Prospective multicentre study on azole resistance in Aspergillus isolates from surveillance cultures in haematological patients in Italy.

OBJECTIVES: This study was conducted to assess the prevalence of azole resistance in Aspergillus isolates from patients with haematological malignancies or who were undergoing haematopoietic stem cell transplantation and to identify the molecular mechanism of resistance. METHODS: In this 28-month prospective study involving 18 Italian centres, Aspergillus isolates from surveillance cultures were collected and screened for azole resistance, and mutations in the cyp51A gene were identified. Resistant isolates were genotyped by microsatellite analysis, and the allelic profiles were compared with those of resistant environmental and clinical isolates from the same geographical area that had been previously genotyped. RESULTS: There were 292 Aspergillus isolates collected from 228 patients. The isolates belonged mainly to the section Fumigati (45.9%), Nigri (20.9%), Flavi (16.8%) and Terrei (4.8%). Three isolates showed itraconazole resistance: Aspergillus fumigatus sensu stricto, Aspergillus lentulus (section Fumigati) and Aspergillus awamori (section Nigri). The itraconazole resistance rates were 1% and 1.48% considering all Aspergillus spp. isolates and the Aspergillus section Fumigati, respectively. The prevalence of azole resistance among all the patients was 1.3%. Among patients harbouring A. fumigatus sensu stricto isolates, the resistance rate was 0.79%. The A. fumigatus isolate, with the TR34/L98H mutation, was genotypically distant from the environmental and clinical strains previously genotyped. CONCLUSIONS: In this study, the Aspergillus azole resistance rate was 1% (3/292). In addition to A. fumigatus sensu stricto, A. lentulus and A. awamori azole-resistant isolates were identified. Therefore, it is important have a correct identification at the species level to address a rapid therapy better, quickly understand the shift towards cryptic species and have an updated knowledge of the local epidemiology.

Evolution of fungemia in an Italian region.

BACKGROUND: Fungemia represents a public health concern. Knowing aetiology and activity of the antifungals is critical for the management of bloodstream infections. Therefore, surveillance on local/international levels is desirable for a prompt administration of appropriate therapy. METHODS: Data on fungi responsible for fungemia and antifungal susceptibility profiles were collected from a laboratory-based surveillance over 2016-2017 in 12 hospitals located in Lombardia, Italy. The trend of this infection in twenty years was analysed. RESULTS: A total of 1024 episodes were evaluated. Rate of candiaemia progressively increased up to 1.46/1000 admissions. C.albicans was the most common species (52%), followed by C. parapsilosis (15%) and C glabrata (13%). As in the previous surveys the antifungal resistance is rare (echinocandins<2%, fluconazole 6%, amphotericin B 0.6%). Fungi other than Candida were responsible for 18 episodes: Cryptococcus neoformans (5 cases), Fusarium spp. (4), Magnusiomyces clavatus (3), Saccharomyces cerevisiae (3), Rhodotorula spp. (2), Exophiala dermatitidis (1). All fungi, except S.cerevisiae, were intrinsically resistant to echinocandins. Some isolates showed also elevated azole MIC. CONCLUSIONS: No particular changes in terms of species distribution and antifungal susceptibility patterns was noted. However, surveillance programs are needed to monitor trends in antifungal resistance, steer stewardship activities, orient empirical treatment.

Human infections due to Schizophyllumcommune: Case report and review of the literature.

Schizophyllumcommune is an environmental basidiomycetous fungus, causing occasional, predominantly respiratory, infections in humans. Although S. commune is considered an emerging pathogen, some authors pointed out the possibility that the increase in the diagnosed cases may be also due to recent advances in diagnostic technologies now allowing a more prompt and precise identification at the species level. Here we describe the first Italian case of chronic non-invasive fungal rhinosinusitis due to S. commune in an immunocompetent subject and update the literature review on S. commune sinusitis published between 2012-2019. A timely diagnosis is important to avoid local and systemic complications due to infection with this fungus. In our case, prompt identification at species level was only possible with the use of MALDI-TOF mass spectrometry and confirmed by sequence analysis of ribosomal DNA ITS regions, due to the difficulty in achieving a correct and rapid identification using routine morphological analysis.

Method-Dependent Epidemiological Cutoff Values for Detection of Triazole Resistance in Candida and Aspergillus Species for the Sensititre YeastOne Colorimetric Broth and Etest Agar Diffusion Methods.

Although the Sensititre Yeast-One (SYO) and Etest methods are widely utilized, interpretive criteria are not available for triazole susceptibility testing of Candida or Aspergillus species. We collected fluconazole, itraconazole, posaconazole, and voriconazole SYO and Etest MICs from 39 laboratories representing all continents for (method/agent-dependent) 11,171 Candida albicans, 215 C. dubliniensis, 4,418 C. glabrata species complex, 157 C.guilliermondii (Meyerozyma guilliermondii), 676 C. krusei (Pichia kudriavzevii), 298 C.lusitaniae (Clavispora lusitaniae), 911 C.parapsilosissensu stricto, 3,691 C.parapsilosis species complex, 36 C.metapsilosis, 110 C.orthopsilosis, 1,854 C.tropicalis, 244 Saccharomyces cerevisiae, 1,409 Aspergillus fumigatus, 389 A.flavus, 130 A.nidulans, 233 A.niger, and 302 A.terreus complex isolates. SYO/Etest MICs for 282 confirmed non-wild-type (non-WT) isolates were included: ERG11 (C. albicans), ERG11 and MRR1 (C. parapsilosis), cyp51A (A. fumigatus), and CDR2 and CDR1 overexpression (C. albicans and C. glabrata, respectively). Interlaboratory modal agreement was superior by SYO for yeast species and by the Etest for Aspergillus spp. Distributions fulfilling CLSI criteria for epidemiological cutoff value (ECV) definition were pooled, and we proposed SYO ECVs for S. cerevisiae and 9 yeast and 3 Aspergillus species and Etest ECVs for 5 yeast and 4 Aspergillus species. The posaconazole SYO ECV of 0.06 µg/ml for C. albicans and the Etest itraconazole ECV of 2 µg/ml for A. fumigatus were the best predictors of non-WT isolates. These findings support the need for method-dependent ECVs, as, overall, the SYO appears to perform better for susceptibility testing of yeast species and the Etest appears to perform better for susceptibility testing of Aspergillus spp. Further evaluations should be conducted with more Candida mutants.

Fungemia due to Saprochaete capitata in a non-neutropenic patient hospitalized in an intensive care unit after cardiac surgery.

The majority of invasive fungal infections observed in non-neutropenic patients hospitalized in an intensive care unit are caused by Candida spp and current guidelines recommend echinocandins as the first-line treatment. Fungemias caused by filamentous or arthrosporic fungi such as Saprochaete capitata (previously named Geotrichum capitatum) are extremely rare. In fact, invasive infections due to S. capitata have been reported almost exclusively in neutropenic oncohematological patients. In this report, we describe a case of fungemia caused by S. capitata in a non-neutropenic patient hospitalized in an intensive care unit after aortic valve replacement. The prompt identification of S. capitata is extremely important because of its intrinsic resistance to echinocandins.

European Confederation of Medical Mycology (ECMM) epidemiological survey on invasive infections due to Fusarium species in Europe.

In order to better understand the epidemiology of fusariosis in Europe, a survey collecting information on the clinical characteristics of the patients infected by Fusarium as well as on the infecting isolates was launched. A total of 76 cases of invasive fusariosis occurring from January 2007 to June 2012 were collected and Fusarium isolates were identified by sequencing the translation elongation factor 1α (TEF) gene. Also, antifungal susceptibility was tested by broth microdilution according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the Etest. Disseminated disease was considered proven in 46 cases and probable in 17 cases. Localised infection was seen in 13 cases. Gibberella fujikuroi species complex (SC), including Fusarium verticillioides and F. proliferatum, and F. solani SC were the most frequent aetiology of disseminated and localised infections, respectively. The crude mortality rate was 46 %, the highest associated with F. solani SC (67 %) and F. proliferatum (62.5 %). A wide range of antifungal susceptibilities was observed. Amphotericin B was the most potent antifungal in vitro, and itraconazole the least effective. The azoles exhibited lower minimum inhibitory concentrations (MICs) against F. verticillioides strains, with posaconazole having a slightly better performance, while F. solani SC isolates were resistant to all three azoles tested. The essential agreement between the Etest and the EUCAST method was 100 % for itraconazole and voriconazole, and 96 % for amphotericin B and posaconazole. In conclusion, we confirm that fusariosis is a rare but severe event in Europe, that G. fujikuroi SC is the predominant cause of deep infections and that different species have different antifungal in vitro susceptibility patterns.

Candida glabrata and pre-term premature rupture of membrane complicating in vitro pregnancy: case report and confirmation of mother to neonate transmission.

CASE REPORT: We report a case of Candida glabrata infection in an in vitro fertilization-assisted pregnancy complicated by pre-term pre-labor rupture of the membranes. We found C. glabrata in gastric fluid, amniotic fluid and maternal vaginal swab. Neonatal and maternal isolates showed indistinguishable molecular patterns analyzed by fingerprint DNA multilocus. DISCUSSION: Strong consideration should be given to perform a screening test C. glabrata. Multiple treatments, even in pregnancy, should be considered in women who have positive cultures results.

Invasive fungal sinusitis due to Bipolaris hawaiiensis.

A phaeohyphomycotic sinusitis due to Bipolaris hawaiiensis, observed in an immunologically competent patient, is reported and a review of data from the literature is given. The patient was successfully treated by surgical drainage and amphotericin B.

Acid tolerance and fecal recovery following oral administration of Saccharomyces cerevisiae.

Probiotic microorganisms to be used as biotherapeutic agents have to resist the rigors of the upper human gastrointestinal tract. In this study we evaluated the acid tolerance in vitro and the fecal recovery in vivo after oral administration of a Saccharomyces cerevisiae strain to healthy volunteers. At the lowest pH value (pH 1.0) the yeast load in tablets decreased slightly. From pH 1.0 to pH 7.0 the release of S. cerevisiae in buffer solutions increased. The selected yeast strain showed good tolerance to low pH which mimic the gastric environment. After one month of treatment at a dose of 100 million cells per day, S. cerevisiae grew from the feces of 6 (37.5%) of the 16 healthy, treated volunteers. Based on the results of the present experiments the yeast studied can be considered a strain that tolerates adverse conditions comparable to those of the human gastrointestinal tract, and when administered orally may colonize the bowel of healthy volunteers and even replace resident Candida species.

European Confederation of Medical Mycology (ECMM) prospective survey of candidaemia: report from one Italian region.

An ECMM epidemiological prospective survey of candidaemia was performed in one Italian region (Lombardy; population: 8 924 870) by the National Society of Medical Mycology (FIMUA) from September 1997 to December 1999. In total, 569 episodes were reported with an overall rate of 0.38/1000 admissions, 4.4/100000 patient days. Predisposing factors included presence of an intravascular catheter (89%), antibiotic treatment (88%), surgery (56%), intensive care (45%), solid tumour (28%), steroid treatment (15%), haematological malignancy (7%), HIV infection (6%), fetal immaturity (4%). Mucous membrane colonization preceded candidaemia in 83% of patients. Candida albicans was identified in 58% of cases, followed by Candida parapsilosis (15%), Candida glabrata (13%), Candida tropicalis (6%). Septic shock occurred in 95 patients. Crude mortality was 35%, the highest in C. tropicalis fungaemia (44%), the elderly (64%) and solid tumour cancer patients (43%). Intravascular catheter removal was associated with higher survival rate (71 vs. 47%). This survey underscores the importance of candidaemia in hospital settings.

Multicenter comparative evaluation of six commercial systems and the national committee for clinical laboratory standards m27-a broth microdilution method for fluconazole susceptibility testing of Candida species.

Fluconazole susceptibility among 800 clinical Candida isolates (60% C. albicans) and two control strains (C. krusei ATCC 6258 and C. parapsilosis ATCC 22019) was tested with the NCCLS M27-A method (gold standard) and six commercial products (Candifast, disk, Etest, Fungitest, Integral System Yeasts, and Sensititre YeastOne). Results were classified as susceptible, susceptible-dose dependent, or resistant using M27-A breakpoints or, for Fungitest, Integral System Yeasts, and Candifast, as susceptible, intermediate, or resistant, according to the manufacturers' instructions. Concordance with NCCLS M27-A results was analyzed with the chi(2) test. Intra- and interlaboratory reproducibility was also evaluated. NCCLS M27-A (90.1%), Etest (93.1%), Sensititre YeastOne (93.1%), disk (96.7%), Fungitest (92.6%), Integral System Yeasts (40.6%), and Candifast (6.0%) classified the indicated percentages of C. albicans isolates as susceptible. Among non-C. albicans strains, the percentages of susceptible isolates were as follows: NCCLS M27-A, 74.0%; Etest, 83.8%; Sensititre YeastOne, 64.1%; disk, 60.6%; Fungitest, 76.6%; Integral System Yeasts, 28.3%; and Candifast, 27.4%. All methods except Candifast and Integral System Yeasts showed good agreement with NCCLS M27-A results for both C albicans and non-C. albicans isolates. Intralaboratory reproducibility was excellent for NCCLS M27-A, Etest, Sensititre YeastOne, disk, and Fungitest (88 to 91%). Similar results emerged from the interlaboratory reproducibility evaluation. Our findings indicate that some commercial methods can be useful for fluconazole susceptibility testing of clinical Candida isolates. Those characterized by a lack of medium standardization and/or objective interpretative criteria should be avoided. Particular caution is necessary when testing is being done for clinical and epidemiological purposes.

Imported and autochthonous histoplasmosis in Bergamo province, Northern Italy.

This paper reviews the Italian literature and illustrates the results of a survey on Histoplasma capsulatum var. capsulatum infection carried out in the Bergamo area of Italy over the last 10 y. During the period January 1989-July 1999, 7 patients were diagnosed as being affected by the H. capsulatum var. capsulatum infection. Until 1999 41 cases have been described in the Italian literature (9 before the 1980s, 6 in the 1980s and 26 in the 1990s). The epidemiological profile of histoplasmosis is well-defined: risk factors are activities such as visiting caves or building sites, travelling and immigration, and it affects both immunocompetent and patients with AIDS. However, in the past, the Italian soil was considered as a low-endemic pabulum for H. capsulatum var. capsulatum and only a few autochthonous cases of histoplasmosis have been reported in Italy, specifically in the Po valley. In the present series, the identification of new cases in the Po valley in Lombardy suggests the possible autochthonous presence of histoplasmosis in Italy.

Primary cutaneous cryptococcosis in an HIV-negative patient.

We report a case of primary cutaneous cryptococcosis in an HIV-negative patient, who presented with painless, ulcerated lesions involving the right forearm (fingers and elbow), which developed over 45 days. On the basis of the clinical appearance, serological and cultural examinations were performed to confirm the diagnosis; the histological evaluation of a skin biopsy showed an acute inflammatory infiltrate containing several PAS + Cryptococci. Subsequently, the patient was treated with fluconazole (400 mg/day for 10 days, then a maintenance therapy of 200 mg/day); after one month, the cutaneous lesions were remarkably improved, but, although a series of further laboratory and clinical examinations was scheduled, the patient repeatedly refused any other re-evaluation, and he was lost from follow-up.

Clinical and microbiological characteristics of symptomatic vulvovaginal candidiasis in HIV-seropositive women.

OBJECTIVES: To evaluate the clinical and microbiological characteristics of symptomatic vaginal candidiasis in Human Immunodeficiency Virus (HIV)-seropositive women attending a gynaecologic outpatient clinic for sexually transmitted diseases (STDs). DESIGN: Vaginal, rectal and oral specimens from cases and controls were cultured for Candida spp. SUBJECTS: Eighty-four consecutive HIV-seropositive and 384 HIV-seronegative women with clinical signs of vulvovaginitis. SETTING: A gynaecological out-patient clinic in Pavia, Italy. RESULTS: The overall prevalence of vaginal candidiasis was 61.9% (52/84) in the cases and 32.3% (124/384; p < .001) in the controls. After adjustment by logistic regression analysis for confounding factors (age at first intercourse, lifetime sex partners, new partner/s in the last 6 months, type of contraceptive used), HIV-seropositive patients were at higher risk for both Candida albicans (odds ratio = 2.5; 95% confidence interval 1.31-4.69; p = 0.006) and Torulopsis glabrata vaginitis (OR = 3.5; 95% CI = 1.05-11.60; p = 0.04) than controls. HIV-seropositive subjects had also increased rates of oral and rectal colonisation with Candida spp. Finally, the time to recurrence of vaginal infection was significantly shorter in HIV-seropositive patients than controls and was correlated with the severity of HIV-induced immunodepression. CONCLUSIONS: Vulvovaginal candidiasis is very common in HIV-seropositive women and its prevalence is correlated with the immunological status of the host. These patients have higher frequencies of Torulopsis glabrata vaginal infection and are more prone to recurrence than HIV-seronegative controls.

Frequency and significance of drug resistance in vulvovaginal candidiasis.

The rates of non-albicans species and susceptibility to the main antimycotic drugs were evaluated in a series of 472 cultures of vulvar or vaginal specimens positive for Candida species. Torulopsis glabrata was associated more frequently than C. albicans with recurrent vulvovaginal candidiasis (18/40 vs. 72/379; p = 0.003, chi 2 test). As evaluated by the results of susceptibility testing, C. albicans isolates were uniformly sensitive to the antifungals tested, whereas 7.5% (3/40) of T. glabrata strains were resistant to imidazole derivatives. All these patients had a history of repeated antimycotic treatments. Previous antimycotic therapies in susceptible individuals could cause a selection of non-albicans species, especially T. glabrata, which is occasionally resistant to the commonly used antimycotic drugs.

Recurrent vaginal candidiasis. Results of a cohort study of sexual transmission and intestinal reservoir.

Yeast cultures from the oral cavity, vagina and rectum were obtained from 125 women experiencing an acute episode of recurrent candidal vaginitis. To investigate the role of sexual transmission, oral, penile and ejaculate cultures were also prepared from all the male sexual partners. The rates of oral and rectal Candida species colonization in the women were 36% (45/125) and 44.8% (56/125), respectively. The male partners' oral cavities were positive in 23.2% (29/125) and the penile coronal sulcus and seminal fluid in 16% (20/125) and 14.4% (18/125), respectively. The susceptibility of the isolated species to the main antimycotic drugs was ascertained with the agar diffusion method. Therapy in the women and the colonized sexual partners was carried out, eliminating the microorganism from every positive site. Control cultures were obtained two weeks after the completion of therapy, and follow-up was continued for one year. The overall clinical and microbiologic cure rate in the study group was 72% (95/125). The rate of relapse was not influenced by the treatment of Candida colonization of the female intestinal tract. The recurrence rate after treatment in the couples in which the man harbored yeast (oral cavity, penile coronal sulcus, seminal fluid) was lower (15.8% vs. 44.8%, P = .0019) than that recorded in the couples without sexual partner involvement. The identification and treatment of the male sexual partner's Candida colonization seems important in the prevention of recurrent vulvovaginitis.

Recurrent Histoplasma capsulatum pneumonia: a case report.

The epidemiological, clinical and therapeutical findings are described in a case of recurrent pulmonary histoplasmosis due to Histoplasma capsulatum. The patient, a bulldozer-operator, worked in Africa for a long period in extremely dusty conditions without any protection. Three different episodes of H. capsulatum pneumonia recurred during eighteen months after his return from Africa. A full dose treatment by Amphotericin B failed to eliminate disease recurrence on three occasions. The high concentration of airborne H. capsulatum spores inhaled could have been the main cause of the difficulty obtaining a rapid sterilization of the microorganism by Amphotericin B and disease recurrence. The late start of the treatment or the unexplained ability of some persons to develop repeated infections even with normal immunological parameters could be another explanation for the reported phenomenon.

Antifungal chemoprophylaxis in cancer children: a prospective randomized controlled study.

We report a prospective, randomized, controlled study of antifungal chemoprophylaxis in forty immunocompromised children with hematologic malignancies, receiving respectively itraconazole, ketoconazole, amphotericin-B and no prophylaxis. Fungal isolates from patients' saliva or stools were obtained from 19/40 patients (Candida albicans from 15 patients). Disseminated infection was never observed. Fungal isolates were significantly less frequent in the ketoconazole group of patients vs any other group. Systemic antifungal chemoprophylaxis with ketoconazole appears more effective even than the recently introduced itraconazole.

Fluconazole vs amphotericin B: "in vitro" comparative evaluation of the minimal inhibitory concentration (MIC) against yeasts isolated from AIDS patients.

Fluconazole, a novel triazole antifungal drug, holds promise as a significant advance in the management of human fungal diseases. This new drug can be both orally and parenterally administered and is reported to be less toxic than other antifungal agents. The Authors studied the in vitro susceptibility of yeasts isolated from AIDS patients to this antimycotic drug, in order to evaluate if fluconazole's MICs were comparable to those of amphotericin B. A sample of 200 yeast strains (100 C.albicans, 20 C.parapsilosis, 20 C.tropicalis, 8 C.guilliermondii, 6 C.krusei, 6 C.pseudotropicalis, 24 T.glabrata and 16 Cr.neoformans) was tested. The results show an average MIC for amphotericin B (MIC90: 3.12 micrograms/ml) ten-fold lower than for fluconazole (MIC90: 100 micrograms/ml). However, C.albicans and Cr.neoformans presented the same MIC value (3.12 micrograms/ml) both for fluconazole and for amphotericin B, suggesting that fluconazole could represent a valid alternative to amphotericin B in the treatment of fungal infections caused by these two agents.

Itraconazole vs amphotericin B: in vitro comparative evaluation of the minimal inhibitory concentration (MIC) against clinically isolated yeasts.

Itraconazole is a triazole compound which, following several clinical trials, has begun to be used for therapy of mycotic infections. This new drug, with a broad-spectrum antifungal activity, can be orally administered. The Authors studied the in vitro susceptibility to amphotericin B and itraconazole of the following clinical isolates of pathogenic yeasts: 100 Candida albicans, 20 C. tropicalis, 20 C. parapsilosis, 8 C. guilliermondii, 6 C. pseudotropicalis, 24 Torulopsis glabrata and 16 Cryptococcus neoformans. Serial two-fold dilution, from 100 micrograms/ml to 0.04 micrograms/ml, of each drug were prepared in Yeast Nitrogen Base + Glucose 5%, after dissolving the itraconazole in dimethylsulfoxide (DMSO) and amphotericin B in 5% glucose solution. Amphotericin B (MIC90: 3.12 micrograms/ml) was found to have an average in vitro MIC six-fold lower than itraconazole (MIC90: 25 micrograms/ml). Thus, even though itraconazole is active, amphotericin B remains one of the most effective of the antifungal drugs.