Infections during mobilizing chemotherapy and following autologous stem cell transplantation.

Autologous peripheral blood stem cells (PBSC), for transplantation following high-dose chemotherapy, are collected using regimens containing cytokines with or without chemotherapy. The added period of neutropenia prior to stem cell transplantation (SCT) in patients receiving chemotherapy mobilization may increase the risk of infections following transplantation. We studied the incidence of culture-positive infections in 107 consecutive patients who were divided into three groups, according to whether they experienced extended neutropenia during chemotherapy for stem cell mobilization as well as post autotransplant. All the patients received antibiotic prophylaxis and hematopoietic growth factors during neutropenia. The total duration of pre-transplant neutropenia differed among the three mobilization schemes (growth factors alone; one cycle; or two cycles of chemotherapy plus growth factor for mobilization) at 0, 6 and 18 days, respectively (median). However the post-autograft time to myeloid engraftment was similar at 10 days (median). The incidence of culture-proven infections in all three groups was similar. Using fluconazole for yeast prophylaxis, 40% patients developed gastrointestinal colonization with yeast, and the majority of speciated isolates were Candida glabrata. Bacteremia developed in 22% and 9% of patients with S. epidermidis and Gram-negative organisms, respectively, while 11% developed C. difficile-associated diarrhea. In conclusion, treatment using none, one or two cycles of mobilizing chemotherapy pre-transplant does not influence the overall incidence of infections among autologous SCT recipients. However, although post-transplant neutropenia is brief, infections remain a significant cause of morbidity.

Randomized clinical trial of ganciclovir vs acyclovir for prevention of cytomegalovirus antigenemia after allogeneic transplantation.

Cytomegalovirus (CMV) disease remains a major cause of morbidity following allogeneic stem cell transplantation (SCT). In a prospective randomized trial, we tested prophylactic therapy with ganciclovir or acyclovir for patients at high risk of disease. Ninety-one CMV seropositive recipients of related (n = 53) and unrelated (n = 38) donor transplants were enrolled. All patients received intravenous (i.v.) ganciclovir 5 mg/kg every 12 h days -7 to -2, followed by acyclovir 10 mg/kg i.v. every 8 h from day -1 until neutrophil engraftment. Patients were then randomly assigned to either ganciclovir (n = 45) or acyclovir (n = 46) until day 100 post transplant. Any degree of antigenemia was treated with ganciclovir 5 mg/kg i.v. twice a day for 2 weeks, followed by 5 mg/kg i.v. each weekday for 6 weeks. At day 100, the cumulative incidence of antigenemia was 31% (95% CI 17-45%) for ganciclovir and 41% (95% CI 26-56%) (P = 0.22) for acyclovir prophylaxis, respectively. The assigned prophylaxis cohort did not predict for CMV antigenemia. The cumulative incidence of CMV disease at 12 months was 13% (95% CI 3-23%) and 17% (95% CI 6-28%) (P = 0.59) for the ganciclovir- and acyclovir-treated groups, respectively. An absolute neutrophil count (ANC)

Infections in recipients of blood and marrow transplantation.

The approach to infections in blood and marrow transplant (BMT) recipients involves an understanding of clinical infection syndromes and the natural history of individual infections, taken in the context of patterns of immunosuppression after transplantation and mechanisms underlying immune system reconstitution over time. The conditioning regimen used to prepare the host is a major determinant of host tissue injury and may lead to mucositis or diarrhea, facilitating transmucosal origin of bloodstream infections. Infectious risk also differs between autologous and allogeneic grafts as a consequence of ongoing immunosuppression from graft-versus-host disease and its therapy. Post-transplant complications may mimic infectious processes, and multiple infections may occur in one patient at the same time. Thus, the BMT patient with suspected infection should be evaluated in the context of pretransplant exposure history (infectious disease serologies), conditioning regimen, available culture data from nonsterile mucosal surfaces, previous and recent infections, contemporary transplant complications, and the current degree and duration of neutropenia, cellular immunodeficiency, and hypogammaglobulinemia.

Infectious complications following unrelated cord blood transplantation.

INTRODUCTION: Infections following cord blood transplantation are just beginning to be defined in the literature. This review will outline infections at death, the epidemiology of individual infections, and the impact of stem cell source. METHODS: A review of studies published since 2000. RESULTS: Based on registry data, most studies demonstrate an approximate rate of infection at death of 30-40% among cord blood recipients. Bacterial infections often occur prior to engraftment and increase among patients with graft failure. In addition, there is delayed recovery of the immune response among patients with graft-versus-host disease that leads to viral infections at later time points. The risk of serious infection among children receiving umbilical cord blood (UCB) grafts is comparable to that of children receiving unmanipulated marrow and is lower than that of recipients of a T-cell-depleted stem cell source. Among adult patients, despite an overall higher incidence of serious infections after UCB transplantation as compared with unrelated donor grafts, non-relapse mortality and overall survival were not significantly different between haematopoietic stem cell sources. CONCLUSIONS: Further studies are needed to confirm these observations and determine whether the risk of infection for cord blood recipients is comparable to that of recipients of unmanipulated marrow.

Posaconazole as salvage therapy for zygomycosis.

Zygomycosis, an infection that is associated with significant morbidity and mortality, is becoming common in immunocompromised patients. Posaconazole is a new extended-spectrum azole antifungal that has demonstrated in vitro and in vivo activity against zygomycetes. This report provides the results from the first 24 patients with active zygomycosis who were enrolled in two open-label, nonrandomized, multicentered compassionate trials that evaluated oral posaconazole as salvage therapy for invasive fungal infections. Posaconazole was usually given as an oral suspension of 200 mg four times a day or 400 mg twice a day. Eleven (46%) of the infections were rhinocerebral. Duration of posaconazole therapy ranged from 8 to 1,004 days (mean, 292 days; median, 182 days). Rates of successful treatment (complete cure and partial response) were 79% in 19 subjects with zygomycosis refractory to standard therapy and 80% in 5 subjects with intolerance to standard therapy. Overall, 19 of 24 subjects (79%) survived infection. Survival was also associated with surgical resection of affected tissue and stabilization or improvement of the subjects' underlying illnesses. Failures either had worsening of underlying illnesses or requested all therapy withdrawn; none of the failures received more than 31 days of posaconazole. Posaconazole oral solution was well tolerated and was discontinued in only one subject due to a drug rash. Posaconazole appears promising as an oral therapy for zygomycosis in patients who receive required surgery and control their underlying illness.

Aspects of fungal pathogenesis in humans.

Fungal diseases have become increasingly important in the past few years. Because few fungi are professional pathogens, fungal pathogenic mechanisms tend to be highly complex, arising in large part from adaptations of preexisting characteristics of the organisms' nonparasitic lifestyles. In the past few years, genetic approaches have elucidated many fungal virulence factors, and increasing knowledge of host reactions has also clarified much about fungal diseases. The literature on fungal pathogenesis has grown correspondingly; this review, therefore, will not attempt to provide comprehensive coverage of fungal disease but focuses on properties of the infecting fungus and interactions with the host. These topics have been chosen to make the review most useful to two kinds of readers: fungal geneticists and molecular biologists who are interested in learning about the biological problems posed by infectious diseases, and physicians who want to know the kinds of basic approaches available to study fungal virulence.

Cytomegalovirus enteritis among hematopoietic stem cell transplant recipients.

Cytomegalovirus (CMV) is a cause of enteritis associated with hematopoietic stem cell transplantation (HSCT), but the natural history is rarely studied and hence poorly understood. CMV infection at this end-organ site is notably less frequent than is pneumonitis. To evaluate the spectrum of CMV enteritis after HSCT, we reviewed the database spanning 11.5 years of 2240 University of Minnesota transplantation patients for cases of gastrointestinal CMV. We identified 46 case-patients. The incidence of CMV enteritis at 2 years following HSCT averaged 2% over the 11.5-year study interval. The median time to diagnosis of CMV enteritis after HSCT was 91 days (range, 17-527 days). The methods used in diagnosis included histopathology (58%) and virology (61%). Viremia was detected in two thirds of patients with CMV prior to the diagnosis of enteritis. Most treatment regimens included ganciclovir. The overall survival rate was 35% at 2 years following the onset of enteritis.

Epidemiology of Aspergillus infections in a large cohort of patients undergoing bone marrow transplantation.

To investigate the incidence, risk factors, and outcome of Aspergillus infections among marrow transplant recipients, records from 2496 patients were reviewed, and 214 patients had Aspergillus organisms identified. Of these, 158 had invasive aspergillosis, 44 were colonized, and 12 had contaminated cultures. The incidence of invasive aspergillosis increased from 5.7% to 11.2% during the study. The onset of infection was bimodal, peaking 16 and 96 days after transplant. For patients within 40 days after transplant, underlying disease, donor type, season, and transplant outside of laminar air flow rooms were associated with significant risk for invasive aspergillosis. For patients >40 days after transplant, age, underlying disease, donor type, graft-versus-host disease, neutropenia, and corticosteroid use were associated with increased risk of aspergillosis. Only 31% of infected patients were neutropenic at the time of diagnosis. The risk factors for aspergillosis depend on the time after marrow transplant and include both host and environmental characteristics.

Phase I study of amphotericin B colloidal dispersion for the treatment of invasive fungal infections after marrow transplant.

Amphotericin B colloidal dispersion (ABCD; Amphocil) was evaluated in a phase I dose-escalation study in 75 marrow transplant patients with invasive fungal infections (primarily Aspergillus or Candida species) to determine the toxicity profile, maximum tolerated dose, and clinical response. Escalating doses of 0.5-8.0 mg/kg in 0.5-mg/kg/patient increments were given up to 6 weeks. No infusion-related toxicities were observed in 32% of the patients; 52% had grade 2 and 5% had grade 3 toxicity. No appreciable renal toxicity was observed at any dose level. The estimated maximum tolerated dose was 7.5 mg/kg, defined by rigors and chills and hypotension in 3 of 5 patients at 8.0 mg/kg. The complete or partial response rate across dose levels and infection types was 52%. For specific types of infections, 53% of patients with fungemia had complete responses, and 52% of patients with pneumonia had complete or partial responses. ABCD was safe at doses to 7.5 mg/kg and had tolerable-infusion-related toxicity and demonstrable antifungal activity.

Panfungal PCR assay for detection of fungal infection in human blood specimens.

A novel panfungal PCR assay which detects the small-subunit rRNA gene sequence of the two major fungal organism groups was used to test whole-blood specimens obtained from a series of blood or bone marrow transplant recipients. The 580-bp PCR product was identified after amplification by panfungal primers and hybridization to a 245-bp digoxigenin-labeled probe. The lower limit of detection of the assay was approximately four organisms per milliliter of blood. Multiple whole-blood specimens from five patients without fungal infection or colonization had negative PCR results. Specimens from 11 infected patients had positive PCR results. Blood from three patients with pulmonary aspergillosis had positive PCR results: one patient's blood specimen obtained in the week prior to the diagnosis of infection by a positive bronchoalveolar lavage fluid culture result was positive by PCR, and blood specimens obtained from two patients 1 to 2 days after lung biopsy and which were sterile by culture were positive by PCR. The blood of four patients with candidemia, three patients with mixed fungal infections, and one patient with fusariosis also had positive PCR signals. The panfungal PCR assay can detect multiple fungal genera and may be used as an adjunct to conventional methods for the detection of fungal infection or for describing the natural history of fungal infection. Further studies are needed to define the sensitivity and specificity of this assay for the diagnosis of fungal infection prior to the existence of other clinical or laboratory indications of invasive fungal infection.

Development of fluconazole resistance in Candida albicans causing disseminated infection in a patient undergoing marrow transplantation.

Oral candidiasis due to azole-resistant Candida albicans is an increasing problem in patients with AIDS who received prolonged periods of fluconazole prophylaxis. Infection with C. albicans is also frequent in patients undergoing transplantation. However, azole resistance has not been appreciated as a major problem for these patients, presumably because they receive a relatively short duration of fluconazole prophylaxis. We describe a case of disseminated candidiasis due to fluconazole-resistant C. albicans in a patient following marrow transplantation. Restriction fragment length polymorphism analysis with use of the C. albicans strain-specific Ca3 probe was performed on sequential isolates. Identical banding patterns were obtained, thereby confirming that a fluconazole-susceptible endogenous C. albicans acquired azole resistance during a brief exposure to the drug and subsequently caused disseminated infection. This observation raises questions regarding the incidence, significance, and mechanism of azole resistance in fungi causing infection in this population.

Granulocyte colony-stimulating factor administered in vivo augments neutrophil-mediated activity against opportunistic fungal pathogens.

Granulocyte colony-stimulating factor (G-CSF) not only increases neutrophil (polymorphonuclear leukocyte, PMNL) production but also modulates PMNL biologic function. To assess the ability of G-CSF administered in vivo to enhance PMNL activity against opportunistic fungal pathogens, the antifungal activity of PMNL obtained from normal human volunteers before and after G-CSF administration was compared. In vivo, G-CSF significantly enhanced PMNL-mediated killing of Aspergillus fumigatus and Rhizopus arrhizus by 4-fold and 15-fold, respectively (P < .05). In contrast, PMNL-mediated killing of Candida albicans was unaffected by G-CSF. The ability of aqueous fungal extracts to induce the PMNL respiratory burst was evaluated by luminol-enhanced chemiluminescence. G-CSF in vivo primed PMNL for sustained chemiluminescence in response to extracts of Candida, Aspergillus, and Rhizopus organisms. These data demonstrate that G-CSF in vivo augments antifungal activities of PMNL, thereby implicating a possible therapeutic role for G-CSF as a biologic response-modifying agent during opportunistic fungal infection.

Modulation of neutrophil-mediated activity against the pseudohyphal form of Candida albicans by granulocyte colony-stimulating factor (G-CSF) administered in vivo.

Renewed interest in neutrophil transfusions has emerged with the development and clinical use of granulocyte colony-stimulating factor (G-CSF). G-CSF not only increases neutrophil (polymorphonuclear leukocyte, PMNL) production but also modulates various physiological properties of PMNL. The effects of G-CSF on PMNL-mediated fungicidal activity were evaluated by administration of G-CSF (300 micrograms/day subcutaneously) to 5 healthy volunteers for 6 days. G-CSF significantly enhanced PMNL-mediated damage of Candida albicans pseudohyphae by 33% (P=.007) on day 2 and by 44% (P=.04) on day 6 at a 10:1 effector:target ratio. In contrast, the ability of PMNL to induce damage of hyphae from either Fusarium solani or Aspergillus fumigatus did not significantly change during the study period. These data demonstrate that G-CSF administered in vivo modulates PMNL-mediated fungicidal activity against the pseudohyphal form of C. albicans, thereby suggesting potential utility of G-CSF as a biologic response-modifying therapy in some opportunistic fungal infections.

Comparison of interferon-gamma, granulocyte colony-stimulating factor, and granulocyte-macrophage colony-stimulating factor for priming leukocyte-mediated hyphal damage of opportunistic fungal pathogens.

Proinflammatory cytokines have been proposed as adjunctive therapeutic agents to enhance the host immune response during infections caused by opportunistic fungi. The study compared the differential in vitro priming effects of interferon-gamma (IFN-gamma), granulocyte colony-stimulating factor (G-CSF), and granulocyte-macrophage colony-stimulating factor (GM-CSF) on hyphal damage of opportunistic fungi mediated by isolated neutrophils (polymorphonuclear leukocytes, PMNL) and buffy coat cells (polymorphonuclear leukocytes/peripheral blood mononuclear cells, PMNL/PBMC) from healthy donors. IFN-gamma (1000 U/mL) effectively primed both PMNL and PMNL/PBMC for enhanced hyphal damage of Aspergillus fumigatus, Fusarium solani, and Candida albicans. G-CSF (100 ng/mL) increased hyphal damage mediated by both PMNL and PMNL/PBMC against F. solani, and GM-CSF (100 ng/mL) augmented the antifungal activity of PMNL/PBMC against hyphal forms of both F. solani and C. albicans. IFN-gamma may be superior to G-CSF or GM-CSF for enhancing the microbicidal activity of PMNL and PMNL/PBMC against opportunistic fungi.

Comparison of six extraction techniques for isolation of DNA from filamentous fungi.

Filamentous fungi have a sturdy cell wall which is resistant to the usual DNA extraction procedures. We determined the DNA extraction procedure with the greatest yield of high quality fungal DNA and the least predilection for cross-contamination of equipment between specimens. Each of six extraction methods was performed using Aspergillus fumigatus hyphae. The six methods were: (1) glass bead pulverization with vortexing; (2) grinding with mortar and pestle followed by glass bead pulverization; (3) glass bead pulverization using 1% hydroxyacetyl trimethyl ammonium bromide (CTAB) buffer in a water bath sonicator; (4) water bath sonication in CTAB buffer; (5) grinding followed by incubation with CTAB; and (6) lyticase enzymatic cell lysis. Genomic DNA yields were measured by spectrophotometry and by visual reading of 2% agarose gels, with shearing assessed by the migration of the DNA on the gel. Genomic fungal DNA yields were highest for Method 1, followed by Methods 5 approximately = to 2 >3 approximately = to 4 approximately = to 6. Methods 2 and 5, both of which involved grinding with mortar and pestle, led to shearing of the genomic DNA in one of two trials each. We conclude that the use of glass beads with extended vortexing is optimal for extraction of microgramme amounts of DNA from filamentous fungal cultures.

Nocardiosis after bone marrow transplantation: a retrospective study.

To evaluate the spectrum of nocardiosis after marrow transplantation, we reviewed the medical records of 27 patients with nocardiosis who were treated at three centers, and we reviewed the findings of three cases reported in the literature. Nocardial involvement was defined as invasive nocardiosis (n = 25), colonization (n = 4), or contamination (n = 1). The median time to the diagnosis of nocardiosis after marrow transplantation was 210 days. Nocardia asteroides complex accounted for 96% of isolates. All 25 invasive infections occurred in allogeneic marrow recipients. Ten (40%) of 25 patients with invasive nocardiosis were receiving double-strength oral trimethoprimsulfamethoxazole twice weekly as prophylaxis for Pneumocystis carinii pneumonia. Treatment regimens for nocardiosis included sulfonamides; synergistic agents were also often added. The overall survival rate at 6 years was 34%; survival from the infection itself was 84%. Two of four nocardiosis-related deaths also involved other pathogens. The incidence of nocardiosis among allogeneic marrow recipients averaged 0.3% over 25 years. We conclude that nocardiosis is a rare infection that occurs later after marrow transplantation than other infections and that is marginally associated with increased mortality among long-term survivors of allogeneic marrow transplantation.

Determinants of human immunodeficiency virus DNA and RNA shedding in the anal-rectal canal of homosexual men.

To define the determinants of anal-rectal shedding of human immunodeficiency virus (HIV) DNA and RNA, 374 HIV-seropositive homosexual men were tested. Factors independently associated with detection of anal-rectal HIV DNA included anal-rectal inflammation and detection of anal human papillomavirus DNA; predictors of HIV RNA included detection of anal-rectal HIV DNA, anal-rectal inflammation, and high plasma HIV RNA levels. The latter (>10,000 copies/mL) was the main determinant of anal-rectal HIV RNA shedding when HIV DNA (e.g., HIV-infected cells) was not detected in the anal-rectal sample. The local presence of HIV-infected cells and local inflammation were the principal determinants of HIV RNA among those with low (<10,000 copies/mL) plasma HIV RNA load. Among those with anal-rectal HIV DNA present, increased HIV RNA plasma load did not increase the risk of shedding of HIV RNA into the anal-rectal canal.

A phase I-II clinical trial to evaluate removal of CD4 cells and partial depletion of CD8 cells from donor marrow for HLA-mismatched unrelated recipients.

We conducted a phase I-II clinical trial to test the hypothesis that removal of CD4 cells from an HLA-mismatched unrelated marrow graft would substantially reduce the risk of grades III-IV graft-versus-host disease (GVHD) and that retention of a specified number of CD8 cells in the graft would be sufficient to prevent rejection. Patients were eligible for this study when an HLA-A, -B, or -DRB1-matched unrelated donor could not be identified. HLA matching of the donor and recipient was based on typing of HLA-A and -B antigens by serologic methods and by typing of HLA-DRB1 alleles by molecular methods, and donors were selected when disparity was limited to a single HLA-DRB1 allele or a single HLA-A or -B antigen. Twenty-seven patients with hematologic malignancy or aplastic anemia were prepared to receive a transplant with conventional regimens of cyclophosphamide and fractionated total body irradiation, and a standard regimen of methotrexate and cyclosporine was given for GVHD prophylaxis. CD4 cells were removed from the donor marrow, and the numbers of CD8 cells were adjusted systematically in graded steps for successive patients, depending on the occurrence of grades III-IV GVHD or graft failure in previously enrolled patients. Removal of CD4 cells did not cause graft rejection or appreciably decrease the risk of grades III-IV GVHD. Depletion of CD8 cells was associated with an increased risk of rejection with either HLA-DRB1 disparity or with HLA-A or -B disparity. With either type of disparity, the risk of grades III-IV GVHD is likely to be higher than 15% at any dose of CD8 cells associated with less than 5% risk of graft failure. The absence of graft failure associated with CD4 depletion supports the hypothesis that donor CD4 cells are not essential for preventing marrow graft rejection in humans. The correlation between graft failure and the number of CD8 cells in the donor marrow supports the hypothesis that donor CD8 cells help to prevent marrow graft rejection.

Randomized, double-blind clinical trial of amphotericin B colloidal dispersion vs. amphotericin B in the empirical treatment of fever and neutropenia.

We conducted a prospective, randomized, double-blind study comparing amphotericin B colloidal dispersion (ABCD) with amphotericin B in the empirical treatment of fever and neutropenia. Patients with neutropenia and unresolved fever after > or = 3 days of empirical antibiotic therapy were stratified by age and concomitant use of cyclosporine or tacrolimus. Patients were then randomized to receive therapy with ABCD (4 mg/[kg.d]) or amphotericin B (0.8 mg/[kg.d]) for < or = 14 days. A total of 213 patients were enrolled, of whom 196 were evaluable for efficacy. Fifty percent of ABCD-treated patients and 43.2% of amphotericin B-treated patients had a therapeutic response (P = .31). Renal dysfunction was less likely to develop and occurred later in ABCD recipients than in amphotericin B recipients (P < .001 for both parameters). Infusion-related hypoxia and chills were more common in ABCD recipients than in amphotericin B recipients (P = .013 and P = .018, respectively). ABCD appeared comparable in efficacy with amphotericin B, and renal dysfunction associated with ABCD was significantly less than that associated with amphotericin B. However, infusion-related events were more common with ABCD treatment than with amphotericin B treatment.