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PubMed2002

Gel shift assay of nuclear extracts from Histoplasma capsulatum demonstrates the presence of several DNA binding proteins.

A gel shift assay was optimized to detect several general DNA binding proteins from Histoplasma capsulatum strain G217B. The electrophoretic mobility shift assay (EMSA) technique also detected protein(s) recognizing a pyrimidine-rich motif found in several Histoplasma promoters. Establishment of EMSA conditions provides an important framework to evaluate regulation of homeostatic or phase-specific genes that may influence virulence in Histoplasma and other dimorphic fungal pathogens.

Bacterial Proteins↗
PubMed1986

Evaluation of cross-reactions in Histoplasma capsulatum serologic tests.

Cross-reactivity in Histoplasma serologic tests was evaluated by using sera from patients with histoplasmosis and other infections. Serum samples from 127 of 134 (95%) patients with histoplasmosis were judged positive by complement fixation tests, and 121 (90%) showed H bands, M bands, or both by immunodiffusion. Of these 134 patients, cross-reactions were seen to Blastomyces dermatitidis in 53 patients (40%), to Coccidioides immitis in 20 patients (16%), and to Aspergillus fumigatus in 3 patients (2%) by complement fixation. Serum samples from 5 of 99 patients (5%) with other fungal infections and from 5 of 46 patients (11%) with tuberculosis had M precipitin bands by the Histoplasma immunodiffusion test, whereas none of the 123 sera from patients with other bacterial, Mycoplasma, or viral infections showed H or M precipitin bands. In the complement fixation test, positive reactions were observed in 16 of 90 patients (18%) with other fungal infections, in 14 of 41 patients (34%) with tuberculosis, and in 18 of 105 patients (17%) with other bacterial, Mycoplasma, or viral infections. Positive reactions were seen by radioimmunoassay in 54 of 110 patients (49%) with other fungal infections, in 23 of 46 patients (50%) with tuberculosis, and in 35 of 123 patients (28%) with with other bacterial, Mycoplasma, or viral infections. These results demonstrate a wider range of cross-reactions in Histoplasma serology than has been previously recognized, and the cross-reactivity was greatest when observed by radioimmunoassay. Caution should be exercised in the interpretation of serologic data from patients with suspected fungal infections.

Antibodies, Fungal↗
PubMed1992

Typing of Histoplasma capsulatum by restriction fragment length polymorphisms in a nuclear gene.

We previously described yps-3, a Histoplasma-specific nuclear gene probe useful in the identification of Histoplasma capsulatum. By using restriction fragment length polymorphisms (RFLPs) of DNA detected by the yps-3 gene and mitochondrial DNA, 76 clinical and soil isolates of H. capsulatum were classified. The majority of North American isolates obtained from endemic regions of the midwestern United States were members of the previously characterized class 2, although four clinical isolates from different patients with AIDS from that region were grouped in class 1 with the temperature-sensitive Downs strain. A Florida soil isolate (FLS1) was placed in class 4 on the basis of RFLP with both probes. Two American Type Culture Collection strains (G184B and G186B) from Panama were grouped into class 3 by this analysis. A group of five H. capsulatum isolates obtained from patients with AIDS in New York City were typed into a new class 5 on the basis of yps-3 polymorphisms; those organisms fell into two broad mitochondrial DNA patterns, designated 5b and 5c. Two new isolates from Panama were also members of this broad yps-3 class 5 group, but they exhibited a distinct mitochondrial DNA profile (class 5a). A sixth class was detected in DNA obtained from a patient with AIDS from Panama; that DNA had unique RFLP profiles with respect to both probes. These observations suggest that the Histoplasma-specific yps-3 gene probe is a sensitive tool for typing H. capsulatum in clinical specimens. Additionally, these studies provide molecular support for the hypothesis that AIDS-associated histoplasmosis in nonendemic areas is due to reactivation of a previously acquired infection.

Acquired Immunodeficiency Syndrome↗
PubMed2002

Utilization and cell-surface binding of hemin by Histoplasma capsulatum.

Histoplasma capsulatum, a dimorphic fungus capable of causing severe respiratory illness in immunocompromised individuals, resides in macrophages during mammalian infection. Previous studies suggest that siderophore-mediated iron transport may be important for the acquisition of iron from transferrin while the organism resides in macrophages. However, iron is also present as hemin in the intracellular environment of the macrophage and may serve as a major source of iron during infection. Thus the ability of H. capsulatum to use hemin and heme-containing compounds was examined. Histoplasma capsulatum G217B was iron-starved by adding the iron chelator deferoxamine mesylate to the culture. The addition of 10 microM hemin in the presence of deferoxamine mesylate restored growth to the levels seen in the absence of the chelator. Histoplasma capsulatum was also cultivated in an iron-limited, chemically defined medium without the addition of chelators and it was determined that the organism could also use hemoglobin as a sole source of iron. The method of iron internalization from heme was examined by measuring hemin binding to the yeast-cell surface. The ability of H. capsulatum to bind hemin was related to the nutritional status of the cells. Cells grown under iron-limited conditions bound more heme to the cell surface than did cells grown in medium without chelator. Pretreatment of iron-starved cells with proteinase K eliminated the ability of the organism to bind hemin. Additionally, the pre-incubation of iron-starved H. capsulatum with hemin eliminated the ability of these cells to remove hemin from the solution, although pre-incubation of cells with the iron-free form of hemin, protoporphyrin IX, only modestly affected the ability of the organism to bind hemin. These results suggest that H. capsulatum uses hemin as a sole source of iron and that one mechanism of iron acquisition involves a cell-surface receptor for hemin.

Culture Media↗
PubMedDate not supplied

[The use of exoantigens for identifying Histoplasma capsulatum].

The exoantigen test has been successfully used in the last years for the immunological identification of pathogenic fungi. Twenty seven straing presumptively identified as Histoplasma capsulatum by morphologic methods, were studied by such test. The exoantigens obtained from each strain were studied by immunoprecipitation technique in front of histoplasmosis control sera experimentally obtained in rabbits. Reference strains of Histoplasma capsulatum. Histoplasma duboisii and Paracoccidioides brasiliensis were used. Of the strains studied, 96% was identified as H. capsulatum by the exoantigen test, a simple, rapid and highly specific technique greatly useful in the laboratories of Microbiology.

Antigens, Fungal↗
PubMed1986

Experimentally induced Histoplasma capsulatum infection in coyotes and a dog.

Captive coyotes (Canis latrans) were inoculated intragastrically (1 coyote), and intratracheally (6 coyotes) with Histoplasma capsulatum to simulate natural routes of exposure. In addition, the infectious organism was inoculated IV into another coyote and into a dog (C familiaris) to simulate hematogenous dissemination of H capsulatum. Isolation of the pathogen from various tissue specimens, using direct plating and mouse inoculation methods, provided mycologic evidence of infection. The presence of anti-Histoplasma serum antibodies also was monitored. Of the 6 coyotes challenged intratracheally, one developed a benign pulmonary form of histoplasmosis. This coyote developed a transient cough, and intracellular yeasts typical of H capsulatum were observed microscopically in pulmonary tissue specimens. The pathogen was isolated from specimens of pulmonary tissue and associated lymph nodes. The coyote that was fed Histoplasma-infected mice (intragastric challenge) did not develop clinical disease. The coyote and the dog inoculated IV developed a multifocal, nonsuppurative infection, without clinical evidence of histoplasmosis. Pathologic findings in the coyote and dog were similar; both animals had splenitis, lymphadenitis, and petechiation of the lungs. The liver and spleen were considered the primary extrapulmonary target organs on the basis of mycologic and histologic findings.

Animals↗
PubMed1985

Cerebrospinal fluid Histoplasma antibodies in central nervous system histoplasmosis.

We have evaluated the Histoplasma antibody response in the cerebrospinal fluid (CSF) in nine patients with central nervous system histoplasmosis and 98 controls. While the CSF Histoplasma antibody response identified eight of the nine patients, CSF cultures were positive in only two. Of controls with histoplasmosis but without meningitis (13 patients), or without histoplasmosis (85 patients), elevated CSF antibodies were detected by complement fixation in seven, by IgG radioimmunoassay in 17, and by IgM radioimmunoassay in five. Measurement of the CSF Histoplasma antibody response appears useful for identifying meningitis in patients with histoplasmosis, although cross-reactions occur in half of patients with other forms of chronic fungal meningitis. Patients with these other infections can usually be identified by tests for CSF Coccidioides antibodies, or cryptococcal antigens.

Antibody Formation↗
PubMedDate not supplied

[Digestive disorders in histoplasmosis. Report of a case of ulcerous enterocolitis caused by Histoplasma capsulatum (author's transl)].

Report of a new American histoplasmosis caused by Histoplasma capsulatum in a young adult Haitian, and review of the digestive disorders as reported in the previous medical papers. In the present case, the first localization detected was a granulomatous hepatitis which lead to the disclosure of an ulcerous entercolitis. Tuberculosis and Crohn disease were first suspected, but their treatment induced a worse turn of the patient's condition and compelled to a laparotomy, which shown ulcerous entercolitis with associated lymph-nodes. Per operative pathological examination gave evidence of Histoplasma capsulatum yeasts. Treatment, first with miconazole and later on with amphotericin B could not prevent the death of the patient affected by a deficiency of cell mediated immunity. Disseminated histoplasmosis is rare: 1 out of 5,000 cases of American histoplasmosis-digestive disorders are generally detected by surgery or post-mortem examination. They occur in 20 p. 100 of the cases envolving the whole digestive tract from mouth to anus, giving mucous ulcerations, difficult to detect by X ray. Endoscopic control with biopsies has therefore a great value. In Histoplasma duboisii, histoplasmosis digestive disorders are very unusual and localized to intestine. They may be detected by surgery, or by endoscopies. In the reported case, in spite of the geographical origin of the patient, the diagnosis of American histoplasmosis was not retained because of the negativity of the immunological tests and of the biopsies performed before surgery. An afterward checking of these biopsy specimens gave evidence of a few yeasts. Special dyeing technique may have made the diagnosis somewhat earlier; this is important because new non toxic imidazole drugs seem to be active against these yeasts.

Adult↗
PubMed1980

[A case of disseminated histoplasmosis due to Histoplasma capsulatum (author's transl)].

A case is reported of American histoplasmosis due to Histoplasma capsulatum presenting as a hepatic granulomatosis. The patient was a young adult from Haiti and investigations revealed the presence of an ulcerous enterocolitic suggesting tuberculosis or Crohn's disease. The condition became worse after antituberculous therapy and corticoids, and operation was necessary. Extemporaneous histological examination, of the colonic ulcerations showed Histoplasma capsultaum yeasts. The patient, who had a cellular immunity deficiency, died during antifungal treatment with amphotericin B of the disseminated histoplasmosis is rare: 1 case in 5000 American histoplasmosis cases. Digestive tract lesions, usually detected during operation or autopsy, are found in about 20% of cases. In the present observations the diagnosis of American histoplasmosis was based on the country of origin of the patient, but this was not confirmed: the immunological reactions were and remained negative and no yeasts were demonstrated after staining of pre-operative biopsy specimens (colon and liver particularly). Also, serial sections conducted after the diagnosis had been established demonstrated the presence of Histoplasma capsulatum in the colon sample removed during colonoscopy.

Adult↗
PubMed1962

Conversion in strains of Histoplasma capsulatum.

Pine, Leo (Duke University, Durham, N. C.) and Robert E. Webster. Conversion in strains of Histoplasma capsulatum. J. Bacteriol. 83:149-157. 1962-Photographs of mycelial elements taken at various time intervals from a single strain of Histoplasma capsulatum have shown the direct conversion of microconidia to yeast cells by polar and nonpolar budding, the formation of yeast cells by budding of hyphal cells, and the probable conversion of hyphal cells to yeast phase by monilial chain formation. In addition, studies of fixed-slide preparations of a second strain have indicated the formation of stalked yeast cells arising in a manner similar to that observed for microconidia formation but, apparently, by-passing the microconidial stage. These observations are discussed briefly in regard to temperature-induced changes in cell-wall synthesis during conversion.

Histoplasma↗
PubMed1962

In vitro conversion of Histoplasma capsulatum.

Northey, W. T. (Arizona State University, Tempe) and L. D. Brooks. In vitro conversion of Histoplasma capsulatum. J. Bacteriol. 84:577-580. 1962.-The in vitro fungus-yeast conversion of the diphasic fungus, Histoplasma capsulatum, has been accomplished using enzymatic digests of casein. Of these casein digests, the most rapid and complete conversion was found when Ferm-Amine Types I, II, and III were used. Technical grades of these products were found to stimulate conversion to a greater extent than the reagent grades. The use of these media as research and diagnostic aids is suggested.

Caseins↗
PubMed1963

SEROLOGICAL CHARACTERIZATION OF PATHOGENIC FUNGI BY MEANS OF FLUORESCENT ANTIBODIES. I. ANTIGENIC RELATIONSHIPS BETWEEN YEAST AND MYCELIAL FORMS OF HISTOPLASMA CAPSULATUM AND BLASTOMYCES DERMATITIDIS.

Kaufman, Leo (Communicable Disease Center, Atlanta, Ga.) and William Kaplan. Serological characterization of pathogenic fungi by means of fluorescent antibodies. I. Antigenic relationships between yeast and mycelial forms of Histoplasma capsulatum and Blastomyces dermatitidis. J. Bacteriol. 85:986-991. 1963.-Four antiglobulins prepared against the yeast and mycelial forms of Histoplasma capsulatum and Blastomyces dermatitidis were labeled with fluorescein isothiocyanate. Cross-staining and adsorption techniques enabled the discernment of five antigenic factors in the fungi studied. The yeast-phase cells of H. capsulatum and B. dermatitidis contained the respective distinct antigens B and D, and a common factor E. In addition, the yeast-phase cells of these two species shared antigen C with the mycelial elements of H. capsulatum. The mycelial cells of H. capsulatum and B. dermatitidis shared antigen A with the yeast-phase cells of these two organisms and contained no distinct antigens.

Antigens↗
PubMed1964

FLUORESCENT-ANTIBODY STUDIES OF THE MYCELIAL FORM OF HISTOPLASMA CAPSULATUM AND MORPHOLOGICALLY SIMILAR FUNGI.

Kaufman, Leo (Communicable Disease Center, Atlanta, Ga.), and Brenda Brandt. Fluorescent-antibody studies of the mycelial form of Histoplasma capsulatum and morphologically similar fungi. J. Bacteriol. 87:120-126. 1964.-Fluorescent-antibody reagents were developed to differentiate the mycelial form of the pathogen, Histoplasma capsulatum, from several morphologically similar but saprophytic species of the genera Sepedonium and Chrysosporium. Cross-staining and adsorption procedures revealed antigenic relationships among certain species of these three genera and other fungi. The significance of these observations to the diagnostician and taxonomist is discussed.

Coloring Agents↗
PubMed1964

INTRACELLULAR BEHAVIOR OF HISTOPLASMA CAPSULATUM.

Howard, Dexter H. (University of California, Los Angeles). Intracellular behavior of Histoplasma capsulatum. J. Bacteriol. 87:33-38. 1964.-The tinctorial characteristics and intracellular generation time of Histoplasma capsulatum within mouse histiocytes maintained in tissue culture were studied. The appearance of viable and nonviable cells of H. capsulatum was observed after staining with the May Grunwald-Giemsa, Methenamine Silver, and Periodic acid-Schiff (PAS) procedures. Death of the fungus, either induced by heat or occurring naturally in saline suspension, caused a loss of nuclear differentiation. This loss was not accompanied by changes in the PAS or Silver staining capacities of the cell walls of the fungus. There was no tinctorial evidence for intracellular death of H. capsulatum in normal mouse histiocytes. The intracellular generation time of the fungus was estimated to be 10.3 +/- 1.5 hr, and was not affected by the age of the histiocytes. The generation time was approximately the same in histiocytes from Webster-Swiss and A/J mice.

Animals↗
PubMed1965

INTRACELLULAR GROWTH OF HISTOPLASMA CAPSULATUM.

Howard, Dexter H. (University of California, Los Angeles). Intracellular growth of Histoplasma capsulatum. J. Bacteriol. 89:518-523. 1965.-Under a variety of experimental conditions, the generation time of Histoplasma capsulatum in mammalian histiocytes was remarkably constant. The rate of intracellular growth of five different isolates of H. capsulatum was very similar. The generation time of the fungus within guinea pig cells was nearly the same as it was within mouse cells under comparable conditions. Preliminary exposure of yeast cells to specific antibodies and complement did not affect intracellular proliferation of the fungus during the first 24 hr after phagocytosis by mouse histiocytes. The rate of growth of the fungus within cells from immunized mice and guinea pigs was comparable to that in cells from normal animals. This latter result was not altered by preliminary exposure of the fungus to hyperimmune serum and complement.

Animals↗
PubMed1988

In situ localization of antigens of Histoplasma capsulatum using colloidal gold immune electron microscopy.

Histoplasma capsulatum contains multiple antigens, among them the H antigen and M antigen, which are useful in serologic testing for histoplasmosis. We prepared 7 mouse monoclonal antibodies (5 IgG, 2 IgM) to histoplasmin, and compared these with polyclonal histoplasmin antibodies raised in rabbits and mice. Both monoclonal and polyclonal antibodies were high titered by ELISA. Colloidal gold immune electron microscopy (CGIEM) showed that polyclonal antibodies to histoplasmin or H antigen bound at multiple sites in the cell wall, cytoplasm, and nucleus of Histoplasma yeast cells. In contrast, antibodies to M antigen selectively label the cell membrane and antibodies to alkali soluble cell wall antigen label only the cell wall. Polyclonal antibodies cross reacted extensively with other fungi, both by ELISA and CGIEM. Monoclonal antibodies stained only cytoplasmic epitopes, but also cross reacted with other fungi by electron microscopy. Only periodate treated H antigen elicited polyclonal antibodies which were more specific than those of untreated H antigen or histoplasmin.

Animals↗
PubMed1986

Immunocytochemical staining of Histoplasma capsulatum at the electron microscopic level.

Rabbits were immunized with histoplasmin emulsified in Freund's complete adjuvant. Antibody raised in these rabbits was exposed to Histoplasma capsulatum yeast cells, either in tissue culture medium, or after in vitro or in vivo phagocytosis by mouse macrophages. The sites of antibody binding were identified using an immunoperoxidase technique. At least two sites of antibody binding were identified, one to the fungal cell wall and the other to the outer cell membrane. Within 6 h after phagocytosis by macrophages, fungal cell walls appeared roughened, with what appeared to be cell wall antigen released into the phagolysosome, appearing associated with the phagolysosome membrane, and possibly adjacent macrophage cytoplasm. Similar staining of fungal antigen was noted in alveolar macrophages which had ingested Histoplasma capsulatum after a respiratory challenge. This method may be useful in detailing the host/pathogen interactions which occur in human pulmonary histoplasmosis.

Animals↗
PubMed1980

Non-specific incorporation of nucleic acid precursors in Blastomyces dermatitidis and Histoplasma capsulatum.

Incorporation of thymidine, thymidine monophosphate (TMP), thymidine triphosphate (TTP), uridine and orotic acid into DNA, RNA and protein in Blastomyces dermatitidis and Histoplasma capsulatum was studied utilizing a specific acid hydrolysis technique developed for these fungi. Thymidine was incorporated to the greatest extent (approximately 0.5% of added label) followed by uridine, orotic acid, TMP and TTP. In Blastomyces, uridine and orotic acid labeled primarily RNA. TMP and TTP labeled RNA, DNA and protein at nearly the same level. In Histoplasma RNA was labeled poorly by any of these precursors. TMP and TTP labeled DNA predominately and protein to a slightly lower level. Deoxyadenosine or uridine media supplements of 250 micrograms/ml did not enhance incorporation. All precursors tested were found to be nonspecific in that RNA, DNA and protein were labeled. All data indicate that neither RNA nor DNA synthesis can be specifically measured in whole cells or acid precipitates by any of these precursors. Specific radiometric monitoring with these isotopes therefore requires the separation of these macromolecules.

Animals↗