Robert L. Schaufele

Epidemiology and Outcome of Zygomycosis: A Review of 929 Reported Cases

BACKGROUND Zygomycosis is an increasingly emerging life-threatening infection. There is no single comprehensive literature review that describes the epidemiology and outcome of this disease. METHODS We reviewed reports of zygomycosis in the English-language literature since 1885 and analyzed 929 eligible cases. We included in the database only those cases for which the underlying condition, the pattern of infection, the surgical and antifungal treatments, and survival were described. RESULTS The mean age of patients was 38.8 years; 65% were male. The prevalence and overall mortality were 36% and 44%, respectively, for diabetes; 19% and 35%, respectively, for no underlying condition; and 17% and 66%, respectively, for malignancy. The most common types of infection were sinus (39%), pulmonary (24%), and cutaneous (19%). Dissemination developed in 23% of cases. Mortality varied with the site of infection: 96% of patients with disseminated disease died, 85% with gastrointestinal infection died, and 76% with pulmonary infection died. The majority of patients with malignancy (92 [60%] of 154) had pulmonary disease, whereas the majority of patients with diabetes (222 [66%] of 337) had sinus disease. Rhinocerebral disease was seen more frequently in patients with diabetes (145 [33%] of 337), compared with patients with malignancy (6 [4%] of 154). Hematogenous dissemination to skin was rare; however, 78 (44%) of 176 cutaneous infections were complicated by deep extension or dissemination. Survival was 3% (8 of 241 patients) for cases that were not treated, 61% (324 of 532) for cases treated with amphotericin B deoxycholate, 57% (51 of 90) for cases treated with surgery alone, and 70% (328 of 470) for cases treated with antifungal therapy and surgery. By multivariate analysis, infection due to Cunninghamella species and disseminated disease were independently associated with increased rates of death (odds ratios, 2.78 and 11.2, respectively). CONCLUSIONS Outcome from zygomycosis varies as a function of the underlying condition, site of infection, and use of antifungal therapy.

Detection of Circulating Candida Enolase by Immunoassay in Patients with Cancer and Invasive Candidiasis

BACKGROUND Invasive candidiasis is a major nosocomial infection that is difficult to diagnose. Few biochemically defined markers of invasive candidiasis are known. Initial findings suggested that the presence of candida enolase in the blood may be a novel marker for invasive candidiasis. METHODS We tested 170 patients at high risk for invasive candidiasis for candida enolase antigenemia. All the patients had cancer and neutropenia. We detected antigen using a double-sandwich liposomal immunoassay for candida enolase in serially collected serum samples. Invasive candidiasis was proved by finding candida species in deep nonmucosal tissue, blood cultures, or both. Antigen testing was performed with the investigator blinded to tissue or culture diagnosis. RESULTS Among 24 patients with proved invasive candidiasis, 149 serum samples were tested for enolase antigenemia; 80 were positive and 69 negative (sensitivity per sample, 54 percent). Multiple sampling improved the detection of antigenemia, which was found in 11 of 13 proved cases of deep tissue infection (85 percent) and in 7 of 11 proved cases of fungemia (64 percent). Specificity was 96 percent as measured against control groups including patients with mucosal colonization, bacteremia, and other deep mycoses. Antigenemia was detected in the absence of fungemia in 5 cases of deep tissue candidiasis, but was not detected in 6 cases of fungemia alone. CONCLUSIONS Candida enolase antigenemia is a novel marker for invasive candidiasis. It may be a useful indicator of deep infection in patients with cancer and neutropenia and may complement the diagnostic usefulness of blood cultures.

Experimental pulmonary aspergillosis due to Aspergillus terreus : pathogenesis and treatment of an emerging fungal pathogen resistant to amphotericin B

Aspergillus terreus is an uncommon but emerging fungal pathogen, which causes lethal infections that are often refractory to amphotericin B (AmB). In comparison to Aspergillus fumigatus, A. terreus was resistant to the in vitro fungicidal effects of safely achievable concentrations of AmB. These in vitro findings correlated directly with resistance of A. terreus to AmB in experimental invasive pulmonary aspergillosis. Residual fungal pulmonary burden and galactomannan antigenemia demonstrated persistent infection, despite therapy with deoxycholate AmB or liposomal AmB. By comparison, posaconazole and itraconazole resolved GM antigenemia, reduced residual fungal burden, and improved survival. There were no differences in phagocytic host response to A. terreus versus A. fumigatus; however, the rate of conidial germination of A. terreus was slower. The strain of A. terreus with the highest minimum inhibitory and minimum lethal concentration of AmB also had the lowest membrane ergosterol content. The hyphae of A. terreus in vivo displayed distinctive aleurioconidia, which may be a practical microscopic feature for rapid preliminary diagnosis.

Antifungal Efficacy of Caspofungin (MK-0991) in Experimental Pulmonary Aspergillosis in Persistently Neutropenic Rabbits: Pharmacokinetics, Drug Disposition, and Relationship to Galactomannan Antigenemia

ABSTRACT The antifungal efficacy, pharmacokinetics, and safety of caspofungin (CAS) were investigated in the treatment and prophylaxis of invasive pulmonary aspergillosis due to Aspergillus fumigatus in persistently neutropenic rabbits. Antifungal therapy consisted of 1, 3, or 6 mg of CAS/kg of body weight/day (CAS1, CAS3, and CAS6, respectively) or 1 mg of deoxycholate amphotericin B (AMB)/kg/day intravenously for 12 days starting 24 h after endotracheal inoculation. Prophylaxis (CAS1) was initiated 4 days before endotracheal inoculation. Rabbits treated with CAS had significant improvement in survival and reduction in organism-mediated pulmonary injury (OMPI) measured by pulmonary infarct score and total lung weight (P < 0.01). However, animals treated with CAS demonstrated a paradoxical trend toward increased residual fungal burden (log CFU per gram) and increased serum galactomannan antigen index (GMI) despite improved survival. Rabbits receiving prophylactic CAS1 also showed significant improvement in survival and reduction in OMPI (P < 0.01), but there was no effect on residual fungal burden. In vitro tetrazolium salt hyphal damage assays and histologic studies demonstrated that CAS had concentration- and dose-dependent effects on hyphal structural integrity. In parallel with a decline in GMI, AMB significantly reduced the pulmonary tissue burden of A. fumigatus (P ≤ 0.01). The CAS1, CAS3, and CAS6 dose regimens demonstrated dose-proportional exposure and maintained drug levels in plasma above the MIC for the entire 24-h dosing interval at doses that were ≥3 mg/kg/day. As serial galactomannan antigen levels may be used for therapeutic monitoring, one should be aware that profoundly neutropenic patients receiving echinocandins for aspergillosis might have persistent galactomannan antigenemia despite clinical improvement. CAS improved survival, reduced pulmonary injury, and caused dose-dependent hyphal damage but with no reduction in residual fungal burden or galactomannan antigenemia in persistently neutropenic rabbits with invasive pulmonary aspergillosis.

Detection of Galactomannan Antigenemia in Patients Receiving Piperacillin-Tazobactam and Correlations between In Vitro, In Vivo, and Clinical Properties of the Drug-Antigen Interaction

ABSTRACT Recent case reports describe patients receiving piperacillin-tazobactam who were found to have circulating galactomannan detected by the double sandwich enzyme-linked immunosorbent assay (ELISA) system, leading to the false presumption of invasive aspergillosis. Since this property of piperacillin-tazobactam and galactomannan ELISA is not well understood, we investigated the in vitro, in vivo, and clinical properties of this interaction. Among the 12 reconstituted antibiotics representing four classes of antibacterial compounds that are commonly used in immunocompromised patients, piperacillin-tazobactam expressed a distinctively high level of galactomannan antigen in vitro (P = 0.001). After intravenous infusion of piperacillin-tazobactam into rabbits, the serum galactomannan index (GMI) in vivo changed significantly (P = 0.0007) from a preinfusion mean baseline value of 0.27 to a mean GMI of 0.83 by 30 min to slowly decline to a mean GMI of 0.44 24 h later. Repeated administration of piperacillin-tazobactam over 7 days resulted in accumulation of circulating galactomannan to a mean peak GMI of 1.31 and a nadir of 0.53. Further studies revealed that the antigen reached a steady state by the third day of administration of piperacillin-tazobactam. Twenty-six hospitalized patients with no evidence of invasive aspergillosis who were receiving antibiotics and ten healthy blood bank donors were studied for expression of circulating galactomannan. Patients (n = 13) receiving piperacillin-tazobactam had significantly greater mean serum GMI values (0.74 ± 0.14) compared to patients (n = 13) receiving other antibiotics (0.14 ± 0.08) and compared to healthy blood bank donors (0.14 ± 0.06) (P < 0.001). Five (38.5%) of thirteen patients receiving piperacillin-tazobactam had serum GMI values > 0.5 compared to none of thirteen subjects receiving other antibiotics (P = 0.039) and to none of ten healthy blood bank donors (P = 0.046). These data demonstrate that among antibiotics that are commonly used in immunocompromised patients, only piperacillin-tazobactam contains significant amounts of galactomannan antigen in vitro, that in animals receiving piperacillin-tazobactam circulating galactomannan antigen accumulates in vivo to significantly increased and sustained levels, and that some but not all patients receiving this antibiotic will demonstrate circulating galactomannan above the threshold considered positive for invasive aspergillosis by the recently licensed double sandwich ELISA.

Zygomycosis in Children: A Systematic Review and Analysis of Reported Cases

Background: Zygomycosis has emerged as an increasingly important infection with a high mortality especially in immunocompromised patients. No comprehensive analysis of pediatric zygomycosis cases has been published to date. Methods: We used a PUBMED search for English publications of pediatric (0–18 years) zygomycosis cases and references from major books as well as single case reports or case series. Individual references were reviewed for additional cases. Data were entered into Filemaker-pro database and analyzed by logistic regression analysis. Results: One hundred fifty-seven cases (64% male) were found with median age 5 years (range, 0.16–13). Underlying conditions included neutropenia (18%), prematurity (17%), diabetes mellitus (15%), ketoacidosis (10%), and no apparent underlying condition (14%). The most common patterns of zygomycosis were cutaneous (27%), gastrointestinal (21%), rhinocerebral (18%), and pulmonary (16%). Among 77 culture-confirmed cases, Rhizopus spp. (44%) and Mucor spp. (15%) were most commonly identified. Of 81 patients who were given antifungal therapy, 73% received an amphotericin B formulation only. The remaining patients received mostly amphotericin B in combination with other antifungal agents. Mortality in patients without antifungal therapy was higher than in those with therapy (88% versus 36%, P < 0.0001). Ninety-two (59%) patients underwent surgery. Cerebral, gastrointestinal, disseminated and cutaneous zygomycosis were associated with mortality rates of 100, 100, 88, and 0%, respectively. Independent risk factors for death were disseminated infection (OR: 7.18; 95% CI: 3.02–36.59) and age <1 year (OR: 3.85; 95% CI: 1.05–7.43). Antifungal therapy and particularly surgery reduced risk of death by 92% (OR: 0.07; 95% CI: 0.04–0.25) and 84% (OR: 0.16; 95% CI: 0.09–0.61), respectively. Conclusions: Zygomycosis is a life-threatening infection in children with neutropenia, diabetes mellitus, and prematurity as common predisposing factors, and there is high mortality in untreated disease, disseminated infection, and age <1 year. Amphotericin B and surgery significantly improve outcome.

Host-Dependent Patterns of Tissue Injury in Invasive Pulmonary Aspergillosis

Invasive pulmonary aspergillosis (IPA) is an important cause of morbidity and mortality in neutropenic, nonneutropenic, and other immunocompromised patients. We therefore compared the patterns of infection and inflammation among 3 cohorts of immunocompromised patients with profound neutropenia, nonneutropenic immunosuppression, and hematopoietic stem cell transplantation. Lesions of IPA in neutropenic patients and hematopoietic stem cell transplant (HSCT) recipients were similar and consisted predominantly of angioinvasion and intraalveolar hemorrhage. The frequency of these histologic findings in neutropenic patients and HSCT recipients differed significantly from those of nonneutropenic patients (P < .05). It is noteworthy that even if HSCT recipients have normal peripheral blood neutrophil counts, there may be no influx into sites of infection. In the nonneutropenic cohort, lesions of IPA consisted mainly of neutrophilic and monocytic infiltrates and inflammatory necrosis. Thus, the status of innate host defenses contributes significantly to the histologic patterns observed in IPA.

Triazole-Polyene Antagonism in Experimental Invasive Pulmonary Aspergillosis: In Vitro and In Vivo Correlation

Combination antifungal therapy is increasingly used in the treatment of invasive aspergillosis. Whether the interaction between amphotericin B and triazoles is antagonistic against invasive aspergillosis is a controversial issue that is not likely to be resolved through a randomized clinical trial. Here, we found both in vitro and in vivo antagonism between liposomal amphotericin B and ravuconazole in simultaneous treatment of experimental invasive pulmonary aspergillosis in persistently neutropenic rabbits. Bliss independence-based drug-interaction modeling showed significant antagonism in vitro and in vivo, with the observed drug effects being 20%-69% lower than would be expected if the drugs were acting independently. These in vitro and in vivo findings of antagonism were consistent with the findings from Loewe additivity-based drug-interaction modeling. No pharmacokinetic interaction was found. The combination of a triazole and polyene may be antagonistic in the treatment of invasive pulmonary aspergillosis.

Characterization and Comparison of Galactomannan Enzyme Immunoassay and Quantitative Real-Time PCR Assay for Detection of Aspergillus fumigatus in Bronchoalveolar Lavage Fluid from Experimental Invasive Pulmonary Aspergillosis

ABSTRACT Bronchoalveolar lavage (BAL) is widely used for evaluation of patients with suspected invasive pulmonary aspergillosis (IPA). However, the diagnostic yield of BAL for detection of IPA by culture and direct examination is limited. Earlier diagnosis may be facilitated by assays that can detect Aspergillus galactomannan antigen or DNA in BAL fluid. We therefore characterized and compared the diagnostic yields of a galactomannan enzyme immunoassay (GM EIA), quantitative real-time PCR (qPCR), and quantitative cultures in experiments using BAL fluid from neutropenic rabbits with experimentally induced IPA defined as microbiologically and histologically evident invasion. The qPCR assay targeted the rRNA gene complex of Aspergillus fumigatus. The GM EIA and qPCR assay were characterized by receiver operator curve analysis. With an optimal cutoff of 0.75, the GM EIA had a sensitivity and specificity of 100% in untreated controls. A decline in sensitivity (92%) was observed when antifungal therapy (AFT) was administered. The optimal cutoff for qPCR was a crossover of 36 cycles, with sensitivity and specificity of 80% and 100%, respectively. The sensitivity of qPCR also decreased with AFT to 50%. Quantitative culture of BAL had a sensitivity of 46% and a specificity of 100%. The sensitivity of quantitative culture decreased with AFT to 16%. The GM EIA and qPCR assay had greater sensitivity than culture in detection of A. fumigatus in BAL fluid in experimentally induced IPA (P ± 0.04). Use of the GM EIA and qPCR assay in conjunction with culture-based diagnostic methods applied to BAL fluid could facilitate accurate diagnosis and more-timely initiation of specific therapy.

Combination Therapy in Treatment of Experimental Pulmonary Aspergillosis: In Vitro and In Vivo Correlations of the Concentration- and Dose- Dependent Interactions between Anidulafungin and Voriconazole by Bliss Independence Drug Interaction Analysis

ABSTRACT We studied the antifungal activity of anidulafungin (AFG) in combination with voriconazole (VRC) against experimental invasive pulmonary aspergillosis (IPA) in persistently neutropenic rabbits and further explored the in vitro and in vivo correlations by using Bliss independence drug interaction analysis. Treatment groups consisted of those receiving AFG at 5 (AFG5 group) and 10 (AFG10 group) mg/kg of body weight/day, VRC at 10 mg/kg every 8 h (VRC group), AFG5 plus VRC (AFG5+VRC group), and AFG10 plus VRC (AFG10+VRC group) and untreated controls. Survival throughout the study was 60% for the AFG5+VRC group, 50% for the VRC group, 27% for the AFG10+VRC group, 22% for the AFG5 group, 18% for the AFG10 group, and 0% for control rabbits (P < 0.001). There was a significant reduction of organism-mediated pulmonary injury, measured by infarct scores, lung weights, residual fungal burdens, and galactomannan indexes, in AFG5+VRC-treated rabbits versus those treated with AFG5 and VRC alone (P < 0.05). In comparison, AFG10+VRC significantly lowered only infarct scores and lung weights in comparison to those of AFG10-treated animals (P < 0.05). AFG10+VRC showed no significant difference in other outcome variables. Significant Bliss synergy was found in vivo between AFG5 and VRC, with observed effects being 24 to 30% higher than expected levels if the drugs were acting independently. These synergistic interactions were also found between AFG and VRC in vitro. However, for AFG10+VRC, only independence and antagonism were observed among the outcome variables. We concluded that the combination of AFG with VRC in treatment of experimental IPA in persistently neutropenic rabbits was independent to synergistic at a dosage of 5 mg/kg/day but independent to antagonistic at 10 mg/kg/day, as assessed by Bliss independence analysis, suggesting that higher dosages of an echinocandin may be deleterious to the combination.