Environmental surveillance of fungi and susceptibility to antifungal agents in tertiary care hospitals.

IMPORTANCE: Saprophytic fungi can cause nosocomial infections in high-risk patients. These infections are related to high mortality and cost. In the current study, different species of filamentous fungi and yeast were isolated from the environment of the studied hospitals. Some species were resistant to antifungal drugs. We suggest that the future work concentrates on the relationship between the level/quantification of saprophytic contamination in the environment of hospitals and fungal infections in patients.

Comparison of in vitro activities of newer triazoles and classic antifungal agents against dermatophyte species isolated from Iranian University Hospitals: a multi-central study.

BACKGROUND: Dermatophytes have the ability to invade the keratin layer of humans and cause infections. The aims of this study were the accurate identification of dermatophytes by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism method and sequencing and comparison between the in vitro activities of newer and established antifungal agents against them. METHODS: Clinical specimens of patients from five Iranian university laboratories were entered in this study. Samples were cultured on sabouraud dextrose agar medium. For molecular identification, extracted DNAs were amplified by the universal fungal primers ITS1 and ITS4, and digested with MvaI enzymes. The antifungal susceptibility test for each isolate to terbinafine, griseofulvin, caspofungin, fluconazole, itraconazole, luliconazole, and isavuconazole was performed, according to the microdilution CLSI M38-A2 and CLSI M61 standard methods. RESULTS: Two hundred and seven fungi species similar to dermatophytes were isolated of which 198 (95.6%) were dermatophytes by molecular assay. The most commonly isolated were Trichophyton mentagrophytes (76/198), followed by Trichophyton interdigitale (57/198), Trichophyton rubrum (34/198), Trichophyton tonsurans (12/198), Microsporum canis (10/198), Trichophyton simii (3/198), Epidermophyton floccosum (3/198), Trichophyton violaceum (2/198), and Trichophyton benhamiae (1/198). The GM MIC and MIC90 values for all the isolates were as follows: terbinafine (0.091 and 1 µg/ml), griseofulvin (1.01 and 4 µg/ml), caspofungin (0.06 and 4 µg/ml), fluconazole (16.52 and 32 µg/ml), itraconazole (0.861 and 8 µg/ml), isavuconazole (0.074 and 2 µg/ml), and luliconazole (0.018 and 0.25 µg/ml). CONCLUSION: Trichophyton mentagrophytes, Trichophyton interdigitale, and Trichophyton rubrum were the most common fungal species isolated from the patients. luliconazole, terbinafine, and isavuconazole in vitro were revealed to be the most effective antifungal agents against all dermatophyte isolates.

Epidemiology and Antifungal Susceptibility of Candida Species Isolated from 10 Tertiary Care Hospitals in Iran.

In recent decades, the incidence of Candida infections has increased in immunocompromised patients. This multicenter study aimed to evaluate in vitro antifungal activities of 8 antifungal agents against the Candida species isolated from 10 university hospitals in Iran. During the period from Dec 2019 to Dec 2021, Candida species were collected from clinical samples of patients. The isolates were identified by PCR restriction fragment length polymorphism and sequencing methods. The antifungal susceptibility tests of each isolate to eight antifungal agents were performed according to the microdilution CLSI M27, M59, and M60 standard methods. A total of 598 Candida strains were isolated from clinical samples. The most commonly isolated Candida species was C. albicans, followed by C. glabrata, C. parapsilosis, Debaryomyces hansenii (Candida famata), C. tropicalis, Pichia kudriavzevii (Candida krusei), C. orthopsilosis, Meyerozyma guilliermondii (Candida guilliermondii), Kluyveromyces marxianus (Candida kefyr), and Clavispora lusitaniae (Candida lusitaniae). MIC90 values in all Candida species were as follows: 0.25 µg/mL for caspofungin and voriconazole; 0.5 µg/mL for amphotericin B and isavuconazole; 2 µg/mL for itraconazole, luliconazole, and posaconazole; and 16 µg/mL for fluconazole. Although 30/285 C. albicans, 15/31 C. hansenii, 3/12 M. guilliermondii, 67/125 C. glabrata, 5/15 P. kudriavzevii, 6/60 C. parapsilosis, and 5/23 C. tropicalis isolates were multiazole resistant with resistance to 2 to 4 azoles, pan-azole resistance was not observed. According to our data, Candida albicans and C. glabrata were the most frequent species isolated from clinical samples in Iran. Caspofungin and voriconazole, with lower MIC90 values, are the most effective than other antifungal agents for the treatment of Candida infections in this region. IMPORTANCE Candida species cause severe invasive infections of the heart, brain, eyes, bones, and other parts of the body. Knowledge of regional distributions of causative Candida agents and their antifungal susceptibility patterns can help to monitor resistance to antifungal agents of various species and support local and national surveillance programs. In the present study, C. albicans and C. glabrata were the most frequently isolated species from clinical samples in Iran. Increasing rates of non-albicans Candida isolates from the Iranian population should be looked at as alarming due to various levels of intrinsic MIC values or resistance to various antifungal drugs. Caspofungin and voriconazole are recommended over fluconazole for the treatment of Candida infections in the study region. However, amphotericin B and isavuconazole are also active against the most common Candida species isolated from patients. Pan azole-resistant Candida species were not observed in the present study.

Antifungal activity of Taurolidine against Mucorales: An in vitro study on clinical isolates.

Background and Purpose: Taurolidine is active against a wide variety of micro-organisms, including bacteria and fungi. Mucormycosis is one of the life-threatening opportunistic fungal infections, especially in immunocompromised patients. Currently, the emergence of Mucormycosis during the COVID-19 pandemic raises public health concerns regarding untoward morbidity and mortality among SARS-CoV-2 patients. It is well-known that delayed and inappropriate antifungal therapy leads to increased morbidity and mortality. This study aimed to investigate the in-vitro antifungal activity of taurolidine to evaluate its effects against clinical isolates of Mucorales. Materials and Methods: This study included previously collected clinical Mucorales isolates. The minimum in vitro inhibitory concentration (MIC) of amphotericin B, caspofungin, voriconazole, posaconazole, and itraconazole was determined using the broth microdilution method. Results: All clinical isolates showed full sensitivity to amphotericin B. Posaconazole MIC range from 8 µg/mL to 0.032 µg/mL. The MIC range of voriconazole and caspofungin were determined to be 2-8 µg/mL and 0.5-16 µg/mL, respectively. Growth of the isolates was entirely inhibited in 1000 µg/mL concentration of taurolidine. In microscopic observations, morphological effects on hyphal growth were observed at 500 µg/mL concentration. Conclusion: In conclusion, this is an updated experience of using taurolidine against Mucorales. However, our in-vitro findings need to be confirmed in well-designed clinical trials aimed at treating invasive Mucormycosis infections.

Role of biomarkers in the diagnosis of invasive aspergillosis in immunocompromised patients.

BACKGROUND: Invasive aspergillosis is one of the important causes of infection in immunocompromised patients. This study aimed to evaluate the roles of biomarkers in the diagnosis of invasive aspergillosis and their relationship with antifungal stewardship programs. METHODS: 190 sera from 52 immunocompromised patients and volunteer individuals were included in this study. 18 immunocompromised volunteers without IA and 34 patients with probable and proven aspergillosis according to the European Organization for Research and Treatment of Cancer and the Mycoses Study Group consensus definitions were entered in this study. The respective sera were evaluated for procalcitonin, soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) levels; white blood cells count (WBC) count, C reactive protein (CRP), lactate dehydrogenase (LDH), and erythrocyte sedimentation rate (ESR) values. Demographic data and clinical characteristics of patients were extracted from their files. RESULTS: The male-to-female ratio and mean age of patients were 22/12 and 38.9 years, respectively. The hematologic disorder was the most predisposing factor (29/34, 85.3%). Sensitivity of biomarkers for diagnosis of invasive aspergillosis was 70.6% (cut off value > 190 pg/mL for sTREM-1, 71% (cut off value > 260 pg/mL) for PCT, 85.3% (cut off value > 193 U/L) for LDH, 94.1% (cut off value > 8 mg/l) for CRP, 64.7% (cut off value < 5200 cells/ml) for WBC, and 85.3% (cut off value > 23 mm/h) for ESR. Twelve patients died, with significantly increased sTREM-1 levels and decreased WBC count in them. CONCLUSION: According to our data, evaluation of the biomarkers can help in the diagnosis, management, and prediction of the severity of Aspergillus infection, and the rational use of antifungal agents in immunocompromised patients.

Phenotype, genotype, and mating type determination in oral Candida albicans isolates from pediatric patients with neutropenia.

BACKGROUND: The most frequent species of Candida to infect and colonize patients with neutropenia is still Candida albicans. This study aimed to provide detailed information on the phenotype, genotype, and mating type of oral C. albicans isolated from neutropenic pediatric patients, and to investigate how these characteristics are related. METHODS: Two hundred fifty-four oral samples from patients under 18 years old with neutropenia and malignancies were collected from January to October 2021. Samples were cultured on CHROMagar Candida. Isolates of C. albicans were identified with the germ tube test, chlamydospore production on cornmeal agar, and PCR-RFLP. Genotyping of C. albicans isolates was carried out by amplifying the 25S rDNA gene with specific CAINT-L and CA-INT-R primers. MTLa1 and MTLα1 primers were used to identify each mating type. Yeast peptone dextrose supplemented with phloxine B was used to identify different phenotypes. RESULTS: Ninety-two (36%) patients were positive for C. albicans. The mean age of patients was 7.85. Fifty-three (58.9%) isolates demonstrated type A, 15 (16.7%) type B, 15 (16.7%) types D/E, and 7 (7.7%) type C. Three isolates each (3.3%) were homozygous for MTLa or homozygous for MTLα. All of the MTL-homozygous isolates were genotype A. There was a significant correlation between patients' underlying disease and genotype (p = 0.036). There was a significant correlation between mating type and genotype (p = 0.000). CONCLUSION: Most of the isolates exhibited a white phenotype, noted in the literature as the most virulent. Moreover, heterozygous strains were frequent and may play a role in Candida colonization.

Population structure, susceptibility profile, phenotypic and mating properties of Candida tropicalis isolated from pediatric patients.

BACKGROUND: Candida tropicalis is one of the most frequently isolated species and is commonly associated with nosocomial infections, hematological malignancy, neutropenia, and urinary tract infections. AIMS: This study aims to genotype C. tropicalis strains isolated from pediatric patients admitted to two hospitals in Ahvaz, Iran. We provide a vision of the genotypes, mating types, enzymatic activity, phenotypes, and antifungal susceptibility profile of these isolates. METHODS: Candida tropicalis isolates were collected from various clinical (Oral, urine, wound, and bronchoalveolar lavage) and environmental sources between November 2020 and November 2021. Primitively, samples were cultured on CHROMagar Candida. All isolates were identified by sequencing the Internal Transcribed Spacer (ITS) region for precise identification. Isolates were genotyped by six microsatellite markers specific for C. tropicalis. Antifungal susceptibility profiles were determined against eight antifungal agents according to CLSI M27 standards. The phenotype of each C. tropicalis isolate was assessed using yeast peptone dextrose agar supplemented with phloxine B. Mating types of C. tropicalis isolates were determined using MTLa1 and MTL2 specific primers. RESULTS: Species identification revealed 46 C. tropicalis strains. Among them, 39 different genotypes were detected that have split into 34 singletons and five clusters. Twenty isolates were the non-wild type for itraconazole and posaconazole. Four isolates were multidrug-resistant. The activity of hemolysin and esterase enzyme was very strong among all isolates. Mating type and phenotype were not significantly correlated with genotypes (p = 0.721 and p = 0.135, respectively). CONCLUSIONS: To conclude, tested populations were moderately differentiated with high gene flow. One cluster of isolates among different hospitals was identified, and three clusters were from different cities.

Evaluation of catalase activity of clinical and environmental isolates of Aspergillus species.

Background and Objectives: Catalases are a good scavenger of H2O2 which degrades hydrogen peroxide into water and oxygen. They are considered as a virulence factor that are present in both spores and hypha of fungi. There is limited data regarding catalase activity in Aspergillus species. This study aimed to assess the mycelial catalase activity of clinical and environmental isolates of Aspergillus niger, A. tubingensis, A. flavus, A. luchuensis, A. piperis and A. terreus. Materials and Methods: Briefly, clinical and environmental Aspergillus species were used in the current study. Catalase activity was assessed for both groups of isolates including 13 A. flavus (12 clinical, 1 environmental), 13 A. terreus (8 clinical, 5 environmental), 26 A. tubingensis (13 clinical, 13 environmental), and 44 A. niger (25 environmental, 19 clinical) species. Fungal balls of mycelia were separated from the liquid culture and were crushed using homogenizer. The supernatants were collected and used for a catalase activity assay. Results: Totally, in our study 98 Aspergillus including 45 environmental and 53 clinical isolates were assessed for catalase activity. High catalase activity was detected among environmental Aspergillus species (Mean= 1.62 mU/ml) and the mean of mycelial catalase activity among clinical A. terreus isolates was higher than environmental strains. Conclusion: In summary, mycelial catalase activity varied among species and environmental isolates demonstrated higher catalase activity. Totally a significant difference was found between clinical and environmental Aspergillus isolates.

Significance of biomarkers in stewardship program in pediatric patients infected with Aspergillus species.

BACKGROUND: The potential use of biomarkers in the diagnosis of fungal infections is a challenge. The aim of this study was to evaluate the role of a biomarker-guided antifungal stewardship program for hospitalized pediatrics suffering from invasive aspergillosis (IA). METHODS: Pediatric patients with suspected probable or proven IA were enrolled in this study. Demographic data were collected from their records. Clinical samples were examined by wet mount KOH smear, culture, galactomannan Ag test, and real-time PCR. Patients' sera were evaluated for procalcitonin (PCT) and soluble-triggering receptor expressed on myeloid cells -1 (sTREM-1) levels by ELISA Kits. RESULTS: A total of 73 children were entered in this study with a mean age of 5 years and the male to female ratio 39/34. The most predisposing factors were hematologic disorders (71.2%). The area under the curves (95% confidence interval) for each biomarker were 0.9 (0.85% to 97%) for lactate de hydrogenase (LDH), 0.9 (0.85% to 0.94%) for C-reactive protein, 0.8 (0.75% to 0.84%) for PCT, 0.8 (0.73% to 0.85%) for erythrocyte sedimentation rate, 0.7 (0.6% to 0.8%) for sTREM-1, and 0.5 (0.45% to 0.58%) for white blood cell count. During the study period, 27.4% patients died. The LDH and sTREM-1 levels were significant increase in died patient (p < 0.05). CONCLUSIONS: According to our data, evaluation of biomarkers along with radiologic and clinical signs and symptoms of pediatric patients can lead to proper antifungal therapy and decreased side effects, antifungal resistance, and cost. The combined measurements could be better than a single marker in the prediction of IA.

Multicenter Study of Susceptibility of Aspergillus Species Isolated from Iranian University Hospitals to Seven Antifungal Agents.

Aspergillus species are a major cause of life-threatening invasive infections and noninvasive diseases. This study seeks to investigate the frequency of Aspergillus species among Iranian patients and their susceptibility to seven antifungals. In a cross-sectional study, 233 Aspergillus isolates were collected from 11 university hospitals in Iran between 2018 and 2021. Aspergillus isolates were identified based on colony morphology, microscopic characteristics, PCR-restriction fragment length polymorphism (RFLP), and sequencing of the beta-tubulin gene. The CLSI M38-A2 reference methodology was used for antifungal susceptibility testing of amphotericin B, voriconazole, posaconazole, itraconazole, luliconazole, isavuconazole, and caspofungin. Members of Aspergillus section Flavi (117/233, 50.2%), Aspergillus section Nigri (77/233, 33.1%), Aspergillus section Fumigati (21/233, 9%), Aspergillus section Terrei (14/233, 6%), Aspergillus pseudodeflectus (2/233, 0.85%), and Aspergillus melleus (2/233, 0.85%) were isolated from the samples. The lowest 0.25 MIC90 values for all isolates tested were for luliconazole (0.016 µg/mL) and isavuconazole (0.250 µg/mL), and the highest value was observed for itraconazole (≥ 8µg/mL). The 90% minimum effective concentration (MEC90) value for caspofungin was 0.125 µg/mL. MIC90 values for voriconazole, amphotericin B, and posaconazole were 1, 2, and 2 µg/mL, respectively. The non-wild-type species were presented for amphotericin B (3%), voriconazole (1.3%), posaconazole (2.6%), luliconazole (1.3%), isavuconazole (1.7%), and caspofungin (4.7%). Positive correlations in the MIC values of azole antifungals were observed, and using one azole increases the MIC value rates of other ones. None of the species were pan-azole resistant. Species of Aspergillus section Flavi were the most common Aspergillus species isolated from Iranian samples. Luliconazole, caspofungin, and isavuconazole present the most effective antifungal agents for treatment of infection due to Aspergillus species. Susceptibility tests should be performed frequently in each region for the best management of patients. IMPORTANCE Aspergillus species are the leading cause of invasive aspergillosis in immunocompromised hosts. The susceptibility of Aspergillus species to antifungal agents might be different. Azole-resistant species have emerged worldwide. Performing susceptibility testing in each region can help in the best management of patients. Here, we show the epidemiology and distribution of Aspergillus species in Iran and their susceptibility patterns for seven antifungal agents. The significant points of the present study are that species of Aspergillus section Flavi are the most prevalent Aspergillus species isolated from 11 university hospitals. Luliconazole, caspofungin, and isavuconazole were effective antifungal agents against all Aspergillus species.

ABC typing and extracellular enzyme production of Candida albicans isolated from Candida vulvovaginitis.

BACKGROUND: Candida albicans is the most common and virulent genus Candida. Detection of virulence factors in this species plays an important role in the better understanding of pathogenesis and antifungal treatment. Molecular typing investigations are important in the epidemiological interpretation of infection. This study aimed to evaluate extracellular enzyme activity and genotyping of C. albicans species isolated from vulvovaginal samples. METHODS: One hundred and three vaginal C. albicans isolates were tested for esterase, phospholipase, proteinase, and hemolysin activities by specific media. Besides, the DNA of C. albicans isolates was extracted and amplified for ABC genotyping. RESULTS: The highest enzyme production of C. albicans isolates was for proteinase (97.1%) and esterase (95.2%), whereas 59.2% of C. albicans isolates were negative for hemolysin secretion. Genotype C (83.5%) was the most frequent genotype followed by genotype B (12.6%) and genotype A (3.9%). CONCLUSION: It is concluded that genotype C was the predominant genotype in all examined vulvovaginal C. albicans isolates. Also, there was a significant difference between enzyme production in each genotype (except for proteinase).

Impact of antifungal stewardship interventions on the susceptibility of colonized Candida species in pediatric patients with malignancy.

There is a worldwide concern regarding the antimicrobial resistance and the inappropriate use of antifungal agents, which had led to an ever-increasing antifungal resistance. This study aimed to identify the antifungal susceptibility of colonized Candida species isolated from pediatric patients with cancer and evaluate the clinical impact of antifungal stewardship (AFS) interventions on the antifungal susceptibility of colonized Candida species. Candida species colonization was evaluated among hospitalized children with cancer in a tertiary teaching hospital, Shiraz 2017-2018. Samples were collected from the mouth, nose, urine, and stool of the patients admitted to our center and cultured on sabouraud dextrose agar. The isolated yeasts identified by polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP). DNA Extracted and PCR amplification was performed using the ITS1 and ITS4 primer pairs and Msp I enzyme. The broth microdilution method was used to determine the minimum inhibitory concentrations (MICs) for amphotericin B, caspofungin, and azoles. The prevalence of Candida albicans in the present study was significantly higher than other Candida species. Candida albicans species were completely susceptible to the azoles. The susceptibility rate of C. albicans to amphotericin B and caspofungin was 93.1% and 97.1%, respectively. The fluconazole MIC values of Candida albicans decreased significantly during the post-AFS period (P < 0.001; mean difference: 72.3; 95% CI of the difference: 47.36-98.62). We found that 52.5% (53/117) of the isolated C. albicans were azole-resistant before AFS implementation, while only 1.5% (2/102) of the isolates were resistant after implementation of the AFS program (P < 0.001). C. albicans fluconazole and caspofungin resistant rate also decreased significantly (P < 0.001) after implementation of the AFS program [26 (32.9%) versus 0 (0.0%) and 11 (10.9%) versus 1 (0.9%), respectively]. Besides, fluconazole use (p < 0.05) and fluconazole expenditure reduced significantly (about one thousand US$ per year) after the AFS program. Our results confirm the positive effect of optimized antifungal usage and bedside intervention on the susceptibility of Candida species after the implementation of the AFS program. C. albicans and C. glabrata exhibited a significant increase in susceptibility after the execution of the AFS program.

Genotyping, antifungal susceptibility, enzymatic activity, and phenotypic variation in Candida albicans from esophageal candidiasis.

BACKGROUND: Esophageal candidiasis is the most frequent form of esophagitis. The pathogenicity of Candida spp. is related to a combination of microbial factors, hydrolytic enzyme secretion and phenotypic switching. This study was designed to investigate esophageal candidiasis, antifungal activity, enzymatic activity patterns, phenotyping, and genotyping profiles of Candida albicans species. METHODS: Nine hundred thirty-three visited patients were evaluated, and esophageal biopsies from patients were included in this study during 2019-2020. Direct smear, Gram staining, and culture on CHROMagar were performed for each sample. Isolated species were identified with conventional procedures and PCR-RFLP. Susceptibility to antifungals was determined according to CLSI guidelines. ABC typing, phenotype switching, hemolysin, proteinase, phospholipase, and esterase activity were also determined with the appropriate protocols. RESULTS: Twenty-three (2.5%) patients (mean age 55.2 years) were diagnosed with esophageal candidiasis. The species isolated were 19(82.6%) C. albicans, 3(13.1%) C. glabrata, and 1(4.3%) C. tropicalis. Genotype A (57.9%) was the predominant type in C. albicans isolates. 50% of C. albicans isolates exhibited a white phenotype. A high level of phospholipase (47.4%), hemolysin (68.4%), and proteinase activity (36.8%) was observed in the C. albicans isolates. Only three C. glabrata isolates displayed non-wild type susceptibility to voriconazole and itraconazole. CONCLUSION: This study shows that C. albicans are still the most frequent isolates from patients with esophageal candidiasis. The predominance of genotype A, the white phenotype, and strong hemolysin activity may indicate a high prevalence of pathogenicity in these isolates. Sensitivity to antifungal drugs was greatest for amphotericin and fluconazole.

Molecular epidemiology of zygomycosis and their related factors in tertiary referral centers in southern Iran.

INTRODUCTION: For the best management of the zygomycosis in immunocompromised patients, the present study aims to detect and identify the etiologic agents by DNA sequencing method and their related factors in clinical samples of patients. METHODOLOGY: Clinical samples from 1,058 patients admitted in 11 university hospitals in Shiraz, Southern Iran were collected between July 2015 and July 2018. All samples (bronchoalveolar lavage, sputum, blood, tissue) were examined by routine microscopic and culture tests for zygomycetes. The etiologic agents were identified by the molecular method and sequencing. RESULTS: Direct microscopic examinations or pathology smear, culture, and PCR were positive in 61 (5.8%), 15 (1.4%), and 103 (9.7%) patients, respectively. According to EORTC/MSG criteria, the rates of proven, probable, and possible zygomycosis were 59.2% (61/103), 14.6% (15/103), and 26.2% (27/103 patients), respectively. The most prevalent etiologic agents according to sequencing were Rhizopus oryzae (44 cases), Rhizopus microsporus (31 cases), Rhizopus stolonifer (15 cases). Twenty-two patients (21.4%) with positive PCR died. There were significant relations between zygomycosis and the underlying disease (p = 0.043) and prior antifungal therapy (p = 0.023). White blood cell count was in the normal range in 14.1% of patients, and the means of erythrocyte sedimentation rate (ESR) and C reactive protein (CRP) were 65 mm/hour and 57 mg/L, respectively. CONCLUSIONS: Molecular methods and sequencing may have considered as suitable tools to diagnose zygomycosis. Identification of the etiologic agents may be considered as the future antifungal therapy and management of the respective patients.

Cerebral and pulmonary aspergillosis, treatment and diagnostic challenges of mixed breakthrough invasive fungal infections: case report study.

BACKGROUND: Breakthrough invasive fungal infections (bIFIs) are an area of concern in the scarcity of new antifungals. The mixed form of bIFIs is a rare phenomenon but could be potentially a troublesome challenge when caused by azole-resistant strains or non-Aspergillus fumigatus. To raise awareness and emphasize diagnostic challenges, we present a case of mixed bIFIs in a child with acute lymphoblastic leukemia. CASE PRESENTATION: A newly diagnosed 18-month-old boy with acute lymphoblastic leukemia was complicated with prolonged severe neutropenia after induction chemotherapy. He experienced repeated episodes of fever due to extended-spectrum beta-lactamase-producing Escherichia coli bloodstream infection and pulmonary invasive fungal infection with Aspergillus fumigatus (early-type bIFIs) while receiving antifungal prophylaxis. Shortly after pulmonary involvement, his condition aggravated by abnormal focal movement, loss of consciousness and seizure. Cerebral aspergillosis with Aspergillus niger diagnosed after brain tissue biopsy. The patient finally died despite 108-day antifungal therapy. CONCLUSIONS: Mixed bIFIs is a rare condition with high morbidity and mortality in the patients receiving immunosuppressants for hematological malignancies. This case highlights the clinical importance of Aspergillus identification at the species level in invasive fungal infections with multiple site involvement in the patients on antifungal prophylaxis.

Development a hydrolysis probe-based quantitative PCR assay for the specific detection and quantification of Candida auris.

BACKGROUND AND PURPOSE: Candida auris is an emerging multidrug-resistant pathogen. The identification of this species with the conventional phenotypic or biochemical mycological methods may lead to misidentification. Molecular-based species-specific identification methods such as quantitative real-time polymerase chain reaction (qPCR) facilitate a more reliable identification of C. auris than mycological methods. Regarding this, the present study aimed to develop a hydrolysis probe-based qPCR assay for the rapid, accurate identification of C. auris. MATERIALS AND METHODS: The internal transcribed spacer 2 regions in the nuclear ribosomal DNA of C. auris and other related yeasts were assayed to find a specific PCR target for C. auris. A 123-base-pair target was selected, and primers and a probe were designed for hydrolysis probe-based real-time PCR with TaqMan chemistry. Ten-fold serial dilutions of C. auris ranging from 106 to 100 CFU/mL were prepared to establish a standard curve to quantify the yeast. RESULTS: The qPCR assay was able to identify and quantify C. auris with a detection limit of 1 C. auris CFU per reaction. Specificity was confirmed by the non-amplification of the sequences belonging to other Candida species, yeasts, molds, bacteria, or human DNAs. The standard curve of the assay showed a highly significant linearity between threshold values and dilution rates (R2=0.99; slope=-3.42). CONCLUSION: The applied qPCR assay facilitated the rapid and accurate identification and quantification of emerging opportunistic C. auris. Therefore, considering the promising test validation results, we succeeded to develop a rapid and accurate hydrolysis probe- based qPCR assay for the screening and identification of C. auris.

Investigation of the Physical, Chemical Characteristics and Microbial Contamination of the Indoor Swimming Pools

Objective: The aim of this study was to investigate the physical, chemical and microbiological contamination of indoor swimming pools. Methods: Pool water specimens were collected using a plastic polypropylene sterilized bottle. The physical and chemical qualities of the waters were analyzed in terms of temperature, turbidity, pH, and free residual chlorine, with the standard methods for the examination of water. Bacteriological (routine methods) and parasitological (molecular methods) tests were carried out on pools water. Results: The mean temperature, pH, and residual chlorine of the indoor pools were 31.2 °C, 7.6 and 1.5 mg/L, respectively. Turbidity was not observed in any of the pools. The pH and temperature values were in standard ranges in 92.3% and 15.4% of the waters of swimming pools, respectively. The prevalence rates of bacterial and amoebic contaminations of the water in the swimming pools were 53.8% and 46.2%, respectively. One pool (7.7%) was contaminated with both bacteria and amoeba. Streptococcus viridans, Staphylococcus epidermidis, Pseudomonas stutzeri, Cryptosporidium and Bacillus spp. were isolated from the pool waters. Conclusion: In this study, some microorganisms were identified from the water pools. Effective management of swimming pools and proper control of the physical, chemical and microbiological property of water pools can produce the healthy recreational activity.

Changing face of Candida colonization pattern in pediatric patients with hematological malignancy during repeated hospitalizations, results of a prospective observational study (2016-2017) in shiraz, Iran.

BACKGROUND: Surveillance of current changes in the epidemiology of Invasive Fungal Diseases (IFDs) as an important component of the antifungal stewardship programs (ASP), requires careful regular monitoring, especially in high-risk settings such as oncology centers. This study aimed to examine Candida colonization status and corresponding current changes in children with malignancy during repeated admissions and also investigate the possible epidemiological shifts after the implementation of ASP. METHODS: In this prospective observational study, all eligible patients younger than 18 years were recruited during 2016-2017 at Amir Medical Oncology Center (AMOC) in Shiraz, Iran. Totally, 136 patients were enrolled and 482 samples were collected from different sites (oral/nasal discharges, urine and stool). Weekly regular sampling was carried out during hospitalization. Candida colonization status and epidemiological changes were monitored during repeated admissions. Samples were cultivated on Sabouraud Dextrose agar medium and identified by Polymerase Chain Reaction -Restriction Fragment Length Polymorphism (PCR-RFLP). RESULTS: Estimated Candida colonization incidence was 59.9% (82/136) in our patients. Candida colonization was found to be higher in oral cavity and rectum than that in nasal cavity. Among those long-term follow ups and repetitive hospitalizations, a significant number of patients exhibited changes in their colonization patterns (37.7%). Candida colonization did not reveal any significant relationship with age, sex, oncologic diseases and degree of neutropenia. C. albicans (72.0%) was found as the most common Candida species in colonized patients, followed by C. krusei, C. kefyr, C. glabrata and C. parapsilosis. CONCLUSION: Given the high incidence of Candida infections in children with cancers, close monitoring of epidemiologic changes is essential for judicious management, based on local surveillance data and improvement of overall quality of care in high risk patients.

Antifungal effect of sesame medicinal herb on Candida Species: original study and mini-review

The aim of this study was to evaluate the antifungal susceptibility patterns of three antifungals, methanolic extracts and N-hexane oil of sesame seeds on C. albicans and C. glabrata, isolated from oral cavity of liver transplant recipients. The results were compared with other reports to develop a mini review as well. Candida species were isolated from liver transplant recipients. To evaluate the antifungal activity of sesame seed oil and methanolic extract, fluconazole, caspofungin and nystatin, the corresponding minimum inhibitory concentrations were determined by CLSI M27-A3 standard method. Minimum fungicidal concentration was also evaluated. The most prevalent species was C. albicans, followed by C. glabrata. Findings indicated sensitivity to antifungal agents and resistance to methanolic extract and N-hexane oil for all C. albicans and C. glabrata isolates. The rate of Candida colonization in the oral cavity of liver transplant recipients was high. Our results revealed that the methanolic and N-hexan extracts of sesame seeds are not effective on C. albicans and C. glabrata species, isolated from the patients. The sesame seed oil pulling and mouthwash cannot effectively cleanse and remove the Candida species in the mouth. Investigation of other medicinal plants or other parts of sesame like leaves and roots are suggested.