Meningitis secondary to Cryptococcus gattii,an emerging pathogen affecting immunocompetent hosts

BACKGROUND:

Meningitis continues to be one of the most important infections diagnosed and treated by emergency physicians. Despite the advent of anti-infective therapy, meningitis carries a mortality rate of 20%–40%. In this study, we describe the first reported emergency medicine case of meningitis associated with Cryptococcus gattii to alert providers of this insidious, emerging global pathogen infecting immunocompetent individuals.

METHODS:

We provided a case report and accompanying review of the literature. A MEDLINE search for the term Cryptoccocus gattii was performed to obtain background information on Cryptococcus gattii.

RESULTS:

After two months of hospitalization, the patient was eventually discharged neurologically intact except for a continued mild bilateral hearing deficit.

CONCLUSION:

Cryptococcus gattii is an emerging world pathogen, which affects otherwise healthy, immunocompetent patients and requires timely identification and treatment in order to prevent severe neurological sequelae.KEY WORDS: Meningitis, Cryptococcus gattii     

Cryptococcus neoformans-Cryptococcus gattii Species Complex: an International Study of Wild-Type Susceptibility Endpoint Distributions and Epidemiological Cutoff Values for Fluconazole,Itraconazole, Posaconazole,and Voriconazole

Epidemiological cutoff values (ECVs) for the Cryptococcus neoformans-Cryptococcus gattii species complex versus fluconazole, itraconazole, posaconazole, and voriconazole are not available. We established ECVs for these species and agents based on wild-type (WT) MIC distributions. A total of 2,985 to 5,733 CLSI MICs for C. neoformans (including isolates of molecular type VNI [MICs for 759 to 1,137 isolates] and VNII, VNIII, and VNIV [MICs for 24 to 57 isolates]) and 705 to 975 MICs for C. gattii (including 42 to 260 for VGI, VGII, VGIII, and VGIV isolates) were gathered in 15 to 24 laboratories (Europe, United States, Argentina, Australia, Brazil, Canada, Cuba, India, Mexico, and South Africa) and were aggregated for analysis. Additionally, 220 to 359 MICs measured using CLSI yeast nitrogen base (YNB) medium instead of CLSI RPMI medium for C. neoformans were evaluated. CLSI RPMI medium ECVs for distributions originating from at least three laboratories, which included ≥95% of the modeled WT population, were as follows: fluconazole, 8 μg/ml (VNI, C. gattii nontyped, VGI, VGIIa, and VGIII), 16 μg/ml (C. neoformans nontyped, VNIII, and VGIV), and 32 μg/ml (VGII); itraconazole, 0.25 μg/ml (VNI), 0.5 μg/ml (C. neoformans and C. gattii nontyped and VGI to VGIII), and 1 μg/ml (VGIV); posaconazole, 0.25 μg/ml (C. neoformans nontyped and VNI) and 0.5 μg/ml (C. gattii nontyped and VGI); and voriconazole, 0.12 μg/ml (VNIV), 0.25 μg/ml (C. neoformans and C. gattii nontyped, VNI, VNIII, VGII, and VGIIa,), and 0.5 μg/ml (VGI). The number of laboratories contributing data for other molecular types was too low to ascertain that the differences were due to factors other than assay variation. In the absence of clinical breakpoints, our ECVs may aid in the detection of isolates with acquired resistance mechanisms and should be listed in the revised CLSI M27-A3 and CLSI M27-S3 documents.     

Heteroresistance to Itraconazole Alters the Morphology and Increases the Virulence of Cryptococcus gattii

Cryptococcus gattii is the main etiological agent of cryptococcosis in immunocompetent individuals. The triazole drug itraconazole is one of the antifungals used to treat patients with cryptococcosis. Heteroresistance is an adaptive mechanism to counteract the stress of increasing drug concentrations, and it can enhance the ability of a microorganism to survive under antifungal pressure. In this study, we evaluated the ability of 11 C. gattii strains to develop itraconazole heteroresistance. Heteroresistant clones were analyzed for drug susceptibility, alterations in cell diameter, capsule properties, and virulence in a murine model. Heteroresistance to itraconazole was intrinsic in all of the strains analyzed, reduced both the capsule size and the cell diameter, induced molecular heterogeneity at the chromosomal level, changed the negatively charged cells, reduced ergosterol content, and improved the antioxidant system. A positive correlation between surface/volume ratio of original cells and the level of heteroresistance to itraconazole (LHI) was observed in addition to a negative correlation between capsule size of heteroresistant clones and LHI. Moreover, heteroresistance to itraconazole increased the engulfment of C. gattii by macrophages and augmented fungal proliferation inside these cells, which probably accounted for the reduced survival of the mice infected with the heteroresistant clones and the higher fungal burden in lungs and brain. Our results indicate that heteroresistance to itraconazole is intrinsic and increases the virulence of C. gattii. This phenomenon may represent an additional mechanism that contributes to relapses of cryptococcosis in patients during itraconazole therapy.     

Multicenter Study of Isavuconazole MIC Distributions and Epidemiological Cutoff Values for the Cryptococcus neoformans-Cryptococcus gattii Species Complex Using the CLSI M27-A3 Broth Microdilution Method

Epidemiological cutoff values (ECVs) of isavuconazole are not available for Cryptococcus spp. The isavuconazole ECVs based on wild-type (WT) MIC distributions for 438 Cryptococcus neoformans nongenotyped isolates, 870 isolates of genotype VNI, and 406 Cryptococcus gattii isolates from six laboratories and different geographical areas were 0.06, 0.12, and 0.25 μg/ml, respectively. These ECVs may aid in detecting non-WT isolates with reduced susceptibilities to isavuconazole.     

The Investigational Fungal Cyp51 Inhibitor VT-1129 Demonstrates Potent In Vitro Activity against Cryptococcus neoformans and Cryptococcus gattii

The in vitro activities of the novel fungal Cyp51 inhibitor VT-1129 were evaluated against a large panel of Cryptococcus neoformans and Cryptococcus gattii isolates. VT-1129 demonstrated potent activities against both Cryptococcus species as demonstrated by low MIC₅₀ and MIC₉₀ values. For C. gattii, the in vitro potency was maintained against all genotypes. In addition, significantly lower geometric mean MICs were observed for VT-1129 than for fluconazole against C. neoformans, including isolates with reduced fluconazole susceptibility.     

Azole Resistance in Cryptococcus gattii from the Pacific Northwest: Investigation of the Role of ERG11

Cryptococcus gattii is responsible for an expanding epidemic of serious infections in Western Canada and the Northwestern United States (Pacific Northwest). Some patients with these infections respond poorly to azole antifungals, and high azole MICs have been reported in Pacific Northwest C. gattii. In this study, multiple azoles (but not amphotericin B) had higher MICs for 25 Pacific Northwest C. gattii than for 34 non-Pacific Northwest C. gattii or 20 Cryptococcus neoformans strains. We therefore examined the roles in azole resistance of overexpression of or mutations in the gene (ERG11) encoding the azole target enzyme. ERG11/ACT1 mRNA ratios were higher in C. gattii than in C. neoformans, but these ratios did not differ in Pacific Northwest and non-Pacific Northwest C. gattii strains, nor did they correlate with fluconazole MICs within any group. Three Pacific Northwest C. gattii strains with low azole MICs and 2 with high azole MICs had deduced Erg11p sequences that differed at one or more positions from that of the fully sequenced Pacific Northwest C. gattii strain R265. However, the azole MICs for conditional Saccharomyces cerevisiae erg11 mutants expressing the 5 variant ERG11s were within 2-fold of the azole MICs for S. cerevisiae expressing the ERG11 gene from C. gattii R265, non-Pacific Northwest C. gattii strain WM276, or C. neoformans strains H99 or JEC21. We conclude that neither ERG11 overexpression nor variations in ERG11 coding sequences was responsible for the high azole MICs observed for the Pacific Northwest C. gattii strains we studied.     

The Production of Reactive Oxygen Species Is a Universal Action Mechanism of Amphotericin B against Pathogenic Yeasts and Contributes to the Fungicidal Effect of This Drug

Amphotericin B (AMB) is an antifungal drug that binds to ergosterol and forms pores at the cell membrane, causing the loss of ions. In addition, AMB induces the accumulation of reactive oxygen species (ROS), and although these molecules have multiple deleterious effects on fungal cells, their specific role in the action mechanism of AMB remains unknown. In this work, we studied the role of ROS in the action mechanism of AMB. We determined the intracellular induction of ROS in 44 isolates of different pathogenic yeast species (Candida albicans, Candida parapsilosis, Candida glabrata, Candida tropicalis, Candida krusei, Cryptococcus neoformans, and Cryptococcus gattii). We also characterized the production of ROS in AMB-resistant isolates. We found that AMB induces the formation of ROS in all the species tested. The inhibition of the mitochondrial respiratory chain by rotenone blocked the induction of ROS by AMB and provided protection from the killing action of the antifungal. Moreover, this phenomenon was absent in strains that displayed resistance to AMB. These strains showed an alteration in the respiration rate and mitochondrial membrane potential and also had higher catalase activity than that of the AMB-susceptible strains. Consistently, AMB failed to induce protein carbonylation in the resistant strains. Our data demonstrate that the production of ROS by AMB is a universal and important action mechanism that is correlated with the fungicidal effect and might explain the low rate of resistance to the molecule. Finally, these data provide an opportunity to design new strategies to improve the efficacy of this antifungal.     

Genetic Diversity of the Cryptococcus Species Complex Suggests that Cryptococcus gattii Deserves to Have Varieties

The Cryptococcus species complex contains two sibling taxa, Cryptococcus neoformans and Cryptococcus gattii. Both species are basidiomycetous yeasts and major pathogens of humans and other mammals. Genotyping methods have identified major haploid molecular types of C. neoformans (VNI, VNII, VNB and VNIV) and of C. gattii (VGI, VGII, VGIII and VGIV). To investigate the phylogenetic relationships among these haploid genotypes, we selected 73 strains from 2000 globally collected isolates investigated in our previous typing studies, representing each of these genotypes and carried out multigene sequence analyses using four genetically unlinked nuclear loci, ACT1, IDE, PLB1 and URA5. The separate or combined sequence analyses of all four loci revealed seven clades with significant support for each molecular type. However, three strains of each species revealed some incongruence between the original molecular type and the sequence-based type obtained here. The topology of the individual gene trees was identical for each clade of C. neoformans but incongruent for the clades of C. gattii indicating recent recombination events within C. gattii. There was strong evidence of recombination in the global VGII population. Both parsimony and likelihood analyses supported three major clades of C. neoformans (VNI/VNB, VNII and VNIV) and four major clades of C. gattii (VGI, VGII, VGIII and VGIV). The sequence variation between VGI, VGIII and VGIV was similar to that between VNI/VNB and VNII. MATa was for the first time identified for VGIV. The VNIV and VGII clades are basal to the C. neoformans or the C. gattii clade, respectively. Divergence times among the seven haploid monophyletic lineages in the Cryptococcus species complex were estimated by applying the hypothesis of the molecular clock. The genetic variation found among all of these haploid monophyletic lineages indicates that they warrant varietal status.     

Heteroresistance of Cryptococcus gattii to Fluconazole

We analyzed 71 clinical and environmental Cryptococcus gattii strains that had been isolated before or after the advent of azole antifungals to determine their level of heteroresistance to fluconazole (LHF). All strains of C. gattii manifested heteroresistance, with LHFs that ranged between 4 μg/ml and 32 μg/ml. A considerably higher proportion of the C. gattii strains (86%) than Cryptococcus neoformans strains (46%) exhibited LHFs that were ≥16 μg/ml. No significant correlation was observed between the molecular type or serotypes of strains and their respective LHF. The strains which expressed a higher LHF were also more resistant to xenobiotics than the strains with a low LHF, and the level of resistance to xenobiotics was significantly higher than that reported for C. neoformans. The heteroresistant subpopulation, whose level of drug resistance had been raised in a stepwise manner to 64 μg/ml, reverted to the original LHF upon daily transfers in drug-free medium. Importantly, the strains with high LHFs were significantly more virulent than those with low LHFs. Since all the clinical isolates that had not been exposed to azole drugs as well as the environmental strains manifested heteroresistance to fluconazole, heteroresistance of C. gattii to azoles is an intrinsic mechanism as in C. neoformans and is associated with the strain''s virulence.Cryptococcosis is caused by two species, Cryptococcus neoformans and Cryptococcus gattii (15); fluconazole (FLC), a triazole, is widely used for the treatment of cryptococcosis regardless of the causative species (29). Fluconazole-resistant strains of C. neoformans have been increasingly reported from cases of therapy failure in AIDS patients undergoing long-term maintenance therapy (1-3, 27, 36). In 1999, we reported heteroresistance to fluconazole, an adaptive mode of drug resistance, in strains of C. neoformans recovered from two patients. One patient had AIDS and suffered from a series of recurrent cryptococcosis episodes during azole maintenance therapy, while the other was a non-AIDS patient who had never been treated with azole drugs (26). Our recent expanded characterization of heteroresistance using over 100 strains of C. neoformans that had been isolated at least a decade prior to the advent of azole antifungals revealed that C. neoformans is intrinsically heteroresistant to fluconazole. The innate level of heteroresistance to fluconazole (LHF) ranged from 4 μg/ml to 64 μg/ml, and 50% of serotype A strains and 21% of serotype D strains manifested high LHFs (≥16 μg/ml). Importantly, the LHF was observed to correlate with both the virulence of C. neoformans strains in mice and resistance to xenobiotics unrelated to fluconazole (31).Strains of C. gattii infect immunocompetent patients more often than C. neoformans strains (25), and animal studies have revealed that some strains of C. gattii are more virulent than strain H99, a well-characterized highly virulent C. neoformans strain of serotype A (10). Although the clinical manifestations of the disease caused by C. neoformans and C. gattii are not significantly different, the two species can readily be differentiated by their biochemical, genetic, ecological, and epidemiological characteristics (14). In light of the fact that strains of C. gattii infect immunocompetent individuals more often than C. neoformans strains and the two species are biologically and genetically distinguishable, we investigated the LHFs of C. gattii strains to compare them with those of the C. neoformans strains.In this study, we have analyzed the LHFs of 71 clinical and environmental strains of C. gattii, which include serotypes B and C and are comprised of four major molecular types, VGII, VGIII, VGI, and VGIV. The strains were collected from different geographical areas, and all serotype B and the majority of serotype C clinical strains had been isolated at least 10 years prior to the availability of triazole antifungals for therapy of mycosis. As with C. neoformans, all strains of C. gattii manifested heteroresistance regardless of their chronology or source of isolation. Interestingly, a considerably higher percentage of the C. gattii strains (86%) expressed LHFs of ≥16 μg/ml than that for C. neoformans (46%) (27). The strains with high LHFs were more virulent in mice and more resistant to xenobiotics that are unrelated to fluconazole, as reported for C. neoformans. However, the degree of resistance to xenobiotics in the C. gattii strains with high LHFs was significantly higher than that for the strains of C. neoformans with high LHFs.     

In Vitro Antifungal Susceptibilities and Amplified Fragment Length Polymorphism Genotyping of a Worldwide Collection of 350 Clinical,Veterinary, and Environmental Cryptococcus gattii Isolates

The in vitro susceptibilities of a worldwide collection of 350 Cryptococcus gattii isolates to seven antifungal drugs, including the new triazole isavuconazole, were tested. With amplified fragment length polymorphism (AFLP) fingerprinting, human, veterinary, and environmental C. gattii isolates were subdivided into seven AFLP genotypes, including the interspecies hybrids AFLP8 and AFLP9. The majority of clinical isolates (n = 215) comprised genotypes AFLP4 (n = 76) and AFLP6 (n = 103). The clinical AFLP6 isolates had significantly higher geometric mean MICs for flucytosine and fluconazole than the clinical AFLP4 isolates. Of the seven antifungal compounds examined in this study, isavuconazole had the lowest MIC₉₀ (0.125 μg/ml) for all C. gattii isolates, followed by a 1 log₂ dilution step increase (MIC₉₀, 0.25 μg/ml) for itraconazole, voriconazole, and posaconazole. Amphotericin B had an acceptable MIC₉₀ of 0.5 μg/ml, but fluconazole and flucytosine had relatively high MIC₉₀s of 8 μg/ml.The basidiomycetous yeast Cryptococcus gattii is responsible for life-threatening invasive disease in apparently healthy humans and animals (7, 19). A typical C. gattii infection is acquired through the respiratory tract, from which it can further disseminate to the central nervous system, resulting in fatal meningitis (7, 19, 32). Cryptococcosis caused by the primary pathogenic yeast C. gattii was, until a decade ago, a rarely encountered infection outside tropical and subtropical regions (17, 26, 27). However, this changed due to an unprecedented outbreak that emerged in the temperate climate of Vancouver Island (British Columbia, Canada) that subsequently expanded farther into the Pacific Northwest (1, 8, 10, 16). Its sibling species, Cryptococcus neoformans, differs ecologically and epidemiologically from C. gattii since it occurs on a global scale and is linked with disease occurring in immunocompromised individuals, such as HIV-positive patients and transplant patients who receive immune-suppressive medicines (7, 10, 18, 19, 31).Cryptococcus gattii can be discerned from C. neoformans using a wide range of microbiological and molecular techniques (7, 20). A convenient method is the use of canavanine-glycine-bromothymol blue (CGB) medium, which allows C. gattii but not C. neoformans to grow and which changes the pH indicator in the medium from green-yellowish to blue (18). With the increasing use of molecular techniques, such as PCR fingerprinting, restriction fragment length polymorphism (RFLP) analysis of the PLB1 and URA5 loci, and amplified fragment length polymorphism (AFLP) fingerprint analysis, as well as several multilocus sequence typing (MLST) approaches, it became clear that C. gattii could be divided into five distinct genotypes, named AFLP4/VGI, AFLP5/VGIII, AFLP6/VGII, AFLP7/VGIV, and AFLP10 (the last one of which is a recently observed novel genotype) (2, 6, 7, 13, 16, 20, 21, 23). Until recently, a serotype agglutination assay was widely used to distinguish C. neoformans (serotypes A and D) from C. gattii (serotypes B and C) (7, 27). In general, serotype B strains are found in each of the five C. gattii AFLP genotypes, but it seems that C. gattii serotype C strains are restricted to genotypes AFLP5/VGIII and AFLP7/VGIV (2, 6, 16, 21, 27).In addition, it was found that C. gattii and C. neoformans can form interspecies hybrids, named genotype AFLP8 (C. neoformans var. neoformans AFLP2/VNIII serotype D × C. gattii AFLP4/VGI serotype B) and AFLP9 (C. neoformans var. grubii AFLP1/VNI serotype A × C. gattii AFLP4/VGI serotype B). These interspecies hybrids have, until now, been isolated only from clinical samples, and they might have a higher virulence potential than regular C. gattii or C. neoformans isolates (4, 5; F. Hagen, K. Tintelnot, and T. Boekhout, unpublished data).Treatment of cryptococcosis depends on, besides the immune status of the patient, the severity and localization of the infection (11). Severe cases of cryptococcosis in immunocompetent and -compromised patients are treated according to the guidelines of the Infectious Diseases Society of America, according to which treatment consists of an induction therapy for 2 weeks with a combination of amphotericin B and flucytosine, followed by a 10-week consolidation therapy with fluconazole (11, 24).Cryptococcus neoformans has been extensively studied for its in vitro susceptibility to a wide variety of antifungal compounds, including the new triazoles posaconazole, voriconazole, ravuconazole, and isavuconazole (12, 14, 28, 29, 33). Despite the ongoing C. gattii outbreak, only a few studies using relatively small sets of C. gattii isolates have been performed to investigate their in vitro susceptibilities to amphotericin B, flucytosine, fluconazole, and the new triazole antifungals (12, 15, 28-30). A few studies divided the C. gattii isolates into groups according to their serotype or genotype (15, 29).Therefore, we studied the in vitro susceptibilities of each of the C. gattii genotypes from a large worldwide collection, subdivided by AFLP genotyping, to amphotericin B, flucytosine, fluconazole, itraconazole, voriconazole, posaconazole, and the new experimental broad-spectrum antifungal triazole isavuconazole.     

Fluconazole Susceptibility in Cryptococcus gattii Is Dependent on the ABC Transporter Pdr11

Cryptococcus gattii isolates from the Pacific Northwest have exhibited higher fluconazole MICs than isolates from other sites. The mechanism of fluconazole resistance in C. gattii is unknown. We sought to determine the role of the efflux pumps Mdr1 and Pdr11 in fluconazole susceptibility. Using biolistic transformation of the parent isolate, we created a strain lacking Mdr1 (mdr1Δ) and another strain lacking Pdr11 (pdr11Δ). Phenotypic virulence factors were assessed by standard methods (capsule size, melanin production, growth at 30 and 37°C). Survival was assessed in an intranasal murine model of cryptococcosis. Antifungal MICs were determined by the M27-A3 methodology. No differences in key virulence phenotypic components were identified. Fluconazole susceptibility was unchanged in the Mdr1 knockout or reconstituted isolates. However, fluconazole MICs decreased from 32 μg/ml for the wild-type isolate to <0.03 μg/ml for the pdr11Δ strain and reverted to 32 μg/ml for the reconstituted strain. In murine models, no difference in virulence was observed between wild-type, knockout, or reconstituted isolates. We conclude that Pdr11 plays an essential role in fluconazole susceptibility in C. gattii. Genomic and expression differences between resistant and susceptible C. gattii clinical isolates should be assessed further in order to identify other potential mechanisms of resistance.     

Antifungal Therapy in a Murine Model of Disseminated Infection by Cryptococcus gattii

We have evaluated the efficacy of posaconazole (PSC), voriconazole (VRC), and amphotericin B (AMB) in a murine model of systemic infection by Cryptococcus gattii using immunocompromised animals and three clinical strains of the fungus. AMB was the most effective drug in prolonging the survival of mice and also in reducing tissue burden in all organs tested. To a lesser degree, VRC at 60 mg/kg of body weight in lung tissue and PSC at 40 mg/kg also in spleen demonstrated good efficacy in reducing the fungal load. The PSC and VRC levels in serum and brain tissue, determined by an agar diffusion bioassay method at 4 h after the last dose of the therapy, were above the corresponding MIC values. However, these drugs were not able to reduce the fungal load in brain tissue. Our results demonstrated that PSC and, to a lesser degree, VRC, have fungistatic activity and potential for the treatment of human pulmonary cryptococcosis.Cryptococcosis is an emerging infection commonly involving the lungs, from which it can disseminate to different tissues, usually the central nervous system (CNS) (20, 23). Cryptococcus neoformans and Cryptococcus gattii are the main agents responsible for this disease, which can affect both immunosuppressed and healthy individuals. Despite antifungal therapies, this infection still has mortality rates near 20% (20).The first choice in the primary therapy of CNS infections remains fungicidal drugs, with amphotericin B (AMB) alone or in combination with flucytosine being the most widely used (20, 23). Fungistatic drugs like itraconazole and fluconazole, with less toxicity, are also used in the maintenance of the therapy and in pulmonary cryptococcosis, but their use in CNS infections has been less than satisfactory. In addition, the extended duration of the therapy with these azoles increases the risk of developing drug resistance (11, 23, 26). It has been suggested that that C. gattii has a higher pathogenicity than C. neoformans (27), which emphasizes the importance of the correct species identification and makes it necessary to improve and search for alternatives to the current therapy.On the basis of the promising results obtained with posaconazole (PSC) and voriconazole (VRC) against C. neoformans in animal models (1, 19, 24) and also in a clinical setting (9, 15, 21, 22), we have evaluated in this study the efficacy of PSC, VRC, and AMB in a murine model of disseminated infection by C. gattii.     

Antifungal susceptibilities among different serotypes of Cryptococcus gattii and Cryptococcus neoformans

We measured antifungal activity against 128 cryptococcal isolates (86 of C. neoformans and 42 of C. gattii) to determine if differences in serotype susceptibility exist. Contrary to previous results, we found no serotype susceptibility differences. Isavuconazole, posaconazole, and voriconazole demonstrated excellent potency against each isolate and serotype, including isolates with reduced fluconazole susceptibilities.     

Cryptococcus endocarditis: A case report and review of the literature

IntroductionCryptococcus neoformans is known to be a cause of meningitis. However, as cryptococcal endocarditis is rare, it is not well understood. Here, we describe a case with Implantable Cardioverter Defibrillator associated endocarditis and meningitis caused by Cryptococcus neoformans and we review the literature associated cryptococcal endocarditis.Case presentationA 72 years old Japanese male presented in emergency department with non-productive cough and respiratory discomfort. His past medical history was ischemic heart disease four years ago and ICD was implanted. Physical examination was unremarkable. Chest computer tomography revealed ground glass opacity in the right lung. He received a diagnosis of amiodarone-induced interstitial pneumonitis and high dose steroid pulse therapy. Septic shock and acute respiratory failure occurred after steroid therapy. Cryptococcus neoformans was identified by blood culture and cerebral spinal fluid. Intravenous liposomal Amphotericin B and oral flucytosine were initiated. Transesophageal echocardiography revealed vegetation on the lead of the ICD. Diagnosis of cryptococcal endocarditis was made. The patient died despite antifungal therapy was continued.DiscussionWe analyzed our case and 8 cases of cryptococcal endocarditis in the literature for 40 years. Almost all of the patients had previous valve replacement surgery or immunocompromised state. Three cases had meningitis. Surgery performed in 3 cases. The overall mortality rate were 44.4%.ConclusionsCryptococcal endocarditis is rare and carries a high mortality. Almost all of the patients had underlying diseases. Diagnosis needs repeating blood culture and echocardiogram, sometimes. Cryptococcal endocarditis needs lumber puncture for rule out meningitis.     

Cryptococcus gattii as an important fungal pathogen of western North America

Cryptococcus gattii, a pathogenic fungus historically appreciated to be endemic to tropical regions, was recognized to emerge in a more temperate zone of North America in the 1990s. Early reports focused on an outbreak that was first apparent on Vancouver Island (BC, Canada), involving both the veterinary and human population. More recently, it has been recognized that this organism is endemic to a wider geography in western North America, with recognized disease caused by unique molecular subtypes in both healthy and immunosuppressed human hosts and a variety of domestic and wild animals. A number of cases of disease caused by C. gattii isolates that are unrelated to the Vancouver Island–Pacific Northwest outbreak strains have also been recognized in different parts of the USA. As microbiology laboratories have historically not identified these organisms to the species level, our current understanding of the scope of this infection is probably an underestimate. Ongoing public health epidemiologic efforts will be facilitated by increased attention towards culture-confirmed diagnosis and species identification in clinical microbiology laboratories. Early experience presents a strong rationale for increasing diagnostic attention, with multiple clinical features that are unique to this infection, including variability in antifungal susceptibilities and a heightened need for aggressive management of inflammatory responses. Larger prospective studies to evaluate and optimize clinical management are needed.     

Combination therapy of voriconazole and terbinafine for disseminated fusariosis: case report and literature review

Fusarium spp. cause a broad spectrum of infection and are relatively resistant to most antifungal agents, leading to unfavorable prognosis, especially in immunocompromised patients. Several reports have shown synergism among amphotericin B, voriconazole (VRC), terbinafine (TRB), and other antifungal agents in vitro, but the most efficacious combination remains to be elucidated. We report the first case of disseminated Fusarium solani infection successfully treated by combination therapy of VRC and TRB accompanied by surgical resection of endocardial lesions. We also review 15 case reports of combination antifungal therapy for fusariosis and 6 case reports of Fusarium endocarditis.     

First Reported Case of Cryptococcus gattii in the Southeastern USA: Implications for Travel-Associated Acquisition of an Emerging Pathogen

In 2007, the first confirmed case of Cryptococcus gattii was reported in the state of North Carolina, USA. An otherwise healthy HIV negative male patient presented with a large upper thigh cryptococcoma in February, which was surgically removed and the patient was started on long-term high-dose fluconazole treatment. In May of 2007, the patient presented to the Duke University hospital emergency room with seizures. Magnetic resonance imaging revealed two large CNS lesions found to be cryptococcomas based on brain biopsy. Prior chest CT imaging had revealed small lung nodules indicating that C. gattii spores or desiccated yeast were likely inhaled into the lungs and dissemination occurred to both the leg and CNS. The patient's travel history included a visit throughout the San Francisco, CA region in September through October of 2006, consistent with acquisition during this time period. Cultures from both the leg and brain biopsies were subjected to analysis. Based on phenotypic and molecular methods, both isolates were C. gattii, VGI molecular type, and distinct from the Vancouver Island outbreak isolates. Based on multilocus sequence typing of coding and noncoding regions and virulence in a heterologous host model, the leg and brain isolates are identical, but the two differed in mating fertility. Two clinical isolates, one from a transplant recipient in San Francisco and the other from Australia, were identical to the North Carolina clinical isolate at all markers tested. Closely related isolates that differ at only one or a few noncoding markers are present in the Australian environment. Taken together, these findings support a model in which C. gattii VGI was transferred from Australia to California, possibly though an association with its common host plant E. camaldulensis, and the patient was exposed in San Francisco and returned to present with disease in North Carolina.     

Efficacy of NS-718, a Novel Lipid Nanosphere-Encapsulated Amphotericin B, against Cryptococcus neoformans

In vitro and in vivo efficacies of NS-718, a lipid nanosphere-encapsulated amphotericin B (AMPH-B), have been studied. Of the tested AMPH-B formulations, NS-718 had the lowest MIC for Cryptococcus neoformans. In a murine model, low-dose therapy (0.8 mg/kg of body weight) with NS-718 showed higher efficacy than that with AmBisome. High-dose therapy (2.0 mg/kg) with NS-718 was much more effective than those with Fungizone and AmBisome. In mice treated with a high dose of NS-718, only a few yeast cells had grown in lung by 7 days after inoculation. A pharmacokinetic study showed higher concentrations of AMPH-B in lung following administration of NS-718 than after administration of AmBisome. Our results indicated that NS-718, a new AMPH-B formulation, is a promising antifungal agent for treatment of pulmonary cryptococcosis and could be the most effective antifungal agent against C. neoformans infections.     

Cryptococcal endocarditis of native valves without immunodeficiency or drug abuse: a case report

Cryptococcal endocarditis has rarely been reported. Most patients with this condition are associated with risk factors, such as structural heart disease/valve replacement, immunodeficiency/immunosuppression or drug abuse. We report a case of cryptococcal endocarditis of the native valves without any risk factors. A 50-year-old Chinese man was admitted to hospital with fever for 1 month without any underlying heart disease, immunodeficiency, or drug use. He was diagnosed as having Cryptococcus neoformans infective endocarditis and was discharged after valve replacement surgery and long-term antifungal therapy.     

Combination antifungals: an update

Invasive fungal infections are an important cause of morbidity and mortality in specific patient populations. There has been an impressive increase in the antifungal armamentarium, yet optimal therapies for many invasive fungal infections remain unknown. Genomic sequencing of a number of pathogenic fungi will pave the way to discovering additional newer targets for antifungal drug design. These new discoveries, plus the existing repertoire of antifungal agents, create the need to effectively model single and combination antifungal agents. Future therapies may also include the use of cell-stress pathway inhibitors in combination with existing antifungal agents. This review focuses on combination antifungal therapy against Cryptococcus neoformans, Candida and Aspergillus species. Combination therapy is only supported by randomized clinical trials for cryptococcal meningitis. We review data from in vitro and animal model studies as well as insights from clinical trials to discuss current thoughts and highlight the gaps in our knowledge surrounding combination antifungal therapy.