Features of cryptococcosis among 652 HIV-seronegative individuals in France: a cross-sectional observational study (2005-2020).

OBJECTIVES: We aimed to describe features and outcomes of cryptococcosis among HIV-seronegative individuals in a large surveillance network for cryptococcosis in France. METHODS: We included incident cases of cryptococcosis in HIV-seronegative individuals from 2005 to 2020. We compared patient characteristics, disease presentations, cryptococcal antigen results, and induction antifungal treatments according to underlying disease. We examined factors associated with 90-day mortality. Among patients with disseminated infections, we investigated whether receipt of flucytosine and polyene combination was associated with lower mortality. RESULTS: Among 652 individuals, 209 (32.1%) had malignancy, 130 (19.9%) were solid-organ transplant recipients, 204 (31.3%) had other immunocompromising conditions, and 109 (16.7%) had no reported underlying factor. The commonest presentations were disseminated infections (63.3%, 413/652) and isolated pulmonary infections (25.3%, 165/652). Solid-organ transplant patients were most likely to have disseminated infections and a positive serum cryptococcal antigen result. Patients with malignancy were older and less likely to receive a flucytosine-containing regimen for disseminated infections than others (58.7%, 78/133 vs. 73.2%, 194/265; p 0.029). The crude 90-day case-fatality ratio was 27.2% (95% CI, 23.5%-31.1%). Age ≥60 years (aOR: 2.75 [1.78-4.26]; p < 0.001), meningitis/fungaemia (aOR: 4.79 [1.80-12.7]; p 0.002), and malignancy (aOR: 2.4 [1.14-5.07]; p 0.02) were associated with higher 90-day mortality. Receipt of flucytosine and polyene combination was associated with lower 90-day mortality (aOR: 0.40 [0.23-0.71]; p 0.002) in multivariable analysis and inverse probability of treatment weighted analysis (aOR: 0.45 [0.25-0.80]; p 0.006). DISCUSSION: HIV-seronegative individuals with cryptococcosis comprise a wide range of underlying conditions with different presentations and outcomes, requiring a tailored approach to diagnosis and management.

Cryptococcus neoformans Infections Differ Among Human Immunodeficiency Virus (HIV)-Seropositive and HIV-Seronegative Individuals: Results From a Nationwide Surveillance Program in France.

Among 1107 cryptococcosis cases from the French surveillance network (2005-2020), the proportion of HIV-seronegative individuals has recently surpassed that of HIV-seropositive individuals. We observed marked differences in patient characteristics, disease presentations, cryptococcal antigen results, infecting species, and mortality according to HIV serostatus.

Caspofungin sequestration in a polyacrylonitrile-derived filter: Increasing the dose does not mitigate sequestration.

OBJECTIVES: Critically ill patients frequently require continuous renal replacement therapy. Echinocandins are recommended as first-line treatment of candidemia. Preliminary results suggested echinocandin sequestration in a polyacrylonitrile filter. The present study aimed to determine whether increasing the dose might balance sequestration. METHODS: An STX filter (Baxter-Gambro) was used. A liquid chromatography-mass spectrometry method was used for dosage of caspofungin. In vitro drug disposition was evaluated by NeckEpur (Neckepur, Versailles, France) technology using a crystalloid medium instead of diluted/reconstituted blood, focusing on the disposition of the unbound fraction of drugs. Two concentrations were assessed. RESULTS: At the low dose, the mean measured initial concentration in the central compartment (CC) was 5.1 ± 0.6 mg/L. One hundred percent of the initial amount was eliminated from the CC within the 6-h session. The mean total clearance from the CC was 9.6 ± 2.5 L/h. The mean percentages of elimination resulting from sequestration and diafiltration were 96.0 ± 5.0 and 4.0 ± 5.2%, respectively. At high dose, the mean measured initial concentration in the CC was 13.1 mg/L. One hundred percent of the initial amount was eliminated from the CC within the 6-h session. The mean total clearance from the CC was 9.5 L/h. The mean percentages of elimination resulting from sequestration and filtration were 88.5% and 11.5%, respectively. CONCLUSION: Increasing the dose does not mitigate caspofungin sequestration in the STX filter. The results raise caution about the simultaneous use of caspofungin and polyacrylonitrile-derived filters. Intermittent modes of renal replacement therapy might be considered. For sensitive species, fluconazole might be an alternative.

Candida haemulonii complex, an emerging threat from tropical regions?

BACKGROUND: Candida haemulonii complex-related species are pathogenic yeasts closely related to Candida auris with intrinsic antifungal resistance, but few epidemiological data are available. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed clinical and demographic characteristics of patients with fungemia due to C. haemulonii complex and related species (C. pseudohaemulonii, C. vulturna) reported in France during 2002-2021, and compared them to data of C. parapsilosis fungemia, as they all can be commensal of the skin. We also conducted a study on adult inpatients and outpatients colonized by C. haemulonii complex, managed at the University Hospital of Martinique during 2014-2020. Finally, we performed a literature review of fungemia due to C. haemulonii complex and related species reported in Medline (1962-2022). In total, we identified 28 fungemia due to C. haemulonii complex in France. These episodes were frequently associated with bacterial infection (38%) and high mortality rate (44%), and differed from C. parapsilosis fungemia by their tropical origin, mainly from Caribbean and Latin America. All isolates showed decreased in vitro susceptibility to amphotericin B and fluconazole. In Martinique, we found that skin colonization was frequent in the community population, while colonization was strongly associated with the presence of foreign devices in ICU patients. The literature review identified 274 fungemia episodes, of which 56 were individually described. As in our national series, published cases originated mainly from tropical regions and exhibited high crude mortality. CONCLUSIONS/SIGNIFICANCE: Multidrug-resistant C. haemulonii complex-related species are responsible for fungemia and colonization in community and hospital settings, especially in tropical regions, warranting closer epidemiological surveillance to prevent a potential C. auris-like threat.

Coregulation of extracellular vesicle production and fluconazole susceptibility in Cryptococcus neoformans.

Resistance to fluconazole (FLC), the most widely used antifungal drug, is typically achieved by altering the azole drug target and/or drug efflux pumps. Recent reports have suggested a link between vesicular trafficking and antifungal resistance. Here, we identified novel Cryptococcus neoformans regulators of extracellular vesicle (EV) biogenesis that impact FLC resistance. In particular, the transcription factor Hap2 does not affect the expression of the drug target or efflux pumps, yet it impacts the cellular sterol profile. Subinhibitory FLC concentrations also downregulate EV production. Moreover, in vitro spontaneous FLC-resistant colonies showed altered EV production, and the acquisition of FLC resistance was associated with decreased EV production in clinical isolates. Finally, the reversion of FLC resistance was associated with increased EV production. These data suggest a model in which fungal cells can regulate EV production in place of regulating the drug target gene expression as a first line of defense against antifungal assault in this fungal pathogen. IMPORTANCE Extracellular vesicles (EVs) are membrane-enveloped particles that are released by cells into the extracellular space. Fungal EVs can mediate community interactions and biofilm formation, but their functions remain poorly understood. Here, we report the identification of the first regulators of EV production in the major fungal pathogen Cryptococcus neoformans. Surprisingly, we uncover a novel role of EVs in modulating antifungal drug resistance. Disruption of EV production was associated with altered lipid composition and changes in fluconazole susceptibility. Spontaneous azole-resistant mutants were deficient in EV production, while loss of resistance restored initial EV production levels. These findings were recapitulated in C. neoformans clinical isolates, indicating that azole resistance and EV production are coregulated in diverse strains. Our study reveals a new mechanism of drug resistance in which cells adapt to azole stress by modulating EV production.

Active Surveillance Program to Increase Awareness on Invasive Fungal Diseases: the French RESSIF Network (2012 to 2018).

The French National Reference Center for Invasive Mycoses and Antifungals leads an active and sustained nationwide surveillance program on probable and proven invasive fungal diseases (IFDs) to determine their epidemiology in France. Between 2012 and 2018, a total of 10,886 IFDs were recorded. The incidence increased slightly over time (2.16 to 2.36/10,000 hospitalization days, P = 0.0562) in relation with an increase of fungemia incidence (1.03 to 1.19/10,000, P = 0.0023), while that of other IFDs remained stable. The proportion of ≥65-year-old patients increased from 38.4% to 45.3% (P < 0.0001). Yeast fungemia (n = 5,444) was due mainly to Candida albicans (55.6%) with stable proportions of species over time. Echinocandins became the main drug prescribed (46.7% to 61.8%), but global mortality rate remained unchanged (36.3% at 1 month). Pneumocystis jirovecii pneumonia (n = 2,106) was diagnosed mostly in HIV-negative patients (80.7%) with a significantly higher mortality than in HIV-positive patients (21.9% versus 5.4% at 1 month, P < 0.0001). Invasive aspergillosis (n = 1,661) and mucormycosis (n = 314) were diagnosed mostly in hematology (>60% of the cases) with a global mortality rate of 42.5% and 59.3%, respectively, at 3 months and significant changes in diagnosis procedure over time. More concurrent infections were also diagnosed over time (from 5.4% to 9.4% for mold IFDs, P = 0.0115). In conclusion, we observed an aging of patients with IFD with a significant increase in incidence only for yeast fungemia, a trend toward more concurrent infections, which raises diagnostic and therapeutic issues. Overall, global survival associated with IFDs has not improved despite updated guidelines and new diagnostic tools. IMPORTANCE The epidemiology of invasive fungal diseases (IFDs) is hard to delineate given the difficulties in ascertaining the diagnosis that is often based on the confrontation of clinical and microbiological criteria. The present report underlines the interest of active surveillance involving mycologists and clinicians to describe the global incidence and that of the main IFDs. Globally, although the incidence of Pneumocystis pneumonia, invasive aspergillosis, and mucormycosis remained stable over the study period (2012 to 2018), that of yeast fungemia increased slightly. We also show here that IFDs seem to affect older people more frequently. The most worrisome observation is the lack of improvement in the global survival rate associated with IFDs despite the increasing use of more sensitive diagnostic tools, the availability of new antifungal drugs very active in clinical trials, and a still low/marginal rate of acquired in vitro resistance in France. Therefore, other tracks of improvement should be investigated actively.

Echinocandins Susceptibility Patterns of 2,787 Yeast Isolates: Importance of the Thresholds for the Detection of FKS Mutations.

Since echinocandins are recommended as first line therapy for invasive candidiasis, detection of resistance, mainly due to alteration in FKS protein, is of main interest. EUCAST AFST recommends testing both MIC of anidulafungin and micafungin, and breakpoints (BPs) have been proposed to detect echinocandin-resistant isolates. We analyzed MIC distribution for all three available echinocandins of 2,787 clinical yeast isolates corresponding to 5 common and 16 rare yeast species, using the standardized EUCAST method for anidulafungin and modified for caspofungin and micafungin (AM3-MIC). In our database, 64 isolates of common pathogenic species were resistant to anidulafungin, according to the EUCAST BP, and/or to caspofungin, using our previously published threshold (AM3-MIC ≥ 0.5 mg/L). Among these 64 isolates, 50 exhibited 21 different FKS mutations. We analyzed the capacity of caspofungin AM3-MIC and anidulafungin MIC determination in detecting isolates with FKS mutation. They were always identified using caspofungin AM3-MIC and the local threshold while some isolates were misclassified using anidulafungin MIC and EUCAST threshold. However, both methods misclassified four wild-type C. glabrata as resistant. Based on a large data set from a single center, the use of AM3-MIC testing for caspofungin looks promising in identifying non-wild-type C. albicans, C. tropicalis and P. kudiravzevii isolates, but additional multicenter comparison is mandatory to conclude on the possible superiority of AM3-MIC testing compared to the EUCAST method.

Cryptococcus gattii Species Complex as an Opportunistic Pathogen: Underlying Medical Conditions Associated with the Infection.

The Cryptococcus gattii species complex has often been referred to as a primary pathogen due to its high infection frequency among apparently immunocompetent patients. In order to scrutinize the immune status of patients and the lineages of etiologic agents, we analyzed patient histories and the molecular types of etiologic agents from 135 global C. gattii cases. Eighty-six of 135 patients had been diagnosed as immunocompetent, although some of them had underlying medical issues, and 49 were diagnosed as immunocompromised with risk factors similar to those seen in Cryptococcus neoformans infection. We focused on the 86 apparently immunocompetent patients and were able to obtain plasma from 32 (37%) to analyze for the presence of autoantibodies against the granulocyte-macrophage colony-stimulating factor (GM-CSF) since these antibodies have been reported as a hidden risk factor for C. gattii infection. Among the 32 patients, 25 were free from any known other health issues, and 7 had various medical conditions at the time of diagnosis for cryptococcosis. Importantly, plasma from 19 (76%) of 25 patients with no recognized underlying medical condition showed the presence of GM-CSF autoantibodies, supporting this antibody as a major hidden risk factor for C. gattii infection. These data indicate that seemingly immunocompetent people with C. gattii infection warrant detailed evaluation for unrecognized immunologic risks. There was no relationship between molecular type and underlying conditions of patients. Frequency of each molecular type was related to its geographic origin exemplified by the overrepresentation of VGIV in HIV-positive (HIV+) patients due to its prevalence in Africa. IMPORTANCE The C. neoformans and C. gattii species complex causes cryptococcosis. The C. neoformans species complex is known as an opportunistic pathogen since it primarily infects immunocompromised patients. C. gattii species complex has been referred to as a primary pathogen due to its high infection frequency in apparently immunocompetent people. We analyzed 135 global cases of C. gattii infection with documented patient history. Eighty-six of 135 patients were originally diagnosed as immunocompetent and 49 as immunosuppressed with similar underlying conditions reported for C. neoformans infection. A significant number of C. gattii patients without known underlying conditions possessed autoantibodies against granulocytes-macrophage colony-stimulating factor (GM-CSF) in their plasma, supporting the presence of GM-CSF antibodies as a hidden risk factor for C. gattii infection. No relationship was found between C. gattii lineages and the underlying conditions except for overrepresentation of the molecular type VGIV among HIV+ patients due to the prevalence of VGIV in Africa.

Earliest case of Candida auris infection imported in 2007 in Europe from India prior to the 2009 description in Japan.

Candida auris is an emerging pathogen frequently associated with multidrug resistance and involved in many worldwide outbreaks. We here report the first European imported case in France due to isolate belonging of the South Indian clade I and the importance of prevention measure to avoid fungal spreading.

No Impact of Fluconazole to Echinocandins Replacement as First-Line Therapy on the Epidemiology of Yeast Fungemia (Hospital-Driven Active Surveillance, 2004-2017, Paris, France).

Replacement of fluconazole by echinocandins as the first-line therapy for yeast-related fungemia could have an impact on both the mortality rate and the epidemiology of yeast species responsible for candidemia. We analyzed the individual clinical and microbiological data collected through the active surveillance program on yeast fungemia (YEASTS program, 2004-2016, Paris area, France) within 14 University Hospitals. The cohort included 3,092 patients [male:female ratio: 1.56; median age 61.0 years (IQR: 23.8)]. The mean mortality rate within 30 days was 38.5% (1,103/2,868) and significantly higher in intensive care units (690/1,358, 50.8%) than outside (413/1,510, 27.4%, p < 0.0001) without significant change over time. The yeast species distribution [Candida albicans (n = 1,614, 48.0%), Candida glabrata (n = 607, 18.1%), Candida parapsilosis (n = 390, 11.6%), Candida tropicalis (n = 299, 8.9%), Candida krusei (n = 96, 2.9%), rare species (n = 357, 10.6%)], minimal inhibitory concentration distribution, and the distribution between the patient populations (hematological malignancies, solid tumors, without malignancy) did not change either while the proportion of patients ≥60-years increased from 48.7% (91/187) in 2004 to 56.8% (133/234) in 2017 (p = 0.0002). Fluconazole as first-line therapy dramatically decreased (64.4% in 2004 to 27.7% in 2017, p < 0.0001) with a corresponding increase in echinocandins (11.6% in 2004 to 57.8% in 2017, p < 0.0001). Survival rates did not differ according to the first antifungal therapy. The progressive replacement of fluconazole by echinocandins as the first-line antifungal therapy was not associated with change in global mortality, regardless of species involved and antifungal susceptibility profiles. Other factors remain to be uncovered to improve the prognosis of yeast fungemia.

Azole Susceptibility Profiles of More than 9,000 Clinical Yeast Isolates Belonging to 40 Common and Rare Species.

Invasive yeast infections represent a major global public health issue, and only few antifungal agents are available. Azoles are one of the classes of antifungals used for treatment of invasive candidiasis. The determination of antifungal susceptibility profiles using standardized methods is important to identify resistant isolates and to uncover the potential emergence of intrinsically resistant species. Here, we report data on 9,319 clinical isolates belonging to 40 pathogenic yeast species recovered in France over 17 years. The antifungal susceptibility profiles were all determined at the National Reference Center for Invasive Mycoses and Antifungals based on the EUCAST broth microdilution method. The centralized collection and analysis allowed us to describe the trends of azole susceptibility of isolates belonging to common species, confirming the high susceptibility for Candida albicans (n = 3,295), Candida tropicalis (n = 641), and Candida parapsilosis (n = 820) and decreased susceptibility for Candida glabrata (n = 1,274) and Pichia kudriavzevii (n = 343). These profiles also provide interesting data concerning azole susceptibility of Cryptococcus neoformans species complex, showing comparable MIC distributions for the three species but lower MIC50s and MIC90s for serotype D (n = 208) compared to serotype A (n = 949) and AD hybrids (n = 177). Finally, these data provide useful information for rare and/or emerging species, such as Clavispora lusitaniae (n = 221), Saprochaete clavata (n = 184), Meyerozyma guilliermondii complex (n = 150), Candida haemulonii complex (n = 87), Rhodotorula mucilaginosa (n = 55), and Wickerhamomyces anomalus (n = 36).

Cryptococcus gattii in Patients with Lymphoid Neoplasms: An Illustration of Evolutive Host-Fungus Interactions.

Recent outbreaks of Cryptococcus gattii (CG) infections in North America have sparked renewed interest in the pathogenic potential of CG, and have underscored notable differences with Cryptococcus neoformans in terms of geographic distribution, pathogen virulence, and host susceptibility. While cases of CG are increasingly reported in patients with a wide variety of underlying conditions, only very few have been reported in patients with lymphoid neoplasms. Herein, we report a case of autochthonous CG meningitis in a patient receiving ibrutinib for chronic lymphocytic leukemia in France, and review available data on the clinical epidemiology of CG infections in patients with lymphoid neoplasms. We also summarise recent data on the host responses to CG infection, as well as the potential management pitfalls associated with its treatment in the haematological setting. The clinical epidemiology, clinical presentation, and course of disease during infections caused by CG involve complex interactions between environmental exposure to CG, infecting genotype, pathogen virulence factors, host susceptibility, and host immune responses. Future treatment guidelines should address the challenges associated with the management of antifungal treatments in the onco-haematological setting and the potential drug-drug interactions.

Kazachstania slooffiae: An unexpected journey to a human pleural sample.

We report a case of a 50-year-old shepherd hospitalized in intensive care unit for hiatal hernia complicated by an occlusive syndrome. In post-surgery, an acute respiratory distress occurs due to mediastinitis with large pleural effusion. At the laboratory, direct examination of the pleural sample revealed the presence of pseudohyphae. Kazachstania slooffiae was identified by Mass Spectrometry and confirmed by DNA sequencing. This uncommon yeast has never been previously described in human infections. Although its pathogenicity is not well known, K. slooffiae should be considered in the case of critically ill patients.

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.

A Short-Tandem-Repeat Assay (MmySTR) for Studying Genetic Variation in Madurella mycetomatis.

Madurella mycetomatis is the major causative agent of eumycetoma, a neglected tropical infection characterized by painless subcutaneous lesions, inflammation, and grains draining from multiple sinuses. To study the epidemiology of mycetoma, a robust discriminatory typing technique is needed. We describe the use of a short-tandem-repeat assay (MmySTR) for genotyping of M. mycetomatis isolates predominantly from Sudan. Eleven microsatellite markers (3 dinucleotides, 4 trinucleotide repeats, and 4 tetranucleotide repeats) were selected from the M. mycetomatis MM55 genome using the Tandem Repeats Finder software. PCR amplification primers were designed for each microsatellite marker using primer3 software and amplified in a multicolor multiplex PCR approach. To establish the extent of genetic variation within the population, a collection of 120 clinical isolates from different regions was genotyped with this assay. The 11 selected MmySTR markers showed a large genotypic heterogeneity. From a collection of 120 isolates, 108 different genotypes were obtained. Simpson's diversity index (D) value for individual markers ranged from 0.081 to 0.881, and the combined panel displayed an overall D value of 0.997. The MmySTR assay demonstrated high stability, reproducibility, and specificity. The MmySTR assay is a promising new typing technique that can be used to genotype isolates of M. mycetomatis Apart from the possible contribution of host factors, the genetic diversity observed among this group of isolates might contribute to the different clinical manifestations of mycetoma. We recommend that the MmySTR assay be used to establish a global reference database for future study of M. mycetomatis isolates.

Case Report: Emergence of Candida auris in the Indian Ocean Region.

Candida auris is an emerging multidrug-resistant yeast that can cause severe infections and spread easily between hospitalized patients, leading to outbreaks in hospital. Here, we report the first four cases of colonization and invasive infection with C. auris reported in the Indian Ocean region. All cases were observed in the French overseas Reunion Island, a very popular destination for European travelers. Three patients had urinary tract or skin colonization, and one had a fatal invasive infection. In three cases, including that of the infected patient, the yeast was not initially identified as C. auris, preventing specific hygiene measures to be implemented as suggested in the December 2016 clinical alert to European healthcare facilities. The infected patient likely acquired C. auris in the intensive care unit from the first colonized patient. This is the first case of C. auris infection and the first potential case of nosocomial transmission of the pathogen to be reported in the French overseas Reunion Island.

Comparison of MultiLocus Sequence Typing (MLST) and Microsatellite Length Polymorphism (MLP) for Pneumocystis jirovecii genotyping.

Pneumocystis jirovecii is an atypical fungus responsible for severe respiratory infections, often reported as local outbreaks in immunocompromised patients. Epidemiology of this infection, and transmission risk emphasises the need for developing genotyping techniques. Currently, two methods have emerged: Multilocus Sequence typing (MLST) and microsatellite length polymorphism (MLP). Here we compare an MLST strategy, including 2 nuclear loci and 2 mitochondrial loci, with an MLP strategy including 6 nuclear markers using 37 clinical PCR-positive respiratory samples from two French hospitals. Pneumocystis jirovecii MLST and MLP provided 30 and 35 different genotypes respectively. A higher number of mixed infections was detected using MLP (48.6% vs. 13.5% respectively; p = 0.002). Only one MLP marker (STR279) was statistically associated with the geographical origin of samples. Haplotype network inferred using the available genotypes yielded expanded network for MLP, characterized by more mutational steps as compared to MLST, suggesting that the MLP approach is more resolutive to separate genotypes. The correlation between genetic distances calculated based on MLST and MLP was modest with a R 2 value = 0.32 (p < 0.001). Finally, both genotyping methods fulfilled important criteria: (i) a discriminatory power from 97.5% to 99.5% and (ii) being quick and convenient genotyping tools. While MLP appeared highly resolutive regarding genotypes mixture within samples, using one genotyping method rather than the other may also depend on the context (i.e., MLST for investigation of suspected clonal outbreaks versus MLP for population structure study) as well as local facilities.

Saprochaete clavata Outbreak Infecting Cancer Center through Dishwasher.

Saprochaete clavata is a pathogenic yeast responsible for rare outbreaks involving immunocompromised patients, especially those with hematologic malignancies. During February 2016-December 2017, we diagnosed S. clavata infections in 9 patients (8 with fungemia), including 3 within 1 month, at a cancer center in Marseille, France. The patients (median age 58 years), 4 of 9 of whom had acute myeloid leukemia, were hospitalized in 3 different wards. Ten environmental samples, including from 2 dishwashers and 4 pitchers, grew S. clavata, but no contaminated food was discovered. The outbreak ended after contaminated utensils and appliances were discarded. Whole-genome sequencing analysis demonstrated that all clinical and environmental isolates belonged to the same phylogenetic clade, which was unrelated to clades from previous S. clavata outbreaks in France. We identified a dishwasher with a deficient heating system as the vector of contamination.

The genus Madurella: Molecular identification and epidemiology in Sudan.

Eumycetoma (mycotic mycetoma) is the fungal form of mycetoma, a subcutaneous infection occurring in individuals living in endemic areas of the disease. The Sudan is hyperendemic for mycetoma, with the highest incidence being reported from Gezira State, Central Sudan. The present study was conducted at the Gezira Mycetoma Center and aimed to determine the cause of black-grain eumycetoma in the state and describe its epidemiology. Black-grain specimens were collected during the surgical operation and direct detection of the causative agent was performed using M. mycetomatis species-specific PCR and ITS PCR followed by sequencing. Black-grain was reported from 93.3% of all confirmed mycetoma cases (n = 111/119), with a prevalence in young males. Of the 91 samples subjected to direct PCR, 90.1% (n = 82) gave positive results. The predominant species (88.2%) was Madurella mycetomatis. One sample was identified as M. fahalii, one as M. tropicana, and one matched the phytopathogenic species Sphaerulina rhododendricola. The highest endemic zones were Southern Gezira (76.6%) and Northern Sinnar (23.4%). The study confirmed that direct molecular detection on grains provides rapid and specific diagnosis of agents of eumycetoma.