Paracoccidioides spp. and Histoplasma capsulatum: Current and New Perspectives for Diagnosis and Treatment.

The thermally-dimorphic systemic fungal group includes several important human pathogens: Blastomyces dermatitides, Coccidioides immitis and C. posadasii, Histoplasma capsulatum, Paracoccidioides brasiliensis, P. lutzii, and Talaromyces (Penicillium) marneffei. They usually are geographically restricted and have natural habitats in soil or in plants, and when fungal propagules invade mammalian host by inhalation, they initiate an inflammatory reaction that can result in self-resolution of the infection or cause an acute or chronic disease. In the setting of the AIDS pandemic and the developments in modern medicine, such as immunosuppressive therapy in cancer surgery patients and in transplantation and autoimmune diseases, the incidence of endemic mycoses has progressively increased. Another important factor of the increased incidence of systemic mycoses in certain regions is the progressive devastation of tropical and subtropical forests. In this review, we focus on two of the most important systemic mycoses: paracoccidioidomycosis and histoplasmosis, and their major characteristics in epidemiology, clinical aspects and laboratorial diagnosis.

Ciprofloxacin shows synergism with classical antifungals against Histoplasma capsulatum var. capsulatum and Coccidioides posadasii.

This study evaluated the in vitro interaction between ciprofloxacin (CIP) and classical antifungals against Histoplasma capsulatum var. capsulatum in mycelial (n = 16) and yeast-like forms (n = 9) and Coccidioides posadasii in mycelial form (n = 16). This research was conducted through broth microdilution and macrodilution, according to Clinical Laboratory Standards Institute. Inocula were prepared to obtain from 0.5 × 10(3) to 2.5 × 10(4) cfu ml(-1) for H. capsulatum and from 10(3) to 5 × 10(3) cfu ml(-1) for C. posadasii. Initially, minimum inhibitory concentration (MIC) for each drug alone was determined. Then, these MICs were used as the highest concentration for each drug during combination assays. The procedures were performed in duplicate. For all combination assays, MICs were defined as the lowest concentration capable of inhibiting 80% of visible fungal growth, when compared to the drug-free control. Drug interaction was evaluated by paired sample t-Student test. The obtained data showed a significant MIC reduction for most tested combinations of CIP with antifungals, except for that of CIP and voriconazole against yeast-like H. capsulatum. This study brings potential alternatives for the treatment of histoplasmosis and coccidioidomycosis, raising the possibility of using CIP as an adjuvant antifungal therapy, providing perspectives to delineate in vivo studies.

Evaluation of berries of Phytolacca dodecandra for growth inhibition of Histoplasma capsulatum var. farciminosum and treatment of cases of epizootic lymphangitis in Ethiopia.

OBJECTIVE: To evaluate the berries of Phytolacca dodecandra (P. dodecandra) for its effect on Histoplasma capsulatum var. farciminosum (HCF) and for the treatment of cases of epizootic lymphangitis (EL). METHODS: Samples were collected from un-ruptured nodules of cases of EL at Debre Zeit and Akaki (central Ethiopia). Mycological culture and isolation of HCF were performed at the Aklilu Lemma Institute of Pathobiology. Phytochemical screening was done for n-butanol extract of P. dodecandra to detect alkaloids, saponins, phenolic compounds and flavonoids. The minimum inhibitory concentrations (MICs) and minimum fungicidal concentrations (MFCs) of aqueous and n-butanol extracts of P. dodecandra against HCF were determined by agar dilution assay. For the in vivo trial, 5% simple ointment was prepared from n-butanol extract and applied topically to 24 (twelve early and twelve moderate) cases of EL. RESULTS: Phytochemical screening showed that n-butanol extract of P. dodecandra was positive for alkaloids, saponins and phenolic compounds but negative for flavonoids. The MICs of n-butanol and aqueous extracts of P. dodecandra were (0.039%-0.078%) and (0.625%-1.250%), respectively. The MFCs of n-butanol and aqueous extracts of P. dodecandra were (0.078%-0.156%) and (1.250%-2.500%), respectively. The MIC and MFC of ketoconazole (positive control) was (1.200×10(-5)%-2.500×10(-5)%) and (5.000×10(-5)%-1.000×10(-4)%), respectively while growth was observed on free medium (negative control). From the total of 24 treated cases of EL, 14 (58.3%) responded to treatment; however, 10 (41.7%) did not respond to treatment. There was no significant difference in the degree of response to treatment between early and moderate cases (χ(2)=0.686; P=0.408). CONCLUSIONS: It can be concluded that n-butanol extract of P. dodecandra demonstrates antifungal effects while the aqueous extract shows no antifungal activity.

Combined therapy with amphotericin B and caspofungin in an experimental model of disseminated histoplasmosis.

OBJECTIVE: To assess the effect of amphotericin B and caspofungin, as well as their combinations in the therapy of experimental disseminated histoplasmosis. MATERIAL AND METHODS: BALB/c mice were intraperitoneally infected with four different strains of Histoplasma capsulatum and given to antifungal treatments. The response to intraperitoneal therapy with amphotericin B (0.5, 1.0, and 2.0 mg/kg of body weight) or caspofungin (10 mg/kg of body weight) and their combinations, was evaluated by the quantification of yeast colony-forming units (CFU) per gram of spleen or lung, from each animal. Additionally, the pathogen was monitored histopathologically in the excised organs. Data were analyzed with the Kruskall-Wallis and Tukey tests. RESULTS: Caspofungin was more effective than amphotericin B in reducing the CFU/ g. A synergistic effect was observed when caspofungin (10 mg/ kg) was combined with amphotericin B (0.5 or 1.0 mg/kg). Significant differences in CFU values, H = 119.78 (P = 0.00001), were found among the treatment groups. However, statistical analyses did not reveal significant differences, H = 2.837 (P = 0.428), in the therapeutic responses with the four H. capsulatum strains tested. CONCLUSION: Combined therapy with amphotericin B and caspofungin could represent an alternative treatment to be explored in severe human histoplasmosis.

Emergence of resistance to fluconazole as a cause of failure during treatment of histoplasmosis in patients with acquired immunodeficiency disease syndrome.

In sequential clinical trials of treatment for histoplasmosis in patients with acquired immunodeficiency syndrome, therapy with fluconazole failed in a higher proportion of patients than did therapy with itraconazole. To determine the cause for failure with fluconazole, antifungal susceptibility testing that used modified National Committee on Clinical Laboratory Standards procedures was performed on all baseline and failure isolates. Failure occurred more frequently in patients with baseline isolates with fluconazole minimum inhibitory concentrations (MICs) > or =5 microg/mL versus lower MICs; 29% versus 3%, respectively. There was at least a 4-fold increase in fluconazole MIC in the isolates from 10 (59%) of 17 patients for whom paired pretreatment and failure or relapse isolates were available. Cross-resistance to itraconazole was not seen. In conclusion, fluconazole is less active than itraconazole for Histoplasma capsulatum and induces resistance during therapy, which accounted for treatment failure in some patients.

Chloroquine and the fungal phagosome.

The antimalarial drug chloroquine accumulates inside the macrophage phagolysosome by ion trapping where it exerts potent antifungal activity against Histoplasma capsulatum and Cryptococcus neoformans by distinct mechanisms. Chloroquine inhibits growth of H. capsulatum by pH-dependent iron deprivation, whereas it is directly toxic to C. neoformans. Clearly, clinical studies are required to document the potential therapeutic efficacy of chloroquine or related congeners as adjuvant therapy in fungal disease. Moreover, the diversity of pathogenic microorganisms inhibited and/or killed by chloroquine makes this drug an attractive candidate for prophylactic therapy.

Comparison of the echinocandin caspofungin with amphotericin B for treatment of histoplasmosis following pulmonary challenge in a murine model.

Twenty clinical isolates of Histoplasma capsulatum were tested for their in vitro susceptibilities to caspofungin in comparison to those to amphotericin B by following National Committee for Clinical Laboratory Standards guidelines for yeasts. The mean MICs were 16.6 microgram/ml (range, 8 to 32 microgram/ml) for caspofungin and 0.56 microgram/ml (range, 0.5 to 1.0 microgram/ml) for amphotericin B. Survival experiments used a 10(5) dose in a pulmonary challenge model with B6C3F(1) mice. All mice that received amphotericin B at 2 mg/kg of body weight every other day (q.o.d.), 30% of mice that received caspofungin at 8 mg/kg/day, and 20% of mice that received caspofungin at 4 mg/kg/day survived to day 15, while mice that received caspofungin at 2 mg/kg/day and all control mice that received the vehicle died by day 14. Amphotericin B at 2 mg/kg q.o.d. markedly reduced the fungal burden in the lungs and spleens, as measured by Histoplasma antigen detection techniques and quantitative cultures, for each comparison. Caspofungin at 10 mg/kg twice a day (b.i.d.) did not reduce the fungal burden, as measured by antigen detection techniques, but slightly reduced the levels of fungi in both the lungs and spleens, as determined by quantitative cultures. Caspofungin at 5 mg/kg b.i.d. did not affect fungal burden. Overall, caspofungin had only a slight effect on survival or fungal burden.

Practice guidelines for the management of patients with histoplasmosis. Infectious Diseases Society of America.

OBJECTIVE: The objective of this guideline is to provide recommendations for treating patients with the more common forms of histoplasmosis. PARTICIPANTS AND CONSENSUS PROCESS: A working group of 8 experts in this field was convened to develop this guideline. The working group developed and refined the guideline through a series of conference calls. OUTCOMES: The goal of treatment is to eradicate the infection when possible, although chronic suppression may be adequate for patients with AIDS and other serious immunosuppressive disorders. Other important outcomes are resolution of clinical abnormalities and prevention of relapse. EVIDENCE: The published literature on the management of histoplasmosis was reviewed. Controlled trials have been conducted that address the treatment of chronic pulmonary and disseminated histoplasmosis, but clinical experience and descriptive studies provide the basis for recommendations for other forms of histoplasmosis. VALUE: Value was assigned on the basis of the strength of the evidence supporting treatment recommendations, with the highest value assigned to controlled trials, according to conventions established for developing practice guidelines. BENEFITS AND COSTS: Certain forms of histoplasmosis cause life-threatening illnesses and result in considerable morbidity, whereas other manifestations cause no symptoms or minor self-limited illnesses. The nonprogressive forms of histoplasmosis, however, may reduce functional capacity, affecting work capacity and quality of life for several months. Treatment is clearly beneficial and cost-effective for patients with progressive forms of histoplasmosis, such as chronic pulmonary or disseminated infection. It remains unknown whether treatment improves the outcome for patients with the self-limited manifestations, since this patient population has not been studied. Other chronic progressive forms of histoplasmosis are not responsive to pharmacologic treatment. TREATMENT OPTIONS: Options for therapy for histoplasmosis include ketoconazole, itraconazole, fluconazole, amphotericin B (Fungizone; Bristol-Meyer Squibb, Princeton, NJ), liposomal amphotericin B (AmBisome; Fujisawa, Deerfield, IL), amphotericin B colloidal suspension (ABCD, or Amphotec; Seques, Menlo Park, CA), and amphotericin B lipid complex (ABLC, or Abelcet; Liposome, Princeton, NJ).

Comparison of a new triazole antifungal agent, Schering 56592, with itraconazole and amphotericin B for treatment of histoplasmosis in immunocompetent mice.

A murine model of intratracheally induced histoplasmosis was used to evaluate a new triazole antifungal agent, Schering (SCH) 56592, for treatment of histoplasmosis. MICs were determined for SCH 56592, amphotericin B, and itraconazole by testing yeast-phase isolates from 20 patients by a macrobroth dilution method. The MICs at which 90% of the isolates are inhibited were for 0.019 microgram/ml for SCH 56592, 0.5 microgram/ml for amphotericin B, and < or = 0.019 microgram/ml for itraconazole. Survival studies were done on groups of 10 B6C3F1 mice with a lethal inoculum of 10(5). All mice receiving 5, 1, or 0.25 mg of SCH 56592 per kg of body weight per day, 2.5 mg of amphotericin B per kg every other day (qod), or 75 mg of itraconazole per kg per day survived to day 29. Only 44% of mice receiving 5 mg of itraconazole/kg/day survived to day 29. Fungal burden studies done in similar groups of mice with a sublethal inoculum of 10(4) showed a reduction in CFUs and Histoplasma antigen levels in lung and spleen tissue in animals treated with 2 mg of amphotericin B/kg qod, 1 mg of SCH 56592/kg/day, and 75 mg of itraconazole/kg/day, but not in those treated with lower doses of the study drugs (0.2 mg of amphotericin B/kg qod, 0.1 mg of SCH 56592/kg/day, or 10 mg of itraconazole/kg/day). Serum drug concentrations were measured 3 and 24 h after the last dose in mice (groups of five to seven mice), each treated for 7 days with SCH 56592 (10 and 1 mg/kg/day) and itraconazole (75 and 10 mg/kg/day). Mean levels measured by bioassay were as follows: SCH 56592, 10 mg/kg/day (2.15 micrograms/ml at 3 h and 0.35 microgram/ml at 24 h); SCH 56592, 1 mg/kg/day (0.54 microgram/ml at 3 h and none detected at 24 h); itraconazole, 75 mg/kg/day (22.53 micrograms/ml at 3 h and none detected at 24 h); itraconazole, 10 mg/kg/day (1.33 micrograms/ml at 3 h and none detected at 24 h). Confirmatory results were obtained by high-pressure liquid chromatography assay. These studies show SCH 56592 to be a promising candidate for studies of treatment of histoplasmosis in humans.

Treatment of histoplasmosis with MK-991 (L-743,872).

BALB/c nu/+ immunocompetent and athymic (nu/nu) mice were infected intravenously with yeast cells of Histoplasma capsulatum. Mice were either given water (controls) intraperitoneally (i.p.) or given MK-991 i.p. once daily or twice daily. Protection was measured as prolonged survival or reduction in tissue counts. MK-991 was protective in immunocompetent mice, prolonging survival and reducing counts in spleen and livers at a dose as low as 0.05 mg/kg of body weight/day. MK-991 was modestly effective in athymic mice at a higher dose, 5 mg/kg/day. These studies suggest that MK-991 may be appropriate for clinical development in histoplasmosis.

Hypothesis on the mechanism of resistance to fluconazole in Histoplasma capsulatum.

An AIDS patient with disseminated histoplasmosis who improved during treatment with fluconazole but remained fungemic and subsequently relapsed is described. Isolates obtained from blood during therapy showed a progressive increase in fluconazole MIC from 0.625 to 20 micrograms/ml. The pretreatment, or parent, isolate and the posttreatment, or relapse, isolate demonstrated identical genetic patterns by PCR fingerprinting with three different primers. Fluconazole was less potent inhibitor of the growth of the relapse isolate than of the pretreatment isolate (50% inhibitory concentration [IC50] = 11.7 microM), while itraconazole was more potent (relapse isolate IC50 = 0.0011 microM versus pretreatment isolate IC50 = 0.0064 microM). Neither the increased sensitivity to itraconazole nor the decreased activity of fluconazole on the growth of the relapse isolate results from changes in the intracellular content of these agents. To reach 50% inhibition of ergosterol synthesis in both the parent and relapse isolates, about 2 nM itraconazole was needed; with fluconazole, 50% inhibition was achieved at 20.9 microM and 55.5 microM, respectively. Resistance to fluconazole may develop during treatment and results from decreased sensitivity of ergosterol synthesis.

Compuestos triazólicos en el tratamiento de la histoplasmosis diseminada experimental del hamster / Triazoles compound in the treatment of disseminated experimental histoplasmosis in hamsters

Se llevó a cabo un estudio comparativo de la eficacia de cuatro compuestos triazólicos en el tratamiento de la histoplasmosis experimental del hamster. Fueron utilizados en total 110 hamsters, de ambos sexos. Estos animales se inocularon por vía intracardíaca con una suspensión de la fase levaduriforme del Histoplasma capsulatum. Los tratamientos comenzaron 1 semana después de la infección y se prolongaron por 3 semanas. Todas las drogas fueron administradas por gastroclisis una vez por día, a las dosis de 8 y 16 mg/kg de peso. Tres grupos de hamsters fueron empleados como control y recibieron los solventes de los antifúngicos estudiados. Fueron administrados los siguientes triazoles: itraconazol, fluconazol, Sch 39.304 y Bay r 3783. Todos los animales se sacrificaron una semana después de terminados los tratamientos. La evaluación de los resultados se realizó en base a los siguientes criterios: 1)Aspecto macroscópico de higado y bazo, 2)examen microscópico de frotis teñidos con Giemsa de los mismos órganos, 3)cortes histológicos de bazo teñidos por el método de P.A.S.y 4)cultivos de bazo (cultivos masivos de un homogeneizado del órgano y determinación de U.F.C/g). El itraconazol y el Sch 39.304 redujeron en forma muy marcada las alteraciones macroscópicas, la positividad de los exámenes microscópicos y los cultivos. Los animales tratados con fluconazol y con Bay r 3783 a la dosis de 16 mg/kg/día, sólo mostraron escasa reducción del número de U.F.C/g de bazo, en relación a los controles. El itraconazol es una droga de probada eficacia en la histoplasmosis humana. El Sch 39.304 parece ser un antifúngico interesante en el tratamiento de esta enfermedad, en especial si se tienen en cuenta sus propiedades fármaco-cinéticas distintas de las itraconazol lo que facilitaría su aplicación en otras localizaciones de la histoplasmosis diseminada

Compuestos triazólicos en el tratamiento de la histoplasmosis diseminada experimental del hamster / Triazoles compound in the treatment of disseminated experimental histoplasmosis in hamsters

Se llevó a cabo un estudio comparativo de la eficacia de cuatro compuestos triazólicos en el tratamiento de la histoplasmosis experimental del hamster. Fueron utilizados en total 110 hamsters, de ambos sexos. Estos animales se inocularon por vía intracardíaca con una suspensión de la fase levaduriforme del Histoplasma capsulatum. Los tratamientos comenzaron 1 semana después de la infección y se prolongaron por 3 semanas. Todas las drogas fueron administradas por gastroclisis una vez por día, a las dosis de 8 y 16 mg/kg de peso. Tres grupos de hamsters fueron empleados como control y recibieron los solventes de los antifúngicos estudiados. Fueron administrados los siguientes triazoles: itraconazol, fluconazol, Sch 39.304 y Bay r 3783. Todos los animales se sacrificaron una semana después de terminados los tratamientos. La evaluación de los resultados se realizó en base a los siguientes criterios: 1)Aspecto macroscópico de higado y bazo, 2)examen microscópico de frotis teñidos con Giemsa de los mismos órganos, 3)cortes histológicos de bazo teñidos por el método de P.A.S.y 4)cultivos de bazo (cultivos masivos de un homogeneizado del órgano y determinación de U.F.C/g). El itraconazol y el Sch 39.304 redujeron en forma muy marcada las alteraciones macroscópicas, la positividad de los exámenes microscópicos y los cultivos. Los animales tratados con fluconazol y con Bay r 3783 a la dosis de 16 mg/kg/día, sólo mostraron escasa reducción del número de U.F.C/g de bazo, en relación a los controles. El itraconazol es una droga de probada eficacia en la histoplasmosis humana. El Sch 39.304 parece ser un antifúngico interesante en el tratamiento de esta enfermedad, en especial si se tienen en cuenta sus propiedades fármaco-cinéticas distintas de las itraconazol lo que facilitaría su aplicación en otras localizaciones de la histoplasmosis diseminada

In vitro and in vivo activities of Sch 39304, fluconazole, and amphotericin B against Histoplasma capsulatum.

The antifungal activities of amphotericin B and two triazoles, Sch 39304 and fluconazole, were tested against Histoplasma capsulatum. In this study Sch 39304 compared favorably with amphotericin B in treating histoplasmosis in normal and leukopenic mice, whereas fluconazole was much less active. The differences in the efficacies of the triazoles appeared to be due to differences in their pharmacokinetics and the dosage schedule that was used. For amphotericin B there was a good correlation between in vitro and in vivo efficacy, but this was not true of the triazole derivatives. These results further demonstrate that, with the methods used in this study, in vitro susceptibility testing of triazoles may not be predictive of in vivo activity against isolates of H. capsulatum.

Treatment of systemic mycoses with ketoconazole: in vitro susceptibilities of clinical isolates of systemic and pathogenic fungi to ketoconazole.

Ketoconazole was tested in vitro in three different media against 69 isolates of pathogenic fungi by using a macro-broth dilution procedure. The dimorphic systemic pathogens were highly susceptible, with most isolates of Blastomyces dermatitidis and Histoplasma capsulatum being inhibited and killed by concentrations less than or equal to 0.39 micrograms of ketoconazole/ml. Most isolates of Coccidioides immitis were also inhibited or killed by 0.39 micrograms of ketoconazole/ml; however, several were not killed by 100 micrograms/ml. Isolates of Cryptococcus neoformans and Sporothrix schenckii appeared to be less susceptible, with many isolates being resistant to less than or equal to 1.56 micrograms of ketoconazole/ml. There were 19 isolates of B. dermatitidis, C. immitis, and H. capsulatum recovered from 12 patients either during or following treatment with ketoconazole. Evidence for selection of secondary resistance to ketoconazole in these isolates was not observed. Results of these in vitro studies correlated poorly with the clinical responses to ketoconazole observed in the patients from whom the isolates were recovered.

In-vitro studies with four new antifungal agents: BAY n 7133, bifonazole (BAY h 4502), ICI 153,066 and Ro 14-4767/002.

Four new antifungal agents were compared in vitro with miconazole and ketoconazole. The agents were BAY n 7133 and ICI 153,066, two orally active triazoles, and bifonazole (BAY h 4502) and Ro 14-4767/002, both topical agents. While all four were found to be broad spectrum antifungal agents they also demonstrated certain gaps in their spectra. In general, Ro 14-4767/002 was the most active agent tested whereas bifonazole and BAY n 7133 were the least active. Noteworthy activities included that of Ro 14-4767/002 against Candida albicans, the dermatophytes and Sporothrix schenckii and that of ICI 153,066 against Torulopsis glabrata.

Selenocystine-resistant mutants of Histoplasma capsulatum.

Cysteine metabolism has been thought to be important to the phenomenon of dimorphism in Histoplasma capsulatum. We sought mutants with genetic blocks in the metabolism of cysteine by selection of colonies resistant to the toxic analogue, selenocystine. The 22 resistant strains thus obtained were all deficient in uptake of cystine from the surrounding medium but were normally able to convert from mycelium to yeast and back again. Furthermore, they had normal quantities of NADH-dependent cystine reductase when this enzyme was measured. We conclude that mutants defective in cystine uptake can be readily obtained by selection of colonies resistant to selenocystine, and that a lesion in cystine-uptake does not appear to affect the phenomenon of dimorphism in this organism.

Suppression of cryptococcosis and histoplasmosis by ketoconazole in athymic nude mice.

Congenitally athymic nude (nu/nu) mice are more susceptible to disseminated cryptococcosis and histoplasmosis than their heterozygous (nu/ + ) thymus-bearing litter-mates. The therapeutic efficacy of ketoconazole, an orally absorbable antifungal agent, was evaluated in nu/nu and nu/ + mice infected intraperitoneally with Cryptococcus neoformans and Histoplasma capsulatum. Two- to five-week courses of ketoconazole significantly prolonged the survival of nu/nu mice infected with either fungus in dose-dependent fashion, but death eventually followed discontinuance of therapy. More significant protection was seen in nu/ + mice infected with C. neoformans, and markedly lower fungal counts in organs, with some negative cultures, were seen in ketoconazole-treated nu/ + mice infected with H. capsulatum. These studies indicate that ketoconazole is effective against both fungi, although results of treatment are much better in the immunologically intact nu/ + host.