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.

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.

Photoinactivation by UVA radiation and visible light of Candida auris compared to other fungi.

In addition to the rising number of patients affected by viruses and bacteria, the number of fungal infections has also been rising over the years. Due to the increase in resistance to various antimycotics, investigations into further disinfection options are important. In this study, two yeasts (Candida auris and Saccharomyces cerevisiae) and a mold (Cladosporium cladosporioides) were irradiated at 365, 400, and 450 nm individually. The resulting log 1 reduction doses were determined and compared with other studies. Furthermore, fluorescence measurements of C. auris were performed to detect possible involved photosensitizers. A roughly exponential photoinactivation was observed for all three fungi and all irradiation wavelengths with higher D90 doses for longer wavelengths. The determined log 1 reduction doses of C. auris and S. cerevisiae converged with increasing wavelength. However, S. cerevisiae was more photosensitive than C. auris for all irradiation wavelengths and is therefore not a suitable C. auris surrogate for photoinactivation experiments. For the mold C. cladosporioides, much higher D90 doses were determined than for both yeasts. Concerning potential photosensitizers, flavins and various porphyrins were detected by fluorescence measurements. By excitation at 365 nm, another, so far unreported fluorophore and potential photosensitizer was also observed. Based on its fluorescence spectrum, we assume it to be thiamine.Graphic abstract.

From Child to Old Man: A Slowly Evolving Case of Chromoblastomycosis Caused by Cladosporium cladosporioides.

Chromoblastomycosis is a chronic granulomatous mycosis of the skin and subcutaneous tissue caused by traumatic inoculation with dematiaceous fungi. This disease primarily affects agricultural workers, who are mostly men. We present a case of chromoblastomycosis in a 63-year-old male farmer patient with dermatosis over 50 years of evolution, with warty, erythematous, and scaly plaques that predominate on the left hemithorax. Direct examination with potassium hydroxide (KOH) revealed numerous fumagoid cells. Amplification and sequencing of the internal transcribed spacer (ITS) and translation elongation factor 1-alpha (TEF-1a) gene revealed that chromoblastomycosis was caused by Cladosporium cladosporioides. The chromoblastomycosis was treated with itraconazole and fluconazole without any improvement, and amphotericin B was administered with partial improvement.

High energy density storage, antifungal activity and enhanced bioimaging by green self-doped heteroatom carbon dots.

Self-heteroatom-doped N-carbon dots (N-CDs) with a 2.35 eV energy gap and a 65.5% fluorescence quantum yield were created using a one-step, efficient, inexpensive, and environmentally friendly microwave irradiation method. FE-SEM, EDX, FT-IR, XRD, UV-VIS spectroscopy, FL spectroscopy, and CV electrochemical analysis were used to characterise the produced heteroatom-doped N-CDs. The graphitic carbon dot surface is doped with heteroatom functional groups such (S, P, K, Mg, Zn) = 1%, in addition to the additional passivating agent (N), according to the EDX surface morphology and the spontaneous heteroatom doping was caused by the heterogeneous chemical composition of pumpkin seeds. These spontaneous heteroatom-doped N-CDs possess quasispherical amorphous graphitic structure with an average size of less than 10 nm and the interplaner distance of 0.334 nm. Calculations utilising cyclic voltammetry showed that the heteroatom-doped N-CDs placed on nickel electrodes had a high specific capacitance value of 1044 F/g at a scan rate of 10 mV/s in 3 M of KOH electrolyte solution. Furthermore, it demonstrated a high energy and power density of 28.50 Wh/kg and 3350 W/kg, respectively. The higher value of specific capacitance and energy density were attributed to the fact that the Ni/CDs electrode material possesses both EDLC and PC properties due to the sufficient surface area and the multiple active sites of the prepared N-CDs. Furthermore, the heteroatom N-CDs revealed the antifungal action and bioimaging of the "Cladosporium cladosporioides" mould, which is mostly accountable for economic losses in agricultural products. The functional groups of nitrogen, sulphur, phosphorus, and zinc on the surface of the CDs have strong antibacterial and antifungal properties as well as fluorescence enhanced bioimaging.

Cadavers found outdoor in whom fungal growth was observed on the body surface: Consideration of the role of mycology in forensic medicine.

In forensic case work, fungal growth is occasionally observed on decomposed cadavers, however, the isolation of these organisms is not always carried out routinely. The usefulness of investigating fungi isolated from corpses in forensic settings, has been reported, such as providing trace evidence, estimating the time since death, and ascertaining the time of deposition. A 32-year-old male missing for 4 months, was found in a car in a forest far from his home and had died from blood loss, having been stabbed with a kitchen knife in the left side of his chest. Whitish or white greenish colonies ranging 5 cm to 8 cm in size were observed over his whole body. Penicillium commune and Cladosporium cladosporioides were isolated from the colonies. A 49-year-old male missing for one month, and was found dead at 500 m elevation in the mountains. An autopsy was conducted, but the cause of death could not be determined. Dark greenish or whitish colonies were found on his face and neck, and Epicoccum nigrum, Mucor sp.. Cladosporium sp., and Pestalotiopsis sp. were isolated. Penicillium sp. and Cladosporium sp. are major indoor fungi, and Epicoccum sp. and Pestalotiopsis sp. are plant pathogens found in outdoor environments. There was no discrepancy between the police statement and findings of the corpse such as place discovered, types of isolated fungi, and estimated time since death. Identifying fungi on cadavers may aid in forensic casework, and further studies are needed to develop for the use of fungi as a forensic tool.

First Report of Cladosporium cladosporioides Causing Leaf Spot of Aralia cordata var. continentalis in Korea.

Aralia cordata var. continentalis (Kitag), commonly known as Japanese spikenard, is an upright herbaceous perennial medicinal plant effective in relieving pain. It is also consumed as a leafy vegetable. Leaf spots and blight symptoms on A. cordata resulting in defoliation were observed in July 2021 from a research field with a disease incidence of nearly 40-50% from 80 plants in Yeongju, Korea. Brown spots with chlorotic halos first appear on the upper leaf surface (Fig. 1A). In the later stage, spots enlarge and coalesce; resulting in the leaves to dry-off (Fig. 1B). To isolate the causal agent, small pieces of diseased leaves displaying the lesion were surface-sterilized by 70% ethanol for 30 s and rinsed twice with sterile distilled water (SDW). Later, the tissues were crushed in a sterile 2.0-ml Eppendorf tube with a rubber pestle in SDW. The suspension was serially diluted and spread on potato dextrose agar (PDA) medium, incubated at 25°C for 3 days. A total of 3 isolates were obtained from the infected leaves. Pure cultures were obtained by the monosporic culture technique (Choi et al. 1999). After 2 to 3 days of incubation with a 12-h photoperiod, the fungus initially produced gray mold colonies in olive color, and the edges of the mold appeared white with a velvety texture after 20 days (Fig. 1C). Microscopic observations revealed small, single-celled, rounded, and pointed conidia that measured 6.67 ± 0.23 µm × 4.18 ± 0.12 µm (length × width) (n=40 spores) (Fig. 1D). On the basis of its morphology, the causal organism was identified as Cladosporium cladosporioides (Torres et al. 2017). For molecular identification, pure colonies of three single-spore isolates were used for DNA extraction. A fragment of the ITS, ACT, and TEF1-α were amplified using the primers ITS1/ITS4 (Zarrin et al. 2016), ACT-512F/ACT-783R, and EF1-728F/EF1-986R, respectively, by PCR (Carbone et al. 1999). The DNA sequences from all three isolates (GYUN-10727, GYUN-10776, and GYUN-10777) were identical. The resulting ITS (ON005144), ACT (ON014518), and TEF1-α (OQ286396) sequences from the representative isolate GYUN-10727 were 99 to 100% identical to the C. cladosporioides (ITS: KX664404, MF077224; ACT: HM148509; TEF1-α: HM148268, HM148266). The phylogenetic dendrogram was constructed from the comparative analysis of ITS, ACT, and TEF1-α gene sequences, showing the relationship between Cladosporium cladosporioides and related Cladosporium species (Fig. 2). The isolate GYUN-10727 has been deposited in Korean Agricultural Culture Collection (KACC 410009), and used as a representative strain in this study. For the pathogenicity test, healthy fresh leaves (3 leaves per plant) of 3-months-old A. cordata plants in pots were spray inoculated with conidial suspensions (1 × 104 conidia/mL) of GYUN-10727, which was obtained from a 7-day-old PDA culture. Leaves sprayed with SDW were considered as control. After 15 days of incubation at 25°C ± 5°C under greenhouse conditions, necrotic lesions were observed on the inoculated A. cordata leaves, while control leaves did not develop any disease symptoms. The experiment was performed twice with three replicates (pots) per treatment. The pathogen was re-isolated from the symptomatic A. cordata leaves, but not from control plants, to fulfill Koch's postulates. The re-isolated pathogen was identified by PCR. Cladosporium cladosporioides has been reported to cause diseases in sweet pepper (Krasnow et al. 2022) and garden peas (Gubler et al. 1999). To our knowledge, this is the first report of C. cladosporioides causing leaf spots of A. cordata in Korea. The identification of this pathogen will help develop strategies to efficiently control the disease in A. cordata.

Four dodecanoic acid derivatives from the cold-seep-derived fungus Cladosporium cladosporioides 8-1.

Two new compounds, cladospolides I (1) and J (2), together with two new naturally occurring ones, methyl 11-hydroxy-4-oxododecanoate (3) and 11-hydroxy-4-oxododecanoic acid (4), were isolated from the culture extract of the cold-seep sediment-derived fungus Cladosporium cladosporioides 8-1. Their structures and configurations were established by analysis of 1D/2D NMR, MS, ECD, and specific optical rotation data. Compound 3 was possibly formed by methyl esterification of 4 during the purification process due to the utilization of methanol. All compounds were evaluated for inhibition of four marine phytoplankton species and five marine-derived bacteria.

Integration of agricultural residues as biomass source to saccharification bioprocess and for the production of cellulases from filamentous fungi.

The production of second-generation bioethanol has several challenges, among them finding cheap and efficient enzymes for a sustainable process. In this work, we analyzed two native fungi, Cladosporium cladosporioides and Penicillium funiculosum, as a source of cellulolytic enzyme production, and corn stover, wheat bran, chickpeas, and bean straw as a carbon source in two fermentation systems: submerged and solid fermentation. Corn stover was selected for cellulase production in both fermentation systems, because we found the highest enzymatic activities when carboxymethyl cellulase activity (CMCase) was assessed using CMC as substrate. C. cladosporioides showed the highest CMCase activity (1.6 U/mL), while P. funiculosum had the highest filter paper activity (Fpase) (0.39 U/mL). The ß-glucosidase activities produced by both fungi were similar in submerged fermentation using corn stover as substrate. Through in-gel zymography, three polypeptides with cellulolytic activities were identified in each fungus: with molecular weights of ~ 38, 45 and 70 kDa in C. cladosporioides and ~ 21, 63 and 100 kDa in P. funiculosum. The best results for saccharification (10.11 g/L of reducing sugars) of diluted acid pretreated corn stover were obtained after 36 h of the hydrolytic process at pH 5 and 50 °C using the enzyme extract of P. funiculosum. This is the first report of cellulase identification in C. cladosporioides and the saccharification of corn stover using enzymes of this fungus. Enzymatic extracts of C. cladosporioides and P. funiculosum obtained from low-cost lignocellulosic biomass have great potential for use in the production of second-generation bioethanol.

Cladosporium cladosporioides causes leaf blight on garden pea in Telangana, India.

Garden pea (Pisum sativum L.) is an important vegetable and pulse crop grown worldwide. Leaf blight symptoms with chlorotic lesions were observed on the pea genotypes, 2006-2008A and Arkel in field conditions at Asian Vegetable Research and Development Centre (AVRDC), Hyderabad (17°23' N; 78°29' E) and greenhouse conditions at University of Hyderabad, Hyderabad, India (17° 27' N; 78° 19' E) since 2010. The blight symptoms showed black lesions coated by a fine velvety layer of olive-grey conidia. The disease incidence on field and greenhouse pea plants ranged from 3-6%. The Cladosporium isolates, Gp01 and Gp02 were isolated from pea genotypes, Arkel and 2006-2008A in greenhouse and field conditions respectively. One fungus was predominantly isolated from infected leaf tissue plated onto potato dextrose agar (PDA), and it was identified as Cladosporium sp. based on colony morphology and conidial appearance. Single-spored cultures grown at 26±2 °C on PDA showed olive green mycelia on the obverse-side and dark green on reverse-side (Fig. 1 a,b). The conidia were limoniform with dimensions ranging from 5.91-7.33 x 3.19-4.58 µm (Table 1) and were produced on solitary conidiophores containing 1-2 septa, with no constriction whereas the aerial hyphae were septate and unbranched or rarely branched (Fig.1 c-d). To verify the pathogenicity of the fungus, detached leaf assay was carried on 30-day old Arkel leaves by point inoculation with 20 µL (106 spores/ml) spore suspension using a needle-less syringe to infiltrate the spores into the abaxial surface and the inoculated leaves were incubated at 26±2 °C for 3 days in petri plates containing moistened autoclaved absorbent cotton. Pea leaves inoculated with water served as controls. The experiment was repeated with a minimum of three times with five leaves per replication and three replications per experiment for each of the isolates. The appressorial germ tube formation was observed by scanning electron microscopy (SEM) at 12 hours post inoculation (hpi) (Fig.1 e-f). Thirty-day old pea plants were spray-inoculated with a spore suspension of 1-2 ml per plant with a concentration of 106 spores/ ml while the control plants were mock-inoculated (sprayed) with water. 5 plants per replicate per isolate with three replications per experiment and a minimum of three experiments were carried. The inoculated plants were covered with polythene zip-lock bags for two days post inoculation (dpi) and maintained in greenhouse at 26±2 °C with natural photoperiod of 14 hours. Wilting and necrosis symptoms accompanied by chlorotic lesions were observed on pea plants at 7 days post inoculation (dpi) (Fig. 2). Complying with Koch's postulates, the pathogen, which was re-isolated from the first appearance at 3 dpi from infected pea showed similar morphological characteristics as those used for inoculation. Sequencing analysis of the internal transcribed spacer (ITS) region using ITS1 and ITS4 primers (White et al. 1990), D1-D2 region sequencing (Kwiatkowski et al. 2012), Translation elongation factor 1-alpha (TEF) and Actin (ACT) gene sequencing (Nam et al. 2015) of Gp01 and Gp02 showed significant similarity with C. cladosporioides (Table S1). Further, concatenated phylogenetic trees constructed by maximum likelihood method using all the four (ITS, D1-D2, TEF and ACTIN) gene sequences of these isolates along with other reported Cladosporium species (Fig. S1a) and specific isolates of C. cladosporiodes which are reported as "sensu stricto" (Bensch et al. 2010) collected from different substrata (Fig. S1b) confirmed them as C. cladosporioides. Gp01 and Gp02 were also confirmed by Microbial Type Culture Collection (MTCC, India) as C. cladosporioides with Acc. No. MTCC 9994 and MTCC 9995. C. cladosporioides has been previously reported on Dalbergia sp. and Eucalyptus sp in India and on garden pea in other countries (Bensch et al. 2010). However, owing to the impact of this pathogen on growth and productivity of pea crop, further in-depth study needs to be carried. To our understanding this is the first report of Cladosporium cladosporioides infecting garden pea in India.

Cladamide: a new ceramide from the endophytic fungus Cladosporium cladosporioides.

A new ceramide, named cladamide (1), in addition to cinnamic acid (2), para-coumaric acid (3), stigmasterol-3-O-ß-D-glucoside (4), and uracil (5), was isolated from the white beans culture of Cladosporium cladosporioides, a marine-derived endohpytic fungus isolated from the leaves of the mangrove, Avicennia marina (Forssk.) Vierh. Structure elucidation of compound 1 was established on the basis of extensive 1D and 2D NMR spectroscopic techniques in combination with HR-ESI-MS. The ability of the isolated compounds to inhibit acetylcholine esterase was evaluated. Compound 3 showed the highest acetylcholine esterase inhibitory activity (IC50 = 0.057 ± 0.003 µM), followed by compound 4 (IC50 = 0.068 ± 0.003 µM) and compound 1 (IC50 = 0.099 ± 0.005 µM) compared to donepezil, the positive control, (IC50 = 0.044 ± 0.002 µM). Compounds 2 and 5 showed lower activity (IC50 = 0.182 ± 0.009 and 0.236 ± 0.012 µM, respectively). The results were further validated by molecular docking study.

Porcine embryo development and inactivation of microorganisms after ultraviolet-C irradiation at 228 nm.

It is important to prevent contamination inside the incubator as a method of preventing microbial infections during the embryo culture. In the present study, we examined the effects of ultraviolet-C (UV-C) irradiation, used for microorganism inactivation, on embryo development and the growth of bacteria, including Escherichia coli and Staphylococcus aureus, and the fungus Cladosporium cladosporioides. In the embryo irradiation experiment, we examined the effects of the plastic lid of the culture dish, irradiation distances (10, 20, and 25 cm), and different irradiation wavelengths (228 and 260 nm) during embryo culture for 7 days on the development and quality of porcine in vitro-fertilized embryos. None of the embryos cultured in dishes without plastic lids developed into blastocysts after irradiation with 228 nm UV-C. When porcine embryos were cultured in a culture dish with lids, the 228 nm UV-C irradiation decreased blastocyst formation rates of the embryos but not their quality, irrespective of the UV-C irradiation distance. Moreover, irradiation with 260 nm UV-C, even with plastic lids, had more detrimental effects on embryo development than irradiation with 228 nm UV-C. Investigation of the inactivating effects of UV-C irradiation at 228 nm and 260 nm on the growth of the bacteria and fungus showed that 260 nm UV-C reduced the viability to a greater extent than 228 nm UV-C. Moreover, the disinfection efficacy for the bacteria increased when the irradiation duration increased and the distance decreased. In conclusion, porcine embryos can develop into blastocysts without loss of quality even after continuous long-duration irradiation (7 days) with 228 nm UV-C, which can inactivate the growth of bacteria and the tested fungus; however, the development rate of the embryo is reduced.

Isolation, Identification and Hyperparasitism of a Novel Cladosporium cladosporioides Isolate Hyperparasitic to Puccinia striiformis f. sp. tritici, the Wheat Stripe Rust Pathogen.

Wheat rust outbreaks have caused significantly economic losses all over the world. Puccinia striiformis f. sp. tritici (Pst) is an obligate biotrophic fungus causing stripe rust on wheat. Application of fungicides may cause environmental problems. The effects of hyperparasites on plant pathogens are the basis for biological control of plant pathogenic fungi and parasites of Pst have great value in biological agents development. Here, we report the isolation and characterization of isolate of Cladosporium cladosporioides from Pst based on morphological characterization and analysis of molecular markers. The hyperparasitic isolate was isolated from taupe-colored uredinia of Pst. Upon artificial inoculation, the hyperparasitic isolate was able to reduce the production and germination rate of Pst urediospores, and Pst uredinia changed color from yellow to taupe. Scanning electron microscopy demonstrated that the strain could efficiently colonize Pst urediospores. Therefore, the isolate has the potential to be developed into a biological control agent for managing wheat stripe rust.

Cladosporioles A and B, Two New Indole Derivatives from the Deep-Sea-Derived Fungus Cladosporium cladosporioides 170056.

Two new (cladosporioles A and B, 1 and 2) and fourteen known (3-16) compounds were isolated from the deep-sea-derived fungus Cladosporium cladosporioides 170056. The relative structures of the new compounds were elucidated mainly by detailed analysis of their NMR and HR-ESI-MS spectroscopic data. Their absolute configurations were determined by comparison of the experimental and calculated electronic circular dichroism (ECD) spectra. All isolates were tested for antimicrobial activity against Vibrio parahaemolyticus. Compound 15 exhibited weak effect with the MIC value of 156.25 µg/mL.

Possible Antagonism between Cladosporium cladosporioides and Microcystis aeruginosa in a Freshwater Lake during Bloom Seasons.

To ensure drinking-water safety, it is necessary to understand the factors that regulate harmful cyanobacterial blooms (HCBs) and the toxins they produce. One controlling factor might be any relationship between fungi and the cyanobacteria. To test this possibility, water samples were obtained from Harsha Lake in southwestern Ohio during the 2015, 2016, and 2017 bloom seasons, i.e., late May through September. In each water sample, the concentration of the filamentous fungus Cladosporium cladosporioides was determined by quantitative PCR (qPCR) assay, and Microcystis aeruginosa microcystin-gene transcript copy number (McyG TCN) was quantified by reverse-transcriptase qPCR (RT-qPCR) analyses. The results showed that during each bloom season, the C. cladosporioides concentration and McyG TCN appeared to be interrelated. Therefore, C. cladosporioides concentrations were statistically evaluated via regression on McyG TCN in the water samples for lag times of 1 to 7 days. The regression equation developed to model the relationship demonstrated that a change in the C. cladosporioides concentration resulted in an opposing change in McyG TCN over an approximately 7-day interval. Although the interaction between C. cladosporioides and McyG TCN was observed in each bloom season, the magnitude of each component varied yearly. To better understand this possible interaction, outdoor Cladosporium spore-count data for the Harsha Lake region were obtained for late May through September of each year from the South West Ohio Air Quality Agency. The average Cladosporium spore count in the outdoor air samples was significantly greater in 2016 than in either 2015 or 2017, and the M. aeruginosa McyG TCN was significantly lower in Harsha Lake water samples in 2016 compared to 2015 or 2017. These results suggest that there might be a "balanced antagonism" between C. cladosporioides and M. aeruginosa during the bloom season.

Innate and mild Th17 cutaneous immune responses elicited by subcutaneous infection of immunocompetent mice with Cladosporium cladosporioides.

Cladosporium cladosporioides is a dematiaceous hyphomycete that is pathogenic in the superficial and deep tissues of both immunodeficient and immunocompetent humans and animals. Our aim was to evaluate the antifungal immune responses elicited by C. cladosporioides in immunocompetent mice. Hence, we subcutaneously injected suspensions of C. cladosporioides spores into immunocompetent mice to investigate the anti-fungal immune responses in the skin. We collected skin tissue samples for histopathological examination, immunofluorescence staining, and quantitative real-time polymerase chain reaction analysis. We observed subcutaneous abscesses in mice after subcutaneous injection of C. cladosporioides. A large number of inflammatory cells, including dendritic cells, macrophages, and neutrophils, infiltrated the focal abscess, with comparatively few infiltrating inflammatory cells in the epidermal and dermal layers of the skin. We detected the expression of CD54 in the abscesses and the skin. Gene expression of the pattern recognition receptors Dectin-1 and TLR-2 was higher in infected mice than in controls. Gene expression of the cytokines IL-6, IL-1ß, and IL-17A also increased after infection, suggesting that the Th17 signaling pathway may be involved in the anti-fungal response. Although the pathogenicity of C. cladosporioides in healthy mice was weak after subcutaneous infection, resulting in few serious pathological phenomena, it appears that innate and Th17 immune responses play important roles in the cutaneous host response to C. cladosporioides. These findings lay a foundation for further study of the pathogenic mechanism and treatment of C. cladosporioides infection.

First Report of Fruit Rot of Grapes (Vitis vinifera) Caused by Cladosporium cladosporioides in Xinjiang, China.

Xinjiang Province accounts for nearly 20% of the total grape (Vitis Vinifera L.), proles orientalis) (wine, table and raisin combined) production, being the largest production area in China. Fruit rot is the most common disease that impacts grape quality and yield. A new disease where the ripe grape berry surfaces were coated with brownish mildew was observed, and the disease mainly occurred on whole clusters or most of the berries in the cluster. In September 2019 and 2020, 125 diseased grape clusters were collected from 10 locations in northern Xinjiang where the disease incidence was 15.3% - 27.4% ((diseased clusters/ total clusters)*100). To identify the pathogen, symptomatic grape berries were disinfected with 1% NaClO for 2 min, followed by 70% ethanol for 30 s, and rinsed thrice in sterile distilled water. Three pieces of ~0.5 cm2 diseased grape skin with partial exocarp were placed on potato dextrose agar (PDA) amended with streptomycin sulfate and kanamycin (50 µg/mL each). The PDA plates were then incubated at 25℃ under light condition with the luminous intensity 3500 Lux for 7 days. Fungal colonies emerging from the plated tissue were subcultured and single-spored three times to obtain pure cultures. From 20 strains with similar colony phenotype and grey olive hue, flocculent, felt-like surface, six (Cc-Vivi-3, 7, 9, 11, 13 and 19) isolates were chosen for further characterization after 7 days of incubation. Conidia were either single or grew in chains, with around 4 conidia per chain. Conidia were ovoid, nearly spindle or globose with slightly smooth or irregular reticulate surface. Conidiophores were solitary, smooth, septate, erect or geniculate. These characteristics were consistent with the descriptions for Cladosporium cladosporioides. To confirm this identification, PCR was performed on the genomic DNA of the selected strains using primers for internal transcribed spacer (ITS) region ITS1/ITS4, actin (ACT) and translation elongation factor (TEF) (Schubert et al., 2007; Braun et al., 2003). Amplified ITS sequences provided a 100% match to C. cladosporioides (AY213641) in NCBI. Homology of ACT sequences to C. cladosporioides (HM148527 and MH047330) was 99.57% and 100%, respectively; and the homology of TEF sequences with C. cladosporioides (HM148258, HM148289, HM148260 and HM148266) was 97.56% ~ 100%. To further confirm the evolutionary relationship of strains from grapes with Cladosporium spp., phylogenetic analyses based on ITS, ACT and TEF conjoint sequences from the six experimental isolates, five C. cladosporioides strains, eight proximal Cladosporium species were analyzed. The phylogenetic tree showed that the six isolates from grapes clustered with C. cladosporioides strains, but not other proximal Cladosporium species. This confirmed that all six isolates evaluated were C. cladosporioides. Pathogenicity tests with one C. cladosporioides isolate (Cc-Vivi-3; accession No. ITS: MW556429, ACT: MW567144, TEF: MW567143) were carried out as follows: ripe and healthy grape clusters from cultivars Xinyu and Munag when total soluble solids were 20-21°Bx and 19-20 °Bx, respectively, were detached from the vines. Five berries of three clusters of each cultivar were punctured with a sterile syringe, then inoculated with a 20 µL conidial suspension (107 conidia/mL). And uninoculated, punctured berries in clusters treated with sterilized water served as controls. The experiment was repeated three times. Symptoms were recorded 15 days after incubation at 80% relative humidity and 25℃ with a 14 h light/10 h dark cycle. The olive green or blackish green mildew layer was produced on all inoculated berries. No symptoms were observed on the uninoculated berries. Koch's postulates were fulfilled by reisolating C. cladosporioides from all symptomatic tissues and identifying them by PCR targeting the ACT gene. This is the first description of C. cladosporioides causing grape fruit rot in Xinjiang, China. In recent years, worldwide reports of Cladosporium spp. damaging crops are increasing (Briceño et al., 2008; Walker et al., 2016; Meneses et al., 2018; Robles-Yerena et al., 2019; Ding et al., 2019; Yang et al., 2021). However, relatively few methods of management including some fungicides and biocontrol agents are available in different crops (Wang et al., 2018; Addrah et al., 2019). In view of the important role of Xinjiang in China agricultural production, that should arouse strong attention.

New Cladosporium Species from Normal and Galled Flowers of Lamiaceae.

A series of isolates of Cladosporium spp. were recovered in the course of a cooperative study on galls formed by midges of the genus Asphondylia (Diptera, Cecidomyidae) on several species of Lamiaceae. The finding of these fungi in both normal and galled flowers was taken as an indication that they do not have a definite relationship with the midges. Moreover, identification based on DNA sequencing showed that these isolates are taxonomically heterogeneous and belong to several species which are classified in two different species complexes. Two new species, Cladosporium polonicum and Cladosporium neapolitanum, were characterized within the Cladosporium cladosporioides species complex based on strains from Poland and Italy, respectively. Evidence concerning the possible existence of additional taxa within the collective species C. cladosporioides and C. pseudocladosporioides is discussed.

Complete mitochondrial genome of Cladosporium cladosporioides YFCC 8621 isolated from a salt mine in Yunnan, southwestern China.

Cladosporium cladosporioides is one of the most isolated species in the genus Cladosporium and has a wide medical and industrial usage. Here, we first report the complete mitogenome of C. cladosporioides based on the Illumina sequencing data. The circular mitogenome is 36,768 bp in length, containing 14 protein-coding genes (PCGs), 2 ribosomal RNA (rns and rnl) genes, 2 ORFs (ORF214 and ORF240), and a set of 28 transfer RNA (tRNA) genes. The overall base composition is 35.7% A, 34.0% T(U), 15.3% C, 15.0% G, with a GC content of 30.3%. Phylogenetic analysis shows that C. cladosporioides is clustered in the order Capnodiales and is closely related to the congeneric species Cladosporium zixishanense of Cladosporiaceae.

First Report of Cladosporium Blossom Blight Caused by Cladosporium cladosporioides on Calliandra haematocephala in China.

Calliandra haematocephala Hassk., commonly called red powder puff, is widely cultivated as an ornamental plant in Taiwan, Hainan, Guangdong and Fujian in China (CAS, 1988). The flowers are dark crimson with conspicuous stamens, which give them the appearance of powder-puffs. Blossom blight on C. haematocephala was first observed in early January 2019 on plants grown on the university campus as well as in parks in Fuzhou city, with nearly 80% of flowers on individual plants infected. At various locations in the city, disease incidence was 100%. Symptoms appeared as grayish green fungal growth on the stamens with the entire flower eventually turning black and covered with masses of fungal spores. Fifteen single spore isolates obtained from nine necrotic stamen samples were purified and cultured on Potato dextrose agar (PDA) plates at 24 ℃.The resultant fungal colonies were olivaceous-green to olivaceous-brown and had a velvet-like appearance. Conidiophores were smooth-walled, solitary, non-nodulose, and measuring 40 to 340 × 3 to 4 µm (n=50). Ramoconidia were cylindrical-oblong or slightly curved with 0 to 3 septa, and measuring 10 to 25 × 3 to 4 µm (n=50). Conidia were smooth-walled and prolifically produced in long chains. Terminal conidia were aseptate, subglobose, ovoid to limoniform, measuring 3 to 6 × 2 to 2.5 µm (n=50). Intercalary conidia were elliptical to limoniform or subcylindrical, aseptate, measuring 5 to 12 × 2.5 to 3 µm (n=50). On the basis of its morphology, the causal organism was identified as Cladosporium cladosporioides (Bensch et al. 2010). For molecular identification, pure cultures of five single-spore isolates were used for DNA extraction. A fragment in the ITS regions of the fungal rDNA, the ACT and the TEF1-α, was amplified using the primers ITS1/ITS4, ACT-512F/ACT-783R, and EF1728 F/EF1-986R. The DNA sequences obtained from all five isolates were identical. The resulting ITS (MK720012) and ACT (MN013164), and TEFl-α (MK752020) sequences from a representative isolate MRCIM19 were 98-100% identical to the C. cladosporioides accessions (ITS: MH863979, MG228421; ACT: HM148509, JF499878, HM148532; TEFl-α: JF499872). To test pathogenicity, a spore suspension (1×105 conidia/mL) was prepared from a seven- day- old culture of isolate MRCIM19 and 10 mL of the suspension was sprayed onto six flowers on each of three C. haematocephala plants. Sterile distilled water was sprayed onto three flowers of two plants as control. The inoculated flowers were covered with plastic bags which were removed two days post inoculation. Disease symptoms were recorded on each flower at 10 days post inoculation. Based on the morpho-molecular characters, the re-isolated fungus from the inoculated flowers was C. cladosporioides. This fungus was previously reported to cause blossom blight in strawberry in the USA and Korea (Gubler et al. 1999; Nam et al. 2015). Although it has been reported from many plants (Zhang 2003) in China, this is the first report of C. cladosporioides on C. haematocephala worldwide. References Bensch, K. et al. 2010. Stud Mycol. 67:1-94. Chinese Academy of Sciences (CAS), 1988. Flora Republicae Popularis Sinicae Editorial Committee, Beijing Sci. Press., 39: 38. Gubler, W. D. et al. 1999. Plant Dis. 83:400. Nam, M. H. et al. 2015. Microbiol. 43: 354-359. Zhang Z., Ed. 2003. Flora fungorum sinicorum, Vol. 14. Cladosporium, Fusicladium, Pyricularia. Beijing Science Press. 297.