Accumulated melanin in molds provides wavelength-dependent UV tolerance.

Fungal contamination poses a serious threat to public health and food safety because molds can grow under stressful conditions through melanin accumulation. Although ultraviolet (UV) irradiation is popular for inhibiting microorganisms, its effectiveness is limited by our insufficient knowledge about UV tolerance in melanin-accumulating molds. In this study, we first confirmed the protective effect of melanin by evaluating the UV sensitivity of young and mature spores. Additionally, we compared UV sensitivity between spores with accumulated melanin and spores prepared with melanin biosynthesis inhibitors. We found that mature spores were less UV-sensitive than young spores, and that reduced melanin accumulation by inhibitors led to reduced UV sensitivity. These results suggest that melanin protects cells against UV irradiation. To determine the most effective wavelength for inhibition, we evaluated the wavelength dependence of UV tolerance in a yeast (Rhodotorula mucilaginosa) and in molds (Aspergillus fumigatus, Cladosporium halotolerans, Cladosporium sphaerospermum, Aspergillus brasiliensis, Penicillium roqueforti, and Botrytis cinerea). We assessed UV tolerance using a UV-light emitting diode (LED) irradiation system with 13 wavelength-ranked LEDs between 250 and 365 nm, a krypton chlorine (KrCl) excimer lamp device, and a low pressure (LP) Hg lamp device. The inhibition of fungi peaked at around 270 nm, and most molds showed reduced UV sensitivity at shorter wavelengths as they accumulated pigment. Absorption spectra of the pigments showed greater absorption at shorter wavelengths, suggesting greater UV protection at these wavelengths. These results will assist in the development of fungal disinfection systems using UV, such as closed systems of air and water purification.

Changes in Electron Paramagnetic Resonance Parameters Caused by Addition of Amphotericin B to Cladosporium cladosporioides Melanin and DOPA-Melanin-Free Radical Studies.

Cladosporium cladosporioides are the pigmented soil fungi containing melanin. The aim of this work was to determine the influence of amphotericin B on free radicals in the natural melanin isolated from pigmented fungi Cladosporium cladosporioides and to compare it with the effect in synthetic DOPA-melanin. Electron paramagnetic resonance (EPR) spectra were measured at X-band (9.3 GHz) with microwave power in the range of 2.2-70 mW. Amplitudes, integral intensities, linewidths of the EPR spectra, and g factors, were analyzed. The concentrations of free radicals in the tested melanin samples were determined. Microwave saturation of EPR lines indicates the presence of pheomelanin in addition to eumelanin in Cladosporium cladosporioides. o-Semiquinone free radicals in concentrations ~1020 [spin/g] exist in the tested melanin samples and in their complexes with amphotericin B. Changes in concentrations of free radicals in the examined synthetic and natural melanin point out their participation in the formation of amphotericin B binding to melanin. A different influence of amphotericin B on free radical concentration in Cladosporium cladosporioides melanin and in DOPA-melanin may be caused by the occurrence of pheomelanin in addition to eumelanin in Cladosporium cladosporioides. The advanced spectral analysis in the wide range of microwave powers made it possible to compare changes in the free radical systems of different melanin polymers. This study is important for knowledge about the role of free radicals in the interactions of melanin with drugs.

Truncation of a novel C-terminal domain of a ß-glucanase improves its thermal stability and specific activity.

Enzymes that degrade ß-glucan play important roles in various industries, including those related to brewing, animal feed, and health care. Csph16A, an endo-ß-1,3(4)-glucanase encoded by a gene from the halotolerant, xerotolerant, and radiotrophic black fungus Cladosporium sphaerospermum, was cloned and expressed in Pichia pastoris. Two isoforms (Csph16A.1 and Csph16A.2) are produced, arising from differential glycosylation. The proteins were predicted to contain a catalytic Lam16A domain, along with a C-terminal domain (CTD) of unknown function which exhibits minimal secondary structure. Employing PCR-mediated gene truncation, the CTD of Csph16A was excised to assess its functional impact on the enzyme and determine potential alterations in biotechnologically relevant characteristics. The truncated mutant, Csph16A-ΔC, exhibited significantly enhanced thermal stability at 50°C, with D-values 14.8 and 23.5 times greater than those of Csph16A.1 and Csph16A.2, respectively. Moreover, Csph16A-ΔC demonstrated a 20%-25% increase in halotolerance at 1.25 and 1.5 M NaCl, respectively, compared to the full-length enzymes. Notably, specific activity against cereal ß-glucan, lichenan, and curdlan was increased by up to 238%. This study represents the first characterization of a glucanase from the stress-tolerant fungus C. sphaerospermum and the first report of a halotolerant and engineered endo-ß-1,3(4)-glucanase. Additionally, it sheds light on a group of endo-ß-1,3(4)-glucanases from Antarctic rock-inhabiting black fungi harboring a Lam16A catalytic domain and a novel CTD of unknown function.

Cladoic Acid, an Anti-Trypanosoma cruzi Polyketide from Cladosporium sp. TP-F2020.

A new polyketide, cladoic acid, was isolated from a fungus of the genus Cladosporium. The structure of the highly oxygenated trans-decalin ring with an all-E triene side chain was elucidated by extensive spectroscopic analysis. The unique chair/twist-boat conformation of the trans-decalin core and the flexibility of the B-ring were demonstrated by computer-aided conformational analysis. Cladoic acid was active against Trypanosoma cruzi and inhibited the proliferation of amastigotes and epimastigotes with IC50 values of 27 and 46 µM, respectively, but it did not show any appreciable activity against P388 murine leukemia cells, bacteria, or fungi, indicating it is a potential candidate for drug development against Chagas disease.

Early diagnosis of Cladosporium fulvum in greenhouse tomato plants based on visible/near-infrared (VIS/NIR) and near-infrared (NIR) data fusion.

Plant diseases can inflict varying degrees of damage on agricultural production. Therefore, identifying a rapid, non-destructive early diagnostic method is crucial for safeguarding plants. Cladosporium fulvum (C. fulvum) is one of the major diseases in tomato growth. This work presents a method of data fusion using two hyperspectral imaging systems of visible/near-infrared (VIS/NIR) and near-infrared (NIR) spectroscopy for the early diagnosis of C. fulvum in greenhouse tomatoes. First, hyperspectral images of samples at health and different times of infection were collected. The average spectral data of the image regions of interest were extracted and preprocessed for subsequent spectral datasets. Then different classification models were established for VIS/NIR and NIR data, optimized through various variable selection and data fusion methods. The principal component analysis-radial basis function neural network (PCA-RBF) model established using low-level data fusion achieved optimal results, achieving accuracies of 100% and 99.3% for calibration and prediction, respectively. Moreover, both the macro-averaged F1 (Macro-F1) values reached 1, and the geometric mean (G-mean) values reached 1 and 1, respectively. The results indicated that it was feasible to establish a PCA-RBF model by using the hyperspectral technique with low-level data fusion for the early detection of C. fulvum in greenhouse tomatoes.

Resistance to adverse conditions and characterization of Cladosporium species from marine and terrestrial Antarctic samples.

Microbial adaptations to extreme environments can lead to biotechnological applications. This study aimed to evaluate the resistance of Antarctic Cladosporium to adverse conditions (temperature, salinity, UV radiation, and nutrients) and refine their taxonomy. Sequencing and phylogenetic analysis using ITS-act markers resulted in a more accurate taxonomic identification, revealing the presence of five different species, belonging to the complexes C. cladosporioides and C. sphaerospermum. The growth at different temperatures indicates that the soil isolates LAMAI 564 and 1800 (phylogenetically closely related) and LAMAI 2541 are psychrophilic, while the other isolates are psychrotolerant. The fungi isolated from the saline samples LAMAI 595, 616, and 1369 showed better growth results at higher salinity (15%). The fungi most resistant to UV radiation were isolated from terrestrial and marine samples (LAMAI 595, 616, 1800, and 564). LAMAI 595 and 616 (phylogenetically closely related and isolated from the same kind of sample) showed the capacity of nutritional versatility, growing well in both rich and poor-nutrient media. The fungus LAMAI 595 was the most promising for biotechnological application, exceeding the other isolates in the harsh conditions studied. The resistance of the Antarctic Cladosporium to adverse conditions opens new perspectives in the field of applied microbiology of extremophiles.

Experimental evaluation of a full-scale in-duct UV germicidal irradiation system for bioaerosols inactivation.

Bioaerosols control techniques, especially ultraviolet germicidal irradiation (UVGI) are gaining attention due to increasing needs for controlling of health risk caused by airborne biocontaminants. The effectiveness of a full-scale in-duct UVGI air disinfection system was investigated. One bacterium, a wild type Escherichia coli, and three fungal spores, Penicillium aragonense, Rhodotorula glutinis, and Cladosporium sp., were selected as test organisms and their inactivation under different conditions representative of a real application in HVAC systems were investigated. The results demonstrated that inactivation of airborne E. coli by the UVGI system was extremely effective, with >99.5 % of the input E. coli inactivated at a residence time lower than 0.36 s in the disinfection section. Airborne fungal spores were less susceptible to UV irradiation than E. coli. Under same conditions, viable counts reduction of P. aragonense, R. glutinis, and Cladosporium sp. spores were 53 %, 63 % and 73 %, respectively. The effect of UV light intensity, air flowrate and relative humidity were analyzed separately. A simplified model based on redefinition of the parameters in the classical inactivation kinetic equation was used to simulate the inactivation of airborne contaminants in the in-duct system under different conditions. The results showed that the simplified model was adequate to estimate disinfection efficacy of different bioaerosols by the UVGI system which could be useful for system design. Overall, this study shows that such in-duct UVGI systems can provide significant control of bioaerosols.

Cladosporium psychrotolerans strain T01 enhances plant biomass and also exhibits antifungal activity against pathogens.

An increasing number of microorganisms are being identified to enhance plant growth and inhibit phytopathogens. Some Cladosporium species form beneficial associations with plants, either as endophytes or by colonizing the rhizosphere. Herein, we evaluated the influence of the Cladosporium psychrotolerans (T01 strain) fungus on the in vitro growth of Arabidopsis thaliana plantlets through direct and split interactions. After 9 days post-inoculation with C. psychrotolerans, Arabidopsis plantlets exhibited a notable increase in fresh weight and lateral roots, particularly in split interactions. Chlorophyll content increased in both plant-fungus interaction conditions, whereas the primary root was inhibited during direct interaction. We observed an increase in the GUS signal from the Arabidopsis auxin-inducible DR5:uidA marker in lateral root tips in both contact and split fungal interactions, and primary root tips in a split interaction. Arabidopsis and tomato plants cultivated in soil pots and inoculated with C. psychrotolerans (T01 strain) showed a positive effect on biomass production. GC/MS analysis detected that the T01 strain emitted volatile organic compounds (VOCs), predominantly alcohols and aldehydes. These VOCs displayed potent inhibitory effects, with a 60% inhibition against Botrytis cinerea and a 50% inhibition against C. gloeosporioides. Our study demonstrates that C. psychrotolerans T01 has the potential to enhance biomass production and inhibit pathogens, making it a promising candidate for green technology applications.

Bioengineering secreted proteases converts divergent Rcr3 orthologs and paralogs into extracellular immune co-receptors.

Secreted immune proteases "Required for Cladosporium resistance-3" (Rcr3) and "Phytophthora-inhibited protease-1" (Pip1) of tomato (Solanum lycopersicum) are both inhibited by Avirulence-2 (Avr2) from the fungal plant pathogen Cladosporium fulvum. However, only Rcr3 acts as a decoy co-receptor that detects Avr2 in the presence of the Cf-2 immune receptor. Here, we identified crucial residues in tomato Rcr3 that are required for Cf-2-mediated signaling and bioengineered various proteases to trigger Avr2/Cf-2-dependent immunity. Despite substantial divergence in Rcr3 orthologs from eggplant (Solanum melongena) and tobacco (Nicotiana spp.), minimal alterations were sufficient to trigger Avr2/Cf-2-mediated immune signaling. By contrast, tomato Pip1 was bioengineered with 16 Rcr3-specific residues to initiate Avr2/Cf-2-triggered immune signaling. These residues cluster on one side of the protein next to the substrate-binding groove, indicating a potential Cf-2 interaction site. Our findings also revealed that Rcr3 and Pip1 have distinct substrate preferences determined by two variant residues and that both are suboptimal for binding Avr2. This study advances our understanding of Avr2 perception and opens avenues to bioengineer proteases to broaden pathogen recognition in other crops.

Ultra-rapid development of 'solid' aerobic granular sludge by stable transition/filling of inoculated 'hollow' mycelial pellets in hypersaline wastewater.

To overcome the long start-up period in cultivating aerobic granular sludge (AGS) under hypersaline environment, mycelial pellets (MPs) of halotolerant fungus Cladosporium tenuissimum NCSL-XY8 were inoculated to try to realize the ultra-rapid development of salt-tolerant AGS by stable transition of 'hollow' MPs into 'solid' AGS without apparent fragmentation. The granules directly met the standard of AGS after inoculating MPs (Day 0), and it basically satisfied relatively strict standards of AGS (SVI30 < 50 mL/g, D50 > 300 µm, D10 > 200 µm and SVI30/SVI5 > 0.9) under anaerobic/aerobic mode during whole cultivation processes. Microstructure of the granular cross section clarified that MPs with hollow/loose inner layer transitioned into solid/dense AGS under anaerobic/aerobic mode within 7 days, while formed skin-like floating pieces and unstable double-layer hollow granules under aerobic mode. Organics removal reached relatively stable within 13 days under anaerobic/aerobic mode, 6 days faster than aerobic mode. This study provided a strategy for ultra-rapid and stable development of AGS, which showed the shortest granulation period in various AGS-cultivation strategies.

In vitro evaluation of antifungal combinations against neurotropic dematiaceous fungi associated with primary cerebral phaeohyphomycosis.

Primary cerebral phaeohyphomycosis is a life-threatening disease caused by neurotropic dematiaceous fungi. At present, there are no consensus guidelines regarding optimal antifungal therapy in such cases. Generally, a combination of antifungal agents is recommended for treatment. However, the activities of antifungal combinations against these fungi have not been investigated. In this study, we evaluated the in vitro activities of 13 double and five triple antifungal combinations against clinical isolates of Cladophialophora bantiana (n = 7), Fonsecaea monophora (n = 2), and Cladosporium cladosporioides (n = 1), using a simplified checkerboard procedure. The minimum inhibitory concentrations (MICs) of nine antifungal drugs were determined by the broth microdilution method, and the interaction between antifungal agents in each combination was assessed by the fractional inhibitory concentration index. Excellent activity was observed for posaconazole and itraconazole. Flucytosine had potent activity against C. bantiana but was ineffective against F. monophora, and C. cladosporioides. The echinocandins demonstrated high MICs for all the isolates. Synergistic interactions were observed for all the double combinations, except when itraconazole was combined with either amphotericin B or flucytosine. The combination of amphotericin B with caspofungin showed synergistic interactions against 40% of the isolates. Antagonism was observed with isavuconazole-flucytosine combination against two C. bantiana isolates. The triple combinations of caspofungin and flucytosine with amphotericin B or posaconazole were synergistic against one isolate of F. monophora. For C. cladosporioides, synergy was observed for the triple combination of amphotericin B with caspofungin and flucytosine. Our results indicate that combination of caspofungin with amphotericin B or a triazole, with or without 5-flucytosine has great potential against neurotropic dematiaceous fungi.IMPORTANCEThis research uses a modified version of the checkerboard assay to standardize the in vitro testing of double and triple combinations of antifungal agents against neurotropic dematiaceous fungi. Antifungal combination therapy is associated with improved outcomes in cerebral phaeohyphomycosis. In this study, we demonstrate that posaconazole is the single most active antifungal drug against this group of fungi. The double combination of amphotericin B with caspofungin or a trizole, and the triple combinations of caspofungin and flucytosine with amphotericin B or posaconazole might hold promise in the treatment of cerebral phaeohyphomycosis. Our findings will guide in developing optimal therapeutic strategies for these refractory infections.

Mycodegradation of low-density polyethylene by Cladosporium sphaerospermum, isolated from platisphere.

Plastic accumulation is a severe threat to the environment due to its resistivity to thermal, mechanical and biological processes. In recent years, microbial degradation of plastic waste disposal is of interest because of its eco-friendly nature. In this study, a total of 33 fungi were isolated from the plastisphere and out of which 28 fungal species showed halo zone of clearance in agarized LDPE media. The fungus showing highest zone of clearance was further used to evaluate its degradation potential. Based on morphological and molecular technique, the fungus was identified as Cladosporium sphaerospermum. The biodegradation of LDPE by C. sphaerospermum was evaluated by various methods. The exposure of LDPE with C. sphaerospermum resulted in weight loss (15.23%) in seven days, higher reduction rate (0.0224/day) and lower half-life (30.93 days). FTIR analysis showed changes in functional group and increased carbonyl index in LDPE treated with C. sphaerospermum. SEMimages evidenced the formation of pits, surface aberrations and grooves on the LDPE film treated with the fungus whereas the untreated control LDPE film showed no change. AFM analysis confirmed the surface changes and roughness in fungus treated LDPE film. This might be due to the extracellular lignolytic enzymes secreted by C. sphaerospermum grown on LDPE. The degradation of polyethylene by Short chain alkanes such as dodecane, hexasiloxane and silane were identified in the extract of fungus incubated with LDPE film through GC-MS analysis which might be due to the degradation of LDPE film by C. sphaerospermum. This was the first report on the LDPE degradation by C. sphaerospermum in very short duration which enables green scavenging of plastic wastes.

Characterization and removal mechanism of fluoroquinolone-bioremediation by fungus Cladosporium cladosporioides 11 isolated from aquacultural sediments.

Antibiotics have been widely detected in aquatic environments, and fungal biotransformation receives considerable attention for antibiotic bioremediation. Here, a fungus designated Cladosporium cladosporioides 11 (CC11) with effective capacity to biotransform fluoroquinolones was isolated from aquaculture pond sediments. Enrofloxacin (ENR), ciprofloxacin (CIP) and ofloxacin (OFL) were considerably abated by CC11, and the antibacterial activities of the fluoroquinolones reduced significantly after CC11 treatment. Transcriptome analysis showed the removal of ENR, CIP and OFL by CC11 is a process of enzymatic degradation and biosorption which consists well with ligninolytic enzyme activities and sorption experiments under the same conditions. Additionally, CC11 significantly removed ENR in zebrafish culture water and reduced the residue of ENR in zebrafish. All these results evidenced the potential of CC11 as a novel environmentally friendly process for the removal of fluoroquinolones from aqueous systems and reduce fluoroquinolone residues in aquatic organisms.

Isolation of undescribed cladosporols and spirobisnaphthalenes from a plant pathogen Cladosporium cladosporioides F-10-2-A.

Two undescribed cladosporol derivatives, cladosporols J-K (1-2), and three previously unreported spirobisnaphthalenes, urnucratins D-F (3-5), as well as eleven known cladosporols (6-16), were characterized from Cladosporium cladosporioides (Cladosporiaceae), a common plant pathogen isolated from the skin of Chinese toad. Cladosporols J-K (1-2) with a single double bond have been rarely reported, while urnucratins D-F (3-5) featured an unusual benzoquinone bisnaphthospiroether skeleton, contributing to an expanding category of undiscovered natural products. Their structures and absolute configurations were determined using extensive spectroscopic methods, including NMR, HRESIMS analyses, X-ray single crystal diffraction, as well as through experimental ECD analyses. Biological assays revealed that compounds 1 and 2 exhibited inhibitory activity against A549 cells, with IC50 values of 30.11 ± 3.29 and 34.32 ± 2.66 µM, respectively.

Identification and characterization of a fungal cutinase-like enzyme CpCut1 from Cladosporium sp. P7 for polyurethane degradation.

Plastic degradation by biological systems emerges as a prospective avenue for addressing the pressing global concern of plastic waste accumulation. The intricate chemical compositions and diverse structural facets inherent to polyurethanes (PU) substantially increase the complexity associated with PU waste management. Despite the extensive research endeavors spanning over decades, most known enzymes exhibit a propensity for hydrolyzing waterborne PU dispersion (i.e., the commercial Impranil DLN-SD), with only a limited capacity for the degradation of bulky PU materials. Here, we report a novel cutinase (CpCut1) derived from Cladosporium sp. P7, which demonstrates remarkable efficiency in the degrading of various polyester-PU materials. After 12-h incubation at 55°C, CpCut1 was capable of degrading 40.5% and 20.6% of thermoplastic PU film and post-consumer foam, respectively, while achieving complete depolymerization of Impranil DLN-SD. Further analysis of the degradation intermediates suggested that the activity of CpCut1 primarily targeted the ester bonds within the PU soft segments. The versatile performance of CpCut1 against a spectrum of polyester-PU materials positions it as a promising candidate for the bio-recycling of waste plastics.IMPORTANCEPolyurethane (PU) has a complex chemical composition that frequently incorporates a variety of additives, which poses significant obstacles to biodegradability and recyclability. Recent advances have unveiled microbial degradation and enzymatic depolymerization as promising waste PU disposal strategies. In this study, we identified a gene encoding a cutinase from the PU-degrading fungus Cladosporium sp. P7, which allowed the expression, purification, and characterization of the recombinant enzyme CpCut1. Furthermore, this study identified the products derived from the CpCut1 catalyzed PU degradation and proposed its underlying mechanism. These findings highlight the potential of this newly discovered fungal cutinase as a remarkably efficient tool in the degradation of PU materials.

Diversity and enzymatic, biosurfactant and phytotoxic activities of culturable Ascomycota fungi present in marine sediments obtained near the South Shetland Islands, maritime Antarctica.

We studied the culturable fungal community recovered from deep marine sediments in the maritime Antarctic, and assessed their capabilities to produce exoenzymes, emulsifiers and metabolites with phytotoxic activity. Sixty-eight Ascomycota fungal isolates were recovered and identified. The most abundant taxon recovered was the yeast Meyerozyma guilliermondii, followed by the filamentous fungi Penicillium chrysogenum, P. cf. palitans, Pseudeurotium cf. bakeri, Thelebolus balaustiformis, Antarctomyces psychrotrophicus and Cladosporium sp. Diversity indices displayed low values overall, with the highest values obtained at shallow depth, decreasing to the deepest location sampled. Only M. guilliermondii and P. cf. palitans were detected in the sediments at all depths sampled, and were the most abundant taxa at all sample sites. The most abundant enzymes detected were proteases, followed by invertases, cellulases, lipases, carrageenases, agarases, pectinases and esterases. Four isolates showed good biosurfactant activity, particularly the endemic species A. psychrotrophicus. Twenty-four isolates of P. cf. palitans displayed strong phytotoxic activities against the models Lactuca sativa and Allium schoenoprasum. The cultivable fungi recovered demonstrated good biosynthetic activity in the production of hydrolytic exoenzymes, biosurfactant molecules and metabolites with phytotoxic activity, reinforcing the importance of documenting the taxonomic, ecological and biotechnological properties of fungi present in deep oceanic sediments of the Southern Ocean.

Bio-oxidation of progesterone by Penicillium oxalicum CBMAI 1185 and evaluation of the cytotoxic activity.

We report the biotransformation of progesterone 1 by whole cells of Brazilian marine-derived fungi. A preliminary screening with 12 fungi revealed that the strains Penicillium oxalicum CBMAI 1996, Mucor racemous CBMAI 847, Cladosporium sp. CBMAI 1237, Penicillium oxalicum CBMAI 1185 and Aspergillus sydowii CBMAI 935 were efficient in the biotransformation of progesterone 1 in the first days of the reaction, with conversion values ranging from 75 % to 99 %. The fungus P. oxalicum CBMAI 1185 was employed in the reactions in quintuplicate to purify and characterize the main biotransformation products of progesterone 1. The compounds testololactone 1a, 12ß-hydroxyandrostenedione 1b and 1ß-hydroxyandrostenedione 1c were isolated and characterized by NMR, MS, [α]D and MP. In addition, the chromatographic yield of compound 1a was determined by HPLC-PDA in the screening experiments. In this study, we show a biotransformation pathway of progesterone 1, suggesting the presence of several enzymes such as Baeyer-Villiger monooxygenases, dehydrogenases and cytochrome P450 monooxygenases in the fungus P. oxalicum CBMAI 1185. In summary, the results obtained in this study contribute to the synthetic area and have environmental importance, since the marine-derived fungi can be employed in the biodegradation of steroids present in wastewater and the environment. The cytotoxic results demonstrate that the biodegradation products were inactive against the cell lines, in contrast to progesterone.

Investigating the inoculum dynamics of Cladosporium on the surface of raspberry fruits and in the air.

Raspberry production is under threat from the emerging fungal pathogenic genus Cladosporium. We used amplicon-sequencing, coupled with qPCR, to investigate how fruit age, fruit location within a polytunnel, polytunnel location and sampling date affected the fruit epiphytic microbiome. Fruit age was the most important factor impacting the fungal microbiome, followed by sampling date and polytunnel location. In contrast, polytunnel location and fruit age were important factors impacting the bacterial microbiome composition, followed by the sampling date. The within-tunnel location had a small significant effect on the fungal microbiome and no effect on the bacterial microbiome. As fruit ripened, fungal diversity increased and the bacterial diversity decreased. Cladosporium was the most abundant fungus of the fruit epiphytic microbiome, accounting for nearly 44% of all fungal sequences. Rotorod air samplers were used to study how the concentration of airborne Cladosporium inoculum (quantified by qPCR) varied between location (inside and outside the polytunnel) and time (daytime vs. nighttime). Quantified Cladosporium DNA was significantly higher during the day than the night and inside the polytunnel than the outside. This study demonstrated the dynamic nature of epiphytic raspberry fruit microbiomes and airborne Cladosporium inoculum within polytunnels, which will impact disease risks on raspberry fruit.

Unveiling biological activities of biosynthesized starch/silver-selenium nanocomposite using Cladosporium cladosporioides CBS 174.62.

BACKGROUND AND OBJECTIVES: Microbial cells capability to tolerate the effect of various antimicrobial classes represent a major worldwide health concern. The flexible and multi-components nanocomposites have enhanced physicochemical characters with several improved properties. Thus, different biological activities of biosynthesized starch/silver-selenium nanocomposite (St/Ag-Se NC) were assessed. METHODOLOGY: The St/Ag-Se NC was biosynthesized using Cladosporium cladosporioides CBS 174.62 (C. cladosporioides) strain. The shape and average particle size were investigated using scanning electron microscope (SEM) and high-resolution transmission electron microscope (HR-TEM), respectively. On the other hand, the St/Ag-Se NC effect on two cancer cell lines and red blood cells (RBCs) was evaluated and its hydrogen peroxide (H2O2) scavenging effect was assessed. Moreover, its effects on various microbial species in both planktonic and biofilm growth forms were examined. RESULTS: The St/Ag-Se NC was successfully biosynthesized with oval and spherical shape and a mean particle diameter of 67.87 nm as confirmed by the HR-TEM analysis. St/Ag-Se NC showed promising anticancer activity toward human colorectal carcinoma (HCT-116) and human breast cancer (MCF-7) cell lines where IC50 were 21.37 and 19.98 µg/ml, respectively. Similarly, little effect on RBCs was observed with low nanocomposite concentration. As well, the highest nanocomposite H2O2 scavenging activity (42.84%) was recorded at a concentration of 2 mg/ml. Additionally, Staphylococcus epidermidis (S. epidermidis) ATCC 12,228 and Candida albicans (C. albicans) ATCC 10,231 were the highly affected bacterial and fungal strains with minimum inhibitory concentrations (MICs) of 18.75 and 50 µg/ml, respectively. Moreover, the noticeable effect of St/Ag-Se NC on microbial biofilm was concentration dependent. A high biofilm suppression percentage, 87.5% and 68.05%, were recorded with S. epidermidis and Staphylococcus aureus (S. aureus) when exposed to 1 mg/ml and 0.5 mg/ml, respectively. CONCLUSION: The biosynthesized St/Ag-Se NC showed excellent antioxidant activity, haemocompatibility, and anti-proliferative effect at low concentrations. Also, it exhibited promising antimicrobial and antibiofilm activities.

Biotransformation of Patchouli Alcohol by Cladosporium cladosporioides and the Anti-Influenza Virus Activities of Biotransformation Products.

The biotransformation of patchouli alcohol by Cladosporium cladosporioides afforded 31 products, including 21 new ones (1-3, 5, 6, 8-14, and 17-25). Their structures were determined by extensive spectroscopic data analysis (1H and 13C NMR, HSQC, HMBC, 1H-1H COSY, ROESY, and HRESIMS), and the absolute configuration of compounds 1, 2, 8, 9, and 17 was determined by single-crystal X-ray diffraction using Cu Kα radiation. Structurally, compounds 21-24 were patchoulol-type norsesquiterpenoids without Me-12. Among them, a Δ3(4) double bond existed in compounds 21 and 22; a three-membered ring was formed between C-4, C-5, and C-6 in compound 23; an epoxy moiety appeared between C-3 and C-4 in compound 24. Furthermore, the biotransformation products 9, 10, 12, and 25 showed potent anti-influenza virus activity with EC50 values of 2.11, 7.94, 20.87, and 3.45 µM, respectively.