The Aspergillus galactomannan Ag VIRCLIA® Monotest and the sõna Aspergillus galactomannan lateral flow assay show comparable performance for the diagnosis of invasive aspergillosis.

BACKGROUND: Rapid galactomannan tests, such as the sõna Aspergillus GM Lateral Flow Assay (GM-LFA) and the Aspergillus Galactomannan Ag VIRCLIA® Monotest (GM-Monotest), which are suitable for the analysis of single samples, have the potential to accelerate diagnosis of invasive aspergillosis (IA). OBJECTIVES: To compare the performance of the GM-Monotest and the GM-LFA for the diagnosis of IA. PATIENTS/METHODS: Two patient cohorts were analysed: adults who had received an allogeneic haematopoietic stem-cell transplant (alloHSCT-cohort) and patients with proven/probable IA from a 5-year period (cross-sectional IA-cohort). In the alloHSCT-cohort, weekly serum samples were tested, whereas in the cross-sectional IA-cohort sera and bronchoalveolar lavage fluids were analysed. The diagnostic performance was calculated using two definitions for positivity: (1) a single positive GM result and (2) at least two positive GM results from consecutive samples. IA classification followed EORTC/MSG 2019. RESULTS: The alloHSCT-cohort included 101 patients. Four had proven/probable IA, 26 possible IA and 71 no IA. The specificity for one positive serum and two consecutively positive sera was 88.7% and 100% (GM-Monotest) and 85.9% and 98.6% (GM-LFA). Comparison of ROC curves in the alloHSCT-cohort showed no significant difference. The cross-sectional IA-cohort included 59 patients with proven/probable IA. The sensitivity for one positive sample and two consecutively positive samples was 83.1% and 55.1% (GM-Monotest) and 86.4% and 71.4% (GM-LFA). CONCLUSIONS: Both assays showed comparable diagnostic performance with a higher sensitivity for the GM-LFA if two consecutive positive samples were required for positivity. However, due to poor reproducibility, positive GM-LFA results should always be confirmed.

An immunocompetent lady with invasive aspergillosis presenting as disseminated lesions: a case report.

BACKGROUND: Invasive Aspergillosis is a fungal infection caused by Aspergillus species, typically posing life-threatening risks to immunocompromised individuals. While occurrences in immunocompetent hosts are rare, a recent case report documented fulminant pulmonary aspergillosis in an immunocompetent patient during autopsy. Here, we present a case of invasive aspergillosis in an immunocompetent woman, manifesting with disseminated lesions. CASE PRESENTATION: A 29-year-old Asian woman presented to our hospital in March 2022, reporting chest pain and shortness of breath persisting for two months. Upon examination, she appeared thin and unwell, with no notable abnormalities otherwise. Radiographic imaging revealed an ill-defined lesion in her left lung. Subsequent bronchoscopy and lavage were performed, followed by initiation of empirical antibiotic therapy. Lavage results were negative for gram staining, culture, and ZN staining for AFB, but revealed numerous septate hyphae on fungal smear. Histopathological examination indicated chronic granulomatous inflammation with septal fungal hyphae, indicative of aspergillosis. Subsequent culture confirmed Aspergillus species, prompting initiation of voriconazole therapy. Remarkably, the patient exhibited significant improvement, with weight gain and restored appetite observed within a short period. Within 2 months of treatment, her symptoms resolved, and she resumed near-normal daily activities. CONCLUSION: This case highlights the diagnosis of aspergillosis in an immunocompetent individual presenting with disseminated nodular lesions across the lungs, mediastinum, and abdomen. Clinicians should maintain a high index of suspicion for aspergillosis in cases of non-resolving pneumonia and disseminated nodular lesions, even in patients lacking traditional predisposing factors.

Performance of the IMMY® sona Aspergillus lateral flow assay for the detection of galactomannan in tracheal aspirate samples from Brazilian patients with COVID-19-associated pulmonary aspergillosis: Cross-sectional and systematic review of literature.

During the COVID-19 pandemic, many patients in intensive care units (ICUs) were affected by invasive fungal infections, including aspergillosis, contributing to a high mortality rate. Diagnosing proven COVID-19-associated pulmonary aspergillosis (CAPA) requires clinical and radiological evaluations, along with laboratory testing of bronchoalveolar lavage samples or lung biopsies. However, these procedures and equipment are often inaccessible in developing countries or regions with limited resources, including Brazil. Consequently, alternative diagnostic methods, such as measuring Aspergillus galactomannan (GM) in tracheal aspirate (TA), have been explored for CAPA diagnosis. Nonetheless, research on the efficacy of TA-based diagnostic tests is limited. This study aimed to assess the performance of the IMMY® Sona Aspergillus lateral flow assay (LFA) for GM detection in TA samples from 60 ICU patients with suspected CAPA at two tertiary hospitals in Campo Grande, Brazil. The ELISA method (Platelia Aspergillus AG, Bio-Rad®) was used to detect Aspergillus GM in TA samples, serving as the microbiological criterion and reference test. Fifteen patients (12.4%) were identified as having possible CAPA. The overall accuracy of LFA was 94%, and the tests demonstrated an agreement of 93.1% (Cohen's kappa of 0.83). Based on our findings, the LFA for Aspergillus GM detection in TA samples exhibited excellent performance, proving to be a valuable diagnostic tool for potential CAPA. In a systematic review, two studies were included, and the meta-analysis revealed pooled estimates provided a sensitivity of 86% (95% CI, 80%-91%) and specificity of 93% (95% CI, 86%-97%). The diagnostic odds ratio (DOR) for identification of Aspergillus using LFA was 103.38 (95% CI, 38.03-281.03). Despite its lower sensitivity compared to our study, the LFA appears to be a promising diagnostic option for CAPA, particularly in suspected cases that have not received antifungal therapy. This enables timely antifungal treatment and could reduce mortality rates in regions where bronchoscopy is unavailable or limited.

Aspergillosis coinfection in patients with proven mucormycosis.

Although research on aspergillosis and mucormycosis confection is important to optimize antifungal therapy, data on this issue is scarce. Thus, we systematically investigated aspergillosis coinfection in patients with proven mucormycosis. Medical records of adult patients with proven mucormycosis whose formalin-fixed paraffin-embedded (FFPE) tissue sections were available, in a tertiary hospital from August 2007 to July 2023 were retrospectively reviewed to assess coinfection with aspergillosis. We noted cultures of fungi from sterile and non-sterile sites and performed polymerase chain reaction (PCR) assays on FFPE tissues to detect Aspergillus- and Mucorales-specific DNA. Sixty-seven patients with proven mucormycosis, including 12 (18%) with a positive culture of the mucormycosis agent from sterile site cultures, were enrolled. Fungal cultures from sterile and non-sterile sites revealed Aspergillus spp. growth in nine (13%) of the 67 patients, including two sterile and seven non-sterile cultures. The fungal PCR analysis from the FFPE sections was positive for Aspergillus-specific PCR in five (7%) and positive for both Aspergillus- and Mucorales-specific PCR results in eight (12%). Overall, 21 (31%) of the 67 patients with proven mucormycosis had microbiologic and/or molecular evidence of aspergillosis coinfection. Positive blood or bronchoalveolar lavage fluid galactomannan results were more common in the coinfection group (67% [14/21]) than in the mucormycosis group (37% [17/46], P = .024). No significant difference in mortality between the two groups was observed. Approximately one-third of patients with proven mucormycosis exhibited molecular and/or microbiologic evidence of aspergillosis coinfection. Further research is needed to identify patients with aspergillosis and mucormycosis coinfections, for optimal antifungal therapy.

The study aims to investigate the coinfection between mucormycosis and aspergillosis. Key findings reveal that approximately 31% of patients demonstrated evidence of coinfection, which emphasizes the importance of considering both pathogens in diagnosis and treatment decisions.

[SWOT analysis of risk prevention and control of microorganism pollution in shopping malls and supermarkets].

OBJECTIVE: To understand the status and problems of microbial pollution in shopping malls and supermarkets in China. METHODS: Microbial pollution in shopping malls and supermarkets was assessed by literature search, key information extraction and analysis. The strengths, weaknesses, opportunities and threats(SWOT) of risk control of pathogenic microorganisms in shopping malls and supermarkets were analyzed by SWOT analysis. RESULTS: Common bacteria in the indoor air of shopping malls and supermarkets included staphylococcus and Bacillus, and common fungi include Aspergillus and Penicillium. The bacteria detected in dust samples, escalator surfaces and floor surfaces were mainly Proteobacteria and Actinomyces. The complete public places laws and regulations, standards and health supervision system were the advantages of the risk prevention and control countermeasures of microbial contamination in shopping malls and supermarkets. At the same time, it also had the disadvantages of incomplete microbial-related indexes in the premises, and insufficiently detailed countermeasures for prevention and control in the premises. There were opportunities for multi-sectoral participation and post-licensing risk prevention, and it was also facing challenges brought by many factors affecting the health microenviroment and over-disinfection. CONCLUSION: The main sites for microbial risk prevention and control in superstore-type public places included high-frequency contact areas, key public supplies and utensils, indoor air, etc. , which could be prevented and controlled through a variety of measures such as controlling the release of the source, dilution and reduction, disinfection and denaturation, etc. , and exploring a comprehensive prevention and control system that involves the autonomy of the organization, industry self-regulation, collaboration of multi-government departments, and participation of the whole society.

Comparison of galactomannan lateral flow assay and enzyme immunoassay to identify Aspergillus spp. in bronchoalveolar lavage fluid.

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.

Susceptibility Testing of Environmental and Clinical Aspergillus sydowii Demonstrates Potent Activity of Various Antifungals.

The genus Aspergillus consists of a vast number of medically and environmentally relevant species. Aspergillus species classified in series Versicolores are ubiquitous in the environment and include the opportunistic pathogen Aspergillus sydowii, which is associated with onychomycosis and superficial skin infections. Despite frequent clinical reports of A. sydowii and related series Versicolores species, antifungal susceptibility data are scarce, hampering optimal treatment choices and subsequent patient outcomes. Here, we employed antifungal susceptibility testing (AFST) based on microbroth dilution on a set of 155 series Versicolores strains using the common antifungals amphotericin B, itraconazole, voriconazole, posaconazole, isavuconazole and micafungin with the addition of luliconazole and olorofim. All strains were identified using partial calmodulin gene sequencing, with 145 being A. sydowii, seven A. creber and three A. versicolor, using the latest taxonomic insights. Overall, tested antifungals were potent against the entire strain collection. In comparison to A. fumigatus, azole and amphotericin B MICs were slightly elevated for some strains. AFST with luliconazole and olorofim, here reported for the first time, displayed the highest in vitro activity, making these antifungals interesting alternative drugs but clinical studies are warranted for future therapeutic use.

Comparative Analysis of the Clarus Aspergillus Galactomannan Enzyme Immunoassay Prototype for the Diagnosis of Invasive Pulmonary Aspergillosis in Bronchoalveolar Lavage Fluid.

BACKGROUND: Galactomannan (GM) testing using Platelia Aspergillus enzyme immunoassay (Platelia AGM) from bronchoalveolar lavage fluid (BALF) aids in early diagnosis of invasive pulmonary aspergillosis (IPA). Globally, only a minority of laboratories have the capability to perform on-site GM testing, necessitating accessible and affordable alternatives. Hence, we conducted a comparative evaluation of the new clarus Aspergillus GM enzyme immunoassay prototype (clarus AGM prototype) with Platelia AGM using BALF samples. METHODS: This is a single-center, prospective, cross-sectional study, where Platelia AGM testing was routinely performed followed by clarus AGM prototype testing in those with true positive or true negative AGM test results according to the 2020 EORTC/MSG and the 2024 FUNDICU consensus definitions. Descriptive statistics, ROC curve analysis, and Spearman's correlation analysis were used to evaluate analytical performance of the clarus AGM prototype assay. RESULTS: This study enrolled 259 adult patients, of which 53 (20%) were classified as probable IPA, while 206 did not fulfill IPA-criteria. Spearman's correlation analysis revealed a strong correlation between the two assays (rho = 0.727, p < 0.001). The clarus AGM prototype had a sensitivity of 96% (51/53) and a specificity of 74% (153/206) for differentiating probable versus no IPA when using the manufacturer recommended cut-off. ROC curve analysis showed an AUC of 0.936 (95% CI 0.901-0.971) for the clarus AGM prototype, while the Platelia AGM yielded an AUC of 0.918 (95% CI 0.876-0.959). CONCLUSIONS: Clarus AGM prototype demonstrated a strong correlation and promising test performance, comparable to Platelia AGM, rendering it a viable alternative in patients at risk of IPA.

The Utility of Real-Time PCR, Metagenomic Next-Generation Sequencing, and Culture in Bronchoalveolar Lavage Fluid for Diagnosis of Pulmonary Aspergillosis.

Timely detection of Aspergillus infection is crucial given the high mortality rate of pulmonary aspergillosis (PA). Here, the diagnostic performances for PA of mycological culture, Aspergillus real-time PCR, and metagenomic next-generation sequencing (mNGS) assay from bronchoalveolar lavage fluid, were evaluated. In total, 139 patients with suspected fungal pneumonia were enrolled between December 2021 and July 2023, collecting 139 bronchoalveolar lavage fluid samples for real-time PCR and culture, with 87 undergoing mNGS assay. The sensitivity, specificity, positive predictive value, negative predictive value, and area under the curve with 95% CIs of these assays for PA were as follows: 35.3% (14.2%-61.7%), 100.0% (94.0%-100.0%), 100.0% (54.1%-100.0%), 84.5% (79.3%-88.6%), and 0.676 (0.560-0.779), respectively, for culture; 82.4% (56.6%-96.2%), 98.3% (91.1%-100.0%), 93.3% (66.4%-99.0%), 95.2% (87.6%-98.2%), and 0.903 (0.815-0.959), respectively, for same diagnostic performance of real-time PCR and mNGS; and 94.1% (71.3%-99.9%), 96.7% (88.5%-99.6%), 88.9% (67.1%-96.9%), 98.3% (89.6%-99.7%), and 0.954 (0.880-0.989), respectively, for real-time PCR combining mNGS; real-time PCR, mNGS, and their combination significantly improved in area under the curve values over culture (P < 0.001), but real-time PCR testing and mNGS had no significant difference with each other and their combination. Overall, the performance of culture was limited by low sensitivity; both real-time PCR and mNGS assays as single diagnostic tests are promising compared with culture and combined tests.

Analysis of composition and molecular characterization of mycobiota occurring on surface of cheese ripened in Dossena's mine.

Accurate identification of the fungal community spontaneously colonizing food products, aged in natural and not controlled environments, provides information about potential mycotoxin risk associated with its consumption. Autochthonous mycobiota colonizing cheese aging in Dossena mines, was investigated and characterized by two approaches: microbial isolations and metabarcoding. Microbial isolations and metabarcoding analysis were conducted on cheese samples, obtained by four batches, produced in four different seasons of the year, aged for 90 and 180 days, by five dairy farms. The two approaches, with different taxonomical resolution power, highlighted Penicillium biforme among filamentous fungi, collected from 58 out of 68 cheeses, and Debaryomyces hansenii among yeasts, as the most abundant species (31 ÷ 65%), none representing a health risk for human cheese consumption. Shannon index showed that the richness of mycobiota increases after 180 days of maturation. Beta diversity analysis highlighted significant differences in composition of mycobiota of cheese produced by different dairy farms and aged for different durations. Weak negative growth interaction between P. biforme and Aspergillus westerdijkiae by in vitro analysis was observed leading to hypothesize that a reciprocal control is possible, also affected by natural environmental conditions, possibly disadvantageous for the last species.

Increased Dissemination of Aflatoxin- and Zearalenone-Producing Aspergillus spp. and Fusarium spp. during Wet Season via Houseflies on Dairy Farms in Aguascalientes, Mexico.

Crops contamination with aflatoxins (AFs) and zearalenone (ZEA) threaten human and animal health; these mycotoxins are produced by several species of Aspergillus and Fusarium. The objective was to evaluate under field conditions the influence of the wet season on the dissemination of AF- and ZEA-producing fungi via houseflies collected from dairy farms. Ten dairy farms distributed in the semi-arid Central Mexican Plateau were selected. Flies were collected in wet and dry seasons at seven points on each farm using entomological traps. Fungi were isolated from fly carcasses via direct seeding with serial dilutions and wet chamber methods. The production of AFs and ZEA from pure isolates was quantified using indirect competitive ELISA. A total of 693 Aspergillus spp. and 1274 Fusarium spp. isolates were obtained, of which 58.6% produced AFs and 50.0% produced ZEA (491 ± 122; 2521 ± 1295 µg/kg). Houseflies and both fungal genera were invariably present, but compared to the dry season, there was a higher abundance of flies as well as AF- and ZEA-producing fungi in the wet season (p < 0.001; 45.3/231 flies/trap; 8.6/29.6% contaminated flies). These results suggest that rainy-weather conditions on dairy farms increase the spread of AF- and ZEA-producing Aspergillus spp. and Fusarium spp. through houseflies and the incorporation of their mycotoxins into the food chain.

Comparative performance of the Platelia Aspergillus Antigen and Aspergillus Galactomannan antigen Virclia Monotest immunoassays in serum and lower respiratory tract specimens: a "real-life" experience.

The Platelia Aspergillus Antigen immunoassay is the "gold standard" for Aspergillus galactomannan (GLM) measurement in sera and bronchoalveolar lavage (BAL) for the diagnosis of invasive pulmonary aspergillosis (IPA). We evaluated the performance of the Aspergillus GLM antigen Virclia Monotest compared to the Platelia assay. A total of 535 specimens [320 sera, 86 bronchial aspirates (BAs), 70 BAL, and 59 tracheal aspirates (TAs)] from 177 adult patients (72 hematological, 32 Intensive Care Unit, and 73 hospitalized in other wards) were processed for GLM testing upon clinical request. One patient had proven IPA, and 11 had probable disease. After excluding indeterminate Virclia results (n = 38), 396 specimens yielded concordant results (56 positive and 340 negative) and 101 discordant results (Virclia positive/Platelia negative, n = 95). The overall agreement between immunoassays was higher for sera (κ 0.56) than for BAL (κ ≤ 0.24) or BAS and TA (κ ≤ 0.22). When considering all specimen types in combination, the overall sensitivity and specificity of the Virclia assay for the diagnosis of proven/probable IPA were 100% and 65%, respectively, and for the Platelia immunoassay, sensitivity and specificity were 91.7% and 89.4%, respectively. The correlation between index values by both immunoassays was strong for serum/BAL (ρ = 0.73; P < 0.001) and moderate for BAS/TA (Rho = 0.52; P = 0.001). The conversion of Virclia index values into the Platelia index could be derived by the formula y = (11.97 * X)/3.62 + X). Data from GLM-positive serum/BAL clinical specimens fitted the regression model optimally (R2 = 0.94), whereas that of BAS and TA data did not (R2 = 0.11). Further studies are needed to determine whether the Virclia assay may be an alternative to the Platelia assay for GLM measurement in sera and lower respiratory tract specimens.IMPORTANCEGalactomannan detection in serum or bronchoalveolar fluid specimens is pivotal for the diagnosis of invasive pulmonary aspergillosis (IPA). The Platelia Aspergillus Antigen immunoassay has become the "gold standard" for Aspergillus GLM measurement. Here, we provide data suggesting that the Virclia Monotest assay, which displays several operational advantages compared with the Platelia assay, may become an alternative to the Platelia assay, although further studies are needed to validate this assumption. We also provide a formula allowing the conversion of Virclia index values into Platelia values. The study may contribute toward positioning the Virclia assay within the diagnostic algorithm of IPA.

Endogenous Endophthalmitis Caused by Aspergillus lentulus in an Immunocompromised Patient with Lung Cancer

A 78-year-old man with a history of lung cancer, chemotherapy, radiotherapy, and coronavirus disease 2019 infection experienced visual deterioration of two-weeks' duration in his right eye. There was multifocal, yellowish-white retinitis foci, vascular engorgement, and scattered intraretinal hemorrhages extending from posterior pole to retinal periphery in the right eye, whereas the left eye was normal. Intravitreal vancomycin, ceftazidime, clindamycin, and dexamethasone were given for endogenous endophthalmitis initially. Vitreous culture confirmed the presence of Aspergillus lentulus, and he was treated with intravitreal amphotericin-B and voriconazole injections together with systemic amphotericin-B, voriconazole, posaconazole, and micafungin therapy. During follow-up, vitreoretinal surgery was performed because of rhegmatogenous retinal detachment, and he received one additional cycle of chemotherapy due to recurrence of the cancer. Although the retina was attached, enucleation was eventually required due to painful red eye. Atypical squamous cells beneath the neurosensory retina suggesting metastasis were noted on histopathological examination. Timely ocular examination is crucial for any immunocompromised patient having ocular symptoms. High level of suspicion for a fungal etiology is a must in these patients.

Mycobiota contaminating some market cake samples with reference to their toxin and enzyme.

Fungi can spoil the majority of baked products. Spoilage of cake during storage is commonly associated with fungi. Therefore, this study aimed to assess the quality of different types of cakes sold in the market. The most predominant fungal genera in the tested cake samples (14 samples) were Aspergillus spp., and Penicillium spp. On Potato Dextrose Agar (PDA), the medium fungal total count was 43.3 colonies /g. Aspergillus was the most dominant genus and was isolated from six samples of cake. Aspergillus was represented by 3 species namely, A. flavus, A. niger, and A. nidulans, represented by 13.32, 19.99, and 3.33 colonies /g respectively. On Malt Extract Agar (MEA) Medium, the fungal total count was 123.24 colonies / g. Aspergillus was the most dominant isolated genus from 11 samples of cake and was represented by 5 species, namely, A. flavus, A. niger, A. ochraceous, A. terreus, and A. versicolor (26. 65, 63.29, 3.33, 6.66, and 3.33 colonies / g , respectively). Twenty-four isolates (88.88 %) of the total tested twenty-seven filamentous fungi showed positive results for amylase production. Ten isolates (37.03%) of the total tested filamentous fungi showed positive results for lipase production, and finally eleven isolates (40.74 %) of the total fungal isolates showed positive results for protease production. Aflatoxins B1, B2, G1, G2, and ochratoxin A were not detected in fourteen collected samples of cake. In this study, clove oil was the best choice overpeppermint oil and olive oil for preventing mold development when natural agents were compared. It might be due to the presence of a varietyof bioactive chemical compounds in clove oil, whose major bioactive component is eugenol, which acts as an antifungal reagent. Therefore, freshly baked cake should be consumed within afew days to avoid individuals experiencing foodborne illnesses.

Successful endoscopic closure of alveolar-pleural fistula due to Aspergillus empyema using a ventricular septal defect occluder.

We report a patient with fever and cough for 2 months who was finally given a diagnosis of alveolar-pleural fistula due to aspergillus empyema. We successfully closed the alveolar-pleural fistula with a ventricular septal defect occluder through bronchoscopy. Endoscopic closure of an alveolar-pleural fistula with ventricular septal defect occluder is worth being explored.

Molecular and chemical evaluation of patulin production of Aspergillus and Penicillium-like species isolated from Hungarian apples.

Mycotoxins are secondary fungal metabolites harmful to humans and animals. Patulin (PAT) is a toxin found in different food products but especially in apples and their derivative products. The most common fungi producers of this compound are Aspergillus clavatus and Penicillium expansum. The production of patulin, as other mycotoxins, can be impacted by diverse phenomena such as water and nutrient availability, UV exposure, and the presence of antagonistic organisms. Consequently, gaining a comprehensive understanding of climate and environmental conditions is a crucial step in combating patulin contamination. In this study, moulds were isolated from 40 apple samples collected from seven locations across Hungary: Csenger, Damak, Pallag, Lövopetri, Nagykálló, and Újfehértó. A total of 183 moulds were morphologically identified, with 67 isolates belonging to the Alternaria, 45 to the Aspergillus, and 13 to the Penicillium groups. The location possessed a higher influence than farming method on the distribution of mould genera. Despite the requirement of higher temperature, Aspergillus species dominated only for the region of Újfehértó with approximately 50% of the isolates belonging to the genus. Four of the seven locations assessed: Csenger, Debrecen-Pallag, Nyírtass and Nagykálló, were dominated by Alternaria species. All isolates belonging to the genera Aspergillus and Penicillium were tested for the presence of the isoepoxidone dehydrogenase (idh) gene, a key player in the patulin metabolic pathway. To guarantee patulin production, this ability was confirmed with TLC assays. The only Aspergillus strain that presented a positive result was the strain Aspergillus clavatus B9/6, originated from the apple cultivar Golden Reinders grown in Debrecen-Pallag by integrated farming. Of the Penicillium isolates only one strain, B10/6, presented a band of the right size (500-600 bp) for the idh gene. Further sequencing of the ITS gene showed that this strain should be classified as Talaromyces pinophilus. The TLC tests confirmed this microorganism as the only patulin producer under the studied conditions for its cluster.

Screening of mycoflora and ochratoxin A on common culinary herbs and spices in Kenya.

The study aimed to screen fungal diversity and ochratoxin A levels on culinary spice and herb samples sold in open-air markets and supermarkets in Nairobi County, Kenya. All herbs were grown in Kenya, while locally-produced and imported spices were purchased from both types of retail outlet. The results showed a high frequency of Aspergillus and Penicillium species contaminating the samples. The isolated species included Aspergillus ochraceous, Aspergillus nomiae, Aspergillus niger, Aspergillus flavus, Aspergillus ustus, Aspergillus terrus, Aspergillus nidulans, Aspergillus clavutus, Penicillium crustosum, Penicillium expansum, Penicillium brevicompactum, Penicillium glabrum, Penicillium thomii, Penicillium citrinum, Penicillium polonicum, and Cladosporium cladosporioides. Total fungal count on spice and herb samples collected from various sources varied between 6 and 7 CFU/mL. Of imported spices, garlic had the highest fungal diversity, while cardamom had the least. For spices from both open market and supermarket outlets, cloves had the highest fungal diversity, while white pepper had the least. For the herbs sampled from the open markets, basil was the most contaminated, while sage was the least. In supermarket samples, parsley, sage, and mint had the highest fungal diversity, and bay had the least. The results indicate the contamination of spices and herbs with OTA at high concentrations. The calibration curve was saturated at 40 µg/kg; with samples of garlic, cinnamon, red chili, basil, thyme, mint, sage, and parsley having levels above this. Of the spices, imported ginger had the highest OTA levels (28.7 µg/kg), while turmeric from the open market had the least, 2.14 µg/kg. For herb samples, parsley from the open market had the highest OTA levels at 29.4 µg/kg, while marjoram from the open market had the lowest at 6.35 µg/kg. The results demonstrate the presence of mycotoxigenic fungi and OTA contamination of marketed culinary herbs and spices beyond acceptable limits. Hence, there is a need for informed and sustainable mitigation strategies aimed at reducing human exposure in Kenya to OTA mycotoxicosis through dietary intake of spices and herbs.

Performance of the clarus Aspergillus galactomannan enzyme immunoassay prototype for the diagnosis of invasive pulmonary aspergillosis in serum.

BACKGROUND: Serum galactomannan (GM) testing is essential for diagnosing invasive aspergillosis (IA), particularly in immunocompromised individuals. The global lack of on-site GM testing capacities necessitates cost-effective alternatives, such as .the clarus Aspergillus GM enzyme immunoassay prototype (clarus AGM prototype). METHODS: This single-centre, cross-sectional study compared the diagnostic performance of the clarus AGM prototype (IMMY, Norman, Oklahoma) with the serological gold standard (=Platelia AGM assay; Bio-Rad, Marnes-la-Cocquette, France). IA was classified according to modified 2020 EORTC/MSG consensus and 2024 FUNDICU criteria. In total, 300 prospectively (May-Dec 2023) and retrospectively (2012-2015) collected samples were included. RESULTS: Among 300 samples from 232 patients, 49 (16%) were classified as proven (n = 1) or probable IA (n = 48). In non-IA cases (n = 250), one patient was classified as possible IA. With the manufacturer recommended cut-off of ≥0.2, sensitivity and specificity of the clarus AGM prototype were 27% (13/49; 95% confidence interval [CI]: 15%-41%) and 99% (248/250; 95% CI: 97%-100%), respectively, while sensitivity and specificity were 78% and 79% when using the optimised Youden's cut-off of 0.0045 ODI. ROC curve analysis demonstrated an area under the curve (AUC) of 0.829 (95% CI: 0.760-0.898) for the clarus AGM prototype in distinguishing between proven/probable IA and non-IA. The AUC for the Platelia AGM was 0.951 (95% CI: 0.909-994). Spearman's correlation analysis showed a weak correlation between the two assays (0.382; p < .001). CONCLUSIONS: The weak correlation between the clarus AGM prototype and Platelia AGM highlights the need for further investigation into the clinical performance of the clarus AGM prototype, giving the different antigen epitopes addressed.

Investigation of antifungal susceptibility of Aspergillus species isolated from systemic clinical specimens by different methods.

PURPOSE: Due to the potential for Aspergillus species to cause lethal infections and the rising rates of antifungal resistance, the significance of antifungal susceptibility tests has increased. We aimed to assess the sensitivities of Aspergillus species to amphotericin B (AMB), voriconazole (VOR), itraconazole (ITZ), and caspofungin (CAS) using disk diffusion (DD) and gradient diffusion (GD) methods and compare them with broth microdilution (BMD) as the reference susceptibility method. METHODS: The study involved 62 Aspergillus fumigatus, 28 Aspergillus flavus, and 16 Aspergillus terreus isolates, totaling 106 Aspergillus isolates. BMD and DD methods were performed in accordance with CLSI M38-A2 and CLSI M51-A documents, respectively. The GD method utilized nonsupplemented Mueller Hinton agar (MHA) as the medium. RESULTS: In the BMD method, the lowest minimal inhibitory concentration (MIC)90 or minimal effective concentration (MEC)90 values were observed for VOR and CAS (0.5 µg/mL and 0.06 µg/mL, respectively). AMB and ITZ MIC90 values were both 2 µg/mL. In our comparison of the GD method with the BMD method at ±2 dilution, we observed essential agreement rates of 91.6%, 99.1%, 100%, and 38.6% for AMB, VOR, ITZ, and CAS, respectively. When comparing DD and BMD methods, we found categorical agreement rates of 65.1%, 99.1%, 77.3%, and 100% for AMB, VOR, ITZ, and CAS, respectively. For GD and BMD methods, these rates were 79.2%, 99.1%, 87.8%, and 100%. CONCLUSIONS: Given the high essential and categorical agreement rates, we posit that the GD method is a viable alternative to the BMD method for AMB, ITZ and VOR but not for CAS. In addition, the use of nonsupplemented MHA in the GD method proves advantageous due to its cost-effectiveness and widespread availability compared to other growth media.