibitory activity against nitric oxide production in lipopolysaccharide (LPS)-activated BV-2 microglia cells, with inhibition rates of 34.2%, 30.7%, 32.9%, 38.6%, and 58.2%, respectively. Of particularly note is compound 22, which exhibited the most remarkable inhibitory activity, with an IC50 value of 426.2 nM.

Structural optimization of natural product fusaric acid to discover novel T3SS inhibitors of Salmonella.

Type III secretion system (T3SS) plays a critical role in host cell invasion and pathogenesis of Salmonella. We recently identified the mycotoxin fusaric acid (FA) as a T3SS inhibitor of Salmonella. Herein, twenty-two diphenylsulfane derivatives were designed and synthesized using FA as a lead compound through scaffold hopping. Among them, SL-8 and SL-19 possessing strong anti-T3SS and anti-invasion activity were identified as T3SS inhibitors with improvement in potency as compared to FA. The inhibitory mechanisms on SPI-1 did not depend on the HilD-HilC-RtsA-HilA or PhoP-PhoQ pathway or the assembly of T3SS needle complex. Accordingly, we proposed that the inhibitory effects of SL-8 and SL-19 on SPI-1 probably influence the formation of SicA/InvF-effector complex or other related proteins.

Fusaric acid instigates the invasion of banana by Fusarium oxysporum f. sp. cubense TR4.

Fusaric acid (FSA) is a phytotoxin produced by several Fusarium species and has been associated with plant disease development, although its role is still not well understood. Mutation of key genes in the FSA biosynthetic gene (FUB) cluster in Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) reduced the FSA production, and resulted in decreased disease symptoms and reduced fungal biomass in the host banana plants. When pretreated with FSA, both banana leaves and pseudostems exhibited increased sensitivity to Foc TR4 invasion. Banana embryogenic cell suspensions (ECSs) treated with FSA exhibited a lower rate of O2 uptake, loss of mitochondrial membrane potential, increased reactive oxygen species (ROS) accumulation, and greater nuclear condensation and cell death. Consistently, transcriptomic analysis of FSA-treated ECSs showed that FSA may induce plant cell death through regulating the expression of genes involved in mitochondrial functions. The results herein demonstrated that the FSA from Foc TR4 functions as a positive virulence factor and acts at the early stage of the disease development before the appearance of the fungal hyphae in the infected tissues.

Evaluation of the cytotoxic and genotoxic effects of mycotoxin fusaric acid.

Fusaric acid (FA) is produced by several Fusarium species and is commonly found in grains. This investigation was performed to evaluate the cytotoxic and genotoxic effects of FA either in human cervix carcinoma (HeLa) cell line using 3-(4,5-dimethylthiazolyl-2)-2,5 diphenyltetrazolium bromide (MTT) assay and in human lymphocytes using chromosome aberrations (CAs), sister chromatid exchanges (SCEs), micronuclei (MN) as well as comet assay in vitro. The cells were treated with 0.78, 1.56, 3.125, 6.25, 12.50, 25, 50, 100, 200, and 400 µg/mL concentrations of FA. It has potent cytotoxic effect on HeLa cell line measured by MTT assay especially at higher concentrations (200, 400 µg/mL). The half of inhibitory concentration (IC50) evidenced by FA in the HeLa cells was 200 µg/mL at 24 h and between 200 and 400 µg/mL at 48 h. It was also observed that FA produced a significant decrease in mitotic index (MI) at 12.50 µg/mL compared to solvent control. Furthermore, it indicated a cytotoxic effect at the concentrations ranging from 25 to 400 µg/mL in human lymphocytes. The results of this research point out that being exposed to FA at high concentrations show cytotoxicity. Besides FA induced comet tail intensity at 3.125, 6.25, and 12.50 µg/mL concentrations in isolated human lymphocytes. On the other hand, no genotoxic effects were seen in human lymphocytes in vitro using CA, SCE and MN assays.

Concise Total Synthesis and Antifungal Activities of Fusaric Acid, a Natural Product.

The total synthesis of a natural product alkaloid fusaric acid (FA), which exhibits herbicide, fungicide, insecticide and even diverse notable pharmacological activities, was accomplished in four steps using commercially available materials. The synthesis, based on a unified and flexible strategy using 6-bromonicotinaldehyde as a common intermediate, is concise, convergent, practical and can be carried out on a two-gram scale. This approach could be readily applicable to the synthesis of its analogues. In addition, FA had a wide range of inhibitory activities against 14 plant pathogenic fungi in this study, which demonstrated that as a leading compound, and it has great potential to be further developed as an agricultural fungicide.

Fusaric Acid Induces DNA Damage and Post-Translational Modifications of p53 in Human Hepatocellular Carcinoma (HepG2 ) Cells.

Fusaric acid (FA), a common fungal contaminant of maize, is known to mediate toxicity in plants and animals; however, its mechanism of action is unclear. p53 is a tumor suppressor protein that is activated in response to cellular stress. The function of p53 is regulated by post-translational modifications-ubiquitination, phosphorylation, and acetylation. This study investigated a possible mechanism of FA induced toxicity in the human hepatocellular carcinoma (HepG2 ) cell line. The effect of FA on DNA integrity and post-translational modifications of p53 were investigated. Methods included: (a) culture and treatment of HepG2 cells with FA (IC50 : 580.32 µM, 24 h); (b) comet assay (DNA damage); (c) Western blots (protein expression of p53, MDM2, p-Ser-15-p53, a-K382-p53, a-CBP (K1535)/p300 (K1499), HDAC1 and p-Ser-47-Sirt1); and (d) Hoechst 33342 assay (apoptosis analysis). FA caused DNA damage in HepG2 cells relative to the control (P < 0.0001). FA decreased the protein expression of p53 (0.24-fold, P = 0.0004) and increased the expression of p-Ser-15-p53 (12.74-fold, P = 0.0126) and a-K382-p53 (2.24-fold, P = 0.0096). This occurred despite the significant decrease in the histone acetyltransferase, a-CBP (K1535)/p300 (K1499) (0.42-fold, P = 0.0023) and increase in the histone deacetylase, p-Ser-47-Sirt1 (1.22-fold, P = 0.0020). The expression of MDM2, a negative regulator of p53, was elevated in the FA treatment compared to the control (1.83-fold, P < 0.0001). FA also inhibited cell proliferation and induced apoptosis in HepG2 cells as evidenced by the Hoechst assay. Together, these results indicate that FA is genotoxic and post-translationally modified p53 leading to HepG2 cell death. J. Cell. Biochem. 118: 3866-3874, 2017. © 2017 Wiley Periodicals, Inc.

Microbial Resistance Mechanisms to the Antibiotic and Phytotoxin Fusaric Acid.

Fusaric acid (FA) produced by Fusarium oxysporum plays an important role in disease development in plants, including cotton. This non-specific toxin also has antibiotic effects on microorganisms. Thus, one expects a potential pool of diverse detoxification mechanisms of FA in nature. Bacteria and fungi from soils infested with Fusarium and from laboratory sources were evaluated for their ability to grow in the presence of FA and to alter the structure of FA into less toxic compounds. None of the bacterial strains were able to chemically modify FA. Highly FA-resistant strains were found only in Gram-negative bacteria, mainly in the genus of Pseudomonas. The FA resistance of the Gram-negative bacteria was positively correlated with the number of predicted genes for FA efflux pumps present in the genome. Phylogenetic analysis of predicted FA resistance proteins (FUSC, an inner membrane transporter component of the efflux pump) revealed that FUSC proteins having high sequence identities with the functionally characterized FA resistance protein FusC or Fdt might be the major contributors of FA resistance. In contrast, most fungi converted FA to less toxic compounds regardless of the level of FA resistance they exhibited. Five derivatives were detected, and the detoxification of FA involved either oxidative reactions on the butyl side chain or reductive reactions on the carboxylic acid group. The production of these metabolites from widely different phyla indicates that resistance to FA by altering its structure is highly conserved. A few FA resistant saprophytic or biocontrol strains of fungi were incapable of altering FA, indicating a possible involvement of efflux transporters. Deployment of both efflux and derivatization mechanisms may be a common feature of fungal FA resistance.

Fusaric acid induces a notochord malformation in zebrafish via copper chelation.

Over a thousand extracts were tested for phenotypic effects in developing zebrafish embryos to identify bioactive molecules produced by endophytic fungi. One extract isolated from Fusarium sp., a widely distributed fungal genus found in soil and often associated with plants, induced an undulated notochord in developing zebrafish embryos. The active compound was isolated and identified as fusaric acid. Previous literature has shown this phenotype to be associated with copper chelation from the active site of lysyl oxidase, but the ability of fusaric acid to bind copper ions has not been well described. Isothermal titration calorimetry revealed that fusaric acid is a modest copper chelator with a binding constant of 4.4 × 10(5) M(-1). These results shed light on the toxicity of fusaric acid and the potential teratogenic effects of consuming plants infected with Fusarium sp.

[Inhibition of Amaranthus retroflexus by wasp gut fungal isolate Fusarium oxysporum MF06].

OBJECTIVE: To study the inhibitory effect of wasp gut fungus against the radicle growth of Amaranthus retroflexus for the development of herbicides from microorganisms. METHODS: Eleven strains were isolated from wasp gut. Among them the fermentation broth of strain MF06 showed potent herbicidal activity against A. retroflexus. MF06 was identified by morphological observation and molecular biology identification. Fermentation product was isolated and purified by silica column chromatography, TLC and Sephadex LH-20 column chromatography. Metabolite 1 was obtained from fermentation product, and the inhibitory effect of metabolite 1 against the radicle growth of A. retrofexus was studied. The structure was determined by mass spectrum and nuclear magnetic resonance analyses. RESULTS: By the morphological observation and ITS sequence analysis, MF06 was identified as Fusarium oxysporum. The ethyl acetate extract of MF06 had strong activity against A. retrofexus with inhibition rate of more than 68% under the concentration of 100 µg/mL. It is better than other polarities crude extracts. Metabolite 1 was separated from ethyl acetate extract, and it was determined as a mixture of fusaric acid and 9 ,10-dehydrofusaric acid. The mixture is co-crystallizing in a 1:1 molar stoichiometry. It inhibited radical growth of A. retroflexus with IC50 value of (0.51 ± 0.18) µg/mL, comparable to that of 2-( 2,4- dichlorophenoxy) acetic acid (0.30 ± 0.14 µg/mL) used as a positive control. CONCLUSION: Strain MF06 could be potentially developed as a microbial herbicide.

Fusaric acid modulates Type Three Secretion System of Salmonella enterica serovar Typhimurium.

Natural small-molecule products are promising lead compounds for developing a generation of novel antimicrobials agents to meet the challenge of antibiotic-resistant pathogens. To facilitate the search for novel anti-virulence agents, we chose a virulence factor of Type Three Secretion System (T3SS) as a drug target to screen candidates from a small-molecule library in our laboratory. This study demonstrated fusaric acid had dramatically inhibitory effects on secretion of Salmonella island 1 (SPI-1) effector proteins and invasion of Salmonella into HeLa cells. Moreover, fusaric acid had no inhibitory effects on bacterial growth and viability of host cells. Protein HilA is a key regulator of SPI-1 in Salmonella, which affects transcription of SPI-1 effectors and SPI-1 apparatus genes. In this study, fusaric acid (FA) did not affect secretion of SPI-1 effectors in HilA over-expressed strain, suggesting it did not affect the transcription of SPI-1. In addition, fusaric acid did not affect the protein level of apparatus protein PrgH in SPI-1 needle complex. As a result, we proposed fusaric acid had an inhibitory effect on SPI-1 probably depending on its influence on SicA/InvF. In summary, fusaric acid is a novel inhibitor of T3SS with potential for further developing novel anti-virulence agents.

Identification of toxin inhibitors using a magnetic nanosensor-based assay.

A magnetic nanosensor-based method is described to screen a library of drugs for potential binding to toxins. Screening is performed by measuring changes in the magnetic relaxation signal of the nanosensors (bMR nanosensors) in aqueous suspension upon addition of the toxin. The Anthrax lethal factor (ALF) is selected as a model toxin to test the ability of our bMR nanosensor-based screening method to identify potential inhibitors of the toxin. Out of 30 molecules screened, sulindac, naproxen and fusaric acid are found to bind LF, with dissociation constants in the low micromolar range. Further biological analysis of the free molecules in solution indicate that sulindac and its metabolic products inhibited LF cytotoxicity to macrophages with IC50 values in the micromolar range. Meanwhile, fusaric acid is found to be less effective at inhibiting LF cytotoxicity, while naproxen does not inhibit LF toxicity. Most importantly, when the sulindac and fusaric acid-bMR nanosensors themselves are tested as LF inhibitors, as opposed to the corresponding free molecules, they are stronger inhibitors of LF with IC50 values in the nanomolar range. Taken together, these studies show that a bMR nanosensors-based assay can be used to screen known drugs and other small molecules for inhibitor of toxins. The method can be easily modified to screen for inhibitors of other molecular interactions and not only the selected free molecule can be study as potential inhibitors but also the bMR nanosensors themselves achieving greater inhibitory potential.

Exogenous application of mycotoxin fusaric acid improve the morphological, cytogenetic, biochemical and anatomical parameters in salt (NaCl) stressed Allium cepa L.

Salinity is one of the most important abiotic stress factors that negatively affect plant growth and development. In contrast, fusaric acid (FA), a mycotoxin produced by Fusarium and Giberella fungal genera, has biological and metabolic effects in various plants. In this study, it was aimed to investigate the protective effect of externally applied FA (0.1 nM) against the damage caused by salt (0.15 M NaCl) stress in onion (Allium cepa L.) plant. Salt stress resulted in an increase in the chromosomal aberrations (CAs) and micronucleus (MN) frequency, a decrease in the mitotic index (MI), fresh weight, root number, germination percentage, and root length. It promoted CAs such as irregular mitosis, bilobulated nuclei, chromosome loss, bridge, unequal seperation of chromosome, vagrant chromosome and polar slip in root meristem cells. In addition, salt stress caused a enhancement in free proline (PR), catalase (CAT), superoxide dismutase (SOD) and malondialdehyde (MDA) contents in the roots of onion plant. Moreover, it revealed damage and changes that include the accumulation of some chemical substances such as proline and sugars in epidermis and cortex layer cells, epidermal cell injury, flattening of the cell nucleus, wall thickening in cortex cells, necrotic areas and indistinct transmission tissue in the anatomical structure of onion roots. On the other hand, FA application promoted bulb germination and mitotic activity, strengthened the antioxidant defense system, and reduced chromosome and anatomical structure damages. In conclusion; it has been revealed that exogenous FA application may have a positive effect on increasing the resistance of onion plants to salt stress.

New insights into the anti-inflammatory and anti-melanoma mechanisms of action of azelaic acid and other Fusarium solani metabolites via in vitro and in silico studies.

Metabolites exploration of the ethyl acetate extract of Fusarium solani culture broth that was isolated from Euphorbia tirucalli root afforded five compounds; 4-hydroxybenzaldehyde (1), 4-hydroxybenzoic acid (2), tyrosol (3), azelaic acid (4), malic acid (5), and fusaric acid (6). Fungal extract as well as its metabolites were evaluated for their anti-inflammatory and anti-hyperpigmentation potential via in vitro cyclooxygenases and tyrosinase inhibition assays, respectively. Azelaic acid (4) exhibited powerful and selective COX-2 inhibition followed by fusaric acid (6) with IC50 values (2.21 ± 0.06 and 4.81 ± 0.14 µM, respectively). As well, azelaic acid (4) had the most impressive tyrosinase inhibitory effect with IC50 value of 8.75 ± 0.18 µM compared to kojic acid (IC50 = 9.27 ± 0.19 µM). Exclusive computational studies of azelaic acid and fusaric acid with COX-2 were in good accord with the in vitro results. Interestingly, this is the first time to investigate and report the potential of compounds 3-6 to inhibit cyclooxygenase enzymes. One of the most invasive forms of skin cancer is melanoma, a molecular docking study using a set of enzymes related to melanoma suggested pirin to be therapeutic target for azelaic acid and fusaric acid as a plausible mechanism for their anti-melanoma activity.

Alkaline pH, Low Iron Availability, Poor Nitrogen Sources and CWI MAPK Signaling Are Associated with Increased Fusaric Acid Production in Fusarium oxysporum.

Fusaric acid (FA) is one of the first secondary metabolites isolated from phytopathogenic fungi belonging to the genus Fusarium. This molecule exerts a toxic effect on plants, rhizobacteria, fungi and animals, and it plays a crucial role in both plant and animal pathogenesis. In plants, metal chelation by FA is considered one of the possible mechanisms of action. Here, we evaluated the effect of different nitrogen sources, iron content, extracellular pH and cellular signalling pathways on the production of FA siderophores by the pathogen Fusarium oxysporum (Fol). Our results show that the nitrogen source affects iron chelating activity and FA production. Moreover, alkaline pH and iron limitation boost FA production, while acidic pH and iron sufficiency repress it independent of the nitrogen source. FA production is also positively regulated by the cell wall integrity (CWI) mitogen-activated protein kinase (MAPK) pathway and inhibited by the iron homeostasis transcriptional regulator HapX. Collectively, this study demonstrates that factors promoting virulence (i.e., alkaline pH, low iron availability, poor nitrogen sources and CWI MAPK signalling) are also associated with increased FA production in Fol. The obtained new insights on FA biosynthesis regulation can be used to prevent both Fol infection potential and toxin contamination.

Screening Biocontrol Agents for Cash Crop Fusarium Wilt Based on Fusaric Acid Tolerance and Antagonistic Activity against Fusarium oxysporum.

Fusarium wilt, caused by Fusarium oxysporum, is one of the most notorious diseases of cash crops. The use of microbial fungicides is an effective measure for controlling Fusarium wilt, and the genus Bacillus is an important resource for the development of microbial fungicides. Fusaric acid (FA) produced by F. oxysporum can inhibit the growth of Bacillus, thus affecting the control efficacy of microbial fungicides. Therefore, screening FA-tolerant biocontrol Bacillus may help to improve the biocontrol effect on Fusarium wilt. In this study, a method for screening biocontrol agents against Fusarium wilt was established based on tolerance to FA and antagonism against F. oxysporum. Three promising biocontrol bacteria, named B31, F68, and 30833, were obtained to successfully control tomato, watermelon, and cucumber Fusarium wilt. Strains B31, F68, and 30833 were identified as B. velezensis by phylogenetic analysis of the 16S rDNA, gyrB, rpoB, and rpoC gene sequences. Coculture assays revealed that strains B31, F68, and 30833 showed increased tolerance to F. oxysporum and its metabolites compared with B. velezensis strain FZB42. Further experiments confirmed that 10 µg/mL FA completely inhibited the growth of strain FZB42, while strains B31, F68, and 30833 maintained normal growth at 20 µg/mL FA and partial growth at 40 µg/mL FA. Compared with strain FZB42, strains B31, F68, and 30833 exhibited significantly greater tolerance to FA.

A MAPK signaling cascade regulates the fusaric acid-induced cell death in Arabidopsis.

Mycotoxin contamination of foods and feeds is a global problem. Fusaric acid (FA) is a mycotoxin produced by Fusarium species that are phytopathogens of many economically important plant species. FA can cause programmed cell death (PCD) in several plant species. However, the signaling mechanisms of FA-induced cell death in plants are largely unknown. Here we showed that FA induced cell death in the model plant Arabidopsis thaliana, and MPK3/6 phosphorylation was triggered by FA in Arabidopsis. Both the acid nature and the radical of FA are required for its activity in inducing MPK3/6 activation and cell death. Expression of the constitutively active MKK5DD resulted in the activation of MPK3/6 and promoted the FA-induced cell death. Our work demonstrates that the MKK5-MPK3/6 cascade positively regulates FA-induced cell death in Arabidopsis and also provides insight into the mechanisms of how cell death is induced by FA in plants.

Fusaric acid inhibits proliferation and induces apoptosis through triggering endoplasmic reticulum stress in MCF-7 human breast cancer cells.

Breast cancer has replaced lung cancer to be the leading cancer in the world. Currently, chemotherapy is still the major method for breast cancer therapy, but its overall effect remains unsatisfactory. Fusaric acid (FSA), a mycotoxin derived from fusarium species, has shown potency against the proliferation of several types of cancer cells, but its effect on breast cancer cells has not been examined. Therefore, we explored the possible effect of FSA on the proliferation of MCF-7 human breast cancer cells and uncovered the underlying mechanism in the present study. Our results showed that FSA has a strong anti-proliferative effect on MCF-7 cells through inducing ROS production, apoptosis and arresting cell cycle at G2/M transition phase. Additionally, FSA triggers endoplasmic reticulum (ER) stress in the cells. Notably, the cell cycle arrest and apoptosis inducing effect of FSA can be attenuated by ER stress inhibitor, tauroursodeoxycholic acid. Our study provide evidence that FSA is a potent proliferation inhibition and apoptosis inducing agent against human breast cancer cells, and the possible mechanism involves the activation of ER stress signaling pathways. Our study may highlight that FSA is promising for the future in vivo study and development of potential agent for breast cancer therapy.

Ethylene-dependent regulation of oxidative stress in the leaves of fusaric acid-treated tomato plants.

The mycotoxin fusaric acid (FA) induces rapid oxidative burst leading to cell death in plants. At the same time, plant defence reactions are mediated by several phytohormones for instance ethylene (ET). However, previously conducted studies leave research gaps on how ET plays a regulatory role under mycotoxin exposure. Therefore, this study aims to the time-dependent effects of two FA concentrations (0.1 mM and 1 mM) were explored on the regulation of reactive oxygen species (ROS) in leaves of wild-type (WT) and ET receptor mutant Never ripe (Nr) tomatoes. FA induced superoxide and H2O2 accumulation in both genotypes in a mycotoxin dose- and exposure time-dependent pattern. 1 mM FA activated NADPH oxidase (+34% compared to the control) and RBOH1 transcript levels in WT leaves. However, superoxide production was significantly higher in Nr with 62% which could contribute to higher lipid peroxidation in this genotype. In parallel, the antioxidative defence mechanisms were also activated. Both peroxidase and superoxide dismutase activities were lower in Nr but ascorbate peroxidase showed one-fold higher activity under 1 mM FA stress than in WT leaves. Interestingly, catalase (CAT) activity decreased upon FA in a time- and concentration-dependent manner and the encoding CAT genes were also downregulated, especially in Nr leaves at 20%. Ascorbate level was decreased and glutathione remained lower in Nr than WT plants under FA exposure. Conclusively, Nr genotype showed more sensitivity to FA-induced ROS suggesting that ET serves defence reactions of plants by activating several enzymatic and non-enzymatic antioxidants to detoxify excess ROS accumulation.

In vitro acanthamoebicidal activity of fusaric acid and dehydrofusaric acid from an endophytic fungus Fusarium sp. Tlau3.

Acanthamoeba is a genus of free-living protozoa that can cause sight- and life-threatening diseases in man. Its control is still problematic due to the lack of effective and nontoxic acanthamoebicidal agents. Herein, we report the first finding of an in vitro killing effect of fusaric acid and dehydrofusaric acid, isolated from metabolites of the Fusarium fujikuroi species complex Tlau3, on Acanthamoeba trophozoites isolated from two clinical (AS, AR) and two soil (S3, S5) samples. AS, AR, and S3 were classified as members of the T4 genotype, whereas S5 belongs to T5. The fungal extract was found to exhibit acanthamoebicidal activity, and activity-guided fractionation led to the isolation and identification of active principles, fusaric acid and dehydrofusaric acid. Their effects were in concentration- and time-dependent manners. Fusaric acid and dehydrofusaric acid showed IC50 values against AS trophozoites of 0.31 and 0.34 µM, respectively. Commercial fusaric acid displayed the same acanthamoebicidal activity as that of the isolated fusaric acid, and therefore, commercial fusaric acid was used throughout this study. IC50 values of commercial fusaric acid against AR, S3, and S5 trophozoites were 0.33, 0.33, and 0.66 µM, respectively. Fusaric acid calcium salt has a history of usage as a hypotensive agent in humans with no observed toxicity. The present study suggests that fusaric acid may serve as a starting point for the development towards therapeutic and environmental acanthamoebicides with low toxicity to humans.

Biosensor approach for electrochemical quantitative assessment and qualitative characterization of the effect of fusaric acid on a culture-receptor.

Fusaric acid (FA) is a secondary fungal metabolite, which is widespread on corn and corn-based feed and food; FA has non-specific toxicity. Biosensor method is an express and easy-to-use method for quantitative and qualitative assessment of FA effect. Search for cultures has been performed for the formation of laboratory models of FA biosensor with the Clark-type oxygen electrode as transducer: respiration intensity of chosen cultures changed in the presence of FA. Resting cells of Fusarium oxysporum f. sp. vasinfectum and Bacillus subtilis were used as receptors of the amperometric biosensor for FA determination in aqueous solution. To enhance the sensitivity of detection, induction by substrate was performed for Bacillus subtilis. Response-concentration linear dependencies were obtained in a range of 0.5-500 FA mg/L. Biosensor models were applied to characterize influence of FA on microbial cells and investigate some features of FA transport. The dependences of the cells' response to FA on FA concentration were obtained; the kinetic parameters S0.5 and Vmax were determined for each culture. Inhibition-threshold FA (Sit) concentrations were similar for both studied cultures. At concentrations lower than Sit, the process of simple diffusion governed FA transport into cells and caused the cells' response to FA for non-induced culture.