The incidence of invasive fungal infections

AN EPIDEMIOLOGIC UPDATE ON INVASIVE FUNGAL INFECTIONS * Michael A. Pfaller, MD ABSTRACT *Based on a presentation given by Dr Pfaller at a symposium held in conjunction with the 43rd Interscience Conference on Antimicrobial Agents and Chemotherapy. Professor of Pathology, Director, Molecular Epidemiology and Fungus Testing Laboratory, Division of Medical Microbiology, Department of Pathology, University of Iowa College of Medicine, Iowa City, Iowa. Address correspondence to: Michael A. Pfaller, MD, Department of Pathology, Medical Microbiology Division, University of Iowa College of Medicine, 200 Hawkins Drive, Room C606GH, Iowa City, IA 52242. E-mail: michael-pfaller@uiowa.edu. Aspergillus and Candida are the 2 major sources of invasive mycoses, and deaths due to these infections have been increasing overall during the past 2 decades. Antimicrobial resistance is an important concern, but the extent of resistance among fungi remains largely undefined. Assessing resistance patterns is essential to determine optimal treatment strategies. For invasive Candida infections, the echinocandins and broad-spectrum triazoles are more potent than fluconazole and amphotericin B against significant clinical isolates of Candida, including the increasing number of fluconazole-resistant isolates (ie, C glabrata and C krusei). For invasive aspergillosis, 2 new approaches have the potential to improve on the hitherto abysmal mortality figures. Voriconazole has potent activity against Aspergillus and appears to be fungicidal, at least in neutropenic rabbit models of invasive aspergillosis. Voriconazole is active against all Aspergillus species and has been shown to be superior to amphotericin B in clinical trials of invasive aspergillosis. Caspofungin treatment of Aspergillus results in decreased angioinvasion and decreased pulmonary infarcts in animal models of invasive aspergillosis. However, its use as a primary therapy or as combination therapy is an outstanding question that still remains to be answered. To correlate in vitro and in vivo data, we suggest the 90-60 rule : (bacterial or fungal) infections due to susceptible isolates respond to appropriate therapy about 90% of the time, and infections due to resistant isolates or those treated with inappropriate antimicrobials respond about 60% of the time. Our recommendations involve a stepwise approach to using in vitro susceptibility data for selection of fungal therapy. (Adv Stud Med. 2004;4(4A):S269-S275) The incidence of invasive fungal infections is increasing for several reasons: the increase in the number of patients at risk for developing these infections due to advances in chemotherapy regimens, increased use of high-dose corticosteroids, advances in solid-organ and bone marrow transplantation, and increased immunosuppression due to other infections. 1 Many of these factors prolong the survival of critically ill patients and increase the number of susceptible patients. A recent analysis of the National Center for Health Statistics database for multiple causes of death found that deaths due to mycoses increased from the tenth to the seventh most common cause between 1980 and 1997. 2 During this same time period, the number of deaths due to invasive mycoses (based on death certificate data) increased from 1557 to 6534, which translates to a 3.4-fold rate increase, or from 0.7 to 2.4 deaths per 100 000 in the general population. Given that only the information on the death certificate was available, these are almost surely underestimates from the failure to clinically diagnose mycotic infection in severely ill patients. 2 Advanced Studies in Medicine S269

The 2 major sources of fungal infections are Aspergillus and Candida. Deaths due to candidiasis have decreased dramatically from their peak during the study period, 2 but the majority of the deaths were in non- HIV infected patients. This downward trend is most likely, therefore, due to decreases in candidemia and invasive candidiasis, rather than the more characteristic Candida infections in HIV-associated disease (ie, oropharyngeal candidiasis and esophagitis). Of note, non-albicans Candida is emerging among high-risk patients, 2 the ramifications of which will be discussed in this article. Antimicrobial resistance is an important concern for mycoses as well as bacterial and viral infections, but the extent of the resistance problems among fungi remain largely undefined. Yet, it is critical to assess resistance patterns to determine optimal treatment strategies in these fragile patients. Resistance occurs through innate and acquired mechanisms in yeasts and molds. Table 1 lists the current and pending antifungal treatments and a brief description of their mechanisms of action. 1 49% in 2001, with no significant increase in crude mortality. 8,9 Of note, the patients with candidemia in the earlier study period were primarily neutropenic, whereas in the later period, almost all of the candidemia infections were in nonneutropenic patients (ie, mostly intensive care unit [ICU] patients). 8,9 The patient population becoming infected in 2001 was actually less sick overall than the population from 1986, so the impact of infection was even greater. The top 5 causative species for candidemia are Candida albicans, C glabrata, C parapsilosis, C tropicalis, and C krusei. Fluconazole is used extensively in the treatment of candidal infections. Resistance to fluconazole is now well documented in AIDS and patients with oropharyngeal candidiasis, but remains uncommon among bloodstream infections, with the exception of C glabrata and C krusei. C albicans, C tropicalis, and C parapsilosis remain very susceptible (97%, 98%, and 99% susceptible, respectively) to fluconazole, assuming a breakpoint of 8 µg/ml. 10 C glabrata is the second most common species in the United States, yet it is only 60% CANDIDA Candidemia is arguably the most important of the invasive mycoses. Occurrence varies based on the study population but ranges from 0.5 to 10 per 1000 hospital admissions and 6 to 10 episodes per 100 000 in the general population. Candida accounts for 8% to 10% of all nosocomial bloodstream infections. 3-7 As noted earlier, until 1997, the mortality due to candidemia had been decreasing. However, at the University of Iowa, case-controlled studies in 2 separate time periods (1983-1986 and 1997-2001) show that the excess mortality due to candidemia is actually increasing. In these studies, case patients were closely matched with control patients during the same time period according to age, sex, date of hospital admission, underlying disease(s), length of time at risk, and surgical procedure(s). The results show that the excess attributable mortality between the 2 groups increased from 38% in 1986 to Table 1. Selected Systemic Antifungal Agents Drug Formulation Mechanism of action Binds to ergosterol, the major sterol Amphotericin B deoxycholate IV in fungal cytoplasmic membranes; Amphotericin B binding creates channels, thus increasing lipid complex IV permeability and causing death Amphotericin B through leakage of essential nutrients colloidal dispersion IV Liposomal amphotericin B IV POLYENE Inhibits fungal cytochrome P45014DM Itraconazole Oral, IV (lanosterol 14-alpha-demethylase, which Voriconazole Oral, IV catalyzes a late step in ergosterol biosynthesis); Posaconazole* Oral precursors to ergosterol therefore accumulate, Ravuconazole* Oral resulting in abnormalities in membrane permeability, membrane-bound enzyme activity, and coordination of chitin synthesis TRIAZOLE Interferes with cell wall biosynthesis by Caspofungin IV noncompetitive inhibition of 1,3-beta-D- Anidulafungin* IV glucan synthase; 1,3-beta-D-glucan is an Micafungin* IV essential cell wall polysaccharide ECHINOCANDIN IV = intravenous. *Under investigation for US Food and Drug Administration approval. Adapted with permission from Steinbach WJ, Stevens DA. Review of newer antifungal and immunomodulatory strategies for invasive aspergillosis. Clin Infect Dis. 2003;37(suppl 3):S157-S187. 1 2003 The Infectious Disease Society of America. S270 Vol. 4 (4A) April 2004

susceptible, and 7% are highly resistant. C glabrata infections treated with fluconazole must have optimal dosing for efficacy. Of note, C krusei does not respond adequately (5% susceptible) to fluconazole and so it should be considered universally resistant to this drug. 10 There has been a notable increase in bloodstream infections with C glabrata in the last half of the 1990s. 11-15 This is due in part to suboptimal dosing of fluconazole (ie, <400 mg per day) and for poor indications (eg, in a surgical ICU, using fluconazole 100 mg daily for poorly defined indications). 16 Suboptimal dosing has important consequences beyond poor outcome. By first exposing the Candida species in the gut flora to fluconazole, strains with decreased susceptibility will naturally be selected (ie, C glabrata). Subinhibitory concentrations of fluconazole in these species will induce CDR efflux pumps, which render the fungus not only fluconazole resistant but also panazole resistant. For this reason, C glabrata should be the focus of surveillance efforts. Amphotericin B has historically been considered to be the gold standard for treating Candida infections, but its antifungal activity is species dependent. Using the Etest method of susceptibility testing, the differences among C albicans, C glabrata, and C krusei become apparent (Table 2). 10 Using 1 µg/ml as a point of comparison, resistance to amphotericin B by C glabrata is significantly increased. As supported by the Infectious Diseases Society of America recommendations for treating candidiasis, breakthrough fungemias occur more often with C glabrata and C krusei when standard doses of amphotericin B are used. Doses clearly need to be higher for effective treatment, but the optimal dose is not yet known, and the effect of different formulations of amphotericin B and dosing has also not been studied. 17 Voriconazole is one of the newer triazoles, although it has not yet been approved for use in treating Candida. In analyzing more than 6200 bloodstream isolates of Candida, and stratifying them by susceptibility to fluconazole, it appears that voriconazole as well as ravuconazole are potent antifungal agents against fluconazole-susceptible isolates; 10 their potency extends to more than 90% susceptibility at 1 µg/ml, which is a concentration that can easily be achieved or exceeded with current dosing regimens. Approximately half of the fluconazole-resistant Candida species will have low minimum inhibitory concentrations (MICs) to these new triazoles, which is essentially accounted for by C krusei. Notably, 98% to 100% of C krusei isolates were susceptible to ravuconazole or voriconazole (MIC 1 µg/ml), irrespective of their level of resistance to fluconazole. In fact, Table 2. Susceptibility to Amphotericin B Across Select Candida Species Cumulative % inhibited at MIC*: Species 0.25 0.5 1.0 2.0 4.0 8.0 (n tested) C albicans 11 61 95 >99 >99 >99 (3727) C glabrata 4 16 47 84 97 99 (796) C krusei 1 2 7 37 83 97 (106) *By Etest method. MIC = minimum inhibitory concentration. Adapted with permission from the American Society for Microbiology. Pfaller MA, Messer SA, Hollis RJ, Jones RN, Diekema DJ. In vitro activities of ravuconazole and voriconazole compared with those of four approved systemic antifungal agents against 6,970 clinical isolates of Candida spp. Antimicrob Agents Chemother. 2002;46(6):1723-1727. 10 Table 3. Caspofungin Has Excellent In Vitro Activity Against Candida % Susceptible at MIC (µg/ml) of: Candida Species 0.25 0.5 1 2 4 8 C albicans 97 99 99 99 99 99 C glabrata 91 99 99 99 99 99 C parapsilosis 6 25 60 87 95 99 C tropicalis 86 96 98 98 98 98 C dubliniensis 83 98 100 C krusei 10 42 99 100 C lusitaniae 23 81 96 100 N = 3959. Overall MIC 90 = 1 µg/ml. MIC = minimum inhibitory concentration. Adapted with permission from the American Society for Microbiology. Pfaller MA, Diekema DJ, Messer SA, Hollis RJ, Jones RN. In vitro activities of caspofungin compared with those of fluconazole and itraconazole against 3,959 clinical isolates of Candida spp, including 157 fluconazole-resistant isolates. Antimicrob Agents Chemother. 2003;47(3):1068-1071. 18 Advanced Studies in Medicine S271

of the 4 triazoles tested in this study (including fluconazole and itraconazole), only ravuconazole and voriconazole were reliably active against C krusei, indicating a wider spectrum of protection with these newer agents. The other half that remain nonsusceptible to the other triazoles are due to panazole resistance by fluconazole-resistant C glabrata. 10 Caspofungin has broad-spectrum activity against both Candida and Aspergillus infections and has recently received US Food and Drug Administration (FDA) approval for treating Candida bloodstream infection, esophageal candidiasis, and certain other Candida infections, as well as salvage therapy for aspergillosis. Analysis of nearly 4000 isolates of Candida species obtained from more than 95 different medical centers worldwide show that this drug is a potent antifungal agent, maintaining strong activity in fluconazole-resistant isolates. As shown in Table 3, almost all isolates exhibit 95% or greater susceptibility to caspofungin at an MIC of 1 µg/ml. 18 C parapsilosis has a somewhat lower susceptibility in vitro but responds to caspofungin in vivo. 19 Caspofungin also exhibits potency in fluconazole-resistant strains, with more than 90% susceptibility at 1 µg/ml (Table 4), so infections involving fluconazole-resistant C glabrata can be treated by this broad-spectrum agent. 18 The pharamacokinetics and pharmacodynamics of caspofungin for Candida infections is fairly well documented. Peak concentrations can be achieved in excess of 16 µg/ml with the recommended dose of 1 mg/kg per day. There is concentration-dependent killing with caspofungin with the optimal peak-to-mic ratio of about 4:1. Given the MIC data discussed above, this can be achieved for virtually all of the Candida species. Also, there is a post-antifungal effect of more than 12 hours with caspofungin. 20,21 For invasive Candida infections, the echinocandins and broad-spectrum triazoles are more potent than fluconazole and amphotericin B against significant clinical isolates of Candida (ie, C glabrata and C krusei). In particular, the in vivo data support the in vitro data regarding the efficacy of the echinocandins in the treatment of invasive candidiasis, including among the increasing number of fluconazole-resistant isolates. ASPERGILLUS Aspergillosis is also a devastating infection. Case fatality rates associated with invasive aspergillosis Table 4.Antifungal Susceptibility of Candida Species to Caspofungin by Fluconazole Susceptibility Cumulative % Inhibited at MIC (µg/ml) of: Fluconazole Tested Susceptibility (n) 0.12 0.25 0.5 1 2 4 8 Susceptible 3479 64 82 88 93 96 97 98 Susceptibility 323 57 82 90 95 95 96 96 Dose-Dependent Resistant 157 47 68 82 99 99 99 99 MIC = minimum inhibitory concentration. Adapted with permission from the American Society for Microbiology. Pfaller MA, Diekema DJ, Messer SA, Hollis RJ, Jones RN. In vitro activities of caspofungin compared with those of fluconazole and itraconazole against 3,959 clinical isolates of Candida spp, including 157 fluconazole-resistant isolates. Antimicrob Agents Chemother. 2003;47(3):1068-1071. 18 Table 5. Species of Aspergillus Isolated from Hospitalized Patients with Signs and Symptoms of Pneumonia: SENTRY Objective F, 2000-2001 Species n %* A fumigatus 256 67.3 A flavus 39 7.8 A niger 29 7.6 A versicolor 21 5.5 A terreus 16 4.2 Other species 27 7.6 *% of 382 total isolates Data from Diekema et al. 23 Table 6. In Vitro Susceptibilities of Aspergillus Species to Established and Investigational Agents Agent MIC 90 1 µg/ml (%) Caspofungin (MEC) 0.06 98 Posaconazole 0.5 98 Voriconazole 1.0 96 Ravuconazole 1.0 96 Amphotericin B 1.0 91 Itraconazole 2.0 83 N = 382: A fumigatus (n = 257); A flavus (n = 30); A niger (n = 29); A versicolor (n = 21); A terreus (n = 16), other Aspergillus (n = 29). MEC = minimum effective concentration. Data from Diekema et al. 23 S272 Vol. 4 (4A) April 2004

hover at 60%, increasing up to 87% in bone marrow transplant recipients. 22 Even with the use of amphotericin B deoxycholate and its lipid formulations, preventing mortality was achieved in only one third to one half of patients. 22 Approximately 5 species of Aspergillus are found in hospitalized patients with signs and symptoms of pneumonia, the most common of which is A fumigatus (Table 5). 23 However, as with Candida, some of the less common species cause the most concern because of their susceptibility patterns to commonly used antifungal agents. Of particular concern is A terreus. Again using 1 µg/ml as a point of comparison (because this dose can be exceeded by the dosing practices for all of the drugs tested), all of the newer agents are active against Aspergillus spp (Table 6). 23 These data include A terreus, and as shown in Table 7, susceptibility to A terreus is 100% for all agents except amphotericin B, for which almost two thirds of A terreus isolates have MICs of 2 µg/ml or higher. 23 Echinocandins inhibit growth by inhibiting glucan synthesis at growing hyphal tips and branch points. They do not kill the entire mold, so the rest of the hyphal mass may remain viable and appear as trailing growth. To assess the in vitro efficacy of caspofungin against Aspergillus, the microscopic morphology of the organism needs to be observed. As such, efficacy is measured by the minimum effective concentration (MEC), which is the first concentration at which a prominent visual change in the growth pattern and, microscopically, a transition from healthy organisms to sick aberrant forms is observed. Data for the in vitro activity of echinocandins against molds are often referred to as MEC, rather than MIC, indicating this unique fungistatic activity. Caspofungin treatment of Aspergillus results in decreased angioinvasion and decreased pulmonary infarcts in animal models of invasive aspergillosis. However, its use as a primary therapy or as combination therapy is a question that remains to be answered. Voriconazole has potent activity against Aspergillus and appears to be fungicidal, at least in neutropenic rabbit models of invasive aspergillosis. 24 Voriconazole is active against all Aspergillus species and has been shown to be superior to amphotericin B in clinical trials of invasive aspergillosis. 25 These 2 new approaches to invasive aspergillosis have potential to improve the hitherto abysmal mortality figures. ANTIMICROBIAL SUSCEPTIBILITY TESTING: CORRELATING IN VITRO AND IN VIVO DATA Infectious disease specialists have an imperfect means of determining the extent to which organisms will respond clinically based on laboratory susceptibility testing. We published a review of studies to determine the expected correlation between in vitro and in vivo data. According to the 90-60 rule, (bacterial or fungal) infections due to susceptible isolates respond to appropriate therapy about 90% of the time, and infections due to resistant isolates or those treated with inappropriate antimicrobials respond about 60% of Table 7. In Vitro Susceptibilities of Aspergillus terreus (n = 16) to Established and Investigational Agents MIC/MEC (µg/ml), % Agent 50 90 1 µg/ml Amphotericin B 2 2 37.5 Caspofungin 0.03 0.06 100 Itraconazole 0.5 0.5 100 Posaconazole 0.12 0.25 100 Ravuconazole 0.25 0.5 100 Voriconazole 0.25 1 100 MIC = minimum inhibitory concentration; MEC = minimum effective concentration. Data from Diekema et al. 23 Table 8.The 90-60 Rule : Correlation of Susceptibility Testing with Outcome for Fungal and Bacterial Infections Cases with Successful Outcome by Susceptibility Class,* % (n/total) Studies Patients (n) (n) Susceptible Resistant P value Fungi 13 1197 91 (828/923) 48 (121/274) <.001 Bacteria 12 5447 89 (4521/5081) 59 (215/366) <.001 *Antifungal testing performed according to NCCLS M27-A2. Includes mucosal, fungemia, meningitis, and disseminated infections due to Candida, C neoformans, and Histoplasma capsulatum treated with fluconazole, itraconazole, or ketoconazole. Data from Rex et al. 26 Advanced Studies in Medicine S273

the time. 26 A summary of this review, shown in Table 8, illustrates this rule. While it is not strictly followed, it does show that drug susceptibility test results can be used to guide therapy choices. RECOMMENDATIONS Our recommendations (see Sidebar, below) use a stepwise approach to using in vitro data for selection of fungal therapy. 26 First, it is important to identify the isolate to species. For Candida species from sterile sites, we recommend that routine testing should be done for fluconazole in particular (especially for C glabrata), and possibly for 5-fluorocytosine as well. The susceptibility of species other than C glabrata to fluconazole can be predicted based on previously published data. 10 For Candida species at mucosal sites, routine testing is not necessary, but susceptibility testing to fluconazole and itraconazole are useful for sorting out microbial resistance versus drug-delivery problems with a suboptimal response. We also recommend testing organism drug combinations as an adjunct in patients who fail initial therapy. Candida species, in particular, should be tested against amphotericin B, and Cryptococcus neoformans should be tested against fluconazole, 5-fluorocytosine, or amphotericin because some treatment failures have been associated with the development of drug resistance. There are limited data on Histoplasmosis capsulatum suggesting that fluconazole use may lead to resistance. 27,28 Organisms with a high rate of intrinsic resistance need not be tested; an alternative agent should automatically be selected based on the published literature. Organisms with significant rates of acquired resistance should be monitored closely for treatment failure, and susceptibility testing should be performed if possible. Species with high rates of resistance are listed in Table 9. CONCLUSION The spectrum of predominant yeasts and molds is changing, but several new broad-spectrum antifungal agents have become available, and several apparently are on the brink of FDA approval. Future concerns for invasive fungal infections are moving away from C albicans and A fumigatus, the strains historically most notorious for fatality. Drug resistance, including multidrug resistance, is apparent in some of the emerging Table 9. Species with High Rates of Resistance to Antifungal Agents Drugs (to which there is a high frequency Fungus of resistance) Class of Resistance Aspergillus terreus Amphotericin B Intrinsic Candida glabrata Azoles Intrinsic and acquired Amphotericin B Acquired C krusei Fluconazole Intrinsic 5-fluorocytosine Intrinsic Amphotericin B Acquired C lusitaniae Amphotericin B Intrinsic and acquired Histoplasma capsulatum Fluconazole Acquired Scedosporium apiospermum Amphotericin B Intrinsic S prolificans Amphotericin B Intrinsic Trichosporon beigelii Amphotericin B Intrinsic A Stepwise Approach to the Use of Fungal Identification and Antifungal Susceptibility Testing in the Selection of Antifungal Therapy 1. Identify isolate to species level 2. Candida spp from sterile sites test routinely fluconazole 5-fluorocytosine 3. Candida spp from mucosal sites Routine testing not necessary Test fluconazole and itraconazole to sort out microbial resistance vs drug-delivery problems 4. Test selected organism/drug combinations as an adjunct in patients failing initial therapy Candida spp and amphotericin B (Etest) C neoformans and fluconazole, 5-fluorocytosine, or amphotericin B (Etest) Histoplasma capsulatum and fluconazole 5. Isolates of species with high rates of intrinsic resistance need not be tested Alternative agent 6. Isolates of species with significant rates of acquired resistance Monitor closely for failure Perform susceptibility testing if possible Data from Rex et al. 26 S274 Vol. 4 (4A) April 2004

opportunistic molds, as it is with bacteria. Much can be learned from taking the time for susceptibility testing, as long as the clinician understands the limitations of the information provided by tests (ie, the 90-60 rule ). REFERENCES 1. Steinbach WJ, Stevens DA. Review of newer antifungal and immunomodulatory strategies for invasive aspergillosis. Clin Infect Dis. 2003;37(suppl 3):S157-S187. 2. McNeil MM, Nash SL, Hajjeh RA, et al. Trends in mortality due to invasive mycotic diseases in the United States, 1980-1997. Clin Infect Dis. 2001;33(5):641-647. 3. Eggimann P, Garbino J, Pittet D. Epidemiology of Candida species infections in critically ill non-immunosuppressed patients. Lancet Infect Dis. 2003;3(11):685-702. 4. Pfaller MA, Diekema DJ. Role of sentinel surveillance of candidemia: trends in species distribution and antifungal susceptibility. J Clin Microbiol. 2002;40(10):3551-3557. 5. Diekema DJ, Messer SA, Brueggemann AB, et al. 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