Human phagocytic cell responses to Scedosporium apiospermum (Pseudallescheria boydii): variable susceptibility to oxidative injury

Scedosporium apiospermum (Pseudallescheria boydii) is an emerging opportunistic filamentous fungus that causes serious infections in both immunocompetent and immunocompromised patients. To gain insight into the immunopathogenesis of infections due to S. apiospermum, the antifungal activities of human polymorphonuclear leukocytes (PMNs), mononuclear leukocytes (MNCs), and monocyte-derived macrophages (MDMs) against two clinical isolates of S. apiospermum were evaluated. Isolate SA54A was amphotericin B resistant and was the cause of a fatal disseminated infection. Isolate SA1216 (cultured from a successfully treated localized subcutaneous infection) was susceptible to amphotericin B. MDMs exhibited similar phagocytic activities against conidia of both isolates. However, PMNs and MNCs responded differently to the hyphae of these two isolates. Serum opsonization of hyphae resulted in a higher level of superoxide anion (O(2)(-)) release by PMNs in response to SA54A (amphotericin B resistant) than that seen in response to SA1216 (amphotericin B susceptible; P < 0.001). Despite this increased O(2)(-) production, PMNs and MNCs induced less hyphal damage to SA54A than to SA1216 (P < 0.001). To investigate the potential mechanisms responsible for these differences, hyphal damage was evaluated in the presence of antifungal oxidative metabolites as well as in the presence of a series of inhibitors and scavengers of antifungal PMN function. Mannose, catalase, superoxide dismutase, dimethyl sulfoxide, and heparin had no effect on PMN-induced hyphal damage to either of the two isolates. However, azide, which inhibits PMN myeloperoxidase activity, significantly reduced hyphal damage to SA1216 (P < 0.01) but not to SA54A. Hyphae of SA1216 were slightly more susceptible to oxidative pathway products, particularly HOCl, than those of SA54A. Thus, S. apiospermum is susceptible to antifungal phagocytic function to various degrees. The selective inhibitory pattern of azide with respect to hyphal damage and the parallel susceptibility to HOCl suggests an important difference in susceptibilities to myeloperoxidase products that may be related to the various levels of pathogenicity and amphotericin B resistance of S. apiospermum.     

Characterization of Scedosporium prolificans clinical isolates by randomly amplified polymorphic DNA analysis.

Fingerprinting by randomly amplified polymorphic DNA (RAPD) analysis was used to differentiate Scedosporium prolificans isolates. A total of 59 arbitrary primers were screened with six unrelated S. prolificans isolates, and a panel of 12 primers was selected. The 12 primers were then used to detect DNA polymorphisms among 17 S. prolificans isolates from 11 patients with systemic S. prolificans infections diagnosed in three hospitals located in geographically different areas of Spain. Eight patients were diagnosed with S. prolificans infection in a single institution over a 6-year period, and two other patients were diagnosed with S. prolificans infection in a different hospital over a 1-year period. No single primer allowed for the discrimination of all the isolates from different patients, but this was possible by combining the RAPD patterns from three primers (UBC 701, AB1.08, and AB1.11 or UBC 701, AB1.08, and UBC 707). However, multiple isolates from the same patient were identical. In this study, we also compared a visual method and a computerized method for the analysis of the RAPD patterns. Both methods were satisfactory and gave few discordances, but given the advantages and disadvantages of each method, both systems should be used together. RAPD analysis provided a fast and economical means of typing S. prolificans isolates, with a high level of discrimination among unrelated isolates. Typing by RAPD analysis confirmed that the S. prolificans infections were epidemiologically unrelated.     

Surviving a recurrent Scedosporium prolificans endocarditis: A case report

Scedosporium prolificans have been reported to be resistant to all antifungals including the newer azoles and echinocandins. We report an unusual case of repeated S. prolificans infection of the heart valves in an immunocompetent patient.     

Fungemia due to Scedosporium prolificans: a description of two cases with fatal outcome

Two cases, probably related, of fungemia due to Scedosporium prolificans are described in two patients with acute leukemia. Both were admitted to the hematological ward in nearby rooms, during building work in the hospital. After a previous bacterial sepsis in the neutropenic phase, which improved with antibiotic treatment, the respiratory status in both patients deteriorated presenting acute dypsnea, with a lung infiltrate in one of them. A few hours later both patients died. Blood cultures were positive for S. prolificans . These two new cases of S. prolificans infection stress the importance of awareness of this emerging pathogen in patients who suffer a hematologic malignancy during the neutropenic phase, especially if building work is taking place in the hospital.     

Human T cell responses to beta-galactosidase.

The peripheral blood of most normal individuals has been shown to contain T cells that respond to beta-galactosidase (beta-Gal), presumably as a result of natural priming. Three T cell clones (clones 1,2,4) specific for beta-Gal were isolated from peripheral blood mononuclear cells (PBMC) after pretreatment with leucine methyl ester (LeuOMe); a fourth clone from the same individual was isolated from untreated cells. All four clones were CD4+ CD8- alpha beta TcR+ and clone 1 was additionally shown to be cytotoxic. Epstein-Barr virus (EBV) transformed B cell lines were derived from LeuOMe-treated or untreated PBMC and used to study the efficiency of presentation of beta-Gal to one of the clones. The results indicated that B cells transformed after LeuOMe treatment presented beta-Gal at lower concentrations than untreated controls. beta-Gal would therefore appear to be a highly suitable model antigen for studies of immunoregulation in humans.     

Ribosomal DNA internal transcribed spacer analysis supports synonomy of Scedosporium inflatum and Lomentospora prolificans.

Scedosporium inflatum is a dematiaceous opportunistic pathogen originally described by D. Malloch and I.F. Salkin (Mycotaxon 21:247-255, 1984). However, E. Gueho and G. S. De Hoog (J. Mycol. Med. 118:3-9, 1991) recently suggested reducing this mold to synonomy with Lomentospora prolificans on the basis of their similar morphological and molecular characteristics. We have investigated the ribosomal DNA internal transcribed spacers (ITS), i.e., ITS I and ITS II, of 18 isolates, including these two fungi and a closely related pathogen, Scedosporium apiospermum, and its telemorph, Pseudallescheria boydii. Identical ITS restriction fragment length polymorphisms were found in eight isolates of S. inflatum and L. prolificans. These results support Gueho and De Hoog's proposal to combine S. inflatum and L. prolificans into the binomial Scedosporium prolificans. However, the ITS I sequence of S. apiospermum and the ITS restriction fragment length polymorphisms of S. apiospermum and P. boydii were found to be significantly different from those of S. inflatum and L. prolificans. The ITS restriction pattern differences may be valuable in clinical settings for distinguishing these fungi.     

Molecular typing of clinical and environmental isolates of Scedosporium prolificans by inter-simple-sequence-repeat polymerase chain reaction.

Invasive infections by Scedosporium prolificans have increased alarmingly in recent years, mainly in immunosuppressed patients. The epidemiology, pathogenesis and the natural habitat of this pathogen are practically unknown. Isolates of S. prolificans were distinguished from one another by inter-simple-sequence-repeat (ISSR) fingerprinting, a technique based on the high degree of polymorphism of the multisatellite genetic markers used. This technique was found useful for typing 84 isolates of S. prolificans from different countries and sources. The assemblage of S. prolificans isolates tested was extremely diverse, with 35 genotypes present. Several patients were found to have been infected or colonized by more than one strain. Overall, this technique facilitates the epidemiological study of S. prolificans infection.     

Human peritoneal macrophage phagocytic, killing, and chemiluminescent responses to opsonized Listeria monocytogenes.

Opsonization with normal human serum, purified immunoglobulin G, or immunoglobulin G-deficient serum promoted phagocytosis of Listeria monocytogenes by human peritoneal macrophages. However, normal human serum was the most effective opsonin in elicting killing and chemiluminescent responses. Macrophages phagocytized and killed almost as much as polymorphonuclear leukocytes but produced considerably less chemiluminescence.     

Disseminated infection caused by Scedosporium prolificans in a patient with acute multilineal leukemia

In this report, we describe a case of disseminated infection caused by Scedosporium prolificans (S. inflatum) in a patient affected by chemotherapy-induced acute multilineal leukemia and neutropenia. For the fungus isolated in four blood cultures, high MICs of currently available antifungal agents were found. Postmortem examination revealed multiorgan involvement.     

Nosocomial outbreak caused by Scedosporium prolificans (inflatum): four fatal cases in leukemic patients.

Four cases of fatal disseminated Scedosporium prolificans (inflatum) infection occurring in neutropenic patients are reported. Because of hospital renovation, the patients were cared for in a temporary hematologic facility. S. prolificans (inflatum) was isolated from blood cultures of these four patients, two of whom underwent full necropsy, and revealed abundant vegetative hyphae and ovoid conida with truncate bases in many organs. In vitro susceptibility testing of fungal strains showed all isolates to be resistant to amphotericin B, flucytosine, miconazole, ketoconazole, fluconazole, and itraconazole, with MICs greater than 16 micrograms/ml. The reported infections, two in each of two rooms, occurred over a period of 1 month, with very similar clinical outcomes. Circumstancial evidence suggested a nosocomial outbreak, but the environmental samples collected from the rooms, corridors, and adjacent areas did not yield S. prolificans (inflatum). Nevertheless, circumstantial evidence suggested a nosocomial outbreak of S. prolificans (inflatum) infection.     

Comparison of the virulence of Scedosporium prolificans strains from different origins in a murine model

Scedosporium prolificans is an emerging opportunist fungus that causes different types of infections in immunocompetent and immunosuppressed people. These infections show an irregular geographical distribution and, generally, disseminated systemic infections are noticed only in specific countries. This study used a murine model of disseminated infection by this fungus to assess if strains from different origins have different virulence. Two strains from each of four different sources (disseminated infection, localised infection, asymptomatic cystic fibrosis patients and the environment) were tested. Two strains of S. apiospermum of clinical origin were also included in the study; these were clearly less virulent than those of S. prolificans. The S. prolificans strains tested were classified in three groups according to their virulence. The groups with higher and lower virulence were represented by only one strain each, and the intermediate group contained six strains. No significant differences were found between the strains from different geographic areas or different forms of disease.     

Fatal meningoencephalitis caused byScedosporium inflatum (Scedosporium prolificans) in a child with lymphoblastic leukemia

A fatal case of meningoencephalitis caused byScedosporium inflatum (Scedosporium prolificans) in a 5-year-old boy with acute myeloblastic leukemia who was given intrathecal treatment is reported. Itraconazole treatment was ineffective. The fungus was identified on brain sections at autopsy and was not observed in any other organ. As no other portal of entry was detected, meningoencephalitis may have originated via direct introduction of the fungus at therapeutic lumbar puncture.     

Scedosporium prolificans Keratouveitis in Association with a Contact Lens Retained Intraocularly over a Long Term

Scedosporium prolificans is a soil saprophyte that is associated with a large variety of infectious processes and with respiratory colonization in immunocompetent and immunocompromised patients. We report the first described case of S. prolificans keratouveitis associated with the intraocular long-term retention of a contact lens in a 76-year-old female patient.     

Clinical, pathologic and epidemiologic features of infection with Scedosporium prolificans: four cases and review

Objective: To describe the features of infection with Scedosporium prolificans and to investigate the possibility of a common source of infection.
Methods: S.prolificans caused invasive infections in four patients at Liverpool Hospital from 1994 to 1997, prompting a pathologic and epidemiologic investigation. Blood cultures were processed by either the BACTEC NR660 or VITAL systems. MIC testing was performed with Etest strips. Strain differentiation of isolates from three of the patients and two soil samples from the home of one of the patients was performed by allozyme electrophoresis.
Results: The four cases represented disseminated infections that arose during prolonged neutropenia, and progressed relentlessly despite treatment, including in one case high-dose liposomal amphotericin B. In two cases, VITAL blood culture bottles contained mycelia of S. prolificans without the reader having signaled. An autopsy was performed in three of the cases. Angio-invasion, tissue necrosis and multiple abscesses were found in each patient. Multiple air samples from the ward were negative for S. prolificans . The organism was grown from two samples of pot plant soil from the home of one patient. Allozyme electrophoresis performed on isolates from three cases and pot plant soil indicated that all strains were unrelated. Furthermore, two patients appeared to harbor at least two different strains of S. prolificans .
Conclusions: S. prolificans causes disseminated infection in neutropenic patients. Antifungal treatment, including high-dose liposomal amphotericin B, is ineffective in overcoming these infections. Genetic techniques discriminated all strains and suggested that an outbreak had not occurred. Multiple strains of S. prolificans were isolated from two patients, indicating the possibility of mixed infections occurring.     

Resistance to the Sterne strain of B. anthracis: phagocytic cell responses of resistant and susceptible mice

Inflammatory responses were compared in vivo, and host phagocytic cell functions compared in vitro, of mice resistant (CBA/J) and susceptible (A/J) to lethal infection with the Sterne strain of Bacillus anthracis. Polymorphonuclear leukocyte (PMN) and macrophage responses at the initial site of infection were slower in A/J mice than in CBA/J mice. Whereas in A/J mice, the number of PMN ultimately responding to infection was equal to, or greater than, that in CBA/J mice, fewer macrophages accumulated. A/J mice failed to clear relatively low doses of the organisms and died. In vitro, chemotactic responses to both serum- and bacteria-derived attractants were similar for macrophages from A/J and CBA/J mice but were reduced for PMN from A/J mice. PMN and macrophages from the two mouse strains phagocytosed and killed spores in vitro to a similar extent, although killing by A/J PMN could be blocked by prior uptake of large numbers of killed spores. Thus susceptibility to lethal infection with Sterne strain correlated with the delayed influx (PMN) and reduced accumulation (macrophages) of phagocytes at the initial site of infection, but not with defective in vitro uptake or killing of spores.     

Chemotactic and phagocytic responses of human alveolar macrophages to activated complement components.

Human alveolar macrophages (AMs) migrated toward and aggregated to C5a but not toward C5a-deficient serum. Human AMs from cigarette smokers migrated significantly farther than did AMs from nonsmokers. Human AMs phagocytized Escherichia coli opsonized with activated C3. Immunoglobulin G was not required for phagocytosis. Human AMs demonstrate chemotaxis and aggregation to C5a. Further, human AMs can phagocytize bacteria coated with activated C3.