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Climate change is associated with increasing challenges for the wild and agricultural ecosystemson which humans depend. In our most diverse forests and in agricultural lands, rising temperatures, shifts in precipitation regimes, and growing pressure from pathogens and pests represent major challenges that will impact the supply of food, fuel, fiber, and ecosystem services worldwide. Although the effects of climate change on plant communities have been studied previously, their particular effects on plant-microbe symbionts are less understood. Plant symbioses with microbes, especially microscopic fungi, are vital for plant resilience, productivity, and survival in all ecosystems. In my dissertation I examine how climate change impacts aspects of plant-fungal associations, with a focus on two main environments: wild tropical forests, which are storehouses of biodiversity; and agricultural systems, the immediate sources of the plant products we need for human use. Specifically, my work focuses on understanding (1) how hurricane disturbances affect the diversity and composition of fungal symbionts associated with roots and leaves of tropical forest trees, with a focus on Puerto Rico; (2) how fungal symbionts can protect cultivated plants against heat stress and disease under a rapidly warming climate, with a focus on Lactuca sativa, an important crop in Arizona; and (3) how gaps in knowledge about plant pathogens merit attention in areas where tropical and agricultural environments meet, and climate change is felt in all sectors, as in the island of Puerto Rico. In quantifying effects of hurricane damage on fungal symbionts of tropical trees, I used a combination of fieldwork, next-generation sequencing, and statistical approaches to study fungal endophytes (fungi that live within healthy plant tissues without causing disease). I found a significant relationship between foliar endophyte richness and hurricane damage, with evidence that severe damage to forests is associ

Aflatoxins are cancer-causing, immuno-suppressive mycotoxins that frequently contaminate important staples in Zambia including maize and groundnut. Managing aflatoxins begins with understanding the distribution of aflatoxins across the target region. Seventeen percent of crops from markets contained aflatoxin concentrations above allowable levels in Zambia, with the frequency of contamination in groundnut and maize highest in warmest regions of the country. Proper management of aflatoxin contamination requires a clear understanding of the etiologic agents of the observed contamination. Several species within Aspergillus section Flavi have been implicated as causal agents of aflatoxin contamination in Africa. In Zambia, A. parasiticus was the main etiologic agent of aflatoxin contamination of maize and groundnut, although fungi with S morphology also caused contamination. Aspergillus flavus L morphotype fungi were associated with reduced aflatoxins, suggesting natural biological control by atoxigenic strains may reduce aflatoxin contamination in Zambia. In addition to maize and groundnut, wild insects, fruits and fish are important sources of food and incomes in Zambia. Unfortunately, both insects and wild plants are susceptible to aflatoxin contamination. To evaluate the safety of wild insects and fruit, concentrations of aflatoxins and presence of aflatoxin-producers were assessed. Some species of wild fruits and insects were found to have unsafe levels of aflatoxins suggesting mitigation efforts should target these important foods of Zambia in addition to crops such as groundnut and maize. New lineages of aflatoxin-producing fungi have been described, and found associated with cases of aflatoxicoses in Kenya and elsewhere. Although A. parasiticus is highly frequent and an important etiologic agent of aflatoxin contamination, it is not known how this fungus is related to similar fungi elsewhere. A multigene phylogenetic analysis revealed at least two new groups div

The cacao swollen shoot disease of Theobroma cacao L. (cacao) is caused by Cacao swollen shoot virus (CSSV; genus, Badnavirus, family, Caulimoviridae). The virus is endemic to West Africa, where it poses a serious threat to cocoa production. Despite efforts to control CSSV spread by replacement of infected trees with tolerant cultivars and mealybug vector management, the disease is widespread in West Africa. In Trinidad, leaf mosaic and vein-banding symptoms have been observed in cacao plants in the field since the 1940s, and recently at the International Cocoa Genebank (ICGT), a custodian of cacao germplasm resources. The strains A and B of the suspect Cacao Trinidad virus (CTV) caused the symptoms, and were thought to be related to CSSV, however, viral causality was not demonstrated, until now. To develop molecular detection methods for CSSV in infected plants, polymerase chain reaction (PCR) amplification of eight regions of the CSSV genome was implemented. The PCR results showed variable amplification frequencies of 19 - 42% at each region, for 124 isolates collected in Cote d'Ivoire and Ghana. Pairwise nucleotide (nt) analyses of the eight regions showed 66-99% shared identities, indicating that CSSV isolates exhibit extensive variability with respect to primer design. The results provided preliminary evidence for the existence of a CSSV complex consisting of four divergent species. The full length genome of 14 CSSV isolates from cacao determined using the Illumina HiSeq platform showed 70-99% shared nt identities. The pairwise nt identities placed CSSV sequences into a group of four distinct species, one of which represented a previously undescribed species. Moreover, the full-length genomes grouped phylogenetically with other badnaviruses and revealed two CSSV subclades with three types of genome arrangements; four, five or six open reading frames (ORFs). Predicted functional protein domains were conserved on each ORF. Two distinct, full-length genome sequenc

Background Alternaria is considered one of the most common saprophytic fungal genera on the planet. It is comprised of many species that exhibit a necrotrophic phytopathogenic lifestyle. Several species are clinically associated with allergic respiratory disorders although rarely found to cause invasive infections in humans. Finally, Alternaria spp. are among the most well known producers of diverse fungal secondary metabolites, especially toxins. Description We have recently sequenced and annotated the genomes of 25 Alternaria spp. including but not limited to many necrotrophic plant pathogens such as A. brassicicola (a pathogen of Brassicaceous crops like cabbage and canola) and A. solani (a major pathogen of Solanaceous plants like potato and tomato), and several saprophytes that cause allergy in human such as A. alternata isolates. These genomes were annotated and compared. Multiple genetic differences were found in the context of plant and human pathogenicity, notably the pro-inflammatory potential of A. alternata. The Alternaria genomes database was built to provide a public platform to access the whole genome sequences, genome annotations, and comparative genomics data of these species. Genome annotation and comparison were performed using a pipeline that integrated multiple computational and comparative genomics tools. Alternaria genome sequences together with their annotation and comparison data were ported to Ensembl database schemas using a self-developed tool (EnsImport). Collectively, data are currently hosted using a customized installation of the Ensembl genome browser platform. Conclusion Recent efforts in fungal genome sequencing have facilitated the studies of the molecular basis of fungal pathogenicity as a whole system. The Alternaria genomes database provides a comprehensive resource of genomics and comparative data of an important saprophytic and plant/human pathogenic fungal genus. The database will be updated regularly with new genomes when t

Aflatoxins are carcinogenic mycotoxins most frequently associated with the filamentous fungus, Aspergillus flavus. These potent toxins pose serious health threats and their concentrations in foods are widely regulated. Maize, a critical staple of billions, is frequently contaminated with aflatoxins. Development of commercial maize hybrids with superior resistance to aflatoxin contamination has been sought for over 30 years without success. Analyses of native maize land races (MLRs) from Mexico revealed several accessions with significant resistance to both aflatoxin contamination and fungal reproduction. Physical barriers are important components of MLRs resistance. Traditional use of MLRs may reduce human exposure to aflatoxins. MLRs may contribute resistant genes of significant value in breeding for aflatoxin resistance. In Mexico, maize is produced from<10 to over 2,000 masl. Elevation had only minor influence on community compositions of aflatoxin-producing fungi associated with maize over three years in Sonora, Mexico. Most variation in community structure occurred between years. Dominant vegetative compatibility groups (VCGs) differed among years but were detected in similar frequencies across four agroecological zones. Multiple locations and multiple years must be sampled to obtain realistic assessments of the most successful VCGs. Elevated frequencies of VCG YV150 throughout Sonora during 2006 led to investigate genetic diversity within this VCG using microsatellite loci. The 2006 increases were attributed to rapid increase of a single clone. Examination of YV150 isolates collected over 20 years in the US and Mexico revealed several haplotypes and two genetically distinct populations, which were composed of isolates containing only a population specific mating-type idiomorph. Microsatellite loci in each population were in gametic equilibrium. Gene flow between isolates with different idiomorphs was not detected. VCG YV36, to which the biocontrol agent AF36 b

Research regarding the distribution and structure of fungal communities in caves is lacking. The current study examines fungal communities in Kartchner Caverns, a mineralogically diverse cave located in the Whetstone Mountains, Arizona, USA. The first study examines culturable fungal diversity from speleothem surfaces. Twenty-one fungal genera represented by 43 genotypes and 53 distinct morphological taxonomic units (MTU) were recovered from 15 speleothems. Analysis of DGGE profiles indicated a significant effect of sampling site on community structure. The second study examined fungal diversity from speleothem and rock wall surfaces using the 454 FLX Titanium sequencing platform using the rDNA internal transcribed spacer 1 (ITS1) as a genetic marker. Fungal diversity was estimated and compared between speleothem and rock wall surfaces and its variation with distance from the natural entrance of the cave was quantified. Effects of environmental factors and nutrient concentrations in speleothem drip water at different sample sites on fungal diversity were also examined. Sequencing revealed 2219 fungal operational taxonomic units (OTUs) at 95% similarity. Speleothems supported a higher fungal richness and diversity than rock walls, but community membership and the taxonomic distribution of fungal OTUs did not differ significantly. OTU richness and diversity were negatively correlated with distance from the natural cave entrance. Community membership and taxonomic distribution of fungal OTUs differed significantly between the front and back of the cave. There was no observed effect of drip water nutrient concentration on fungal community structure. The third study examined fungal community structure from bat guano over the course of a year. There was no significant difference in fungal OTU richness, diversity, or community membership and taxonomic affiliations among sampling times. There were no significant differences in nutrient concentrations of guano samples among

The hypothesis of horizontal gene transfer of a hybrid gene from bacteria to the filamentous Ascomycota was tested using constrained phylogenetic analyses and tests of topological congruence. Results suggest that the hybrid gene was acquired from bacteria by a single transfer before the radiation of the Leotiomyceta. The phylogenetic relationship among Nimbya and Embellisia was investigated using both morphological and molecular data. Examination of conidia morphology revealed that Nimbya and Embellisia comprise two and four distinct morphological groups, respectively. Molecular analyses support all morphological groups of Nimbya and Embellisia and reveal that both genera are polyphyletic. Results suggest the circumscription of these genera is based upon convergent morphological characters. To further understand the evolutionary relationship among Alternaria and closely related genera, ten protein-coding genes were sequenced across 176 species. Three genes possessed significant substitution saturation and two other genes did not possess sufficient phylogenetic signal to assess relationships among the asexual Alternaria. The remaining five loci revealed strong support for asexual Alternaria and the order of divergence among eight asexual Alternaria species-groups. The current polyphyly of Alternaria was resolved using morphological and molecular data. Morphological examination revealed that most members of the infectoria clade (sexual Alternaria) produce diagnostic colony characters on dichloran rose bengal yeast extract sucrose agar (DRYES) and weak potato dextrose agar (WPDA) that are fundamentally different as compared to other small-spored Alternaria species. These data also revealed that all members of the infectoria clade produce arachnoid vegetative hyphae with multiple primary conidiophores, whereas other small-spored Alternaria species do not. Phylogenetic analyses revealed that the sexual infectoria clade clusters with other sexual genera phylogenetically d

This work encompasses studies on the development of biocontrol strategies to manage the disease lettuce drop, caused by the fungi Sclerotinia sclerotiorum and S. minor, using the mycoparasitic fungus Coniothyrium minitans, and to better understand interactions during mycoparasitism at the biochemical level. Results from field experiments revealed that two applications of C. minitans at manufacturer recommended rates significantly reduced the incidence of lettuce drop caused by S. sclerotiorum but not by S. minor. Applications of other biocontrol products tested did not significantly reduce disease incidence caused by either pathogen. Sclerotium population studies revealed that soil populations of S. sclerotiorum in lettuce production fields ranged from 0.08 to 2.9 sclerotia/100g of soil and were generally aggregated in their distribution. Continued field studies revealed that there was no significant effect of irrigation (sprinkler vs furrow) on either the impact of sclerotium density or the efficacy of C. minitans. Studies on the evaluation of different application rates of Contans against S. minor revealed that two applications of Contans at 5 X manufactures recommended rates significantly reduced the disease incidence. Examination of sclerotial exudates of Sclerotinia spp. revealed that crude exudates from both Sclerotinia spp. stimulated C. minitans spore germination and the stimulation was due to compounds within the polar fraction. Studies on the role of lectin-carbohydrate binding during fungal-mycoparasite interactions revealed that many plant lectins as well as crude proteins extracted from sclerotia of either Sclerotinia spp. induced agglutination of C. minitans spores in vitro. Spore germination of C. minitans stimulated by sclerotial exudates of either Sclerotinia spp. was significantly inhibited in the presence of the lectin Con A but not other plant lectins. In vitro studies on the directional growth of C. minitans preceding mycoparasitism revealed the

Phytostabilization is an emerging technology for the remediation of mine tailings sites. In arid and semiarid environments, mine tailings disposal sites are a major source of environmental pollution as they are subject to eolian dispersion and water erosion. Mine tailings are acidic to neutral, high in metal content, and nutrient poor. Furthermore, these sites remain unvegetated even after decades of no additional mining activity. In arid and semiarid regions, climatic variables such as high winds, salinity, and drought exacerbate the problem. The Klondyke mine tailings site is a model site for studying plant establishment in mine tailings within semiarid regions. It was a lead and zinc ore- processing operation from 1948 to 1958 and is similar in physicochemical characteristics to other acidic pyritic mine tailings.In a greenhouse study, a native drought tolerant halophyte, Atriplex lentiformis (Torr.) S. Wats., was evaluated for its potential as a phytostabilization candidate in compost-amended tailings from the Klondyke site. Germination, plant growth, and metal uptake of A. lentiformis were examined, and the microbial community was monitored by enumeration of autotrophic iron- and sulfur-oxidizing bacteria as well as heterotrophic bacteria. Results demonstrated that with 10 to 15% compost addition, growth of A. lentiformis was not affected and shoot metal concentrations were generally not a concern for foraging animals. Furthermore, the heterotrophic bacterial community is severely stressed but recovers with compost addition and successful plant growth. Therefore, A. lentiformis is a good candidate for phytostabilization of mine tailings with compost amendments.Poor revegetation of mine tailings has been attributed to the microbial community involved in acidifying tailings; however, no thorough microbial studies have been conducted. The second study characterizes the bacterial community of the Klondyke site and compares it to an offsite control sample. Results d