Novel Approach to Designing Primers for Identification and Distinction of the Human Pathogenic Fungi Coccidioides immitis and Coccidioides posadasii by PCR Amplification

ABSTRACTRodents are a natural host for the dimorphic pathogenic fungiCoccidioides immitisandCoccidioides posadasii, and mice are a good model for human infection. Humans and rodents both express Dectin-1 and Toll-like receptor 2 (TLR2) on myeloid cells, and those receptors collaborate to maximize the cytokine/chemokine responses to spherules (the tissue form of the fungi) and to formalin-killed spherules (FKS). We showed that Dectin-1 is necessary for resistance to pulmonary coccidioidomycosis, but the importance of TLR2in vivois uncertain. Myeloid differentiation factor 88 (MyD88) is the adapter protein for TLR2 and -4, interleukin-1R1 (IL-1R1), and IL-18R1. MyD88/TRIF−/−and MyD88−/−mice were equally susceptible toC. immitisinfection, in contrast to C57BL/6 (B6) controls. Of the four surface receptors, only IL-1R1 was required for resistance toC. immitis, partially explaining the susceptibility of MyD88−/−mice. We also found that FKS stimulated production of IL-1Ra by bone marrow-derived dendritic cells (BMDCs), independent of MyD88 and Dectin-1. There also was a very high concentration of IL-1Ra in the lungs of infected B6 mice, supporting the potential importance of this regulatory IL-1 family protein in the largely ineffective response of B6 mice to coccidioidomycosis. These results suggest that IL-1R1 signaling is important for defense againstC. immitisinfection.

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Biochemistry and function of cutin and suberin

Canadian Journal of Botany ◽

10.1139/b84-391 ◽

1984 ◽

Vol 62 (12) ◽

pp. 2918-2933 ◽

Cited By ~ 201

Author(s):

P. E. Kolattukudy

Keyword(s):

Hydrolytic Enzyme ◽

Pathogenic Fungi ◽

Catalytic Triad ◽

Nucleotide Sequencing ◽

Hydroxyl Groups ◽

Serine Hydrolase ◽

Tissue Slices ◽

Novel Approach ◽

Copy Dna ◽

And Function

Cutin, the structural component of plant cuticle, is a biopolyester composed of hydroxy- and hydroxyepoxy-fatty acids. The major monomers are a 16-hydroxy C16 acid, a 10,16-dihydroxy C16 acid together with its positional isomers, 18-hydroxy C18 acids, 18-hydroxy-9,10-epoxy C18 acids, and 9,10,18-trihydroxy C18 acids. The hydroxylation, epoxidation, and epoxide hydration reactions postulated to be involved in the biosynthesis of these monomers have been demonstrated in tissue slices and in cell-free preparations. The synthesis of the polymer occurs by the enzymatic transfer of the hydroxyacyl groups from CoA to the free hydroxyl groups in cutin primer. Natural and wound periderms and a variety of internal barrier layers contain a somewhat analogous polymer called suberin. This polymer is probably composed of aromatic domains somewhat similar to those found in lignin and aliphatic polyester domains somewhat similar to cutin. The chemical composition and biosynthesis of this polymer is discussed. Pathogenic fungi use a hydrolytic enzyme, cutinase, to gain entry into the plant through the cuticle. The fungal cutinase has been isolated from a variety of pathogenic fungi and characterized. This enzyme is a "serine hydrolase" containing the characteristic catalytic triad. The primary structure of this enzyme has been determined using both amino acid and nucleotide sequencing of the cloned copy DNA. Inhibition of cutinase was shown to prevent fungal infection of plants. This novel approach to fungal control is described.

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Development of a rapid, cost-effective TaqMan Real-Time PCR Assay for identification and differentiation of Coccidioides immitis and Coccidioides posadasii

Medical Mycology ◽

10.1080/13693780903218990 ◽

2009 ◽

pp. 1-4 ◽

Cited By ~ 1

Author(s):

Kelly Sheff ◽

Emily York ◽

Elizabeth Driebe ◽

Bridget Barker ◽

Steven Rounsley ◽

...

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Cost Effective ◽

Coccidioides Immitis ◽

Coccidioides Posadasii ◽

Pcr Assay

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Differences in Host Innate Responses among Coccidioides Isolates in a Murine Model of Pulmonary Coccidioidomycosis

Eukaryotic Cell ◽

10.1128/ec.00122-15 ◽

2015 ◽

Vol 14 (10) ◽

pp. 1043-1053 ◽

Cited By ~ 8

Author(s):

Eric R. G. Lewis ◽

Victoria R. David ◽

Adina L. Doyle ◽

Khadijeh Rajabi ◽

Jeffrey A. Kiefer ◽

...

Keyword(s):

Murine Model ◽

Host Response ◽

Early Time ◽

Lavage Fluid ◽

Host Responses ◽

Coccidioides Immitis ◽

Coccidioides Posadasii ◽

Hybrid Strain ◽

Susceptible Host ◽

Content Type

ABSTRACTCoccidioides immitisandCoccidioides posadasiiare soil-dwelling fungi and the causative agents of coccidioidomycosis, a mycosis endemic to certain semiarid regions in the Americas. The most common route of infection is by inhalation of airborneCoccidioidesarthroconidia. Once a susceptible host inhales the conidia, a transition to mature endosporulated spherules can occur within the first 5 days of infection. For this study, we examined the host response in a murine model of coccidioidomycosis during a time period of infection that has not been well characterized. We collected lung tissue and bronchoalveolar lavage fluid (BALF) from BALB/c mice that were infected with aC. immitispure strain, aC. immitishybrid strain, or aC. posadasiistrain as well as uninfected mice. We compared the host responses to theCoccidioidesstrains used in this study by assessing the level of transcription of selected cytokine genes in lung tissues and characterized host and fungal proteins present in BALF. Host response varied depending on theCoccidioidesstrain that was used and did not appear to be overly robust. This study provides a foundation to begin to dissect the host immune response early in infection, to detect abundantCoccidioidesproteins, and to develop diagnostics that target these early time points of infection.

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Coccidioidomicosis en caninos y felinos: hallazgos clínicos, diagnóstico y tratamiento

Analecta Veterinaria ◽

10.24215/15142590e023 ◽

2018 ◽

Vol 38 (1) ◽

pp. 33-44

Author(s):

Galo Ernesto Martínez Cepeda ◽

Paola Revelo Ruales

Keyword(s):

Coccidioides Immitis ◽

Coccidioides Posadasii ◽

Sobre Todo

La coccidioidomicosis es una enfermedad endémica de las zonas áridas de América. Se han identificado dos especies de este género, Coccidioides immitis y Coccidioides posadasii. La variedad de especies susceptibles, incluyendo a los humanos, hace que su diagnóstico tenga una gran relevancia. Esta enfermedad no presenta una sintomatología específica que facilite su identificación. Tanto en caninos como en felinos, se puede presentar como una enfermedad pulmonar leve o una enfermedad multisistémica diseminada que puede causar la muerte. El diagnóstico se basa en el hallazgo histopatológico de microabscesos (frescos) en muestras del tracto respiratorio u otros órganos con la respectiva identificación de esférulas que contengan endosporas. No obstante, a causa del desafío que representa su diagnóstico sobre todo en regiones donde no es endémica, es necesario recurrir a pruebas complementarias, como el cultivo y aislamiento, bajo las condiciones de bioseguridad adecuadas, y una posterior confirmación mediante pruebas moleculares.

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Dual UMIs and Dual Barcodes With Minimal PCR Amplification Removes Artifacts and Acquires Accurate Antibody Repertoire

Frontiers in Immunology ◽

10.3389/fimmu.2021.778298 ◽

2021 ◽

Vol 12 ◽

Author(s):

Qilong Wang ◽

Huikun Zeng ◽

Yan Zhu ◽

Minhui Wang ◽

Yanfang Zhang ◽

...

Keyword(s):

Pcr Amplification ◽

Antibody Repertoire ◽

Consensus Building ◽

Chain Reaction ◽

Single Nucleotide ◽

False Discovery ◽

Novel Approach ◽

Repertoire Sequencing ◽

Polymerase Chain ◽

Antibody Repertoires

Antibody repertoire sequencing (Rep-seq) has been widely used to reveal repertoire dynamics and to interrogate antibodies of interest at single nucleotide-level resolution. However, polymerase chain reaction (PCR) amplification introduces extensive artifacts including chimeras and nucleotide errors, leading to false discovery of antibodies and incorrect assessment of somatic hypermutations (SHMs) which subsequently mislead downstream investigations. Here, a novel approach named DUMPArts, which improves the accuracy of antibody repertoires by labeling each sample with dual barcodes and each molecule with dual unique molecular identifiers (UMIs) via minimal PCR amplification to remove artifacts, is developed. Tested by ultra-deep Rep-seq data, DUMPArts removed inter-sample chimeras, which cause artifactual shared clones and constitute approximately 15% of reads in the library, as well as intra-sample chimeras with erroneous SHMs and constituting approximately 20% of the reads, and corrected base errors and amplification biases by consensus building. The removal of these artifacts will provide an accurate assessment of antibody repertoires and benefit related studies, especially mAb discovery and antibody-guided vaccine design.

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The endozoan, small-mammal reservoir hypothesis and the life cycle of Coccidioides species

Medical Mycology ◽

10.1093/mmy/myy039 ◽

2019 ◽

Vol 57 (Supplement_1) ◽

pp. S16-S20 ◽

Cited By ~ 16

Author(s):

John W Taylor ◽

Bridget M Barker

Keyword(s):

Small Mammals ◽

Small Mammal ◽

Alternative Hypothesis ◽

Pathogenic Fungi ◽

Gene Families ◽

Comparative Genomic ◽

Soil Parameters ◽

Plant Cell Walls ◽

Coccidioides Immitis ◽

Growth And Reproduction

AbstractThe prevailing hypothesis concerning the ecology of Coccidioides immitis and C. posadasii is that these human pathogenic fungi are soil fungi endemic to hot, dry, salty regions of the New World and that humans and the local, small-mammal fauna are only accidental hosts. Here we advance an alternative hypothesis that Coccidioides spp. live in small mammals as endozoans, which are kept inactive but alive in host granulomas and which transform into spore-producing hyphae when the mammal dies. The endozoan hypothesis incorporates results from comparative genomic analyses of Coccidioides spp. and related taxa that have shown a reduction in gene families associated with deconstruction of plant cell walls and an increase in those associated with digestion of animal protein, consistent with an evolutionary shift in substrate from plants to animals. If true, the endozoan hypothesis requires that models of the prevalence of human coccidioidomycosis account not only for direct effects of climate and soil parameters on the growth and reproduction of Coccidioides spp. but also consider indirect effects on these fungi that come from the plants that support the growth and reproduction of the small mammals that, in turn, support these endozoic fungi.

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Characterization of the tRNA ligases of pathogenic fungi Aspergillus fumigatus and Coccidioides immitis

RNA ◽

10.1261/rna.057455.116 ◽

2016 ◽

Vol 22 (10) ◽

pp. 1500-1509 ◽

Cited By ~ 5

Author(s):

Barbara S. Remus ◽

Beate Schwer ◽

Stewart Shuman

Keyword(s):

Aspergillus Fumigatus ◽

Pathogenic Fungi ◽

Coccidioides Immitis

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The Novel Fungal Cyp51 Inhibitor VT-1598 Is Efficacious in Experimental Models of Central Nervous System Coccidioidomycosis Caused by Coccidioides posadasii and Coccidioides immitis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02258-17 ◽

2018 ◽

Vol 62 (4) ◽

Cited By ~ 16

Author(s):

Nathan P. Wiederhold ◽

Lisa F. Shubitz ◽

Laura K. Najvar ◽

Rosie Jaramillo ◽

Marcos Olivo ◽

...

Keyword(s):

Central Nervous System ◽

Nervous System ◽

Plasma Concentrations ◽

Coccidioides Immitis ◽

Coccidioides Posadasii ◽

Content Type ◽

Fungal Burden ◽

Content Model ◽

Survival Studies

ABSTRACT Coccidioidal meningitis can cause significant morbidity, and lifelong antifungal therapy is often required. VT-1598 is a fungus-specific Cyp51 inhibitor that has potent in vitro activity against Coccidioides species. We evaluated the in vivo efficacy of VT-1598 in murine models of central nervous system coccidioidomycosis caused by C. posadasii and C. immitis . Infection was introduced via intracranial inoculation, and therapy began 48 h postinoculation. Oral treatments consisted of vehicle control, VT-1598, and positive controls of fluconazole in the C. immitis study and VT-1161 in the C. posadasii study. Treatment continued for 7 and 14 days in the fungal-burden and survival studies, respectively. Fungal burden was assessed in brain tissue collected 24 to 48 h posttreatment in the fungal-burden studies, on the days the mice succumbed to infection, or at prespecified endpoints in the survival studies. VT-1598 plasma concentrations were also measured in the C. posadasii study. VT-1598 resulted in significant improvements in survival in mice infected with either species. In addition, the fungal burden was significantly reduced in the fungal-burden studies. Plasma concentrations 48 h after dosing stopped remained above the VT-1598 MIC against the C. posadasii isolate, although levels were undetectable in the survival study after a 4-week washout. Whereas fungal burden remained suppressed after a 2-week washout in the C. immitis model, a higher fungal burden was observed in the survival arm of the C. posadasii model. This in vivo efficacy supports human studies to establish the utility of VT-1598 for the treatment of coccidioidomycosis.

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Coccidioides Species

Mayo Clinic Infectious Diseases Board Review ◽

10.1093/med/9780199827626.003.0017 ◽

2012 ◽

pp. 168-170

Author(s):

Janis E Blair

Keyword(s):

New Mexico ◽

Western Hemisphere ◽

South American ◽

Coccidioides Immitis ◽

Coccidioides Posadasii ◽

Small Areas ◽

Northern Mexico ◽

Restricted Region ◽

Life Zone ◽

Parasitic Phase

Coccidioidomycosis is endemic only in the Western Hemisphere, mainly in southern Arizona, New Mexico, Texas, and much of central and southern California, and in small areas in South American countries, including Argentina, Brazil, Colombia, Paraguay, and Venezuela. It is highly endemic in northern Mexico. The organism grows in soil in a semiarid, geographically restricted region, the Lower Sonoran Life Zone. Two nearly identical species cause coccidioidomycosis: Coccidioides immitis and Coccidioides posadasii. Upon inhalation, the fungus enters the lungs and converts to the tissue parasitic phase, which consists of alternating spherules and endospores. Infection can be asymptomatic or can cause flulike symptoms, meningitis, or skin, bone or joint manifestations. Diagnosis and treatment are also reviewed.

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