Julie A. Ribes

Zygomycetes in Human Disease

The Zygomycetes represent relatively uncommon isolates in the clinical laboratory, reflecting either environmental contaminants or, less commonly, a clinical disease called zygomycosis. There are two orders of Zygomycetes containing organisms that cause human disease, the Mucorales and the Entomophthorales. The majority of human illness is caused by the Mucorales. While disease is most commonly linked to Rhizopus spp., other organisms are also associated with human infection, including Mucor, Rhizomucor, Absidia, Apophysomyces, Saksenaea, Cunninghamella, Cokeromyces, and Syncephalastrum spp. Although Mortierella spp. do cause disease in animals, there is no longer sufficient evidence to suggest that they are true human pathogens. The spores from these molds are transmitted by inhalation, via a variety of percutaneous routes, or by ingestion of spores. Human zygomycosis caused by the Mucorales generally occurs in immunocompromised hosts as opportunistic infections. Host risk factors include diabetes mellitus, neutropenia, sustained immunosuppressive therapy, chronic prednisone use, iron chelation therapy, broad-spectrum antibiotic use, severe malnutrition, and primary breakdown in the integrity of the cutaneous barrier such as trauma, surgical wounds, needle sticks, or burns. Zygomycosis occurs only rarely in immunocompetent hosts. The disease manifestations reflect the mode of transmission, with rhinocerebral and pulmonary diseases being the most common manifestations. Cutaneous, gastrointestinal, and allergic diseases are also seen. The Mucorales are associated with angioinvasive disease, often leading to thrombosis, infarction of involved tissues, and tissue destruction mediated by a number of fungal proteases, lipases, and mycotoxins. If the diagnosis is not made early, dissemination often occurs. Therapy, if it is to be effective, must be started early and requires combinations of antifungal drugs, surgical intervention, and reversal of the underlying risk factors. The Entomophthorales are closely related to the Mucorales on the basis of sexual growth by production of zygospores and by the production of coenocytic hyphae. Despite these similarities, the Entomophthorales and Mucorales have dramatically different gross morphologies, asexual reproductive characteristics, and disease manifestations. In comparison to the floccose aerial mycelium of the Mucorales, the Entomophthorales produce a compact, glabrous mycelium. The asexually produced spores of the Entomophthorales may be passively released or actively expelled into the environment. Human disease with these organisms occurs predominantly in tropical regions, with transmission occurring by implantation of spores via minor trauma such as insect bites or by inhalation of spores into the sinuses. Conidiobolus typically infects mucocutaneous sites to produce sinusitis disease, while Basidiobolus infections occur as subcutaneous mycosis of the trunk and extremities. The Entomophthorales are true pathogens, infecting primarily immunocompetent hosts. They generally do not invade blood vessels and rarely disseminate. Occasional cases of disseminated and angioinvasive disease have recently been described, primarily in immunocompromised patients, suggesting a possible emerging role for this organism as an opportunist.

Zygomycosis (mucormycosis): emerging clinical importance and new treatments.

Purpose of review New importance has been given to zygomycosis, as what was uncommon is no longer. Zygomycosis (mucormycosis) typically occurs in patients with leukemia, with solid-organ transplants or bone marrow transplants, with diabetic ketoacidosis, in those who have received steroids or are neutropenic, and after desferioxamine therapy. Often, both diagnostic and therapeutic measures are performed too late and are inadequate. Mortality rates may be as high as 80% in infected transplant recipients. Zygomycosis also appears to have made a subtle increase in incidence: up to 8% in autopsied patients with leukemia, and 2% in allogenic bone marrow transplant patients. Most infections are acquired by inhalation, ingestion, or trauma. They rapidly infarct blood vessels, resulting in necrosis of surrounding tissue. Over the past few years, new diagnostic procedures, susceptibility tests, and drugs have entered the clinic, and these advances are discussed in the review. Recent findings With the rise in number of cases of ‘zygomycosis’, new scrutiny has been directed at the terms ‘zygomycosis’ and ‘mucormycosis’. This review explains their differences and the attending relevance for the clinician. Diagnostic methods include new molecular detection assays and new susceptibility testing options. New treatment options will soon exist with triazole antifungal agents. The first one expected to enter clinical practice is posaconazole in 2005. Its metabolism, pharmacokinetics, in-vitro and in-vivo activity, and clinical study results are described. Finally, we present our approach to zygomycosis. Summary This review discusses key elements to laboratory diagnostic and susceptibility procedures and new treatment options.

Fibrin induces release of von Willebrand factor from endothelial cells.

Addition of fibrinogen to human umbilical vein endothelial cells in culture resulted in release of von Willebrand factor (vWf) from Weibel-Palade bodies that was temporally related to formation of fibrin in the medium. Whereas no release occurred before gelation, the formation of fibrin was associated with disappearance of Weibel-Palade bodies and development of extracellular patches of immunofluorescence typical of vWf release. Release also occurred within 10 min of exposure to preformed fibrin but did not occur after exposure to washed red cells, clot liquor, or structurally different fibrin prepared with reptilase. Metabolically labeled vWf was immunopurified from the medium after release by fibrin and shown to consist of highly processed protein lacking pro-vWf subunits. The contribution of residual thrombin to release stimulated by fibrin was minimized by preparing fibrin clots with nonstimulatory concentrations of thrombin and by inhibiting residual thrombin with hirudin or heating. We conclude that fibrin formed at sites of vessel injury may function as a physiologic secretagogue for endothelial cells causing rapid release of stored vWf.

Six-Year Study of the Incidence of Herpes in Genital and Nongenital Cultures in a Central Kentucky Medical Center Patient Population

ABSTRACT Herpes infections are among the most common sexually transmitted diseases and are the most common cause of genital ulcer disease in the United States. This study addresses the changing distribution of herpes simplex virus type 1 (HSV-1) and HSV-2 in patients presenting for evaluation of herpetic infections. Viral culture results from the University of Kentucky Clinical Microbiology Laboratory were reviewed for a 6-year period (1994 through 1999). Data were collected on patient sex, site of culture, and culture result. These data were analyzed statistically to identify yearly trends. Of the 4,498 cultures analyzed, nearly equal proportions of HSV-1 (13.3%) and HSV-2 (12.0%) were detected for an overall culture positivity rate of 25.3%. Approximately two-thirds of all positive cultures were from women. Although HSV-2 remained the predominant type of genital herpes, over the 6-year span of this study, there was a trend toward increasing proportions of HSV-1 genitalis, with 31.8% of male patients and 44.8% of female patients demonstrating HSV-1 genitalis by 1999. The majority of patients with HSV in nongenital sites grew HSV-1. Although there was significant yearly variation, HSV-2 was isolated from only 9.4% of patients with nongenital HSV for the entire 6-year period. This study therefore concludes that HSV-2 remains primarily a genital pathogen, while HSV-1 is taking on an increasingly important role in causing genital ulcer disease in addition to being the primary nongenital HSV.

Evaluation of Methods To Increase the Sensitivity and Timeliness of Detection of Streptococcus agalactiae in Pregnant Women

ABSTRACT Direct culture of rectovaginal specimens on Granada agar was compared to culture on sheep blood agar plate (SBAP) and AccuProbe detection of group B streptococcus from overnight LIM broth enhancement cultures (LIM-SBAP). Both broth-enhanced methods demonstrated excellent sensitivity (97.5% for LIM-SBAP and 93.5% for AccuProbe), while Granada agar demonstrated a sensitivity of only 40.3%.

Mediation of fibrin-induced release of von Willebrand factor from cultured endothelial cells by the fibrin beta chain.

The exposure of endothelial cells (EC) to fibrin has been shown to stimulate the rapid release of von Willebrand factor (vWf) from storage sites in Weibel-Palade bodies. We have now investigated the fibrin structural features required for stimulation of release. The role of fibrinopeptide cleavage was examined by preparing fibrin with thrombin to remove both fibrinopeptide A (FPA) and fibrinopeptide B (FPB) and with reptilase or Agkistrodon contortrix procoagulant to selectively remove FPA or FPB, respectively. vWf release was found to require FPB cleavage, whereas removal of FPA and Factor XIIIa cross-linking of fibrin were without effect. The dependence of release on FPB cleavage suggested that a site involving the NH2 terminus of the beta chain could mediate vWf secretion. To test this hypothesis, B beta chain derivatives were prepared and examined for their capacity to induce release. Purified B beta chain had no effect on release at a concentration of 20 nM but stimulated release from 26 +/- 6% of cells at 200 nM, the maximum solubility. However, after thrombin cleavage of FPB, release occurred from 36 +/- 9% of cells at 20 nM and from 60 +/- 7% at 200 nM, both significantly greater than before cleavage. FPB and B beta 1-42 showed no activity, whereas beta 15-42, representing the NH2 terminus of the thrombin cleaved beta chain, stimulated significant release at concentrations of 0.1 and 1 mM. We conclude that FPB cleavage from fibrin is required for stimulation of vWf release from EC and that this is mediated by a site that includes the NH2 terminus of the beta chain.

Fatal Rhizopus Pneumonia in Allogeneic Stem Cell Transplant Patients Despite Posaconazole Prophylaxis: Two Cases and Review of the Literature

Posaconazole is a triazole with broad spectrum of activity against multiple fungi including members of the fungal order Mucorales. This activity has been shown both in clinical and in vitro studies, which are critically reviewed here. It has become very popular in prophylaxis in acute myelogenous leukemia (AML) induction and in the graft-versus-host disease (GVHD) settings after 2 recent prospective trials that showed advantage of posaconazole prophylaxis compared to fluconazole or itraconazole. In this report, 2 patients are presented, in whom, despite posaconazole prophylaxis, invasive and ultimately fatal Rhizopus pulmonary infections developed. These cases are similar to a previously reported case of Rhizopus infection in a stem cell transplant recipient who also received posaconazole, indicating a potential newly recognized pattern of breakthrough infections in patients receiving posaconazole prophylaxis.

Invasive Enteric Infections in Hospitalized Patients With Underlying Strongyloidiasis

Disseminated strongyloidiasis is often associated with enteric bacterial infections. This study was undertaken to determine if enteric organisms caused extraintestinal infections in patients infected with Strongyloides stercoralis but without apparent dissemination. The medical records of hospitalized patients from central Kentucky with strongyloidiasis (1993-2003) were examined to determine the occurrence of extraintestinal infections with enteric organisms. Of 30 patients with S stercoralis, 16 had invasive infections, including sepsis, meningitis, pneumonia, peritonitis, and endocarditis caused by enteric bacteria and Candida organisms. Infections were seen in 8 (62%) of 13 patients with disseminated strongyloidiasis and 8 (47%) of 17 with disease apparently limited to the gastrointestinal tract. Fifteen patients were receiving corticosteroids or other immunosuppressive therapy. Peripheral eosinophilia was seen in only 23% (7/30). Infection with S stercoralis, even without obvious dissemination, may predispose to invasive infections caused by enteric organisms. In Strongyloides-endemic areas, patients with invasive infections caused by enteric organisms should be examined for coinfection with S stercoralis.

Comparative Performance of Herpes Simplex Virus Type 2-Specific Serologic Assays from Meridian Diagnostics and MRL Diagnostics

ABSTRACT MRL Diagnostics and Meridian Diagnostics have recently designed herpes simplex virus type 2 (HSV-2)-specific enzyme immunoassays for HSV-2 antibody detection. Blood donor sera were assayed for HSV-2 antibodies by both methods. The sensitivity, specificity, and efficiency were 97.9, 95.4, and 95.9% for the MRL assay and 83.2, 98.2, and 95.5% for the Meridian assay, respectively.

Performance characteristics of VIDAS and directigen respiratory syncytial virus (RSV) antigen detection assays and culture for the identification of RSV in respiratory specimens.

ABSTRACT In a comparison of the Directigen and VIDAS respiratory syncytial virus antigen detection assays with viral culture, the sensitivity, specificity, positive and negative predictive values, and testing efficiency were 86, 93.1, 82.7, 94.6, and 91.2% for Directigen; 96.1, 90.8, 80.3, 98.3, and 92.3% for VIDAS; and 88.2, 100, 100, 95.7, and 96.8% for viral culture, respectively.