Detection Of Antifungal Resistance Research Articles

A study of Candida albicans from raw bovine milk: Phenotypic and molecular characterisation, evaluation of the thermotolerance and the effects on pH

Milk serves as a suitable medium for the growth of foodborne pathogens, including Candida albicans, which may possess virulence properties. Therefore, understanding the virulence characteristics and thermotolerance of C. albicans isolated from milk is important for ensuring safety and quality in the dairy industry. This study aims to identify Candida species in raw milk and detect thermotolerance and pH‐changing features of C. albicans by indicating antifungal resistance, virulence and biofilm‐forming capability via phenotypic and genotypic researching. Identification of Candida species and detection of antifungal resistance and virulence markers in C. albicans were carried out via Polymerase Chain Reaction. Thermal tolerance of C. albicans isolates was determined through microbial counts on specific parameters (63°C, 68°C and 72°C). Each experiment was replicated three times to ensure reliability. Post hoc analyses were performed to compare the observed differences. The prevalence of Candida albicans (15%), Candida parapsilosis (8%), Candida krusei (3%), Candida lusitaniae (2%) and Candida glabrata (1%) was revealed in 100 milk samples. C. albicans isolates exhibited significant virulence, antifungal resistance and biofilm‐producing properties. Low‐temperature long‐time pasteurisation (63°C for 30 min) provided a 3.18 log reduction of C. albicans from an initial 4 log population, while high‐temperature short‐time pasteurisation (72°C for 15 s) resulted in a 0.83 log reduction. Determination of the thermotolerance of C. albicans with phenotypically and genotypically determined virulence properties will guide the optimisation of the pasteurisation process to prevent economic, safety and quality concerns in dairy industry. This investigation will also provide valuable insights into the scientific understanding of the dynamics of virulent C. albicans in milk and the efficacy of heat treatments in reducing its load.

PERFIL DE RESISTÊNCIA DE AGENTES DE MICOSES OPORTUNISTAS NO BRASIL

O uso indiscriminado de antifúngicos vem contribuindo com a mudança no perfil epidemiológico de fungos resistentes, elevando a taxa de mortalidade nos pacientes acometidos. No Brasil, devido à ausência de dados oficiais, não existem relatos precisos sobre a primeira detecção de resistência a antifúngicos. Entretanto, dentre os agentes causadores de micoses oportunistas a espécie mais isolada é a Candida albicans, além de outras espécies como Cryptococcus neoformans, Sporothrix schenckii e Aspergillus spp. têm sido descritas. Este estudo buscou evidenciar o progressivo aumento dos índices de resistência de fungos oportunistas aos antifúngicos comumente utilizados e ressaltar a importância do conhecimento do perfil de sensibilidade antes do início do tratamento, pois o tratamento incorreto destas infecções agrava as falhas terapêuticas e leva à remissão das micoses. Realizou-se revisão de literatura científica com base no banco de dados Bireme, Scielo, Pubmed e Google Acadêmico, utilizando como descritores: Candida albicans, Sporothrix schenckii, Cryptococcus neoformans, Aspergillus spp., micoses oportunistas, resistência, resistência em micoses oportunistas, perfil de resistência e resistência fúngica no Brasil. Os mesmos foram pesquisados em português, inglês e espanhol. Com o crescente número de pacientes imunocomprometidos os índices de infecções oportunistas aumentaram. Patógenos fúngicos oportunistas, pertencentes a estes gêneros, mostraram-se resistentes aos poliênicos (anfotericina B), imidazóis (cetoconazol, miconazol, econazol), flucitosina, triazóis (voriconazol, fluconazol, itraconazol) e multiazóis em muitos estudos. A utilização inadequada de alguns antifúngicos vem contribuindo para o aumento de taxas de incidência de infecções oportunistas, principalmente por cepas resistentes, piorando o prognóstico destes pacientes.

Antifungal Resistance Testing and Implications for Management

Antifungal agents are the mainstay in the management of patients with invasive fungal disease. However, resistance to current antifungal agents can develop with clinical use, which may negatively impact clinical outcomes. We review the strengths and weaknesses of antifungal susceptibility testing and how the detection of resistance, either phenotypically or molecularly, correlates with clinical outcomes. Phenotypic resistance is associated with worse outcomes, although this must be taken in context with other patient factors. Newer molecular assays have been developed that have shown promising results for the detection of resistance mechanisms, including azole resistance in Aspergillus fumigatus and echinocandin resistance in different Candida species. Further work is needed to improve the clinical utility of these assays for faster turn-around-time and direct use on specimens. Detection of antifungal resistance may provide useful information for the treatment of invasive fungal disease.

Diagnosis of Antifungal Drug Resistance Mechanisms in Fungal Pathogens: Transcriptional Gene Regulation

Several fungal species can cause mild to severe diseases in humans. Antifungal strategies have been made possible by the development of several drugs with antifungal activity against these pathogenic fungi. Fungi have counteracted antifungal agents in several cases by developing resistance mechanisms. These mechanisms are based on the modifications of drug target genes and on the regulation of drug-resistance genes. Their regulation is controlled by different transcription factors, and recently several mutations in these factors were demonstrated to be responsible for the development of antifungal drug resistance. Given the availability of this genome information, the detection of antifungal resistance could be realized by specific diagnostic tools with DNA-based detection technologies. Here we review the current understanding of transcriptional regulation of drug-resistance genes from several fungal pathogens and prospective diagnostic options.

High-frequency triazole resistance found In nonculturable Aspergillus fumigatus from lungs of patients with chronic fungal disease.

Oral triazole therapy is well established for the treatment of invasive (IPA), allergic (ABPA), and chronic pulmonary (CPA) aspergillosis, and is often long-term. Triazole resistance rates are rising internationally. Microbiological diagnosis of aspergillosis is limited by poor culture yield, leading to uncertainty about the frequency of triazole resistance. Using an ultrasensitive real-time polymerase chain reaction (PCR) assay for Aspergillus spp., we assessed respiratory fungal load in bronchoalveolar lavage (BAL) and sputum specimens. In a subset of PCR-positive, culture negative samples, we further amplified the CYP51A gene to detect key single-nucleotide polymorphisms (SNPs) associated with triazole resistance. Aspergillus DNA was detected in BAL from normal volunteers (4/11, 36.4%) and patients with culture or microscopy confirmed IPA (21/22, 95%). Aspergillus DNA was detected in sputum in 15 of 19 (78.9%) and 30 of 42 (71.4%) patients with ABPA and CPA, compared with 0% and 16.7% by culture, respectively. In culture-negative, PCR-positive samples, we detected triazole-resistance mutations (L98H with tandem repeat [TR] and M220) within the drug target CYP51A in 55.1% of samples. Six of 8 (75%) of those with ABPA and 12 of 24 (50%) with CPA had resistance markers present, some without prior triazole treatment, and in most despite adequate plasma drug concentrations around the time of sampling. The very low organism burdens of fungi causing infection have previously prevented direct culture and detection of antifungal resistance in clinical samples. These findings have major implications for the sustainability of triazoles for human antifungal therapy.

Using PCR to Detect Aspergillus and Assess Resistance

Aspergillus species are associated with several clinical syndromes, including invasive pulmonary aspergillosis (IPA), chronic pulmonary aspergillosis (CPA), and allergic bronchopulmonary aspergillosis (ABPA). Triazole antifungals are recommended for these syndromes, but resistance to these agents is increasing. Currently, detection of antifungal resistance requires a positive culture. In a study partially funded by the assay manufacturer, researchers examined …

Red Regional de Laboratorios para la Vigilancia de las Infecciones Fúngicas Invasoras y Susceptibilidad a los Antifúngicos

This article describes the general objectives of the Regional Laboratory Network for Surveillance of Invasive Fungal Infections and Antifungal Susceptibility in Latin America. Formation of the Network was coordinated by the Essential Medicines, Vaccines, and Health Technologies Unit of the Pan American Health Organization, with the technical and financial support of the National Center for Microbiology of the Carlos III Health Institute (Spain), and the technical support of the Microbiology Department of the Dr. C. Malbrán National Institute on Infectious Diseases (Argentina) and the Microbiology Unit of the Parasitology Service of the Adolfo Lutz Institute (Brazil). The Network's principle objectives are epidemiological surveillance of invasive fungal infections through detection of antifungal resistance and identification of emergent, invasive fungal infections; establishment of norms and common protocols for early diagnosis of mycoses; and strengthening coordination, communications, and transference mechanisms among countries. The Network must be gradually implemented and must include staff training, a systematic process for sharing technology, evaluation of diagnostic techniques, identification of fungal species, and standardized tests for antifungal susceptibility.

High prevalence of non-albicans yeasts and detection of anti-fungal resistance in the oral flora of patients with advanced cancer.

Oral fungal infections frequently develop in individuals with advanced cancer. This study examined the oral mycological flora of 207 patients receiving palliative care for advanced malignant disease. Demographic details and a clinical history were documented from each participant. A tongue swab was collected and cultured on CHROMAgar Candida (CHROMAgar Paris, France). All yeasts were identified by germ tube test, API ID 32C profiles and, for Candida dubliniensis, by species-specific PCR. Susceptibility to fluconazole and itraconazole was determined by a broth microdilution assay according to the National Committee for Clinical Laboratory Standards (NCCLS). At time of sampling, 54 (26%) of the 207 subjects had clinical evidence of a fungal infection and yeasts were isolated from 139 (67%) individuals. In total, 194 yeasts were isolated, of which 95 (49%) were Candida albicans. There was a high prevalence of Candidia glabrata (47 isolates) of which 34 (72%) were resistant to both fluconazole and itraconazole. All nine isolates of C. dubliniensis recovered were susceptible to both azoles. No relationship was established between anti-fungal usage in the preceding three months and the presence of azole resistant yeasts. This study of patients with advanced cancer has demonstrated a high incidence of oral colonization with non-C. albicans yeasts, many of which had reduced susceptibility to fluconazole and itraconazole. The role of improved oral care regimes and novel anti-fungal drugs merits further attention, to reduce the occurrence of fungal infection in these patients.