A gain-of-function mutation in zinc cluster transcription factor Rob1 drives Candida albicans adaptive growth in the cystic fibrosis lung environment.

Candida albicans chronically colonizes the respiratory tract of patients with Cystic Fibrosis (CF). It competes with CF-associated pathogens (e.g. Pseudomonas aeruginosa) and contributes to disease severity. We hypothesize that C. albicans undergoes specific adaptation mechanisms that explain its persistence in the CF lung environment. To identify the underlying genetic and phenotypic determinants, we serially recovered 146 C. albicans clinical isolates over a period of 30 months from the sputum of 25 antifungal-naive CF patients. Multilocus sequence typing analyses revealed that most patients were individually colonized with genetically close strains, facilitating comparative analyses between serial isolates. We strikingly observed differential ability to filament and form monospecies and dual-species biofilms with P. aeruginosa among 18 serial isolates sharing the same diploid sequence type, recovered within one year from a pediatric patient. Whole genome sequencing revealed that their genomes were highly heterozygous and similar to each other, displaying a highly clonal subpopulation structure. Data mining identified 34 non-synonymous heterozygous SNPs in 19 open reading frames differentiating the hyperfilamentous and strong biofilm-former strains from the remaining isolates. Among these, we detected a glycine-to-glutamate substitution at position 299 (G299E) in the deduced amino acid sequence of the zinc cluster transcription factor ROB1 (ROB1G299E), encoding a major regulator of filamentous growth and biofilm formation. Introduction of the G299E heterozygous mutation in a co-isolated weak biofilm-former CF strain was sufficient to confer hyperfilamentous growth, increased expression of hyphal-specific genes, increased monospecies biofilm formation and increased survival in dual-species biofilms formed with P. aeruginosa, indicating that ROB1G299E is a gain-of-function mutation. Disruption of ROB1 in a hyperfilamentous isolate carrying the ROB1G299E allele abolished hyperfilamentation and biofilm formation. Our study links a single heterozygous mutation to the ability of C. albicans to better survive during the interaction with other CF-associated microbes and illuminates how adaptive traits emerge in microbial pathogens to persistently colonize and/or infect the CF-patient airways.

Deciphering Unexpected Vascular Locations of Scedosporium spp. and Lomentospora prolificans Fungal Infections, France.

Scedosporium spp. and Lomentospora prolificans are emerging non-Aspergillus filamentous fungi. The Scedosporiosis/lomentosporiosis Observational Study we previously conducted reported frequent fungal vascular involvement, including aortitis and peripheral arteritis. For this article, we reviewed 7 cases of Scedosporium spp. and L. prolificans arteritis from the Scedosporiosis/lomentosporiosis Observational Study and 13 cases from published literature. Underlying immunosuppression was reported in 70% (14/20) of case-patients, mainly those who had solid organ transplants (10/14). Osteoarticular localization of infection was observed in 50% (10/20) of cases; infections were frequently (7/10) contiguous with vascular infection sites. Scedosporium spp./Lomentospora prolificans infections were diagnosed in 9 of 20 patients ≈3 months after completing treatment for nonvascular scedosporiosis/lomentosporiosis. Aneurysms were found in 8/11 aortitis and 6/10 peripheral arteritis cases. Invasive fungal disease--related deaths were high (12/18 [67%]). The vascular tropism of Scedosporium spp. and L. prolificans indicates vascular imaging, such as computed tomography angiography, is needed to manage infections, especially for osteoarticular locations.

Deep cutaneous mycoses in kidney transplant recipients: Diagnostic and therapeutic challenges.

Deep cutaneous mycoses (DCMs) are rare infections that extend throughout the dermis and subcutis, often occurring after inoculation with pathogenic fungi. Trends toward a growing incidence have been observed that may be partially related to an increasing population of solid organ transplant patients. The aim of this study is to describe the diagnostics and the outcomes of DCM among kidney transplant recipients so as to optimize their management. We performed a retrospective review of cases of DCM occurring among kidney transplant recipients in our institution over 12 years. Twenty cases were included. Lesions were only located on the limbs and presented mainly as single (10/20, 50%) nodular lesions (15/20, 75%), with a mean size of 3 cm. Direct mycological examination was positive for 17 patients (17/20, 85%) and the cultures were consistently positive. Thirteen different fungal species were observed, including phaehyphomycetes (n = 8), hyalohyphomycetes (n = 3), dermatophytes (n = 1), and mucorale (n = 1). The (1-3) beta-D-glucan antigen (BDG) was also consistently detected in the serum (20/20, 100%). Systematic imaging did not reveal any distant infectious lesions, but locoregional extension was present in 11 patients (11/14, 79%). Nineteen patients received antifungal treatment (19/20, 95%) for a median duration of 3 months, with surgery for 10 (10/20, 50%). There is a great diversity of fungal species responsible for DCMs in kidney transplant recipients. The mycological documentation is necessary to adapt the antifungal treatment according to the sensitivity of the species. Serum BDG positivity is a potentially reliable and useful tool for diagnosis and follow-up.

Primary Invasive Cutaneous Fusariosis in Patients with STAT3 Hyper-IgE Syndrome.

Dominant negative (DN) mutations in signal transducer and activator of transcription 3 (STAT3) are known to cause hyper-IgE syndrome, a rare primary immunodeficiency. STAT3 DN patients are prone to develop fungal infections, including chronic mucocutaneous candidiasis due to impaired IL-17-mediated immunity, and pulmonary aspergillosis. Despite having preserved phagocyte functions, STAT3 DN patients present connective tissue abnormalities and a defect in the immunological skin barrier. Fusarium species are ubiquitous molds, whose potential to infect humans depends on the host's innate and cellular immune status. Our aim was to describe four STAT3 DN patients with fusariosis confined to the skin. Medical records were reviewed and summarized. Four patients, aged 4, 11, 30, and 33 years, presented with chronic skin lesions which started in the extremities. Two patients had remote lesions, and none had systemic involvement. Skin biopsies showed mycelial threads with deep inflammatory-occasionally granulomatous-infiltrates, reaching the dermis; cultures grew Fusarium solani. Response to treatment was heterogeneous, often requiring multimodal therapies, including topical antifungal preparations. In this work, we describe primary invasive cutaneous fusariosis as a syndromic entity in four STAT3 DN patients.

Invasive bone and joint infections from the French Scedosporiosis/lomentosporiosis Observational Study (SOS) cohort: no mortality with long-term antifungal treatment and surgery.

Little is known about localized osteoarticular Scedosporiosis (LOS). Most data come from case reports and small case series. Here we present an ancillary study of the nationwide French Scedosporiosis Observational Study (SOS), describing 15 consecutive cases of LOS diagnosed between January 2005 and March 2017. Adult patients diagnosed with LOS defined by osteoarticular involvement without distant foci reported in SOS were included. Fifteen LOS were analyzed. Seven patients had underlying disease. Fourteen patients had prior trauma as potential inoculation. Clinical presentation was arthritis (n = 8), osteitis (n = 5), and thoracic wall infection (n = 2). The most common clinical manifestation was pain (n = 9), followed by localized swelling (n = 7), cutaneous fistulization (n = 7), and fever (n = 5). The species involved were Scedosporium apiospermum (n = 8), S. boydii (n = 3), S. dehoogii (n = 1), and Lomentospora prolificans (n = 3). The species distribution was unremarkable except for S. boydii, which was associated with healthcare-related inoculations. Management was based on medical and surgical treatment for 13 patients. Fourteen patients received antifungal treatment for a median duration of 7 months. No patients died during follow-up. LOS exclusively occurred in the context of inoculation or systemic predisposing factors. It has a non-specific clinical presentation and is associated with an overall good clinical outcome, provided there is a prolonged course of antifungal therapy and adequate surgical management.

Localized osteoarticular scedosporiosis mostly occurs following direct inoculation. Management was most often based on voriconazole therapy and concomitant surgery. Unlike other invasive scedosporiosis, no patient died during follow-up.

Evaluation of Serum Mucorales Polymerase Chain Reaction (PCR) for the Diagnosis of Mucormycoses: The MODIMUCOR Prospective Trial.

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 = 0·15, 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.

Arthritis Caused by Nannizziopsis obscura, France.

Nannizziopsis spp., fungi responsible for emerging diseases, are rarely involved in human bone and joint infections. We present a rare case of septic arthritis with necrotizing cellulitis caused by N. obscura in a patient in France who had undergone kidney transplant. Rapid, aggressive medical and surgical management led to a favorable outcome.

Detection of circulating DNA for the diagnosis of invasive fusariosis: retrospective analysis of 15 proven cases.

Fusarium spp. are plant pathogens and opportunistic pathogens in severely immunocompromised (hematological malignancy, neutropenia, solid organ transplantation, etc.) and severely burned patients. Invasive fusariosis often disseminates and mortality remains high partly due to delayed diagnosis in the absence of a positive culture. The aim of our study is to design a quantitative PCR (qPCR) assay and evaluate the detection of Fusarium spp. DNA for early diagnosis of invasive infection. A qPCR assay was designed and optimized to identify all Fusarium species complex and secondarily evaluated on patient samples. A total of 81 blood samples from 15 patients diagnosed with proven invasive fusariosis from 9 centers in France were retrospectively tested. Circulating DNA was detected in 14 patients out of 15 (sensitivity of 93% [95% Confidence Interval (CI95), 70.1-99.7]). Detection was possible up to 18 days (median 6 days) before the diagnosis was confirmed by positive blood culture or biopsy. By comparison serum galactomannan and ß-D-glucan were positive in 7.1 and 58.3% of patients respectively. qPCR was negative for all patients with other invasive fungal diseases (IFD) tested (n = 12) and IFD-free control patients (n = 40). No cross-reactions were detected using DNA extracted from 81 other opportunistic fungi. We developed and validated a pan-Fusarium qPCR assay in serum/plasma with high sensitivity, specificity, and reproducibility that could facilitate early diagnosis and treatment monitoring of invasive fusariosis. LAY ABSTRACT: Fusariosis ranks third among invasive mould infections. It is frequently diagnosed late due to the lack of specific tools. We designed and evaluated a new qPCR assay with high sensitivity and specificity allowing detection of Fusarium DNA in serum samples up to 18 days before conventional diagnosis.

Invasive Aspergillosis Due to Aspergillus Section Usti: A Multicenter Retrospective Study.

BACKGROUND: Aspergillus spp. of section Usti (A. ustus) represent a rare cause of invasive aspergillosis (IA). This multicenter study describes the epidemiology and outcome of A. ustus infections. METHODS: Patients with A. ustus isolated from any clinical specimen were retrospectively identified in 22 hospitals from 8 countries. When available, isolates were sent for species identification (BenA/CaM sequencing) and antifungal susceptibility testing. Additional cases were identified by review of the literature. Cases were classified as proven/probable IA or no infection, according to standard international criteria. RESULTS: Clinical report forms were obtained for 90 patients, of whom 27 had proven/probable IA. An additional 45 cases were identified from literature review for a total of 72 cases of proven/probable IA. Hematopoietic cell and solid-organ transplant recipients accounted for 47% and 33% cases, respectively. Only 8% patients were neutropenic at time of diagnosis. Ongoing antimold prophylaxis was present in 47% of cases. Pulmonary IA represented 67% of cases. Primary or secondary extrapulmonary sites of infection were observed in 46% of cases, with skin being affected in 28% of cases. Multiple antifungal drugs were used (consecutively or in combination) in 67% of cases. The 24-week mortality rate was 58%. A. calidoustus was the most frequent causal agent. Minimal inhibitory concentrations encompassing 90% isolates (MIC90) were 1, 8, >16, and 4 µg/mL for amphotericin B, voriconazole, posaconazole, and isavuconazole, respectively. CONCLUSIONS: Aspergillus ustus IA mainly occurred in nonneutropenic transplant patients and was frequently associated with extrapulmonary sites of infection. Mortality rate was high and optimal antifungal therapy remains to be defined.

Regulators of commensal and pathogenic life-styles of an opportunistic fungus-Candida albicans.

The yeast Candida albicans is primarily a commensal of humans that colonizes the mucosal surfaces of the gastrointestinal and genital tracts. Yet, C. albicans can under certain circumstances undergo a shift from commensalism to pathogenicity. This transition is governed by fungal factors such as morphological transitions, environmental cues for instance relationships with gut microbiota and the host immune system. C. albicans utilizes distinct sets of regulatory programs to colonize or infect its host and to evade the host defense systems. Moreover, an orchestrated iron acquisition mechanism operates to adapt to specific niches with variable iron availability. Studies on regulatory networks and morphogenesis of these two distinct modes of C. albicans growth, suggest that both yeast and hyphal forms exist in both growth patterns and the regulatory circuits are inter-connected. Here, we summarize current knowledge about C. albicans commensal-to-pathogen shift, its regulatory elements and their contribution to human disease.

Poor outcome and high prevalence of invasive fungal infections in patients with adult T-cell leukemia/lymphoma exposed to zidovudine and interferon alfa.

Patients treated for adult T-Cell leukemia/lymphoma (ATL) have a poor prognosis and are prone to infectious complications which are poorly described. As the French reference center for ATL, we retrospectively analyzed 47 consecutive ATL (acute, n = 23; lymphoma, n = 14; chronic, n = 8; smoldering, n = 2) patients between 2006 and 2016 (median age 51 years, 96% Afro-Caribbean origin). The 3-year overall survival (OS) was 15.8%, 11.3%, and 85.7% for acute, lymphoma, and indolent (chronic and smoldering) forms respectively. Among aggressive subtypes, 20 patients received, as frontline therapy, high dose of zidovudine and interferon alfa (AZT-IFN⍺) resulting in an overall response rate (ORR) of 39% (complete response [CR] 33%) and 17 chemotherapy resulting of an ORR of 59% (CR 50%). Ninety-five infections occurred in 38 patients, most of whom had an acute subtype (n = 73/95; 77%). During their follow-up, patients receiving frontline chemotherapy or frontline AZT-IFNα developed infections in 74% (n = 14/19) and 89% (n = 24/27) of the cases respectively. Sixty-four (67%) of infections were microbiologically documented. Among them, invasive fungal infections (IFI, n = 11) included 2 Pneumocystis jirovecii pneumonia, 5 invasive aspergillosis, and 4 yeast fungemia. IFI exclusively occurred in patients with acute subtype mostly exposed to AZT-IFNα (n = 10/11) and experiencing prolonged (> 10 days) grade 4 neutropenia. Patients with aggressive subtype experiencing IFI had a lower OS than those who did not (median OS 5.4 months versus 18.4 months, p = 0.0048). ATL patients have a poor prognosis even in the modern era. Moreover, the high rate of infections impacts their management especially those exposed to AZT-IFNα.

Central nervous system candidiasis beyond neonates: Lessons from a nationwide study.

Though candidiasis is the most frequent invasive fungal infection, Candida spp. central nervous system (CNS) infections are rare but severe. To further describe clinico-patho-radiological presentations of this entity, we report a retrospective study from January 2005 to December 2018 including patients aged ≥ 28 days with proven or probable CNS candidiasis in France. Twenty-four patients were included. Seventeen patients (70%) had CNS localization secondary to disseminated candidiasis (10 with hematologic malignancies [HM]; the seven other patients had infective endocarditis [IE]). Among patients with HM, seven previously had lumbar puncture for intrathecal chemotherapy, the three others had IE. Among patients with disseminated infection, magnetic resonance imaging (MRI) evidenced meningitis (17%), micro-abscesses (58%), or vascular complications (67%). Seven patients (30%) had isolated CNS involvement related to neurosurgery (n = 2), CARD9 deficiency (n = 2), intravenous drug use, diabetes mellitus, or no identified predisposing condition (n = 1 each). All evaluated patients with isolated CNS involvement had meningitis on cerebrospinal fluid (CSF) and intracranial hypertension. For the latter patients, MRI evidenced meningitis (71%) or abscesses (57%). Among all patients, cerebrospinal fluid (CSF) culture grew Candida spp. in 31% of cases. CSF ßDGlucan or mannan Ag were positive in respectively 86% and 80% of cases. Mortality attributed to CNS candidiasis was 42%: 53% in case of disseminated infection (70% for HM) and 14% in case of localized infection. CNS candidiasis are isolated or occur during disseminated infection in patients with HM and lumbar puncture for intrathecal chemotherapy or during IE. Clinical, radiological finding and outcome highly vary according to CNS localized versus disseminated candidiasis. LAY SUMMARY: Candida is a yeast and is the most common cause of fungal infections worldwide. Candida central nervous system (CNS) infections are rare, severe, and poorly described. We report a retrospective study from January 2005 to December 2018 including patients aged ≥ 28 days with proven or probable CNS candidiasis in France. Twenty-four patients were included (14 men, median age 51 years). Seventeen patients had CNS localization secondary to disseminated candidiasis from blood to CNS (10 with hematologic malignancies [HM], the seven other patients had infective endocarditis [IE]). Seven patients had isolated CNS involvement related to neurosurgery (n = 2), CARD9 deficiency (n = 2), intravenous drug use (n = 1), diabetes mellitus (n = 1), or no identified risk factor (n = 1).During Candida CNS infections, brain lesions were meningitis abscesses or vascular complications. Cerebrospinal fluid (CSF) culture grew Candida spp. in 31% of cases. Forty-two percent of patients died from infection: 53% in case of disseminated infection (70% for HM) and 14% in case of localized infection.

Epidemiological and clinical study of microsporidiosis in French kidney transplant recipients from 2005 to 2019: TRANS-SPORE registry.

INTRODUCTION: Microsporidiosis is an emerging opportunistic infection in renal transplantation (RT) recipients. We aimed to describe its clinical presentation and treatment. MATERIALS AND METHODS: We collected microsporidiosis cases identified in RT recipients between 2005 and 2019 in six French centers from the Crystal, Divat and Astre prospective databases. RESULTS: We report 68 RT recipients with intestinal microsporidiosis; the patients were predominantly male (61.8%), with a median age of 58 (46-69) years. Infection occurred at a median time of 3 (0.8-6.8) years posttransplant. Only Enterocytozoon bieneusi was found. Microsporidiosis manifested as diarrhea (98.5% of patients) with weight loss (72.1%) and acute renal injury (57.4%) without inflammatory biological parameters. The therapeutic approaches were no treatment (N = 9), reduction of the immunosuppressive regimen (∆IS) (N = 22), fumagillin alone (N = 9), fumagillin and ∆IS (N = 19), and albendazole or nitazoxanide and ∆IS (N = 9). Overall clinical remission was observed in 60 patients (88.2%). We observed no acute kidney rejection, renal transplant failure, or death within 6 months after microsporidiosis. CONCLUSION: E. bieneusi is an underestimated opportunistic pathogen in RT recipients, and infection with E. bieneusi leads to diarrhea with important dehydration and acute renal injury. The treatment is based on the reduction of the immunosuppressive regimen and the administration of fumagillin if available.

A review of significance of Aspergillus detection in airways of ICU COVID-19 patients.

It is now well known that patients with SARS-CoV-2 infection admitted in ICU and mechanically ventilated are at risk of developing invasive pulmonary aspergillosis (IPA). Nevertheless, symptomatology of IPA is often atypical in mechanically ventilated patients, and radiological aspects in SARS-CoV-2 pneumonia and IPA are difficult to differentiate. In this context, the significance of the presence of Aspergillus in airway specimens (detected by culture, galactomannan antigen or specific PCR) remains to be fully understood. To decipher the relevance of the detection of Aspergillus, we performed a comprehensive review of all published cases of respiratory Aspergillus colonisation and IPA in COVID-19 patients. The comparison of patients receiving or not antifungal treatment allowed us to highlight the most important criteria for the decision to treat. The comparison of surviving and non-surviving patients made it possible to unveil criteria associated with mortality that should be taken into account in the treatment decision.

Large-scale genome mining allows identification of neutral polymorphisms and novel resistance mutations in genes involved in Candida albicans resistance to azoles and echinocandins.

BACKGROUND: The genome of Candida albicans displays significant polymorphism. Point mutations in genes involved in resistance to antifungals may either confer phenotypic resistance or be devoid of phenotypic consequences. OBJECTIVES: To catalogue polymorphisms in azole and echinocandin resistance genes occurring in susceptible strains in order to rapidly pinpoint relevant mutations in resistant strains. METHODS: Genome sequences from 151 unrelated C. albicans strains susceptible to fluconazole and caspofungin were used to create a catalogue of non-synonymous polymorphisms in genes involved in resistance to azoles (ERG11, TAC1, MRR1 and UPC2) or echinocandins (FKS1). The potential of this catalogue to reveal putative resistance mutations was tested in 10 azole-resistant isolates, including 1 intermediate to caspofungin. Selected mutations were analysed by mutagenesis experiments or mutational prediction effect. RESULTS: In the susceptible strains, we identified 126 amino acid substitutions constituting the catalogue of phenotypically neutral polymorphisms. By excluding these neutral substitutions, we identified 22 additional substitutions in the 10 resistant strains. Among these substitutions, 10 had already been associated with resistance. The remaining 12 were in Tac1p (n = 6), Upc2p (n = 2) and Erg11p (n = 4). Four out of the six homozygous substitutions in Tac1p (H263Y, A790V, H839Y and P971S) conferred increases in azole MICs, while no effects were observed for those in Upc2p. Additionally, two homozygous substitutions (Y64H and P236S) had a predicted conformation effect on Erg11p. CONCLUSIONS: By establishing a catalogue of neutral polymorphisms occurring in genes involved in resistance to antifungal drugs, we provide a useful resource for rapid identification of mutations possibly responsible for phenotypic resistance in C. albicans.

Aspergillus felis in Patient with Chronic Granulomatous Disease.

We report a case of Aspergillus felis infection in a patient with chronic granulomatous disease who had overlapping features of invasive pulmonary aspergillosis and allergic bronchopulmonary aspergillosis. Identifying the species responsible for aspergillosis by molecular methods can be crucial for directing patient management and selection of appropriate antifungal agents.

Systematic gene overexpression in Candida albicans identifies a regulator of early adaptation to the mammalian gut.

Candida albicans is part of the human gastrointestinal (GI) microbiota. To better understand how C. albicans efficiently establishes GI colonisation, we competitively challenged growth of 572 signature-tagged strains (~10% genome coverage), each conditionally overexpressing a single gene, in the murine gut. We identified CRZ2, a transcription factor whose overexpression and deletion respectively increased and decreased early GI colonisation. Using clues from genome-wide expression and gene-set enrichment analyses, we found that the optimal activity of Crz2p occurs under hypoxia at 37°C, as evidenced by both phenotypic and transcriptomic analyses following CRZ2 genetic perturbation. Consistent with early colonisation of the GI tract, we show that CRZ2 overexpression confers resistance to acidic pH and bile salts, suggesting an adaptation to the upper sections of the gut. Genome-wide location analyses revealed that Crz2p directly modulates the expression of many mannosyltransferase- and cell-wall protein-encoding genes, suggesting a link with cell-wall function. We show that CRZ2 overexpression alters cell-wall phosphomannan abundance and increases sensitivity to tunicamycin, suggesting a role in protein glycosylation. Our study reflects the powerful use of gene overexpression as a complementary approach to gene deletion to identify relevant biological pathways involved in C. albicans interaction with the host environment.

Why are so many cases of invasive aspergillosis missed?

Invasive aspergillosis (IA) incidence is increasing in several countries like France, and numerous cases are indeed missed and still only diagnosed at autopsy as evidenced by recently published data. Such missed diagnoses are obviously encountered when appropriate diagnostic tools are not available especially in low resource areas or when biologists have not been trained enough in medical mycology (i.e., microscopic examination and culture in most of those areas). Besides logistical issues, which are indeed critical, IA may not be recognized because clinicians failed to consider that risk factors are evolving with the IA burden now observed among patients with chronic lymphoid malignancies or receiving new biotherapies, with diabetes mellitus or liver cirrhosis and/or acute alcoholic hepatitis, with patients from the intensive care unit (ICU) and among patients with some predisposing primary immune deficiencies now reaching the adult's age. This is also the case for human immunodeficiency virus (HIV)-infected patients who failed to meet the classical definitions of IA. From the radiology perspective, new entities of IA have also emerged which absolutely need to be recognized especially bronchial-based-IA among allogeneic stem cell transplant recipients. Finally, from the laboratory side, contribution and limits of indirect blood biomarkers should be integrated to the clinical life in order not to miss IA cases. To conclude, several diagnostic tools should be combined and a constant dialog between laboratory and clinics is crucial to appropriately diagnose IA.