Michael A. Pfaller

Epidemiology of Invasive Mycoses in North America

The incidence of invasive mycoses is increasing, especially among patients who are immunocompromised or hospitalized with serious underlying diseases. Such infections may be broken into two broad categories: opportunistic and endemic. The most important agents of the opportunistic mycoses are Candida spp., Cryptococcus neoformans, Pneumocystis jirovecii, and Aspergillus spp. (although the list of potential pathogens is ever expanding); while the most commonly encountered endemic mycoses are due to Histoplasma capsulatum, Coccidioides immitis/posadasii, and Blastomyces dermatitidis. This review discusses the epidemiologic profiles of these invasive mycoses in North America, as well as risk factors for infection, and the pathogens’ antifungal susceptibility.

Invasive Fungal Pathogens: Current Epidemiological Trends

Patient characteristics, antifungal prophylaxis, and other factors appear to have contributed to a change in the spectrum of invasive fungal pathogens. Infections with Candida glabrata, Aspergillus terreus, and non-Aspergillus moulds appear to be on the rise, at least among certain populations. These species are resistant or less susceptible to some commonly used antifungal agents. Non-Aspergillus moulds are particularly lethal. This article reviews the spectrum of invasive mycoses and risk factors for infection with these pathogens.

Progress in Antifungal Susceptibility Testing of Candida spp. by Use of Clinical and Laboratory Standards Institute Broth Microdilution Methods, 2010 to 2012

ABSTRACT Antifungal susceptibility testing of Candida has been standardized and refined and now may play a useful role in managing Candida infections. Important new developments include validation of 24-h reading times for all antifungal agents and the establishment of species-specific epidemiological cutoff values (ECVs) for the systemically active antifungal agents and both common and uncommon species of Candida. The clinical breakpoints (CBPs) for fluconazole, voriconazole, and the echinocandins have been revised to provide species-specific interpretive criteria for the six most common species. The revised CBPs not only are predictive of clinical outcome but also provide a more sensitive means of identifying those strains with acquired or mutational resistance mechanisms. This brief review serves as an update on the new developments in the antifungal susceptibility testing of Candida spp. using Clinical and Laboratory Standards Institute (CLSI) broth microdilution (BMD) methods.

Overview of the Epidemiological Profileand Laboratory Detection of Extended-Spectrum β-Lactamases

Extended-spectrum beta-lactamases (ESBLs) are plasmid-mediated bacterial enzymes that confer resistance to a broad range of beta-lactams. They are descended by genetic mutation from native beta-lactamases found in gram-negative bacteria, especially infectious strains of Escherichia coli and Klebsiella species. Genetic sequence modifications have broadened the substrate specificity of the enzymes to include third-generation cephalosporins, such as ceftazidime. Because ESBL-producing strains are resistant to a wide variety of commonly used antimicrobials, their proliferation poses a serious global health concern that has complicated treatment strategies for a growing number of hospitalized patients. Another resistance mechanism, also common to Enterobacteriaceae, results from the overproduction of chromosomal or plasmid-derived AmpC beta-lactamases. These organisms share an antimicrobial resistance pattern similar to that of ESBL-producing organisms, with the prominent exception that, unlike most ESBLs, AmpC enzymes are not inhibited by clavulanate and similar beta-lactamase inhibitors. Recent technological improvements in testing and in the development of uniform standards for both ESBL detection and confirmatory testing promise to make accurate identification of ESBL-producing organisms more accessible to clinical laboratories.

Azole Resistance in Aspergillus fumigatus Isolates from the ARTEMIS Global Surveillance Study Is Primarily Due to the TR/L98H Mutation in the cyp51A Gene

ABSTRACT We surveyed 497 isolates of Aspergillus fumigatus collected from 2008 to 2009 as part of the ARTEMIS global surveillance study for elevated MIC values to itraconazole, voriconazole, and posaconazole. Sequencing of the cyp51A gene revealed that 8/29 isolates with elevated MIC values to one or more triazoles, all originating in China, contained the TR/L98H mutation associated with resistant European isolates of A. fumigatus. This is the first time the TR/L98H mutation has been identified outside Europe.

Geographic variation in the frequency of isolation and fluconazole and voriconazole susceptibilities of Candida glabrata: an assessment from the ARTEMIS DISK Global Antifungal Surveillance Program

Geographic differences in frequency and azole resistance among Candida glabrata may impact empiric antifungal therapy choice. We examined geographic variation in isolation and azole susceptibility of C. glabrata. We examined 23 305 clinical isolates of C. glabrata during ARTEMIS DISK global surveillance. Susceptibility testing to fluconazole and voriconazole was assessed by disk diffusion, and the results were grouped by geographic location: North America (NA) (2470 isolates), Latin America (LA) (2039), Europe (EU) (12 439), Africa and the Middle East (AME) (728), and Asia-Pacific (AP) (5629). Overall, C. glabrata accounted for 11.6% of 201 653 isolates of Candida and varied as a proportion of all Candida isolated from 7.4% in LA to 21.1% in NA. Decreased susceptibility (S) to fluconazole was observed in all geographic regions and ranged from 62.8% in AME to 76.7% in LA. Variation in fluconazole susceptibility was observed within each region: AP (range, 50-100% S), AME (48-86.9%), EU (44.8-88%), LA (43-92%), and NA (74.5-91.6%). Voriconazole was more active than fluconazole (range, 82.3-84.2% S) with similar regional variation. Among 22 sentinel sites participating in ARTEMIS from 2001 through 2007 (84 140 total isolates, 8163 C. glabrata), the frequency of C. glabrata isolation increased in 14 sites and the frequency of fluconazole resistance (R) increased in 11 sites over the 7-year period of study. The sites with the highest cumulative rates of fluconazole R were in Poland (22% R), the Czech Republic (27% R), Venezuela (27% R), and Greece (33% R). C. glabrata was most often isolated from blood, normally sterile body fluids and urine. There is substantial geographic and institutional variation in both frequency of isolation and azole resistance among C. glabrata. Prompt species identification and fluconazole susceptibility testing are necessary to optimize therapy for invasive candidiasis.

Characterization of biofilms formed by Candida parapsilosis, C. metapsilosis, and C. orthopsilosis.

Infections due to Candida parapsilosis have been associated with the ability of this fungus to form biofilms on indwelling medical devices. Recently, C. parapsilosis isolates were reclassified into 3 genetically non-identical classes: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Little information is available regarding the ability of these newly reclassified species to form biofilms on biomedical substrates. In this study, we characterized biofilm formation by 10 clinical isolates each of C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Biofilms were allowed to form on silicone elastomer discs to early (6h) or mature (48 h) phases and quantified by tetrazolium (XTT) and dry weight assays. Surface topography and three-dimensional architecture of the biofilms were visualized using scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM), respectively. Metabolic activity assay revealed strain-dependent biofilm forming ability of the 3 species tested, while biomass determination revealed that all 3 species formed equivalent biofilms (P>0.05 for all comparisons). SEM analyses of representative isolates of these species showed biofilms with clusters of yeast cells adherent to the catheter surface. Additionally, confocal microscopy analyses showed the presence of cells embedded in biofilms ranging in thickness between 62 and 85 microm. These results demonstrate that similar to C. parapsilosis, the 2 newly identified Candida species (C. orthopsilosis and C. metapsilosis) were able to form biofilms.

Rapid Detection of Antibiotic-Resistant Organism Carriage for Infection Prevention

Rapid detection of multidrug-resistant organism (MDRO) carriers could help reduce MDRO infections by allowing for faster institution of prevention measures. However, improving the turnaround time (TAT) of a test requires attention to more than the analytic TAT, and will only occur if postanalytic processes (test reporting and care interventions) are also rapid and efficient. Obstacles to rapid MDRO test development include complex evolving resistance mechanisms, performance directly on mixed samples (eg, nares, stool), and adaptation of new methods for routine clinical diagnostic use. Existing data to support the clinical utility of rapid detection (vs standard culture methods) are scant. For these reasons, rapid detection of MDRO carriers remains a work in progress. Future efforts should be on developing rapid tests to detect multidrug-resistant gram-negative rods, particularly those harboring β-lactamases, and on performing clinical trials to determine how best to incorporate rapid detection of MDRO carriage into healthcare-associated infection prevention efforts.

Validation of 24-Hour Posaconazole and Voriconazole MIC Readings versus the CLSI 48-Hour Broth Microdilution Reference Method: Application of Epidemiological Cutoff Values to Results from a Global Candida Antifungal Surveillance Program

ABSTRACT We performed 24- and 48-h MIC determinations of posaconazole and voriconazole against more than 16,000 clinical isolates of Candida species. By using the 24- and 48-h epidemiological cutoff values (ECVs), the categorical agreement between the 24-h and reference 48-h broth microdilution results ranged from 97.1% (C. parapsilosis and voriconazole) to 99.8% (C. krusei and voriconazole), with 0.0 to 2.9% very major discrepancies (VMD). The essential agreement (within 2 log2 dilutions) between the 24- and 48-h results was 99.6% for both posaconazole and voriconazole. The MIC results obtained for both posaconazole and voriconazole after only 24 h of incubation may be used to determine the susceptibilities of Candida spp. to these important antifungal agents. The applications of ECVs to this large collection of Candida isolates suggests the potential to develop 24-h species-specific clinical breakpoints for both posaconazole and voriconazole.

A global evaluation of voriconazole activity tested against recent clinical isolates of Candida spp

Voriconazole susceptibility testing was performed on 7191 Candida spp. from 78 centers worldwide between 2004 and 2007. Voriconazole was very active in vitro (MIC(50)/MIC(90), 0.008/0.25 microg/mL; 98% susceptible). In comparison to 5866 Candida spp. isolates collected during global surveillance from 1997 to 2001, there were no changes in voriconazole mean MIC or MIC distribution.