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At least 37 records · Page 2

The lipid composition and its alteration during the growth stage in pathogenic fungus, epidermophyton floccosum

Qualitative and quantitative changes of lipid components during the growth stages were studied in E. floccosum. The acyl group components of total lipids of Trichophyton rubrum and Microsporum cookei were also examined. The lipids of E. floccosum amounted to approximately 4% of the dry cell weight. Neutral lipids mainly consisted of triglycerides and sterols, and major polar lipids were phosphatidylcholine, phosphatidylethanolamine, and an unknown lipid X. The fatty acids in tryglycerides and phospholipids were palmitic, palmitoleic, stearic, oleic, and linoleic acids. The unknown polar lipid X which appeared between phosphatidylethanolamine and cardiolipin on thin layer chromatography plates contained no phosphorus. There was no significant change in the fatty acid components of E. floccosum and T. rubrum during the cell growth, whereas profound changes occurred in M. cookei. The sterol components of E. floccosum showed striking changes depending on the growth stage.

Yamada, T.

Total Dose Effects (TDE) of heavy ionizing radiation in fungus spores and plant seeds: Preliminary investigations

The opportunity to compare cosmic radiation effects caused during long and short duration exposure flights in biological objects are limited until now, and data obtained so far are very rare and insufficient. Because of the very long exposure of the experiment during the Long Duration Exposure Facility (LDEF) mission (approximately 2000 days) structural changes of the hardware material can be expected which will influence its biocompatibility and, thus, will interact with the radiobiological effects. The aim of the experiment flown on LDEF was a detailed investigation of biological effects caused by cosmic radiation especially of particles of high atomic number Z and high energy. The flight hardware consisted of standard BIOSTACK containers; in these containers a special sandwich construction consisted of visual plastic detectors with seed rsp. spore layers interlocked.

Kranz, A. R.

Kinetic studies on the control of the bean rust fungus (Uromyces phaseoli L.) by an inhibitor of polyamine biosynthesis

alpha-Difluoromethylornithine (DFMO), a specific and irreversible inhibitor of the polyamine biosynthetic enzyme ornithine decarboxylase, effectively inhibits mycelial growth of several phytopathogenic fungi on defined media in vitro and provides systemic protection of bean plants against infection by Uromyces phaseoli L. race 0 (MV Rajam, AW Galston 1985 Plant Cell Physiol 26: 683-692; MV Rajam et al. 1985 Proc Natl Acad Sci USA 82: 6874-6878). We now find that application of 0.5 millimolar DFMO to unifoliolate leaves of Pinto beans up to 3 days after inoculation with uredospores of U. phaseoli completely inhibits the growth of the pathogen, while application 4 or 5 days after inoculation results in partial protection against the pathogen. Spores do not germinate on the surface of unifoliolate leaves treated with DFMO 1 day before infection, but addition of spermidine to the DFMO treatments partially reverses the inhibitory effect. The titer of polyamines in bean plants did not decline after DFMO treatment; rather, putrescine and spermidine contents actually rose, probably due to the known but paradoxical stimulation of arginine decarboxylase activity by DFMO.

Non-NASA Center

Phycomyces in space: A problem in bioengineering

Sustaining life with total automation is a difficult problem for GAS canisters. The length of time between setting the experiment and flight, the conditions of a completely sealed container, no guarantee on launch delay, orientation and the possibility of contamination all tend to exclude experiments with living matter. This experiment examines the growth of a nontoxic, everyday fungus, Phycomyces, in a microgravity environment. Data from this experiment will help define the mechanism by which plants determine the direction of gravity. The bioengineering problems were solved only after numerous tests and design changes. Phycomyces normally have a shelf life of approximately one week. Storing the fungus for two months, activating the fungus for growth and precise timing were the major obstacles. Solutions were found for storage by drying the fungus spores onto pieces of filter paper. Activation occurs when this filter paper is dropped onto the growth medium, via a solenoid system. The problem of timing is partially solved by growing more than one chamber of the fungus at different time intervals. This experiment proves that the simpler a design, the better it works.

Schneringer, Julie A.

Effects of artificial defoliation of pines on the structure and physiology of the soil fungal community of a mixed pine-spruce forest

Loss of photosynthetic area can affect soil microbial communities by altering the availability of fixed carbon. We used denaturing gradient gel electrophoresis (DGGE) and Biolog filamentous-fungus plates to determine the effects of artificial defoliation of pines in a mixed pine-spruce forest on the composition of the fungal community in a forest soil. As measured by DGGE, two fungal species were affected significantly by the defoliation of pines (P < 0.001); the frequency of members of the ectomycorrhizal fungus genus Cenococcum decreased significantly, while the frequency of organisms of an unidentified soil fungus increased. The decrease in the amount of Cenococcum organisms may have occurred because of the formation of extensive hyphal networks by species of this genus, which require more of the carbon fixed by their host, or because this fungus is dependent upon quantitative differences in spruce root exudates. The defoliation of pines did not affect the overall composition of the soil fungal community or fungal-species richness (number of species per core). Biolog filamentous-fungus plate assays indicated a significant increase (P < 0.001) in the number of carbon substrates utilized by the soil fungi and the rate at which these substrates were used, which could indicate an increase in fungal-species richness. Thus, either small changes in the soil fungal community give rise to significant increases in physiological capabilities or PCR bias limits the reliability of the DGGE results. These data indicate that combined genetic and physiological assessments of the soil fungal community are needed to accurately assess the effect of disturbance on indigenous microbial systems.

Pinus/growth & development/physiology

Prevalence and diversity of TAL effector-like proteins in fungal endosymbiotic Mycetohabitans spp.

EndofungalMycetohabitans(formerlyBurkholderia) spp. rely on a type III secretion system to deliver mostly unidentified effector proteins when colonizing their host fungus,Rhizopus microsporus. The one known secreted effector family fromMycetohabitansconsists of homologues of transcription activator-like (TAL) effectors, which are used by plant pathogenicXanthomonasandRalstoniaspp. to activate host genes that promote disease. These ‘BurkholderiaTAL-like (Btl)’ proteins bind corresponding specific DNA sequences in a predictable manner, but their genomic target(s) and impact on transcription in the fungus are unknown. Recent phenotyping of Btl mutants of twoMycetohabitansstrains revealed that the single Btl in oneMycetohabitans endofungorumstrain enhances fungal membrane stress tolerance, while others in aMycetohabitans rhizoxinicastrain promote bacterial colonization of the fungus. The phenotypic diversity underscores the need to assess the sequence diversity and, given that sequence diversity translates to DNA targeting specificity, the functional diversity of Btl proteins. Using a dual approach to maximize capture of Btl protein sequences for our analysis, we sequenced and assembled nineMycetohabitansspp. genomes using long-read PacBio technology and also mined available short-read Illumina fungal–bacterial metagenomes. We show thatbtlgenes are present across diverseMycetohabitansstrains from Mucoromycota fungal hosts yet vary in sequences and predicted DNA binding specificity. Phylogenetic analysis revealed distinct clades of Btl proteins and suggested thatMycetohabitansmight contain more species than previously recognized. Within our data set, Btl proteins were more conserved acrossM. rhizoxinicastrains than acrossM. endofungorum, but there was also evidence of greater overall strain diversity within the latter clade. Overall, the results suggest that Btl proteins contribute to bacterial–fungal symbioses in myriad ways.

Genetics & Heredity

Study of coatings for improved fire and decay resistance of mine timbers

The purpose of this program was to find a fire- and rot-retardant polymer/fungicide reaction product for coating mine timbers. Fire-retardant polymers were screened as films and coatings on fir wood. Curable polyimide appeared to be flame retardant and evolved a minimum of fumes when exposed to a flame. Several organic and metal, low toxicity, fungicides were reacted with the polyimide in-situ on the wood. These coated samples were screened for fungus resistance. All formulations rated well - even the polyimide film without additives was fungicidal. The fir wood control itself resisted internal damage during the ten weeks of fungus exposure. A more severe test for fungus resistance will be required.

Baum, B.

Mutualism in a Reduced Gravity Environment (MuRGE)

MuRGE (Mutualism in a Reduced Gravity Environment) is a NASA flight-research experiment to investigate the microgravity effects associated with cell-cell communication and beneficial microbe-host interactions using a plant-fungal model system. This investigation will use a clinostat, an instrument that slowly rotates the plants to negate the effects of gravitational pull on plant growth (gravitropism) and development, to simulate microgravity. I will be using the endophytic fungus Piriformospora indica (Pi) and the model plant species Arabidopsis thaliana (At). P. indica has been shown to colonize roots of various plant species, including A. thaliana, and to increase plant growth and resistance to stress. The fungus has the ability to grow from spores or in axenic cultures without the presence of a host. P. indica spores and P. indica extract will be used to inoculate Arabidopsis seeds germinated on a clinostat in order to determine if simulated microgravity affects the interaction between the fungus and its plant host.

Patel, Karishma K.

[Mutualism in a Reduced Gravity Environment (MuRGE)]

MuRGE (Mutualism in a Reduced Gravity Environment) is a NASA flight-research experiment to investigate the microgravity effects associated with cell-cell communication and beneficial microbe-host interactions using a plant-fungal model system. This investigation will use a clinostat, an instrument that slowly rotates the plants to negate the effects of gravitational pull on plant growth (gravitropism) and development, to simulate microgravity. I will be using the endophytic fungus Piriformospora indica (Pi) and the model plant species Arabidopsis thaliana (At). P. indica has been shown to colonize roots of various plant species, including A. thaliana, and to increase plant growth and resistance to stress. The fungus has the ability to grow from spores or in axenic cultures without the presence of a host. P. indica spores and P. indica extract will be used to inoculate Arabidopsis seeds germinated on a clinostat in order to determine if simulated microgravity affects the interaction between the fungus and its plant host.

Patel, Karishma

Eucalyptus grandis MYB‐Like and RAN‐Like Zinc Finger Proteins Display Dual Roles in Regulating Plant Immunity and Symbiosis Pathways

Plant roots live in constant contact with diverse microbes in the soil. Plant fitness, therefore, relies on signaling pathways that mount an effective immune response against pathogens while fostering mutualistic symbioses. Plant pathways, and specifically immune genes that may act as "switches," discriminating between pathogenic or mutualistic fungi, remain largely unknown. Using Eucalyptus grandis as a model system, we investigate alterations to the root transcriptomic landscape during pre-symbiosis with either the pathogen Armillaria luteobubalina or the mutualistic fungus Pisolithus microcarpus. Comparative analyses identified three strongly counter-regulated genes that may act as immune switches to accommodate or to repress fungal colonization. We characterized two of these, a MYB-like and RAN-like zinc finger protein, using a transgenic approach and demonstrated that they have bifunctional roles in the regulation of cell death and a hypersensitive-like response, depending on the lifestyle of the associated fungus. Using co-expression network analysis, we identified hypothetical pathways correlated to these genes. We functionally validated these predictions using plants with transgenic roots with increased or decreased transcription of these genes, thereby showing the power of co-expression networks as an a priori approach to identify key immune response pathways in plants. Overall, our results demonstrate that prior to physical contact with microbes, MYB-like and RAN-like zinc finger proteins are key regulators of plant immune signaling that respond to fungal signals and enable or repress symbiotic establishment.

mycorrhizal fungi

A novel regulator of the fungal phosphate starvation response revealed by transcriptional profiling and DNA affinity purification sequencing

Cells must accurately sense and respond to nutrients to compete for resources and establish growth. Phosphate is a critical nutrient source necessary for signaling, energy metabolism, and synthesis of nucleic acids, phospholipids, and cellular metabolites. During phosphate limitation, fungi import phosphate from the environment and liberate phosphate from phosphate-containing molecules in the cell. In the model filamentous fungus Neurospora crassa, the phosphate starvation response is regulated by the conserved transcription factor NUC-1. The activity of NUC-1 is repressed by a complex of the cyclin-dependent kinase MDK-1 and the cyclin PREG when phosphate is plentiful. When phosphate is limiting, NUC-1 repression by MDK-1/PREG is relieved by the cyclin-dependent kinase inhibitor NUC-2. We investigated the global response of N. crassa to phosphate starvation. During phosphate starvation, NUC-1 directly activated the expression of genes encoding phosphatases, nucleases, and a phosphate transporter and directly repressed genes associated with the ribosome. Additionally, NUC-1 indirectly activated the expression of an uncharacterized transcription factor, which we named nuc-3. NUC-3 directly repressed the expression of genes involved in phosphate acquisition and liberation after an extended period of phosphate starvation. Additionally, NUC-3 directly repressed the expression of the cyclin-dependent kinase inhibitor nuc-2. Thus, through the combination of NUC-3 direct repression of genes in the phosphate starvation response and nuc-2, an activator of the phosphate starvation response, NUC-3 serves to act as a brake on the phosphate starvation response after an extended period of phosphate starvation. This braking mechanism could reduce transcription, a phosphate-intensive process, under conditions of extended phosphate limitation.IMPORTANCEFungi have evolved regulatory networks to respond to available nutrients. Phosphate is often a limiting nutrient for fungi that is critical for many cellular functions, including nucleic acid and phospholipid biosynthesis, cell signaling, and energy metabolism. The fungal response to phosphate limitation is important in interactions with plants and animals. We investigated the global transcriptional response to phosphate starvation and the role of a major transcriptional regulator, NUC-1, in the model filamentous fungus Neurospora crassa. Our data show that NUC-1 is a bifunctional transcription factor that directly activates phosphate acquisition genes, while directly repressing genes associated with phosphate-intensive processes. NUC-1 indirectly regulates an uncharacterized transcription factor, which we named nuc-3. NUC-3 directly represses phosphate acquisition genes and nuc-2, an activator of the phosphate starvation response, during extended periods of phosphate starvation. Thus, NUC-3 acts as a brake on the phosphate starvation response to reduce phosphate-intensive activities, like transcriptional activation, when phosphate starvation persists.

DNA affinity purification sequencing

Engineering Anti-Microbial Peptide Production in Fungi and Bioprocess Development (CRADA 502)

Crop Protection, specifically controlling agricultural diseases and pests, is essential for crop production at every agricultural scale. Biologics hold enormous potential in this equation to decrease use of more toxic pesticides. Currently, the widespread use of biologics in crop protection is severely hampered by the lack of available tools for their mass production. To ensure biologics are regulated under EPA Biopesticide regulations, the following must be true: The biopesticide is a naturally occurring substance or structurally similar and functionally identical to a naturally occurring substance with a history of exposure to humans and the environment demonstrating minimal toxicity. Since this pertains to the marketed product it is also essential that fermentation hosts have a history of exposure and safe use by humans This research focuses on developing a novel platform for the production and delivery of the cghSAMP peptide using the filamentous fungus Aspergillus niger. A. niger is a well-established industrial microorganism, widely recognized for its strong protein secretion capabilities, high-yield production of enzymes, and classification as a generally recognized as safe (GRAS) organism. By genetically modifying A. niger to express the cghSAMP peptide transgene, we aim to leverage the fungus's robust fermentation system to produce a high volume of the antimicrobial agent. This engineered system represents a potential breakthrough in sustainable disease management, offering an environmentally friendly method for localized application or large-scale production of a biopesticide designed to mitigate the effects of citrus greening disease. The main objectives of this project were: 1. Developing a high-yield and cost-efficient bioprocess to produce an AMP in a current ABF host or readily onboarded hosts. 2. Scale up the process in stirred tank reactors and produce at least 100 grams of AMP at 1- 5g/L

60 APPLIED LIFE SCIENCES

Microbiological test results of the environmental control and life support systems vapors compression distillation subsystem recycle tank components following various pretreatment protocols

Microbiological samples were collected from the recycle tank of the vapor compression distillation (VCD) subsystem of the water recovery test at NASA MSFC following a 68-day run. The recycle tank collects rejected urine brine that was pretreated with a commercially available oxidant (Oxone) and sulfuric acid and pumps it back to the processing component of the VCD. Samples collected included a water sample and two swab samples, one from the particulate filter surface and a second from material floating on the surface of the water. No bacteria were recovered from the water sample. Both swab samples contained a spore-forming bacterium, Bacillus insolitus. A filamentous fungus was isolated from the floating material. Approximately 1 month after the pretreatment chemicals were changed to sodium hypochlorite and sulfuric acid, a swab of the particulate filter was again analyzed for microbial content. One fungus was isolated, and spore-forming bacteria were observed. These results indicate the inability of these pretreatments to inhibit surface attachment. The implications of the presence of these organisms are discussed.

Huff, Tim

Chamber for Growing and Observing Fungi

A chamber has been designed to enable growth and observation of microcolonies of fungi in isolation from the external environment. Unlike prior fungus-growing apparatuses, this chamber makes it possible to examine a fungus culture without disrupting it. Partly resembling a small picture frame, the chamber includes a metal plate having a rectangular through-thethickness opening with recesses for a top and a bottom cover glass, an inlet for air, and an inlet for water. The bottom cover glass is put in place and held there by clips, then a block of nutrient medium and a moisture pad are placed in the opening. The block is inoculated, then the top cover glass is put in place and held there by clips. Once growth is evident, the chamber can be sealed with tape. Little (if any) water evaporates past the edges of the cover glasses, and, hence there is little (if any) need to add water. A microscope can be used to observe the culture through either cover glass. Because the culture is sealed in the chamber, it is safe to examine the culture without risking contamination. The chamber can be sterilized and reused.

Pierson, Duane L.

Post-Flight Microbial Analysis of Samples from the International Space Station Water Recovery System and Oxygen Generation System

The Regenerative, Environmental Control and Life Support System (ECLSS) on the International Space Station (ISS) includes the the Water Recovery System (WRS) and the Oxygen Generation System (OGS). The WRS consists of a Urine Processor Assembly (UPA) and Water Processor Assembly (WPA). This report describes microbial characterization of wastewater and surface samples collected from the WRS and OGS subsystems, returned to KSC, JSC, and MSFC on consecutive shuttle flights (STS-129 and STS-130) in 2009-10. STS-129 returned two filters that contained fluid samples from the WPA Waste Tank Orbital Recovery Unit (ORU), one from the waste tank and the other from the ISS humidity condensate. Direct count by microscopic enumeration revealed 8.38 x 104 cells per mL in the humidity condensate sample, but none of those cells were recoverable on solid agar media. In contrast, 3.32 x lOs cells per mL were measured from a surface swab of the WRS waste tank, including viable bacteria and fungi recovered after S12 days of incubation on solid agar media. Based on rDNA sequencing and phenotypic characterization, a fungus recovered from the filter was determined to be Lecythophora mutabilis. The bacterial isolate was identified by rDNA sequence data to be Methylobacterium radiotolerans. Additional UPA subsystem samples were returned on STS-130 for analysis. Both liquid and solid samples were collected from the Russian urine container (EDV), Distillation Assembly (DA) and Recycle Filter Tank Assembly (RFTA) for post-flight analysis. The bacterium Pseudomonas aeruginosa and fungus Chaetomium brasiliense were isolated from the EDV samples. No viable bacteria or fungi were recovered from RFTA brine samples (N= 6), but multiple samples (N = 11) from the DA and RFTA were found to contain fungal and bacterial cells. Many recovered cells have been identified to genus by rDNA sequencing and carbon source utilization profiling (BiOLOG Gen III). The presence of viable bacteria and fungi from WRS and OGS subsystems demonstrates the need for continued monitoring of ECLSS during future ISS operations and investigation of advanced antimicrobial controls.

Birmele, Michele N.

Investigating Meteorological Conditions Associated with Valley Fever Outbreaks in Southwestern United States

According to CDC, a sharp increase in reported Valley fever cases (Coccidioidomycosis) has been found between 2009 and 2012. Environmental conditions play an important role for Valley fever outbreaks. For example, dust storms can significantly increase the amount of the fungus Coccidioides in the air and associated strong winds can transport the fungus to other areas. In last year’s AGU session, we reported a preliminary study to investigate hydrometeorological conditions and their connection with dust storm activities in southwestern United States. We found wind is a major contributing factor for the seasonal variation of dust storm activities. Interannual variation of the regional hydrometeorological conditions are closely linked to the large-scale environment such as the Pacific Decadal Oscillation (PDO). Strong winds are linked with a number of weather events such cold front passages, thunderstorms that produce downbursts and strong winds, the Santa Ana winds, etc. In this presentation, we will report the results of our latest investigation on meteorological conditions associated with Valley fever outbreaks in southwestern United States, using NLDAS (North American Land Data Assimilation System), GPM (Global Precipitation Measurement), and MERRA-2 (Modern Era Retrospective-analysis for Research and Applications, Version 2) hourly datasets, from the NASA Goddard Earth Sciences (GES) Data and Information Services Center (DISC).

Liu, Zhong

Serendipita indica Drives Sulfur-Related Microbiota in Enhancing Growth of Hyperaccumulator Sedum alfredii and Facilitating Soil Cadmium Remediation

Endophytic fungus Serendipita indica can bolster plant growth and confer protection against various biotic and abiotic stresses. However, S. indica -reshaped rhizosphere microecology interactions and root-soil interface processes in situ at the submicrometer scale remain poorly understood. Here, we combined amplicon sequencing and high-resolution nano X-ray fluorescence (nano-XRF) imaging of the root-soil interface to reveal cadmium (Cd) rhizosphere processes. S. indica can successfully colonize the roots of Sedum alfredii Hance, which induces a remarkable increase in shoot biomass by 211.32% and Cd accumulation by 235.72%. Nano-XRF images showed that S. indica colonization altered the Cd distribution in the rhizosphere and facilitated the proximity of more Cd and sulfur (S) to enter the roots and transport to the shoot. Furthermore, the rhizosphere-enriched microbiota demonstrated a more stable network structure after the S. indica inoculation. Keystone species were strongly associated with growth promotion and Cd absorption. For example, Comamonadaceae are closely related to the organic acid cycle and S bioavailability, which could facilitate Cd and S accumulation in plants. Meanwhile, Sphingomonadaceae could release auxin and boost plant biomass. In summary, we construct a mutualism system for beneficial fungi and hyperaccumulation plants, which facilitates high-efficient remediation of Cd-contaminated soils by restructuring the rhizosphere microbiota.

60 APPLIED LIFE SCIENCES