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Bruce S. Klein - One of the best experts on this subject based on the ideXlab platform.
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Blastomyces Dermatitidis serine protease dipeptidyl peptidase iva dppiva cleaves elr cxc chemokines altering their effects on neutrophils
Cellular Microbiology, 2017Co-Authors: Jenna Lorenzini, Scott J Fites, Jeniel E Nett, Bruce S. KleinAbstract:Summary Blastomycosis elicits a pyogranulomatous inflammatory response that involves a prominent recruitment of neutrophils to the site of infection. Although neutrophils are efficiently recruited to the site of infection, this event is paradoxically coupled with the host's inability to control infection by Blastomyces Dermatitidis, the causative agent. The mechanisms underlying this characteristic pyogranulomatous response and inability of neutrophils to kill the yeast are poorly understood. We recently reported that the fungal protease dipeptidyl peptidase IVA (DppIVA) promotes B. Dermatitidis virulence by cleaving a dipeptide from the N-terminus of C–C chemokines and granulocyte/macrophage-colony stimulating factor, thereby inactivating them. Herein, we present evidence that DppIVA can also truncate the N-terminus of members of the ELR+CXC chemokine family, which are known to modulate neutrophil function. We show that the DppIVA cleaved form of human (h) CXCL-2, for example, hCXCL-2 (3-73), is a more potent neutrophil chemoattractant than its intact counterpart, but hCXCL-2 (3-73) is conversely impaired in its ability to prime the reactive oxygen species response of neutrophils. Thus, DppIVA action on ELR+CXC chemokines may promote the pyogranulomatous response that is typical of blastomycosis, while also explaining the inability of neutrophils to control infection.
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Analysis of non-adhesive pathogenic mechanisms of BAD1 on Blastomyces Dermatitidis
2016Co-Authors: Beatriz Finkel-jimenez, Tristan T Brandhorst, Hanna I. Filutowicz, Tom Warner, Bruce S. KleinAbstract:The adhesin BAD1 is required for virulence of Blastomyces Dermatitidis in a pulmonary model of infection. Herein, we explored mechanisms by which BAD1 enhances pathogenicity of the fungus. Isogenic strains with and without BAD1 exhibited similar phenotypic differences in virulence by pulmonary and intravenous routes of infection, indicating that BAD1 may exert virulence beyond adherence to respiratory lining cells. Non-adhesive mechanisms including maintenance of intrinsic resistance of yeast against phagocyte responses and products were excluded. A shift in the balance of type 1 and 2 cytokines and in the cellular profile of the inflammatory response after the first week of pulmonary infection was associated with BAD1. By the second week of infection, infection with wild-type yeast was associated with less IL-12 and IFN-g, and more IL-10, and an influx of inflammatory cells rich in neutrophils and poor in T-cells, when compared to infection with the BAD1 null strain. Taken together with previously reported BAD1 perturbations of TNF-a and TGF-b, these data suggest that BAD1 contributes significantly to the pathogenicity of B. Dermatitidis by also deviating host adaptive immunity, and leukocyte responses
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mannose receptor is required for optimal induction of vaccine induced t helper type 17 cells and resistance to Blastomyces Dermatitidis infection
The Journal of Infectious Diseases, 2016Co-Authors: Huafeng Wang, Bruce S. Klein, Vanessa Lebert, Tassanee Lerksuthirat, Kevin Galles, Marcel WuthrichAbstract:We investigated how innate sensing by the mannose receptor (MR) influences the development of antifungal immunity. We demonstrate that MR senses mannan on the surface of attenuated Blastomyces Dermatitidis vaccine yeast and that MR(-/-) mice demonstrate impaired vaccine immunity against lethal experimental blastomycosis, compared with wild-type control mice. Using naive Blastomyces-specific transgenic CD4(+) T cells, we found that MR regulates differentiation of naive T cells into T-helper type 17 (Th17) effector cells, which are essential in vaccine immunity against systemic dimorphic fungi. Thus, MR regulates differentiation of Th17 cells and is required to induce vaccine immunity against lethal pulmonary blastomycosis.
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the c type lectin receptor mcl mediates vaccine induced immunity against infection with Blastomyces Dermatitidis
Infection and Immunity, 2016Co-Authors: Huafeng Wang, Bruce S. Klein, Tassanee Lerksuthirat, Marcel WuthrichAbstract:C-type lectin receptors (CLRs) are essential in shaping the immune response to fungal pathogens. Vaccine-induced resistance requires Dectin-2 to promote differentiation of antifungal Th1 and Th17 cells. Since Dectin-2 and MCL heterodimerize and both CLRs use FcRγ as the signaling adaptor, we investigated the role of MCL in vaccine immunity to the fungal pathogen Blastomyces Dermatitidis. MCL(-/-) mice showed impaired vaccine resistance against B. Dermatitidis infection compared to that of wild-type animals. The lack of resistance correlated with the reduced recruitment of Th17 cells to the lung upon recall following experimental challenge and impaired interleukin-17 (IL-17) production by vaccine antigen-stimulated splenocytes in vitro. Soluble MCL fusion protein recognized and bound a water-soluble ligand from the cell wall of vaccine yeast, but the addition of soluble Dectin-2 fusion protein did not augment ligand recognition by MCL. Taken together, our data indicate that MCL regulates the development of vaccine-induced Th17 cells and protective immunity against lethal experimental infection with B. Dermatitidis.
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the unappreciated intracellular lifestyle of Blastomyces Dermatitidis
Journal of Immunology, 2015Co-Authors: Alana K Sterkel, Marcel Wuthrich, Robert C Mettelman, Bruce S. KleinAbstract:Blastomyces Dermatitidis, a dimorphic fungus and the causative agent of blastomycosis, is widely considered an extracellular pathogen, with little evidence for a facultative intracellular lifestyle. We infected mice with spores, that is, the infectious particle, via the pulmonary route and studied intracellular residence, transition to pathogenic yeast, and replication inside lung cells. Nearly 80% of spores were inside cells at 24 h postinfection with 10 4 spores. Most spores were located inside of alveolar macrophages, with smaller numbers in neutrophils and dendritic cells. Real-time imaging showed rapid uptake of spores into alveolar macrophages, conversion to yeast, and intracellular multiplication during in vitro coculture. The finding of multiple yeast in a macrophage was chiefly due to intracellular replication rather than multiple phagocytic events or fusion of macrophages. Depletion of alveolar macrophages curtailed infection in mice infected with spores and led to a 26-fold reduction in lung CFU by 6 d postinfection versus nondepleted mice. Phase transition of the spores to yeast was delayed in these depleted mice over a time frame that correlated with reduced lung CFU. Spores cultured in vitro converted to yeast faster in the presence of macrophages than in medium alone. Thus, although advanced B. Dermatitidis infection may exhibit extracellular residence in tissue, early lung infection with infectious spores reveals its unappreciated facultative intracellular lifestyle.
Marcel Wuthrich - One of the best experts on this subject based on the ideXlab platform.
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mannose receptor is required for optimal induction of vaccine induced t helper type 17 cells and resistance to Blastomyces Dermatitidis infection
The Journal of Infectious Diseases, 2016Co-Authors: Huafeng Wang, Bruce S. Klein, Vanessa Lebert, Tassanee Lerksuthirat, Kevin Galles, Marcel WuthrichAbstract:We investigated how innate sensing by the mannose receptor (MR) influences the development of antifungal immunity. We demonstrate that MR senses mannan on the surface of attenuated Blastomyces Dermatitidis vaccine yeast and that MR(-/-) mice demonstrate impaired vaccine immunity against lethal experimental blastomycosis, compared with wild-type control mice. Using naive Blastomyces-specific transgenic CD4(+) T cells, we found that MR regulates differentiation of naive T cells into T-helper type 17 (Th17) effector cells, which are essential in vaccine immunity against systemic dimorphic fungi. Thus, MR regulates differentiation of Th17 cells and is required to induce vaccine immunity against lethal pulmonary blastomycosis.
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the c type lectin receptor mcl mediates vaccine induced immunity against infection with Blastomyces Dermatitidis
Infection and Immunity, 2016Co-Authors: Huafeng Wang, Bruce S. Klein, Tassanee Lerksuthirat, Marcel WuthrichAbstract:C-type lectin receptors (CLRs) are essential in shaping the immune response to fungal pathogens. Vaccine-induced resistance requires Dectin-2 to promote differentiation of antifungal Th1 and Th17 cells. Since Dectin-2 and MCL heterodimerize and both CLRs use FcRγ as the signaling adaptor, we investigated the role of MCL in vaccine immunity to the fungal pathogen Blastomyces Dermatitidis. MCL(-/-) mice showed impaired vaccine resistance against B. Dermatitidis infection compared to that of wild-type animals. The lack of resistance correlated with the reduced recruitment of Th17 cells to the lung upon recall following experimental challenge and impaired interleukin-17 (IL-17) production by vaccine antigen-stimulated splenocytes in vitro. Soluble MCL fusion protein recognized and bound a water-soluble ligand from the cell wall of vaccine yeast, but the addition of soluble Dectin-2 fusion protein did not augment ligand recognition by MCL. Taken together, our data indicate that MCL regulates the development of vaccine-induced Th17 cells and protective immunity against lethal experimental infection with B. Dermatitidis.
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the unappreciated intracellular lifestyle of Blastomyces Dermatitidis
Journal of Immunology, 2015Co-Authors: Alana K Sterkel, Marcel Wuthrich, Robert C Mettelman, Bruce S. KleinAbstract:Blastomyces Dermatitidis, a dimorphic fungus and the causative agent of blastomycosis, is widely considered an extracellular pathogen, with little evidence for a facultative intracellular lifestyle. We infected mice with spores, that is, the infectious particle, via the pulmonary route and studied intracellular residence, transition to pathogenic yeast, and replication inside lung cells. Nearly 80% of spores were inside cells at 24 h postinfection with 10 4 spores. Most spores were located inside of alveolar macrophages, with smaller numbers in neutrophils and dendritic cells. Real-time imaging showed rapid uptake of spores into alveolar macrophages, conversion to yeast, and intracellular multiplication during in vitro coculture. The finding of multiple yeast in a macrophage was chiefly due to intracellular replication rather than multiple phagocytic events or fusion of macrophages. Depletion of alveolar macrophages curtailed infection in mice infected with spores and led to a 26-fold reduction in lung CFU by 6 d postinfection versus nondepleted mice. Phase transition of the spores to yeast was delayed in these depleted mice over a time frame that correlated with reduced lung CFU. Spores cultured in vitro converted to yeast faster in the presence of macrophages than in medium alone. Thus, although advanced B. Dermatitidis infection may exhibit extracellular residence in tissue, early lung infection with infectious spores reveals its unappreciated facultative intracellular lifestyle.
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interleukin 1 enhances vaccine induced antifungal t helper 17 cells and resistance against Blastomyces Dermatitidis infection
The Journal of Infectious Diseases, 2013Co-Authors: Marcel Wuthrich, Vanessa Lebert, Kevin Galles, Jane Huli, S Z Bensasson, William E Paul, Bruce S. KleinAbstract:Vaccine-induced T-helper 17 (Th17) cells are necessary and sufficient to protect against fungal infection. Although live fungal vaccines are efficient in driving protective Th17 responses and immunity, attenuated fungi may not be safe for human use. Heat-inactivated formulations and subunit vaccines are safer but less potent and require adjuvant to increase their efficacy. Here, we show that interleukin 1 (IL-1) enhances the capacity of weak vaccines to induce protection against lethal Blastomyces Dermatitidis infection in mice and is far more effective than lipopolysaccharide. While IL-1 enhanced expansion and differentiation of fungus-specific T cells by direct action on those cells, cooperation with non-T cells expressing IL-1R1 was necessary to maximize protection. Mechanistically, IL-17 receptor signaling was required for the enhanced protection induced by IL-1. Thus, IL-1 enhances the efficacy of safe but inefficient vaccines against systemic fungal infection in part by increasing the expansion of CD4+ T cells, allowing their entry into the lungs, and inducing their differentiation to protective Th17 cells.
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discordant influence of Blastomyces Dermatitidis yeast phase specific gene bys1 on morphogenesis and virulence
Infection and Immunity, 2010Co-Authors: Theerapong Krajaejun, Thomas D. O'sullivan, Marcel Wuthrich, Gregory M Gauthier, Thomas F Warner, Bruce S. KleinAbstract:ABSTRACT Blastomyces Dermatitidis is a thermally induced dimorphic fungus capable of causing lung and systemic infections in immunocompetent animal hosts. With the publication of genomic sequences from three different strains of B. Dermatitidis and the development of RNA interference as a gene-silencing tool, it has become possible to easily ascertain the virulence and morphological effects of knocking down the expression of candidate genes of interest. BYS1 (Blastomyces yeast-phase-specific 1), first identified by Burg and Smith, is expressed at high levels in yeast cells and is undetectable in mold. The deduced protein sequence of BYS1 has a putative signal sequence at its N terminus, opening the possibility that the BYS1-encoded protein is associated with the yeast cell wall. Herein, strains of B. Dermatitidis with silenced expression of BYS1 were engineered and tested for morphology and virulence. The silenced strains produced rough-surfaced cultures on agar medium and demonstrated a propensity to form pseudohyphal cells on prolonged culture in vitro and in vivo, as measured in the mouse lung. Tests using a mouse model of blastomycosis with either yeast or spore inocula showed that the bys1-silenced strains were as virulent as control strains. Thus, although silencing of BYS1 alters morphology at 37°C, it does not appear to impair the pathogenicity of B. Dermatitidis.
G M Scalarone - One of the best experts on this subject based on the ideXlab platform.
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Blastomyces Dermatitidis yeast lysate antigen combinations antibody detection in dogs with blastomycosis
Veterinary Medicine International, 2013Co-Authors: Alex R Boyd, Jamie L Vandyke, G M ScalaroneAbstract:The systemic fungal infection, blastomycosis, which infects both humans and animals has presented a diagnostic challenge for clinicians for many years. The aim of this study was to evaluate the diagnostic sensitivity of Blastomyces Dermatitidis yeast lysate antigens with respect to antibody detection in dogs with blastomycosis. Lysate antigens were prepared from B. Dermatitidis isolates T-58 and T-66 (dogs, Tennessee) and WI-R and WI-J (dogs, Wisconsin). Based on results obtained from a preliminary comparative study, five combinations of these isolates and one individual isolate were tested against 92 serum specimens from dogs with culture-proven or histologically-confirmed blastomycosis, using the indirect enzyme-linked immunosorbent assay (ELISA). Mean absorbance values obtained from the sera ranged from 0.905 with the individual T-58 antigen to 1.760 using an antigen combination (T-58 + T-66 + WI-R). All of the 6 antigenic preparations were able to detect antibody in the serum specimens, but the antigen combinations detected antibody to a higher degree than the individual antigen. This study provides evidence that combinations of the yeast lysate reagents seem to be more efficacious for antibody detection in dog sera, but our laboratory is continuing to evaluate antigen lysate combinations for detection of antibodies in blastomycosis.
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Blastomyces Dermatitidis stability studies on different yeast lysate antigens
Open Journal of Immunology, 2013Co-Authors: Tiffany R Allison, Joshua C Wright, G M ScalaroneAbstract:In Trial 1, 19 lots of Blastomyces Dermatitidis (T-58; Tennessee dog isolate) were assayed to determine the stability of the reagents following storage. The reactivity of the antigens, produced from 1989 to 2012 and stored at 4°C, was determined by comparing antibody detection (enzyme-linked immunosorbent assay; ELISA) in 12 serum specimens from immunized rabbits. All of the 19 reagents produced during this 23-year period exhibited a high degree of stability and were able to detect antibody in the sera. Mean absorbance values ranged from 0.798 (1989) to 0.827 (2012) and a mean value for all 19 antigens of 0.728. In a related evaluation, Trial 2, B. Dermatitidis lysate antigens prepared from 8 isolates (dog, human, soil) at two different time periods were assayed as above to determine reactivity. The time of storage between the first and second reagents varied from 4 to 17 years. The results indicated that all 16 of the lysate antigens detected antibody in the 15 rabbit serum specimens with mean absorbance values ranging from 0.346 to 0.682, but variations in reactivity were observed depending on the lysate and the serum specimen assayed. This comparative study provided evidence that the antigenic reagents do exhibit some lot-to-lot variation in reactivity, but they did not lose any appreciable potency during prolonged storage.
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sensitivity and specificity determinations with isoelectric focusing fractions of Blastomyces Dermatitidis for antibody detection in serum specimens from infected dogs
Open Journal of Veterinary Medicine, 2012Co-Authors: Joshua C Wright, Terrick E Harrild, G M ScalaroneAbstract:Blastomycosis and histoplasmosis manifest as lung and systemic fungal infections in mammals caused by Histoplasma capsulatum, and Blastomyces Dermatitidis. These infections exhibit cross reactivity of antibodies which makes a correct diagnosis potentially elusive. The purpose of this study was to gain an understanding of which isoelectric focusing fractions (RotoforTM) of B. Dermatitidis were reactive or cross reactive with serum specimens from dogs infected with B. Dermatitidis, H. capsulatum, and Cryptococcus neoformans. Three serum specimens from dogs that were infected with B. Dermatitidis, two dogs infected with H. capsulatum, and one dog infected with C. neoformans were assayed against the 20 B. Dermatitidis RotoforTM fractions. Reactivity was determined using the indirect enzyme linked immunoassay (ELISA). Reactivity with B. Dermatitidis was found predominantly in the protein fractions 1 - 6, and cross reactivity with H. capsulatum, and C. neoformans sera was found within the B. Dermatitidis protein fractions 15 - 19.
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isoelectric focusing and elisa evaluation of a Blastomyces Dermatitidis human isolate
Mycopathologia, 2007Co-Authors: Jack F Shurley, G M ScalaroneAbstract:Blastomyces Dermatitidis is a dimorphic fungal organism and the causative agent of blastomycosis. This organism is endemic east of the Mississippi river as is the fungal organism Histoplasma capsulatum. This study was performed to determine if sensitive and specific antigens from the B. Dermatitidis yeast phase lysate (human isolate 592) could be separated using isoelectric focusing (IEF) to eliminate antigens that are cross-reactive with H. capsulatum. Indirect enzyme linked immunosorbent assays were performed to test for reactivity and cross-reactivity and indicate that certain fractions (4-6) were highly reactive. Fraction 16 exhibited a high degree of cross-reactivity with H. capsulatum. This study indicates that IEF may be a useful method for the separation of B. Dermatitidis proteins.
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Blastomyces Dermatitidis antigen detection in urine specimens from dogs with blastomycosis using a competitive binding inhibition elisa
Mycopathologia, 2005Co-Authors: Jack F Shurley, A M Legendre, G M ScalaroneAbstract:A competitive binding inhibition enzyme linked immunosorbent assay (ELISA) was used to detect Blastomyces Dermatitidis antigens in urine specimens from dogs with blastomycosis. Sera from rabbits immunized with B. Dermatitidis killed whole yeast cells were used as the primary antibody in the competitive ELISA. This initial study was performed to determine if B. Dermatitidis antigen detection was possible and to test the efficacy of the rabbit sera as a primary antibody. An indirect ELISA was also performed to compare antigen detection in urine to antibody detection in the sera of the infected dogs. The results indicate 100% (36/36 specimens) detection of both antigen and antibody. Cross reactivity with Histoplasma capsulatum, as well as non-specific binding with the normal urine specimens, was observed with the competitive binding inhibition ELISA.
Elizabeth M. Brown - One of the best experts on this subject based on the ideXlab platform.
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Correction: Phylogenetic Analysis Reveals a Cryptic Species Blastomyces gilchristii, sp. nov. within the Human Pathogenic Fungus Blastomyces Dermatitidis
2016Co-Authors: Elizabeth M. Brown, Sean X. Zhang, Lisa R. Mctaggart, Donald E. Low, David A. Stevens, Susan E. RichardsonAbstract:Correction: Phylogenetic Analysis Reveals a Cryptic Species Blastomyces gilchristii, sp. nov. within the Human Pathogenic Fungus Blastomyces Dermatitidis
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Effective method for the heat inactivation of Blastomyces Dermatitidis.
Medical mycology, 2014Co-Authors: Elizabeth M. Brown, Lisa R. Mctaggart, Donald E. Low, Susan E. RichardsonAbstract:Manipulation of Blastomyces Dermatitidis requires the use of containment level 3 (CL3) practices. However, access to CL3 laboratories is limited and working conditions are restrictive. We describe the validation of a "heat-killing" method to inactivate B. Dermatitidis, thus allowing cellular material to be removed from the CL3 laboratory for subsequent DNA isolation that is suitable for genetic applications.
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phylogenetic analysis reveals a cryptic species Blastomyces gilchristii sp nov within the human pathogenic fungus Blastomyces Dermatitidis
PLOS ONE, 2013Co-Authors: Sean X. Zhang, Elizabeth M. Brown, Lisa R. Mctaggart, Donald E. Low, David A. Stevens, Susan E. RichardsonAbstract:Background Analysis of the population genetic structure of microbial species is of fundamental importance to many scientific disciplines because it can identify cryptic species, reveal reproductive mode, and elucidate processes that contribute to pathogen evolution. Here, we examined the population genetic structure and geographic differentiation of the sexual, dimorphic fungus Blastomyces Dermatitidis, the causative agent of blastomycosis. Methodology/Principal Findings Criteria for Genealogical Concordance Phylogenetic Species Recognition (GCPSR) applied to seven nuclear loci (arf6, chs2, drk1, fads, pyrF, tub1, and its-2) from 78 clinical and environmental isolates identified two previously unrecognized phylogenetic species. Four of seven single gene phylogenies examined (chs2, drk1, pyrF, and its-2) supported the separation of Phylogenetic Species 1 (PS1) and Phylogenetic Species 2 (PS2) which were also well differentiated in the concatenated chs2-drk1-fads-pyrF-tub1-arf6-its2 genealogy with all isolates falling into one of two evolutionarily independent lineages. Phylogenetic species were genetically distinct with interspecific divergence 4-fold greater than intraspecific divergence and a high Fst value (0.772, P<0.001) indicative of restricted gene flow between PS1 and PS2. Whereas panmixia expected of a single freely recombining population was not observed, recombination was detected when PS1 and PS2 were assessed separately, suggesting reproductive isolation. Random mating among PS1 isolates, which were distributed across North America, was only detected after partitioning isolates into six geographic regions. The PS2 population, found predominantly in the hyper-endemic regions of northwestern Ontario, Wisconsin, and Minnesota, contained a substantial clonal component with random mating detected only among unique genotypes in the population. Conclusions/Significance These analyses provide evidence for a genetically divergent clade within Blastomyces Dermatitidis, which we use to describe a novel species, Blastomyces gilchristii sp. nov. In addition, we discuss the value of population genetic and phylogenetic analyses as a foundation for disease surveillance, understanding pathogen evolution, and discerning phenotypic differences between phylogenetic species.
David A. Stevens - One of the best experts on this subject based on the ideXlab platform.
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Correction: Phylogenetic Analysis Reveals a Cryptic Species Blastomyces gilchristii, sp. nov. within the Human Pathogenic Fungus Blastomyces Dermatitidis
2016Co-Authors: Elizabeth M. Brown, Sean X. Zhang, Lisa R. Mctaggart, Donald E. Low, David A. Stevens, Susan E. RichardsonAbstract:Correction: Phylogenetic Analysis Reveals a Cryptic Species Blastomyces gilchristii, sp. nov. within the Human Pathogenic Fungus Blastomyces Dermatitidis
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phylogenetic analysis reveals a cryptic species Blastomyces gilchristii sp nov within the human pathogenic fungus Blastomyces Dermatitidis
PLOS ONE, 2013Co-Authors: Sean X. Zhang, Elizabeth M. Brown, Lisa R. Mctaggart, Donald E. Low, David A. Stevens, Susan E. RichardsonAbstract:Background Analysis of the population genetic structure of microbial species is of fundamental importance to many scientific disciplines because it can identify cryptic species, reveal reproductive mode, and elucidate processes that contribute to pathogen evolution. Here, we examined the population genetic structure and geographic differentiation of the sexual, dimorphic fungus Blastomyces Dermatitidis, the causative agent of blastomycosis. Methodology/Principal Findings Criteria for Genealogical Concordance Phylogenetic Species Recognition (GCPSR) applied to seven nuclear loci (arf6, chs2, drk1, fads, pyrF, tub1, and its-2) from 78 clinical and environmental isolates identified two previously unrecognized phylogenetic species. Four of seven single gene phylogenies examined (chs2, drk1, pyrF, and its-2) supported the separation of Phylogenetic Species 1 (PS1) and Phylogenetic Species 2 (PS2) which were also well differentiated in the concatenated chs2-drk1-fads-pyrF-tub1-arf6-its2 genealogy with all isolates falling into one of two evolutionarily independent lineages. Phylogenetic species were genetically distinct with interspecific divergence 4-fold greater than intraspecific divergence and a high Fst value (0.772, P<0.001) indicative of restricted gene flow between PS1 and PS2. Whereas panmixia expected of a single freely recombining population was not observed, recombination was detected when PS1 and PS2 were assessed separately, suggesting reproductive isolation. Random mating among PS1 isolates, which were distributed across North America, was only detected after partitioning isolates into six geographic regions. The PS2 population, found predominantly in the hyper-endemic regions of northwestern Ontario, Wisconsin, and Minnesota, contained a substantial clonal component with random mating detected only among unique genotypes in the population. Conclusions/Significance These analyses provide evidence for a genetically divergent clade within Blastomyces Dermatitidis, which we use to describe a novel species, Blastomyces gilchristii sp. nov. In addition, we discuss the value of population genetic and phylogenetic analyses as a foundation for disease surveillance, understanding pathogen evolution, and discerning phenotypic differences between phylogenetic species.
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molecular epidemiology of Blastomyces Dermatitidis
Clinical Infectious Diseases, 2000Co-Authors: David A. Stevens, Arthur F. Disalvo, Michael Mccullough, Karl V Clemons, Pilsang ParkAbstract:The inhalation of conidia of Blastomyces Dermatitidis, a fungus found in soil, causes disease in humans and animals. We studied the genetic diversity of this pathogen by extracting DNA yeasts and analyzing them with a polymerase chain reaction (PCR)-based typing system we developed, which used restriction fragment analysis of amplicons from the regions between the rDNA repeats and allowed us to class isolates into 3 major groups. Strains were further differentiated by use of PCR fingerprinting with 3 different primers. Fifty-nine isolates collected over 35 years from 15 regions (United States, India, Africa, Canada) were analyzed. Genotypic groups A, B, and C contained 17, 23, and 19 isolates, which were divided into 5, 15, and 12 types, respectively. All 16 isolates from North America in group A were from the upper midwestern United States or Canada, whereas 0 of 20 isolates from the southeastern United States were in group A. Studies of the largest collection from 1 locale (Eagle River, WI), revealed that the soil isolates studied were not responsible for the majority of cases in this outbreak, as previously proposed, and that >1 strain was present in the environment and in patients. Overall, these results provide a tool for the epidemiological study of blastomycosis and illuminate the genetic and geographic diversity of this important pathogen.
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variable colonial phenotypic expression and comparison to nuclei number in Blastomyces Dermatitidis
Medical Mycology, 1991Co-Authors: Karl V Clemons, Steven Michael Hurley, L G Treatclemons, David A. StevensAbstract:We examined colonial phenotypes of five isolates of Blastomyces Dermatitidis at 33, 35 and 37°C on four growth media. Three different colony types were identified: yeast, mycelial, and a mixed type consisting of both yeast and mycelia. Each isolate varied in its ability to grow on the different media and at different temperatures, and in the types of colonies it produced in the various temperaturemedia combinations. Quantification of the number of nuclei per yeast cell by fluorescent staining revealed no correlation between the number of nuclei per cell and the colonial phenotype. These results indicate that the colonial phenotype of B. Dermatitidis varies with the isolate as well as with temperature and culture medium, but is not correlated with the number of nuclei per yeast. These findings could provide a start towards typing B. Dermatitidis isolates.