Ibrexafungerp is efficacious in a neutropenic murine model of pulmonary mucormycosis as monotherapy and combined with liposomal amphotericin B.

Ibrexafungerp (formerly SCY-078) is the first member of the triterpenoid class that prevents the synthesis of the fungal cell wall polymer ß-(1,3)-D-glucan by inhibiting the enzyme glucan synthase. We evaluated the in vivo efficacy of ibrexafungerp against pulmonary mucormycosis using an established murine model. Neutropenic mice were intratracheally infected with either Rhizopus delemar or Mucor circinelloides. Treatment with placebo (diluent control), ibrexafungerp (30 mg/kg, PO BID), liposomal amphotericin B (LAMB 10 mg/kg IV QD), posaconazole (PSC 30 mg/kg PO QD), or a combination of ibrexafungerp plus LAMB or ibrexafungerp plus PSC began 16 h post-infection and continued for 7 days for ibrexafungerp or PSC and through day 4 for LAMB. Ibrexafungerp was as effective as LAMB or PSC in prolonging median survival (range: 15 days to >21 days) and enhancing overall survival (30%-65%) vs placebo (9 days and 0%; P < 0.001) in mice infected with R. delemar. Furthermore, median survival and overall percent survival resulting from the combination of ibrexafungerp plus LAMB were significantly greater compared to all monotherapies (P ≤ 0.03). Similar survival results were observed in mice infected with M. circinelloides. Monotherapies also reduce the lung and brain fungal burden by ~0.5-1.0log10 conidial equivalents (CE)/g of tissue vs placebo in mice infected with R. delemar (P < 0.05), while a combination of ibrexafungerp plus LAMB lowered the fungal burden by ~0.5-1.5log10 CE/g compared to placebo or any of the monotherapy groups (P < 0.03). These results are promising and warrant continued investigation of ibrexafungerp as a novel treatment option against mucormycosis.

Broad antifungal resistance mediated by RNAi-dependent epimutation in the basal human fungal pathogen Mucor circinelloides.

Mucormycosis-an emergent, deadly fungal infection-is difficult to treat, in part because the causative species demonstrate broad clinical antifungal resistance. However, the mechanisms underlying drug resistance in these infections remain poorly understood. Our previous work demonstrated that one major agent of mucormycosis, Mucor circinelloides, can develop resistance to the antifungal agents FK506 and rapamycin through a novel, transient RNA interference-dependent mechanism known as epimutation. Epimutations silence the drug target gene and are selected by drug exposure; the target gene is re-expressed and sensitivity is restored following passage without drug. This silencing process involves generation of small RNA (sRNA) against the target gene via core RNAi pathway proteins. To further elucidate the role of epimutation in the broad antifungal resistance of Mucor, epimutants were isolated that confer resistance to another antifungal agent, 5-fluoroorotic acid (5-FOA). We identified epimutant strains that exhibit resistance to 5-FOA without mutations in PyrF or PyrG, enzymes which convert 5-FOA into the active toxic form. Using sRNA hybridization as well as sRNA library analysis, we demonstrate that these epimutants harbor sRNA against either pyrF or pyrG, and further show that this sRNA is lost after reversion to drug sensitivity. We conclude that epimutation is a mechanism capable of targeting multiple genes, enabling Mucor to develop resistance to a variety of antifungal agents. Elucidation of the role of RNAi in epimutation affords a fuller understanding of mucormycosis. Furthermore, it improves our understanding of fungal pathogenesis and adaptation to stresses, including the evolution of drug resistance.

Microwave prompted solvent-free synthesis of new series of heterocyclic tagged 7-arylidene indanone hybrids and their computational, antifungal, antioxidant, and cytotoxicity study.

In this study, we report the expeditious synthesis of ten new antifungal and antioxidant agents containing heterocyclic linked 7-arylidene indanone moiety. The solvent-free microwave technique, ample substrate scope, superfast synthesis, and very simple operation are noteworthy features of this protocol. Antifungal activities of the newly synthesized compounds were evaluated against four fungal strains namely Rhizophus oryzae, Mucor mucido, Aspergillus niger, and Candida albicans. Most of the compounds were shown strong inhibition of the investigated fungal agents. In vitro, antioxidant potential against DPPH and OH radicals affirmed that the synthesized compounds are good to excellent radicals scavenging agents. The cytotoxicity data of the synthesized compounds towards HL-60 cells uncovered that the synthesized compounds display very low to negligible cytotoxicity. The structural and quantum chemical parameters of the synthesized compounds were explored by employing density functional theory (DFT) at B3LYP functional using 6-311G(d,p) basis set. The compound 3a is discussed in detail for the theoretical and experimental correlation. Time-dependent density functional theory (TD-DFT) at CAM-B3LYP functional with 6-311G(d,p) basis set was used for the electronic absorption study in the gas phase and indichloromethane and benzene solvents. The UV-Visible absorption peaks and fundamental vibrational wavenumbers were computed and a good agreement between observed and theoretical results has been achieved. From the DFT and antifungal activity correlation, it has been found that the 7-heteroarylidene indanones with more stabilized LUMO energy levels display good antifungal potential.

Invasive Mucormycosis in Children With Malignancies: Report From the Infection Working Group of the Hellenic Society of Pediatric Hematology-Oncology.

Mucormycosis is an invasive, life-threatening fungal infection that mainly affects immunocompromised hosts. We collected data of pediatric mucormycosis cases from all 7 Greek Hematology-Oncology Departments for the years 2008-2017. Six cases of invasive mucormycosis diagnosed during treatment for malignancies were included in the study. In 4 children (66%) mucormycosis occurred within the first 20 days after diagnosis of the underlying disease. Two cases were classified as proven mucormycosis and 4 as probable. The most frequently recorded species was Rhizopus arrhizus (2 patients), followed by Mucor spp (1), and Lichtheimia spp (1). All patients received liposomal amphotericin B. Combined antifungal treatment was used in 5 cases. Surgical excision was performed in 4 cases (66%). Two patients died at 6 and 12 months after the diagnosis, respectively, 1 (17%) because of mucormycosis. Our data suggest that mucormycosis may occur early after the initiation of intensive chemotherapy in children with malignancies.

Frequency of occurrence, seasonal variation and antifungal susceptibility of opportunistic Mucorales isolated from hospital soils in Iran.

BACKGROUND: Mucorales are opportunistic pathogens that can cause life-threatening diseases predominantly in immunocompromised patients. OBJECTIVES: This study aimed to investigate the frequency, seasonal variation and antifungal susceptibility of pathogenic Mucorales in the soil collected from seven hospitals in Urmia, Iran, between November 2017 and July 2018 in four different seasons. METHODS: Mucorales isolates obtained from soil were characterised based on conventional and molecular assays. In addition, in vitro antifungal susceptibility was performed using the CLSI M38Ed3 procedure. RESULTS: Out of 196 tested soil samples, 80 (40.8%) samples were positive for mucoralean fungi. Rhizopus arrhizus var. arrhizus (n = 47) was the most frequent species followed by Mucor circinelloides (n = 21) and Cunninghamella echinulata (n = 6). A seasonal variation in the frequency of Mucorales in soil was detected with a maximum of culture-positive soil samples detected in wet autumn (43.2%) followed by winter (23.4%), summer (19.7%) and spring (13.6%). In vitro antifungal susceptibility testing for 80 environmental isolates exhibited MIC of ≤2 µg/ml for amphotericin B indicating the smallest range of MIC variation among the tested Mucorales (range: 0.125-2 µg/ml). Among the azoles, posaconazole was the most effective antifungals (GM MIC, 0.724 µg/ml). CONCLUSIONS: We considered associations of species and seasonal frequencies between soil mucoralean fungi and mucormycosis. The effect of opportunistic Mucorales dominating in the soil and prevalent causative agents of mucormycosis in Iran reported in the literatures but more comprehensive studies are needed to confirm this conclusion.

Two New Triterpenes from Basidiomata of the Medicinal and Edible Mushroom, Laetiporus sulphureus.

In the search for novel anti-infectives from natural sources, fungi, in particular basidiomycetes, have proven to still harbor so much potential in terms of secondary metabolites diversity. There have been numerous reports on isolating numerous secondary metabolites from genus Laetiporus. This study reports on two new triterpenoids, laetiporins C and D, and four known triterpenes from the fruiting body of L. sulphureus. The structures of the isolated compounds were elucidated based on their 1D and 2D nuclear magnetic resonance (NMR) spectroscopic data in combination with high-resolution electrospray mass spectrometric (HR-ESIMS) data. Laetiporin C exhibited weak antifungal activity against Mucor hiemalis. Furthermore, the compounds showed weak antiproliferative activity against the mouse fibroblast L929 and human cancer cell lines, including KB-3-1, A431, MCF-7, PC-3 and A549.

Alteration of Fermentative Metabolism Enhances Mucor circinelloides Virulence.

The fungus Mucor circinelloides undergoes yeast-mold dimorphism, a developmental process associated with its capability as a human opportunistic pathogen. Dimorphism is strongly influenced by carbon metabolism, and hence the type of metabolism likely affects fungus virulence. We investigated the role of ethanol metabolism in M. circinelloides virulence. A mutant in the adh1 gene (M5 strain) exhibited higher virulence than the wild-type (R7B) and the complemented (M5/pEUKA-adh1+) strains, which were nonvirulent when tested in a mouse infection model. Cell-free culture supernatant (SS) from the M5 mutant showed increased toxic effect on nematodes compared to that from R7B and M5/pEUKA-adh1+ strains. The concentration of acetaldehyde excreted by strain M5 in the SS was higher than that from R7B, which correlated with the acute toxic effect on nematodes. Remarkably, strain M5 showed higher resistance to H2O2, resistance to phagocytosis, and invasiveness in mouse tissues and induced an enhanced systemic inflammatory response compared with R7B. The mice infected with strain M5 under disulfiram treatment exhibited only half the life expectancy of those infected with M5 alone, suggesting that acetaldehyde produced by M. circinelloides contributes to the toxic effect in mice. These results demonstrate that the failure in fermentative metabolism, in the step of the production of ethanol in M. circinelloides, contributes to its virulence, inducing a more severe tissue burden and inflammatory response in mice as a consequence of acetaldehyde overproduction.

A Case Series and Literature Review of Isavuconazole Use in Pediatric Patients with Hemato-oncologic Diseases and Hematopoietic Stem Cell Transplantation.

We analyzed the use of isavuconazole (ISA) as treatment or prophylaxis for invasive fungal disease (IFD) in children with hemato-oncologic diseases. A multicentric retrospective analysis was performed among centers belonging to the Italian Association for Pediatric Hematology and Oncology (AIEOP). Pharmacokinetic (PK) monitoring was applied by a high-performance liquid chromatography-tandem mass spectrometry (HLPC-MS/MS) assay. Twenty-nine patients were studied: 10 during chemotherapy and 19 after allogeneic hematopoietic stem cell transplantation (HSCT). The patients consisted of 20 males and 9 females with a median age of 14.5 years (age range, 3 to 18 years) and a median body weight of 47 kg (body weight range, 15 to 80 kg). ISA was used as prophylaxis in 5 patients and as treatment in 24 cases (20 after therapeutic failure, 4 as first-line therapy). According to European Organization for Research and Treatment of Cancer (EORTC) criteria, we registered 5 patients with proven IFD, 9 patients with probable IFD, and 10 patients with possible IFD. Patients with a body weight of <30 kg received half the ISA dose; the others received ISA on the adult schedule (a 200-mg loading dose every 8 h on days 1 and 2 and a 200-mg/day maintenance dose); for all but 10 patients, the route of administration switched from the intravenous route to the oral route during treatment. ISA was administered for a median of 75.5 days (range, 6 to 523 days). The overall response rate was 70.8%; 12 patients with IFD achieved complete remission, 5 achieved partial remission, 5 achieved progression, and 3 achieved stable IFD. No breakthrough infections were registered. PK monitoring of 17 patients revealed a median ISA steady-state trough concentration of 4.91 mg/liter (range, 2.15 to 8.54 mg/liter) and a concentration/dose (in kilograms) ratio of 1.13 (range, 0.47 to 3.42). Determination of the 12-h PK profile was performed in 6 cases. The median area under the concentration-time curve from 0 to 12 h was 153.16 mg·h/liter (range, 86.31 to 169.45 mg·h/liter). Common Terminology Criteria for Adverse Events grade 1 to 3 toxicity (increased transaminase and/or creatinine levels) was observed in 6 patients, with no drug-drug interactions being seen in patients receiving immunosuppressants. Isavuconazole may be useful and safe in children with hemato-oncologic diseases, even in the HSCT setting. Prospective studies are warranted.

Antifungal drug resistance evoked via RNAi-dependent epimutations.

Microorganisms evolve via a range of mechanisms that may include or involve sexual/parasexual reproduction, mutators, aneuploidy, Hsp90 and even prions. Mechanisms that may seem detrimental can be repurposed to generate diversity. Here we show that the human fungal pathogen Mucor circinelloides develops spontaneous resistance to the antifungal drug FK506 (tacrolimus) via two distinct mechanisms. One involves Mendelian mutations that confer stable drug resistance; the other occurs via an epigenetic RNA interference (RNAi)-mediated pathway resulting in unstable drug resistance. The peptidylprolyl isomerase FKBP12 interacts with FK506 forming a complex that inhibits the protein phosphatase calcineurin. Calcineurin inhibition by FK506 blocks M. circinelloides transition to hyphae and enforces yeast growth. Mutations in the fkbA gene encoding FKBP12 or the calcineurin cnbR or cnaA genes confer FK506 resistance and restore hyphal growth. In parallel, RNAi is spontaneously triggered to silence the fkbA gene, giving rise to drug-resistant epimutants. FK506-resistant epimutants readily reverted to the drug-sensitive wild-type phenotype when grown without exposure to the drug. The establishment of these epimutants is accompanied by generation of abundant fkbA small RNAs and requires the RNAi pathway as well as other factors that constrain or reverse the epimutant state. Silencing involves the generation of a double-stranded RNA trigger intermediate using the fkbA mature mRNA as a template to produce antisense fkbA RNA. This study uncovers a novel epigenetic RNAi-based epimutation mechanism controlling phenotypic plasticity, with possible implications for antimicrobial drug resistance and RNAi-regulatory mechanisms in fungi and other eukaryotes.

A Revised Species Concept for Opportunistic Mucor Species Reveals Species-Specific Antifungal Susceptibility Profiles.

Recently, the species concept of opportunistic Mucor circinelloides and its relatives has been revised, resulting in the recognition of its classical formae as independent species and the description of new species. In this study, we used isolates of all clinically relevant Mucor species and performed susceptibility testing using the EUCAST reference method to identify potential species-specific susceptibility patterns. In vitro susceptibility profiles of 101 mucoralean strains belonging to the genus Mucor (72), the closely related species Cokeromyces recurvatus (3), Rhizopus (12), Lichtheimia (10), and Rhizomucor (4) to six antifungals (amphotericin B, natamycin, terbinafine, isavuconazole, itraconazole, and posaconazole) were determined. The most active drug for all Mucorales was amphotericin B. Antifungal susceptibility profiles of pathogenic Mucor species were specific for isavuconazole, itraconazole, and posaconazole. The species formerly united in M. circinelloides showed clear differences in their antifungal susceptibilities. Cokeromyces recurvatus, Mucor ardhlaengiktus, Mucor lusitanicus (M. circinelloides f. lusitanicus), and Mucor ramosissimus exhibited high MICs to all azoles tested. Mucor indicus presented high MICs for isavuconazole and posaconazole, and Mucor amphibiorum and Mucor irregularis showed high MICs for isavuconazole. MIC values of Mucor spp. for posaconazole, isavuconazole, and itraconazole were high compared to those for Rhizopus and the Lichtheimiaceae (Lichtheimia and Rhizomucor). Molecular identification combined with in vitro susceptibility testing is recommended for Mucor species, especially if azoles are applied in treatment.

Role of Arf-like proteins (Arl1 and Arl2) of Mucor circinelloides in virulence and antifungal susceptibility.

Mucor circinelloides is an etiologic agent of mucormycosis, a fungal infection produced by Mucorales often associated with mortality due to unavailability of antifungal drugs. Arl proteins belong to the Arf family and are involved in vesicle trafficking and tubulin assembly. This study identified two Arl (Arf-like)-encoding genes, arl1 and arl2, in M. circinelloides and explored their function in morphogenesis, virulence, and antifungal susceptibility. Although Arl1 and Arl2 proteins shared 55% amino acid sequence identity, arl1 and arl2 genes showed distinct transcriptional expression patterns. arl1 was expressed at higher levels than arl2 and induced in mycelia, suggesting a role in morphological transitions. Disruption of the arl1 and arl2 genes led to heterokaryon (Δarl1(+)(-)) and homokaryon (Δarl2) genotypes, respectively. The incapacity to generate homokaryon mutants for arl1 suggested that it is essential for growth of M. circinelloides. Deletion of each gene reduced the expression of the other, suggesting the existence of a positive cross-regulation between them. Thus, deletion of arl2 resulted in a ~60% reduction of arl1 expression, whereas the Δarl1(+)(-) showed ∼90% reduction of arl1 expression. Mutation of arl2 showed no phenotype or a mild phenotype between Δarl1(+)(-) and wild-type (WT), suggesting that all observed phenotypes in both mutant strains corresponded to arl1 low expression. The Δarl1(+)(-) produced a small amount of spores that showed increased sensitivity to dodecyl-sulfate and azoles, suggesting a defect in the cell wall that was further supported by decrease in saccharide content. These defects in the cell wall were possibly originated by abnormal vesicle trafficking since FM4-64 staining of both mutants Δarl1(+)(-) and Δarl2 revealed less well-localized endosomes compared to the WT. Moreover, aberrant vesicle trafficking may be responsible for the secretion of specific virulence-related proteins since cell-free medium from Δarl1(+)(-) were found to increase killing of Caenorhabditis elegans compared to WT.

Galleria mellonella as a model system to study virulence potential of mucormycetes and evaluation of antifungal treatment.

Mucorales can cause cutaneous to deep-seated infections, mainly in the immunocompromised host, resulting in high mortality rates due to late and inefficient treatment. In this study, Galleria mellonella larvae were evaluated as a heterologous invertebrate host to study pathogenicity of clinically relevant mucormycetes (Rhizopus spp., Rhizomucor spp., Lichtheimia spp., Mucor spp.). All tested species were able to infect G. mellonella larvae. Virulence potential was species-specific and correlated to clinical relevance. Survival of infected larvae was dependent on (a) the species (growth speed and spore size), (b) the infection dose, (c) the incubation temperature, (d) oxidative stress tolerance, and (e) iron availability in the growth medium. Moreover, we exploited the G. mellonella system to determine antifungal efficacy of liposomal amphotericin B, posaconazole, isavuconazole, and nystatin-intralipid. Outcome of in vivo treatment was strongly dependent upon the drug applied and the species tested. Nystatin-intralipid exhibited best activity against Mucorales, followed by posaconazole, while limited efficacy was seen for liposomal amphotericin B and isavuconazole. Pharmacokinetic properties of the tested antifungals within this alternative host system partly explain the limited treatment efficacy. In conclusion, G. mellonella represents a useful invertebrate infection model for studying virulence of mucormycetes, while evaluation of treatment response was limited.

Development of antifungal ingredients for dairy products: From in vitro screening to pilot scale application.

Biopreservation represents a complementary approach to traditional hurdle technologies for reducing microbial contaminants (pathogens and spoilers) in food. In the dairy industry that is concerned by fungal spoilage, biopreservation can also be an alternative to preservatives currently used (e.g. natamycin, potassium sorbate). The aim of this study was to develop antifungal fermentates derived from two dairy substrates using a sequential approach including an in vitro screening followed by an in situ validation. The in vitro screening of the antifungal activity of fermentates derivating from 430 lactic acid bacteria (LAB) (23 species), 70 propionibacteria (4 species) and 198 fungi (87 species) was performed against four major spoilage fungi (Penicillium commune, Mucor racemosus, Galactomyces geotrichum and Yarrowia lipolytica) using a cheese-mimicking model. The most active fermentates were obtained from Lactobacillus brevis, Lactobacillus buchneri, Lactobacillus casei/paracasei and Lactobacillus plantarum among the tested LAB, Propionibacterium jensenii among propionibacteria, and Mucor lanceolatus among the tested fungi. Then, for the 11 most active fermentates, culture conditions were optimized by varying incubation time and temperature in order to enhance their antifungal activity. Finally, the antifungal activity of 3 fermentates of interest obtained from Lactobacillus rhamnosus CIRM-BIA1952, P. jensenii CIRM-BIA1774 and M. lanceolatus UBOCC-A-109193 were evaluated in real dairy products (sour cream and semi-hard cheese) at a pilot-scale using challenge and durability tests. In parallel, the impact of these ingredients on organoleptic properties of the obtained products was also assessed. In semi-hard cheese, application of the selected fermentates on the cheese surface delayed the growth of spoilage molds for up to 21 days, without any effect on organoleptic properties, P. jensenii CIRM-BIA1774 fermentate being the most active. In sour cream, incorporation of the latter fermentate at 2 or 5% yielded a high antifungal activity but was detrimental to the product organoleptic properties. Determination of the concentration limit, compatible with product acceptability, showed that incorporation of this fermentate at 0.4% prevented growth of fungal contaminants in durability tests but had a more limited effect against M. racemosus and P. commune in challenge tests. To our knowledge, this is the first time that the workflow followed in this study, from in vitro screening using dairy matrix to scale-up in cheese and sour cream, is applied for production of natural ingredients relying on a large microbial diversity in terms of species and strains. This approach allowed obtaining several antifungal fermentates which are promising candidates for dairy products biopreservation.

In vitro analysis of growth patterns of invasive fungal species on commonly used endonasal hemostatic agents.

OBJECTIVE: Previous studies have not examined the potential role of endonasal hemostatic agents in facilitating growth of fungal species. We aim to determine the possibility of these to serve as a nutrient source for fungal growth. METHODS: Cultures of Aspergillus, Fusarium, and Mucor were harvested and placed in solution in sterile saline at standardized high and low concentrations. Thrombin gelatin matrix, carboxyl methylcelluose, and potato starch derivative agents were prepared following manufacturer instructions and applied to two separate Petri dishes per agent. Each substrate was then inoculated with either high or low concentrations of fungal species. Negative and positive control plates with each organism were included. Dishes were sealed, incubated, and examined daily for fourteen days for microscopic and macroscopic growth. RESULTS: Thrombin gelatin matrix was relatively resilient to growth, although Fusarium growth was noted on all packing material by day three. Carboxyl methylcellulose also supported growth of high-concentration Mucor appreciated on day five. The potato starch derivative supported fulminant growth of all fungal species. CONCLUSIONS: Endonasal hemostatic agents may be nutrient sources that facilitate growth of fungal species. This may be a consideration in a surgeon's decision to use a hemostatic agent. Prompt initial post-operative debridement may be warranted in select patients. Our findings serve as a model for further testing of fungal growth on other hemostatic materials. Future studies are needed to confirm the clinical significance of these findings in vivo.

Mucor sp.-An endophyte of Brassicaceae capable of surviving in toxic metal-rich sites.

The aim of this study was to assess the biodiversity of endophytic fungi from Arabidopsis arenosa growing on a post mining waste dump and to evaluate their role in plant adaptation to metal toxicity. Severeal of the fungi were beneficial for the plant. Among them, a fungus belonging to the Mucor genus, was found to interact with a broad range of plants, including Brassicaceae metallophytes. Mucor sp. was shown to be highly tolerant to elevated levels of Zn, Cd, and Pb and to accelerate plant-host growth under either toxic-metal stress or control conditions. When inoculated with Mucor sp., A. arenosa under toxic-metal stress acquired more N and showed significantly down-regulated catalase activity, which suggests suppression of toxic-metal-induced oxidative stress. We used the model plant-A. thaliana to evaluate the dynamics of plant-tissue colonization by the fungus as monitored with qPCR and to analyze the host's transcriptome response during early stages of the interaction. The results revealed the induction of a plant-defense and stress-related response on the 5th day of co-culture, which was in accord with the decrease of fungal abundance in shoots on the 6th day of interaction. Presented results demonstrate the importance of endophytic fungi in plant toxic-metal tolerance.

[The 471st case: duodenal ulcer, mucor infection, and elevated IgG(4)].

Mucor infection is rarely reported in non-immunocompromised population, especially in isolated gastrointestinal tracts. IgG(4)-related diseases (IgG(4)-RD) have been recognized in recent years, but secondary causes of IgG(4) elevation should be differentiated. We reported a young man with duodenal mass and ulcer and high serum IgG(4) level. Histological biopsy of the mass revealed positive mucor mycelium and infiltration of IgG(4) positive plasma cells. Serum IgG(4) decreased to normal range after surgical resection and systemic antifungal treatment. This case suggests that isolated mucor mycosis infection can develop in the digestive tract and mimics as IgG(4)-related disease.

Biocontrol and plant stimulating potential of novel strain Bacillus sp. PPM3 isolated from marine sediment.

In the current study, the biocontrol potential of a novel strain Bacillus sp. PPM3 isolated from marine sediment from the Red Sea in Hurghada, Egypt is recognized. This novel strain was selected out of 32 isolates based on its ability to suppress the growth of four plant pathogenic fungi: Aspergillus flavus, Fusarium graminearum, Mucor sp. and Alternaria sp. The new marine strain was identified and characterized by phenotypic and molecular approaches. The culture filtrate of Bacillus sp. PPM3 suppressed the growth and spore germination of all tested fungi in vitro with the highest value of inhibition reported for Mucor sp. (97.5%). The antifungal effect of the culture filtrate from the strain PPM3 was due to production of highly stable secondary metabolites resistant to extreme pH, temperature and enzymatic treatments. A PCR analysis confirmed the expression of genes involved in the synthesis of antifungal lipopeptides: iturin, bacillomycin D, mycosubtilin and surfactin. In a greenhouse experiment strain PPM3 effectively reduced disease incidence of F. graminearum in maize plants and displayed additional plant growth stimulating effect. The results show that novel marine strain PPM3 could have a potential in commercial application as biocontrol agent for treatment of various plant diseases caused by soil-borne and postharvest pathogenic fungi.

Pestynol, an Antifungal Compound Discovered Using a Saccharomyces cerevisiae 12geneΔ0HSR-iERG6-Based Assay.

The multidrug-sensitive budding yeast, Saccharomyces cerevisiae 12geneΔ0HSR-iERG6, is very useful in antifungal screens. A novel compound, named pestynol (1), was discovered from a culture of the fungus Pestalotiopsis humus FKI-7473 using the multidrug-sensitive yeast. The structure of 1 was elucidated by NMR studies and modified Mosher's method as (1 R,2 R,3 R,4 R)-( E)-5-(7,11-dimethyl-3-methylenedodeca-6,10-dien-1-yn-1-yl)cyclohex-5-ene-1,2,3,4-tetraol. Compound 1 showed antimicrobial activity against the Gram-positive bacteria, Klebsiella pneumoniae, and S. cerevisiae 12geneΔ0HSR-iERG6 and Mucor racemosus, but displayed only weak cytotoxicity against various human cancer cell lines. Compound 1 displayed antifungal activities against S. cerevisiae 12geneΔ0HSR-iERG6 and Mucor racemosus at 10 µg/disc.

A protein complex bearing an oxidase with napthalene dihydrodiol dehydrogenase activity is induced in Mucor circinelloides strain YR-1 during growth on polycyclic aromatic compounds.

Fungi are organisms capable of growing in a myriad of conditions and respond to counteract environmental cues. Several locations in the world are polluted with oil and its derivatives, and some microorganisms tolerant to these compounds have been isolated. Some fungi can grow in the presence of molecules such as polycyclic aromatic hydrocarbons as sole carbon sources. In this report, we further characterized the induced enzymes with phenanthrene from Mucor circinelloides YR-1 strain, isolated from a polluted field near a petrochemical facility in México. We identified a putative oxidase that is induced when growth with phenanthrene as sole carbon source at a pH of 8.5 and is NADP+ dependent. We show that this enzyme bears naphthalene dihydrodiol dehydrogenase activity with substrate preference for the cis-naphthalene over the trans-naphthalene, with an optimal pH in the range of 8-10. Mass spectrometry analysis revealed that the induced enzyme belongs to the NADP+ oxidase family enzymes with the typical Rossmann-fold for NADP+ binding. This enzyme seems to form a high molecular weight structure (~ 541 kDa) and with a monomer of 57 kDa, suggesting that the multimer is constituted of 10 subunits. Our findings contribute to understanding of the roles that dihydrodiol dehydrogenases have in organisms exposed to toxic compounds in the environment and can regulate their expression.

Activity of Salvia dolomitica and Salvia somalensis Essential Oils against Bacteria, Molds and Yeasts.

Essential oils (EOs) from Salvia dolomitica and Salvia somalensis, widely employed in the cosmetic and perfume industry, were analyzed for composition and tested against bacterial and fungal pathogens isolated from clinical and environmental specimens. The analyses were carried out against Staphylococcus aureus, Staphylococcus pseudointermedius, Pseudomonas aeruginosa, Escherichia coli, Streptococcus canis, Streptococcus pyogenes, Klebsiella pneumoniae, Proteus mirabilis, Microsporum canis, Microsporum gypseum, Trichophyton mentagrophytes, Aspergillus niger, Aspergillus flavus, Candida albicans, Candida krusei, Mucor sp. and Trichothecium roseum. Both EOs showed similar percentages of total monoterpenes and sesquiterpene hydrocarbons. The main constituents were 1,8-cineole and ß-caryophyllene in S. dolomitica and bornyl acetate and camphor in S. somalensis. The selected EOs have no relevant antifungal or antibacterial activities if compared to conventional drugs.