DYNAMYC MYCOLOGY

BACKGROUND: Data on incidence of ventilator-associated pneumonia (VAP) and invasive pulmonary aspergillosis in patients with severe SARS-CoV-2 infection are limited. METHODS: We conducted a monocenter retrospective study comparing the incidence of VAP and invasive aspergillosis between patients with COVID-19-related acute respiratory distress syndrome (C-ARDS) and those with non-SARS-CoV-2 viral ARDS (NC-ARDS). RESULTS: We assessed 90 C-ARDS and 82 NC-ARDS patients, who were mechanically ventilated for more than 48 h. At ICU admission, there were significantly fewer bacterial coinfections documented in C-ARDS than in NC-ARDS: 14 (16%) vs 38 (48%), p < 0.01. Conversely, significantly more patients developed at least one VAP episode in C-ARDS as compared with NC-ARDS: 58 (64%) vs. 36 (44%), p = 0.007. The probability of VAP was significantly higher in C-ARDS after adjusting on death and ventilator weaning [sub-hazard ratio = 1.72 (1.14-2.52), p < 0.01]. The incidence of multi-drug-resistant bacteria (MDR)-related VAP was significantly higher in C-ARDS than in NC-ARDS: 21 (23%) vs. 9 (11%), p = 0.03. Carbapenem was more used in C-ARDS than in NC-ARDS: 48 (53%), vs 21 (26%), p < 0.01. According to AspICU algorithm, there were fewer cases of putative aspergillosis in C-ARDS than in NC-ARDS [2 (2%) vs. 12 (15%), p = 0.003], but there was no difference in Aspergillus colonization. CONCLUSIONS: In our experience, we evidenced a higher incidence of VAP and MDR-VAP in C-ARDS than in NC-ARDS and a lower risk for invasive aspergillosis in the former group.

The invertebrate Galleria mellonella has increasingly and widely been used in the last few years to study complex host–microbe interactions. Aspergillus fumigatus is one of the most pathogenic fungi causing life-threatening diseases in humans and animals. Galleria mellonella larvae has been proven as a reliable model for the analysis of pathogenesis and virulence factors, enable to screen a large number of A. fumigatus strains. This review describes the different uses of G. mellonella to study A. fumigatus and provides a comparison of the different protocols to trace fungal pathogenicity. The review also includes a summary of the diverse mutants tested in G. mellonella, and their respective contribution to A. fumigatus virulence. Previous investigations indicated that G. mellonella should be considered as an interesting tool even though a mammalian model may be required to complete and verify initial data.

Bacteriological diagnosis is traditionally based on culture. However, this method may be limited by the difficulty of cultivating certain species or by prior exposure to antibiotics, which justifies the resort to molecular methods, such as Sanger sequencing of the 16S rRNA gene (Sanger 16S). Recently, shotgun metagenomics (SMg) has emerged as a powerful tool to identify a wide range of pathogenic microorganisms in numerous clinical contexts. In this study, we compared the performance of SMg to Sanger 16S for bacterial detection and identification. All patients' samples for which Sanger 16S was requested between November 2019 and April 2020 in our institution were prospectively included. The corresponding samples were tested with a commercial 16S semi-automated method and a semi-quantitative pan-microorganism DNA- and RNA-based SMg method. Sixty-seven samples from 64 patients were analyzed. Overall, SMg was able to identify a bacterial etiology in 46.3% of cases (31/67) vs. 38.8% (26/67) with Sanger 16S. This difference reached significance when only the results obtained at the species level were compared (28/67 vs. 13/67). This study provides one of the first evidence of a significantly better performance of SMg than Sanger 16S for bacterial detection at the species level in patients with infectious diseases for whom culture-based methods have failed. This technology has the potential to replace Sanger 16S in routine practice for infectious disease diagnosis.

Azole resistance in the fungal pathogen Aspergillus fumigatus is an emerging problem and may develop during azole therapy in humans and animals or exposure to azole fungicides in the environment. To assess the potential risk of azole-resistance emergence in avian farms where azole compounds are used for the control of avian mycoses, we conducted a drug susceptibility study including A. fumigatus isolates from birds and avian farms in France and Southern China. A total number of 175 isolates were analyzed: 57 isolates were collected in France in avian farms where chemoprophylaxis with parconazole was performed; 51 isolates were collected in southern China in avian farms where no chemoprophylaxis was performed; and 67 additional isolates came from the collection of a mycology laboratory. No resistant isolate was detected, and the distribution of minimum inhibitory concentrations was similar for isolates collected in farms with or without azole chemoprophylaxis. For 61 randomly selected isolates, the full coding sequence of the Cyp51A gene was determined to detect mutations. Nine amino acid alterations were found in the target enzyme, 3 of which were new.

Earlier studies have shown that the outer layers of the conidial and mycelial cell walls of Aspergillus fumigatus are different. In this work, we analyzed the polysaccharidome of the resting conidial cell wall and observed major differences within the mycelium cell wall. Mainly, the conidia cell wall was characterized by (i) a smaller amount of α-(1,3)-glucan and chitin; (ii) a larger amount of β-(1,3)-glucan, which was divided into alkali-insoluble and water-soluble fractions, and (iii) the existence of a specific mannan with side chains containing galactopyranose, glucose, and N-acetylglucosamine residues. An analysis of A. fumigatus cell wall gene mutants suggested that members of the fungal GH-72 transglycosylase family play a crucial role in the conidia cell wall β-(1,3)-glucan organization and that α-(1,6)-mannosyltransferases of GT-32 and GT-62 families are essential to the polymerization of the conidium-associated cell wall mannan. This specific mannan and the well-known galactomannan follow two independent biosynthetic pathways.

The mite Sarcoptes scabiei is responsible for scabies, a pruritic and contagious skin disease in humans. S. scabiei is also responsible for mange in a wide range of animal species. The treatment of S. scabiei infection is hampered by an under-effectiveness of the few available drugs. The objective of this work was to evaluate the in vitro acaricide activity of a large number of plant essential oils (EOs) against S. scabiei. EOs were selected mainly on the basis of traditional treatments for dermatological infections in Madagascar. The sarcoptes originating from a porcine animal model were tested at concentrations ranging from 10 to 0.1%. The viability of sarcoptes was assessed by stereomicroscopic observation at 5 min, 15 min, 30 min, 45 min and then every hour until 6 h after treatment. Estimates of lethal time and lethal concentration producing 50% mortality were generated using a probit analysis. The survival curves were estimated using the Kaplan Meier method. A total of 31 EOs from different plants were tested. Cinnamomum zeylanicum (cinnamom) and Ocimum sanctum (tulsi) oils were the most active for all concentrations tested. They may be included in in vivo studies, in order to further assess their potential interest as topical treatments.

Paecilomyces spp. are emerging fungal pathogens, where Paecilomyces lilacinus and Paecilomyces variotii are the most reported species. Taxonomic and phylogenetic revisions in this genus have shown that P. variotii represents a species complex, whereas P. lilacinus is related to another genus called Purpureocillium. The aims of this study were to identify clinical isolates of Paecilomyces spp. at the species level, and to determine their antifungal susceptibility profiles. 70 clinical Paecilomyces spp. isolates were identified by MALDI-TOF Mass Spectrometry (MS) and by multilocus rDNA genes sequencing including ITS and the D1/D2 genes. Among the 70 Paecilomyces spp. isolates, 28 were identified as P. lilacinum, 26 as P. variotii stricto sensu, and 16 as P. maximus. For antifungal susceptibility testing, Minimal Inhibitory Concentrations (MICs) or Minimal Effective Concentrations (MECs) were determined for 8 antifungals. All P. lilacinum isolates had high MICs and MECs of amphotericin B and echinocandins, respectively, unlike P. variotii and P. maximus. For azole drugs, MICs were molecule- and species- dependent. The differences in in vitro susceptibility to antifungals underline the importance of accurate species identification. The MALDI–TOF MS can be a good alternative in routine laboratory to ensure fast identification of Paecilomyces spp. and P. lilacinum.

Invasive fungal infections have recently been recognized by the WHO as a major health, epidemiological, and economic issue. Their high mortality rates and the emergence of drug resistance have driven the development of new molecules, including olorofim, an antifungal belonging to a new family of compounds, the orotomides. A review was conducted on the PubMed database and the ClinicalTrials.gov website to summarize the microbiological profile of olorofim and its role in the treatment of filamentous fungal infections. Twenty-four articles were included from the search and divided into two groups: an “in vitro” group focusing on minimum inhibitory concentration (MIC) results for various fungi and an “in vivo” group evaluating the pharmacokinetics (PK), pharmacodynamics (PD), efficacy, and tolerability of olorofim in animal models of fungal infection and in humans. Olorofim demonstrated in vitro and in vivo activity against numerous filamentous fungi, including azole-resistant Aspergillus fumigatus, various dermatophytes, and endemic and dimorphic fungi. in vitro results showed higher MICs for certain Fusarium species and dematiaceous fungi Alternaria alternata and Exophiala dermatitidis; further in vivo studies are needed. Published PK-PD data in humans are limited. The results of the ongoing phase III clinical trial are eagerly awaited to evaluate olorofim’s clinical impact.

Kidney transplant recipients develop atypical infections in their epidemiology, presentation and outcome. Among these, meningitis and meningoencephalitis require urgent and adapted anti-infectious therapy, but published data is scarce in KTRs. The aim of this study was to describe their epidemiology, presentation and outcome, in order to improve their diagnostic and management. We performed a retrospective, multicentric cohort study in 15 French hospitals that included all 199 cases of M/ME in KTRs between 2007 and 2018 (0.9 case per 1,000 KTRs annually). Epidemiology was different from that in the general population: 20% were due to Cryptococcus neoformans , 13.5% to varicella-zoster virus, 5.5% to Mycobacterium tuberculosis , and 4.5% to Enterobacteria (half of which produced extended spectrum beta-lactamases), and 5% were Post Transplant Lymphoproliferative Disorders. Microorganisms causing M/ME in the general population were infrequent (2%, for Streptococcus pneumoniae ) or absent ( Neisseria meningitidis ). M/ME caused by Enterobacteria, Staphylococci or filamentous fungi were associated with high and early mortality (50%–70% at 1 year). Graft survival was not associated with the etiology of M/ME, nor was impacted by immunosuppression reduction. Based on these results, we suggest international studies to adapt guidelines in order to improve the diagnosis and the probabilistic treatment of M/ME in SOTRs.

The clouded leopard (Neofelis nebulosa) is classified as vulnerable on the International Union for the Conservation of Nature Red List of Threatened Species. However, diseases affecting this species across zoo populations are not well documented. The primary objective of this retrospective study was to identify common and significant causes of morbidity and mortality in captive-bred clouded leopards from European, Asian, and Australian institutions. Medical records from 44 zoological parks that held 271 clouded leopards from 1934 to 2017 were reviewed. Major causes of mortality in the dead leopards (n = 141) were respiratory disease (17%), maternal neglect and starvation (12%), generalized infectious disease (10%), digestive disease (10%), and trauma (10%). Six animals lived more than 20 yr and two were older than 22 yr. Diseases were recorded 344 times (average of two per leopard) in 166 living leopards. The body systems most frequently affected by disease in these 166 individuals were, in order of frequency, integumentary (prevalence = 21%), digestive (21%), respiratory (16%), musculoskeletal (12%), and urinary (10%) systems. Neoplasia (7%) was less frequent, followed by cardiovascular (5%), genital (3%), and viral (3%) disorders. Extensive, self-induced alopecia on the tail and dorsum was the most frequently reported dermatological disease, which is proposed to be called the “clouded leopard alopecia syndrome.” The most common neoplasm was pheochromocytoma (1%), followed by squamous cell carcinoma of the paw pads, pleural mesothelioma and multicentric lymphomas (<1% each). Dilated cardiomyopathy (2%) was the most common cardiovascular disease. Bronchopneumonia (7%), enteritis (4%), and nephritis (4%) were the most frequently reported respiratory, digestive, and renal diseases, respectively. Diagnosed disease incidence was significantly higher in Europe. This paper reports the results of a comprehensive study of the causes of morbidity and mortality in European, Asian, and Australian clouded leopard zoo populations.

Aspergillosis is pervasive in bird populations, especially those under human care. Its management can be critically impacted by exposure to high levels of conidia and by resistance to azole drugs. The fungal contamination in the environment of a Humboldt penguin (Spheniscus humboldti) group, housed in a French zoological park next to numerous large crop fields, was assessed through three serial sessions of surface sampling in nests, in 2018-20: all isolates were counted and characterized by sequencing. When identified as Aspergillus fumigatus, they were systematically screened for resistance mutations in the cyp51A gene and tested for minimal inhibitory concentrations (MICs) determination. At the same time, the clinical incidence of aspergillosis was evaluated in the penguin population by the means of systematic necropsy and mycological investigations. A microsatellite-based analysis tracked the circulation of A. fumigatus strains. Environmental investigations highlighted the substantial increase of the fungal load during the summer season (>12-fold vs. the other timepoints) and a large overrepresentation of species belonging to the Aspergillus section Fumigati, ranging from 22.7 to 94.6% relative prevalence. Only one cryptic species was detected (A. nishimurae), and one isolate exhibited G138S resistance mutation with elevated MICs. The overall incidence of aspergillosis was measured at ∼3.4% case-years, and mostly in juveniles. The analysis of microsatellite polymorphism revealed a high level of genetic diversity among A. fumigatus clinical isolates. In contrast, one environmental strain appeared largely overrepresented during the summer sampling session. In all, the rural location of the zoo did not influence the emergence of resistant strains.

Guttural pouch mycosis (GPM) is a rare but potentially life-threatening condition in horses. GPM is caused by a fungal invasion into the mucosal lining of the guttural pouches and, frequently, the associated neurovascular structures. Although several species of fungi have been associated with this disease, Aspergillus spp. appear to be the most common isolated from the guttural pouches. However, it remains unclear which are the predisposing factors leading to the development of the infection. The objectives of the present study were to experimentally reproduce an infection by Aspergillus fumigatus and to follow the natural evolution of the mycosis. Eight guttural pouches from four horses were experimentally infected by endoscopy-guided intrapouch inoculation of A. fumigatus culture. Horses were monitored for clinical signs and development of fungal plaques through endoscopic examination. Mycotic lesions were observed in all the horses and a spontaneous regression was observed within 15-28 days. No development of clinical signs was noticed. In conclusion, we were able to induce the development of mycotic lesions and to observe a natural regression of these lesions without clinical signs.

Introduction Monitoring fungal aerocontamination is an essential measure to prevent severe invasive aspergillosis (IA) infections in hospitals. One central block among 32 blocks of Edouard Herriot Hospital (EHH) was entirely demolished in 2015, while care activities continued in surrounding blocks. The main objective was to undertake broad environmental monitoring and clinical surveillance of IA cases to document fungal dispersion during major deconstruction work and to assess clinical risk. Methods and analysis A daily environmental survey of fungal loads was conducted in eight wards located near the demolition site. Air was collected inside and outside selected wards by agar impact samplers. Daily spore concentrations were monitored continuously by volumetric samplers at a flow rate of 10 L.min -1 . Daily temperature, wind direction and speed as well as relative humidity were recorded by the French meteorological station Meteociel. Aspergillus fumigatus strains stored will be genotyped by multiple-locus, variable-number, tandem-repeat analysis. Antifungal susceptibility will be assessed by E-test strips on Roswell Park Memorial Institute medium supplemented with agar. Ascertaining the adequacy of current environmental monitoring techniques in hospital is of growing importance, considering the rising impact of fungal infections and of curative antifungal costs. The present study could improve the daily management of IA risk during major deconstruction work and generate new data to ameliorate and redefine current guidelines. Ethics and dissemination This study was approved by the clinical research and ethics committees of EHH.

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In vitro susceptibility of 933 Candida isolates, from 16 French hospitals, to micafungin was determined using the Etest in each center. All isolates were then sent to a single center for determination of MICs by the EUCAST reference method. Overall essential agreement between the two tests was 98.5% at ±2 log2 dilutions and 90.2% at ±1 log2 dilutions. Categorical agreement was 98.2%. The Etest is a valuable alternative to EUCAST for the routine determination of micafungin MICs in medical mycology laboratories.

Aspergillosis is a fungal infection caused mainly by Aspergillus fumigatus that often results in respiratory disease in birds. Aspergillosis is a major cause of morbidity and mortality in captive-bred penguin species. Currently, there is no registered vaccine to prevent aspergillosis. Recent research demonstrated that oral administration of gram-negative bacteria expressing high levels of galactose-α-1,3-galactose (α-Gal) modulates anti-α-Gal immunity and protects turkeys from clinical aspergillosis caused by experimental A. fumigatus infection. The role of anti-α-Gal immunity in penguins has not been studied. Here, we tested the distribution of α-1,3-galactosyltransferase (α1,3GT) genes in the fecal microbiome of Humboldt penguins ( Spheniscus humboldti ). The occurrence of natural anti-α-Gal antibodies (Abs) in sera and eggs of healthy Humboldt penguins was also assessed. A trial was then conducted to test whether oral administration of Escherichia coli Nissle, expressing high α-Gal levels, modulates anti-α-Gal immunity in a colony of Humboldt penguins. Animals in the vaccination and placebo groups were evaluated before the trial and followed for one year for aspergillosis detection using a diagnostic panel including computed tomography scans, capillary zone electrophoresis, 3-hydroxybutyrate levels, and anti- A. fumigatus Abs. Anti-α-Gal Abs were detected in sera (IgM and IgY) and eggs (IgY) of healthy penguins. Microbiota analysis and functional predictions revealed the presence of α1,3GT genes in the microbiota of Humboldt penguins and other penguin species. A strong decrease in anti-α-Gal IgM levels was observed in all animals in the placebo group three months after vaccination protocol. This decrease was not observed in E. coli Nissle-treated penguins. After the vaccination protocol, we found a positive correlation between anti- E. coli IgY and anti-α-Gal IgY in the E. coli Nissle group, suggesting a correlation between the presence of the bacteria and these Abs. During the study period, three penguins exhibited respiratory signs consistent with aspergillosis. Two were from the placebo group whose symptoms resolved with specific treatments, while a single vaccinated individual developed fatal respiratory aspergillosis eight months after the trial. We conclude that E. coli Nissle represents a safe potential probiotic with a protective effect against aspergillosis in Humboldt penguins that deserves to be further explored for therapeutic uses in these animals.

In the context of the recent re-emergence of mpox worldwide, the French Society for Hospital Hygiene (SF2H) performed a literature review of the transmission paths and proposed specific recommendations for healthcare workers (HCWs) caring for patients with suspected or confirmed MPXV. In developed countries, the risk of contamination among HCWs in healthcare facilities seemed to be very low, limited to contamination through needle stick injuries. Two additional contamination cases were reported and not fully explained. Beyond healthcare settings, the analysis of the literature highlighted (i) a main contamination route during sexual intercourse, mainly among men who have sex with men, and (ii) a very low secondary attack rate in other contexts, such as schools or jails. Numerous studies have reported molecular or virus identification on surfaces or in the air surrounding patients, without any association with the low secondary case incidence; moreover, the minimum infectious dose through air or mucosal exposure is still unknown. Owing to the lack of evidence of MPXV respiratory transmission in the healthcare setting, the SF2H recommends the implementation of standard and contact precautions combined with medical/surgical mask use. Owing to the lack of evidence of transcutaneous contamination, the SF2H recommends the use of gloves only if contact with cutaneous lesions or mucous membranes occurs. Regarding the risk of contamination from the environment in healthcare facilities, additional studies must be conducted to investigate this.

In recent years, the incidence of fungal infections in humans has increased dramatically, accompanied by an expansion in the number of species implicated as etiological agents, especially environmental fungi never involved before in human infection. Among fungal pathogens, Candida species are the most common opportunistic fungi that can cause local and systemic infections, especially in immunocompromised individuals. Candida albicans (C. albicans) is the most common causative agent of mucosal and healthcare-associated systemic infections. However, during recent decades, there has been a worrying increase in the number of emerging multi-drug-resistant non-albicans Candida (NAC) species, i.e., C. glabrata, C. parapsilosis, C. tropicalis, C. krusei, C. auris, and C. ciferrii. In particular, Candida ciferrii, also known as Stephanoascus ciferrii or Trichomonascus ciferrii, is a heterothallic ascomycete yeast-like fungus that has received attention in recent decades as a cause of local and systemic fungal diseases. Today, the new definition of the S. ciferrii complex, which consists of S. ciferrii, Candida allociferrii, and Candida mucifera, was proposed after sequencing the 18S rRNA gene. Currently, the S. ciferrii complex is mostly associated with non-severe ear and eye infections, although a few cases of severe candidemia have been reported in immunocompromised individuals. Low susceptibility to currently available antifungal drugs is a rising concern, especially in NAC species. In this regard, a high rate of resistance to azoles and more recently also to echinocandins has emerged in the S. ciferrii complex. This review focuses on epidemiological, biological, and clinical aspects of the S. ciferrii complex, including its pathogenicity and drug resistance.

Abstract Objectives EUCAST changed the definition of the ‘intermediate’ (I) category in 2019, now defined as ‘susceptible, increased exposure’. This new definition could lead to an increased prescription of antibiotics still reported as ‘S’, compared with those now reported as ‘I’. The objective of this study was to evaluate the influence of this definition on the use of overly broad-spectrum antibiotics for the treatment of infections caused by WT Pseudomonas aeruginosa. Methods A retrospective observational multicentre study was conducted, involving five hospitals. Two 15 month study periods were defined, before and after the implementation of the new definition. All patients with an infection caused by WT P. aeruginosa treated by β-lactams were included. The main endpoint was the proportion of patients treated by an overly broad-spectrum antibiotic treatment by meropenem or ceftolozane/tazobactam. Results Two hundred and ninety-one patients were included. No difference between groups was found, in terms of infection, microbiology or demographic characteristics. Two overly broad-spectrum antibiotic treatments by meropenem or ceftolozane/tazobactam were observed in Period 1 (1.2%), versus 13 in Period 2 (10.8%; P &amp;lt; 0.001). No overly broad-spectrum treatment was observed when the antimicrobial stewardship team had given advice. Conclusions This new definition can cause a negative impact on the use of overly broad-spectrum antibiotic treatment due to misunderstanding by clinicians. Its successful implementation requires adaptation of software for reporting antibiotic susceptibility, a sustained strong information campaign by microbiologists and support by an antimicrobial stewardship team.

INTRODUCTION: Because patients with a "suspicion of Lyme borreliosis (LB)" may experience medical wandering and difficult care paths, often due to misinformation, multidisciplinary care centers were started all over Europe a few years ago. The aim of our study was to prospectively identify the factors associated with the acceptance of diagnosis and management satisfaction of patients, and to assess the concordance of the medical health assessment between physicians and patients 12 months after their management at our multidisciplinary center. METHODS: We included all adults who were admitted to the Tick-Borne Diseases Reference Center of Paris and the Northern Region (TBD-RC) (2017-2020). A telephone satisfaction survey was conducted 12 months after their first consultation. It consisted of 5 domains and 13 items rated between 0 (lowest) and 10 (highest grade): (1)Reception; (2)Care and quality of management; (3)Information/explanations given to the patients; (4)Current medical condition and acceptance of the final diagnosis; (5)Overall appreciation. Factors associated with diagnosis acceptance and management satisfaction at 12 months were identified using logistic regression models. The concordance of the health status as assessed by doctors and patients was calculated using a Cohen's kappa test. RESULTS: Of the 569 patients who consulted, 349 (61.3%) answered the questionnaire. Overall appreciation had a median rating of 9 [8;10] and 280/349 (80.2%) accepted their diagnoses. Patients who were "very satisfied" with their care paths at TBD-RC (OR = 4.64;CI95%[1.52-14.16]) had higher odds of diagnosis acceptance. Well-delivered information was strongly associated with better satisfaction with the management (OR = 23.39;CI95%[3.52-155.54]). The concordance between patients and physicians to assess their health status 12 months after their management at TBD-RC was almost perfect in the groups of those with confirmed and possible LB (κ = 0.99), and moderate in the group with other diagnoses (κ = 0.43). CONCLUSION: Patients seemed to approve of this multidisciplinary care organization for suspected LB. It helped them to accept their final diagnoses and enabled a high level of satisfaction with the information given by the doctors, confirming the importance of shared medical decisions, which may help to reduce health misinformation. This type of structure may be useful for any disease with a complex and controversial diagnosis.