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Aspergillus species can colonize and infect immunocompetent and immunocompromised hosts. Conventional fungal identification depends on microscopic analysis and microorganism medium growth. Other diagnostic methods, non-growth dependent, to invasive fungal infections, are the biomarkers that detect circulating polysaccharides, for example, 1-3-ß-d-Glucan and galactomannan. Both are polysaccharides present on the external layer of fungi cell wall and can be detected in clinical samples during the growth of the fungus in the patient. This study aimed to compare the galactomannan detection of Lateral Flow Assay and Enzyme Immunoassay methods in Bronchoalveolar Lavage Fluid. The galactomannan antigen in Bronchoalveolar Lavage Fluid was measured using Enzyme Immunoassay according to the manufacturer's instructions (PLATELIA ASPERGILLUS™ BioRad) and, using a Lateral Flow Assay according to the manufacturer's instructions (Galactomannan LFA IMMY©). The 71 samples were Bronchoalveolar Lavage Fluid of patients hospitalized at Unicamp Clinical Hospital between 2019 and 2021; of these samples 12/71 (16.9 %) resulted in positive Galactomannan-Lateral Flow Assay. In contrast, Galactomannan-Enzyme Immunoassay resulted as positive in 9/71 (12.6 %) samples, a difference that showed not significant statistically (p-value = 0.36) Comparing both assays' results identified 8 divergences between them, about 11 % of the total sample. The Sensitivity (73.3 %), Specificity (92.35 %), Positive Predictive Value (62.85 %) and Negative Predictive Value (95.15 %) of Lateral Flow Assay were calculated using the Galactomannan Enzyme Immunoassay as standard. The Lateral Flow Assay demonstrated good results when compared with the Enzyme Immunoassay.

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INTRODUCCIÓN: Desde el inicio de la pandemia por COVID-19 se han registrado casos de infecciones de aspergilosis pulmonar asociada a esta infección, la cual tiene características diferentes a la aspergilosis pulmonar clásica y, por lo tanto, han significado un desafío diagnóstico. OBJETIVO: Validar una reacción de polimerasa en cadena (RPC) en tiempo real (sigla en inglés RT-PCR) comercial, como herramienta diagnóstica alternativa a la técnica de galactomanano (GM) en el diagnóstico de aspergilosis pulmonar asociada a COVID-19 (sigla en inglés CAPA). PACIENTES Y MÉTODO: Se analizaron resultados de RT-PCR de Aspergillus spp y GM en lavado bronco-alveolar (LBA) de 72 pacientes, hospitalizados por COVID-19 de Clínica Dávila entre los años 2020 y 2021. De estos pacientes, 33 presentaron CAPA. RESULTADOS: La RT-PCR de Aspergillus y GM presentaron una correlación positiva (r = 0,6351, valor p < 0,0001). La técnica de RT-PCR presentó una sensibilidad (S), especificidad (E), valor predictor positivo (VPP) y valor predictor negativo (VPN) de 100, 44, 66 y 100%, respectivamente, mientras que en GM fueron de 64, 89, 84 y 73%, respectivamente para LBA. Al utilizar ambas técnicas en combinación se obtuvo una S, E, VPP y VPN de 100, 82, 88 y 100%, respectivamente. CONCLUSIÓN: Este estudio concluyó que usar una técnica de RT-PCR de Aspergillus y GM en conjunto en LBA mejoraron los parámetros de desempeño de ambas técnicas usadas de manera individual para diagnosticar CAPA. Se requieren más estudios para evaluar el desempeño de técnicas combinadas en otros tipos de aspergilosis.

BACKGROUND: Since the beginning of the COVID-19 pandemic, there have been cases of pulmonary aspergillosis infections associated with this infection, which has different characteristics from classical pulmonary aspergillosis and therefore, have been diagnostic challenges. AIM: To validate a commercial real-time PCR (RT-PCR) method as an alternative diagnostic tool to the galactomannan (GM) technique in the diagnosis of COVID-19-associated pulmonary aspergillosis (CAPA). METHODS: Results of RT-PCR of Aspergillus spp and GM in broncho-alveolar lavage (BAL) of 72 patients hospitalized for COVID-19 at Clínica Dávila between 2020 and 2021 were analyzed. Of these patients, 33 presented CAPA. RESULTS: The RT-PCR for Aspergillus and GM showed a positive correlation (r = 0.6351, p-value < 0.0001). The RT-PCR for Aspergillus technique presented a sensitivity (S), specificity (S), positive predictive value (PPV) and negative predictive value (NPV) of 100, 44, 66 and 100% respectively, while the GM technique presented 64, 89, 84 and 73%, respectively for BAL. Using both techniques in combination a S, E, PPV and NPV of 100, 82, 88 and 100% were obtained respectively. CONCLUSION: This study concluded that using RT-PCR and GM techniques in combination in BAL improved the performance parameters of both techniques from those used individually to diagnose CAPA. Further studies are required to evaluate the performance of combined techniques in other aspergillosis focus.

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BACKGROUND: A clear cutoff value of galactomannan (GM) has not been established for chronic pulmonary aspergillosis (CPA) and is frequently extrapolated from invasive pulmonary aspergillosis. We performed a systematic review and meta-analysis to evaluate the diagnostic performance of serum and bronchoalveolar lavage (BAL) GM, and to propose a cutoff. METHODS: We extracted from the studies the cutoff of serum or/and BAL GM associated with true positives, false positives, true negatives, and false negatives. We performed a multi-cutoff model and a non-parametric random effect model. We estimated the optimal cutoff and the area under the curve (AUC) for GM in serum and BAL samples. RESULTS: Nine studies from 1999 to 2021 were included. Overall, the optimal cutoff of serum GM was 0.96 with a sensitivity of 0.29 (95%CI: 0.14-0.51); specificity of 0.88 (95%CI: 0.73-0.95); and AUC of 0.529 (with a CI: [0.415-0.682] [0.307-0.713]). The AUC for the non-parametric ROC model was 0.631. For BAL GM the cutoff was 0.67 with a sensitivity of 0.68 (95%CI: 0.51-0.82), specificity of 0.84 (95%CI: 0.70-0.92), and AUC of 0.814 (with a CI: [0.696-0.895] [0.733-0.881]). The AUC for the non-parametric model was 0.789. CONCLUSION: The diagnosis of CPA requires the assessment of a combination of mycological and serological factors, as no single serum and/or BAL GM antigen test is adequate. BAL GM performed better than serum, with better sensitivity and excellent accuracy.

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Aiming of self-sustainable production, the search for biodegradable and biocompatible materials has brought with it the need to know the physicochemical and dielectric characteristics of polysaccharide-based composite structures, which can be used as important and promising raw materials for biotechnology and electronic industries. Galactomannans are polysaccharides, extracted from seeds and microbiological sources, consisting of mannose and galactose. In this context, this work aimed to extract, purify and characterize by XRD, FTIR and impedance spectroscopy galactomannan obtained from seeds of Adenanthera pavonina L. The purification process was made with ethyl alcohol at concentrations of 70, 80 and 90 %. Polymeric films were prepared by solvent slow evaporation at low temperatures. XRD measurements revealed that Galactomannan from Adenanthera pavonina L., after purification, has a semi-crystalline structure due to the identification of two peaks the first between 5.849° and 6.118° and the second between 20.011° and 20.247°. FTIR spectra showed the functional groups associated with monosaccharides of the galactomannan from Adenanthera pavonina L. seeds, as well as the typical polysaccharide bands and peaks, confirmed by literature data. The impedance results give an increment on the state-of-the-art of this biomaterial by showing the existence of dielectric relaxations, independent of the degree of purification, using the dielectric modulus formalism. The permittivity analysis reveals the presence of water in the structure of the film, whose dipoles contribute to the relatively high value of the dielectric constant. From the results obtained, it can be concluded that purified galactomannan has the potential for possible applications in the electronics industry as a green and eco-friendly dielectric material.

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OBJECTIVE: Our aim was to evaluate the performance of two galactomannan (GM) assays (Platelia Aspergillus EIA, Bio-Rad® , and Aspergillus GM LFA, IMMY® ) in tracheal aspirate (TA) samples of consecutive critically ill patients with COVID-19. METHODS: We included critically ill patients, performed GM-EIA and GM-Lateral Flow Assay (GM-LFA) in TA and followed them until development of COVID-19-associated pulmonary aspergillosis (CAPA) or alternate diagnosis. CAPA was defined according to the modified AspICU criteria in patients with SARS-CoV-2 infection. We estimated sensitivity, specificity, positive and negative predictive values for GM-EIA, GM-LFA, the combination of both or either positive results for GM-EIA and GM-LFA. We explored accuracy using different breakpoints, through ROC analysis and Youden index to identify the optimal cut-offs. We described antifungal treatment and 30-day mortality. RESULTS: We identified 14/144 (9.7%) patients with CAPA, mean age was 50.35 (SD 11.9), the median time from admission to CAPA was 8 days; 28.5% received tocilizumab and 30-day mortality was 57%. ROC analysis and Youden index identified 2.0 OD as the best cut-off, resulting in sensitivity and specificity of 57.1% and 81.5% for GM-EIA and 60% and 72.6% for GM-LFA, respectively. CONCLUSIONS: The diagnostic performance of GM in tracheal aspirates improved after using a cut-off of 2 OD. Although bronchoalveolar lavage testing is the ideal test, centres with limited access to bronchoscopy may consider this approach to identify or rule out CAPA.

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Abstract Even today, an effective diagnostic test for aspergillosis in penguins is unknown, being the gold standard post-mortem examinations. The fungal antigen galactomannan (GM) has been used as a biomarker of disease in humans and is detected by the Platelia Aspergillus EIA (BioRad)®, a commercial kit based on the sandwich ELISA technique. It is standardized for use in neutropenic patients, however studies have demonstrated its usefulness also possible for birds. The aim of our study was to evaluate the effectiveness of Platelia Aspergillus EIA® test (BioRad-US) in the diagnosis of aspergillosis in Magellanic penguins, determining sensitivity, specificity, and positive and negative predictive values for different cut-off points. Were included in the study, blood serum samples (n = 29) Magellanic penguins in captivity that died by aspergillosis. Detection of GM was performed following manufacturer's instructions and the GM index was obtained by dividing the average value of OD of the duplicate of the clinical sample by duplicate OD of the average value of the cut-off sample provided by the kit. Through information database results were obtained for the presence of anti-Aspergillus fumigatus antibodies detected by agar gel immunodiffusion (AGID) for all serum samples. Results were analyzed using chi-square test and Kruskal-Wallis from SPSS 20.0, IBM®. ROC curve was obtained and from this, rates of sensitivity, specificity, positive and negative predictive values were also calculated based on four different cutoff points (0.5, 1.0, 1.5 and 2.0). The serum GM index did not differ between animals of the case and control group (pkw =0.097). In determining the ROC curve for serum GM detection the value of area under the curve was 0.635. From the values ​​determined by the coordinate of the curve, four different cut points (0.5, 1.0, 1.5 and 2.0) were analyzed, resulting in sensitivity rates ranging from 86.2 to 34.5% % and specificity between 87% and 26.1%. By comparing the serum GM index in group case as the presence or absence of antibodies detected by AGID was found p=0.503. The detection of GM the Platelia Aspergillus EIA® test seems is not be useful for the diagnosis of aspergillosis in naturally infected penguins.

Resumo Ainda hoje, um teste diagnóstico eficaz para aspergilose em pinguins não é conhecido, sendo o padrão-ouro os exames post-mortem. O antígeno fúngico galactomanana (GM) tem sido utilizado como biomarcador da doença em humanos, sendo detectado pelo Platelia Aspergillus EIA (BioRad)®, um kit comercial que se baseia na técnica ELISA sanduíche. É padronizado para utilização em pacientes neutropênicos, no entanto estudos tem demonstrado sua possível utilidade também para aves.O objetivo de nosso estudo foi avaliar a eficácia do teste Platelia Aspergillus EIA® (BioRad-US) no diagnóstico da aspergilose em pinguins-de-Magalhães, determinando sensibilidade, especificidade e valores preditivos positivos e negativos em diferentes pontos de corte. Foram incluídas no estudo, amostras de soro sanguíneo (n=29) de pinguins-de-Magalhães em cativeiro que vieram a óbito por aspergilose. A detecção de GM foi realizada seguindo instruções do fabricante e o índice de GM foi obtido dividindo o valor da média da DO da duplicata da amostra clínica pelo valor da média da DO da duplicata da amostra de cut-off fornecida pelo kit. Através de informações em banco de dados foram obtidos resultados sobre a presença de anticorpos anti-Aspergillus fumigatus, detectada por Imunodifusão em gel de ágar (IDGA) em todas as amostras séricas. Os resultados foram analisados utilizando-se teste de qui-quadrado e Kruskal-Wallis a partir do programa estatístico SPSS 20.0, IBM®. Curva ROC foi obtida e a partir desta, taxas de sensibilidade, especificidade, valores preditivo positivo e negativo foram igualmente calculados considerando quatro diferentes pontos de corte (0.5, 1.0, 1.5 e 2.0). O índice de GM sérica não diferiu entre os animais do grupo caso e controle (pKW = 0.097). Na determinação da curva ROC para detecção de GM sérica o valor da área sobre a curva foi de 0.635. A partir dos valores determinados pelas coordenadas da curva, quatro diferentes pontos de corte (0.5, 1.0, 1.5 e 2.0) foram analisados, resultando em taxas de sensibilidade variando de 86.2% a 34.5%, e de especificidade entre 87% e 26.1%. Ao comparar o índice de GM sérica nos animais do grupo caso quanto a presença ou não de anticorpos detectados pela IDGA foi encontrado p=0.503. A detecção de GM pelo teste Platelia Aspergillus EIA® não parece ser útil para o diagnóstico da aspergilose em pinguins naturalmente infectados.

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BACKGROUND: Invasive pulmonary aspergillosis is a life-threatening fungal disease principally caused by the ubiquitous mould Aspergillus fumigatus. This clinical entity is a major cause of morbidity and mortality (principally, but not restricted to, immunocompromised individuals). A few recent reports suggest in vitro fungicidal activity of sertraline against Aspergillus spp., but this activity has not yet been investigated in vivo. OBJECTIVES: To evaluate the antifungal activity of sertraline in two in vivo models of aspergillosis. METHODS: The antifungal activity of sertraline as monotherapy at three different doses (3, 10 and 15 mg/kg) was evaluated in Galleria mellonella and in a murine model of invasive pulmonary aspergillosis. Therapeutic efficacy parameters determined were larval survival and health index score for G. mellonella, whereas pulmonary fungal burden, galactomannan and lung histopathology were assessed in the murine model. RESULTS: Sertraline treatments improved larval survival and health index score, especially at doses of 10 and 15 mg/kg. Moreover, 10 mg/kg sertraline was able to reduce pulmonary fungal burden with an efficacy comparable with that of 3 mg/kg amphotericin B and 10 mg/kg voriconazole. CONCLUSIONS: To the best of our knowledge, this is the first in vivo study that evaluates the antifungal activity of sertraline against A. fumigatus, showing a possible promising option for the adjuvant treatment of pulmonary aspergillosis.

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Even today, an effective diagnostic test for aspergillosis in penguins is unknown, being the gold standard post-mortem examinations. The fungal antigen galactomannan (GM) has been used as a biomarker of disease in humans and is detected by the Platelia Aspergillus EIA (BioRad)®, a commercial kit based on the sandwich ELISA technique. It is standardized for use in neutropenic patients, however studies have demonstrated its usefulness also possible for birds. The aim of our study was to evaluate the effectiveness of Platelia Aspergillus EIA® test (BioRad-US) in the diagnosis of aspergillosis in Magellanic penguins, determining sensitivity, specificity, and positive and negative predictive values for different cut-off points. Were included in the study, blood serum samples (n = 29) Magellanic penguins in captivity that died by aspergillosis. Detection of GM was performed following manufacturer's instructions and the GM index was obtained by dividing the average value of OD of the duplicate of the clinical sample by duplicate OD of the average value of the cut-off sample provided by the kit. Through information database results were obtained for the presence of anti-Aspergillus fumigatus antibodies detected by agar gel immunodiffusion (AGID) for all serum samples. Results were analyzed using chi-square test and Kruskal-Wallis from SPSS 20.0, IBM®. ROC curve was obtained and from this, rates of sensitivity, specificity, positive and negative predictive values were also calculated based on four different cutoff points (0.5, 1.0, 1.5 and 2.0). The serum GM index did not differ between animals of the case and control group (pkw =0.097). In determining the ROC curve for serum GM detection the value of area under the curve was 0.635. From the values ​​determined by the coordinate of the curve, four different cut points (0.5, 1.0, 1.5 and 2.0) were analyzed, resulting in sensitivity rates ranging from 86.2 to 34.5% % and specificity between 87% and 26.1%. By comparing the serum GM index in group case as the presence or absence of antibodies detected by AGID was found p=0.503. The detection of GM the Platelia Aspergillus EIA® test seems is not be useful for the diagnosis of aspergillosis in naturally infected penguins.

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La aspergilosis pulmonar invasora (API) es una infección causada por hongos del género Aspergillus que afecta principalmente a pacientes inmunocomprometidos y corresponde a la forma más grave de aspergilosis. Se asocia a una alta morbi-mortalidad, siendo fundamental un diagnóstico y tratamiento oportuno. Las manifestaciones clínicas son inespecíficas, por lo que un estudio adecuado es importante para el diagnóstico, principalmente en pacientes con factores de riesgo poco habituales. En la actualidad se han establecido categorías diagnósticas que consideran factores del hospedero, laboratorio micológico tradicional y biomarcadores como galactomanano. Éstos, junto a la mejor comprensión e interpretación de las imágenes tomográficas permiten ofrecer un manejo adecuado. En este artículo, se presentan dos casos clínicos de API en pacientes reumatológicos, y se discute la utilidad de los métodos diagnósticos.

Invasive pulmonary aspergillosis (IPA) is an infection caused by fungi of the genus Aspergillus that mainly affects immunocompromised patients and corresponds to the most severe form of aspergillosis. It is associated with high morbidity and mortality, and diagnosis and timely treatment are essential. Clinical manifestations are nonspecific, so an adequate study is important for diagnosis, mainly in patients with unusual risk factors. At present, diagnostic categories have been established that consider factors of the host, traditional mycological laboratory and biomarkers such as galactomannan. These, together with the better understanding and interpretation of the tomographic images, allow us to offer an adequate management. In this article, two clinical cases of API in rheumatological patients are presented, and the usefulness of the diagnostic methods is discussed.

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