Marc Joel Orbach

Interests

Research

Mechanisms of fungal pathogenesis; Genetics and genomics of fungi;

Teaching

Fungal Biology and Fungal Genetics, Host-fungal interactions

Courses

Scholarly Contributions

Chapters

• Dean, R. A., Mitchell, T. K., Kulkarni, R., Kulkarni, R., Donofrio, N. M., Powell, A. J., Oh, Y., Diener, S. E., Pan, H., Brown, D., Deng, J., Carbone, I., Ebbole, D. J., Thon, M. R., Farman, M. L., Orbach, M. J., Soderlund, C., Soderlund, C., Xu, J., , Xu, J., et al. (2008).

  The rice blast story: From genome sequence to function

  . In Exploitation of Fungi. Cambridge University Press. doi:10.1017/CBO9780511902451.003
• Kang, S., Mullins, E., Dezwaan, T. M., & Orbach, M. J. (2000).

  Pathogenesis and Genome Organization of the Rice Blast Fungus

  . In Fungal Pathology. Springer, Dordrecht. doi:10.1007/978-94-015-9546-9_8
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  Plant diseases caused by fungi present a serious challenge to our efforts to feed the world’s rapidly increasing population. Manipulation of plant genes through genetic engineering and traditional breeding provides an opportunity to improve the disease resistance in crops of agricultural significance. In addition, the application of target-based drug design will lead to the identification of various novel chemical agents that will possess significant potential as disease control agents. The success of such approaches will depend heavily on the biological knowledge of fungal pathogens in such areas as mechanisms of virulence and pathogenicity, population structure and dynamics, and the nature and mechanisms of genetic changes underpinning the evolution of new races. As a result of limited resources, coupled with the necessity to study many fungal diseases of economic significance, comprehensive research focused on these aspects of fungal biology has been restricted to a limited number of model systems. Magnaporthe grisea, the causal agent for rice blast disease, has become one such system. Worldwide, rice blast is one of the most economically devastating crop diseases. In addition to its agricultural significance, the rice blast system presents many advantages as an experimental model: (a) Extensive genetic and molecular analyses of host specificity and pathogenicity have been carried out; (b) Molecular and cellular bases of infection processes are well characterized; (c) Rice is a model genetic system for monocots; (d) Well-saturated genetic and physical maps of M. grisea are available; (e) Genome sequencing of both rice and M. grisea is in progress; (f) Genetic and race diversity of fungal populations in many rice-growing areas has been extensively surveyed. In this chapter we aim to provide an overview of the nature of the interaction between M. grisea and its hosts and the corresponding mechanisms employed at each level, from molecules to populations.
• Dioh, W., Lebrun, M., Tharreau, D., Gomez, R., Roumen, E., Notteghem, J., Orbach, M. J., Orbach, M. J., & Gomez, R. L. (1996).

  Mapping avirulence genes in the rice blast fungus Magnaporthe Grisea

  . In Rice Genetics III Part 2. IRRI. doi:https://doi.org/10.1142/9789812814289_0128
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  L'agent pathogene Magnaporthe grisea est implique dans des interactions specifiques avec les cultivars de riz. Les souches du champignon avec les genes d'avirulence ne sont pas pathogenes pour les cultivars de riz ayant les genes de resistance specifiques correspondants. Demeler la base moleculaire de telles interactions demande le clonage des genes d'avirulence et de resistance. Nous avons identifie trois genes d'avirulence genetiquement independants chez M. grisea. L'analyse de la segregation de l'avirulence a travers une collection de cultivars de riz suggere l'occurence du meme gene de resistance dans les cultivars non associes. Pour cloner les genes d'avirulence par arpentage chromosomique, nous avons construit une carte genetique partielle en utilisant 77 descendances aleatoires issues d'un croisement entre des isolats Guy11 et 2/0/3. Cette carte inclut 75 marqueurs RFLP et 25 marqueurs RAPD. Deux genes d'avirulence ont ete cartographies sur l'extremite des chromosomes. Nous avons aussi identifie 5 marqueurs RAPD lies etroitement aux genes d'avirulence en utilisant une analyse de segregation en melange. La plupart de ces marqueurs RAPD lies etaient composes de sequences repetees adjacentes aux sequences simples, qui pourraient etre utilisees pour lancer l'arpentage chromosomique
• Chumley, F. G., Valent, B., Orbach, M. J., Sweigard, J. A., Farrall, L., & Walter, A. (1991).

  Repeated DNA Sequences and the Analysis of Host Specificity in the Rice Blast Fungus

  . In Plant Molecular Biology. Springer, Boston, MA. doi:10.1007/978-1-4615-3304-7_17
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  Field isolates of the heterothallic Ascomycete, Magnaporthe grisea Barr (anamorph, Pyricularia oryzae Cav. or P. grisea), include pathogens of many grasses, with rice (Oryza sativa) being the most important example from an economic perspective. Individual field isolates have a limited host range, parasitizing one or a few grass species (MACKILL and BONMAN 1986). We have recently shown that rice pathogens from around the world contain a family of repeated DNA sequences, MGR sequences, that appear to be absent from or present in low copy number in field isolates that infect grasses other than rice (HAMER et al. 1989a). We have hypothesized that the correlation between MGR sequence conservation and rice pathogenicity is due to genetic isolation and independent evolution of rice pathogens descended from a small ancestral population. As we will discuss, MGR sequences have become useful as tools for strain identification and as genetic markers for cloning host specificity genes of interest.

Journals/Publications

• Cotty, P. J., Orbach, M. J., Ching’anda, C., Atehnkeng, J., Bandyopadhyay, R., Callicott, K., & Mehl, H. L. (2022).

  Spatial and Temporal Population Dynamics of Aspergillus flavus in Commercial Pistachio Orchards in Arizona

  . Plant Health Progress, 23(3), 272-280. doi:10.1094/php-10-21-0128-rs
• Maxwell, L. A., Bandyopadhyay, R., Orbach, M. J., Callicott, K. A., & Cotty, P. J. (2019). Degradation: A hidden mechanism of biocontrol by atoxigenic Aspergillus flavus. Applied and Environmental Microbiology.
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  Paper was rejected on 12/28/19. Under revision for submission to another journal. Interestingly, only one review was obtained. They did not like Dr. Cotty.
• Singh, P., Callicott, K. A., Orbach, M. J., & Cotty, P. J. (2019). Phenotypic differentiation of two morphologically similar aflatoxin producers from West Africa. Applied and Environmental Microbiology.
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  This paper was reviewed and rejected by AEM. Under revision for submission to TOXINS. Expect submission in Spring 2020.
• Singh, P., Callicott, K. A., Orbach, M. J., & Cotty, P. J. (2020). Diversity among S morphology fungi in Aspergillus section Flavi from the United States. targeted for Fungal Diversity.
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  Under internal review. We expect to submit this paper to Fungal Diversity in the beginning of March 2020.
• Maxwell, L. A., Callicott, K. A., Bandyopadhyay, R., Mehl, H. L., Orbach, M. J., & Cotty, P. J. (2021). Degradation of aflatoxins B1 by atoxigenic Aspergillus flavus biocontrol agents. Plant Disease, 105(9).
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  This paper by Lourena Arone Maxwell, who was one of Dr. Orbach's PhD students, was selected as the Editor's pick by Plant Disease for the month of September
• Shubitz, L. F., Powell, D. A., Butkiewicz, C. D., Lewis, M. L., Trinh, H. T., Frelinger, J. A., Orbach, M. J., & Galgiani, J. N. (2021). A Chronic Murine Disease Model of Coccidioidomycosis Using Coccidioides posadasii, Strain 1038. The Journal of Infectious Diseases, 223(1), 166-173.
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  Murine infections with most Coccidioides spp. strains are lethal by 3 weeks, limiting the study of immune responses. Coccidioides posadasii, strain 1038 (Cp1038), while slowly lethal, resulted in protracted survival of C57BL/6 (B6) mice. In resistant (B6D2)F1/J mice, lung fungal burdens stabilized by week 4 without progression through week 16, better modeling human coccidioidal infections after their immunologic control. Immunodeficient tumor necrosis factor (Tnf) α knockout (KO) and interferon (Ifn) γ receptor 1 (Ifn-γr1) KO mice survived a median of 22.5 and 34 days, compared with 70 days in B6 mice (P = .001 and P < .01, respectively), though 14-day lung fungal burden studies showed little difference between Ifn-γr1 KO and B6 mice. B6 mice showed peak concentrations of key inflammatory lung cytokines, including interleukin 6, 23, and 17A, Tnf-α, and Ifn-γ, only after 4 weeks of infection. The slower progression in B6 and the acquired fungal burden stability in B6D2 mice after Cp1038 infection greatly increases the array of possible immunologic studies.
• Shubitz, L. F., Robb, E. J., Powell, D. A., Bowen, R. A., Bosco-Lauth, A. .., Hartwig, A., Porter, S. M., Trinh, H., Moale, H., Bielefeldt-Ohmann, H. .., Hoskinson, J., Orbach, M. J., Frelinger, J. A., & Galgiani, J. N. (2021). Δcps1 vaccine protects dogs against experimentally induced coccidioidomycosis. Vaccine, 39(47), 6894-6901.
• Izquierdo, A., Jaffey, J. A., Szabo, S., Struthers, J., Okwumabua, O., Hostnik, E. T., Ohkura, M., Trinh, H., Shubitz, L. F., Orbach, M. J., & White, M. E. (2020). in a Dog With Cervical Dissemination Complicated by Esophageal Fistula. Frontiers in veterinary science, 7, 285.
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  A 5-year-old male, neutered mixed breed dog with a history of a mass with an associated draining tract on the ventral cervical region was diagnosed with an esophageal fistula. The dog exhibited serosanguinous discharge from the draining tract, with enlarged left superficial cervical and mandibular lymph nodes, and was reported to have difficulty with deglutition of solid foods. Computed tomography revealed a communication of the draining tract with the esophagus along with enlargement of the left lateral retropharyngeal, left medial retropharyngeal, and mandibular lymph nodes. This prompted surgical exploration and debridement of the site, with closure of the esophageal fistula. Histopathology of thyroid gland, skeletal muscle, and adipose tissue obtained during surgical exploration showed spherules consistent with . infection. Antibody titers performed post-operatively were consistent with an active spp. Infection. By fungal culture and subsequent PCR and DNA sequencing, was identified as the species infecting the dog. Over the course of 85 days of antifungal therapy, discharge from the draining tract, lymphadenomegaly, and cutaneous and subcutaneous nodules resolved. In conclusion, this is the first reported case of disseminated coccidioidomycosis to the cervical region of a dog with involvement of the thyroid gland, skeletal muscle, adipose tissue, connective tissue, and secondary esophageal fistula. spp. infections should be considered a differential diagnosis in unusual cases for dogs that live in or have traveled to endemic areas.
• Mead, H. L., Hamm, P. S., Shaffer, I. N., Teixeira, M. M., Wendel, C. S., Wiederhold, N. P., Thompson, G. R., Muñiz-Salazar, R., Castañón-Olivares, L. R., Keim, P., Plude, C., Terriquez, J., Galgiani, J. N., Orbach, M. J., & Barker, B. M. (2020). Differential Thermotolerance Adaptation between Species of. Journal of fungi (Basel, Switzerland), 6(4).
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  Coccidioidomycosis, or Valley fever, is caused by two species of dimorphic fungi. Based on molecular phylogenetic evidence, the genus contains two reciprocally monophyletic species: and . However, phenotypic variation between species has not been deeply investigated. We therefore explored differences in growth rate under various conditions. A collection of 39 C. posadasii and 46 C. immitis isolates, representing the full geographical range of the two species, was screened for mycelial growth rate at 37 °C and 28 °C on solid media. The radial growth rate was measured for 16 days on yeast extract agar. A linear mixed effect model was used to compare the growth rate of and at 37 °C and 28 °C, respectively. grew significantly faster at 37 °C, when compared to ; whereas both species had similar growth rates at 28 °C. These results indicate thermotolerance differs between these two species. As the ecological niche has not been well-described for spp., and disease variability between species has not been shown, the evolutionary pressure underlying the adaptation is unclear. However, this research reveals the first significant phenotypic difference between the two species that directly applies to ecological research.
• Singh, P., Callicott, K. A., Orbach, M. J., & Cotty, P. J. (2020). Molecular Analysis of S-morphology Aflatoxin Producers From the United States Reveals Previously Unknown Diversity and Two New Taxa. Frontiers in microbiology, 11, 1236.
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  Aflatoxins are highly toxic carcinogens that detrimentally influence profitability of agriculture and the health of humans and domestic animals. Several phylogenetically distinct fungi within section have S-morphology (average sclerotial size < 400 μm), and consistently produce high concentrations of aflatoxins in crops. S-morphology fungi have been implicated as important etiologic agents of aflatoxin contamination in the United States (US), but little is known about the diversity of these fungi. The current study characterized S-morphology fungi ( = 494) collected between 2002 and 2017, from soil and maize samples, in US regions where aflatoxin contamination is a perennial problem. Phylogenetic analyses based on sequences of the calmodulin (1.9 kb) and nitrate reductase (2.1 kb) genes resolved S-morphology isolates from the US into four distinct clades: (1) S-morphotype (89.7%); (2) sp. nov. (2.4%); (3) (2.2%); and (4) sp. nov. (5.7%). All four S-morphology species produced high concentrations of aflatoxins in maize at 25, 30, and 35°C, but only the S-morphotype produced unacceptable aflatoxin concentrations at 40°C. Genetic typing of S isolates using 17 simple sequence repeat markers revealed high genetic diversity, with 202 haplotypes from 443 isolates. Knowledge of the occurrence of distinct species and haplotypes of S-morphology fungi that are highly aflatoxigenic under a range of environmental conditions may provide insights into the etiology, epidemiology, and management of aflatoxin contamination in North America.
• Singh, P., Mehl, H. L., Orbach, M. J., Callicott, K. A., & Cotty, P. J. (2020). Phenotypic Differentiation of Two Morphologically Similar Aflatoxin-Producing Fungi from West Africa. Toxins, 12(10).
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  Aflatoxins (AF) are hepatocarcinogenic metabolites produced by several species. Crop infection by these species results in aflatoxin contamination of cereals, nuts, and spices. Etiology of aflatoxin contamination is complicated by mixed infections of multiple species with similar morphology and aflatoxin profiles. The current study investigates variation in aflatoxin production between two morphologically similar species that co-exist in West Africa, and . Consistent distinctions in aflatoxin production during liquid fermentation were discovered between these species. The two species produced similar concentrations of AFB in defined media with either urea or ammonium as the sole nitrogen source. However, production of both AFB and AFG were inhibited ( < 0.001) for in a yeast extract medium with sucrose. Although production of AFG by both species was similar in urea, produced greater concentrations of AFG in ammonium ( = 0.039). Based on these differences, a reliable and convenient assay for differentiating the two species was designed. This assay will be useful for identifying specific etiologic agents of aflatoxin contamination episodes in West Africa and other regions where the two species are sympatric, especially when phylogenetic analyses based on multiple gene segments are not practical.
• Donovan, F. M., Shubitz, L., Powell, D., Orbach, M., Frelinger, J., & Galgiani, J. N. (2019). Early Events in Coccidioidomycosis. Clinical microbiology reviews, 33(1).
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  SUMMARYSince its description nearly 130 years ago, hundreds of studies have deepened our understanding of coccidioidomycosis, also known as valley fever (VF), and provided useful diagnostic tests and treatments for the disease caused by the dimorphic fungi spp. In general, most of the literature has addressed well-established infections and has described patients who have experienced major complications. In contrast, little attention has been given to the earliest consequences of the pathogen-host interaction and its implications for disease manifestation, progression, and resolution. The purpose of this review is to highlight published studies on early coccidioidomycosis, identify gaps in our knowledge, and suggest new or former research areas that might be or remain fertile ground for insight into the early stages of this invasive fungal disease.
• Ohkura, M., Cotty, P., & Orbach, M. J. (2018). Comparative genomics of Aspergillus flavus S and L morphotypes yield insights to niche adaptation. G3 Journal (Genes, Genomes, Genetics).
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  This is an article from Mana's main chapter in her thesis. It was accepted in G3: Genes, Genomes, Genetics in Fall 2018 and was selected as the Featured Article in the December 2018 issue.
• Orbach, M. J., Galgiani, J. N., Trinh, H. T., Shubitz, L. F., Powell, D. A., Orbach, M. J., Lewis, M. L., & Frelinger, J. A. (2018).

  Viable spores of Coccidioides posadasii Δcps1 are required for vaccination and provide long lasting immunity.

  . Vaccine, 36(23), 3375-3380. doi:10.1016/j.vaccine.2018.04.026
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  Coccidioidomycosis is a systemic fungal infection for which a vaccine has been sought for over fifty years. The avirulent Coccidioides posadasii strain, Δcps1, which is missing a 6 kb gene, showed significant protection in mice. These studies explore conditions of protection in mice and elucidate the immune response. Mice were vaccinated with different doses and viability states of Δcps1 spores, challenged with virulent C. posadasii, and sacrificed at various endpoints, dependent on experimental objectives. Tissues from vaccinated mice were harvested for in vitro elucidation of immune response. Vaccination with viable Δcps1 spores was required for protection from lethal challenge. Viable spore vaccination produced durable immunity, lasting at least 6 months, and prolonged survival (≥6 months). The C. posadasii vaccine strain also protected mice against C. immitis (survival ≥ 6 months). Cytokines from infected lungs of vaccinated mice in the first four days after Cp challenge showed significant increases of IFN-γ, as did stimulated CD4+ spleen cells from vaccinated mice. Transfer of CD4+ cells, but not CD8+ or B cells, reduced fungal burdens following challenge. IFN-γ from CD4+ cells in vaccinated mice indicates a Th1 response, which is critical for host control of coccidioidomycosis.
• Shubitz, L. F., Trinh, H. T., Powell, D. A., Lewis, M. L., Orbach, M. J., Frelinger, J. A., & Galgiani, J. N. (2017). Viable spores of Coccidioides posadasii delta-cps1 are required for vaccination and provide long lasting immunity. Vaccine.
• Singh, P., Orbach, M. J., & Cotty, P. J. (2018). Aspergillus texensis: A Novel Aflatoxin Producer with S Morphology from the United States. TOXINS, 10(12).
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  In my sudden role as Pummi's major advisor starting in August 2018, I helped guide the completion of this work and assisted in the writing, editing, submission and revision of this paper.
• Ohkura, M., Fitak, R. R., Wisecaver, J. H., DeBlasio, D., Niazi, F., Egholm, M., Rounsley, S. D., Kodira, C. D., & Orbach, M. J. (2017). Genome Sequence of Ophidiomyces ophiodiicola, an Emerging Fungal Pathogen of Snakes. Genome announcements, 5(30).
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  Ophidiomyces ophiodiicola, which belongs to the order Onygenales, is an emerging fungal pathogen of snakes in the United States. This study reports the 21.9-Mb genome sequence of an isolate of this reptilian pathogen obtained from a black racer snake in Pennsylvania.
• Ohkura, M., Worley, J. J., Hughes-Hallett, J. E., Fisher, J. S., Love, B. C., Arnold, A. E., & Orbach, M. J. (2016). OPHIDIOMYCES OPHIODIICOLA ON A CAPTIVE BLACK RACER (COLUBER CONSTRICTOR) AND A GARTER SNAKE (THAMNOPHIS SIRTALIS) IN PENNSYLVANIA. JOURNAL OF ZOO AND WILDLIFE MEDICINE, 47(1), 341-346.
• Orbach, M. J., Mandel, M. A., Shubitz, L. F., Narra, H. P., Galgiani, J. N., Frelinger, J. A., Trinh, H., Griffin, K., & Buntzman, A. S. (2016). A Coccidioides posadasii CPS1 Deletion Mutant Is Avirulent and Protects Mice from Lethal Infection. Infection and Immunity, 84(10), 3007-3016. doi:doi:10.1128/IAI.00633-16
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  This paper describes our discovery and demonstration that the cps1 mutant of Coccidioides functions as an attenuated vaccine against valley fever in mice.
• Xie, P., Wei, G., Taghavi, S., Rensing, C., Orbach, M. J., Hao, X., & Alwathnani, H. A. (2014).

  Phytoremediation of heavy and transition metals aided by legume-rhizobia symbiosis.

  . International journal of phytoremediation, 16(2), 179-202. doi:10.1080/15226514.2013.773273
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  Legumes are important for nitrogen cycling in the environment and agriculture due to the ability of nitrogen fixation by rhizobia. In this review, we introduce an important and potential role of legume-rhizobia symbiosis in aiding phytoremediation of some metal contaminated soils as various legumes have been found to be the dominant plant species in metal contaminated areas. Resistant rhizobia used for phytoremediation could act on metals directly by chelation, precipitation, transformation, biosorption and accumulation. Moreover, the plant growth promoting (PGP) traits of rhizobia including nitrogen fixation, phosphorus solubilization, phytohormone synthesis, siderophore release, and production of ACC deaminase and the volatile compounds of acetoin and 2, 3-butanediol may facilitate legume growth while lessening metal toxicity. The benefits of using legumes inoculated with naturally resistant rhizobia or recombinant rhizobia with enhanced resistance, as well as co-inoculation with other plant growth promoting bacteria (PGPB) are discussed. However, the legume-rhizobia symbiosis appears to be sensitive to metals, and the effect of metal toxicity on the interaction between legumes and rhizobia is not clear. Therefore, to obtain the maximum benefits from legumes assisted by rhizobia for phytoremediation of metals, it is critical to have a good understanding of interactions between PGP traits, the symbiotic plant-rhizobia relationship and metals.
• Barker, B. M., Orbach, M. J., Galgiani, J. N., Orbach, M. J., Nix, D. E., Nguyen, C., Hoover, S. E., Galgiani, J. N., Frelinger, J. A., Barker, B. M., & Ampel, N. M. (2013).

  Recent advances in our understanding of the environmental, epidemiological, immunological, and clinical dimensions of coccidioidomycosis.

  . Clinical Microbiology Reviews, 26(3), 505-25. doi:10.1128/cmr.00005-13
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  Coccidioidomycosis is the endemic mycosis caused by the fungal pathogens Coccidioides immitis and C. posadasii. This review is a summary of the recent advances that have been made in the understanding of this pathogen, including its mycology, genetics, and niche in the environment. Updates on the epidemiology of the organism emphasize that it is a continuing, significant problem in areas of endemicity. For a variety of reasons, the number of reported coccidioidal infections has increased dramatically over the past decade. While continual improvements in the fields of organ transplantation and management of autoimmune disorders and patients with HIV have led to dilemmas with concurrent infection with coccidioidomycosis, they have also led to advances in the understanding of the human immune response to infection. There have been some advances in therapeutics with the increased use of newer azoles. Lastly, there is an overview of the ongoing search for a preventative vaccine.
• Barker, B. M., Tabor, J. A., Shubitz, L. F., Perrill, R., & Orbach, M. J. (2012). Detection and phylogenetic analysis of Coccidioides posadasii in Arizona soil samples. FUNGAL ECOLOGY, 5(2), 163-176.
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  Reports of coccidioidomycosis are on the rise in the southwestern US. However, the ecology of the pathogen, Coccidioides, remains obscure and there is limited knowledge of the environmental antecedents of disease outbreaks. Detection of the fungus in the environment remains a critical challenge to modeling the source of disease. Using BALB/c mice as a biosensor, 8.9% of soils analyzed from the Tucson area (Pima County, Arizona) were found to contain the pathogen. The genotypes of 66 Coccidioides strains, recovered from 11 soils, were determined with diagnostic microsatellite loci. Comparison of these genotypes to clinical isolates revealed all were Coccidioides posadasii and they grouped with Arizona isolates. Among sites where multiple strains were recovered, two indicated a clonal population, while others yielded a diversity of genotypes. A secondary goal of this research was to assess applicability of PCR, with its potential for high-throughput screening, as a method for identifying Coccidioides-containing soils. (C) 2011 Elsevier Ltd and The British Mycological Society. All rights reserved.
• Kour, A., Greer, K., Valent, B., Orbach, M. J., & Soderlund, C. (2012). MGOS: Development of a Community Annotation Database for Magnaporthe oryzae. MOLECULAR PLANT-MICROBE INTERACTIONS, 25(3), 271-278.
• Neafsey, D. E., Barker, B. M., Sharpton, T. J., Stajich, J. E., Park, D. J., Whiston, E., Hung, C., McMahan, C., White, J., Sykes, S., Heiman, D., Young, S., Zeng, Q., Abouelleil, A., Aftuck, L., Bessette, D., Brown, A., FitzGerald, M., Lui, A., , Macdonald, J. P., et al. (2010). Population genomic sequencing of Coccidioides fungi reveals recent hybridization and transposon control. GENOME RESEARCH, 20(7), 938-946.
• Sharpton, T. J., Stajich, J. E., Rounsley, S. D., Gardner, M. J., Wortman, J. R., Jordar, V. S., Maiti, R., Kodira, C. D., Neafsey, D. E., Zeng, Q., Hung, C., McMahan, C., Muszewska, A., Grynberg, M., Mandel, M. A., Kellner, E. M., Barker, B. M., Galgiani, J. N., Orbach, M. J., , Kirkland, T. N., et al. (2009). Comparative genomic analyses of the human fungal pathogens Coccidioides and their relatives. GENOME RESEARCH, 19(10), 1722-1731.
• Lambou, K., Tharreau, D., Kohler, A., Sirven, C., Marguerettaz, M., Barbisan, C., Sexton, A. C., Kellner, E. M., Martin, F., Howlett, B. J., Orbach, M. J., & Lebrun, M. (2008). Fungi have three tetraspanin families with distinct functions. BMC GENOMICS, 9.
• Barker, B. M., Jewell, K. A., Kroken, S., & Orbach, M. J. (2007). The population biology of Coccidioides - Epidemiologic implications for disease outbreaks. COCCIDIOIDOMYCOSIS: SIXTH INTERNATIONAL SYMPOSIUM, 1111, 147-163.
• Betts, M. F., Tucker, S. L., Galadima, N., Meng, Y., Patel, G., Li, L., Donofrio, N., Floyd, A., Nolin, S., Brown, D., Mandel, M. A., Mitchell, T. K., Xu, J., Dean, R. A., Farman, M. L., & Orbach, M. J. (2007). Development of a high throughput transformation system for insertional mutagenesis in Magnaporthe oryzae. FUNGAL GENETICS AND BIOLOGY, 44(10), 1035-1049.
• Li, L., Ding, S., Sharon, A., Orbach, M., & Xu, J. (2007). Mir1 is highly upregulated and localized to nuclei during infectious hyphal growth in the rice blast fungus. MOLECULAR PLANT-MICROBE INTERACTIONS, 20(4), 448-458.
• Li, L., Schmelz, M., Kellner, E. M., Galgiani, J. N., & Orbach, M. J. (2007). Nuclear labeling of Coccidioides posadasii with green fluorescent protein. COCCIDIOIDOMYCOSIS: SIXTH INTERNATIONAL SYMPOSIUM, 1111, 198-207.
• Mandel, M. A., Barker, B. M., Kroken, S., Rounsley, S. D., & Orbach, M. J. (2007). Genomic and population analyses of the mating type loci in Coccidioides species reveal evidence for sexual reproduction and gene acquisition. EUKARYOTIC CELL, 6(7), 1189-1199.
• Meng, Y., Patel, G., Heist, M., Betts, M. F., Tucker, S. L., Galadima, N., Donofrio, N. M., Brown, D., Mitchell, T. K., Li, L., Xu, J., Orbach, M., Thon, M., Dean, R. A., & Farman, M. L. (2007). A systematic analysis of T-DNA insertion events in Magnaporthe oryzae. FUNGAL GENETICS AND BIOLOGY, 44(10), 1050-1064.
• Tucker, S. L., & Orbach, M. J. (2007). Agrobacterium-mediated transformation to create an insertion library in Magnaporthe grisea.. Methods in molecular biology (Clifton, N.J.), 354, 57-68. doi:10.1385/1-59259-966-4:57
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  Magnaporthe grisea is the causal agent of rice blast disease and represents a model organism for the study of fungal plant-pathogen interactions. Pathogenicity is a complex phenotype, which is carefully orchestrated by the fungus and begins with recognition and infection of the host plant, followed by growth within the plant cells, and finally dissemination to the next host and continuation of the fungal life cycle. Certain genes must condition the ability of a pathogenic fungus to infect and cause disease symptoms. To learn more about the infection process and the genes that are involved in the complex interplay between M. grisea and rice, we used an insertional mutagenesis approach to attempt to randomly disrupt all genes in the fungal genome. Two transformation approaches were used to build a library of insertion strains in M. grisea. Polyethylene glycol/CaCl2-mediated protoplast transformation was the initial method we used and resulted in the generation of just more than 17,000 insertion strain lines. Later Agrobacterium tumefaciens-mediated transformation was adopted and the final number of insertional mutant strains of M. grisea strain 70-15 generated was more than 57,000. Here, we describe the methods used for A. tumefaciens-mediated transformation of M. grisea and the optimized protocols we have developed to enable high-throughput fungal transformation.
• Mandel, M. A., Galgiani, J. N., Kroken, S., & Orbach, M. J. (2006). Coccidioides posadasii contains single chitin synthase genes corresponding to classes I to VII. FUNGAL GENETICS AND BIOLOGY, 43(11), 775-788.
• Orbach, M. J., & Turgeon, B. G. (2006). The XXIII Fungal Genetics Conference, march 15-20, 2005, Asilomar Conference Grounds, Pacific Grove, California. FUNGAL GENETICS AND BIOLOGY, 43(10), 669-678.
• Orsborn, K. I., Shubitz, L. F., Peng, T., Kellner, E. M., Orbach, M. J., Haynes, P. A., & Galgiani, J. N. (2006). Protein expression profiling of Coccidioides posadasii by two-dimensional differential in-gel electrophoresis and evaluation of a newly recognized peroxisomal matrix protein as a recombinant vaccine candidate. INFECTION AND IMMUNITY, 74(3), 1865-1872.
• Park, G., Xue, C., Zhao, X., Kim, Y., Orbach, M., & Xu, J. (2006). Multiple upstream signals converge on the adaptor protein Mst50 in Magnaporthe grisea. PLANT CELL, 18(10), 2822-2835.
• Soderlund, C., Haller, K., Pampanwar, V., Ebbole, D., Farman, M., Orbach, M. J., Wang, G., Wing, R., Xu, J., Brown, D., Mitchell, T., & Dean, R. (2006). MGOS: A resource for studying Magnaporthe grisea and Oryza sativa interactions. MOLECULAR PLANT-MICROBE INTERACTIONS, 19(10), 1055-1061.
• Dean, R. A., Talbot, N. J., Ebbole, D. J., Farman, M. L., Mitchell, T. K., Orbach, M. J., Thon, M., Kulkarni, R., Xu, J., Pan, H., Read, N. D., Lee, Y., Carbone, I., Brown, D., Yeon, Y. O., Donofrio, N., Jun, S. J., Soanes, D. M., Djonovic, S., , Kolomlots, E., et al. (2005). The genome sequence of the rice blast fungus Magnaporthe grisea. Nature, 434(7036), 980-986.
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  PMID: 15846337;Abstract: Magnaporthe grisea is the most destructive pathogen of rice worldwide and the principal model organism for elucidating the molecular basis of fungal disease of plants. Here, we report the draft sequence of the M. grisea genome. Analysis of the gene set provides an insight into the adaptations required by a fungus to cause disease. The genome encodes a large and diverse set of secreted proteins, including those defined by unusual carbohydrate-binding domains. This fungus also possesses an expanded family of G-protein-coupled receptors, several new virulence-associated genes and large suites of enzymes involved in secondary metabolism. Consistent with a role in fungal pathogenesis, the expression of several of these genes is upregulated during the early stages of infection-related development. The M. grisea genome has been subject to invasion and proliferation of active transposable elements, reflecting the clonal nature of this fungus imposed by widespread rice cultivation.
• Donofrio, N., Rajagopalon, R., Brown, D., Diener, S., Windham, D., Nolin, S., Floyd, A., Mitchell, T., Galadima, N., Tucker, S., Orbach, M. J., Patel, G., Farman, M., Pampanwar, V., Soderlund, C., Lee, Y. H., & Dean, R. A. (2005). 'PACLIMS': A component LIM system for high-throughput functional genomic analysis. BMC BIOINFORMATICS, 6.
• Kellner, E. M., Orsborn, K. I., Siegel, E. M., Mandel, M. A., Orbach, M. J., & Galgiani, J. N. (2005). Coccidioides posadasii contains a single 1,3-beta-glucan synthase gene that a appears to be essential for growth. EUKARYOTIC CELL, 4(1), 111-120.
• Tucker, S. L., Figueroa, M., Galadima, N., Soderlund, C., Meng, Y., Farman, M. L., Li, L., Xu, J. -., Donofrio, N., Mitchell, T. K., Dean, R. A., & Orbach, M. J. (2004). A genomics approach to pathogenicity: Saturation insertional mutagenesis in Magnaporthe grisea. PHYTOPATHOLOGY, 94(6), S135-S135.
• Galagan, J. E., Calvo, S. E., Borkovich, K. A., Selker, E. U., Read, N. D., Jaffe, D., FitzHugh, W., Ma, L. J., Smirnov, S., Purcell, S., Rehman, B., Elkins, T., Engels, R., Wang, S. G., Nielsen, C. B., Butler, J., Endrizzi, M., Qui, D. Y., Ianakiev, P., , Pedersen, D. B., et al. (2003). The genome sequence of the filamentous fungus Neurospora crassa. NATURE, 422(6934), 859-868.
• Hamer, L., Pan, H., Adachi, K., Orbach, M. J., Page, A., Ramamurthy, L., & Woessner, J. P. (2001). Regions of microsynteny in Magnaporthe grisea and Neurospora crassa. Fungal Genetics and Biology, 33(2), 137-143.
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  PMID: 11456466;Abstract: A bacterial artificial chromosome (BAC) clone containing 110,467 bp of genomic DNA from Magnaporthe grisea was sequenced, annotated, and compared to the genomes of Neurospora crassa, Candida albicans, and Saccharomyces cerevisiae. Twenty-six open reading frames (ORFs), involved in multiple biochemical pathways, were identified in the BAC sequence. A region of 53 kb, containing 18 of the 26 ORFs, was found to be syntenic to a portion of the N. crassa genome. Subregions of complete colinearity as well as interrupted colinearity were present. No synteny was evident with either C. albicans or S. cerevisiae. The identification of syntenic regions containing highly conserved genes across two genera that have been evolutionarily separated for ∼200 million years elicits many biological questions as to the function and identity of these genes. © 2001 Academic Press.
• Kelkar, H. S., Griffith, J., Case, M. E., Covert, S. F., Hall, R. D., Keith, C. H., Oliver, J. S., Orbach, M. J., Sachs, M. S., Wagner, J. R., Weise, M. J., Wunderlich, J. K., & Arnold, J. (2001). The Neurospora crassa genome: Cosmid libraries sorted by chromosome. Genetics, 157(3), 979-990.
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  PMID: 11238388;PMCID: PMC1461552;Abstract: A Neurospora crassa cosmid library of 12,000 clones (at least nine genome equivalents) has been created using an improved cosmid vector pLorist6Xh, Milch contains a bacteriophage λ origin of replication for lowcopy-number replication in bacteria and the hygromycin phosphotransferase marker for direct selection in fungi. The electrophoretic karyotype of the seven chromosomes comprising the 42.9-Mb N. crassa genome was resolved using two translocation strains. Using gel-purified chromosomal DNAs as probes against the new cosmid library and the commonly used medium-copy-number pMOcosX N. crassa cosmid library in two independent screenings, the cosmids were assigned to chromosomes. Assignments of cosmids to linkage groups on the basis of the genetic map vs. the electrophoretic karyotype are 93 ± 3% concordant. The size of each chromosome-specific subcollection of cosmids was found to be linearly proportional to the size of the particular chromosome. Sequencing of an entire cosmid containing the qa gene cluster indicated a gene density of 1 gene per 4 kbp; by extrapolation, 11,000 genes would be expected to be present in the N. crassa genome. By hybridizing 79 nonoverlapping cosmids with an average insert size of 34 kbp against cDNA arrays, the density of previously characterized expressed sequence tags (ESTs) was found to be slightly
• Abuodeh, R. O., Orbach, M. J., Mandel, M. A., Das, A., & Galgiani, J. N. (2000). Genetic transformation of Coccidioides immitis facilitated by Agrobacterium tumefaciens. Journal of Infectious Diseases, 181(6), 2106-2110.
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  PMID: 10837205;Abstract: Agrobacterium tumefaciens was used to facilitate genetic transformation of Coccidioides immitis. A gene cassette containing the gene encoding hygromycin phosphotransferase (hph) was cloned into a T-DNA vector plasmid and introduced into A. tumefaciens, and the resultant strain was used for cocultivation with germinated arthroconidia. This procedure produced numerous colonies 60- to >500-fold more resistant to hygromycin than untransformed mycelia. Both polymerase chain reaction and Southern blot analysis of the transformants indicated that all contained hph, usually as a single genomic copy. A transformation frequency of 1 per 105 arthroconidia was obtained by varying the germination time prior to cocultivation and altering the bacterium: fungus ratio. This approach requires no special equipment that might complicate biocontainment. Furthermore, transformation does not require digestion of fungal cell walls, further simplifying this procedure. A. tumefaciens-facilitated transformation should make possible the development of tagged mutagenesis and targeted gene disruption technology for C. immitis and perhaps other fungi of medical importance.
• Dioh, W., Tharreau, D., Notteghem, J. L., Orbach, M., & Lebrun, M. H. (2000). Mapping of avirulence genes in the rice blast fungus, Magnaporthe grisea, with RFLP and RAPD markers. MOLECULAR PLANT-MICROBE INTERACTIONS, 13(2), 217-227.
• Mandel, M. A., Grace, G. G., Orsborn, K. I., Schafer, F., Murphy, J. W., Orbach, M. J., & Galgiani, J. N. (2000). The Cryptococcus neoformans gene DHA1 encodes an antigen that elicits a delayed-type hypersensitivity reaction in immune mice. Infection and Immunity, 68(11), 6196-6201.
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  PMID: 11035725;PMCID: PMC97699;Abstract: When mice are vaccinated with a culture filtrate from Cryptococcus neoformans (CneF), they mount a protective cell-mediated immune response as detected by dermal delayed-type hypersensitivity (DTH) to CneF. We have identified a gene (DHA1) whose product accounts at least in part for the DTH reactivity. Using an acapsular mutant (Cap-67) of C. neoformans strain B3501, we prepared a culture filtrate (CneF-Cap67) similar to that used for preparing the commonly used skin test antigen made with C. neoformans 184A (CneF-184A). CneF-Cap67 elicited DTH in mice immunized with CneF-184A. Deglycosylation of CneF-Cap67 did not diminish its DTH activity. Furthermore, size separation by either chromatography or differential centrifugation identified the major DTH activity of CneF-Cap67 to be present in fractions that contained proteins of approximately 19 to 20 kDa. Using N-terminal and internal amino acid sequences derived from the 20-kDa band, oligonucleofide primers were designed, two of which produced a 776-bp amplimer by reverse transcription-PCR (RT-PCR) using RNA from Cap-67 to prepare cDNA for the template. The amplimer was used as a probe to isolate clones containing the full-length DHA1 gene from a phage genomic library prepared from strain B3501. The full-length cDNA was obtained by 5' rapid amplification of cDNA ends and RT-PCR. Analysis of DHA1 revealed a similarity between the deduced open reading frame and that of a developmentally regulated gene from Lentinus edodes (shiitake mushroom) associated with fruiting-body formation. Also, the gene product contained several amino acid sequences identical to those determined biochemically from the purified 20-kDa peptide encoded by DHA1. Recombinant DHA1 protein expressed in Escherichia coli was shown to elicit DTH reactions similar to those elicited by CneF-Cap67 in mice immunized against C. neoformans. Thus, DHA1 is the first gene to be cloned from C. neoformans whose product has been shown to possess immunologic activity.
• Orbach, M. J., Farrall, L., Sweigard, J. A., Chumley, F. G., & Valent, B. (2000). A telomeric avirulence gene determines efficacy for the rice blast resistance gene Pi-ta. Plant Cell, 12(11), 2019-2032.
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  PMID: 11090206;PMCID: PMC152363;Abstract: Genetic mapping showed that the rice blast avirulence gene AVR-Pita is tightly linked to a telomere on chromosome 3 in the plant pathogenic fungus Magnaporthe grisea. AVR-Pita corresponds in gene-for-gene fashion to the disease resistance (R) gene Pi-ta. Analysis of spontaneous avr-pita- mutants indicated that the gene is located in a telomeric 6.5-kb BgIII restriction fragment. Cloning and DNA sequencing led to the identification of a candidate gene with features typical of metalloproteases. This gene is located entirely within the most distal 1.5 kb of the chromosome. When introduced into virulent rice pathogens, the cloned gene specifically confers avirulence toward rice cultivars that contain Pi-ta. Frequent spontaneous loss of AVR-Pita appears to be the result of its telomeric location. Diverse mutations in AVR-Pita, including point mutations, insertions, and deletions, permit the fungus to avoid triggering resistance responses mediated by Pi-ta. A point mutation in the protease consensus sequence abolishes the AVR-Pita avirulence function.
• Peng, T., Orsborn, K. I., Orbach, M. J., & Galgiani, J. N. (1999). Proline-rich vaccine candidate antigen of Coccidioides immitis: Conservation among isolates and differential expression with spherule maturation. JOURNAL OF INFECTIOUS DISEASES, 179(2), 518-521.
• Woo, H. H., Orbach, M. J., Hirsch, A. M., & Hawes, M. C. (1999). Meristem-localized inducible expression of a UDP-glycosyltransferase gene is essential for growth and development in pea and alfalfa. PLANT CELL, 11(12), 2303-2315.
• Mandel, M. A., Crouch, V. W., Gunawardena, U. P., Harper, T. M., & Orbach, M. J. (1997). Physical mapping of the Magnaporthe grisea AVR1-MARA locus reveals the virulent allele contains two deletions. Molecular Plant-Microbe Interactions, 10(9), 1102-1105.
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  Abstract: The avirulence genes that have been identified in Magnaporthe grisea exhibit varying degrees of stability in infection assays. AVR1-MARA is considered one of the stable avirulence genes. In an effort to understand this stability, we analyzed the AVR1-MARA locus by physical mapping and chromosome walking. By walking toward AVR1-MARA from restriction fragment length polymorphism markers on both sides of the locus, we isolated sequences that are inseparable from AVR1-MARA, but we were unable to clone the complete locus. In contrast, the virulent locus avr1-MARA was isolated easily. A detailed comparative map of the two loci was constructed that identifies two deletions in the virulent locus, suggesting that virulence may be due to partial or complete deletion of the gene. Physical mapping also revealed that one progeny strain from a cross between avirulent and virulent parents appears to have spontaneously mutated to virulence.
• DiazPerez, S. V., Crouch, V. W., & Orbach, M. J. (1996). Construction and characterization of a Magnaporthe grisea bacterial artificial chromosome library. FUNGAL GENETICS AND BIOLOGY, 20(4), 280-288.
• Orbach, M. J., Chumley, F. G., & Valent, B. (1996). Electrophoretic karyotypes of Magnaporthe grisea pathogens of diverse grasses. Molecular Plant-Microbe Interactions, 9(4), 261-271.
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  Abstract: We have analyzed the chromosomes of a wide variety of strains of Magnaporthe grisea using pulsed-field gel electrophoresis (PFGE), in combination with Southern hybridization and genetic crosses. Strains analyzed included rice pathogens, field pathogens of grasses other than rice, and fertile laboratory strains. All M. grisea strains examined contain chromosomes in the size range from 2,000 kilobases (kb) to greater than 10,000 kb. Some strains also contain chromosomes, termed "mini-chromosomes," that range in size from 500 to 2,000 kb. Variation in chromosome numbers seems to be mainly among the minichromosomes. Infertile field isolates, including most rice pathogens, have a relatively high number of chromosome length polymorphisms. In contrast, some M. grisea strains from diverse hosts around the world are interfertile and have a relatively uniform karyotype, although translocations are present even among these fertile strains. There appears to be a correlation between the presence of minichromosomes and low levels of sexual fertility, although it is not yet clear if mini-chromosomes are a result or a cause of the low fertility. A mini-chromosome from a Chinese rice pathogen failed to segregate in crosses and it contained at least one DNA sequence that was found in mini-chromosomes of diverse strains, but not found in the larger standard chromosomes from these strains. These mini-chromosomes appear to be nonessential for growth and pathogenicity. Thus, we conclude that many small M. grisea chromosomes are like B-chromosomes found in plants and animals. However, size alone does not predict whether a small chromosome has B-chromosome-like properties. This is demonstrated by the normal Mendelian segregation of a small chromosome present in one parental strain used in an M. grisea RFLP mapping project that was apparently formed through an unequal translocation between two larger chromosomes.
• Orbach, M. J. (1994). A cosmid with a Hy^R marker for fungal library construction and screening. Gene, 150(1), 159-162.
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  PMID: 7959044;Abstract: The construction of a double-cos-site cosmid vector, pMOcosX, for use in making filamentous fungal genomic DNA libraries, is described. The vector has features that allow for selection of clones introduced into fungi by transformation and for efficient chromosome walking experiments. These features include (i) two cos sites allowing for easy construction of libraries without requiring size selection of insert DNA; (ii) an XhoI site for insertion of Sau3AI or MboI partially digested genomic DNA inserts that allows usage of a half-site fill-in method which minimizes the possibility of producing clones containing chimeric inserts; (iii) a bacterial hygromycin phosphotransferase-encoding gene fused to a modified cpc-1 promoter of Neurospora crassa for direct selection of cosmid clones upon introduction into fungal cells; and (iv T7 and T3 bacteriophage promoters and EcoRI, NotI and BamHI restriction sites flanking the cloning site that allow for synthesis of, or isolation of, end-specific probes for chromosome walking. The combination of features in this vector allows for the easy construction and use of high-quality fungal DNA libraries from small amounts of genomic DNA. © 1994.
• Orbach, M. J., Sachs, M. S., & Yanofsky, C. (1990). The Neurospora crassa arg-2 Locus: Structure and expression of the gene encoding the small subunit of arginine-specific carbamoyl phosphate synthetase. Journal of Biological Chemistry, 265(19), 10981-10987.
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  PMID: 2141606;Abstract: We have characterized genomic and cDNA clones for arg-2, the gene encoding the small subunit of the Neurospora crassa arginine-specific carbamoyl phosphate synthetase (CPS-A), and examined its transcriptional regulation. The polypeptide's predicted amino acid sequence (453 residues) is 56% and 36% identical with the sequences of the homologous polypeptides of Saccharomyces cerevisiae and Escherichia coli, respectively. The ARG2 polypeptide has an additional amino-terminal domain with the hallmark features of a mitochondrial signal sequence. The arg-2 mRNA also encodes a 24-residue peptide in the segment upstream of the coding region for the ARG2 polypeptide. This upstream open reading frame (uORF) strongly resembles the uORF in the homologous S. cerevisiae transcript. Northern analyses indicate that arg-2 mRNA levels are reduced by arginine supplementation and increased by amino acid limitation. The large increase in arg-2 mRNA levels that occurs in response to amino acid limitation is not observed in a strain containing the cpc-1 mutation, indicating that the cross-pathway control system participates in arg-2 regulation. Four copies of the sequence TGACTC, the binding site for the CPC1 regulatory protein, are found in the arg-2 genetic region. Two copies are located upstream of the mRNA start sites, and two are present within introns in the arg-2 uORF.
• Sweigard, J. A., Orbach, M. J., Valent, B., & Chumley, F. G. (1990). A miniprep procedure for isolating genomic DNA from Magnoporthe grisea. Fungal Genetics Reports, 37(1). doi:10.4148/1941-4765.1493
• Hamer, J. E., Farrall, L., Orbach, M. J., Valent, B., & Chumley, F. G. (1989). Host species-specific conservation of a family of repeated DNA sequences in the genome of a fungal plant pathogen. Proceedings of the National Academy of Sciences of the United States of America, 86(24), 9981-9985.
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  PMID: 2602385;PMCID: PMC298626;Abstract: We have identified a family of dispersed repetitive DNA sequences in the genome of Magnaporthe grisea, the fungus that causes rice blast disease. We have named this family of DNA sequences 'MGR' for M. grisea repeat. Analysis of five MGR clones demonstrates that MGR sequences are highly polymorphic. The segregation of MGR sequences in genetic crosses and hybridization of MGR probes to separated, chromosome-size DNA molecules of M. grisea shows that this family of sequences is distributed among the M. grisea chromosomes. MGR sequences also hybridize to discrete poly(A)+ RNAs. Southern blot analysis using a MGR probe can distinguish rice pathogens from various sources. However, MGR sequences are not highly conserved in the genomes of M. grisea field isolates that do not infect rice. These results suggest that host selection for a specific pathogen genotype has occurred during the breeding and cultivation of rice.
• MCCLUNG, C. R., PHILLIPS, J. D., ORBACH, M. J., & DUNLAP, J. C. (1989). NEW CLONING VECTORS USING BENOMYL RESISTANCE AS A DOMINANT MARKER FOR SELECTION IN NEUROSPORA-CRASSA AND IN OTHER FILAMENTOUS FUNGI. EXPERIMENTAL MYCOLOGY, 13(3), 299-302.
• Orbach, M. J., Schneider, W. P., & Yanofsky, C. (1988). Cloning of methylated transforming DNA from Neurospora crassa in Escherichia coli.. Molecular and Cellular Biology, 8(5), 2211-2213.
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  PMID: 2968501;PMCID: PMC363403;Abstract: An arg-2 mutant of Neurospora crassa was transformed to prototrophy with a pBR322-N. crassa genomic DNA library. Repeated attempts to recover the integrated transforming DNA or segments thereof by digestion, ligation, and transformation of Escherichia coli, with selection for the plasmid marker ampicillin resistance, were unsuccessful. Analyses of a N. crassa transformant demonstrated that the introduced DNA was heavily methylated at cytosine residues. This methylation was shown to be responsible for our inability to recover transformants in standard strains of E. coli; transformants were readily obtained in a strain which is deficient in the two methylcytosine restriction systems. Restriction of methylated DNA in E. coli may explain the general failure to recover vector or transforming sequences from N. crassa transformants.
• Orbach, M. J., Vollrath, D., Davis, R. W., & Yanofsky, C. (1988). An electrophoretic karyotype of Neurospora crassa.. Molecular and Cellular Biology, 8(4), 1469-1473.
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  PMID: 2967910;PMCID: PMC363304;Abstract: A molecular karyotype of Neurospora crassa was obtained by using an alternating-field gel electrophoresis system which employs contour-clamped homogeneous electric fields. The migration of all seven N. crassa chromosomal DNAs was defined, and five of the seven molecules were separated from one another. The estimated sizes of these molecules, based on their migration relative to Schizosaccharomyces pombe chromosomal DNA molecules, are 4 to 12.6 megabases. The seven linkage groups were correlated with specific chromosomal DNA bands by hybridizing transfers of contour-clamped homogeneous electric field gels with radioactive probes specific to each linkage group. The mobilities of minichromosomal DNAs generated from translocation strains were also examined. The methods used for preparation of chromosomal DNA molecules and the conditions for their separation should be applicable to other filamentous fungi.
• Paluh, J. L., Orbach, M. J., Legerton, T. L., & Yanofsky, C. (1988). The cross-pathway control gene of Neurospora crassa, cpc-1, encodes a protein similar to GCN4 of yeast and the DNA-binding domain of the oncogene v-jun-encoded protein.. Proceedings of the National Academy of Sciences of the United States of America, 85(11), 3728-3732.
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  PMID: 2967496;PMCID: PMC280291;Abstract: Expression of the gene cpc-1 is required for cross-pathway-mediated regulation of amino acid-biosynthetic genes in Neurospora crassa. We have cloned cpc-1 and present an analysis of its structure and regulation. The cpc-1-encoded transcript contains three open reading frames, two of which are located in the 720-nucleotide leader segment preceding the cpc-1 coding region. The two leader open reading frames, if translated, would produce peptides 20 and 41 residues in length. The deduced amino acid sequence of the cpc-1 polypeptide, CPC1, contains segments similar to the DNA-binding and transcriptional activation domains of GCN4, the major cross-pathway regulatory protein of yeast. The structural and functional similarities of CPC1 and GCN4 proteins suggest that cpc-1 encodes the analogous transcriptional activator of N. crassa. Messenger RNA measurements indicate that cpc-1 is transcriptionally regulated in response to amino acid starvation. The segment of CPC1 similar to the DNA-binding domain of GCN4 also is similar to the DNA-binding domains of the avian sarcoma virus oncogene-encoded v-JUN protein and human c-JUN protein.
• Orbach, M. J., Porro, E. B., & Yanofsky, C. (1986). Cloning and characterization of the gene for beta-tubulin from a benomyl-resistant mutant of Neurospora crassa and its use as a dominant selectable marker.. Molecular and Cellular Biology, 6(7), 2452-2461.
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  PMID: 2946938;PMCID: PMC367799;Abstract: We cloned the beta-tubulin gene of Neurospora crassa from a benomyl-resistant strain and determined its nucleotide sequence. The gene encodes a 447-residue protein which shows strong homology to other beta-tubulins. The coding region is interrupted by six introns, five of which are within the region coding for the first 54 amino acids of the protein. Intron position comparisons between the N. crassa gene and other fungal beta-tubulin genes reveal considerable positional conservation. The mutation responsible for benomyl resistance was determined; it caused a phenylalanine-to-tyrosine change at position 167. Codon usage in the beta-tubulin gene is biased, as has been observed for other abundantly expressed N. crassa genes such as am and the H3 and H4 histone genes. This bias results in pyrimidines in the third positions of 96% of the codons in codon families in which there is a choice between purines and pyrimidines in this position. Bias is also evident by the absence of 19 of the 61 sense codons. We demonstrated that benomyl resistance is due to the cloned beta-tubulin gene of strain Bml511(r)a and that this gene can be used as a dominant selectable marker in N. crassa transformation.
• Orbach, M. J., & Jackson, E. N. (1982). Transfer of chimeric plasmids among Salmonella typhimurium strains by P22 transduction. Journal of Bacteriology, 149(3), 985-994.
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  PMID: 6277858;PMCID: PMC216487;

Proceedings Publications

• Robb, E., Hennessey, K., Shubitz, L., Bowen, R., Orbach, M. J., Powell, D., Frelinger, J., & Galgiani, J. (2018, April). Development Pathway and Progress Towards a USDA-Approved Avirulent Live Vaccine to Prevent Coccidioidomycosis in Canines. In 62nd Annual Coccidioidomycosis Study Group, 62.
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  Presentation of our program to develop a vaccine by Ed Robb of Anivive.

Presentations

• Orbach, M. J. (2017, August). Invited Talk: Genetic Manipulation of Coccidioides. Seventh International Coccidioidomycosis Symposium. Stanford University, Stanford, CA: Stanford Center for Continuing Medical Education.
• Orbach, M. J., & Shubitz, L. (2017, August 31). For the Dogs: Bringing a UA-Invented Canine Valley Fever Vaccine Condition. CALS Research and Tech Launch Arizona Presentation. ENR2 Room S215: Tech Launch Arizona.
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  Dr. Shubitz and I presented the details of the discovery and development of our valley fever vaccine. A person from Anivive presented their plan to bring the product to market.

Poster Presentations

• Ohkura, M., Mandel, M. A., Galgiani, J. N., & Orbach, M. J. (2021, April). TRANSCRIPTOMICS OF THE COCCIDIOIDOMYCOSIS VACCINE STRAIN Δcps1 REVEALS INSIGHTS INTO CPS1 FUNCTION. 65th Coccidioidomycosis Study Group. Virtual Meeting.
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  Poster and Poster talk presentation by M. Orbach
• Orbach, M. J., Callicott, K. A., & Singh, P. (2019, August 3-7). Genetic diversity of Aspergillus flavus and biological control of aflatoxin contamination in chilies from Nigeria. APS Annual Meeting. Cleveland, OH: American Phytopathological Society.
• Arone, L., Callicott, K. A., & Orbach, M. J. (2019, August 3-7). PCR assay for rapidly differentiating aflatoxin-producing taxa in Aspergillus section Flavi. APS Annual Meeting. Cleveland, OH: American Phytopathological Society.
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  Published Abstract in Phytopathology:Volume 109, Number 10S / October 2019S2.130
• Ching'anda, C., Atehnkeng, J., Bandyopadhyay, R., Callicott, K. A., Orbach, M. J., & Cotty, P. J. (2019, August 3-7). Interactions among aflatoxigenic species of Aspergillus section Flavi during maize infection. APS Annual Meeting. Cleveland, OH: American Phytopathological Society.
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  Phytopathology: Volume 109, Number 10S / October 2019
• Mandel, M. A., Shubitz, L., Lewis, L., Trinh, H., Buntzman, A. S., Frelinger, J. A., Galgiani, J. N., & Orbach, M. J. (2017, March, 2017). Construction and Efficacy of delta-cps1, a live attenuated vaccine for coccidioidomycosis. 29th Fungal Genetics Conference. Asilomar Conference Center, Pacific Grove, CA: Genetics Society of America.
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  A poster was presented, and was selected for a poster talk in the Concurrent Session "Human Pathogenic Fungi", which was presented by Dr. Mandel.The abstracts are published as a supplement of Fungal Genetics Reports.
• Mandel, M. A., Shubitz, L., Trinh, H., Lewis, M. L., Buntzman, A., Frelinger, J. A., Galgiani, J. N., & Orbach, M. J. (2017, April 1). Construction and efficacy of delta-cps1, a live attenuated vaccine for coccidioidomycosis. 56th Annual ASM Regional Meeting- Southwest Region. University of Arizona: American Society of Microbiology.
• Orbach, M. J., Frelinger, J. A., Shubitz, L., Galgiani, J. N., Trinh, H. T., Lewis, M. L., & Butkiewicz, C. D. (2017, August). Further characterization of the avirulent delta-cps1 Coccidioides vaccine in mice. Seventh International Coccidioidomycosis Symposium. Stanford University, Stanford, CA: Stanford Center for Continuing Medical Education.
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  Poster presentation on further analysis of the duration and protection of the delta-cps1 valley fever vaccine.
• Powell, D. A., Shubitz, L., Lewis, M. L., Trinh, H. T., D, B. C., Orbach, M. J., Galgiani, J. N., & Frelinger, J. A. (2017, August). Determining Mechanisms of Protection in a Live Attenuated Coccidiodes Vaccine. 7th International Coccidioidomycosis Symposium. Stanford University, Stanford, CA: Stanford Center for Continuing Medical Education.
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  poster presentation describing the immune response of mice to our valley fever vaccine.
• Ohkura, M., Cotty, P., & Orbach, M. (2016, July 2016). Comparative genomics of S and L morphotypes of Aspergillus flavus. Annual Meeting of the American-Phytopathological-Society. Tampa, FL: American Phytopathological Society.
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  Published Abstract:Phytopathology. 2016. 106:158-158

Reviews

• Hao, X., Taghavi, S., Xie, P., Orbach, M. J., Alwathnani, H. A., Rensing, C., & Wei, G. (2014. PHYTOREMEDIATION OF HEAVY AND TRANSITION METALS AIDED BY LEGUME-RHIZOBIA SYMBIOSIS(pp 179-202).
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  Legumes are important for nitrogen cycling in the environment and agriculture due to the ability of nitrogen fixation by rhizobia. In this review, we introduce an important and potential role of legume-rhizobia symbiosis in aiding phytoremediation of some metal contaminated soils as various legumes have been found to be the dominant plant species in metal contaminated areas. Resistant rhizobia used for phytoremediation could act on metals directly by chelation, precipitation, transformation, biosorption and accumulation. Moreover, the plant growth promoting (PGP) traits of rhizobia including nitrogen fixation, phosphorus solubilization, phytohormone synthesis, siderophore release, and production of ACC deaminase and the volatile compounds of acetoin and 2, 3-butanediol may facilitate legume growth while lessening metal toxicity. The benefits of using legumes inoculated with naturally resistant rhizobia or recombinant rhizobia with enhanced resistance, as well as co-inoculation with other plant growth promoting bacteria (PGPB) are discussed. However, the legume-rhizobia symbiosis appears to be sensitive to metals, and the effect of metal toxicity on the interaction between legumes and rhizobia is not clear. Therefore, to obtain the maximum benefits from legumes assisted by rhizobia for phytoremediation of metals, it is critical to have a good understanding of interactions between PGP traits, the symbiotic plant-rhizobia relationship and metals.
• Nguyen, C., Barker, B. M., Hoover, S., Nix, D. E., Ampel, N. M., Frelinger, J. A., Orbach, M. J., & Galgiani, J. N. (2013. Recent advances in our understanding of the environmental, epidemiological, immunological, and clinical dimensions of coccidioidomycosis(pp 505-25).
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  Coccidioidomycosis is the endemic mycosis caused by the fungal pathogens Coccidioides immitis and C. posadasii. This review is a summary of the recent advances that have been made in the understanding of this pathogen, including its mycology, genetics, and niche in the environment. Updates on the epidemiology of the organism emphasize that it is a continuing, significant problem in areas of endemicity. For a variety of reasons, the number of reported coccidioidal infections has increased dramatically over the past decade. While continual improvements in the fields of organ transplantation and management of autoimmune disorders and patients with HIV have led to dilemmas with concurrent infection with coccidioidomycosis, they have also led to advances in the understanding of the human immune response to infection. There have been some advances in therapeutics with the increased use of newer azoles. Lastly, there is an overview of the ongoing search for a preventative vaccine.