Dermatophytic Biofilms: Characteristics, Significance and Treatment Approaches.

Microbes are found in the environment, possibly more often as biofilms than in planktonic forms. Biofilm formation has been described for several important fungal species. The presence of a dermatophytoma in a dermatophytic nail infection was the basis for the proposal that dermatophytes form biofilms as well. This could explain treatment failure and recurrent dermatophytic infections. Several investigators have performed in vitro and ex vivo experiments to study the formation of biofilms by dermatophytes and their properties. The nature of the biofilm structure itself contributes to fungal protection mechanisms against many harmful external agents, including antifungals. Thus, a different approach should be carried out regarding susceptibility testing and treatment. Concerning susceptibility testing, methods to evaluate either the inhibition of biofilm formation, or the ability to eradicate it, have been introduced. As for treatment, in addition to classical antifungal agents, some natural formulations, such as plant extracts or biosurfactants, and alternative approaches, such as photodynamic therapy, have been proposed. Studies that connect the results of the in vitro and ex vivo experimentation with clinical outcomes are required in order to verify the efficacy of these approaches in clinical practice.

Diagnostic Capacity for Invasive Fungal Infections in the Greek Paediatric Haematology-Oncology Units: Report from the Infection Working Group of the Hellenic Society of Paediatric Haematology-Oncology.

An audit based on a specific questionnaire was attempted, in order to investigate the mycology laboratory diagnostic capacity for invasive fungal diseases (IFDs) in Greek Paediatric Haematology-Oncology departments/units. The study provided the relevant information for the years 2019 and 2020 and included data from all units, concerning culture-based methods and direct microscopy, phenotypic and molecular identification, sensitivity testing, serology and molecular diagnosis, as well as therapeutic drug monitoring. The target was mostly to reveal the level of laboratory coverage for hospitalised paediatric patients, independently of the possibility of performing the tests in the host hospital, or otherwise to refer the specimens elsewhere. In total, the current study demonstrated that the most important facilities and services regarding the IFD diagnostics for paediatric haematology-oncology patients in Greece are available and relatively easily accessible, with a reasonable turnaround time. Acting as an initial registry for further improvements, the audit can serve as a valuable approach to the actual situation and future perspectives. A national clinical mycology network under the auspices of the relevant scientific societies will probably facilitate collaboration between all the departments (clinical and laboratory) involved in invasive fungal infections and provide an easier approach to any necessary test for any hospitalised patient.

Comparison of Four Methods for the in vitro Susceptibility Testing of Dermatophytes.

OBJECTIVES: Infections caused by dermatophytes affect a high percentage of the population. Antifungal susceptibility testing (AST) can offer useful information about the susceptibility profiles of the pathogens as well as the concomitant documentation of the appropriate treatment. However, the slow growth rate of these fungi and their poor sporulation are factors that can delay and affect the performance of the AST. The proposed methods by the CLSI or the EUCAST are both laborious for the everyday routine. There are alternative applications which propose the use of an inoculum, consisting of a conidia-mycelium mixture or even plain mycelia, as well as the use of resazurin in order to facilitate the reading. The aim of this study was to compare these approaches to the EUCAST method and evaluate their performance. METHODS: Three alternative methods were compared to the EUCAST proposed methodology for conidia forming molds. The last was defined as the reference method. The methods under evaluation were (a) a fragmented mycelia method, (b) the EUCAST method with the addition of resazurin sodium salt solution and (c) the fragmented mycelia method with the addition of resazurin sodium salt solution. Twenty-two isolates (8 Trichophyton interdigitale, 8 T. rubrum, and 6 Microsporum canis) were tested against the antifungal agents of griseofulvin, terbinafine, fluconazole, and itraconazole. RESULTS: The essential agreement between the methods was calculated in percentages and a statistical analysis of the results was performed. Data evaluation revealed sufficient overall agreement of the methods with the addition of resazurin to the initial "uncolored" methods (98.9 and 97.5% for the EUCAST and the fragmented mycelia methods, respectively). The fragmented mycelia method exhibited a relatively sufficient overall agreement in comparison to the EUCAST method (90%) and not a satisfactory correlation, probably as a result of various issues of standardization. CONCLUSION: The EUCAST method was found to be the more reliable one, whereas the addition of resazurin sodium salt solution facilitates the reading and provides a reliable and objective evaluation. The fragmented mycelia method could serve as an alternative that should be applied only in cases of poor or no sporulating dermatophytes.

Correction to: Clinical-scale production of Aspergillus-specific T cells for the treatment of invasive aspergillosis in the immunocompromised host.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Clinical-scale production of Aspergillus-specific T cells for the treatment of invasive aspergillosis in the immunocompromised host.

Invasive aspergillosis (IA) represents a leading cause of mortality in immunocompromised patients. Although adoptive immunotherapy with Aspergillus-specific T cells (Asp-STs) represents a promising therapeutic approach against IA, the complex and costly production limits its broader application. We generated Asp-STs from a single blood draw of healthy individuals or IA patients in only 10 days, by either Aspergillus fumigatus (AF) lysate or peptide stimulation of mononuclear cells. The cells were phenotypically and functionally characterized, and safety was assessed in xenografts. Healthy donor-derived and lysate- or peptide-pulsed Asp-STs presented comparable fold expansion, immunophenotype, and Th1 responses. Upon cross-stimulation, only the lysate-pulsed Asp-STs were empowered to respond to peptide stimulation, although both cell products induced hyphal damage. Importantly, Asp-STs cross-reacted with other fungal species and did not induce alloreactivity in vivo. IA patient-derived T cells displayed an anergic phenotype that prohibited sufficient expansion and yield of meaningful doses of Asp-STs for autologous immunotherapy. Using a rapid and simple process, we generated, from healthy donors but not IA patients, functionally active Asp-STs of broad specificity and at clinically relevant numbers. Such an approach may form the basis for the effective management of IA in the context of allogeneic hematopoietic cell transplantation.

Aspergillus galactomannan detection: Trichoderma as a cause of positive results.

Aspergillus galactomannan immunoassay is a main diagnostic and monitoring tool in medical mycology. However, the specificity of the method can be skewered by the presence of several other fungi. Trying to diagnose a possible fungal infection of the lower respiratory tract in a haematology patient, it appeared that the fungus Trichoderma longibrachiatum is an additional probable cause of positive galactomannan results. Although, that Trichoderma is a rare but emerging pathogen in immunocompromised patients, the above information could be a caution point in the clinical evaluation of diagnostic results.

l-selenomethionine supplementation in children and adolescents with autoimmune thyroiditis: A randomized double-blind placebo-controlled clinical trial.

WHAT IS KNOWN AND OBJECTIVE: Although a beneficial effect of selenium (Se) administration has been proposed in adults with autoimmune thyroiditis (AT), there is a paucity of similar data in children and adolescents. The purpose of the study was to investigate whether administration of a high dose of organic Se (200 µg daily as l-selenomethionine) has an effect on antithyroid antibody titres in children and adolescents with AT. METHODS: Seventy-one (71) children and adolescents, with a mean age of 11.3 ± 0.3 years (range 4.5-17.8), diagnosed with AT (antibodies against thyroid peroxidase [anti-TPO] and/or thyroglobulin [anti-Tg] ≥60 IU/mL, euthyroidism or treated hypothyroidism and goitre in thyroid gland ultrasonography) were randomized to receive 200 µg l-selenomethionine or placebo daily for 6 months. Blood samples were drawn for measurement of serum fT4, TSH, anti-TPO and anti-Tg levels, and thyroid gland ultrasonography was performed at the entry to the study and after 6 months of treatment. RESULTS AND DISCUSSION: At the end of the study, a statistically significantly higher reduction in anti-Tg levels was observed in the Se group compared to the placebo group (Δ: -70.9 ± 22.1 vs -6.7 ± 60.6 IU/mL, P = 0.021). Although anti-TPO levels were also decreased in the Se group, this change was not statistically different from that of the control group (Δ: -116.2 ± 68.4 vs +262.8 ± 255.5 IU/mL, P = 0.219). No significant difference in thyroid gland volume was observed between the two study groups (P > 0.05). WHAT IS NEW AND CONCLUSION: In this original study, organic Se supplementation appears to reduce anti-Tg levels in children and adolescents with AT.

Pulmonary co-infections by Pneumocystis jirovecii and Aspergillus fumigatus in non-HIV patients: A report of two cases and literature review.

Pneumocystis jirovecii is the causative agent of Pneumocystis pneumonia (PcP), a common and often life-threatening opportunistic infection in HIV-infected patients. However, non-HIV, immunocompromised patients are at risk of PcP as well, whereas the mortality appears to be higher among these patients. Pneumocystis co-infections with other microorganisms are less frequent and only sparse reports of combined PcP and invasive pulmonary fungal infections exist in the literature, especially in the non-HIV patients. Two cases of pulmonary co-infections by P. jirovecii and Aspergillus fumigatus are presented. Both patients were non-HIV infected, the first one was suffering from crescentic IgA nephropathy under immunosuppressive treatment and the second from resistant non-Hodgkin lymphoma under chemotherapy. Both patients were treated with intravenous trimethoprim/sulphamethoxazole (TMP/SMX) combined with voriconazole. The first patient showed gradual clinical improvement while the outcome for the second patient was unfavourable. In addition, a literature review of the previous published cases of co-infection by P. jirovecii and other fungi in non-HIV patients was performed. Our target was to provide comprehensive information on this kind of infections, highlighting the importance of clinical suspicion.