The introduction of CFTR modulator therapies (CFTRmt) has changed cystic fibrosis (CF) management. By improving airway rheology and function in people with CF (pwCF), CFTRmt are expected to modify cyto-microbiological features. This French multicentre study aimed to assess changes in airway fungal ecology before and during the CFTRmt era. Data from pwCF followed at CF reference centres in Besançon, Bordeaux, Limoges, and Rennes were collected before CFTRmt use (2014) and after their widespread implementation (2022), including elexacaftor/tezacaftor/ivacaftor (ETI) as well as other CFTR modulator therapies used in France. Mycological outcomes included the total number of yearly cultures and the number of positive cultures per fungus and per patient, regardless of CFTRmt. A total of 1555 and 1400 sputum samples from 438 and 483 pwCF were analysed in 2014 and 2022, respectively. The 2022 population was significantly older, in agreement with French ETI-prescription limited to pwCF aged at least 12 in 2022. Regardless of year, patients with positive fungal cultures were older than those with negative ones. Positive cultures for Aspergillus section Fumigati significantly decreased under CFTRmt at both population and individual levels. Conversely, positive cultures for Aspergillus section Nigri, Penicillium sp., and Candida albicans increased under CFTRmt, in correlation with the type of CFTRmt for Aspergillus section Nigri. CFTR modulators appear to modify the airway mycobiome and fungal ecology depending on CFTRmt type. Among several factors that may account for these mycobiome changes between 2014 and 2022, environmental changes, including climate-related shifts in Aspergillus distribution, may contribute potentially.

PURPOSE OF REVIEW:Mucorales quantitative PCR (qPCR) was first developed and evaluated in 2013, and since then, substantial evidence has accumulated supporting its clinical utility. This review highlights the major advances this technique has brought to the management of mucormycosis. RECENT FINDINGS:Different in-house and commercial assays are now available, targeting a broad range of clinically relevant Mucorales species. Performance depends strongly on preanalytical parameters such as volume of plasma or serum used for DNA extraction, and ratio of DNA input in the PCR reaction mix. Multiplex assays have expanded diagnostic scope to include Aspergillus and other molds, supporting differential diagnosis of invasive mold infections. Serial qPCR monitoring provides prognostic information, with DNA clearance after treatment associated with improved survival. Routine screening in high-risk populations, including hematology and severely burned patients, shortens diagnostic delays and improves outcomes. SUMMARY:Compared with conventional methods, Mucorales qPCR provides a rapid, noninvasive, and sensitive diagnostic tool. Large studies have shown that it enables earlier detection, facilitates timely antifungal treatment, and is associated with improved survival.

ABSTRACT A murine IgG2b monoclonal antibody, named TG11, binding to an extracellular polysaccharide antigen secreted by all Mucorales fungi has been recently developed and integrated into a lateral-flow device (TG11-LFD). The aim of this study was to establish the clinical performance of TG11-LFD on bronchoalveolar lavage (BAL) fluids for the diagnosis of mucormycosis. Thirteen BAL samples from 13 patients with mucormycosis, all of which tested positive for Mucorales qPCR ( Mucor/Rhizopus [ n = 5], Lichtheimia [ n = 2], Rhizomucor [ n = 5], and Cunninghamella [ n = 1]), were used to assess the TG11-LFD. We also selected 49 BAL samples from 25 patients with other invasive fungal infections (IFI) (aspergillosis, Pneumocystis infection, candidiasis, and possible IFI) and from 20 patients without IFI for use as negative controls. The intensities of the test and control lines were recorded using a Cube reader. The diagnostic performance was assessed by analyzing the receiver operating characteristics (ROC) curve with the Jamovi software package (version 2.6.13). The area under the curve of the ROC curve was 0.739. Using a threshold value positivity ≤531 artificial units, the TG11-LFD test has a sensitivity and specificity of 76.92% and 75.51%, respectively, a positive predictive value of 45.45%, and a negative predictive value of 92.5%. In this study, we evaluated the performance of TG11-LFD on clinical samples for the first time and demonstrated its significant potential for enhancing the rapid detection of mucormycosis. Combining antigen detection with qPCR, as successfully applied in the diagnosis of aspergillosis, is likely to yield the most reliable diagnostic approach. IMPORTANCE Mucormycosis is a severe emerging, invasive fungal disease caused by fungi in the order Mucorales . The mortality rate remains high at approximately 50%. Rapid diagnosis and prompt initiation of targeted treatment are associated with an improved prognosis. Gold standard diagnostic procedures have poor sensitivity and long turnaround times. Mucorales polymerase chain reaction in blood and respiratory samples has improved diagnosis, but this technique is not widely available due to high costs and the need for specialist equipment. A prototype lateral-flow device (TG11-LFD) incorporating a mouse monoclonal antibody, which binds to an extracellular polysaccharide antigen specific to Mucorales fungi, has been recently developed. In this study, we evaluated for the first time the performance of the TG11-LFD test on clinical bronchoalveolar lavage fluids for diagnosing mucormycosis. With 76.92% sensitivity and 75.51% specificity, this innovative, simple, and affordable approach shows great potential for improving the rapid diagnosis of mucormycosis.

CONTEXT:Farmer's lung disease (FLD) is a form of hypersensitivity pneumonitis due to regular exposure to microbial antigens present in moldy hay. Serological diagnosis is usually based on immunoprecipitation techniques using 1 to 12 in-house antigens. To improve standardization, recombinant antigens (r-Ags) have been produced from three etiological agents involved in FLD. AIM:The aim of this study was to develop a strategy using an ELISA test for the serological diagnosis of FLD. METHODS:The ELISA strategy was developed on an initial group of patients and then validated using sera from 29 patients with FLD and 54 exposed but non-FLD patients. Using sera from the second group of patients, the ELISA was simplified by using purified antigens from Saccharopolyspora rectivirgula, Lichtheimia corymbifera, and Eurotium amstelodami, each mixed with a corresponding r-Ag. The performance of the ELISA was evaluated according to the antigens used and also associated with an immunoprecipitation technique. RESULTS:The ELISA test using three purified antigens (PAgs) mixed with 3 r-Ags showed a sensitivity of 97 % and a specificity of 83 % (AUC: 0.84). The use of ELISA as a screening technique and double diffusion (using 6 somatic antigens) as a confirmatory technique in cases of moderate or positive ELISAs showed a sensitivity of 93 % and a specificity of 84 % (AUC: 0.86). CONCLUSION:The use of a mixture of purified protein antigens and highly standardized recombinant antigens from three etiological agents involved in FLD offers an effective and innovative strategy for the serological diagnosis of FLD cases. This strategy has been successfully applied in the routine workflow of our laboratory.

Background Microsporum audouinii, an anthropophilic dermatophyte has recently reemerged in several European countries (such as France). Aim To describe the epidemiological and microbiological investigations into several reported cases of dermatophytosis to the public health authorities in May 2022 in a kindergarten in the Doubs department (eastern France). Methods All children with clinical signs underwent a clinical examination and mycological investigation of skin or scalp samples taken onsite at the kindergarten. Several diagnosis methods were used: Wood's lamp examination, classical process (direct examination associated to culture) and molecular techniques. An epidemiological survey by questionnaire was also performed to collect individual exposure data. Results Fifteen children presented lesions during the clinical examination. Therapy was prescribed for nine children based on the 2021 recommendations of the French Society of Dermatology. Microsporum canis was initially diagnosed by the classical process. This identification was not confirmed by epidemiological information (absence of direct contact with animals) and molecular techniques. Finally, Microsporum audouinii was isolated in six children (two positive Wood's lamp examinations) and one staff member. Conclusion Species identification allowed us to (1) update the therapeutic strategy (all children were treated successfully) and (2) be more vigilant regarding the risk of human-to-human transmissions. The management of intrafamilial transmissions requires collaboration with healthcare practitioners to clinically examine family members, prescribe laboratory investigations, and reiterate the hygiene measures. This study confirmed the importance of combining field epidemiological investigations and microbiological examinations when managing dermatophytosis outbreaks.

ABSTRACT The aim of this study was to identify parameters influencing DNA extraction and PCR amplification efficiencies in an attempt to standardize Mucorales qPCR. The Fungal PCR Initiative Mucorales Laboratory Working Group distributed two panels of simulated samples to 26 laboratories: Panel A (six sera spiked with Mucorales DNA and one negative control serum) and Panel B (six Mucorales DNA extracts). Panel A underwent DNA extraction in each laboratory according to the local procedure and were sent to a central laboratory for testing using three different qPCR techniques: one in-house qPCR assay and two commercial assays (MucorGenius and Fungiplex). Panel B DNA extracts were PCR amplified in each laboratory using local procedures: nine in-house qPCR assays and two commercial kits (MucorGenius and MycoGENIE). All data were compiled and anonymously analyzed at the central laboratory. For Panel A, a total of six different automated platforms and five manual extraction methods were used. Positive rates were 64%, 70%, and 89%, for the MucorGenius, Fungiplex, and the in-house qPCR assay, respectively. Using a large volume of serum for DNA extraction provided the highest analytical sensitivity (82.5% for 1 mL compared with 62.7% for smaller volumes, P < 0.01). For Panel B, five in-house qPCR assays and two commercial kits had >78% positivity. Using larger PCR input volumes (≥7 µL) was associated with the highest sensitivity at 95.5% compared to 58.3% when lower input volumes were used ( P < 0.01). Using larger sample volumes for nucleic acid extraction and DNA template volumes for PCR amplification significantly improves the performance of Mucorales qPCR when testing serum. IMPORTANCE Mucormycosis is a life-threatening mold infection affecting immunosuppressed patients but also other patients with diabetes or trauma. Better survival is linked to shorter delays in diagnosis and treatment initiation. Detection of Mucorales-free DNA in serum or plasma using quantitative PCR allows a prompt diagnosis and earlier treatment. Several techniques and protocols of quantitative Mucorales PCR are used in Europe, and improving performance remains a common objective of laboratories participating in the fungal PCR Initiative Working Group. This study, which combined results from 26 laboratories in Europe, showed that the main parameters underpinning sensitivity are the preanalytical variables (volume of serum used for DNA extraction and DNA template volume), irrespective of the extraction platforms and qPCR assay/platform.

Our objective was to determine whether the twice-weekly screening of high-risk hematology patients by Mucorales qPCR on serum affects the prognosis of mucormycosis. Results from all serum Mucorales qPCR tests performed on patients from the hematology unit from January 2017 to December 2022 were analyzed. Patients with positive results were classified as having proven, probable or 'PCR-only' mucormycosis. One-month mortality for the local cohort was compared with that of a national cohort of cases of mucormycosis collected by the French surveillance network for invasive fungal disease ('Reseau de surveillances des infections fongiques invasives en France' (RESSIF)) from 2012 to 2018. From 2017 to 2022, 7825 serum Mucorales qPCR tests were performed for patients from the hematology unit; 107 patients with at least one positive Mucorales qPCR (164 positive samples) were identified. Sixty patients (70 positive samples, median Cq = 40) had no radiological criteria for mucormycosis and were considered not to have invasive fungal disease (70/7825, 0.9% false positives). It was not possible to classify disease status for six patients (12 positive samples, median Cq = 38). Forty-one patients (82 positive samples, median Cq = 35) had a final diagnosis of mucormycosis. In comparison with the RESSIF cohort, the local cohort was independently associated with a 48% lower one-month all-cause mortality rate (age-, sex-, and primary disease-adjusted hazard ratio = 0.52; 95% confidence interval: 0.29-0.94; P 0.03). Proactive screening for invasive mold diseases in high-risk hematology patients, including twice-weekly Mucorales qPCR on serum, was associated with mucormycosis higher survival.

A commercial loop-mediated isothermal amplification (LAMP) assay is available for the detection of Pneumocytis jirovecii (Eazyplex®, Amplex diagnostics, Germany). Few centers currently use this LAMP assay in France. Recently, the commercialization of reagents used to perform the P. jirovecii immunofluorescence assay (IFA) was stopped. This study aimed to assess the position of the commercial LAMP P. jirovecii assay in the diagnostic strategy for Pneumocystis pneumonia.Over 24 months (August 1, 2021, to September 1, 2023), all bronchoalveolar lavage fluid (BALF) samples with a request for P. jirovecii detection were analyzed with the commercial Eazyplex® LAMP assay, using a Genie 2® device (Amplex, diagnostics), in parallel with the techniques used for direct examination. Specific P. jirovecii quantitative real-time PCR (qPCR) was subsequently performed.In total, 346 BALF samples were analyzed by Diff-Quik coloration, IFA, and the commercial Eazyplex® LAMP assay for initial screening.Twenty-six cases of PCP were retained based on radiological, biological and clinical criteria.Among the 26 cases of PCP, 11 BALF samples were positive using the initial screening techniques: four with the three techniques, six by IFA and Eazyplex®, and one only by IFA. The eleven BALF samples were positive with the specific P. jirovecii qPCR assay, with a mean quantification cycle (Cq) of 27 [19-32].The commercial Eazyplex® LAMP assay is able to provide a result in 25 min and its sensitivity is similar to that of BALF direct examination techniques, such as IFA, which is a technique no longer available on the European market. The sensitivity of the commercial Eazyplex® LAMP assay is however clearly inferior to that of the specific P. jirovecii qPCR assay and, therefore, cannot replace the specific qPCR, but may have a place in the diagnostic strategy.

Several lateral flow assays (LFA) capable of detecting Aspergillus fumigatus in serum and broncho-alveolar lavage fluid (BALF) within the hour, thereby potentially accelerating the screening process, are now commercially available. We prospectively compared three LFA targeting A. fumigatus on BALF collected from non-surgical intensive care patients between June 2022 and February 2023. The three LFA tested were Sõna Aspergillus galactomannan LFA (Immy), Fungadia Aspergillus antigen (Gadia), and AspLFD (OLM Diagnostics). We compared the results of these LFA with those of the galactomannan (GM) PlateliaAspergillus enzyme immunoassay (Bio-Rad), culture on Sabouraud medium and Aspergillus qPCR. We tested 97 BALF samples from 92 patients. In total 84 BALF samples tested negative with all three LFA, and four BALF samples tested positive with the AspLFD assay only (OLM). Only one BALF sample tested positive with the three LFA. In addition, three BALF samples tested positive only with the GM Platelia immunoassay. Four diagnosis of probable invasive aspergillosis were retained for the 92 patients tested. This prospective series included very few positive samples. From a practical point of view, the LFA from OLM presented the simplest protocol for use.

Rationale: Early life asthma phenotyping remains an unmet need in pediatric asthma. In France, severe pediatric asthma phenotyping has been done extensively; however, phenotypes in the general population remain under -explored. Based on the course and severity of respiratory/allergic symptoms, we aimed to identify and charac-terize early life wheeze profiles and asthma phenotypes in the general population. Methods: ELFE is a general population based birth cohort; which recruited 18,329 newborns in 2011, from 320 maternity units nationwide. Data was collected using parental responses to modified versions of ISAAC ques-tionnaire on eczema, rhinitis, food allergy, cough, wheezing, dyspnoea and sleep disturbance due to wheezing at 3 time points: post-natal (2 months), infancy (age 1) and pre-school (age 5). We built a supervised trajectory for wheeze profiles and an unsupervised approach was used for asthma phenotypes. Chi squared (chi 2) test or fisher's exact test was used as appropriate (p < 0.05). Results: Wheeze profiles and asthma phenotypes were ascertained at age 5. Supervised wheeze trajectory of 9161 children resulted in 4 wheeze profiles: Persistent (0.8%), Transient (12.1%), Incident wheezers at age 5 (13.3%) and Non wheezers (73.9%). While 9517 children in unsupervised clusters displayed 4 distinct asthma pheno-types: Mildly symptomatic (70%), Post-natal bronchiolitis with persistent rhinitis (10.2%), Severe early asthma (16.9%) and Early persistent atopy with late onset severe wheeze (2.9%). Conclusion: We successfully determined early life wheeze profiles and asthma phenotypes in the general popu-lation of France.

Cockroach allergens have a greater impact on asthma morbidity than those from dust mites, cats, and dogs. The American cockroach (Periplaneta americana) and the German cockroach (Blattella germanica) are most frequently responsible for sensitization. The worldwide prevalence of allergic sensitization has been estimated at 2 to 26 % and is influenced by unfavorable socioeconomic conditions. Exposure is generally measured by determining antigen levels in dust or through insect trapping. We developed a real-time quantitative PCR (qPCR) method to provide an objective measurement of B. germanica levels in dwellings. The specificity of the qPCR primers and TaqMan® hydrolysis probe was validated in silico with 18S rRNA sequences. No amplification was observed for other species of cockroaches, with the exception of Blattella nipponica, which is not common indoors. From 2018 to 2021, exposure to B. germanica was detected and quantified in 27 of 389 dwellings in Bourgogne-Franche-Comté (mean = 333.8; median = 9.1 and maximum = 5304 copy number equivalents) and in 236 of 3193 ELFE cohort dwellings in mainland France in 2011 (mean = 15.6; median < 1 and maximum = 1275 copy number equivalents). The distribution of dwellings testing positive for cockroaches (7 %) differed among the 12 regions of France: <1 % in two regions, between 1 and 5 % in eight regions, 16.5 % in two regions and 35 % around Paris. Exposure measurements by the EDC sampling and qPCR methods are effective ways to assess the exposure to cockroaches in dwellings. A knowledge of the level of exposure to cockroaches is particularly important for asthmatic patients, particularly those not allergic to other common antigens.

Le pronostic des mucormycoses est lié à la rapidité d'instauration du traitement antifongique spécifique. La détection d'ADN circulant par PCR quantitative (qPCR) permet de faire le diagnostic de mucormycose de façon très précoce. Notre objectif était d'analyser l'impact sur le pronostic des mucormycoses d'une stratégie de screening des patients à hauts risque par la qPCR Mucorales sur sérum. Nous avons recherché dans le système informatisé de gestion des résultats de laboratoire tous les résultats de PCR Mucorales positifs sur sérum chez les patients hospitalisés dans le service d'hématologie entre janvier 2019 et décembre 2022. Les données cliniques et radiologiques ont été collectés dans les dossiers, et les cas ont été classés comme mucormycose prouvée ou probable selon les critères EORTC/MSG, ou comme mucormycose « PCR only », tel que défini par le Réseau de Surveillance des Infections fongiques (RESSIF). Au total, 5497 qPCR Mucorales sur sérum ont été réalisées pour 611 patients (moyenne 25 tests/ patient) pendant la période d'étude. 101 résultats positifs (68 patients) ont été observés, incluant 48 résultats positifs chez 40 patients pour lesquels le diagnostic de mucormycose n'a pas été retenu. 34/40 patients ont présenté un seul résultat positif avec une très faible charge en ADN (médiane cycle seuil (Cq) = 40). Ces patients ne présentaient pas d'image radiologique en faveur d'une mucormycose, et pour tous, une vérification du résultat sur un deuxième sérum prélevé dans les 2 jours suivant a montré un résultat négatif. Pour 6/40 patients présentant 1 à 3 sérums positifs (Cq median =38), le diagnostic définitif n'a pas pu être établi – Ces patients présentaient des images non spécifiques (micronodules) et ont reçu un traitement antifongique (Isavuconazole ou posaconazole) 53 résultats positifs (mediane Cq= 35) ont été observés chez 28 patients présentant une mucormycose prouvée (4/28), probable (1/28) ou PCR only (23/28). La pathologie sous-jacente principale était la leucémie aigue myéloïde (16/28), avec une allogreffe dans 8/16 cas. Les genres impliqués étaient Mucor/Rhizopus (16/28), Rhizomucor (7/28) et Lichtheimia (5/28). 25/28 patients ont été traités par amphotericine B liposomale (n=20) ou isavuconazole (n=5). Trois patients en situation palliative n'ont pas reçu de traitement antifongique. Les taux de mortalité à 1 mois et 3 mois étaient de 26.6% (8/28) et 42.8% (12/28), significativement inférieurs à ceux observés chez les patient atteints d'hémopathie dans la cohorte RESSIF (50.5% (95/188) et 67.9% (129/190), respectivement (p < 0.05). Cette étude rétrospective montre l'intérêt de la réalisation précoce de la qPCR Mucorales sur sérum chez les patients à haut risque. De rares faux positifs sont observés (<1%). L'interprétation des résultats doit tenir compte du contexte clinique et de la charge en ADN, et une vérification sur un deuxième prélèvement permet d'augmenter la spécificité, mais cette stratégie permet d'initier le traitement antifongique spécifique plus rapidement et d'améliorer le pronostic des mucormycoses. Liens d'intérêts déclarés : LM : prise en charge de frais déplacement et inscription Congrès / Gilead, Pfizer

Candida tropicalis is the Candida species that is mostly involved in case of acute disseminated candidiasis. We report here a case with whole body dissemination (pulmonary, cutaneous, muscular, hepatic, spinal and cerebral) highlighted by impressive imagery obtained by positron emission tomography scanner in a patient treated for T cell acute lymphocytic leukemia.

L'impact de l'exposition d'intérieur à de multiples micro-organismes, de la naissance à l'enfance, sur le développement ultérieur d'asthme demeure irrésolu. Le manque d'études utilisant un échantillonnage standardisé et une analyse microbienne ciblée explique en partie cela. En 2011, la cohorte Étude longitudinale française depuis l'enfance (ELFE) a défini une procédure d'échantillonnage standardisée, utilisée pour le ciblage qPCR de 3193 logements : les dépoussiéreurs électrostatiques (EDC). La cohorte est suivie depuis 2011 et une étude cas-témoin nichée (60 enfants asthmatiques versus 60 enfants non-asthmatiques) a été conçue pour capter les profils microbiens à l’âge de 5 ans et de de les comparer avec les profils microbiens de la même population à la naissance. Les réponses des parents à l'étude internationale sur l'asthme et les allergies dans les questionnaires de l'enfance ont été utilisées. La sélection cas-témoins a été effectuée dans la population ayant un EDC exploitable à la naissance et à 5 ans. Les cas d'asthmes ont été diagnostiqués selon les critères stricts d'utilisation de médicaments par inhalateur et de crises de respiration sifflante. Les témoins étaient sains et sans symptômes de la naissance à 5 ans. Mille sept cent cinquante-trois (1753) enfants avaient un EDC à la naissance ainsi qu’à 5 ans; 52 % sont des garçons, 15 % asthmatiques et 8 % ont des animaux domestiques. Nos critères ont abouti à 402 témoins et 67 cas, qui ont été appariés individuellement selon la saison de naissance et la région d'habitation. Nous avons détecté 27 % à 88 % de moisissures, 92 % à 98 % de bactéries et 48 % d'acariens. Sur la base de six espèces fongiques, une famille et deux genres de bactéries et un acarien, nous avons réussi à établir six profils d'habitat différents à travers la France. Nous sommes actuellement en train de vérifier les profils microbiologiques des 60 cas et témoins. Les auteurs déclarent ne pas avoir de liens d'intérêts.

CONTEXT:Bird fancier's lung (BFL) is the most prevalent form of hypersensitivity pneumonitis (HP) worldwide. The current techniques used for the serological diagnosis of BFL all use crude extracts from feathers, droppings, and blooms as test antigens, which is associated with a lack of standardization and variability of the results. An antigenic protein, immunoglobulin lambda-like polypeptide-1 (IgLL1), isolated from pigeon droppings, was recently identified to be associated with BFL. We used genetic engineering to produce IgLL1 as a recombinant antigen. AIM:We aimed to prospectively validate the use of an automated ELISA based on recombinant IgLL1 protein (r-IgLL1) as the test antigen for the serological diagnosis of BFL. METHODS:Immunoprecipitation (IP) techniques (immunodiffusion (ID), immunoelectrophoresis (IEP)) and ELISA using r-IgLL1 were performed concomitantly over 10 months on 634 sera from patients with a BFL serodiagnosis request. Questionnaires were sent to obtain details on the avian exposure, clinical data, and final diagnosis. Concordance, sensitivity (Se), and specificity (Sp) of the two techniques were compared. RESULTS:In total, 72 completed questionnaires were returned with 18 cases of BFL diagnosed and 54 of non-BFL. The concordance between the ELISA and ID+IEP precipitation techniques was 71%. The combination of immunoprecipitation techniques showed a Se of 78% and a Sp of 67%. The ELISA using r-IgLL1 showed a Se of 89% and a Sp of 91%. The automated r-IgLL1 ELISA test is sufficiently efficient to be used alone for the diagnosis of patients exposed solely to Columbidae. In cases of other avian exposure, the Se and Sp of the r-IgLL1 ELISA used for screening combined with the immunodiffusion test for confirmation were 89% and 93%, respectively. CONCLUSIONS:The automated ELISA using r-IgLL1 is a promising tool for BFL serodiagnosis. Replacing immunodiffusion by the automated ELISA using r-IgLL1 as a screening technique will be the basis of our future strategy for BFL serodiagnosis.

Abstract Although a high prevalence of COVID-19-associated pulmonary aspergillosis has been reported, it is still difficult to distinguish between colonization with Aspergillus fumigatus and infection. Concomitantly, similarities between severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and hypersensitivity pneumonitis were suggested. The objective of this study was to investigate retrospectively if precipitin assays targeting A. fumigatus could have been useful in the management of SARS-CoV-2 patients hospitalized in an Intensive Care Unit (ICU) in 2020. SARS-CoV-2 ICU patients were screened for Aspergillus co-infection using biomarkers (galactomannan antigen, qPCR) and culture of respiratory samples (tracheal aspirates and bronchoalveolar lavage). For all these patients, clinical data, ICU characteristics and microbial results were collected. Electrosyneresis assays were performed using commercial A. fumigatus somatic and metabolic antigens. ELISA were performed using in-house A. fumigatus purified antigen and recombinant antigens. Our study population consisted of 65 predominantly male patients, with a median ICU stay of 22 days, and a global survival rate of 62%. Thirty-five patients had at least one positive marker for Aspergillus species detection. The number of arcs obtained by electrosyneresis using the somatic A. fumigatus antigen was significantly higher for these 35 SARS-CoV-2 ICU patients (P 0.01, Welch's t-test). Our study showed that SARS-CoV-2 ICU patients with a positive marker for Aspergillus species detection more often presented precipitins towards A. fumigatus. Serology assays could be an additional tool to assess the clinical relevance of the Aspergillus species in respiratory samples of SARS-CoV-2 ICU patients. Lay Summary This study showed retrospectively that precipitin assays, such as electrosyneresis, could be helpful to distinguish between colonization and infection with Aspergillus fumigatus during the management of severe acute respiratory syndrome Coronavirus-2 (SARS CoV-2) patients in an intensive care unit.

We present a case of probable invasive pulmonary aspergillosis due to Aspergillus flavus, in a female patient treated for an acute myeloid leukemia. Two weeks after an allogenic stem cell transplantation a probable invasive pulmonary aspergillosis was diagnosed based on thoracic imaging combined with positive galactomannan antigen and positive in-house mitochondrial Aspergillus qPCR in serum. Although an antifungal treatment was initiated, Aspergillus qPCR and galactomannan antigen remained positive in serum and worsening of the thoracic lesions was observed. The discordance between the negativity of the in-house ribosomal Aspergillus qPCR (specific to A. fumigatus) and the positivity of the in-house mitochondrial Aspergillus qPCR (targeting A. fumigatus and some other Aspergillus) allowed the suspicion of a thermophilic Aspergillus species that was not A. fumigatus. No strain was obtained in culture but the involvement of A. flavus was confirmed using a specific A. flavus qPCR. This case illustrated the usefulness of our original strategy combining two different in-house Aspergillus qPCRs, in addition to galactomannan assay, to diagnose invasive aspergillosis in hematology patients.

Background Early diagnosis and prompt initiation of specific antifungal treatment are essential for improving the prognosis of mucormycosis. We aimed to assess the performance of serum Mucorales quantitative polymerase chain reaction (qPCR) for the early diagnosis and follow-up of mucormycosis. Methods We prospectively enrolled 232 patients with suspicion of invasive mold disease, evaluated using standard imaging and mycological procedures. Thirteen additional patients with proven or probable mucormycosis were included to analyze DNA load kinetics. Serum samples were collected twice-a-week for Mucorales qPCR tests targeting the Mucorales genera Lichtheimia, Rhizomucor, and Mucor/Rhizopus. Results The sensitivity was 85.2%, specificity 89.8%, and positive and negative likelihood ratios 8.3 and 0.17, respectively in this prospective study. The first Mucorales qPCR-positive serum was observed a median of 4 days (interquartile range [IQR], 0-9) before sampling of the first mycological or histological positive specimen and a median of one day (IQR, -2 to 6) before the first imaging was performed. Negativity of Mucorales qPCR within seven days after liposomal-amphotericin B initiation was associated with an 85% lower 30-day mortality rate (adjusted hazard ratio = 0 center dot 15, 95% confidence interval [.03-.73], P = .02). Conclusions Our study argues for the inclusion of qPCR for the detection of circulating Mucorales DNA for mucormycosis diagnosis and follow-up after treatment initiation. Positive results should be added to the criteria for the consensual definitions from the European Organization for the Research and Treatment of Cancer/Mycoses Study Group Education and Research Consortium (EORTC/MSGERC), as already done for Aspergillus PCR. This prospective multicenter study confirms the satisfactory performance of serum Mucorales quantitative polymerase chain reaction (qPCR) for the diagnosis of mucormycosis and argues for addition of this new tool in clinical settings and European Organization for the Research and Treatment of Cancer/Mycoses Study Group Education and Research Consortium (EORTC/MSGERC) consensual definitions to improve the management of mucormycosis.

Azole-treated plant bulbs have already been evoked as a potential explanation of the worldwide spread of azole-resistant Aspergillus fumigatus (ARAf). We previously pointed out the presence of a high rate of ARAf (71% of A. fumigatus detected on azole-supplemented media) in flower beds containing azole-treated bulbs at the hospital's surroundings. We show here that planting organic bulbs can be a solution to reduce ARAf burden (from 71% rate to below 3%). The results suggest that replacing treated bulbs with organic bulbs may be sufficient to regain a population that is predominantly susceptible in just 1 year. LAY SUMMARY Antifungal resistance is increasingly observed in fungal pathogens. This study argues that planting organic bulbs in hospitals' outdoor surroundings could be a good alternative to continue to beautify green spaces, without the risk of dissipating antifungal-resistant fungal pathogens.