Reactive Oxygen Species Localization Programs Inflammation to Clear Microbes of Different Size.

How the number of immune cells recruited to sites of infection is determined and adjusted to differences in the cellular stoichiometry between host and pathogen is unknown. Here, we have uncovered a role for reactive oxygen species (ROS) as sensors of microbe size. By sensing the differential localization of ROS generated in response to microbes of different size, neutrophils tuned their interleukin (IL)-1ß expression via the selective oxidation of NF-κB, in order to implement distinct inflammatory programs. Small microbes triggered ROS intracellularly, suppressing IL-1ß expression to limit neutrophil recruitment as each phagocyte eliminated numerous pathogens. In contrast, large microbes triggered ROS extracellularly, amplifying IL-1ß expression to recruit numerous neutrophils forming cooperative clusters. Defects in ROS-mediated microbe size sensing resulted in large neutrophil infiltrates and clusters in response to small microbes that contribute to inflammatory disease. These findings highlight the impact of ROS localization on signal transduction.

Discovery of highly reactive self-splicing group II introns within the mitochondrial genomes of human pathogenic fungi.

Fungal pathogens represent an expanding global health threat for which treatment options are limited. Self-splicing group II introns have emerged as promising drug targets, but their development has been limited by a lack of information on their distribution and architecture in pathogenic fungi. To meet this challenge, we developed a bioinformatic workflow for scanning sequence data to identify unique RNA structural signatures within group II introns. Using this approach, we discovered a set of ubiquitous introns within thermally dimorphic fungi (genera of Blastomyces, Coccidioides and Histoplasma). These introns are the most biochemically reactive group II introns ever reported, and they self-splice rapidly under near-physiological conditions without protein cofactors. Moreover, we demonstrated the small molecule targetability of these introns by showing that they can be inhibited by the FDA-approved drug mitoxantrone in vitro. Taken together, our results highlight the utility of structure-based informatic searches for identifying riboregulatory elements in pathogens, revealing a striking diversity of reactive self-splicing introns with great promise as antifungal drug targets.

miRNA Mediated Regulation and Interaction between Plants and Pathogens.

Plants have evolved diverse molecular mechanisms that enable them to respond to a wide range of pathogens. It has become clear that microRNAs, a class of short single-stranded RNA molecules that regulate gene expression at the transcriptional or post-translational level, play a crucial role in coordinating plant-pathogen interactions. Specifically, miRNAs have been shown to be involved in the regulation of phytohormone signals, reactive oxygen species, and NBS-LRR gene expression, thereby modulating the arms race between hosts and pathogens. Adding another level of complexity, it has recently been shown that specific lncRNAs (ceRNAs) can act as decoys that interact with and modulate the activity of miRNAs. Here we review recent findings regarding the roles of miRNA in plant defense, with a focus on the regulatory modes of miRNAs and their possible applications in breeding pathogen-resistance plants including crops and trees. Special emphasis is placed on discussing the role of miRNA in the arms race between hosts and pathogens, and the interaction between disease-related miRNAs and lncRNAs.

Searching for New Beneficial Bacterial Isolates of Wild Raspberries for Biocontrol of Phytopathogens-Antagonistic Properties and Functional Characterization.

The threat caused by plants fungal and fungal-like pathogens is a serious problem in the organic farming of soft fruits. The European Commission regulations prohibit some commercially available chemical plant protection products, and instead recommend the use of natural methods for improving the microbial soil status and thus increasing resistance to biotic stresses caused by phytopathogens. The solution to this problem may be biopreparations based on, e.g., bacteria, especially those isolated from native local environments. To select proper bacterial candidates for biopreparation, research was provided to preliminarily ensure that those isolates are able not only to inhibit the growth of pathogens, but also to be metabolically effective. In the presented research sixty-five isolates were acquired and identified. Potentially pathogenic isolates were excluded from further research, and beneficial bacterial isolates were tested against the following plant pathogens: Botrytis spp., Colletotrichum spp., Phytophthora spp., and Verticillium spp. The eight most effective antagonists belonging to Arthrobacter, Bacillus, Pseudomonas, and Rhodococcus genera were subjected to metabolic and enzymatic analyses and a resistance to chemical stress survey, indicating to their potential as components of biopreparations for agroecology.

Invasive cutaneous Neoscytalidium infections in renal transplant recipients: a series of five cases.

BACKGROUND: Neoscytalidium species (formerly Scytalidium species) are black fungi that usually cause cutaneous infections mimicking dermatophytes lesions. Very few publications have reported invasive or disseminated infections. CASE PRESENTATION: In this paper, we report the clinical presentations, treatments and outcomes of five cases of invasive Neoscytalidium infections with cutaneous involvement, including two cases with disseminated infection, in five renal transplant recipients. To our knowledge, this is the first report of a series-albeit small-of renal transplant patients in whom this infection was identified. All cases occurred in a single hospital in Paris, France, between 2001 and 2011. Patients all originate from tropical area. CONCLUSION: Treatments of Neoscytalidium infection varied greatly, underlining the lack of a recommendation for a standardized treatment. All patients were cured after long-term antifungal therapy and/or surgical excision. Interestingly, one patient with disseminated infection involving the left elbow, the right leg, the lungs and the nasal septum was cured by medical therapy only without surgery. This may suggest that in contrast to others mycoses (such as mucormycosis), an adequate medical treatment could be sufficient for treating Neoscytalidium. We also point out the difficulties we had in diagnosing two patients with Kaposi's sarcoma because of the similarity of the lesions. Furthermore, our report underlines the need to check for this rare infection in immunocompromised kidney transplant recipients originating from tropical areas.

Pathogenicity of conidia-based preparations of entomopathogenic fungi against the greenhouse pest aphids Myzus persicae, Aphis gossypii, and Aulacorthum solani (Hemiptera: Aphididae).

Seeking new isolates of entomopathogenic fungi with greater virulence against greenhouse aphid pests than those currently registered in North America for control of these insects, single-dose screening assays of 44 selected fungal isolates and 4 commercially available strains were conducted against first-instar nymphs of Myzus persicae and Aphis gossypii. The assays identified a number of Beauveria and Metarhizium isolates with virulence equal to or greater than that of the commercial strains against the nymphal aphids, but none exhibited exceptionally high virulence. Virulence of Isaria isolates was unexpectedly low (<31% mortality at doses>1000conidia/mm(2)). In dose-response assays, Beauveria ARSEF 5493 proved most virulent against M. persicae and A. gossypii; however, LC50s of this isolate did not differ significantly from those of B. bassiana commercial strain JW-1. Dose-response assays were also conducted with Aulacorthum solani, the first reported evaluations of Beauveria and Metarhizium against this pest. The novel isolate Metarhizium 5471 showed virulence⩾that of Beauveria 5493 in terms of LC25 and LC50, but 5493 produced a steeper dose response (slope). Additional tests showed that adult aphids are more susceptible than nymphs to fungal infection but confirmed that infection has a limited pre-mortem effect on aphid reproduction. Effects of assay techniques and the potential of fungal pathogens as aphid-control agents are discussed.

Complexes of potentially pathogenic microscopic fungi in anthropogenic polluted soils.

This study was undertaken to investigate the species' diversity and structure of potentially pathogenic microscopic fungal complexes in podzolic soils polluted by fluorine, heavy metals (Cu, Ni, Co), oil products (diesel fuel, gas condensate, mazut). Lists of potentially pathogenic fungi isolated from soils are made specifically for north-western part of Russia (Kola Peninsula). The majority of studied fungus species belong to the following genera: Penicillium, Aspergillus, Mucor, Lecanicillium and Phoma. Penicillium miczynskii was identified as the most stable type of fungus with respect to all studied types of oil products. Mucor hiemalis was identified as the most sensitive type. An increase of 15% portion of potentially pathogenic fungi as compared to the background soil in zones of aluminum and copper-nickel plants was revealed. The results indicate an increase of 20-25% of potentially pathogenic fungi in pollution of soil with oil products. The structure of fungal complexes was observed to have changed in the polluted soils and the species number and frequency of occurrence of potentially pathogenic fungi were also increased.

[Formation peculiarities of Quercus robur L. acorn mycobiota under conditions of Kyiv Polissya].

The structure of mycobiota (epiphytic and from inner tissues) of Q. robur acorns (580 samples) under conditions of Kyiv Polissya was studied. Acorn samples were collected from oak trees during vegetation period. The epiphytes and mycobiota isolated from inner tissues of acorns was represented by 38 and 20 species, respectively. The majority of fungi were isolated in July. Cosmopolitan and non-specific species Acremonium sp., Alternaria alternata, Cladosporium cladosporioides, Trichoderma harzianum, T. viride and specific ones - Phoma sp. and Phomopsis sp. were isolated from inner acorn tissues. It was established that potential seed and seedling pathogens infect acorns during the process of their formation. The paper is presented in Ukrainian.

Perspectives on the potential of entomopathogenic fungi in biological control of ticks.

Ticks are serious health threats for humans, and both domestic and wild animals. Ticks are controlled mostly by application of chemical products; but these acaricides have several negative side effects, including toxicity to animals, environmental contamination, and induction of chemical resistance in some tick populations. Entomopathogenic fungi infect arthropods in nature and can occur at enzootic or epizootic levels in their host populations. Laboratory studies clearly demonstrate that these fungi can cause high mortality in all developmental stages of several tick species, and also reduce oviposition of infected engorged females. Tick mortality following application of fungi in the field, however, often is less than that suggested by laboratory tests. This is due to many negative biotic and climatic factors. To increase efficacy of fungal agents for biological control of ticks under natural conditions, several points need consideration: (1) select effective isolates (viz., high virulence; and tolerance to high temperature, ultraviolet radiation and desiccation); (2) understand the main factors that affect virulence of fungal isolates to their target arthropods including the role of toxic metabolites of the fungal isolates; and (3) define with more precision the immune response of ticks to infection by entomopathogenic fungi. The current study reviews recent literature on biological control of ticks, and comments on the relevance of these results to advancing the development of fungal biocontrol agents, including improving formulation of fungal spores for use in tick control, and using entomopathogenic fungi in integrated pest (tick) management programs.

The ectopic expression of a pectin methyl esterase inhibitor increases pectin methyl esterification and limits fungal diseases in wheat.

Cell wall pectin methyl esterification can influence plant resistance because highly methyl-esterified pectin can be less susceptible to the hydrolysis by pectic enzymes such as fungal endopolygalacturonases (PG). Pectin is secreted into the cell wall in a highly methyl-esterified form and, here, is de-methyl esterified by pectin methyl esterase (PME). The activity of PME is controlled by specific protein inhibitors called PMEI; consequently, an increased inhibition of PME by PMEI might modify the pectin methyl esterification. In order to test the possibility of improving wheat resistance by modifying the methyl esterification of pectin cell wall, we have produced durum wheat transgenic lines expressing the PMEI from Actinidia chinensis (AcPMEI). The expression of AcPMEI endows wheat with a reduced endogenous PME activity, and transgenic lines expressing a high level of the inhibitor showed a significant increase in the degree of methyl esterification. These lines showed a significant reduction of disease symptoms caused by the fungal pathogens Bipolaris sorokiniana or Fusarium graminearum. This increased resistance was related to the impaired ability of these fungal pathogens to grow on methyl-esterified pectin and to a reduced activity of the fungal PG to hydrolyze methyl-esterified pectin. In addition to their importance for wheat improvement, these results highlight the primary role of pectin despite its low content in the wheat cell wall.

Antifungal therapy in an athymic murine model of chromoblastomycosis by Fonsecaea pedrosoi.

We have compared the efficacy of terbinafine (TRB) and itraconazole (ITZ), the recommended drugs for the treatment of chromoblastomycosis, with that of posaconazole (PSC) and voriconazole (VRC) in athymic mice infected with the fungus Fonsecaea pedrosoi. Three weeks after challenge, mice were treated for 4 months with PSC at 10 or 20 mg/kg of body weight/day, with VRC at 10 or 20 mg/kg/day, with ITZ at 25 or 50 mg/kg/day, or with TRB at 150 or 250 mg/kg/day. The progress of the infection was evaluated by measuring the size of the lesions, by histopathological studies, and by cultures of the excised tissue. The two doses of PSC followed by the highest doses of ITZ and TRB showed the best results while VRC did not show efficacy. PSC could be an alternative in the treatment of chromoblastomycosis.

Pulmonary fungal infection caused by Neoscytalidium dimidiatum in a Risso's dolphin (Grampus griseus).

Neoscytalidium dimidiatum was isolated from two 12-18 cm abscesses in the lung and the mediastinal lymph nodes of a stranded Risso's dolphin (Grampus griseus). Histopathologic examination of samples of these organs revealed the presence of hyphae and sclerotic body-like fungal elements. Photobacterium damselae subsp. damselae was recovered from the dolphin's organs which also were found to contain numerous Monorygma grimaldii cysts. No histopathological signs of morbillivirus infection were seen. To the best of our knowledge, this is the first report of N. dimidiatum infection in a sea mammal.

Pneumonia and lung infections due to emerging and unusual fungal pathogens.

Invasive mold infections affecting the lungs are increasing in incidence and diversity. Severely immunocompromised patients are particularly vulnerable to infection from unusual, normally nonpathogenic fungi that are found naturally in the environment. Certain fungi such as Scedosporium and the dematiaceous fungi also cause lung disease in hosts without overt immune compromise. The impacts of these emerging pathogens range from airway colonization to locally invasive lung, and disseminated, disease. Diagnosis requires isolation and identification of the etiologic agent by either or both phenotypic and molecular biology methods. Evidence of tissue invasion on histopathology is often required to distinguish infection from colonization. Diagnostic imaging techniques are nonspecific, and there are no reliable serological biomarkers of infection. Many rare molds and yeasts demonstrate reduced in vitro susceptibility to antifungal agents. Although amphotericin B formulations remain clinically useful for many of these infections, voriconazole and posaconazole are more effective for some of these difficult-to-treat pathogens. Surgical resection of diseased tissue and support of the host immune system are often required to optimize outcomes.

Diversity of fungal pathogens associated with loquat and development of novel virulence scales.

Loquat [Eriobotrya japonica (Thunb.) Lindl.] is an important fruit crop in Pakistan; however, a constant decline in its production is noted due biotic and abiotic stresses, particularly disease infestation. Fungal pathogens are the major disease-causing agents; therefore, their identification is necessary for devising management options. This study explored Taxila, Wah-Cantt, Tret, Chatar, Murree, Kalar-Kahar, Choa-Saidan-Shah and Khan-Pur districts in the Punjab and Khyber Paktoon Khawa (KPK) provinces of Pakistan to explore the diversity of fungal pathogens associated with loquat. The samples were collected from these districts and their microscopic characterizations were accomplished for reliable identification. Alternaria alternata, Curvularia lunata, Lasiodiplodia theobromae, Aspergilus flavis, Botrytis cinerea, Chaetomium globosum, Pestalotiopsis mangiferae and Phomopsis sp. were the fungal pathogens infesting loquat in the study area. The isolates of A. alternata and C. lunata were isolated from leaf spots and fruit rot, while the isolates of L. theobromae were associated with twig dieback. The remaining pathogens were allied with fruit rot. The nucleotide evidence of internal transcribed spacer (ITS) regions (ITS1, 5.8S, and ITS2) were computed from all the pathogens and submitted in the database of National Center for Biotechnology Information (NCBI). For multigene analysis, beta-tubulin (BT) gene and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) regions were explored for A. alternata and C. lunata isolates, respectively. The virulence scales of leaf spots, fruit rot, and twig dieback diseases of loquat were developed for the first time through this study. It is the first comprehensive study with morpho-molecular identification, and newly developed virulence scales of the fungal pathogens associated with loquat, which improves the understanding of these destructive diseases.

Sexual reproduction and the evolution of microbial pathogens.

Three common systemic human fungal pathogens--Cryptococcus neoformans, Candida albicans and Aspergillus fumigatus--have retained all the machinery to engage in sexual reproduction, and yet their populations are often clonal with limited evidence for recombination. Striking parallels have emerged with four protozoan parasites that infect humans: Toxoplasma gondii, Trypanosoma brucei, Trypanosoma cruzi and Plasmodium falciparum. Limiting sexual reproduction appears to be a common virulence strategy, enabling generation of clonal populations well adapted to host and environmental niches, yet retaining the ability to engage in sexual or parasexual reproduction and respond to selective pressure. Continued investigation of the sexual nature of microbial pathogens should facilitate both laboratory investigation and an understanding of the complex interplay between pathogens, hosts, vectors, and their environments.

Occupational exposure and sensitization to fungi among museum workers.

BACKGROUND: Museum employees are exposed to fungi and storage mites in the workplace. AIMS: To evaluate the prevalence and risk factors of sensitization to moulds, as well as clinical symptoms associated with allergy in museum workers. METHODS: A total of 103 employees of the Polish National Museum (NM) in Warsaw, potentially exposed to fungi during their work, were assessed using a questionnaire and skin prick tests to common allergens and fungal extracts. The level of total and serum-specific IgE to moulds was evaluated, and spirometry was performed in all subjects. Mycological analysis of the workplace was also performed. RESULTS: Penicillium, Aspergillus, Cladosporium, Alternaria, Trichoderma, Acremonium and Paecilomyces were the most frequent species isolated from investigated exhibits of NM. Thirty per cent of museum employees were sensitized to at least one of the fungal allergens. Logistic regression analysis revealed that duration of occupational exposure lasting >5 years, family history of atopy, presence of a cat at home, sinusitis, allergic rhinitis and a history of frequent respiratory infections were risk factors for the development of sensitization to fungi in this working group. CONCLUSIONS: This study suggests an important role of fungi as occupational allergens for museum workers. The prevalence of allergic symptoms among employees of NM was relatively high. Further studies are necessary to elucidate the importance of particular fungal species in the development of occupational allergy.

Evaluation of soybean genotypes for reaction to natural field infection by Cercospora species causing purple seed stain.

Purple seed stain (PSS) of soybean (Glycine max (L.) Merr.) is a prevalent seed disease. It results in poor seed quality and reduced seed lot market grade, and thus undermines value of soybean worldwide. The objectives of this research were to evaluate the reaction of selected soybean genotypes collected from 15 countries representing maturity groups (MGs) III, IV, and V to PSS, and to identify new sources of resistance to PSS based on three years of evaluation of natural field infection by Cercospora spp. in the Mississippi Delta of the U. S. In this study, 42 soybean genotypes were evaluated in 2010, 2011, and 2012. Seventeen lines including six MG III (PI 88490, PI 504488, PI 417361, PI 548298, PI 437482, and PI 578486), seven MG IV (PI 404173, PI 346308, PI 355070, PI 416779, PI 80479, PI 346307, and PI 264555), and four MG V (PI 417567, PI 417420, PI 381659, and PI 407749) genotypes had significantly lower percent seed infection by Cercospora spp. than the susceptible checks and other genotypes evaluated (P ≤ 0.05). These genotypes of soybean can be used in developing soybean cultivars or germplasm lines with resistance to PSS and for genetic mapping of PSS resistance genes. In addition, among these 17 lines with different levels of resistance to PSS, nine soybean genotypes (PI 417361, PI 504488, PI 88490, PI 346308, PI 416779, PI 417567, PI 381659, PI 417567, and PI 407749) were previously reported as resistant to Phomopsis seed decay. Therefore, they could be useful in breeding programs to develop soybean cultivars with improved resistance to both seed diseases.

Synergistic interactions between chitinase ChiCW and fungicides against plant fungal pathogens.

Antifungal activity of ChiCW and synergistic interactions between ChiCW with fungicides were investigated. Conidial germinations of phytopathogenic fungi, Alternaria brassicicola, Botrytis elliptica, and Colletotrichum gloeoporioides, were inhibited by ChiCW but A. longipes was not. In addition, ChiCW showed synergistic effect with fungicides Switch (cyprodinil+fludioxonil) and tebuconazole to inhibit fungal conidial germinations. The level of synergism of ChiCW with tebuconazole was higher than that with Switch. The results indicate that ChiCW may exhibit a higher level of synergism with fungicides that have a primary effect upon membranes.

Pathogenicity and thermotolerance of entomopathogenic fungi for the control of the scab mite, Psoroptes ovis.

Psoroptes ovis is responsible for a highly contagious skin condition, both in sheep and cattle. This parasite has a marked economical impact in the sheep and cattle industry. Biological control is considered as a realistic alternative to chemotherapeutic control. Laboratory experiments were carried out to evaluate the pathogenicity and the thermotolerance of twelve isolates of entomopathogenic fungi from four genera (Beauveria Vuillemin, Metarhizium Sorokin, Paecilomyces Bainier and Verticillium Nees). The pathogenicity was evaluated by the survival of P. ovis females after exposure to 10(6) to 10(8) conidia ml(-1) in humidity chambers. Results revealed intra- and interspecies differences. All isolates with the exception of B. bassiana IHEM3558 and V. lecanii MUCL8672 induced 50% mortality within 2 days at the highest concentration. At this concentration the entire mite population became infected with all isolates but B. bassiana IHEM3558; however, only four isolates gave rise to 100% infected cadavers at the lowest concentration. The thermotolerance of each isolate was evaluated by measuring its growth on an artificial medium kept between 25 and 37.5 degrees C. All isolates were able to grow up to 30 degrees C but only two, M. anisopliae IHEM18027 and Paecilomyces farinosus MUCL18885, tolerated temperatures up to 35 degrees C. These two isolates could be considered as good candidates for further use as biopesticide taking into account their virulence and thermotolerance. Other critical factors linked with the implementation of this type of biocontrol in P. ovis infected animals are discussed.

Enabling mycelial application of Hirsutella thompsonii for managing the coconut mite.

Laboratory and field studies were conducted to examine the prospect of mycelial application of Hirsutella thompsonii as an alternative to the use of mycelial-conidial formulations of the fungus in the suppression of the coconut mite, Aceria guerreronis. In a series of laboratory experiments, glycerol, yeast extract powder and dehydrated malt extract broth were found to be the best among nine substances investigated as possible adjuvants for use on coconut palms in the field along with H. thompsonii mycelia. H. thompsonii biomass in the presence of adjuvants not only produced more colonies but also yielded more conidia per pellet. In terms of the density of conidia generated on a mycelial mat the treatments varied highly significantly in two methods, with glycerol showing an average of 106% increase over control. Though irradiance with simulated sunlight resulted in reduced conidiogenesis, in general, adjuvant-treated pellets, both exposed and unexposed to simulated sunlight, produced substantial conidiation compared with control, irrespective of the two incubation conditions. Better conidiation was observed under alternating light-dark regime than under total darkness in all the treatments. Glycerol boosted the pathogenicity of H. thompsonii by 16.5% over control. In the field, a newly developed mycelial formulation of H. thompsonii applied after tank-mixing separately with the three selected adjuvants brought down the post-treatment population of the coconut mite by 85.6-97.1%. Application of the fungus in combination with glycerol resulted in a tolerable mean nut damage grade of 2.0 during the pre-harvest stage, compared with an acute score of 4.0 in control palms.