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Alternative splicing (AS) is a key element of eukaryotic gene expression that increases transcript and proteome diversity in cells, thereby altering their responses to external stimuli and stresses. While AS has been intensively researched in plants and animals, its frequency, conservation, and putative impact on virulence, are relatively still understudied in plant pathogenic fungi. Here, we profiled the AS events occurring in genes of Cladosporium fulvum isolates Race 5 and Race 4, during nearly a complete compatible infection cycle on their tomato host. Our studies revealed extensive heterogeneity in the transcript isoforms assembled from different isolates, infections, and infection timepoints, as over 80% of the transcript isoforms were singletons that were detected in only a single sample. Despite that, nearly 40% of the protein-coding genes in each isolate were predicted to be recurrently AS across the disparate infection timepoints, infections, and the two isolates. Of these, 37.5% were common to both isolates and 59% resulted in multiple protein isoforms, thereby putatively increasing proteome diversity in the pathogen by 31% during infections. An enrichment analysis showed that AS mostly affected genes likely to be involved in the transport of nutrients, regulation of gene expression, and monooxygenase activity, suggesting a role for AS in finetuning adaptation of C. fulvum on its tomato host during infections. Tracing the location of the AS genes on the fungal chromosomes showed that they were mostly located in repeat-rich regions of the core chromosomes, indicating a causal connection between gene location on the genome and propensity to AS. Finally, multiple cases of differential isoform usage in AS genes of C. fulvum were identified, suggesting that modulation of AS at different infection stages may be another way by which pathogens refine infections on their hosts., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Zaccaron et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The combined pollution of antibiotics and heavy metals in aqueous environments increases the risk of aquatic ecosystem disruption and the complication of pollutant management. Here, a fungus Cladosporium cladosporioides 11 (CC11) isolated from aquaculture pond sediments possessed effective capacities to biotransform the combined pollution of enrofloxacin (ENR) and copper ion (Cu). ENR and Cu were considerably abated by CC11, and the presence of Cu (30 mg/L) promoted the biotransformation efficiency of ENR. The biotranformation of ENR in ENR-Cu co-contamination was associated to ligninolytic enzyme action. The expression of ligninolytic enzymes was enhanced by ENR and ENR-Cu combined pollution. And the increased activities of ligninolytic enzymes confirmed the significant role of enzymatic transformation. Cu played an important role in increasing the expression and activities of ligninolytic enzymes. The expressions of many genes associated with transporters, phosphate assimilation, oxidative phosphorylation, hyperosmotic stress and pectin metabolism were significantly up-regulated when facing Cu-stress, indicating their important roles in determining Cu removal and enhancing Cu-resistance. Additionally, CC11 significantly biotransformed other antibiotic and heavy metal combined pollution. All these results contributed to the applications of CC11 in aqueous environments., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: Sponge-associated microorganisms are promising resources for the production of bioactive compounds with cytotoxic potential. The main goal of our study is to isolate the fungal endophytes from the Red Sea sponge Hyrtios sp. followed by investigating their cytotoxicity against number of cell lines., Results: The fungal strain UR3 was isolated from the Red Sea sponge using Sabouraud dextrose agar media. It was identified based on partial 18 S rRNA gene and ITS sequence analyses as Cladosporium sp. UR3. The in vitro cytotoxic potential of the ethyl acetate extract of the fungal isolate was evaluated using MTT assay against three cancer cell lines: CACO2, MCF7, and HEPG2. Metabolomics profiling of the obtained ethyl acetate extract using LC-HR-ESI-MS, along with molecular docking and pharmacological network studies for the dereplicated compounds were performed to explore its chemical profile and the possible cytotoxic mechanism of the sponge-associated fungi., Conclusion: These results highlighted the role of sponge-associated fungi as a fruitful resource for the discovery of cytotoxic metabolites., (© 2024. The Author(s).)

Enzymes that degrade β-glucan play important roles in various industries, including those related to brewing, animal feed, and health care. Csph16A, an endo-β-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 β-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-β-1,3(4)-glucanase. Additionally, it sheds light on a group of endo-β-1,3(4)-glucanases from Antarctic rock-inhabiting black fungi harboring a Lam16A catalytic domain and a novel CTD of unknown function., (© 2024 The Author(s). Biotechnology Journal published by Wiley‐VCH GmbH.)

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., Competing Interests: The authors declare no conflict of interest.

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., (© 2024 The Authors. Environmental Microbiology published by John Wiley & Sons Ltd.)

Background: Fungal plant pathogens have dynamic genomes that allow them to rapidly adapt to adverse conditions and overcome host resistance. One way by which this dynamic genome plasticity is expressed is through effector gene loss, which enables plant pathogens to overcome recognition by cognate resistance genes in the host. However, the exact nature of these loses remains elusive in many fungi. This includes the tomato pathogen Cladosporium fulvum, which is the first fungal plant pathogen from which avirulence (Avr) genes were ever cloned and in which loss of Avr genes is often reported as a means of overcoming recognition by cognate tomato Cf resistance genes. A recent near-complete reference genome assembly of C. fulvum isolate Race 5 revealed a compartmentalized genome architecture and the presence of an accessory chromosome, thereby creating a basis for studying genome plasticity in fungal plant pathogens and its impact on avirulence genes., Results: Here, we obtained near-complete genome assemblies of four additional C. fulvum isolates. The genome assemblies had similar sizes (66.96 to 67.78 Mb), number of predicted genes (14,895 to 14,981), and estimated completeness (98.8 to 98.9%). Comparative analysis that included the genome of isolate Race 5 revealed high levels of synteny and colinearity, which extended to the density and distribution of repetitive elements and of repeat-induced point (RIP) mutations across homologous chromosomes. Nonetheless, structural variations, likely mediated by transposable elements and effecting the deletion of the avirulence genes Avr4E, Avr5, and Avr9, were also identified. The isolates further shared a core set of 13 chromosomes, but two accessory chromosomes were identified as well. Accessory chromosomes were significantly smaller in size, and one carried pseudogenized copies of two effector genes. Whole-genome alignments further revealed genomic islands of near-zero nucleotide diversity interspersed with islands of high nucleotide diversity that co-localized with repeat-rich regions. These regions were likely generated by RIP, which generally asymmetrically affected the genome of C. fulvum., Conclusions: Our results reveal new evolutionary aspects of the C. fulvum genome and provide new insights on the importance of genomic structural variations in overcoming host resistance in fungal plant pathogens., (© 2024. The Author(s).)

This study reports the first case of lung and fungus ball lesions caused by Cladosporium subcinereum A48 . A rural diabetic woman infected by C. subcinereum A48 , presented with fever and hemoptysis. CT scans, bronchoalveolar lavage, culture and molecular methods were used to evaluate and confirm the disease. Our patient had not taken insulin during the last few months of her life which probably caused acidosis and a decrease in the patient's immunity level, resulting in penetration and formation of a fungal in the lung.

Cladosporium spp., as one of the largest and most heterogeneous genera of hyphomycetes, are widely distributed worldwide. This genus is usually adaptable to a wide variety of extreme environments. However, only 11 genomes of Cladosporium genus have been publicly released. From 2017, we found for the first time that Cladosporium velox could cause cotton boll disease and lead to stiffness and cracking boll in Xinjiang, China. Herein, we provide a high-quality reference genome for the C. velox strain C4 isolated from cotton boll in Xinjiang, China. The genome size and encoding gene number of the C. velox strain C4 and C. cucumerinum strain CCNX2, which was recently released and caused the cucumber scab, showed minor differences. This resource will contribute to future research that aims to elucidate the genetic basis of C. velox pathogenicity and could expand our knowledge of Cladosporium spp. genomic characteristics that will be valuable for the development of Cladosporium disease control measures., Competing Interests: The author(s) declare no conflict of interest.

Caves are underground and natural environments mainly found in rocky terrain. Caves have a very specific microclimate, which benefits the occurrence of specific fungi. In recent studies, researchers have observed that caves harbour a great diversity of fungi. However, studies on fungal diversity in Brazilian caves are still incipient. In September 2019, airborne spore and soil samples were collected from the Monte Cristo cave, in the Southern Espinhaço Range, Diamantina, Minas Gerais state, Brazil. Two Cladosporium single-spore isolates, among other genera, were obtained from these samples. This study aimed to characterise these two fungal isolates based on their DNA sequence data and morphology. Phylogenetic analyses of the rDNA-ITS, ACT and TEF1-α loci revealed that the isolates belonged to the Cladosporium cladosporioides species complex. Both isolates did not cluster with any known species and were formally described and named herein as C. diamantinense and C. speluncae. This study presents taxonomic novelties and contributes to the knowledge about the fungal diversity in Brazilian caves., (© 2023. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.)

The myrtle rust (MR), caused by Austropuccinia psidii, is a worldwide threat to the cultivated and wild Myrtaceae. Originally from the neotropics, it has spread to North America, Africa, and Asia and has reached geographically isolated areas in the Pacific and Australasia. It is attacking native species in those new ranges and is still spreading and causing great concern for the damage caused to endemic Myrtaceae, and to the environment. Classical biological control is regarded as the most sustainable management option for mitigating such biological invasions. However, there are no examples of introductions of host-specific co-evolved natural enemies of plant pathogens, from their native range, as a management strategy for plant pathogens. In order to explore this neglected approach, a survey of potential fungal natural enemies of A. psidii was initiated recently in the state of Minas Gerais (Brazil). Several purported mycoparasites have been collected from A. Psidii pustules formed on myrtaceous hosts. This included some isolates of dematiaceous fungi recognized as having a Cladosporium-like morphology. Here we present the results of the investigation aimed at elucidating their identity through a polyphasic taxonomic approach. Besides morphological and cultural features, molecular analyses using sequences of translation elongation factor 1-α (EF1) and actin (ACT) were performed. The combination of data generated is presented herein and placed all Cladosporium-like isolates in six species of Cladosporium, namely, Cladosporium angulosum, C. anthropophilum, C. bambusicola, C. benschii, C. guizhouense, and C. macadamiae. None of these have ever been recorded in association with A. psidii. Now, with the identification of these isolates at hand, an evaluation of biocontrol potential of these fungi will be initiated. In contrast with the ready finding of fungicolous (possibly mycoparasitic) fungi on MR in this study, no evidence of those was recorded from Australasia until now., (© 2023. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.)

Despite the fact that Cladosporium sp. are ubiquitous fungi, their viromes have been little studied. By analysing a collection of Cladosporium fungi, two new partitiviruses named Cladosporium cladosporioides partitivirus 1 (CcPV1) and Cladosporium cladosporioides partitivirus 2 (CcPV2) co-infecting a strain of Cladosporium cladosporioides were identified. Their complete genome consists of two monocistronic dsRNA segments (RNA1 and RNA2) with a high percentage of pairwise identity on 5' and 3' end. The RNA directed RNA polymerase (RdRp) of both viruses and the capsid protein (CP) of CcPV1 display the classic characteristics required for their assignment to the Gammapartitivirus genus. In contrast, CcPV2 RNA2 encodes for a 41 KDa CP that is unusually smaller when aligned to CPs of other viruses classified in this genus. The structural role of this protein is confirmed by electrophoresis on acrylamide gel of purified viral particles. Despite the low percentage of identity between the capsid proteins of CcPV1 and CcPV2, their three-dimensional structures predicted by AlphaFold2 show strong similarities and confirm functional proximity. Fifteen similar viral sequences of unknown function were annotated using the CcPV2 CP sequence. The phylogeny of the CP was highly consistent with the phylogeny of their corresponding RdRp, supporting the organization of Gammapartitiviruses into three distinct clades despite stretching the current demarcation criteria. It is proposed that a new subgenus be created within the genus Gammapartitivirus for this new group.

Fungal sinusitis is a widespread infection that affects both healthy and immunocompromised individuals. Reports of sinus fungal infections have increased due to recent advances in diagnosis. Furthermore, susceptible and immune-compromised patients play an important role in increasing the number of reported cases. Infections with lesser-known fungi have been reported infrequently around the world. This paper describes a Cladosporium tenuissimum infection caused by chronic fungal sinusitis in a woman who had traveled to several countries. We used morphological and molecular methods to confirm the infection. The infection is most likely caused by the use of sulfasalazine, which is related to the patient's rheumatism. Sulfasalazine inhibits neutrophilic chemoattractant lipid synthesis in neutrophils, which play a key role in antifungal immunity. The patient is also undergoing root canal therapy and has several upper jaw implants, which may have contributed to the development of sinusitis., (© 2023. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.)

Fulvia fulva and Dothistroma septosporum are closely related apoplastic pathogens with similar lifestyles but different hosts: F. fulva is a pathogen of tomato, whilst D. septosporum is a pathogen of pine trees. In 2012, the first genome sequences of these pathogens were published, with F. fulva and D. septosporum having highly fragmented and near-complete assemblies, respectively. Since then, significant advances have been made in unravelling their genome architectures. For instance, the genome of F. fulva has now been assembled into 14 chromosomes, 13 of which have synteny with the 14 chromosomes of D. septosporum, suggesting these pathogens are even more closely related than originally thought. Considerable advances have also been made in the identification and functional characterization of virulence factors (e.g., effector proteins and secondary metabolites) from these pathogens, thereby providing new insights into how they promote host colonization or activate plant defence responses. For example, it has now been established that effector proteins from both F. fulva and D. septosporum interact with cell-surface immune receptors and co-receptors to activate the plant immune system. Progress has also been made in understanding how F. fulva and D. septosporum have evolved with their host plants, whilst intensive research into pandemics of Dothistroma needle blight in the Northern Hemisphere has shed light on the origins, migration, and genetic diversity of the global D. septosporum population. In this review, we specifically summarize advances made in our understanding of the F. fulva-tomato and D. septosporum-pine pathosystems over the last 10 years., (© 2023 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

Timothy is a forage mainly grown in Min County, Gansu Province, China. In 2021, a leaf spot disease outbreak on timothy grass occurred in Min County, adversely affecting its growth and productivity. Therefore, this study investigated the leaf spot disease incidence in Min County, morphologically and molecularly characterized the disease-causing pathogen, and assessed its effects on the growth, photosynthesis, and biomass of timothy seedlings re-inoculated with the isolated pathogen. In the field, the disease incidence on plants and leaves was 100 and 85%, respectively. Morphologically, young lesions were ellipsoidal-fusiform with dark purple margins and an off-white center, while the mature lesions were eye-shaped spots with a light brown center and dark purple edges. Molecular characterization identified the pathogen as Cladosporium phlei causing Cladosporium eyespot disease. The net photosynthetic rate, transpiration rate, fresh shoot weight, and dry shoot weight of timothy seedlings 14 days after inoculation with the pathogen were decreased by 29.77, 56, 45.45, and 46.42%, respectively, implying that Cladosporium eyespot disease is an important timothy grass disease in Min County. Therefore, developing an integrated control strategy is urgent to lessen the economic loss., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

High tunnels extend the growing season of high value crops, including tomatoes, but the environmental conditions within high tunnels favor the spread of the tomato leaf mold pathogen, Passalora fulva (syn. Cladosporium fulvum ). Tomato leaf mold results in defoliation, and if severe, losses in yield. Despite substantial research, little is known regarding the genetic structure and diversity of populations of P . fulva associated with high tunnel tomato production in the United States. From 2016 to 2019, a total of 50 P . fulva isolates were collected from tomato leaf samples in high tunnels in the Northeast and Minnesota. Other Cladosporium species were also isolated from the leaf surfaces. Koch's postulates were conducted to confirm that P . fulva was the cause of the disease symptoms observed. Race determination experiments revealed that the isolates belonged to either race 0 (six isolates) or race 2 (44 isolates). Polymorphisms were identified within four previously characterized effector genes: Avr2 , Avr4 , Avr4e , and Avr9 . The largest number of polymorphisms were observed for Avr2 . Both mating type genes, MAT1 - 1 - 1 and MAT1 - 2 - 1 , were present in the isolate collection. For further insights into the pathogen diversity, the 50 isolates were genotyped at 7,514 single-nucleotide polymorphism loci using genotyping-by-sequencing. Differentiation by region but not by year was observed. Within the collection of 50 isolates, there were 18 distinct genotypes. Information regarding P . fulva population diversity will enable better management recommendations for growers, as high tunnel production of tomatoes expands.

Closely related species are expected to have similar functional traits due to shared ancestry and phylogenetic inertia. However, few tests of this hypothesis are available for plant-associated fungal symbionts. Fungal leaf endophytes occur in all land plants and can protect their host plant from disease by a variety of mechanisms, including by parasitizing pathogens (e.g., mycoparasitism). Here, we tested whether phylogenetic relatedness among species of Cladosporium, a widespread genus that includes mycoparasitic species, predicts the effect of this endophyte on the severity of leaf rust disease. First, we used congruence among different marker sequences (i.e., genealogical concordance phylogenetic species recognition criterion) to delimit species of Cladosporium. Next, in a controlled experiment, we quantified both mycoparasitism and disease modification for the selected Cladosporium species. We identified 17 species of Cladosporium; all the species reduced rust disease severity in our experiment. Cladosporium phylogeny was a significant predictor of mycoparasitism. However, we did not observe a phylogenetic effect on disease severity overall, indicating that other mechanism/s operating independently of shared ancestry also contributed to endophyte effects on disease severity. Indeed, a second experiment showed that Cladosporium endophyte exudates (no live organism) from divergent species groups equally reduced disease severity. Our results reveal that multiple mechanisms contribute to the protective effects of an endophyte against a plant pathogen, but not all traits underlying these mechanisms are phylogenetically conserved., (© 2022 John Wiley & Sons Ltd.)

Tomato leaf mold disease caused by Cladosporium fulvum ( C. fulvum ) is one of the most common diseases affecting greenhouse tomato production. Cf proteins can recognize corresponding AVR proteins produced by C. fulvum , and Cf genes are associated with leaf mold resistance. Given that there are many physiological races of C. fulvum and that these races rapidly mutate, resistance to common Cf genes (such as Cf-2 , Cf-4 , Cf-5 , and Cf-9 ) has decreased. In the field, Ont7813 plants (carrying the Cf-13 gene) show effective resistance to C. fulvum ; thus, these plants could be used as new, disease-resistant materials. To explore the mechanism of the Cf-13 -mediated resistance response, transcriptome sequencing was performed on three replicates each of Ont7813 ( Cf-13 ) and Moneymaker (MM; carrying the Cf-0 gene) at 0, 9, and 15 days after inoculation (dai) for a total of 18 samples. In total, 943 genes were differentially expressed, specifically in the Ont7813 response process as compared to the Moneymaker response process. Gene ontology (GO) classification of these 943 differentially expressed genes (DEGs) showed that GO terms, including "hydrogen peroxide metabolic process (GO_Process)", "secondary active transmembrane transporter activity (GO_Function)", and "mismatch repair complex (GO_Component)", which were the same as 11 other GO terms, were significantly enriched. An analysis of the Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that many key regulatory genes of the Cf-13 -mediated resistance response processes were involved in the "plant hormone signal transduction" pathway, the "plant-pathogen interaction" pathway, and the "MAPK signaling pathway-plant" pathway. Moreover, during C. fulvum infection, jasmonic acid (JA) and salicylic acid (SA) contents significantly increased in Ont7813 at the early stage. These results lay a vital foundation for further understanding the molecular mechanism of the Cf-13 gene in response to C. fulvum infection.

Cladosporium fulvum is a fungal pathogen that causes leaf mould of tomato. The reference genome of this pathogen was released in 2012 but its high repetitive DNA content prevented a contiguous assembly and further prohibited the analysis of its genome architecture. In this study, we combined third generation sequencing technology with the Hi-C chromatin conformation capture technique, to produce a high-quality and near complete genome assembly and gene annotation of a Race 5 isolate of C. fulvum . The resulting genome assembly contained 67.17 Mb organized into 14 chromosomes (Chr1-to-Chr14), all of which were assembled telomere-to-telomere. The smallest of the chromosomes, Chr14, is only 460 kb in size and contains 25 genes that all encode hypothetical proteins. Notably, PCR assays revealed that Chr14 was absent in 19 out of 24 isolates of a world-wide collection of C. fulvum , indicating that Chr14 is dispensable. Thus, C. fulvum is currently the second species of Capnodiales shown to harbour dispensable chromosomes. The genome of C. fulvum Race 5 is 49.7 % repetitive and contains 14 690 predicted genes with an estimated completeness of 98.9%, currently one of the highest among the Capnodiales. Genome structure analysis revealed a compartmentalized architecture composed of gene-dense and repeat-poor regions interspersed with gene-sparse and repeat-rich regions. Nearly 39.2 % of the C. fulvum Race 5 genome is affected by Repeat-Induced Point (RIP) mutations and evidence of RIP leakage toward non-repetitive regions was observed in all chromosomes, indicating the RIP plays an important role in the evolution of this pathogen. Finally, 345 genes encoding candidate effectors were identified in C. fulvum Race 5, with a significant enrichment of their location in gene-sparse regions, in accordance with the 'two-speed genome' model of evolution. Overall, the new reference genome of C. fulvum presents several notable features and is a valuable resource for studies in plant pathogens.

Vicia faba (faba bean) is one of the most significant leguminous crops. The faba bean is specialized by maximum nutritional value, in energy and protein content, which leads it to be suitable for food and feed production. Diseases caused with fungi are amongst the biotic factors responsible for decreasing in faba bean yields. In this work, Cladosporium isolates were recorded in cultivated faba bean leaves and pods collected from markets in Qena, Upper Egypt; morphological features and molecular characterization based on actin gene were performed. The ability of the pathogens to cause disease in faba bean seedlings and the biocontrol method to avoid the pathogenic effect of Cladosporium were determined. Results showed that Cladosporium is the main genera isolated from faba beans, and the morphological criteria showed presence of three species complex groups of Cladosporium (C. cladosporioides, C. herbarum and C. sphaerospermum) and the confirmation with molecular characterization revealed the existence of four species in the three groups. All the 26 tested strains of Cladosporium were able to cause leaf lesions on Vicia faba seedlings with different levels. Chaetomium globosum is a biocontrol agent could inhibit the growth of the majority strains of Cladosporium., (© 2021. The Author(s).)

This study was conducted at the Agriculture College University of Karbala, Iraq to isolate and morphologically and molecularly diagnose thirteen Cladosporium isolates collected from tomato plant residues present in desert regions of Najaf and Karbala provinces, Iraq. We diagnosed the obtained isolates by PCR amplification using the ITS1 and ITS4 universal primer pair followed by sequencing. PCR amplification and analysis of nucleotide sequences using the BLAST program showed that all isolated fungi belong to Cladosporium sphaerospermum. Analysis of the nucleotide sequences of the identified C. sphaerospermum isolates 2, 6, 9, and 10 showed a genetic similarity reached 99%, 98%, 99%, and 99%, respectively, with those previously registered at the National Center for Biotechnology Information (NCBl). By comparing the nucleotide sequences of the identified C. sphaerospermum isolates with the sequences belong to the same fungi and available at NCBI, it was revealed that the identified C. sphaerospermum isolates 2, 6, 9, and 10 have a genetic variation with those previously recorded at the National Center for Biotechnology Information (NCBl); therefore, the identified sequences of C. sphaerospermum isolates have been registered in GenBank database (NCBI) under the accession numbers MN896004, MN896107, MN896963, and MN896971, respectively.

Effectors are small, secreted proteins that promote pathogen virulence. Although key to microbial infections, unlocking the intrinsic function of effectors remains a challenge. We have previously shown that members of the fungal Avr4 effector family use a carbohydrate-binding module of family 14 (CBM14) to bind chitin in fungal cell walls and protect them from host chitinases during infection. Here, we show that gene duplication in the Avr4 family produced an Avr4-2 paralog with a previously unknown effector function. Specifically, we functionally characterize Pf Avr4-2, a paralog of Pf Avr4 in the tomato pathogen Pseudocercospora fuligena , and show that although it contains a CBM14 domain, it does not bind chitin or protect fungi against chitinases. Instead, Pf Avr4-2 interacts with highly de-esterified pectin in the plant's middle lamellae or primary cell walls and interferes with Ca 2+ -mediated cross-linking at cell-cell junction zones, thus loosening the plant cell wall structure and synergizing the activity of pathogen secreted endo-polygalacturonases., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

Background : Endogenous fungal endophthalmitis is a sight-threatening condition with potentially devastating outcome. Hematogenous spread of the infective seedings is the route of infection. Infected individuals have usually a compromised immune status. The clinical picture of mycotic endogenous endophthalmitis is commonly seen as chorioretinitis. Candida is the most common fungus. Cladosporium causing endogenous endophthalmitis is a rare occurrence, with only a few cases published. Methods : The report includes study and management of a diabetic patient with endogenous cladosporium endophthalmitis mimicking toxoplasma retinochoroiditis. Results : Diagnosis was confirmed as Cladosporium Cladosporioides in vitreous and aqueous aspirate by polymerase chain reaction-based DNA sequencing. Patient was successfully managed with intravitreal and systemic voriconazole. Conclusion : Cladosporium can cause endogenous endophthalmitis and mimic toxoplasma retinochoroiditis. Vitreous biopsy can help in diagnosis in the absence of positive blood culture. Intravitreal voriconazole along with systemic voriconazole shows a good response.

Culture-independent DNA sequencing of fungal internal transcribed spacer 2 (ITS2) region was compared to a culture-dependent morphological identification technique to characterize house dust-borne fungal communities. The abundant genera were Aspergillus, Wallemia, Cladosporium, and Penicillium. Statistically significant between-method correlations were observed for Wallemia and Cladosporium (Spearman's ρ = 0.75 and 0.72, respectively; p < 0.001). Penicillium tended to be detected with much higher (averaged 26-times) relative abundances by the culture-based method than by the DNA-based method, although statistically significant inter-method correlation was observed with Spearman's ρ = 0.61 (p = 0.002). Large DNA sequencing-based relative abundances observed for Alternaria and Aureobasidium were likely due to multicellularity of their spores with large number of per-spore ITS2 copies. The failure of the culture-based method in detectiing Toxicocladosporium, Verrucocladosporium, and Sterigmatomyces was likely due to their fastidiousness growth on our nutrient medium. Comparing between the two different techniques clarified the causes of biases in identifying environmental fungal communities, which should be amended and/or taken into consideration when the methods are used for future fungal ecological studies.

Rcr3 is a secreted protease of tomato that is targeted by fungal effector Avr2, a secreted protease inhibitor of the fungal pathogen Cladosporium fulvum. The Avr2-Rcr3 complex is recognized by receptor-like protein Cf-2, triggering hypersensitive cell death (HR) and disease resistance. Avr2 also targets Rcr3 paralog Pip1, which is not required for Avr2 recognition but contributes to basal resistance. Thus, Rcr3 acts as a guarded decoy in this interaction, trapping the fungus into a recognition event. Here we show that Rcr3 evolved > 50 million years ago (Mya), whereas Cf-2 evolved <6Mya by co-opting the pre-existing Rcr3 in the Solanum genus. Ancient Rcr3 homologs present in tomato, potato, eggplants, pepper, petunia and tobacco can be inhibited by Avr2 with the exception of tobacco Rcr3. Four variant residues in Rcr3 promote Avr2 inhibition, but the Rcr3 that co-evolved with Cf-2 lacks three of these residues, indicating that the Rcr3 co-receptor is suboptimal for Avr2 binding. Pepper Rcr3 triggers HR with Cf-2 and Avr2 when engineered for enhanced inhibition by Avr2. Nicotiana benthamiana (Nb) is a natural null mutant carrying Rcr3 and Pip1 alleles with deleterious frame-shift mutations. Resurrected NbRcr3 and NbPip1 alleles were active proteases and further NbRcr3 engineering facilitated Avr2 inhibition, uncoupled from HR signalling. The evolution of a receptor co-opting a conserved pathogen target contrasts with other indirect pathogen recognition mechanisms.

Background: Invasive fungal rhinosinusitis is associated with high morbidity and mortality. Rapid pathogen identification is mandatory, but fresh tissue is not always available. A polymerase chain reaction method was designed in order to detect fungi in formalin-fixed paraffin-embedded samples. This was applied to a retrospective series of tissue biopsies from Thai patients with invasive fungal rhinosinusitis., Methods: Tissue blocks from 64 cases yielded adequate DNA. Three sequential polymerase chain reaction were performed: ZP3 (housekeeping gene) and panfungal polymerase chain reactions, and a differentiating polymerase chain reaction based on the 5.8s ribosomal RNA and internal transcribed spacer 2 regions. The polymerase chain reaction products were then sequenced., Results: Polymerase chain reaction identified a fungal pathogen in 20 of 64 cases (31 per cent). Aspergillus species was the most common cause of invasive fungal rhinosinusitis (nine cases). Other causes included candida (n = 4), cladosporium (n = 4), mucor (n = 1), alternaria (n = 1) and dendryphiella (n = 1) species., Conclusion: Polymerase chain reaction can provide rapid identification of fungal pathogens in paraffin-embedded tissue, enabling prompt treatment of invasive fungal rhinosinusitis.

Plants trigger immune responses upon recognition of fungal cell wall chitin, followed by the release of various antimicrobials, including chitinase enzymes that hydrolyze chitin. In turn, many fungal pathogens secrete LysM effectors that prevent chitin recognition by the host through scavenging of chitin oligomers. We previously showed that intrachain LysM dimerization of the Cladosporium fulvum effector Ecp6 confers an ultrahigh-affinity binding groove that competitively sequesters chitin oligomers from host immune receptors. Additionally, particular LysM effectors are found to protect fungal hyphae against chitinase hydrolysis during host colonization. However, the molecular basis for the protection of fungal cell walls against hydrolysis remained unclear. Here, we determined a crystal structure of the single LysM domain-containing effector Mg1LysM of the wheat pathogen Zymoseptoria tritici and reveal that Mg1LysM is involved in the formation of two kinds of dimers; a chitin-dependent dimer as well as a chitin-independent homodimer. In this manner, Mg1LysM gains the capacity to form a supramolecular structure by chitin-induced oligomerization of chitin-independent Mg1LysM homodimers, a property that confers protection to fungal cell walls against host chitinases., Competing Interests: The authors declare no conflict of interest exists.

A total of 20 dried date samples, chosen as representative among those available on the Perugia (Umbria, Central Italy) market, were analyzed for the possible occurrence of fungal species and related contamination by fungal secondary metabolites. Twenty-six isolates, representative of the total mycobiota, were obtained and morphologically identified as belonging to the genera Aspergillus, Penicillium and Cladosporium. Inside each genus, molecular characterization (by partial sequencing of ITS region and/or β-tubulin and calmodulin regions for Aspergillus and Penicillium isolates or actin region for Cladosporium isolates) and in vitro mycotoxigenic profile characterization (by LC-MS/MS analysis) showed the presence of the following species: A. flavus, A. tubingensis, P. brevicompactum, P. chrysogenum, P. crustosum, P. glabrum, P. solitum, P. venetum, C. cladosporioides, C. limoniforme and C. halotolerans, with A. tubingensis as the prevalent species and P. crustosum, P. solitum, P. venetum and C. limoniforme first reported here on dates. Date packaging and format showed an effect on the incidence of isolated fungi, with the lowest incidence recovered from whole dates and in hermetic bag packaging. These findings can be useful both for dried dates producers and consumers, guiding them towards choices of packaging and format with a lower risk of mycotoxigenic species presence. However, no fungal metabolites were detected in the dried date samples analyzed, which were therefore regarded as safe for human consumption, underlining the absence of correspondence between fungal isolation and mycotoxin contaminations., Competing Interests: Declaration of competing interest The authors declare they have no competing financial interests or personal relationships that could have influenced the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The molecular interactions between tomato and Cladosporium fulvum have been an important model for molecular plant pathology. Complex genetic loci on tomato chromosomes 1 and 6 harbor genes for resistance to Cladosporium fulvum , encoding receptor like-proteins that perceive distinct Cladosporium fulvum effectors and trigger plant defenses. Here, we report classical mapping strategies for loci in tomato accessions that respond to Cladosporium fulvum effector Ecp5, which is very sequence-monomorphic. We screened 139 wild tomato accessions for an Ecp5-induced hypersensitive response, and in five accessions, the Ecp5-induced hypersensitive response segregated as a monogenic trait, mapping to distinct loci in the tomato genome. We identified at least three loci on chromosomes 1, 7 and 12 that harbor distinct Cf-Ecp5 genes in four different accessions. Our mapping showed that the Cf-Ecp5 in Solanum pimpinellifolium G1.1161 is located at the Milky Way locus. The Cf-Ecp5 in Solanum pimpinellifolium LA0722 was mapped to the bottom arm of chromosome 7, while the Cf-Ecp5 genes in Solanum lycopersicum Ontario 7522 and Solanum pimpinellifolium LA2852 were mapped to the same locus on the top arm of chromosome 12. Bi-parental crosses between accessions carrying distinct Cf-Ecp5 genes revealed putative genetically unlinked suppressors of the Ecp5-induced hypersensitive response. Our mapping also showed that Cf-11 is located on chromosome 11, close to the Cf-3 locus. The Ecp5-induced hypersensitive response is widely distributed within tomato species and is variable in strength. This novel example of convergent evolution could be used for choosing different functional Cf-Ecp5 genes according to individual plant breeding needs., (Copyright © 2020 Iakovidis et al.)

Glucose oxidases are widely used in various industrial processes, including bread baking. In this study, a novel glucose oxidase gene, CngoxA, from Cladosporium neopsychrotolerans SL16, was cloned and expressed in Pichia pastoris. Recombinant CnGOXA exhibited maximal activity at 20 °C and pH 7.0, and was stable at 30 °C and pH 6.0-9.0 for 1 h, with a half-life of 15 min at 40 °C. Compared with CnGOXA, the half-life of its mutant CnGOXA-M1 (Y169C-A211C), at 40 °C increased approximately 48-fold, and was stable at 30 °C and pH 3.0-12.0 for 1 h. The k cat and catalytic efficiency of CnGOXA-M1 were enhanced 0.7- and 1.6-fold, respectively. Both enzymes were cold-adapted and highly resistant to SDS. Furthermore, CnGOXA-M1 had a more significant effect on bread volume than that of GOX from Aspergillus niger. These favorable enzymatic properties of CnGOXA-M1 make it a potentially useful enzyme for many industrial applications., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Cassiae Semen (CS) has been widely used as roasted tea and traditional Chinese medicine for decades. However, CS is easily contaminated by fungi and mycotoxins during pre-harvest and post-harvest process, thus posing a potential threat to consumer health. In this study, we used the Illumina MiSeq PE300 platform and targeted the internal transcribed spacer 2 sequences to survey the occurrence of fungi in raw and roasted CS samples. Results showed the fungal contamination in all 12 test samples. Ascomycota was the prevailing fungus at the phylum level, with the relative abundance of 66.50%-99.42%. At the genus level, Aspergillus, Cladosporium, and Penicillium were the most dominant genera, accounting for 0.66%-85.51%, 0.20%-29.11%, and 0.11%-32.92% of the fungal reads, respectively. A total of 68 species were identified, among which six potential toxigenic fungi belonging to Aspergillus, Penicillium, Candida, and Schizophyllum genera were detected. Moreover, differences in fungal communities were observed in raw and roasted CS samples. In conclusion, amplicon sequencing is feasible for analyzing fungal communities in CS samples, which provides a new approach to investigate the fungal contamination in edible-medicinal herb, thereby ensuring food safety and drug efficacy., Competing Interests: Declaration of competing interest None., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Chromoblastomycosis is a neglected disease characterized by cutaneous, subcutaneous or disseminated lesions. It is considered an occupational infectious disease that affects mostly rural workers exposed to contaminated soil and vegetal matter. Lesions mostly arise after a traumatic inoculation of herpotrichiellaceous fungi from the Chaetothyriales order. However, the environmental niche of the agents of the disease remains obscure. Its association with insects has been predicted in a few studies. Therefore, the present work aimed to analyze if social insects, specifically ants, bees, and termites, provide a suitable habitat for the fungi concerned. The mineral oil flotation method was used to isolate the microorganisms. Nine isolates were recovered and phylogenetic analysis identified two strains as potential agents of chromoblastomycosis, i.e., Fonsecaea pedrosoi CMRP 3076, obtained from a termite nest (n = 1) and Rhinocladiella similis CMRP 3079 from an ant exoskeleton (n = 1). In addition, we also identified Fonsecaea brasiliensis CMRP 3445 from termites (n = 1), Exophiala xenobiotica CMRP 3077 from ant exoskeleton (n = 1), Cyphellophoraceae CMRP 3103 from bees (n = 1), Cladosporium sp. CMRP 3119 from bees (n = 1), Hawksworthiomyces sp. CMRP 3102 from termites (n = 1), and Cryptendoxyla sp. from termites (n = 2). The environmental isolate of F. pedrosoi CMRP 3076 was tested in two animal models, Tenebrio molitor and Wistar rat, for its pathogenic potential with fungal retention in T. molitor tissue. In the Wistar rat, the cells resembling muriform cells were observed 30 d after inoculation., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2020 British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

Background: Palmoplantar pustulosis (PPP) is a distinct, chronic skin disorder characterized by intraepidermal pustules on the palms and soles. It is hypothesized that microorganisms on the skin might induce the symptoms of PPP via inflammatory cell activation. However, the microbiota has not been studied in detail because of the assumption that the pustules in PPP are sterile., Aim: To elucidate the role of microorganisms in pathogenesis of PPP., Methods: PCR analysis was performed of microbial DNA fragments in the pustules of patients with PPP. The sequence of the D1/D2 LSU 26s rRNA gene and that of the 16S rRNA gene was used for fungal and bacterial DNA detection, respectively., Results: In total, 71 samples were carefully collected from the pustules of patients with PPP. Fungal DNA bands were detected in 68 samples, and fungi including Malassezia spp. were identified in 30 of 71 samples (42.3%). Malassezia restricta was the most frequently encountered fungus (14/71; 19.7%). However, bacterial DNA was not detected by the methods used. Furthermore, identical fungal DNA was not detected in the outer lid of the pustules, suggesting that the fungi detected within the pustule did not derive from contamination via the skin surface., Conclusions: In the present study, we demonstrated for the first time that certain pustules in patients with PPP contain fungal DNA fragments, especially those of Malassezia spp. Our findings provide new insights on the role of skin microbiota in the pathogenesis of PPP., (© 2019 British Association of Dermatologists.)

Cladosporium spp. is a cosmopolitan fungal genus. In the literature, it has been reported as a biological agent for controlling several plant diseases, but its mechanism of action has never been clarified. The present study aims to identify Cladosporium spp. based on the DNA phylogeny of nine isolates obtained from the phylloplane of rice and their potential antagonistic activity against the main fungal pathogens that affect rice crop. Nine isolates of Cladosporium spp. were identified based on DNA phylogeny, molecular and morphological characterization, and their antagonistic effects with the rice pathogens C. miyabeanus, M. oryzae, M. albescens and S. oryzae. Four isolates were selected to study lytic enzymes such as β-1,3-glucanase, chitinase and protease, and only one isolate was selected for a conidial germination and appressoria formation assay. The nine isolates were identified as C. cladosporioides, C. tenuissimum and C. subuliforme. Four isolates, identified as C. cladosporioides, inhibited the mycelial growth of rice pathogens such as C1H (68.59%) of S. oryzae, C5 G (74.32%) of C. miyabeanus, C11 G (75.97%) of M. oryzae and C24 G (77.39%) of M. albescens. C24 G showed a high activity of lytic enzymes, was tested against C. miyabeanus and M. oryzae, and inhibited conidial germination and appressorium formation by more than 59.36%. The characterization of C. cladosporioides suggested this species as a potential bioagent for the management of several rice diseases, especially rice blast. This is the first time that a potential biological agent from the genus Cladosporium identified at the species level was isolated from the rice phylloplane, and some of its mechanisms of action were demonstrated, such as increasing lytic enzyme activity against rice pathogens., (Copyright © 2019. Published by Elsevier GmbH.)

Five hybrid polyketides ( 1a , 1b , and 2 - 4 ) containing tetramic acid core including a new hybrid polyketide, cladosin L ( 1 ), were isolated from the marine fungus Cladosporium sphaerospermum SW67, which was isolated from the marine hydroid polyp of Hydractinia echinata . The hybrid polyketides were isolated as a pair of interconverting geometric isomers. The structure of 1 was determined based on 1D and 2D NMR spectroscopic and HR-ESIMS analyses. Its absolute configuration was established by quantum chemical electronic circular dichroism (ECD) calculations and modified Mosher's method. Tetramic acid-containing compounds are reported to be derived from a hybrid PKS-NRPS, which was also proved by analyzing our 13 C-labeling data. We investigated whether compounds 1 - 4 could prevent cell damage induced by cisplatin, a platinum-based anticancer drug, in LLC-PK1 cells. Co-treatment with 2 and 3 ameliorated the damage of LLC-PK1 cells induced by 25 μM of cisplatin. In particular, the effect of compound 2 at 100 μM (cell viability, 90.68 ± 0.81%) was similar to the recovered cell viability of 88.23 ± 0.25% with 500 μM N -acetylcysteine (NAC), a positive control.

This report presents the pathologic findings associated with disseminated infection due to Cladosporium halotolerans in a dog that was simultaneously infected with canine adenovirus-1 (CAdV-1) and canine parvovirus-2 (CPV-2). A 12-year-old, mixed breed dog, with a clinical history of neurological manifestations was submitted for routine autopsy due to poor prognosis. The principal pathologic findings were mycotic necrotizing nephritis, hepatitis, and splenitis with embolic dissemination to the brain resulting in mycotic necrotizing meningoencephalitis, ventriculitis, choroid plexitis, and obstructive hydrocephalus associated with intralesional and intravascular septate pigmented fungi. PCR and sequencing of the ITS region of fungi revealed that the intralesional fungal organisms had 82% nucleotide identity with members of the Cladosporium sphaerospermum complex of organisms. However, a PCR assay and sequencing of the beta tubulin gene confirmed that the organism identified in this dog had 100% nucleotide sequence identity with C. halotolerans. Using immunohistochemistry, intralesional antigens of CAdV-1 were identified within the epithelial cells of the liver and lungs; there was positive immunolabeling for CPV-2 antigens in degenerated cardiomyocytes. These findings confirmed the active participation of C. halotolerans in the development of disseminated cladosporiosis in this dog and represent a rare occurrence of concomitant infection with CAdV-1 and CPV-2.

Perylenequinones are a family of structurally related polyketide fungal toxins with nearly universal toxicity. These photosensitizing compounds absorb light energy which enables them to generate reactive oxygen species that damage host cells. This potent mechanism serves as an effective weapon for plant pathogens in disease or niche establishment. The sugar beet pathogen Cercospora beticola secretes the perylenequinone cercosporin during infection. We have shown recently that the cercosporin toxin biosynthesis (CTB) gene cluster is present in several other phytopathogenic fungi, prompting the search for biosynthetic gene clusters (BGCs) of structurally similar perylenequinones in other fungi. Here, we report the identification of the elsinochrome and phleichrome BGCs of Elsinoë fawcettii and Cladosporium phlei, respectively, based on gene cluster conservation with the CTB and hypocrellin BGCs. Furthermore, we show that previously reported BGCs for elsinochrome and phleichrome are involved in melanin production. Phylogenetic analysis of the corresponding melanin polyketide synthases (PKSs) and alignment of melanin BGCs revealed high conservation between the established and newly identified C. beticola, E. fawcettii and C. phlei melanin BGCs. Mutagenesis of the identified perylenequinone and melanin PKSs in C. beticola and E. fawcettii coupled with mass spectrometric metabolite analyses confirmed their roles in toxin and melanin production., (© 2018 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

Fungal rots in sugar beet roots held in long-term storage can lead to considerable sucrose loss but the incidence and distribution of fungal rots inside sugar beet piles and pathogenicity for some species is poorly understood. Thus, Idaho sugar beet held in five outdoor and two indoor piles in 2014 and 2015 were investigated. The root surface area covered by fungal growth and discolored and healthy tissue were assessed in nine 1-m 2 areas per pile using a stratified random sampling design. Pathogenicity was evaluated indoors via plug inoculation in 2015 and 2016. Botrytis cinerea covered more root surface area inside indoor piles (6 to 22%) than outdoor piles (0 to 3%) (P < 0.0001). No trends were evident for the Athelia-like sp. (0 to 15%) and Penicillium-type spp. (0 to 8%). Penicillium-type isolates comprised the following species: 60% Penicillium expansum, 34% P. cellarum, 3% P. polonicum, and 3% Talaromyces rugulosus. Trace levels (<1% of root surface) of other fungi, including Cladosporium and Fusarium spp., were evident on roots and in isolations. Based on sample location in a pile, there were no trends or differences; however, two outdoor piles (OVP1 and OVP2) had more healthy tissue (90 to 96%) than other piles (28 to 80%) (P < 0.0001). When the pathogenicity tests were analyzed by species, all were significantly different from each other (P < 0.0001), except for P. polonicum and P. expansum: B. cinerea (61 mm of rot), P. polonicum (36 mm), P. expansum (35 mm), P. cellarum (28 mm), Athelia-like sp. (21 mm), T. rugulosus (0 mm; not different from check), and noninoculated check (0 mm). The OVP1 and OVP2 piles had negligible fungal growth on roots after more than 120 days of storage under ambient conditions, which indicates that acceptable storage can be achieved over this time period through covering piles with tarps and cooling with ventilation pipe.

Effectors are microbial-derived secreted proteins with an essential function in modulating host immunity during infections. CfAvr4, an effector protein from the tomato pathogen Cladosporium fulvum and the founding member of a fungal effector family, promotes parasitism through binding fungal chitin and protecting it from chitinases. Binding of Avr4 to chitin is mediated by a carbohydrate-binding module of family 14 (CBM14), an abundant CBM across all domains of life. To date, the structural basis of chitin-binding by Avr4 effector proteins and of recognition by the cognate Cf-4 plant immune receptor are still poorly understood. Using X-ray crystallography, we solved the crystal structure of CfAvr4 in complex with chitohexaose [(GlcNAc)6] at 1.95Å resolution. This is the first co-crystal structure of a CBM14 protein together with its ligand that further reveals the molecular mechanism of (GlcNAc)6 binding by Avr4 effector proteins and CBM14 family members in general. The structure showed that two molecules of CfAvr4 interact through the ligand and form a three-dimensional molecular sandwich that encapsulates two (GlcNAc)6 molecules within the dimeric assembly. Contrary to previous assumptions made with other CBM14 members, the chitohexaose-binding domain (ChBD) extends to the entire length of CfAvr4 with the reducing end of (GlcNAc)6 positioned near the N-terminus and the non-reducing end at the C-terminus. Site-directed mutagenesis of residues interacting with (GlcNAc)6 enabled the elucidation of the precise topography and amino acid composition of Avr4's ChBD and further showed that these residues do not individually mediate the recognition of CfAvr4 by the Cf-4 immune receptor. Instead, the studies highlighted the dependency of Cf-4-mediated recognition on CfAvr4's stability and resistance against proteolysis in the leaf apoplast, and provided the evidence for structurally separating intrinsic function from immune receptor recognition in this effector family., Competing Interests: The authors have declared that no competing interests exist.

The aim of this study was to identify fungal species present in 200 samples of rosemary, fennel, cinnamon, clove, pepperoni, black and white pepper and oregano and evaluate the mycotoxigenic potential of the some Aspergilli isolated. Clove, black and white peppers were analyzed by direct plating. For rosemary, cinnamon, fennel, pepperoni pepper and oregano samples were used spread plate. Mycotoxigenic capacity was verified by the agar plug method. With the exception of clove, all the spices showed high fungal contamination, especially by Aspergillus sp., Penicillium sp. and Cladosporium sp. Frequency of toxigenic Aspergillus spp. was intense in white and black peppers, with presence of Aspergillus flavus (up to 32%), Aspergillus nomius (up to 12%), Aspergillus parasiticus (up to 4%), Aspergillus niger complex (up to 52%), Aspergillus ochraceus (up 12%) and Aspergillus carbonarius (up to 4%). 14,2% of A. flavus isolated from black pepper were aflatoxins producers. In the white pepper, 66.7% of A. flavus isolates and 100% of A. nomius were aflatoxigenic. Oregano showed the highest number of A. niger complex isolates (49), however, only 2.04% produced ochratoxin A. This study showed a huge fungal presence in spices, which could compromise the sensorial quality of these products and represent a hazard for consumers., (Copyright © 2018. Published by Elsevier Ltd.)

The occurrence of Cladosporium in cold ecosystems has been evidenced long before, and most of the knowledge about nutrient utilization of this genus is sporadic. An alpine soil isolate C. neopsychrotolerans SL-16, showing great cold tolerance and significant lignocellulose-degrading capability, was sequenced to form a 35.9 Mb genome that contains 13,456 predicted genes. Functional annotation on predicted genes revealed a wide array of proteins involved in the transport and metabolism of carbohydrate, protein and lipid. Large numbers of transmembrane proteins (967) and CAZymes (571) were identified, and those related to hemicellulose degradation was the most abundant. To undermine the hemicellulose (xyaln as the main component) utilization mechanism of SL-16, the mRNA levels of 23 xylanolytic enzymes were quantified, and representatives of three glycoside hydrolase families were functionally characterized. The enzymes showed similar neutral, cold active and thermolabile properties and synergistic action on xylan degradation (the synergy degree up to 15.32). Kinetic analysis and sequence and structure comparison with mesophilic and thermophilic homologues indicated that these cold-active enzymes employed different cold adaptation strategies to function well in cold environment. These similar and complementary advantages in cold adaptation and catalysis might explain the high efficiency of lignocellulose conversion observed in SL-16 under low temperatures.

Introduction: Airborne fungi are considered important causes of respiratory allergy and diseases. The knowledge of these fungi in a city or region is important for the ecological diagnosis and specific treatment of respiratory manifestations induced by inhalation of fungal agents. Therefore, in recent years bioaerosols and their density in the air is highly regarded. Identifying fungi with common mycological methods as well as molecular about 5% of people are prone to allergic symptoms of the respiratory tract caused by molds during their lives. Fungi are being considered as one of the most common triggers of asthma, allergic rhinitis and other respiratory problems., Aims: We aimed at the investigation of the airborne fungi of Ahvaz, Khuzestan province, Iran, during spring and autumn months., Materials and Methods: A total of 224 Petri dishes with Sabouraud dextrose agar medium containing chloramphenicol were exposed at seven different regions in the city. Two hundred and twenty-four samples (112 samples per season) were taken from outdoor air seven areas of Ahvaz using microbial sampler (Quick Take-30, SKC, USA) with Debbi 14.3L/min during 5minutes directly on Sabouraud dextrose agar medium with Chloramphenicol and Dechlorane. The effects of environmental factors such as temperature, humidity and wind velocity on bioaerosol concentrations were studied., Results: The dishes exposed yielded 1240 fungal colonies of 24 genera. The most predominant were: Cladosporium spp. (37.1%), Alternaria spp. (25.7%), Aspergillus spp. (13.5%), Penicillium spp. (5.9%), Drechslera spp. (3.46%) and Stemphylium spp. (3.46%). The maximum and minimum numbers of airborne fungi were isolated from humid (32.3%) and industrial (17%) environments, respectively. In addition, positive correlation was observed between the number of fungi and relative humidity (r=0.476, P=0.000) in spring and autumn seasons., Conclusion: This study indicated that the incidence of airborne fungi with clinical significance had a direct relationship with the variation of environmental conditions., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Tomato leaf mold disease is caused by the biotrophic fungus Cladosporium fulvum. During infection, C. fulvum produces extracellular small secreted protein (SSP) effectors that function to promote colonization of the leaf apoplast. Resistance to the disease is governed by Cf immune receptor genes that encode receptor-like proteins (RLPs). These RLPs recognize specific SSP effectors to initiate a hypersensitive response (HR) that renders the pathogen avirulent. C. fulvum strains capable of overcoming one or more of all cloned Cf genes have now emerged. To combat these strains, new Cf genes are required. An effectoromics approach was employed to identify wild tomato accessions carrying new Cf genes. Proteomics and transcriptome sequencing were first used to identify 70 apoplastic in planta-induced C. fulvum SSPs. Based on sequence homology, 61 of these SSPs were novel or lacked known functional domains. Seven, however, had predicted structural homology to antimicrobial proteins, suggesting a possible role in mediating antagonistic microbe-microbe interactions in planta. Wild tomato accessions were then screened for HR-associated recognition of 41 SSPs, using the Potato virus X-based transient expression system. Nine SSPs were recognized by one or more accessions, suggesting that these plants carry new Cf genes available for incorporation into cultivated tomato.

The role of different growth media and chemical enhancer on synthesis of secondary metabolites Cladosporium resinae (NRL-6437) was investigated for their in vitro biological activities. Cladosporium resinae (NRL-6437) were grown in various nutrient media (Czapeak-dox Broth (CB), Czapeak Yeast-extract Broth (CYB), Yeast Extract Sucrose (YES), Potato Dextrose Broth (PDB) and Czapeak-dox (supplemented with glucose and starch) Broth (CGSB) for the production of metabolites. Two chemical epigenetic modifiers (suberoyl-anilide hydroxamic acid (SAHA) and 5-azacytidine (5-AZA) were also used for the expression of silent genes for secondary metabolite production. Our results indicated that among different media, Czapeak yeast extract broth produced more secondary metabolites. Application of 15mM of both modifiers was effective for the expressions of silent genes resulting in an increased metabolites production. Secondary metabolites extracted in ethyl acetate and fractionized in n-Hexane were also tested for their biological activity. The secondary metabolites revealed varying degrees of growth inhibitions of the tested organisms. Similarly, these metabolites were also active against brine shrimps and Lemna.

Taxol or paclitaxel, an approved drug by the Food and Drug Administration, is being used for the treatment of human cancers. This study aimed to isolate and determine different species of native endophytic fungi from Iranian Taxus baccata (yew) plants located in the northern forests of Iran. To do so, a novel molecular screening approach was performed for 50 isolated endophytic fungi through amplification of exon No. 1 of taxadine synthase as a key gene in taxol production pathway. We used effective colony-polymerase chain reaction technique for rapid screening of potent taxol-producing fungi instead of genomic DNA extraction. Production of taxol was performed in batch culture by selected fungi individually and produced taxol was assayed quantitatively by high-performance liquid chromatography using standard taxanes. We found that only six fungi could produce taxol and baccatin III. Interestingly, after 7 days of incubation, the highest level of taxol was found to be 129 and that of baccatin 139.2 mg/kg dw for two native isolated Cladosporium sp. named F1 and F3. The fungal taxols could decrease cell viability in MTT assay same as commercial taxol. The fungal taxols semi-quantitatively showed antimitotic effects on MCF-7 cells as human breast cancer cell line. The expression of bcl-2 anti-apoptotic gene, in contrast to bax pro-apoptotic gene, significantly decreased after treatment by standard and fungal taxols. As fungal taxol was produced simpler than other methods and could significantly affect viability and specific genes expression profile, it is recommended that using of taxol-producing fungi from Iranian yew could be a safe and confident procedure to overcome challenges of using other methods.

The occurrence of interaction between insects and fungi is interesting from an ecological point of view, particularly when these interactions involve insect pests and plant pathogens within an agroecosystem. In this study, we aimed to perform an accurate analysis on the fungal microbiota associated to Bactrocera oleae (Rossi) through a metabarcoding approach based on 454 pyrosequencing. From this analysis, we retrieved 43,549 reads that clustered into 128 operational taxonomic units (OTUs), of which 29 resulted in the "core" associate fungi of B. oleae. This fungal community was mainly represented by sooty mould fungi, such as Cladosporium spp., Alternaria spp. and Aureobasidium spp., by plant pathogens like Colletotrichum spp. and Pseudocercospora spp., along with several other less abundant taxa whose ecology is unclear in most of the cases. Our findings lead to new insights into the microbial ecology of this specific ecological niche, enabling the understanding of a complex network of interactions within the olive agroecosystem.