Publications by authors named "Heather H Wilkinson"

24 Publications

  • Page 1 of 1

Neurospora crassa ASM-1 complements the conidiation defect in a stuA mutant of Aspergillus nidulans.

Mycologia 2015 Mar-Apr;107(2):298-306. Epub 2014 Dec 30.

Program for the Biology of Filamentous Fungi, Department of Plant Pathology and Microbiology, Texas A&M University, College Station, Texas 77843

Aspergillus nidulans StuA and Neurospora crassa ASM-1 are orthologous APSES (ASM-1, PHD1, SOK2, Efg1, StuA) transcription factors conserved across a diverse group of fungi. StuA and ASM-1 have roles in asexual (conidiation) and sexual (ascospore formation) development in both organisms. To address the hypothesis that the last common ancestor of these diverse fungi regulated conidiation with similar genes, asm-1 was introduced into the stuA1 mutant of A. nidulans. Expression of asm-1 complemented defective conidiophore morphology and restored conidia production to wild type levels in stuA1. Expression of asm-1 in the stuA1 strain did not rescue the defect in sexual development. When the conidiation regulator AbaA was tagged at its C-terminus with GFP in A. nidulans, it localized to nuclei in phialides. When expressed in the stuA1 mutant, AbaA::GFP localized to nuclei in conidiophores but no longer was confined to phialides, suggesting that expression of AbaA in specific cell types of the conidiophore was conditioned by StuA. Our data suggest that the function in conidiation of StuA and ASM-1 is conserved and support the view that, despite the great morphological and ontogenic diversity of their condiphores, the last common ancestor of A. nidulans and N. crassa produced an ortholog of StuA that was involved in conidiophore development.
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http://dx.doi.org/10.3852/14-079DOI Listing
August 2015

Evaluating the performance of carboxylate platform fermentations across diverse inocula originating as sediments from extreme environments.

Bioresour Technol 2014 Mar 4;155:388-94. Epub 2014 Jan 4.

Department of Plant Pathology and Microbiology, Texas A&M University, College Station, TX 77843-2132, USA. Electronic address:

To test the hypothesis that microbial communities from saline and thermal sediment environments are pre-adapted to exhibit superior fermentation performances, 501 saline and thermal samples were collected from a wide geographic range. Each sediment sample was screened as inoculum in a 30-day batch fermentation. Using multivariate statistics, the capacity of each community was assessed to determine its ability to degrade a cellulosic substrate and produce carboxylic acids in the context of the inoculum sediment chemistry. Conductance of soils was positively associated with production of particular acids, but negatively associated with conversion efficiency. In situ sediment temperature and conversion efficiency were consistently positively related. Because inoculum characteristics influence carboxylate platform productivity, optimization of the inoculum is an important and realistic goal.
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http://dx.doi.org/10.1016/j.biortech.2013.12.105DOI Listing
March 2014

Characterization of isolated yeast growth response to methionine analogs.

J Environ Sci Health B 2013 ;48(12):1112-20

Center for Food Safety, and Department of Food Science, University of Arkansas, Fayetteville, AR 72704-5690, USA.

Methionine is one of the first limiting amino acids in poultry nutrition. The use of methionine-rich natural feed ingredients, such as soybean meal or rapeseed meal may lead to negative environmental consequences. Amino acid supplementation leads to reduced use of protein-rich ingredients. The objectives of this study were isolation of potentially high content methionine-containing yeasts, quantification of methionine content in yeasts and their respective growth response to methionine analogs. Minimal medium was used as the selection medium and the isolation medium of methionine-producing yeasts from yeast collection and environmental samples, respectively. Two yeasts previously collected along with six additional strains isolated from Caucasian kefir grains, air-trapped, cantaloupe, and three soil samples could grow on minimal medium. Only two of the newly isolated strains, K1 and C1, grew in minimal medium supplied with either methionine analogs ethionine or norleucine at 0.5% (w/v). Based on large subunit rRNA sequences, these isolated strains were identified as Pichia udriavzevii/Issatchenkia orientalis. P. kudriavzevii/I. orentalis is a generally recognized as a safe organism. In addition, methionine produced by K1 and C1 yeast hydrolysate yielded 1.3 ± 0.01 and 1.1 ± 0.01 mg g(-1) dry cell. Yeast strain K1 may be suitable as a potential source of methionine for dietary supplements in organic poultry feed but may require growth conditions to further increase their methionine content.
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http://dx.doi.org/10.1080/03601234.2013.824305DOI Listing
April 2014

Comparison of three screening methods to select mixed-microbial inoculum for mixed-acid fermentations.

Bioresour Technol 2013 Feb 13;130:739-49. Epub 2012 Oct 13.

Department of Chemical Engineering, Texas A&M University, College Station, TX 78453, USA.

Using a mixed culture of microorganisms, the carboxylate platform converts biomass into hydrocarbons and chemicals. To develop a method that identifies the highest performing inoculum for carboxylate fermentations, five bacterial communities were screened and ranked by three fermentation performance tests: (1) 30-day batch screen, (2) 28-day continuum particle distribution model (CPDM), and (3) 5-month continuous countercurrent fermentation trains. To screen numerous inocula sources, these tests were used sequentially in an aseptic environment. For the batch-fermentation screen, Inoculum 1 achieved the highest conversion. For the CPDM evaluation, the operating map for Inoculum 1 had the highest performance. For the continuous countercurrent fermentation, the train resulting from Inoculum 1 was among the best performers. This study suggests that the three screens are a useful and predictive method for choosing optimal inocula sources. The bacterial community with optimal performance in these three screens could be considered for use in commercial-scale fermentations.
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http://dx.doi.org/10.1016/j.biortech.2012.10.010DOI Listing
February 2013

A microfluidic microbial fuel cell array that supports long-term multiplexed analyses of electricigens.

Lab Chip 2012 Oct;12(20):4151-9

Department of Electrical and Computer Engineering, Texas A&M University, College Station, TX, USA.

Microbial fuel cells (MFCs) are green energy technologies that exploit microbial metabolism to generate electricity. The widespread implementation of MFC technologies has been stymied by their high cost and limited power. MFC arrays in which device configurations or microbial consortia can be screened have generated significant interest because of their potential for defining aspects that will improve performance featuring high throughput characteristics. However, current miniature MFCs and MFC array systems do not support long-term studies that mimic field conditions, and hence, have limitations in fully characterizing and understanding MFC performances in varieties of conditions. Here, we describe an MFC array device that incorporates microfluidic technology to enable continuous long-term analysis of MFC performance at high throughput utilizing periodic anolyte/catholyte replenishment. The system showed 360% higher power output and 700% longer operating time when compared to MFC arrays without catholyte replenishment. We further demonstrate the utility of the system by reporting its successful use in screening microbial consortia collected from geographically diverse environments for communities that support enhanced MFC performance. Taken together, this work demonstrates that anolyte/catholyte replenishment can significantly improve the long-term performance of microfabricated MFC arrays, and support the characterization of diverse microbial consortia.
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http://dx.doi.org/10.1039/c2lc40405bDOI Listing
October 2012

Mesophilic and thermophilic conditions select for unique but highly parallel microbial communities to perform carboxylate platform biomass conversion.

PLoS One 2012 22;7(6):e39689. Epub 2012 Jun 22.

Department of Soil and Crop Sciences, Texas A&M University, College Station, Texas, United States of America.

The carboxylate platform is a flexible, cost-effective means of converting lignocellulosic materials into chemicals and liquid fuels. Although the platform's chemistry and engineering are well studied, relatively little is known about the mixed microbial communities underlying its conversion processes. In this study, we examined the metagenomes of two actively fermenting platform communities incubated under contrasting temperature conditions (mesophilic 40°C; thermophilic 55 °C), but utilizing the same inoculum and lignocellulosic feedstock. Community composition segregated by temperature. The thermophilic community harbored genes affiliated with Clostridia, Bacilli, and a Thermoanaerobacterium sp, whereas the mesophilic community metagenome was composed of genes affiliated with other Clostridia and Bacilli, Bacteriodia, γ-Proteobacteria, and Actinobacteria. Although both communities were able to metabolize cellulosic materials and shared many core functions, significant differences were detected with respect to the abundances of multiple Pfams, COGs, and enzyme families. The mesophilic metagenome was enriched in genes related to the degradation of arabinose and other hemicellulose-derived oligosaccharides, and the production of valerate and caproate. In contrast, the thermophilic community was enriched in genes related to the uptake of cellobiose and the transfer of genetic material. Functions assigned to taxonomic bins indicated that multiple community members at either temperature had the potential to degrade cellulose, cellobiose, or xylose and produce acetate, ethanol, and propionate. The results of this study suggest that both metabolic flexibility and functional redundancy contribute to the platform's ability to process lignocellulosic substrates and are likely to provide a degree of stability to the platform's fermentation processes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0039689PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3382152PMC
January 2013

Comparison of mixed-acid fermentations inoculated with six different mixed cultures.

Bioresour Technol 2012 Aug 18;118:343-9. Epub 2012 May 18.

Department of Chemical Engineering, Texas A&M University, College Station, TX 77843, USA.

The MixAlco™ process biologically converts biomass to carboxylate salts that may be converted to a variety of chemicals and fuels. This study examines the fermentation performance of six different mixed cultures, and how the performance was affected by the bacterial composition of each community. All six countercurrent fermentations had very similar performance, but were dissimilar in microbial community composition. The acid concentrations varied by only 12% between fermentation trains and the conversions varied only by 6%. The microbial communities were profiled using 16S rRNA tag-pyrosequencing, which revealed the presence of dynamic communities that were dominated by bacteria resembling Clostridia, but they shared few taxa in common. Yue-Clayton similarity calculations of the communities revealed that they were extremely different. The presence of different but functionally similar microbial communities in this study suggests that it is the operating parameters that determine the fermentation end-products.
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http://dx.doi.org/10.1016/j.biortech.2012.05.043DOI Listing
August 2012

Comparative genome sequence analysis underscores mycoparasitism as the ancestral life style of Trichoderma.

Genome Biol 2011 18;12(4):R40. Epub 2011 Apr 18.

Area Gene Technology and Applied Biochemistry, Institute of Chemical Engineering Vienna University of Technology, Getreidemarkt 9, 1060 Vienna, Austria.

Background: Mycoparasitism, a lifestyle where one fungus is parasitic on another fungus, has special relevance when the prey is a plant pathogen, providing a strategy for biological control of pests for plant protection. Probably, the most studied biocontrol agents are species of the genus Hypocrea/Trichoderma.

Results: Here we report an analysis of the genome sequences of the two biocontrol species Trichoderma atroviride (teleomorph Hypocrea atroviridis) and Trichoderma virens (formerly Gliocladium virens, teleomorph Hypocrea virens), and a comparison with Trichoderma reesei (teleomorph Hypocrea jecorina). These three Trichoderma species display a remarkable conservation of gene order (78 to 96%), and a lack of active mobile elements probably due to repeat-induced point mutation. Several gene families are expanded in the two mycoparasitic species relative to T. reesei or other ascomycetes, and are overrepresented in non-syntenic genome regions. A phylogenetic analysis shows that T. reesei and T. virens are derived relative to T. atroviride. The mycoparasitism-specific genes thus arose in a common Trichoderma ancestor but were subsequently lost in T. reesei.

Conclusions: The data offer a better understanding of mycoparasitism, and thus enforce the development of improved biocontrol strains for efficient and environmentally friendly protection of plants.
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http://dx.doi.org/10.1186/gb-2011-12-4-r40DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3218866PMC
February 2012

acon-3, the Neurospora crassa ortholog of the developmental modifier, medA, complements the conidiation defect of the Aspergillus nidulans mutant.

Fungal Genet Biol 2011 Apr 8;48(4):370-6. Epub 2011 Jan 8.

Program for the Biology of Filamentous Fungi, Department of Plant Pathology and Microbiology, Texas A&M University, College Station, TX, USA.

Aspergillus nidulans and Neurospora crassa are ascomycetes that produce asexual spores through morphologically distinct processes. MedA, a protein with unknown function, is required for normal asexual and sexual development in A. nidulans. We determined that the N. crassa ortholog of medA is acon-3, a gene required for early conidiophore development and female fertility. To test hypotheses about the evolutionary origins of asexual development in distinct fungal lineages it is important to understand the degree of conservation of developmental regulators. The amino acid sequences of A. nidulans MedA and N. crassa ACON-3 shared 37% identity and 51% similarity. acon-3 is induced at late time points of conidiation. In contrast, medA is constitutively expressed and MedA protein localizes to nuclei in all tissue types. Nonetheless, expression of acon-3 using its native promoter complemented the conidiation defects of the A. nidulans ΔmedA and medA15 mutants. We conclude that the biochemical activity of the medA orthologs is conserved for conidiation.
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http://dx.doi.org/10.1016/j.fgb.2010.12.008DOI Listing
April 2011

Temporal and spatial regulation of gene expression during asexual development of Neurospora crassa.

Genetics 2010 Dec 27;186(4):1217-30. Epub 2010 Sep 27.

Department of Plant Pathology and Microbiology, Texas A&M University, College Station, TX 77843-2132, USA.

In this study we profiled spatial and temporal transcriptional changes during asexual sporulation in the filamentous fungus Neurospora crassa. Aerial tissue was separated from the mycelium to allow detection of genes specific to each tissue. We identified 2641 genes that were differentially expressed during development, which represents ∼25% of the predicted genes in the genome of this model fungus. On the basis of the distribution of functional annotations of 1102 of these genes, we identified gene expression patterns that define key physiological events during conidial development. Not surprisingly, genes encoding transcription factors, cell wall remodeling proteins, and proteins involved in signal transduction were differentially regulated during asexual development. Among the genes differentially expressed in aerial tissues the majority were unclassified and tended to be unique to ascomycete genomes. This finding is consistent with the view that these genes evolved for asexual development in the Pezizomycotina. Strains containing deletions of several differentially expressed genes encoding transcription factors exhibited asexual development-associated phenotypes. Gene expression patterns during asexual development suggested that cAMP signaling plays a critical role in the transition from aerial growth to proconidial chain formation. This observation prompted us to characterize a deletion of the gene encoding a high-affinity cAMP phosphodiesterase (NCU00478). NCU00478 was determined to be allelic to aconidiate-2, a previously identified genetic locus controlling conidiation.
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http://dx.doi.org/10.1534/genetics.110.121780DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2998306PMC
December 2010

Structure and dynamics of the microbial communities underlying the carboxylate platform for biofuel production.

Appl Microbiol Biotechnol 2010 Sep 31;88(1):389-99. Epub 2010 Jul 31.

Department of Soil and Crop Sciences, Texas A&M University, 370 Olsen Blvd, TAMU 2474, College Station, TX 77843-2474, USA.

The carboxylate platform utilizes a mixed microbial community to convert lignocellulosic biomass into chemicals and fuels. While much of the platform is well understood, little is known about its microbiology. Mesophilic (40 degrees C) and thermophilic (55 degrees C) fermentations employing a sorghum feedstock and marine sediment inoculum were profiled using 16S rRNA tag-pyrosequencing over the course of a 30-day incubation. The contrasting fermentation temperatures converted similar amounts of biomass, but the mesophilic community was significantly more productive, and the two temperatures differed significantly with respect to propionic and butyric acid production. Pyrotag sequencing revealed the presence of dynamic communities that responded rapidly to temperature and changed substantially over time. Both temperatures were dominated by bacteria resembling Clostridia, but they shared few taxa in common. The species-rich mesophilic community harbored a variety of Bacteroidetes, Actinobacteria, and gamma-Proteobacteria, whereas the thermophilic community was composed mainly of Clostridia and Bacilli. Despite differences in composition and productivity, similar patterns of functional class dynamics were observed. Over time, organisms resembling known cellulose degraders decreased in abundance, while organisms resembling known xylose degraders increased. Improved understanding of the carboxylate platform's microbiology will help refine platform performance and contribute to our growing knowledge regarding biomass conversion and biofuel production processes.
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http://dx.doi.org/10.1007/s00253-010-2789-7DOI Listing
September 2010

Shifts in microbial community structure along an ecological gradient of hypersaline soils and sediments.

ISME J 2010 Jun 4;4(6):829-38. Epub 2010 Feb 4.

Department of Soil and Crop Sciences, Texas A&M University, College Station, TX 77843-2474, USA.

Studies of hypersaline ecosystems often yield novel organisms and contribute to our understanding of extreme environments. Soils and sediments from La Sal del Rey, a previously uncharacterized, hypersaline lake located in southern Texas, USA, were surveyed to characterize the structure and diversity of their microbial communities. Samples were collected along a transect that spanned vegetated uplands, exposed lakebed sediments, and water-logged locations, capturing a wide range of environments and physical and chemical gradients. Community quantitative PCR (qPCR) was used in combination with tag-encoded pyrosequencing, 16S rRNA gene cloning, and Sanger sequencing to characterize the lake's soil and sediment microbial communities. Further, we used multivariate statistics to identify the relationships shared between sequence diversity and heterogeneity in the soil environment. The overall microbial communities were surprisingly diverse, harboring a wide variety of taxa, and sharing significant correlations with site water content, phosphorus and total organic carbon concentrations, and pH. Some individual populations, especially of Archaea, also correlated with sodium concentration and electrical conductivity salinity. Across the transect, Bacteria were numerically dominant relative to Archaea, and among them, three phyla--the Proteobacteria, Bacteroidetes, and Firmicutes--accounted for the majority of taxa detected. Although these taxa were detected with similar abundances to those described in other hypersaline ecosystems, the greater depth of sequencing achieved here resulted in the detection of taxa not described previously in hypersaline sediments. The results of this study provide new information regarding a previously uncharacterized ecosystem and show the value of high-throughput sequencing in the study of complex ecosystems.
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http://dx.doi.org/10.1038/ismej.2010.3DOI Listing
June 2010

Comparison of loline alkaloid gene clusters across fungal endophytes: predicting the co-regulatory sequence motifs and the evolutionary history.

Fungal Genet Biol 2007 Oct 13;44(10):1002-10. Epub 2007 Apr 13.

Department of Plant Pathology and Microbiology, Texas A&M University (2132), College Station, TX 77843-2132, USA.

LOL, a fungal secondary metabolite gene cluster found in Epichloë and Neotyphodium species, is responsible for production of insecticidal loline alkaloids. To analyze the genetic architecture and to predict the evolutionary history of LOL, we compared five clusters from four fungal species (single clusters from Epichloë festucae, Neotyphodium sp. PauTG-1, Neotyphodium coenophialum, and two clusters we previously characterized in Neotyphodium uncinatum). Using PhyloCon to compare putative lol gene promoter regions, we have identified four motifs conserved across the lol genes in all five clusters. Each motif has significant similarity to known fungal transcription factor binding sites in the TRANSFAC database. Conservation of these motifs is further support for the hypothesis that the lol genes are co-regulated. Interestingly, the history of asexual Neotyphodium spp. includes multiple interspecific hybridization events. Comparing clusters from three Neotyphodium species and E. festucae allowed us to determine which Epichloë ancestors are the most likely contributors of LOL in these asexual species. For example, while no present day Epichloë typhina isolates are known to produce lolines, our data support the hypothesis that the E. typhina ancestor(s) of three asexual endophyte species contained a LOL gene cluster. Thus, these data support a model of evolution in which the polymorphism in loline alkaloid production phenotypes among endophyte species is likely due to the loss of the trait over time.
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http://dx.doi.org/10.1016/j.fgb.2007.04.003DOI Listing
October 2007

Identification and quantification of methanogenic Archaea in adult chicken ceca.

Appl Environ Microbiol 2007 Jan 3;73(1):353-6. Epub 2006 Nov 3.

University of Arkansas, Department of Food Science, 2650 N. Young Avenue, Fayetteville, AR 72704-5690, USA.

By using molecular methods for the identification and quantification of methanogenic archaea in adult chicken ceca, 16S rRNA genes of 11 different phylotypes, 10 of which were 99% similar to Methanobrevibacter woesei, were found. Methanogen populations, as assessed by cultivation, and the 16S rRNA copy number were between 6.38 and 8.23 cells/g (wet weight) and 5.50 and 7.19 log(10)/g (wet weight), respectively.
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http://dx.doi.org/10.1128/AEM.01931-06DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1797138PMC
January 2007

FSR1 is essential for virulence and female fertility in Fusarium verticillioides and F. graminearum.

Mol Plant Microbe Interact 2006 Jul;19(7):725-33

Department of Plant Pathology and Microbiology, Program for the Biology of Filamentous Fungi, Texas A&M University, College Station 77843-2132, USA.

Fusarium verticillioides (teleomorph Gibberella moniliformis) and F. graminearum (teleomorph G. zeae) are well known to cause devastating diseases on cereal crops. Despite their importance, our understanding of the molecular mechanisms involved in these host-pathogen interactions is limited. The FSR1 locus in F. verticillioides was identified by screening REMI mutants for loss of virulence in maize stalk rot inoculation studies. FSR1 encodes an 823-codon open reading frame interrupted by two introns. The Fsr1 protein shares 60% sequence identity with the Sordaria macrospora Pro11, a multimodular protein with four putative protein-protein binding domains (caveolin-binding domain, coiled-coil structure, calmodulin-binding motif, and seven-WD40 repeats), which plays a regulatory role in cell differentiation and ascocarp development. Our data demonstrate that FSR1 is essential for female fertility and virulence in F. verticillioides. Significantly, targeted disruption of the FSR1 ortholog in F. graminearum (FgFSR1) reduced virulence on barley and deterred perithecia formation. Cross-complementation experiments demonstrated that the gene function is conserved in the two Fusarium species. FSR1 is expressed constitutively, and we hypothesize that Fsr1 regulates virulence by acting as a scaffold for a signal transduction pathway. A survey of available genome databases indicates Fsr1 homologs are present in a number of filamentous fungi and animal systems but not in budding yeast or plants. A maximum likelihood analysis of this gene family reveals well-supported monophyletic clades associated with fungi and animals.
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http://dx.doi.org/10.1094/MPMI-19-0725DOI Listing
July 2006

A mitogen-activated protein kinase pathway essential for mating and contributing to vegetative growth in Neurospora crassa.

Genetics 2005 Jul 31;170(3):1091-104. Epub 2005 Mar 31.

Program for the Biology of Filamentous Fungi, Department of Plant Pathology and Microbiology, Texas A&M University, College Station, Texas 77843, USA.

MAP kinases homologous to Saccharomyces cerevisiae Fus3p/Kss1p have been identified in plant pathogenic fungi and are required for pathogenicity and sexual reproduction. To better understand the role of MAP kinase signaling in Neurospora crassa, and to identify downstream target genes of the pathway, we isolated, cloned, and disrupted the FUS3 homolog mak-2. Ste12p is a transcription factor target of Fus3p that activates genes of the mating pathway in yeast, and we also characterized the N. crassa STE12 homolog pp-1. The mak-2 and pp-1 mutants have reduced growth rate, produce short aerial hyphae, and fail to develop protoperithecia. In addition, ascospores carrying null mutations of either gene are inviable. Subtractive cloning was used to isolate genes having reduced expression in the mak-2 mutant. Expression of some of these genes is protoperithecia specific and three of them are part of a gene cluster potentially involved in the production of a polyketide secondary metabolite. Microarray analysis was used to extend the analysis of gene expression in mak-2 and pp-1 mutants. The role of the MAP kinase pathway in both sexual and asexual development as well as secondary metabolism is consistent with the dual regulation of the mating process and pathogencity observed in fungal pathogens.
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http://dx.doi.org/10.1534/genetics.104.036772DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1451179PMC
July 2005

Gene clusters for insecticidal loline alkaloids in the grass-endophytic fungus Neotyphodium uncinatum.

Genetics 2005 Mar 16;169(3):1403-14. Epub 2005 Jan 16.

Department of Plant Pathology, University of Kentucky, Lexington, 40546-0312, USA.

Loline alkaloids are produced by mutualistic fungi symbiotic with grasses, and they protect the host plants from insects. Here we identify in the fungal symbiont, Neotyphodium uncinatum, two homologous gene clusters (LOL-1 and LOL-2) associated with loline-alkaloid production. Nine genes were identified in a 25-kb region of LOL-1 and designated (in order) lolF-1, lolC-1, lolD-1, lolO-1, lolA-1, lolU-1, lolP-1, lolT-1, and lolE-1. LOL-2 contained the homologs lolC-2 through lolE-2 in the same order and orientation. Also identified was lolF-2, but its possible linkage with either cluster was undetermined. Most lol genes were regulated in N. uncinatum and N. coenophialum, and all were expressed concomitantly with loline-alkaloid biosynthesis. A lolC-2 RNA-interference (RNAi) construct was introduced into N. uncinatum, and in two independent transformants, RNAi significantly decreased lolC expression (P < 0.01) and loline-alkaloid accumulation in culture (P < 0.001) compared to vector-only controls, indicating involvement of lolC in biosynthesis of lolines. The predicted LolU protein has a DNA-binding site signature, and the relationships of other lol-gene products indicate that the pathway has evolved from various different primary and secondary biosynthesis pathways.
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http://dx.doi.org/10.1534/genetics.104.035972DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1449547PMC
March 2005

Transcriptional response to glucose starvation and functional analysis of a glucose transporter of Neurospora crassa.

Fungal Genet Biol 2004 Dec;41(12):1104-19

Program for the Biology of Filamentous Fungi, Department of Plant Pathology and Microbiology, Texas A & M University, College Station, TX 77843, USA.

The response to glucose availability in Neurospora crassa was characterized by measuring transcript levels of 1335 genes represented on a cDNA microarray. We found that 19% of the 1335 genes were regulated at least twofold in response to glucose deprivation. Several sugar transporter homologs were found to be regulated by glucose. Functional analysis of one of these, hgt-1, indicates that it encodes a high-affinity glucose transporter. Comparing N. crassa profiling data with the published diauxic shift data from Saccharomyces cerevisiae revealed that the two fungi share similar, but not identical, transcriptional response patterns. Regulation of transcript levels measured by microarray analysis is consistent with previous measurements of central metabolism enzyme activities for N. crassa. Transcripts of genes for ethanol fermentation and the tricarboxylic acid cycle are abundant in glucose-rich medium, consistent with N. crassa carrying out a high rate of both fermentation and respiration simultaneously.
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http://dx.doi.org/10.1016/j.fgb.2004.08.009DOI Listing
December 2004

Increased conidiation associated with progression along the sterigmatocystin biosynthetic pathway.

Mycologia 2004 Nov-Dec;96(6):1190-8

Department of Plant Pathology and Microbiology, Texas A&M University, College Station, Texas 77845-2132.

The Aspergillus nidulans sterigmatocystin (ST) gene cluster contains both regulatory (aflR) and biosynthetic genes (stc genes) required for ST production. A total of 26 genes are in the cluster, 13 of which have been assigned a known function in the biosynthetic pathway. This complex secondary pathway represents a physiological cost to the fungus. We tested the amount of asexual spore production using a series of isogenic lines of A. nidulans, differing only in a mutation in aflR (resulting in a strain containing no ST intermediates) or a mutation in three stc genes that produced either no ST intermediates (ΔstcJ), an early ST intermediate, norsoloroinic acid (ΔstcE) or a late ST intermediate, versicolorin A (ΔstcU). In two independently replicated experiments we compared the numbers of conidia produced by each of these mutant strains and a wild type ST producer in a neutral (growth media) and a host (corn seed) environment. A stepwise increase in asexual spore production was observed with each progressive step in the ST pathway. Thus, the data suggest that recruitment or loss of these secondary metabolite pathway genes has a selective advantage apart from the physiological activity of the metabolite.
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October 2012

Contig assembly and microsynteny analysis using a bacterial artificial chromosome library for Epichloë festucae, a mutualistic fungal endophyte of grasses.

Fungal Genet Biol 2004 Jan;41(1):23-32

Department of Plant Pathology and Microbiology, Texas A and M University, College Station, TX 77845-2132, USA.

We constructed and characterized a bacterial artificial chromosome (BAC) library for Epichloë festucae, a genetically tractable fungal plant mutualist. The 6144 clone library with an average insert size of 87kb represents at least 18-fold coverage of the 29 Mb genome. We used the library to assemble a 110kb contig spanning the putative ornithine decarboxylase (odc) ortholog and subsequently expanded it to 228kb with a single walking step in each direction. Furthermore, we evaluated conservation of microsynteny between E. festucae and some model filamentous fungi by comparing sequence available from a 43kb region at the end of one BAC to publicly available fungal genome sequences. Orthologs to the 13 contiguous open reading frames (ORFs) identified in E. festucae are syntenic in Neurospora crassa and Magnaporthe grisea occurring in small sets of two, three or four colinear ORFs. This library is a valuable resource for research into traits important for the development and maintenance of a plant-fungus mutualistic symbiosis.
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http://dx.doi.org/10.1016/j.fgb.2003.08.004DOI Listing
January 2004

Expressed sequence tags and genes associated with loline alkaloid expression by the fungal endophyte Neotyphodium uncinatum.

Fungal Genet Biol 2002 Aug;36(3):242-54

Department of Plant Pathology, University of Kentucky, Lexington, KY 40546-0091, USA.

Loline alkaloids (LA), which are 1-aminopyrrolizidines with an oxygen bridge, are produced by Epichloë (anamorph=Neotyphodium) species, endophytes of grasses. LA are insecticidal, thus, helping to protect host plants from insect herbivory. The objective of this study was to identify genes associated with LA biosynthesis. Suppression subtractive hybridization PCR was used to isolate transcripts up-regulated during LA production in cultures of Neotyphodium uncinatum. Subtracted cDNAs were cloned and a lambda-phage cDNA library from an LA-expressing N. uncinatum culture was screened with subtracted cDNA. In BLAST searches, several cDNAs identified had sequence similarities to aspartate kinases and another with O-acetylhomoserine-(thiol)lyase. Differential expression of these two genes in LA-producing cultures of N. uncinatum was confirmed and in a survey of 23 isolates from 21 Neotyphodium and Epichloë species these two genes strictly correlated with LA production. These findings open up the possibility for detailed studies on genes and enzymes involved in LA production.
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http://dx.doi.org/10.1016/s1087-1845(02)00023-3DOI Listing
August 2002

Symbiotic specialization and the potential for genotypic coexistence in a plant-bacterial mutualism.

Oecologia 1996 Oct;108(2):361-367

Department of Biological Sciences, State University of New York, 13902, Binghamton, NY, USA.

Genotypes of the annual legume Amphicarpaea bracteata vary in their degree of specialization toward different nitrogen-fixing bacteria. Plants of lineages "Ib" and "II" are specialized for mutualism with a limited group of bacterial genotypes. Lineage "Ia" plants are symbiotic generalists, interacting with all bacteria associated with specialist (Ib and II) plants, and also with a distinct class of bacteria that only fix nitrogen with lineage Ia plants. The relative performance of generalist and specialist plant lineages was measured in four symbiotic environments: (1) in the absence of nitrogen-fixing bacteria, (2) with bacteria with broad host ranges, (3) with bacteria specialized on lineage Ia hosts, and (4) with a mixture of the bacteria with broad and narrow host ranges. In the presence of bacteria with broad host ranges, the relative performance of different plant lineages was inconsistent among experimental replicates. However, lineage Ia plants had nearly 3 times higher total biomass and 6 times higher seed biomass than lineage Ib or II plants when grown with bacteria specialized on Ia hosts. When exposed to a mixture of bacteria with broad and narrow host ranges, generalist plants had 72% higher total biomass and >100% higher seed biomass than specialist plants. These results imply that in diverse natural populations, where all plants have a choice of symbiotic partners, mutualistic interactions are likely to foster competitive exclusion rather than stable coexistence of different plant lineages.
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http://dx.doi.org/10.1007/BF00334662DOI Listing
October 1996

DIVERGENCE IN SYMBIOTIC COMPATIBILITY IN A LEGUME-BRADYRHIZOBIUM MUTUALISM.

Evolution 1996 Aug;50(4):1470-1477

Department of Biological Sciences, State University of New York, Binghamton, New York, 13902-6000.

Geographic variation in the mutualism between the legume Amphicarpaea bracteata and its nitrogen-fixing root nodule bacteria (Bradyrhizobium sp.) was analyzed by sampling genotypes from 11 sites separated by distances ranging from 0.6 km to more than 1000 km. Cross inoculation experiments revealed that plants were genetically differentiated in traits determining compatibility with mutualist partners from different sites. Combinations of plant and bacterial genotypes native to the same local habitat yielded 26% higher plant growth relative to non-native combinations (range across 4 experiments; 9% to 48%). Among non-native symbioses, plant growth was unrelated to the geographic distance between sites of plant and bacterial origin. However, compatibility varied significantly with the genetic distance among host populations (inferred by multilocus enzyme electrophoresis): genetically similar plants from separate sites showed superior growth with each other's mutualist partners. Nevertheless, the tree structure of population genetic similarity was not congruent in plants versus bacteria. This implies that adaptive variation in symbiotic compatibility has evolved without strictly parallel divergence in the two species.
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http://dx.doi.org/10.1111/j.1558-5646.1996.tb03920.xDOI Listing
August 1996

NONRANDOM GENOTYPIC ASSOCIATIONS IN A LEGUME-BRADYRHIZOBIUM MUTUALISM.

Evolution 1996 Feb;50(1):146-154

Department of Biological Sciences, State University of New York, Binghamton, New York, 13902-6000.

Genetically divergent lineages often coexist within populations of the annual legume Amphicarpaea bracteata. At one site dominated by two such lineages (termed biotypes "C" and "S"), isolates of root-nodule bacteria (Bradyrhizobium sp.) were sampled from both hosts and analyzed by enzyme electrophoresis. Symbiont populations on the two plant biotypes were highly distinct. Out of 15 bacterial multilocus genotypes detected (among 51 isolates analyzed), only one was shared in common by the two plant biotypes. Cluster analysis revealed three bacterial lineages (designated I, II, and III), with lineage I found exclusively on biotype C plants, and the two other lineages almost completely restricted to biotype S hosts. Laboratory inoculation tests indicated that lineage I bacteria were strictly specialized on biotype C hosts, forming few or no nodules on plants of the other host biotype. Bacterial lineages II and III were capable of forming nodules on both kinds of plants, but nodule numbers were often significantly higher on biotype S hosts. The nonrandom association between plant and bacterial lineages at this site implies that genetic diversity of hosts is an important factor in the maintenance of polymorphism within the symbiont population.
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http://dx.doi.org/10.1111/j.1558-5646.1996.tb04481.xDOI Listing
February 1996
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