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J D Nosanchuk

Publications and source records attributed to J D Nosanchuk.

At least 19 recordsLinked to original sources
PubMed2005

Protective antibodies and endemic dimorphic fungi.

The host response to infection is the outcome of a complex interaction between a microbe and a host's innate and adaptive immune system. In this context, the role of antibody in the endemic mycoses is relatively poorly understood. Recently, a monoclonal antibody to a cell surface protein has been shown to be protective in a murine histoplasmosis model. The findings with Histoplasma capsulatum may provide a paradigm for antibody protection against endemic fungi. This paper reviews the recent data on protective antibody in histoplasmosis and previous data supporting a role for antibody in protective responses in other dimorphic fungi.

Animals↗
PubMed2004

Evaluation of acute hematologic and long-term pulmonary toxicities of radioimmunotherapy of Cryptococcus neoformans infection in murine models.

We evaluated acute hematological and long-term pulmonary toxicity of radioimmunotherapy in murine models of Cryptococcus neoformans infection. Activities up to 250 microCi were well tolerated by healthy A/JCr mice for (213)Bi-18B7 and (188)Re-18B7 monoclonal antibodies. In infected mice, doses up to 150 microCi produced only transient toxicity. The lungs of treated mice had no evidence of radiation fibrosis.

Animals↗
PubMed2004

Feasibility of radioimmunotherapy of experimental pneumococcal infection.

Streptococcus pneumoniae is an important cause of community-acquired pneumonia, meningitis, and bacteremia. The problem of pneumococcal disease is exacerbated by increasing drug resistance. Furthermore, patients with impaired immunity are at high risk for invasive pneumococcal infections. Thus, there is an urgent need for new approaches to antimicrobial therapy. Antibody therapies take advantage of the specificity and high affinity of the antigen-antibody interaction to deliver antibacterial compounds to a site of infection in the form of naked or conjugated antibodies. We have recently established that radioimmunotherapy (RIT) can be used to treat experimental fungal infections in mice. In the present study, we investigated the feasibility of applying a RIT approach to the treatment of S. pneumoniae infection by evaluating the susceptibility of S. pneumoniae to radiolabeled antibody in vitro and in an animal infection model. For the specific antibody carrier, we used human monoclonal antibody D11, which binds to pneumococcal capsular polysaccharide 8. We have selected the alpha particle emitter (213)Bi as the radionuclide for conjugation to the antibody. Incubation of serotype 8 S. pneumoniae with (213)Bi-D11 resulted in dose-dependent killing of bacteria. RIT of S. pneumoniae infection in C57BL/6 mice showed that 60% more mice survived in the (213)Bi-D11-treated group (80 micro Ci) than in the untreated group (P < 0.01). The treatment did not cause hematological toxicity, as demonstrated by platelet counts. This feasibility study establishes that RIT can be applied to the treatment of bacterial infections.

Animals↗
PubMed2004

Scytalidium dimidiatum causing recalcitrant subcutaneous lesions produces melanin.

Scytalidium dimidiatum is a pigmented dematiaceous coelomycete that typically causes chronic superficial skin diseases and onychomycosis, as well as deeper infections, such as subcutaneous abscesses, mycetoma, and even fungemia in immunocompromised patients. A second species, Scytalidium hyalinum, has hyaline hyphae and arthroconidia and is considered by some authors to be an albino mutant of S. dimidiatum. This study aimed to confirm the presence of melanin or melanin-like compounds (which have been previously implicated in the virulence of other fungal pathogens) in S. dimidiatum from a patient with multiple subcutaneous nodules. Treatment of the hyphae and arthroconidia with proteolytic enzymes, denaturant, and concentrated hot acid yielded dark particles, which were stable free radicals, consistent with their identification as melanins. Extracted melanin particles from S. dimidiatum cultures were labeled by melanin-binding monoclonal antibodies (MAbs) from Sporothrix schenckii, Aspergillus fumigatus, and Cryptococcus neoformans. Lesional skin from the patient infected with S. dimidiatum contained fungal cells that were labeled by melanin-binding MAbs, and digestion of the tissue yielded dark particles that were also reactive. S. hyalinum was also subjected to the melanin extraction protocol, but no dark particles were yielded.

Aged↗
PubMed2002

Activation of the alternative complement pathway by fungal melanins.

Melanins are complex biological pigments formed by the oxidative polymerization of phenolic and/or indolic compounds. These pigments have been implicated in the pathogenesis of some microbial infections, malignancies, degenerative disorders, and autoimmune diseases. Recent studies have demonstrated that melanins have antigenic and anti-inflammatory properties. These findings led us to further explore the interaction of melanins with the immune system. Melanin particles ("ghosts") were isolated from in vitro-melanized Cryptococcus neoformans cells and Aspergillus niger conidia and then incubated in normal human serum containing (125)I-labeled complement C3. The results demonstrated deposition of C3 fragments onto the melanin ghosts as early as 1 min after incubation, with maximum deposition occurring after 12 min for C. neoformans-derived melanin ghosts and after 25 min for A. niger-derived melanin ghosts. The blocking of classical pathway activation did not affect the kinetics or total deposition of C3 onto the melanin ghosts, indicating that melanins activate complement through the alternative pathway. Immunofluorescence analysis of lungs from BALB/c mice injected intratracheally with C. neoformans-derived melanin ghosts demonstrated deposition of C3 fragments onto the ghosts. Small granulomas were also observed surrounding the ghosts. However, melanization of the C. neoformans cell wall did not alter the kinetics or total deposition of C3 fragments onto the fungal cells. The finding that melanin surfaces can activate the complement system suggests a potential mechanism for the pathogenesis of some degenerative and/or autoimmune processes that involve melanized cells as well as another potential role for melanin in the virulence of melanin-producing microorganisms.

Animals↗
PubMed2001

Glyphosate inhibits melanization of Cryptococcus neoformans and prolongs survival of mice after systemic infection.

Cryptococcus neoformans is a major fungal pathogen and is a relatively common cause of life-threatening meningoencephalitis. Glyphosate is a widely used herbicide that inhibits the synthesis of aromatic amino acids via the shikimate acid pathway. This study investigated the effects of glyphosate on C. neoformans growth, melanization, and murine infection. C. neoformans was relatively resistant to glyphosate, requiring concentrations >250 microg/mL for inhibition. Melanization of C. neoformans in the presence of L-dopa was inhibited by subinhibitory concentrations of glyphosate. Glyphosate inhibited autopolymerization of L-dopa and oxidation of L-epinephrine by cryptococcal cells, which is mediated by a laccase. Administration of glyphosate to mice infected with C. neoformans delayed melanization of yeast cells in vivo and prolonged average mouse survival. The results suggest that inhibition of melanization in vivo may facilitate control of C. neoformans infection.

Animals↗
PubMed2001

Detection of melanin-like pigments in the dimorphic fungal pathogen Paracoccidioides brasiliensis in vitro and during infection.

Melanins are implicated in the pathogenesis of several human diseases, including some microbial infections. In this study, we analyzed whether the conidia and the yeasts of the thermally dimorphic fungal pathogen Paracoccidioides brasiliensis produce melanin or melanin-like compounds in vitro and during infection. Growth of P. brasiliensis mycelia on water agar alone produced pigmented conidia, and growth of yeasts in minimal medium with L-3,4-dihydroxyphenylalanine (L-DOPA) produced pigmented cells. Digestion of the pigmented conidia and yeasts with proteolytic enzymes, denaturant, and hot concentrated acid yielded dark particles that were the same size and shape as their propagules. Immunofluorescence analysis demonstrated reactivity of a melanin-binding monoclonal antibody (MAb) with the pigmented conidia, yeasts, and particles. Electron spin resonance spectroscopy identified the yeast-derived particles produced in vitro when P. brasiliensis was grown in L-DOPA medium as a melanin-like compound. Nonreducing polyacrylamide gel electrophoresis of cytoplasmic yeast extract revealed a protein that catalyzed melanin synthesis from L-DOPA. The melanin binding MAb reacted with yeast cells in tissue from mice infected with P. brasiliensis. Finally digestion of infected tissue liberated particles reactive to the melanin binding MAb that had the typical morphology of P. brasiliensis yeasts. These data strongly suggest that P. brasiliensis propagules, both conidia and yeast cells, can produce melanin or melanin-like compounds in vitro and in vivo. Based on what is known about the function of melanin in the virulence of other fungi, this pigment may play a role in the pathogenesis of paracoccidioidomycosis.

Animals↗
PubMed2000

Isolation and serological analyses of fungal melanins.

Melanins are notoriously difficult to work with because of their unique physical and chemical properties. The study of melanins is hampered by the scarcity of melanin-specific reagents and serological techniques. In this study we describe modifications to the standard method for the isolation of melanins from in vitro-melanized fungal cells and detail the optimization of serological techniques for the study of melanin compounds. The isolation procedure involves the digestion of melanized cells with a combination of proteolytic and glycolytic enzymes, denaturant, organic extractions, and boiling in 6.0 M HCl. Elemental quantitative analyses suggest that this procedure does not significantly affect the relative elemental composition of melanins. For the serological assays, our goal was to achieve a homogenous distribution of melanin particles on a solid support to maximize their recognition by melanin-binding antibodies. The results from enzyme-linked immunosorbent assays (ELISAs) demonstrate that melanins, in general, disperse more efficiently on, and adhere better to, medium-binding polystyrene surfaces, especially in the presence of trace amounts of salt. Blocking the melanin-coated ELISA plates with the commercially available SuperBlock((R)) Blocking Buffer for 4 h was more efficient at reducing non-specific binding of a negative control monoclonal antibody (mAb) compared to blocking with 2% bovine serum albumin (BSA) and 5% milk. Increasing the ionic strength of the antibody solutions reduced binding to the melanins, indicating that binding is in part mediated by electrostatic interactions. These conditions were also applied to immunofluorescence (IF) analyses of melanins, and the results were consistent with those obtained by ELISA.

Antibodies↗
PubMed2000

Melanisation of Cryptococcus neoformans in human brain tissue.

Melanin-specific reagents revealed melanin in cryptococcal cell walls from human brain tissue, and fungal-cell melanin "ghosts" were recovered from infected tissue. The results indicate that Cryptococcus neoformans melanises during human infection.

Amphotericin B↗
PubMed2000

Evidence of zoonotic transmission of Cryptococcus neoformans from a pet cockatoo to an immunocompromised patient.

BACKGROUND: Although cryptococcosis has been associated with birds for almost 50 years, point sources for infection have not been identified. OBJECTIVE: To document zoonotic transmission of Cryptococcus neoformans. DESIGN: Case report. SETTING: A home in Boston, Massachusetts. PATIENT: A 72-year-old woman who received a diagnosis of cryptococcal meningitis in November 1998. The patient, who had been taking immunosuppressant drugs since undergoing renal transplantation in 1989, owned a pet cockatoo. MEASUREMENTS: Cryptococcus neoformans was isolated from the feces of the cockatoo. Isolates from excreta and from the patient were compared by using biochemical profiles, monoclonal antibody binding patterns, restriction fragment length polymorphism analysis, and karyotyping. RESULTS: The isolates from the patient and the cockatoo had identical biochemical profiles, the same monoclonal antibody immunofluorescence patterns, and indistinguishable patterns on restriction fragment length polymorphism analysis and karyotyping. CONCLUSIONS: The indistinguishable patient and cockatoo isolates strongly suggest that the patient's infection resulted from exposure to aerosolized cockatoo excreta. Although the incidence of cryptococcal infection due to such exposure is unknown, it may be prudent to advise immunocompromised patients to avoid pet birds and avian excreta.

Aged↗
PubMed2000

Melanin and virulence in Cryptococcus neoformans.

Melanin synthesis has been associated with virulence for the human pathogenic fungus Cryptococcus neoformans. Recent evidence indicates that C. neoformans cells synthesize melanin during infection and that this pigment protects the fungus against immune defense mechanisms.

Animals↗
PubMed2000

Synthesis of polymerized melanin by Cryptococcus neoformans in infected rodents.

The ability of Cryptococcus neoformans to synthesize polymerized melanin in vitro has been associated with virulence, but it is unclear whether this fungus synthesizes polymerized melanin during infection. To study this question, we used two approaches: one involved the generation of monoclonal antibodies (MAbs) to melanin for use in immunohistochemical studies of C. neoformans-infected rodents, and the other sought to isolate fungal melanin from infected tissues. Digestion of in vitro-melanized C. neoformans cells with proteases, denaturant, and hot concentrated acid yields melanin particles that retain the shape of fungal cells and are therefore called melanin ghosts. BALB/c mice were immunized with melanin ghosts, and two immunoglobulin M MAbs to melanin were generated from the spleen of one mouse. Immunofluorescence analyses of lung and brain tissues of rodents infected with wild-type melanin-producing (Mel(+)) C. neoformans strains demonstrated binding of the MAbs to the fungal cell wall. No binding was observed when infections were performed with mutant albino (Mel(-)) C. neoformans strains. Particles with striking similarity to melanin ghosts were recovered after digestion of lung and brain tissues from Mel(+) C. neoformans-infected rodents and were reactive with the MAbs to melanin. No particles were recovered from tissues infected with Mel(-) C. neoformans. A Mel(+) C. neoformans strain grown on lung or brain homogenate agar became lightly pigmented and also yielded particles similar to melanin ghosts upon digestion, providing additional evidence that lung and brain tissues contain substrate for C. neoformans melanization. These results demonstrate that C. neoformans synthesizes polymerized melanin during infection, which has important implications for pathogenesis and antifungal drug development.

Animals↗
PubMed1999

Amphotericin B and fluconazole affect cellular charge, macrophage phagocytosis, and cellular morphology of Cryptococcus neoformans at subinhibitory concentrations.

Amphotericin B (AmB) and fluconazole (FLU) are the major antifungal drugs used in the treatment of cryptococcosis. Both drugs are believed to exert their antifungal effects through actions on cell membrane sterols. In this study we investigated whether AmB and FLU had other, more subtle effects on C. neoformans that could contribute to their therapeutic efficacy. C. neoformans cells were grown in media with subinhibitory concentrations of either AmB or FLU and analyzed for cellular charge, phagocytosis by macrophages with antibody and complement opsonins, appearance by scanning electron and light microscopies, and release of the capsular polysaccharide glucuronoxylomannan into the culture medium. Growth in the presence of either AmB or FLU resulted in major reductions in cellular charge, as measured by determination of the zeta potential. Phagocytosis studies demonstrated that exposure of C. neoformans to subinhibitory concentrations of AmB or FLU enhanced phagocytosis by macrophages. Scanning electron microscopy revealed that a large proportion of cells had an altered capsular appearance. Cells grown in medium with either AmB or FLU were smaller and released more glucuronoxylomannan into the culture medium than cells grown without antibiotics. The results suggest additional mechanisms of action for AmB and FLU that may be operative in body compartments where drug levels do not achieve the MICs. Furthermore, the results suggest mechanisms by which AmB and FLU can cooperate with humoral and cellular immune defense systems in controlling C. neoformans infections.

Amphotericin B↗
PubMed1999

Melanization of Cryptococcus neoformans in murine infection.

Cryptococcus neoformans is a fungus that is pathogenic in humans and that can produce melanin in vitro. Melanization is associated with virulence, but there is no evidence that melanin is made during infection. Melanins are difficult to study because they are amorphous and insoluble. Melanin-binding peptides from a phage display library were used to demonstrate that C. neoformans makes melanin-like compounds in tissue. Melanin-binding peptides were characterized by a high proportion of positively charged and aromatic residues. Two other methods, demonstration of an antibody response to melanin in mice infected with C. neoformans and analysis of yeast cell walls in infected tissue by light microscopy, were used to support these findings. The demonstration that C. neoformans melanizes in tissue has important implications for pathogenesis and drug discovery.

Animals↗
PubMed1999

Melanin as a potential cryptococcal defence against microbicidal proteins.

Cryptococcus neoformans is an important fungal pathogen that synthesizes melanin when grown in the presence of phenolic substrates. The ability of C. neoformans to produce melanin is associated with virulence, but the specific role of melanin in the pathogenesis of infection is not clear. In this study the ability of C. neoformans melanin to bind proteins and protect against microbicidal peptides was investigated. Melanin was shown to bind a variety of proteins of fungal and mammalian origin. Melanin-protein interactions were dependent on the pH of the solution and on the amount of protein and melanin present. Melanized cells were less susceptible to killing by three microbicidal peptides: a defensin, a protegrin, and a magainin. Incubation of the microbicidal peptides with melanin particles, followed by removal of the melanin, reduced or abolished fungicidal activity, demonstrating interactions between peptides and melanin. The ability of melanin to bind proteins and to protect against microbicidal peptides suggests a protective function for melanin, whereby it sequesters microbicidal peptides and abrogates their activity.

Amino Acid Sequence↗