Tania C. Sorrell

Clinical Practice Guidelines for the Management of Cryptococcal Disease: 2010 Update by the Infectious Diseases Society of America

Cryptococcosis is a global invasive mycosis associated with significant morbidity and mortality. These guidelines for its management have been built on the previous Infectious Diseases Society of America guidelines from 2000 and include new sections. There is a discussion of the management of cryptococcal meningoencephalitis in 3 risk groups: (1) human immunodeficiency virus (HIV)-infected individuals, (2) organ transplant recipients, and (3) non-HIV-infected and nontransplant hosts. There are specific recommendations for other unique risk populations, such as children, pregnant women, persons in resource-limited environments, and those with Cryptococcus gattii infection. Recommendations for management also include other sites of infection, including strategies for pulmonary cryptococcosis. Emphasis has been placed on potential complications in management of cryptococcal infection, including increased intracranial pressure, immune reconstitution inflammatory syndrome (IRIS), drug resistance, and cryptococcomas. Three key management principles have been articulated: (1) induction therapy for meningoencephalitis using fungicidal regimens, such as a polyene and flucytosine, followed by suppressive regimens using fluconazole; (2) importance of early recognition and treatment of increased intracranial pressure and/or IRIS; and (3) the use of lipid formulations of amphotericin B regimens in patients with renal impairment. Cryptococcosis remains a challenging management issue, with little new drug development or recent definitive studies. However, if the diagnosis is made early, if clinicians adhere to the basic principles of these guidelines, and if the underlying disease is controlled, then cryptococcosis can be managed successfully in the vast majority of patients.

Epidemiology and Host- and Variety-Dependent Characteristics of Infection Due to Cryptococcus neoformans in Australia and New Zealand

A prospective population-based study was conducted in Australia and New Zealand during 1994-1997 to elucidate the epidemiology of cryptococcosis due to Cryptococcus neoformans var. neoformans (CNVN) and C. neoformans var. gattii (CNVG) and to relate clinical manifestations to host immune status and cryptococcal variety. The mean annual incidence per 10(6) population was 6.6 in Australia and 2.2 in New Zealand. Of 312 episodes, CNVN caused 265 (85%; 98% of the episodes in immunocompromised hosts) and CNVG caused 47 (15%; 44% of the episodes in immunocompetent hosts). The incidence of AIDS-associated cases in Australia declined annually (P<.001). Aborigines in rural or semirural locations (P<.001) and immunocompetent males (P<.001) were at increased risk of CNVG infection. Cryptococcomas in lung or brain were more common in immunocompetent hosts (P< or =.03) in whom there was an association only between lung cryptococcomas and CNVG. An AIDS-associated genetic profile of CNVN serotype A was confirmed by random amplification of polymorphic DNA analysis. Resistance to antifungal drugs was uncommon. The epidemiology of CNVN infection has changed substantially. Clinical manifestations of disease are influenced more strongly by host immune status than by cryptococcal variety.

Extracellular phospholipase activity is a virulence factor for Cryptococcus neoformans

The human pathogenic fungus Cryptococcus neoformans secretes a phospholipase enzyme that demonstrates phospholipase B (PLB), lysophospholipase hydrolase and lysophospholipase transacylase activities. This enzyme has been postulated to be a cryptococcal virulence factor. We cloned a phospholipase‐encoding gene (PLB1) from C. neoformans and constructed plb1 mutants using targeted gene disruption. All three enzyme activities were markedly reduced in the mutants compared with the wild‐type parent. The plb1 strains did not have any defects in the known cryptococcal virulence phenotypes of growth at 37°C, capsule formation, laccase activity and urease activity. The plb1 strains were reconstituted using the wild‐type locus and this resulted in restoration of all extracellular PLB activities. In vivo testing demonstrated that the plb1 strain was significantly less virulent than the control strains in both the mouse inhalational model and the rabbit meningitis model. We also found that the plb1 strain exhibited a growth defect in a macrophage‐like cell line. These data demonstrate that secretory phospholipase is a virulence factor for C. neoformans.

Molecular typing of global isolates of Cryptococcus neoformans var. neoformans by polymerase chain reaction fingerprinting and randomly amplified polymorphic DNA-a pilot study to standardize techniques on which to base a detailed epidemiological survey.

A total of 356 clinical isolates of the encapsulated basidiomycetous fungus Cryptococcus neoformans var. neoformans, obtained from Australia, Argentina, Brazil, India, Italy, New Zealand, Papua New Guinea, South Africa, Thailand and the USA, were analyzed to lay the basis for a comprehensive evaluation of the global genetic structure of C. neoformans. Two polymerase chain reaction (PCR)‐based typing techniques were standardized: PCR fingerprinting using a single primer specific to minisatellite or microsatellite DNA, and randomly amplified polymorphic DNA (RAPD) analysis using two combinations of three 20‐ to 22‐mer random primers. Previous studies showed that the resultant profiles are reproducible and stable over time. Identical results were obtained in two different laboratories and by different scientists in the same laboratory. Both typing techniques separated the isolates into four major groups (VNI and VNII, serotype A; VNIII, serotype A/D; and VNIV, serotype D). The majority (78%) of isolates belonged to VNI, compared with 18% VNII, 1% VNIII and 3% VNIV. All US isolates could be differentiated by a unique, strain‐specific PCR fingerprint or RAPD pattern in contrast to most of the non‐US isolates, which showed a substantially higher degree of genetic homogeneity, with some clonality, in different parts of the world. Isolates obtained from the same patient at different times and from different body sites, had identical banding patterns. Both typing techniques should provide powerful tools for epidemiological studies of medically important fungi.

Echinocandin Antifungal Drugs in Fungal Infections A Comparison

This review compares the pharmacology, spectrum of antifungal activity, pharmacokinetic and pharmacodynamic properties, safety and clinical efficacy of the three licensed echinocandins: caspofungin, micafungin and anidulafungin. Echinocandins inhibit the synthesis of 1,3-β-D-glucan, an essential component of the fungal cell wall, and represent a valuable treatment option for fungal infections.The echinocandins exhibit potent in vitro and in vivo fungicidal activity against Candida species, including azole-resistant pathogens. For all agents, strains with drug minimum inhibitory concentrations (MICs) of ≤2 mg/mL are considered susceptible; the MIC at which 90% of isolates tested were inhibited (MIC90) values are typically <2 mg/mL but 100-fold higher MIC90 values are seen with Candida parapsilosis (1–2mg/mL) and Candida guilliermondii (1–4 mg/mL). Activity is comparable between the three agents, although limited data indicate that anidulafungin may have low MICs against C. parapsilosis and Candida glabrata strains that demonstrate elevated MICs to caspofungin and micafungin. All three drugs have good fungistatic activity against Aspergillus spp., although minimal effective concentrations of micafungin and anidulfungin are 2- to 10-fold lower than those for caspofungin. Synergistic/additive in vitro effects of echinocandins when combined with a polyene or azole have been observed.Clinical resistance to the echinocandins is rare despite case reports of caspofungin resistance in several Candida spp. Resistance has been attributed to mutations in the FKS1 gene within two hot spot regions, leading to amino acid substitutions, mostly at position 645 (serine), yet not all FKS1 mutants have caspofungin MICs of >2mg/mL. Of the three echinocandins, the in vitro ‘paradoxical effect’ (increased growth at supra-MIC drug concentrations) is observed least often with anidulafungin.All echinocandins have low oral bioavailability, and distribute well into tissues, but poorly into the CNS and eye. Anidulafungin is unique in that it undergoes elimination by chemical degradation in bile rather than via hepatic metabolism, has a lower maximum concentration and smaller steady state under the concentration-time curve but longer half-life than caspofungin or micafungin. In children, dosing should be based on body surface area. Daily doses of caspofungin (but not micafungin and anidulafungin) should be decreased (from 50 to 35 mg) in moderate liver insufficiency. All echinocandins display concentration-dependent fungicidal (for Candida) or fungistatic (for Aspergillus) activity. The postantifungal effect is 0.9–20 hours against Candida and <0.5 hours against Aspergillus. The echinocandins are well tolerated with few serious drug-drug interactions since they are not appreciable substrates, inhibitors or inducers of the cytochrome P450 or P-glycoprotein systems. In parallel with the greater clinical experience with caspofungin, this agent has a slightly higher potential for adverse effects/drug-drug interactions, with the least potential observed for anidulafungin. Caspofungin (but not micafungin or anidulafungin) dosing should be increased if coadministered with rifampicin and there are modest interactions of caspofungin with calcineurin inhibitors.All three agents are approved for the treatment of oesophageal candidiasis, candidaemia and other select forms of invasive candidiasis. Only mica-fungin is licensed for antifungal prophylaxis in stem cell transplantation, whereas caspofungin is approved for empirical therapy of febrile neutro-penia. Caspofungin has been evaluated in the salvage and primary therapy of invasive aspergillosis. Combination regimens incorporating an echinocandin showing promise in the treatment of aspergillosis. However, echinocandins remain expensive to use.

Development and Clinical Application of a Panfungal PCR Assay To Detect and Identify Fungal DNA in Tissue Specimens

ABSTRACT Given the rise in the incidence of invasive fungal infections (IFIs) and the expanding spectrum of fungal pathogens, early and accurate identification of the causative pathogen is essential. We developed a panfungal PCR assay that targets the internal transcribed spacer 1 (ITS1) region of the ribosomal DNA gene cluster to detect fungal DNA in fresh and formalin-fixed, paraffin-embedded (PE) tissue specimens from patients with culture-proven (n = 38) or solely histologically proven (n = 24) IFIs. PCR products were sequenced and compared with sequences in the GenBank database to identify the causal pathogen. The molecular identification was correlated with results from histological examination and culture. The assay successfully detected and identified the fungal pathogen in 93.6% and 64.3% of culture-proven and solely histologically proven cases of IFI, respectively. A diverse range of fungal genera were identified, including species of Candida, Cryptococcus, Trichosporon, Aspergillus, Fusarium, Scedosporium, Exophiala, Exserohilum, Apophysomyces, Actinomucor, and Rhizopus. For five specimens, molecular analysis identified a pathogen closely related to that identified by culture. All PCR-negative specimens (n = 10) were PE tissues in which fungal hyphae were visualized. The results support the use of the panfungal PCR assay in combination with conventional laboratory tests for accurate identification of fungi in tissue specimens.

Galactomannan and PCR versus culture and histology for directing use of antifungal treatment for invasive aspergillosis in high-risk haematology patients: a randomised controlled trial

BACKGROUND Empirical treatment with antifungal drugs is often used in haematology patients at high risk of invasive aspergillosis. We compared a standard diagnostic strategy (culture and histology) with a rapid biomarker-based diagnostic strategy (aspergillus galactomannan and PCR) for directing the use of antifungal treatment in this group of patients. METHODS In this open-label, parallel-group, randomised controlled trial, eligible patients were adults undergoing allogeneic stem-cell transplantation or chemotherapy for acute leukaemia, with no history of invasive fungal disease. Enrolled patients were randomly assigned (1:1) by a computer-generated schedule to follow either a standard diagnostic strategy (based on culture and histology) or a biomarker-based diagnostic strategy (aspergillus galactomannan and PCR) to direct treatment with antifungal drugs. Patients, were followed up for 26 weeks or until death. Masking of the use of different diagnostic tests was not possible for patients, treating physicians, or investigators. The primary endpoint was empirical treatment with antifungal drugs in the 26 weeks after enrolment (for the biomarker-based diagnostic strategy, a single postive galactomannan or PCR result was deemed insufficient to confirm invasive aspergillosis, so treatment in this context was classified as empirical). This outcome was assessed by an independent data review committee from which the study allocations were masked. Analyses were by intention to treat and included all enrolled patients. This study is registered with ClinicalTrial.gov, number NCT00163722. FINDINGS 240 eligible patients were recruited from six Australian centres between Sept 30, 2005, and Nov 19, 2009. 122 were assigned the standard diagnostic strategy and 118 the biomarker-based diagnostic strategy. 39 patients (32%) in the standard diagnosis group and 18 (15%) in the biomarker diagnosis group received empirical antifungal treatment (difference 17%, 95% CI 4-26; p=0·002). The numbers of patients who had hepatotoxic and nephrotoxic effects did not differ significantly between the standard diagnosis and biomarker diagnosis groups (hepatotoxic effects: 21 [17%] vs 12 [10%], p=0·11; nephrotoxic effects: 52 [43%] vs 60 [51%], p=0·20). INTERPRETATION Use of aspergillus galactomannan and PCR to direct treatment reduced use of empirical antifungal treatment. This approach is an effective strategy for the management of invasive aspergillosis in high-risk haematology patients. FUNDING Australian National Health and Medical Research Council, Cancer Council New South Wales, Pfizer, Merck, Gilead Sciences.

Not Just Little Adults: Candidemia Epidemiology, Molecular Characterization, and Antifungal Susceptibility in Neonatal and Pediatric Patients

OBJECTIVE. The purpose of this work was to identify differences in incidence, risk factors, microbiology, treatment, and clinical outcome of candidemia in neonates, children, and adults that might impact on management. PATIENTS AND METHODS. Cases of candidemia in Australia were identified prospectively by blood culture surveillance over 3 years. Episodes of candidemia in neonatal, pediatric, and adult age groups were analyzed and compared. RESULTS. Of 1005 incident cases, 33 occurred in neonates, 110 in children, and 862 in adults. The respective annual age-specific incidences were 4.4, 0.9, and 1.8 per 100 000 population. Prematurity and ICU admission were major risk factors in neonates. Hematologic malignancy and neutropenia were significantly more frequent in children than in neonates and adults. Diabetes, renal disease, hemodialysis, and recent surgery were more common in adults. Candidemia was attributed to a vascular access device in 58% of neonates, 70% of children, and 44% of adults. Candida albicans caused ∼48% of cases in all of the age groups. Candida parapsilosis was significantly more common in neonates and children (42% and 38% vs 15%). Candida glabrata was infrequent in neonates and children (9% and 3% vs 17%). Significantly more isolates from children were susceptible to fluconazole compared with those from adults (95% vs 75%). Fluconazole-resistant candidal isolates were infrequent in all of the age groups. Neonates and children were more likely to receive amphotericin B compared with adults. Adults were more likely to receive fluconazole. Survival rates at 30 days were 78% in neonates, 90% in children, and 70% in adults. CONCLUSIONS. This study identifies significant differences in candidemia in neonates, children, and adults. Neonatologists and pediatricians must consider age-specific differences when interpreting adult studies and developing treatment and prevention guidelines.

Active surveillance for candidemia, Australia.

This infection has a high death rate and is predominantly associated with healthcare.

Candidaemia in adult cancer patients: risks for fluconazole-resistant isolates and death

BACKGROUND Candidaemia in cancer patients is associated with increasing fluconazole resistance. Models for predicting such isolates and their clinical impact are required. METHODS Clinical, treatment and outcome data from a population-based candidaemia survey (2001-2004) were collected at 5 and 30 days after diagnosis. Speciation and antifungal susceptibility testing was performed. RESULTS There were 138 candidaemia episodes (33% Candida albicans) in adults with haematological malignancies and 150 (51% C. albicans) in adults with solid organ malignancies. Thirty-nine isolates had fluconazole MICs of >or=64 mg/L and 40 had MICs of 16-32 mg/L (predominantly Candida glabrata and Candida krusei). By multivariate analysis, triazole therapy, gastrointestinal tract (GIT) surgery in the 30 days before candidaemia and age >65 years were predictive of fluconazole-resistant candidaemia. Thirty day crude mortality was 40% in haematology patients and 45% in oncology patients. Fluconazole-resistant isolates were associated with increased risk of mortality by univariate (P = 0.04) and Kaplan-Meier survival analyses. By Cox proportional hazards modelling, the strongest predictors of mortality at onset of candidaemia were invasive ventilation, elevated creatinine, intensive care unit (ICU) admission and receipt of systemic triazoles or corticosteroids in the previous 30 days. Removal of a central venous access device (CVAD) at or within 5 days of onset was associated with decreased mortality. CONCLUSIONS Risk factors for fluconazole-resistant candidaemia in adults with cancer include fluconazole/triazole exposure and GIT surgery. ICU admission, invasive ventilation, renal impairment, age >65 years and prior exposure to corticosteroids and triazoles are risk factors for death. CVAD removal reduced mortality. These findings should be integrated into surveillance and treatment algorithms.