Newer antifungal therapy for emerging fungal pathogens

Review

Newer antifungal William

J. Steinbach(1,2)

therapy for emerging fungal pathogens

and John R. Perfect(2,3)

As the number of immunocompromised patients increases, there is a concomitant increase in the number and diversity of fungal infections. Fungi that were once considered harmless or contaminants are now pathogenic in the immunocompromised host. Often these emerging pathogens are indistinguishable from the more familiar fungal infections; however, they are generally more recalcitrant to conventional antifungal therapies. With the antifungal armamentarium now expanding, the clinician now has many more options for these difficult-to-treat mycoses. We review many of the newer antifungal agents (second-generation triazoles, echinocandins, etc.) as therapeutic options for the recently emerging fungal pathogens. Int J Infect

Dis 2003;

7: 5-20

As advances are made in the treatment of life-threatening diseases with newer, aggressive chemotherapeutic regimens and the frequent use of bone marrow and organ transplantation, fungal pathogens are expanding in response to the increase in the number of immunocompromised patients.l While Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans remain the most common invasive fungal pathogens in immunocompromised hosts, changing environmental and host conditions, as well as selective antifungal pressure, have created newer patterns of emerging fungal diseases.2 Clinicians must be aware of newer, infrequent pathogens that were once viewed as innocuous contaminants of cultures or ubiquitous harmless environmental inhabitants.3 It is clear that the number of fungal opportunistic pathogens is increasing,4 and the variety of species implicated has broadened, so that now the concept of a non-pathogenic fungus has to be justified in the setting of an immunocompromised host.5 These emerging fungal infections also often mimic the more common fungal infection presentations, and many can be lethal in immunocompromised patients. In addition, these infections can be resistant to conventional antifungal therapy, and, frequently, the numbers of infections treated successfully are so low that it is difficult to create a standard algorithm. The armamentarium of newer antifungals available has also recently increased, and may offer new therapeutic options for both the historically common pathogens and the emerging diseases. There have been several excellent reviews on identifying the emerging fungal

(‘JDivision of Pediatric Infectious Diseases, Duke University; University Mycology Research Unit, Durham and (‘JDivision ious Diseases and International Health, Duke University, NC, USA. Address Infectious Durham,

correspondence to William .I. Steinbach, Division Diseases, Duke University Medical Center, NC 27710, USA. E-mail: [email protected]

Corresponding

Editor:

Jonathan

Cohen,

Brighton,

UK

@)Duke of InfectDurham, of Pediatric Box 3499,

pathogens,2T4Jj but there is no review focusing on the recent available data on the newer antifungals, such as amphotericin B lipid products, extended-spectrum triazoles, and echinocandins. Although analyzing all study results for every emerging fungal pathogen is beyond the scope of any single review, we will focus on a selection of these emerging pathogens and highlight the potential use of newer antifungal therapies.

NON-ALBICANSCANDIDASPECIES Background Candida albicans remains the most common Candida species. However, non-albicans Candida spp. have

emerged as common nosocomial pathogens over the last decade.7 For instance, over a recent 2-year study period, a significant decrease was observed in the number of Candida albicans bloodstream isolates, with a concomitant increase in the non-albicans Candida spp. iso1ated.s Data from the National Cancer Institute revealed that the incidence of non-albicans Candida spp. infections increased from 22% in 1992 to 52% in 1997.” Furthermore, epidemiologic data from over 23 233 yeast isolates in 33 countries showed that Candida albicans was the most common isolate (67.3%), but was followed by substantial numbers of non-albicans Candida spp. such as Candida glabrata (9.5%), Candida tropicalis (5.3%), Candida parapsilosis (4.9%), Candida krusei (2.4%), Candida guilliermondii (0.9%), and Candida lusitaniae (O.5%).8 These new Candida species can be more recalcitrant

to standard antifungal therapy, either innately or due to selective antimicrobial pressure. Several important antifungal susceptibility patterns are emerging.‘” A susceptibility survey showed fluconazole resistance with Candida krusei and Candida glabrata, and less so with Candida parapsilosis and Candida guilliermondii.8 At present, Candida krusei is considered to be intrinsically resistant to fluconazole, while Candida glabrata is highly variable. Candida lusitaniae” and Candida guillier-

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International

Journal

of Infectious

Diseases I Volume 7, Number 1.2003

mondii develop resistance when treated with amphotericin B, and Candida parapsilosis may show tolerance to amphotericin B. The increase in the non-albicans Candida spp. that have been historically more difficult to treat, coupled with their own increased resistance to conventional antifungals, suggests the need for newer antifungals. The latest Candida species is Candida dubliniensis, first named in July 1995 by Sullivan et alI2 after they had isolated it from HIV patients with recurrent candidiasis at Dublin Dental Hospital. Candida dubliniensis isolates are phenotypically very similar to Candida albicans, producing germ tubes and chlamydospores, but otherwise yield unique assimilation profiles. Candida dubliniensis is recovered from the oral cavities of approximately 30% of HIV-infected patients with clinical oral candidiasis.i3 Its emergence is postulated to be due to the pressure of antifungal prophylaxis for Candida albicans in this patient population.6 One study reported that Candida dubliniensis also adheres to Vero cells significantly better in the presence of fluconazole,14 and a neutropenic murine model showed that Candida dubliniensis was more virulent than Candida albicans.15 However, most studies of invasive candidiasis or

Table

1.

In vitro

Species Candida

Candida

Candida

Candida

Candida

Candida

tropicalis

parapsilosis

glabrata

krusei

lusitaniae

guilliermondii

susceptibilities

of non-albicans

Candida

AMB

FCZ

93 75 16 86 129 130

0.125

8 4to

93 75 16 86 129 130

0.5 0.5-I

1 0.5-2

0.5 0.13-0.5

1.0

2

.

0.25

0.2a

;.32-0.64a

0.02a

93 75 16 86 129 130

ICZ

>64

In vitro analyses The majority of comparative studies with newer antifungals against Cundida spp. are in vitro susceptibility analyses. Although in vitro analyses may not predict in vivo outcome, and testing for some of the agents is not standardized, the results do give some general comparative information. In vitro susceptibilities reported for newer antifungals against non-albicans Candida spp. are summarized in Table 1. The MICs were generally lower for the newer triazoles than for the echinocandins, but fungicidal activity was generally greater with the echinocandins.i6 Animal or clinical experience There are few murine model studies with the less virulent non-albicans Candidu spp. that have tested newer antifungals; caspofungin has shown poor steriliza-

species

Ref.

0.5-l

candidemia have not identified Candida dubliniensis as a cause. Therefore, it remains unclear whether this species has any different response to antifungal therapy for invasive disease.

vcz

PCZ

0.5 0.5 to z-8

0.2

32 2 40

0.5

8 0.25-I

2

64 8 40

2

1 0.5-I

64 >64

0.8=

16 32 40a

75 16 86 129 130

0.5

1

0.1=

2 2 0.32-0.64

93 16 86 130

0.25

4

0.4"

2a s.o=

All values given as MICW (pg/mL) at 72 h of incubation, AMB, amphotericin B; FCZ, fluconazole; ICZ, itraconazole;

2

2 0.2a

>8.0

1.0

0.25

0.5

8

0.4

1 0.25-I

0.125 0.062Sto

1.0

>4

2

1.0

0.5 1

0.8a

0.4a

0.0313-0.06256

0.06-0.13 0.25

2

2

2 0.8a

>8.0

0.06

0.02a

2 0.299

1.0=

2

0.5

0.06 2

2

0.0156 <0.0078-0.0156

1.0 1.0

0.5 0.4

0.25

1.6

93 75 16 86 129 130

0.5

1

4

0.4

1.0

0.25-2 0.5

64 2-8

AND

0.25

0.8

1 0.25-0.5

MIC 0.0313 0.0313-0.0625

0.25 1.0

CAS

0.25" 0.2a

2"

2*

0.06a 1.6=

unless MIC range given, or specified as geometric mean VCZ, voriconazole; PCZ, posaconazole; CAS, caspofungin;

MICa MIC, micafungin;

AND,

anidulafungin.

Newer antifungal therapy for emerging fungal pathogens I Steinbach and Perfect tion of the kidney with Candida parapsilosis, but good activity against Candida tropicalis, Candida glabrata, Candida lusitaniae, and Candida krusei.17g18Comparative clinical studies with amphotericin B and fluconazole for the treatment of candidemia have suggested that huconazole treatment outcome is similar to that with amphotericin B. 19,20However, there have always been many fewer cases with non-albicans Candida spp. groups than with Candida albicans. Additionally, because of reduced in vitro susceptibility of non-albicans Candida spp., many clinicians will use amphotericin B as initial therapy for candidemia until the Candida species is identified. A recent large double-blind comparative study of amphotericin B versus caspofungin for treatment of candidemia and invasive candidiasis has been reported.21 In this study, caspofungin compared favorably with amphotericin B in the treatment of nonalbicans Candida infections, in addition to causing substantially less drug toxicity. Candiduria with these yeasts remains common, and it is difficult to assess proper management. Candida tropicalis candiduria is difficult to eradicate with fluconazole,22 and the new triazoles and echinocandins do not have ideal pharmacokinetics for use in the treatment of urinary tract infections. Thus it is unlikely that new agents will help in the management of candiduria, and further prospective strategies for this condition will be necessary. Recommendations Fluconazole remains the treatment of choice for candidemia with susceptible isolates, since its activity appears to be equivalent to that of amphotericin B, with less toxicity. Several non-albicans Candida spp. are considered to be increasingly resistant to fluconazole (Candida krusei) or amphotericin B (Candida lusitaniae), while others are relatively resistant.23 Guidelines now include treatment with fluconazole for Candida lusitaniae, due to increasing resistance to amphotericin B.24*25 The broad-spectrum anti-Candida activities, fungicidal properties, reduced drug interactions and safety may make caspofungin and other echinocandinlike drugs attractive therapeutic choices for these emerging yeast infections. The extended-spectrum triazoles such as voriconazole and posaconazole also have potent in vitro activity against many of the nonalbicuns Candida spp., and initial open trials suggest that they possess therapeutic activity for invasive candidiasis, but comparative trials have yet to report the precise success of these agents. FUSARIUM

SPECIES

Background Fusarium spp. have been important soil-borne plant pathogens for years, 26 but are also becoming regular

7

causes of keratitis and colonization of burn patient eschars. In immunocompromised patients they cause respiratory and disseminated infection and have become the second most common filamentous fungal pathogens after Aspergillus. The most common species of this ubiquitous mould include E solani (approximately 50% of Fusarium isolates), E moniliforme, E oxysporum, E dimerum, and E proliferatum.6 Disseminated invasive fusariosis was first reported in 1973.27 It shares many pathogenic characteristics with invasive aspergillosis, including risk factors such as neutropenia and steroid use as well as a tropism for angio-invasion leading to hemorrhagic infarction.2 Fusarium infections are acquired through respiratory inhalation and are seen in the lungs and sinuses, but are also increasingly being isolated from neutropenic patients with onychomycosis and concomitant toe or finger cellulitis as a source for disseminated disease.2,26,28 Furthermore, one study recovered Fusarium spp. from 57% of hospital water samples and used molecular methods to identify the hospital water system as a potential reservoir for exposure and disease.29 In the largest published review, 91% (39/43) of patients presented with characteristic nodular cutaneous lesions,28 in which biopsy often showed extensive necrosis around hyaline branching septated hyphae.2 Blood cultures were also positive in approximately one-half of cases. It is felt that the high rate of bloodstream recovery is due to the production of adventitious unicellular propagules, which, in conjunction with the vasotropism of Fusarium, might continuously release individual fungal cells into the bloodstream and seed the skin.30 Most Aspergillus species, except Aspergillus terreus, will not produce these propagules in tissue, and this represents an important difference in clinical presentation between aspergillosis and fusariosis. In vitro analyses The in vitro susceptibility data on newer antifungals for Fusarium infections are summarized in Table 2, and combination drug studies have also been reported. A checkerboard microdilution technique was used to investigate the activities of amphotericin B, amphotericin B +5fluorocytosine (5-FC) and amphotericin B +rifampin against R solani. One of five strains showed in vitro synergy with both amphotericin B+S-FC and amphotericin B+rifampin, with the remaining strains showing indifferent interactions. A checkerboard study with six Fusarium isolates and amphotericin B + caspofungin showed synergistic or additive interactions for most strains, with no antagonism.31 Finally, a combination study with 22 isolates of Fusarium spp. investigated the interaction between amphotericin B and the protein synthesis inhibitor and antibacterial azithromycin. The majority (85%) of isolates demonstrated amphotericin B resistance (MIC rl mg/mL), but when it was combined with azithromycin, MICs

8

International Journal of Infectious Diseases I Volume 7, Number

Table 2. In vitro

susceptibilities

of

Fusarium species

Species

Ref.

AMB

ICZ

vcz

Fusarium moniliforme

83 75 79 131

4 l-2 >16a I=

>8 2 to >8 4a

2

83 113 16 132 87 73 75 79 51 131 133 39

1.0 1.0

>8 >8

8

2a

>16a

8=

0.25 to >64 >8 8to>32 32a

0.5-2.0

Fusarium oxysporum

Fusarium solani

All values

given

AMB, amphotericin MIC, micafungin;

(Fg/mL)

PCZ

RCZ

CAS

M/C

AND

TRB

UR-9825

8a

8a

4a

8a

>I28 2a 0.5-2

4.16a

>16a

>16a

8” 75.78a

83 113 16 132 87 73 75 85 79 51 131 133 93 39 as MI&

1,2003

0.5-2 4to>16 I= 0.5-2 4a 5.65a

22.6’ 19.21”

4 4

s8 >8

>8

I=

>16a

8”

>64 >8 >I6 8 to >32 32a

1.040

>I28 l-4 4a 0.25-2 2”

>16a

>16=

>16a

>16a 59.46=

2gm 2 1 to >I6 Ia 0.5-2 4.39= 0.25a 17.52a

>16a >88 20a

at 72 h of incubation,

8; ICZ, itraconazole; AND, anidulafungin;

z-128 4 2-8 4a 0.25-2 4.16=

VCZ, voriconazole; TRB, terbinafine.

>64a

unless

MIC range

PCZ, posaconazole;

were reduced by a mean of 3.2-fold. More importantly, the MICs were reduced from levels of resistance to levels achievable in serum.32 Animal or clinical experience There are several studies evaluating drugs in controlled animal models of fusariosis. In a murine model, no differences were seen between controls, amphotericin B monotherapy, amphotericin B +5-FC, or amphotericin B+rifampin.33 This confirms a previous report of lack of amphotericin B activity in a murine mode1.34 In an immunocompetent murine model of fusariosis (E soluni), posaconazole showed a significant and dosedependent effect on survival at 14 days, and therapy also resulted in complete clearance of the organism from the liver.35 A murine model showed that voriconazole was efficacious, being comparable to amphotericin B for fungal burden reduction in the kidney, and superior to amphotericin B for fungal burden reduction in the liver and brain.36 In another murine fusariosis model, high doses (20 mg/kg/day) of liposomal amphotericin B increased survival and reduced tissue burden compared with amphotericin B deoxycholate (1.5 mg/kg/day) and control groups.37

given,

or specified

as geometric

RCZ, ravuconazole;

mean

MICa

CAS, caspofungin;

Neutropenia is a prime risk factor for disseminated infection, and without recovery of neutrophils, Fusarium infection is fata1.28,38 The largest clinical review, including 43 cases of invasive fusariosis at one center and 54 cases from the literature, reported responses to therapy of 30% and 48%, respectively. Cure occurred uniformly only in those patients showing resolution of myelosuppression. 28 It is important to recognize this concept, and thus attention to immune reconstitution is necessary. In some cases, this may require granulocyte transfusions, but in most cases it will be necessary to attempt a rapid reversal of neutropenia with colonystimulating growth factors. In the clinic, Fusarium infections can be relatively resistant to most standard antifungals3’ and a retrospective survey of 31 human cases revealed nine different antifungal treatments used, with none being superior to the others. 4o Since invasive fusariosis often resembles aspergillosis, many patients have been treated with amphotericin B, which shows reduced in vitro activity against Fusarium spp.28,34 High doses of lipid formulations of amphotericin B can sometimes achieve stabilization of disease,4*,42 and one review showed improvement in 9 of 11 patients with 5 mg/kg per day of amphotericin B lipid complex.41 There have also been a

Newer antifungal therapy for emerging fungal pathogens I Steinbach and Perfect few clinical cases in which voriconazole has been used successfully for Fusarium spp. infections. For instance, a 16-year-old girl with E solani keratitis after swimming in a lake showed no response with itraconazole therapy, and was changed to voriconazole at 4 mg/kg twice daily with clinical improvement, but relapsed after 10 days. The voriconazole dose was then increased to 6 mg/kg twice daily, the eye surgically debrided, and the anterior chamber irrigated with voriconazole, which finally led to clinical resolution.43 A patient with Fusarium peritonitis during continuous ambulatory peritoneal dialysis who failed catheter removal and amphotericin B treatment was cured with voriconazole treatment.44 In a multicenter study, voriconazole showed a positive response in approximately 50% of cases with fusariosis,45 and posaconazole showed a positive response in four of five patients with refractory fusariosis.46 Recommendations In summary, invasive fusariosis can be extremely difficult to diagnose and treat. In the immunocompromised patient, the clinical presentation often mimics invasive aspergillosis, but with some unique features. Standard doses of amphotericin B will have few consistent positive effects on infection. As with aspergillosis, attention to recovery of neutrophils is paramount, and control of the underlying disease is essential to a favorable outcome. Newer antifungal agents, such as high doses of lipid products of amphotericin B and the new extendedspectrum triazoles, may help control invasive infection in some patients until immune reconstitution has occurred, and they may represent a significant advance in the management of this difficult-to-manage mycosis. SCEDOSPORIUM

SPECIES

Background Scedosporium is a worldwide ubiquitous fungus recovered from soil, sewage, and animal manure. There are two medically important species: S. upiospermum is the asexual stage (anamorph) of Pseudallescheria boydii, and S. prolificans (formerly S. inflatum) is related to Pseudallescheria boydii.2 In normal hosts, these hyaline moulds produce localized disease after penetrating trauma or aspiration of polluted water with development of pneumonia or meningitis. However, in immunocompromised patients, they can parallel the clinical manifestations of aspergillosis or fusariosis and lead to pulmonary or disseminated infection.47@ Previously, S. apiospermum was associated with ‘whitegrain’ mycetoma, and S. pro1iJican.s with subcutaneous infections and a predilection for cartilage and joint areas, but in recent years they have been recognized as important pathogens in neutropenic patients.49 The spores infect an individual via inhalation, ulcerative lesions in the gastrointestinal tract, surgical wounds, or

9

inoculation from trauma. Spores reach target organs and produce hyphae that eventually sporulate in tissue and disseminate through the bloodstream to other organs, particularly the kidneys, lungs, and brain.50 Many disseminated infections are fatal despite antifungal therapy, but recovery from neutropenia may lead to survival. In vitro analyses The in vitro susceptibilities of Scedosporium infections to newer antifungals are summarized in Tables 3 and 4. However, there are several studies worth highlighting in which a new triazole UR-9825 (Uriach Laboratories, Barcelona, Spain) showed the best activity against S. prolijicans,6~48~49~51 offering a potential agent for a species that is very resistant to antifungal agents. Some clinicians advocate combination therapy for this infection, and the combination of itraconazole and terbinafine has been examined.52 One study, using the checkerboard technique, showed in vitro interactions between itraconazole and terbinafine when used against 20 clinical isolates of S. prolificarts. While each drug showed general inactivity, synergy was seen in 95% (19/20) of isolates after 48 h, using drug levels attainable in blood.53 Another study showed synergistic in vitro interactions between voriconazole and terbinafine when used against S. prolificans.54

An in vitro study examining checkerboard macrodilution to characterize combinations of amphotericin B and various azoles when used against 22 isolates of Pseudallescheria boydii showed additive or synergistic interactions and no antagonism for 67% of isolates. Amphotericin B +fluconazole displayed the greatest synergy (mean fractional inhibitory concentration index (FICI), 0.61), followed by amphotericin B+miconazole, and amphotericin B +itraconazole.55 Another study reported poor in vitro activity with amphotericin B alone, but synergistic activity with amphotericin B and pentamidine when used against clinical isolates oi S. prolificarts. 56 However, overshadowing all the in vitro combination reports is a review of 16 patients with scedosporiosis treated with amphotericin B + 5-FC, fluconazole or itraconazole in which all patients who did not recover from neutropenia died.50 Animal or clinical experience Few animal models have been studied for this infection, but one murine model of Pseudallescheria boydii infection showed that itraconazole was ineffective, while posaconazole was slightly more effective than fluconazole, in survival and fungal burden reduction.s7 Most early reported clinical cases of S. prolificarts infection represented colonization or contaminated wound infection, but the number of disseminated cases is increasing, and disseminated infection is usually fatal.58 A prospective sputum fungal culture screening study of

10

International Journal of Infectious Diseases I Volume 7, Number 1,2003

Table

3.

In vitro

susceptibilities

Species Scedosporium apiospermum

Scedosporium prolificans

All values

as Ml&

AMB

ICZ

83 113 132 48 134 87 73 49 85 51

8 8, 4 4a 16 16

2 0.5, 2a 4 16

>16 8 2a

0.12 >I6 4 32=

(Pg/mL)

55 75 16 86.

1.1= 2 2.6a 2.6a

All values AMB, AND;

given

amphotericin anibulafungin.

0.5a 0.5 2 to >64

>I6 >16* >16

>32 >16a >I6

4 >16= 16

16 8 2a 16a

>32 >64 >16 0.25 I= 232

M/C

AND

TRB

UR-9825

>32

2

2a

0.25

0.125

1

>I6 >8

4-8

>32*

2*

32

4

4

>16a 8.83a 4 to >64

4.0 4

16

2

1= 32a

at 72 h of incubation,

>32

unless

VCZ, voriconazole; TRB, terbinafine.

of Pseudallescheria

MIC range

given,

PCZ, posaconazole;

or specified RCZ,

as geometric

ravuconazole;

mean

16a

MICa

CAS, caspofungin;

boydii

0.45a so.01 5-0.25 0.76a 0.76a

-

vcz

PCZ

CAS

0.33a 0.333

1 .o=

1.3=

MIC

AND

128

at 72 h of incubation,

B; ICZ, itraconazole;

16

0.5

ICZ

as MICgO (FglmL)

CA.5

2

0.12-0.5 0.25 0.5 0.5= >8 >8

susceptibilities

AMB

RCZ

0.5

>8 >8

Table Ref.

PCZ

1.0

>8

B; ICZ, itraconazole; AND, anidulafungin;

In vitro

vcz

0.38a

AMB, amphotericin MIC, micafungin; 4.

species

Ref.

83 113 16 48 132 134 87 53 73 49 75 79 51 given

of Scedosporium

VCZ,

voriconazole;

unless

MIC

range

PCZ, posaconazole;

128 children with cystic fibrosis showed S. apiospermum in 8.6% of cultures,59 a markedly higher level than the < 1% prevalence in the general population.60 Scedosporium spp. were usually recovered from the lungs of cystic fibrosis patients already colonized with Aspergillus fumigatus, and in most cases colonization was not associated with disease, raising the issue of appropriate management. However, two patients presented with symptoms of allergic bronchopulmonary disease, which leads to comparisons with allergic aspergillosis in cystic fibrosis and the potential use of antifungals to reduce symptoms. Case reports of success with older antifungal agents against scedosporiosis are confounded by adjunctive surgical management and varying recovery of host defenses.48 An immunocompetent patient with S. apiospermkm pulmonary disease had an initial response with itraconazole and later showed relapse, but was cured after changing to terbinafine.61 A woman with asthma on chronic steroid therapy developed an S. apiospermum lung abscess, and new lesions developed

given,

or specified CAS, caspofungin;

2.5a

as geometric MIC,

mean

MIC.a

micafungin;

while she was on itraconazole therapy. Treatment was changed to voriconazole, which cured the infection after 63 days of therapy. 47 Another patient with relapsed acute myeloid leukemia developed disseminated disease with S. apiospermum despite empirical febrile neutropenia treatment with amphotericin B lipid complex (3 mg/kg/day). Therapy was switched to voriconazole after in vitro susceptibilities revealed activity, and despite persistent neutropenia, the patient’s skin lesion decreased in size and fever was eliminated; however, he later developed intestinal perforation and died.‘j2 An B-year-old boy with chronic granulomatous disease (CGD) and an S. apiospermum pulmonary abscess was initially treated with miconazole, then oral capsular itraconazole (30 mg/kg/day), and finally oral voriconazole plus subcutaneous interferon-gamma (IFN-y). Computed tomography showed marked improvement only after treatment with voriconazole. The same report documented a 13-year-old boy with CGD and an S. apiospermum lung abscess who was treated with miconazole for 15 days with no improve-

Newer antifungal therapy for emerging fungal pathogens I Steinbach and Perfect

11

ment; oral capsular itraconazole (20 mg/kg/day) and IFN-7 cured his pulmonary disease.63 Voriconazole has also been successfully used to treat a Pseudullescheria boydii central nervous system (CNS) infection after failure with amphotericin B lipid complex, miconazole, and itraconazole. 64 A relatively larg e series of scedosporiosis patients (36) treated with voriconazole has been reported. Over 60% of S. apiospermum infections had a positive clinical response, but fewer than 30% of S. prolificans infections had a positive clinical response.6s

activity of amphotericin B and fluconazole.68 TWO comparative in vitro studies showed terbinafine and itraconazole to have similar activity,72 and itraconazole and voriconazole to have identical activity.73 In another study, posaconazole showed better in vitro activity than itraconazole and amphotericin B against four isolates.74 Micafungin showed the best in vitro activity when compared to fluconazole, itraconazole, miconazole, and amphotericin B. 75 In one combination antifungal study, nikkomycin Z + itraconazole showed in vitro additivity against Penicillium spp.76

Recommendations

Animal or clinical experience

There is no proven effective therapy for disseminated Scedosporium disease. The therapeutic approach to Scedosporium infections involves complete surgical resection, with the role of antifungals probably being important, including a notable lack of response to amphotericin B.M There is a clear trend for better results with S. apiospermum than with S. prolificans. S. prolificans is more resistant to treatment compared to S. apiospermum, both in vitro, in a murine mode1,‘j7 and in clinical experience. It appears that few antifungal agents have consistent activity against S. prolificarts, but the extended-spectrum triazoles such as voriconazole or posaconazole may become agents of choice for S. apiospermum infections.

Amphotericin B has been used as successful initial management of disseminated penicilliosis in immunocompromised patients, but requires prolonged intravenous therapy, and relapse is common. Amphotericin B followed by itraconazole is also effective (97.3%)y1 but itraconazole takes an average of 57 days to clear fungemia.77 A review of 74 clinical isolates showed that itraconazole generally possessed better in vitro activity than amphotericin B, fluconazole, and ketoconazole, and it is generally recommended for treatment.68 However, voriconazole has been used to successfully treat 9 of 10 cases of disseminated penicillinosis.45

PENICILLIUM

MARNEFFEI

Background Penicillium marrteffei, the only dimorphic and human pathogenic Penicillium species, has emerged as the third most common AIDS-defining illness in parts of Southeast Asia.68 The incidence of this disease has increased markedly since the first reported natural infection in 1973.6y From approximately 30 cases in 1973-90, the incidence of this infection rose to over 160 reported cases by 1995. 7” In approximately 80% of cases4 disseminated disease can begin as numerous necrotic papulonodular lesions2 on the face, upper trunk, pinnae, and arms. 7o The disease can also present with non-productive cough, generalized lymphadenopathy, fever, anemia, and weight 10ss.~~,~iApproximately half of the cases involve fungemia.70 Penicillium marneffei can mimic and often be misdiagnosed as other infections, such as tuberculosis, Pneumocystis carinii pneumonia, cryptococcosis, and histoplasmosis. The conidia in the environment probably enter through the respiratory tract and convert to the yeast form, against which pulmonary alveolar macrophages appear to be the primary pulmonary host defense.

In vitro analyses An in vitro study of 30 isolates revealed susceptibility to itraconazole and ketoconazole, and intermediate

Recommendations The increasing incidence of Penicillium marneffei appears to be linked to the HIV pandemic. Unlike some other emerging fungal pathogens, this organism remains sensitive to many antifungals. For severe cases, initial induction treatment with amphotericin B may be necessary before switching to an oral azole, such as itraconazole. Although itraconazole is presently the azole of choice, newer triazoles are attractive and also offer oral formulations for chronic suppressive therapy. ACREMONIUM

SPECIES

Background species cause hyalohyphomycoses, which are mycoses caused by filamentous fungi which have colorless septate hyphae in tissue. They are common in soil, plant debris, and rotting mushrooms, and Acremonium kiliense is the most important among the 10 species reported as causing infections in vertebrates. The immunocompetent host can form a mycetoma or develop an ocular infection after sustaining a penetrating injury.78 Acremonium spp. can lead to a wide range of infections, from keratitis to disseminated disease, with the lung and gastrointestinal tract as apparent portals of entry.2,4 The first case of Acremonium spp. infection other than mycetoma or ocular infection was described in 1932, but most have arisen since 1985.78The largest review of 36 localized or disseminated cases showed that the majority of disseminated cases occurred in immunocompromised Acremonium

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patients, and only approximately half could be cured, with complete resolution of disease dependent on neutrophil recovery. 78 Recognition of disseminated Acremonium infection is difficult, because specific signs and symptoms do not exist.78 Patients may present with a cutaneous nodule, papular skin rash, or fungemia.

CNS phaeohyphomycosis, including patients who may have no apparent immunosuppression.82 CNS disease commonly presents with a headache and a focal neurologic deficit, frequently a hemiparesis.82 Bipolaris spp. represent the most common cause of phaeomycotic sinusitis, but may also cause pneumonia, fungemia, and disseminated infections.2

In vitro analyses

In one study, itraconazole had no activity and voriconazole had a marginally effective MIC for Acremonium kiZiense.73 Another study showed that voriconazole had the best in vitro activity compared to amphotericin B, fluconazole and itraconazole against Acremonium alabamensis and Acremonium strictum.79 Anidulafungin has no activity against Acremonium spp., but posaconazole and caspofungin both possess some fungistatic activity against Acremonium strictum.16 A recent study showed the activities of voriconazole and ravuconazole to be similar, and superior to that of posaconazole, while the strains tested were resistant to amphotericin B and itraconazole.80 Animal or clinical experience

The rarity of Acremonium infections makes it difficult to adequately assess the best treatment regimen. Amphotericin B was previously recommended as initial therapy,81 and despite the in vitro resistance to azole treatment, the combination of surgery and azoles has often been successful.78 Voriconazole appears to have the greatest in vitro activity against Acremonium, but clinical reports of its effectiveness are lacking. Recommendations

As with other fungal pathogens, the status of the host defense system is crucial. Acremonium infections are still rare but deadly, and the limited amount of clinical experience makes firm recommendations difficult. The echinocandins appear to have limited activity, while the newer triazoles may constitute beneficial adjunctive therapy in concert with boosting the host immune response. PHAEOHYPHOMYCOSIS: CLADOPHIALOPHORA AND BIPOLARIS SPP.

BANTIANA

Bacl&ound

Phaeohyphomycosis comprises a heterogeneous group of fungal diseases characterized by dematiaceous (darkly pigmented) hyphal forms in tissue involving the skin, subcutaneous and deep tissue, and CNS. Owing to space limitations, we will limit our discussion to several prominent species that represent this group. Cladophialophora bantiana is the cause of the majority of

In vitro analyses

In vitro studies with Cladophialophora bantiana show similar low MICs for voriconazole, itraconazole, amphotericin B,83 and terbinafine.84 One study showed the activity of posaconazole to be slightly worse than that of itraconazole, but much better than those of either anidulafungin or caspofungin.16 Another study showed itraconazole MICs to be better than those of voriconazole against seven isolates,73 while in another set of experiments voriconazole had superior in vitro activity to itraconazole and amphotericin B.85 The in vitro activity of drugs against Bipolaris species is variable. Amphotericin B possessed better activity than voriconazole and itraconazole against Bipolaris hawaiiensis and Bipolaris spicifera.83 However, in another study, voriconazole showed better in vitro activity than amphotericin B, fluconazole, and itraconazole.79 Voriconazole was also generally better than itraconazole against Bipolaris australiensis.73 In contrast, a study with six Bipolaris spp. isolates showed itraconazole to have the best activity, followed closely by voriconazole and amphotericin B,*(j and another study with 43 isolates showed that itraconazole and terbinafine both had high in vitro activity.84 In the evaluation of the echinocandins, itraconazole showed the best activity, with both voriconazole and posaconazole displaying good activity, and all were far better than caspofungin or anidulafungin.16 Comparing all available triazoles, posaconazole had the most activity, followed by itraconazole, ravuconazole, and voriconazole.80 An in vitro combination study with a nikkomycin Z and fluconazole combination showed additive effects.76 Animal or clinical experience

Including only culture-positive cases of CNS phaeohyphomycosis, the survival rate for this infection is only 35% .82CNS lesions may be best managed surgically, and the largest series of 30 cases revealed that those patients with single, encapsulated lesions did much better than those with multifocal disease. However, no patient who did not undergo surgery survived.Whether patients underwent neurosurgery alone or with antifungal therapy, the most important factor for cure was resectability of the lesion; antifungal therapy itself was not associated with improved survival.s2 This has been our experience with six to eight cases. Recently, we treated one patient with multiple inoperable Cladophialophora bantiana brain lesions and a severe

Newer antifungal therapy for emerging fungal pathogens I Steinbach and Perfect congenital immunodeficiency with voriconazole + WC, with some resolution of infection radiographically. HOWever, several months into treatment he did eventually succumb to his infection. Recommendations CNS phaeohyphomycosis continues to cause a high mortality without surgical resection, and this may be the case for ail localized phaeohyphomycoses. The new extended-spectrum triazoles, such as voriconazole and posaconazole, show good in vitro activity against Cladophialophora bantiana, Bipolaris spp., and several other dematiaceous fungi, with some positive correlating clinical experience. 4s These agents may become attractive as adjunctive therapy for these phaeohyphomycoses. Although the triazoles are probably more effective, the echinocandins do have some limited activity,87 and these agents will probably be tried in refractory cases as combination therapy. TRICHOSPORON

SPECIES

Background The incidence of fungemia caused by Trichosporon spp. is increasings8 in patients with hematologic malignancies and neutropenia, and this fungemia has been reported as the second most common yeast infection in cancer patients after Candida ~pp.~s One study showed that 10% of all fungemias at one institution were caused by Trichosporon spp. 88The taxonomy of Trichosporon has been revised several times recently. The most frequently reported species are Trichosporon asahii (formerly Trichosporon beigelii and Trichosporon cutaneum), Trichosporon pullulans, and Trichosporon capitatum (formerly Blastoschizomyces capitatus).88,89 Culture identification of Trichosporon spp. may be difficult, as it is often morphologically confused with Candida spp., which is worrisome, as Trichosporon has decreased susceptibility to amphotericin B, the drug often empirically used for suspected Candida infections. In vitro analyses, and animal and clinical experience There are limited in vitro studies and clinical reports for Trichosporon spp. One study of 39 Trichosporon clinical isolates showed high MICs for amphotericin B, with those of fluconazole and itraconazole being relatively lower. Ravuconazole, posaconazole and voriconazole appeared to be more active than amphotericin B or fluconazole, and similar to itraconazole, with voriconazole and posaconazole being slightly more active than ravuconazole. There were also potential fungicidal effects with the three newer triazoles.‘” In vitro studies with Trichosporon spp. showed that voriconazole was more active than itraconazole, amphotericin B, and fluconazole.79+86191

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Notably, Trichosporon pullulans has been found to be more resistant to antifungals, including total resistance to amphotericin B and fluconazole.” Further in vitro work showed posaconazole and amphotericin B to be similar, and superior to itraconazole and fluconazole.74 Lipid formulations of amphotericin B had higher MICs against Trichosporon spp. than amphotericin B, liposomal nystatin, and itraconazole.92 Posaconazole showed good activity against Trichosporon beigelii, and the echinocandins caspofungin and anidulafungin showed no activity.i6 This was echoed in further studies with micafungin75%93 and anidulafungin. 94There are some reports of clinical isolates that are resistant to both amphotericin B and azoles,95,96leaving fewer antifungal class options. Clinically, the genus often causes catheter-related fungemia, but there are reports of endocarditis,97 peritonitis9s meningitis,99 and pneumonitis.loO Trichosporon spp. have limited susceptibility to amphotericin B,99 and earlier clinical studies and animal models showed that azoles were effective,‘Ol and azoles plus polyenes were potentially additive.‘(n In one study, 5 of 12 reported cases occurred while the patient was on itraconazole prophylaxis. Additionally, 11 of the 12 patients died despite amphotericin B therapy.88 The mortality of hematogenous trichosporonisis was higher than that of candidiasis in that study, confirmed by a review of the published literature of the last 20 years, showing a Trichosporon spp. mortality of 64-80%s8 and central venous catheters to be the most frequent risk factor for fungemia. Invasive Trichosporon beigelii infection has also developed in a bone marrow transplant patient who was receiving caspofungin prophylaxis against Aspergillus infection, lo3 highlighting the in vitro inactivity of this new class of antifungals against Trichosporon.

Recommendations Trichosporon spp. cause very high mortality in immunocompromised patients, and, as with most emerging fungi, resolution of immunosuppression is essential. Unfortunately, the genus is generally less susceptible to amphotericin B, the drug usually used in suspected yeast infections. This, coupled with the difficulty in distinguishing the morphology of Candida and Trichosporon spp., may lead to delays in starting effective therapy. Azoles are generally more effective, and evidence exists to show that the newer triazoles might be the new treatment of choice. RHODOTORULA

RUBRA

Background Rhodotorula rubra is found naturally in soil, water, and plants, and is a constituent of the normal human respiratory, gastrointestinal and genitourinary flora.104

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Previously, this organism was considered to be only a harmless colonizer when isolated from healthy individuals, but it is now being reported in immunocompromised patients, causing fungemia, endocarditis, meningitis, and peritonitis.lo5 In vitro analyses, and animal and clinical experience In vitro testing of 35 clinical samples of Rhodotor& spp. showed low MICs of amphotericin B, ketoconazole, flucytosine, and itraconazole. The azoles were generally less effective than amphotericin B, and fluconazole showed an MIC of >32 mg/mL against all isolates.86J06J07 In a review of all 43 reported cases from 1960 to 2000, the most common risk for infection was central venous catheter insertion.104,108 It appears that the yeast is already an inhabitant of the skin and mucosa, and is introduced into the bloodstream following a disruption of normal anatomic barriers. In contrast to Trichosporon, mortality with Rhodotorula is uncommon, In a review, 10 of 36 survivors of infection were not treated with antifungals, and appeared to respond to removal of the catheter. In a series of 23 cases of fungemia, the outcome was favorable with the use of various treatment strategies, including line removal alone, its removal with amphotericin B treatment, or antifungal therapy alone.ro5 Only two patients in this group of patients with Rhodotorula fungemia were neutropenic. Recommendations Although the need for antifungal therapy is not always clear for Rhodotorulu, if catheters are removed we would still be conservative and recommend antifungal therapy in the immunocompromised patient. However, it is clear that there is a predilection for catheter involvement and that fluconazole is probably not the optimal antifungal for use in this infection. TRICHODERMA

LONGIBRACHIATUM

Background Trichoderma spp. are ubiquitous in soil, with five species of the genus Trichoderma having been identified as etiologic agents of infection in immunocompromised hosts: Trichoderma Longibrachiatum, Trichoderma Trichoderma koningii, Trichoderma harzianum, pseudokoningii, and Trichoderma viride.5 Trichoderma longibrachiatum causes virtually all human infections, just as Penicillium marneffei is virtually the only pathogenic Penicillium species, and has been reported to

cause pulmonary, cerebral, soft tissue, and disseminated disease in immunocompromised patients.2 It has been suggested that Trichoderma longibrachiatum can be acquired through the gastrointestinal tract, and prolonged therapy with fluconazole and antibacterial agents may selectively favor this filamentous fungus.5

In vitro analyses, and animal and clinical experience Itraconazole displays the best in vitro activity among the older antifungals in several studies.5J09-112 However, other studies showed that itraconazole had no activity, and amphotericin B had better activity than voriconazo1e.83J13 The first case of Trichoderma infection in an immunocompromised patient was reported in 1976, and a recent review lists only 10 cases reported in immunocompromised hosts.5 Resistance to amphotericin B used for treatment of this infection has been common in these reported clinical cases. Recommendations The incidence of Trichoderma infections appears to be slowly increasing in immunocompromised patients. Extended-spectrum triazoles should be considered in preference to the polyenes for treatment, but recovery from immunosuppressive events appears to be crucial to success. Stool surveillance cultures for early identification of this fungus remain unproven for clinical management, but may play a role in future studies. PAECILOMYCES

SPECIES

Background Puecilomyces spp. are common soil saprophytes and occasional airborne laboratory contaminants. They are often confused with the Penicillium spp., and are clinically difficult to identify because their presentation can mimic that of better-known fungi such as dematiaceous fungi and certain hyalohyphomycoses.lr4 They can cause keratitis, endophthalmitis and subcutaneous infections in the healthy host.4J14 In immunocompromised patients, they can manifest as cutaneous disease, catheter-related fungemia, sinusitis, and disseminated disease.2J14 Many immunocompromised patients become infected through integument breaks with intravascular catheters or chemotherapy.4 In fact, contaminated skin lotion has caused a nosocomial outbreak of infection in a bone marrow transplant unit.i15 Fungemia can be seen, as with Fusarium spp. and Acremonium spp. When Fusarium spp., Paecilomyces spp. and Acremonium spp. invade tissue, they produce hyphae as well as adventitious structures similar to microconidia that hematogenously disseminate and are discovered on blood culture.l16 Aspergillus does not do this, except for Aspergillus terreus, which can elaborate lateral conidia in vitro and in vivo.2

In vitro analyses Amphotericin B is very potent against Paecilomyces varioti, but has poor in vitro activity against the most common species, Paecilomyces lilacinus. Voriconazole

Newer antifungal therapy for emerging fungal pathogens I Steinbach and Perfect MICs are much lower than those of itraconazole for Paecilomyces lilacinus, including superior fungicidal activity,8” which was confirmed in another study, but the reverse was true for Paecilomyces varioti, with itraconazole activity being better than that of voriconazole.” In a study with posaconazole, it was found to have similar activity to that of itraconazole and better activity than that of amphotericin B against Paecilomyces varioti. Posaconazole displayed significantly better activity against Paecilomyces lilucinus compared to itraconazole, while amphotericin B showed no activity against that species.74 Comparing all triazoles, posaconazole had the most activity, followed by itraconazole, ravuconazole, and voriconazole.xO Caspofungin was inactive in vitro against Paecilomyces lilacinus, but showed excellent activity against Paecilomyces varioti.87 On the other hand, micafungin showed good in vitro activity against Paecilomyces lilacinus and Paecilomyces varioti, and better activity than fluconazole, itraconazole, miconazole, and amphotericin B.75 Nikkomycin Z + itraconazole combination therapy showed additivity in vitro against Paecilomyces sp~.‘~ Animal and clinical experience, and recommendations

The cornerstone of therapy has been surgical resection and removal of any foreign objects.l14 In the clinic, host factors and surgery play an important role in the cure of this infection. Recent in vitro data suggest that the newer triazoles might be effective in treating Paecilomyces infections, but large data sets of clinical experience are lacking to support the recommendation, and it remains even less clear whether the echinocandins have any place in the management of these infections. ZYGOMYCETES Background

15

to flucytosine.‘20,121 One in vitro study with 37 clinical isolates of zygomycetes showed posaconazole to be the most active azole.‘22 Animal

and clinical experience

Clinical manifestations of zygomycosis can be divided into the following categories: sinonasal, rhinocerebral, cerebral, pulmonary, cardiac, gastrointestinal, cutaneous, and disseminated.l*” Each category of disease requires a three-part management strategy: (1) early and aggressive surgical excision of the necrotic lesions; (2) restoration of immune function or control of underlying disease; and (3) intense antifungal therapy. Because tissue infarction is a prominent feature of infection with this angioinvasive fungus, removal of devitalized tissue is critical and can be curative for localized cutaneous zygomycoses associated with occlusive bandages, burns, or wounds, which rarely extend beyond local involvement.124 Surgical resection has been successful in cases of pulmonary disease.]*” Amphotericin B has been the mainstay of antifungal therapy to control residual infection after surgery for any early micrometastasis. Experience with lipid products of amphotericin B has been generally positive, with one study reporting successful outcome in 71% of patients with zygomycosis treated with amphotericin B lipid complex. 126The ability to give large doses of these formulations with reduced toxicity is attractive for treatment of these infections. The final management strategy is control of the underlying disease. In diabetic patients, this requires careful metabolic control during treatment, and cure is frequently obtainable. On the other hand, the mortality rate among patients with malignancy and zygomycosis continues to be very high, in contrast to the relatively good prognosis for patients with non-malignant underlying diseases. 127In these cancer patients, a favorable outcome correlates with non-pulmonary involvement, surgical debridement, and neutrophil recovery.l18

Zygomycosis (formerly mucormycosis or phycomycosis) is a rare infection caused by fungi in the orders Mucorales (class Zygomycetes) and Entomophthorales, with the most common causative organisms being Mucor, Rhizopus, Absidia, and Rhizomucor.117~118 They are ubiquitous saprophytes found in the air, soil, and food. In a large review of 361 cases of human zygomycosis, the genus and species were reported in 156 cases: 61% Rhizopus spp., 12% Mucor spp., and ~1% Rhizomucor spp. ’ I’)

Surgical removal of affected tissue is a hallmark of therapy. Amphotericin B or its lipid formulations are used for invasive infection, and human studies will be necessary to determine whether posaconazole has any role in the management of zygomycosis. Control of the underlying disease or immunosuppression is essential for successful management of this infection.

In vitro analyses

CONCLUSION

In vitro studies examining the activities of various antifungals against clinical isolates of zygomycetes have shown that the most active drug is amphotericin B. With the exception of itraconazole inhibiting isolates from the genus Absidia, the early-generation azole compounds were generally inactive, and all isolates were resistant

The landscape of medical mycology is changing. Emerging pathogens that were once irritating contaminants now lead to lethal disseminated disease in immunocompromised patients, and the once paltry list of antifungals available to combat this is increasing. However, coupled with the excitement of newer treat-

Recommendations

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ment choices is the realization that nearly all of the evaluation of newer antifungals in these emerging pathogens comprises in vitro analyses, with limited animal and human experience. This is a crucial first step in establishing new treatments, but several caveats must be expressed. First, the science of in vitro antifungal susceptibility testing and clinical correlation is in its infancy, only recently establishing standards for yeasts and continuing studies into filamentous fungi.128 Additionally, many in vitro studies test a limited number of isolates. Without further in vivo models or clinical reports, it is also difficult to ascertain the true pharmacokinetics, interactions and toxicities of these newer agents. Finally, the few anecdotal case reports published often include colony-stimulating factors or granulocyte transfusions as well as numerous other variables that could influence outcome, including improvement or deterioration in the underlying disease. We have attempted to compile available in vitro, in vivo and clinical data on the newer antifungal therapies available for some emerging fungal pathogens. Many of the fungal opportunists of today and tomorrow are innately or selectively resistant to conventional antifungals, so we desperately need to increase experience with these newer agents. As can be seen from this review, there are some baseline data for these emerging infections and newer drugs, but for the clinician it will take a creative individual approach to treat these infections. The issues will be to select a single antifungal agent or combination of antifungal agents, improve the patient’s immunity, and control the underlying disease. Success against these fungi will be determined by this three-pronged attack.

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Newer antifungal therapy for emerging fungal pathogens I Steinbach and Perfect the treatment of candidiasis. Clin Infect Dis 2000; 30:662678. 25. Pietrucha-Dilanchian P, Lewis RE, Ahmad H, et al. Candida lusitaniae catheter-related sepsis.Ann Pharmacother 2001; 351570-1574. 26. Nelson PE, Dignani MC, Anaissie EJ.Taxonomy, biology, and clinical aspects of Fusarium species.Clin Microbial Rev 1994; 7:479-504. 27. Cho CT, Vats TS, Lowman JT, et al. Fusarium solani infection during treatment for acute leukemia. J Pediatr 1973; 83:1028-1031. 28. Boutati EI, Anaissie EJ. Fusarium, a significant emerging pathogen in patients with hematologic malignancy: ten years’ experience at a cancer center and implications for management. Blood 1997; 90:999-1008. 29. Anaissie EJ, Kuchar RT, Rex JH, et al. Fusariosis associated with pathogenic species colonization of a hospital water system: a new paradigm for the epidemiology of opportunistic mold infections. Clin Infect Dis 2001; 33:1871-1878. 30. Schell WA. New aspects of emerging fungal pathogens: a multifaceted challenge. Clin Lab Med 1995; 15:365-387. 31, Arikan S, Lozano-Chiu M, Paetznick V. et al. In vitro synergy studies with caspofungin and amphotericin B against Aspergillus and Fusarium spp.Antimicrob Agents Chemother 2002; 461245-247. 32. Clancy C, Nguyen MH. The combination of amphotericin B and azithromycin as a potential new therapeutic approach to fusariosis. J Antimicrob Chemother 1998; 41:127-730. 33. Guarro J, Pujol I, Mayayo E. In vitro and in vivo experimental activities of antifungal agents against Fusarium solani. Antimicrob Agents Chemother 1999;43:12561257. 34. Anaissie EJ, Hachem R, Legrand C, et al. Lack of activity of amphotericin B in systemic murine fusarial infection. J Infect Dis 1992; 165:1155-1157. 35. Lozano-Chiu M,Arikan S,Paetznick VL, et alTreatment of murine fusariosis with SCH 56592. Antimicrob Agents Chemother 1999; 43:589-591. 36. Reyes GH, Long L, Hossain M, et al. Evaluation of voriconazole efficacy in the treatment of murine fusariosis [Abstract J-16041. In: 41st Interscience Conference on Antimicrobial Agents and Chemotherapy, Chicago, IL. Washington, DC: American Society for Microbiology, 2001. 37. Ortoneda M, Pastor FJ, Pujol C, et al. Successful treatment of murine fusariosis with high doses of liposomal amphotericin B [Abstract J-18301. In: 41st Interscience Conference on Antimicrobial Agents and Chemotherapy, Chicago, IL. Washington, DC: American Society for Microbiology, 2001. 38. Musa MO, Al Eisa A, Halim M, et al. The spectrum of Fusarium infection in immunocompromised patients with haematological malignancies and in non-immunocompromised patients: a single institution experience over 10 years. Br J Haematol2000; 108544-548. 39. Pujol I, Guarro J, Gene J, et al. In-vitro antifungal susceptibility of clinical and environmental Fusarium spp. strains. J Antimicrob Chemother 1997; 39: 163-167. 40. Hennequin C. Lavarde V, Poirot J, et al. Invasive Fusarium infections: a retrospective survey of 31 cases. The French Groupe d’Etudes des Mycoses Opportunistes’ GEMO. J Med Vet Mycol 1997; 35:107-114.

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