Divergent effects of emm types 1 and 12 on invasive group A streptococcal infections-results of a retrospective cohort study, Germany 2023.

As a potential side effect of the severe acute respiratory syndrome coronavirus type 2 pandemic, invasive group A Streptococcus (iGAS) infections in Europe have increased dramatically in both children and adults in the end of 2022. This epidemiological and molecular study describes the distributions of streptococcal genes encoding the M antigen (emm types) and superantigens in patients with invasive and non-invasive GAS infections. From December 2022 to December 2023, a total of 163 GAS isolates were collected from sterile and non-sterile sites of patients at five hospitals in Germany including two tertiary care centers. Genes encoding M protein and superantigens were determined following the guidelines of CDC Streptococcus laboratory. Patients' characteristics were reviewed retrospectively. Correlations of clinical factors, emm types, and superantigens with rates of invasive infections were analyzed. Of the 163 included GAS cases, 112 (69%) were considered as invasive. In total, 33 different emm types were observed, of which emm1.0 (n = 49; 30%), emm89.0 (n = 15; 9%), and emm12.0 (n = 14; 9%) were most prevalent. In total, 70% of emm1.0 isolates belonged to M1UK lineage. No difference in invasive infections was observed for the M1UK lineage compared with other emm1.0 isolates. However, the emm1.0 type, presence of speA1-3, speG, or speJ, as well as adulthood were significantly associated with invasive infections. In contrast, emm12.0 isolates were significantly less associated with invasive infections. Multivariable analysis confirmed a significant influence of speJ and adulthood on iGAS infections. This study underlines the importance of continuous monitoring of genomic trends and identification of emerging GAS variants. This may aid in delineating pathogenicity factors of Streptococcus pyogenes that propel invasive infections.

Key summary of German national guideline for adult patients with nosocomial pneumonia- Update 2024 Funding number at the Federal Joint Committee (G-BA): 01VSF22007.

PURPOSE: This executive summary of a German national guideline aims to provide the most relevant evidence-based recommendations on the diagnosis and treatment of nosocomial pneumonia. METHODS: The guideline made use of a systematic assessment and decision process using evidence to decision framework (GRADE). Recommendations were consented by an interdisciplinary panel. Evidence analysis and interpretation was supported by the German innovation fund providing extensive literature searches and (meta-) analyses by an independent methodologist. For this executive summary, selected key recommendations are presented including the quality of evidence and rationale for the level of recommendation. RESULTS: The original guideline contains 26 recommendations for the diagnosis and treatment of adults with nosocomial pneumonia, thirteen of which are based on systematic review and/or meta-analysis, while the other 13 represent consensus expert opinion. For this key summary, we present 11 most relevant for everyday clinical practice key recommendations with evidence overview and rationale, of which two are expert consensus and 9 evidence-based (4 strong, 5 weak and 2 open recommendations). For the management of nosocomial pneumonia patients should be divided in those with and without risk factors for multidrug-resistant pathogens and/or Pseudomonas aeruginosa. Bacterial multiplex-polymerase chain reaction (PCR) should not be used routinely. Bronchoscopic diagnosis is not considered superior to´non-bronchoscopic sampling in terms of main outcomes. Only patients with septic shock and the presence of an additional risk factor for multidrug-resistant pathogens (MDRP) should receive empiric combination therapy. In clinically stabilized patients, antibiotic therapy should be de-escalated and focused. In critically ill patients, prolonged application of suitable beta-lactam antibiotics should be preferred. Therapy duration is suggested for 7-8 days. Procalcitonin (PCT) based algorithm might be used to shorten the duration of antibiotic treatment. Patients on the intensive care unit (ICU) are at risk for invasive pulmonary aspergillosis (IPA). Diagnostics for Aspergillus should be performed with an antigen test from bronchial lavage fluid. CONCLUSION: The current guideline focuses on German epidemiology and standards of care. It should be a guide for the current treatment and management of nosocomial pneumonia in Germany.

Establishment of a Novel Short Tandem Repeat Typing Method for Exophiala dermatitidis.

The opportunistic black yeast-like fungus Exophiala dermatitidis frequently colonizes the respiratory tract of cystic fibroses (CF) patients. Additionally, it can cause superficial, systemic, and cerebral forms of phaeohyphomycoses. The objective of this study was to develop and apply a microsatellite or short tandem repeat (STR) genotyping scheme for E. dermatitidis. In total, 82 E. dermatitidis isolates from various geographic origins (environmental = 9, CF = 63, invasive isolates = 9, melanin-deficient mutant = 1) were included in this study. After next-generation sequencing of a reference strain and sequence filtering for microsatellites, six STR markers were selected and amplified in two multiplex PCR reactions. The included isolates were discriminated in a genetic cluster analysis using the Pearson algorithm to reveal the relatedness of the isolates. The E. dermatitidis isolates clustered on basis of both, their source and their origin. The invasive isolates from Asia were unrelated to isolates from CF. Nearly all environmental isolates were grouped separately from patients' isolates. The Simpson index was 0.94. In conclusion, we were able to establish a STR genotyping scheme for investigating population genomics of E. dermatitidis.

Targeting the Granulocytic Defense against A. fumigatus in Healthy Volunteers and Septic Patients.

Neutrophil granulocytes (NGs) are among the key players in the defense against Aspergillus fumigatus (A. fumigatus). To better elucidate a pathophysiological understanding of their role and functions, we applied a human cell model using NGs from healthy participants and septic patients to evaluate their inhibitory effects on the growth of A. fumigatus ex vivo. Conidia of A. fumigatus (ATCC® 204305) were co-incubated with NGs from healthy volunteers or septic patients for 16 h. A. fumigatus growth was measured by XTT assays with a plate reader. The inhibitory effect of NGs on 18 healthy volunteers revealed great heterogeneity. Additionally, growth inhibition was significantly stronger in the afternoon than the morning, due to potentially different cortisol levels. It is particularly interesting that the inhibitory effect of NGs was reduced in patients with sepsis compared to healthy controls. In addition, the magnitude of the NG-driven defense against A. fumigatus was highly variable among healthy volunteers. Moreover, daytime and corresponding cortisol levels also seem to have a strong influence. Most interestingly, preliminary experiments with NGs from septic patients point to a strongly diminished granulocytic defense against Aspergillus spp.

Inhibition of azole-resistant Aspergillus fumigatus biofilm at various formation stages by antifungal drugs, including olorofim.

OBJECTIVES: Interest in aspergillosis has increased over the past decades. An increase in the incidence of azole-resistant Aspergillus fumigatus strains has been reported; therefore, the need for novel therapeutic approaches is urgent. The formation of biofilms contributes to pathogen resistance. We investigated the biofilm formation capabilities of azole-resistant A. fumigatus and analysed the susceptibility of biofilms at various developmental stages to three antifungal agents. METHODS: Biofilm formation of 19 clinical A. fumigatus strains (3 azole-susceptible and 16 azole-resistant strains) was determined by crystal violet staining and by an XTT assay over a period of 48 h. We measured antibiofilm activity of voriconazole, amphotericin B and olorofim. These agents were added before adhesion, after adhesion, after germination and to mature fungal biofilm. Antibiofilm activity was assessed in an XTT assay and in confocal laser scan microscopy. Additionally, a growth-kinetic assay with planktonic A. fumigatus was performed. RESULTS: Each of the antifungal agents inhibited the metabolic activity of A. fumigatus biofilms when applied at early stages of biofilm formation. The mature biofilms were more resistant. Olorofim and voriconazole showed promising effects against A. fumigatus adhesion and germination, whereas the mature biofilm was not affected by treatment. In contrast, the biofilm of A. fumigatus showed amphotericin B susceptibility throughout the entire developmental process. The planktonic cells were susceptible to all three antifungal drug classes with an inhibition peak at 12 h after incubation. CONCLUSIONS: This is the first known study to demonstrate the antibiofilm activity of olorofim, voriconazole and amphotericin B against azole-resistant A. fumigatus.

COVID-19-associated pulmonary aspergillosis in ICU patients in a German reference centre: Phenotypic and molecular characterisation of Aspergillus fumigatus isolates.

BACKGROUND: COVID-19-associated invasive pulmonary aspergillosis (CAPA) is associated with increased mortality. Cases of CAPA caused by azole-resistant Aspergillus fumigatus strains have been reported. OBJECTIVES: To analyse the twelve-month CAPA prevalence in a German tertiary care hospital and to characterise clinical A. fumigatus isolates from two German hospitals by antifungal susceptibility testing and microsatellite genotyping. PATIENTS/METHODS: Retrospective observational study in critically ill adults from intensive care units with COVID-19 from 17 February 2020 until 16 February 2021 and collection of A. fumigatus isolates from two German centres. EUCAST broth microdilution for four azole compounds and microsatellite PCR with nine markers were performed for each collected isolate (N = 27) and additional for three non-COVID A. fumigatus isolates. RESULTS: welve-month CAPA prevalence was 7.2% (30/414), and the rate of azole-resistant A. fumigatus isolates from patients with CAPA was 3.7% with detection of one TR34/L98H mutation. The microsatellite analysis revealed no major clustering of the isolates. Sequential isolates mainly showed the same genotype over time. CONCLUSIONS: Our findings demonstrate similar CAPA prevalence to other reports and a low azole-resistance rate. Genotyping of A. fumigatus showed polyclonal distribution except for sequential isolates.

Sphingosine as a New Antifungal Agent against Candida and Aspergillus spp.

This study investigated whether sphingosine is effective as prophylaxis against Aspergillus spp. and Candida spp. In vitro experiments showed that sphingosine is very efficacious against A. fumigatus and Nakeomyces glabrataa (formerly named C. glabrata). A mouse model of invasive aspergillosis showed that sphingosine exerts a prophylactic effect and that sphingosine-treated animals exhibit a strong survival advantage after infection. Furthermore, mechanistic studies showed that treatment with sphingosine leads to the early depolarization of the mitochondrial membrane potential (Δψm) and the generation of mitochondrial reactive oxygen species and to a release of cytochrome C within minutes, thereby presumably initiating apoptosis. Because of its very good tolerability and ease of application, inhaled sphingosine should be further developed as a possible prophylactic agent against pulmonary aspergillosis among severely immunocompromised patients.

COVID-19 Co-infection with Legionella pneumophila in 2 Tertiary-Care Hospitals, Germany.

We describe screening results for detection of co-infections with Legionella pneumophila in patients infected with severe acute respiratory syndrome coronavirus 2. In total, 93 patients were tested; 1 was positive (1.1%) for L. pneumophila serogroup 1. Co-infections with L. pneumophila occur in coronavirus disease patients and should not be missed.

Orbital aspergillosis: a case report and review of the literature.

BACKGROUND: Orbital aspergillosis is a rare sight- and life-threatening fungal infection affecting immunocompromised or otherwise healthy patients. It is often misdiagnosed due to its unspecific clinical and radiologic appearance. Therapeutic delay can have dramatic consequences. However, progress in microbiological diagnostic techniques and therapeutic experience from case series help improve the management of this disease. CASE PRESENTATION: A 78-year-old immunocompetent woman presented at an eye clinic for subacute swelling, reddening, and ptosis of her left upper eyelid. Based on radiologic and histologic considerations, she was treated for idiopathic orbital inflammation, but her condition worsened. After a second biopsy of the orbital mass, aspergillosis was diagnosed. Her condition improved promptly after initiation of an oral voriconazole treatment. Additionally, using a polymerase chain reaction (PCR) assay, A. fumigatus was identified on tissue of both biopsies and its azole susceptibility was examined simultaneously. CONCLUSIONS: In the case described here, oral antifungal treatment was sufficient for the therapy of invasive orbital aspergillosis. Performing fungal PCR on orbital tissue can accelerate the diagnostic process and should be performed in ambiguous cases of slowly growing orbital mass. Finally, interdisciplinary management is the key to optimal treatment of orbital tumours and infections.

Antibiofilm activity of antifungal drugs, including the novel drug olorofim, against Lomentospora prolificans.

OBJECTIVES: Patients with immunodeficiency or cystic fibrosis frequently suffer from respiratory fungal infections. In particular, biofilm-associated fungi cause refractory infection manifestations, linked to increased resistance to anti-infective agents. One emerging filamentous fungus is Lomentospora prolificans. Here, the biofilm-formation capabilities of L. prolificans isolates were investigated and the susceptibility of biofilms to various antifungal agents was analysed. METHODS: Biofilm formation of L. prolificans (n = 11) was estimated by crystal violet stain and antibiofilm activity was additionally determined via detection of metabolically active biofilm using an XTT assay. Amphotericin B, micafungin, voriconazole and olorofim were compared with regard to their antibiofilm effects when added prior to adhesion, after adhesion and on mature and preformed fungal biofilms. Imaging via confocal laser scanning microscopy was carried out to demonstrate the effect of drug treatment on the fungal biofilm. RESULTS: Antibiofilm activities of the tested antifungal agents were shown to be most effective on adherent cells whilst mature biofilm was the most resistant. The most promising antibiofilm effects were detected with voriconazole and olorofim. Olorofim showed an average minimum biofilm eradication concentration (MBEC) of 0.06 mg/L, when added prior to and after adhesion. The MBECs of voriconazole were ≤4 mg/L. On mature biofilm the MBECs of olorofim and voriconazole were higher than the previously determined MICs against planktonic cultures. In contrast, amphotericin B and especially micafungin did not exhibit sufficient antibiofilm activity against L. prolificans. CONCLUSIONS: To our knowledge, this is the first study demonstrating the antibiofilm potential of olorofim against the human pathogenic fungus L. prolificans.

Evaluation of two lateral flow assays in BAL fluids for the detection of invasive pulmonary aspergillosis: A retrospective two-centre study.

BACKGROUND: Commonly, the application of radiological and clinical criteria and the determination of galactomannan (GM) in respiratory samples are used as a diagnostic tool for the detection of invasive pulmonary aspergillosis (IPA). MATERIALS/METHODS: In this study, two lateral flow assays, OLM Aspergillus lateral flow device (LFD) and IMMY sona Aspergillus Galactomannan lateral flow assay (LFA), were evaluated at two tertiary hospitals in Germany. A total of 200 bronchoalveolar lavage (BAL) samples from patients with suspicion of IPA were analysed retrospectively. LFD and LFA were evaluated against four different criteria: Blot, EORTC/MSG, Schauwvlieghe and extended Blot criteria and additionally against GM. RESULTS: The evaluation of four algorithms for the diagnosis of IPA showed that there exist good diagnostic tools to rule out an IPA even before results of Aspergillus culture are available. Sensitivities and negative predictive values are generally higher for the LFA than for the LFD in all four criteria. Specificity and positive predictive values varied depending on the classification criteria. The total agreement between the GM and the LFA cube reader (cut-off = 1) was 84%. The correlation between the GM and LFA was calculated with r = 0.8. CONCLUSION: The here presented data indicate that a negative LFA result in BAL fluid can reliable rule out an IPA in a heterogeneous group of ICU patients based on the original Blot criteria. LFA seems to be a promising immunochromatographic test exhibiting a good agreement with positive GM values.

Evaluation of a commercial Loop-mediated Isothermal Amplification (LAMP) assay for rapid detection of Pneumocystis jirovecii.

BACKGROUND: Various tools are obtainable for the detection of Pneumocystis jirovecii, among them qPCR promising highest sensitivity. A novel molecular method is commercially available, the loop-mediated isothermal amplification (LAMP) assay. OBJECTIVES: We compared the performance of the LAMP eazyplex® Pneumocystis jirovecii with the RealStar Pneumocystis jirovecii PCR 1.0 qPCR. MATERIAL/METHODS: Overall, 162 lower respiratory tract specimens from 146 critically ill patients were investigated. LAMP assay and qPCR were carried out according to the manufacturer's recommendations. Positive results of the LAMP were described as time to positivity (TTP). The limit of detection (LOD) of the LAMP was analysed using 10-fold serial dilutions of a high positive P jirovecii respiratory sample. For each serial dilution, TTP of the LAMP was plotted against cycle threshold (Ct) values of the qPCR. RESULTS: The LOD of the LAMP was determined to be approximately 4 × 103 copies/mL. While the LAMP revealed 28 (17%) positive signals from 20 patients, by using qPCR 41 (25%) positive samples from 28 patients were identified. Overall agreement with qPCR was 92%. Five false-negative, one false-positive and nine invalid results were detected by the LAMP. Positive and negative predictive values were 96% each, and sensitivity and specificity were 84% and 99%, respectively. There was a low correlation between the TTP and the fungal load. CONCLUSION: The LAMP is a time-saving and easy-to-perform method. It can be used as an alternative diagnostic method. However, for quantification purposes the qPCR is still the gold standard.

Comparison of four commercially available chromogenic media to identify Candida albicans and other medically relevant Candida species.

BACKGROUND: The number of invasive Candida infections has significantly increased in recent decades. For the successful treatment of fungal infections, rapid identification at the species level, particularly in polyfungal infections, is a key factor. In this study, four commercially available chromogenic media, CandiSelect™ 4 (CS4), chromID™ Candida Agar (CCA), BBL™ CHROMagar™ Candida Medium (BBL) and Brilliance™ Candida Agar (BCA) were evaluated for Candida identification. MATERIAL/METHODS: Overall, 181 bronchial secretion samples from intensive care patients were analysed prospectively. In addition, 18 primarily sterile materials, previously tested positive for Candida, were investigated retrospectively. All samples were cultured as recommended by the manufacturer and visually inspected after 24 and 48 hours by three independent investigators. As a control, colonies were identified by MALDI-TOF MS. Specificity and sensitivity were determined for C albicans identification prospectively. RESULTS: CS4 and BCA showed the best overall consensus with the identification results reached by MALDI-TOF MS for Candida albicans and species. A clear differentiation between the species could be ascertained via easily identifiable, species-specific coloration in contrast to BBL and CCA. Sensitivity for C albicans (n = 73) identification varied between 32% (BCA) and 69% (CS4 and CCA) after 24 hours and 68% (BBL) and 82% (BCA) after 48 hours incubation, while specificity ranged between 62% (BBL) and 81% (CCA) after 24 hours and 82% (BBL) and 85% (CS4) after 48 hours. CONCLUSION: CS4 and BCA are recommended for routine identification of Candida species in human samples.

European confederation of medical mycology expert consult-An ECMM excellence center initiative.

OBJECTIVES: Difficult-to-treat invasive fungal infections require infectious diseases expert consultation to improve treatment outcome and increase survival rates. METHODS: The European Confederation of Medical Mycology (ECMM) intends to provide expert help free of charge by a newly founded ECMM Expert Consultation Service for medical centres around the globe seeking advice when there is no fungal infection consultant available. The expert consult will provide recommendations and broad expertise on difficult-to-treat invasive fungal infections (eg azole-resistant Aspergillus species, Candida auris, mucormycosis) to improve diagnostic and therapeutic management and outcome. RESULTS: The initiative plans global outreach through video conferencing between ECMM Excellence Centers and treating physicians. FungiScope® registries will be used to structure case information and to evaluate the impact of the collegial advice system at regular intervals. Advice will follow recent guidelines, and EQUAL Scores will be used to measure guideline adherence. CONCLUSIONS: Infectious diseases expert consultation should be an integral component of care for patients with difficult-to-treat invasive fungal infections. The ECMM Expert Consult will attend to this matter on a global scale.

Disseminated Emergomycosis in a Person with HIV Infection, Uganda.

We describe emergomycosis in a patient in Uganda with HIV infection. We tested a formalin-fixed, paraffin-embedded skin biopsy to identify Emergomyces pasteurianus or a closely related pathogen by sequencing broad-range fungal PCR amplicons. Results suggest that emergomycosis is more widespread and genetically diverse than previously documented. PCR on tissue blocks may help clarify emergomycosis epidemiology.

Comparison of genotyping methods for Cunninghamella bertholletiae.

BACKGROUND: Invasive fungal infections caused by filamentous fungi of the order Mucorales are serious complications in immunocompromised patients and often associated with fatal outcome. As a member of this order, Cunninghamella bertholletiae is a saprophytic fungus with naturally exhibited high minimum inhibitory concentrations against common antifungal drugs and with the potential for outbreaks in clinical settings. OBJECTIVES AND METHODS: In a proof-of-principle study, we evaluated the performance of microsatellite markers for the discrimination of thirteen C. bertholletiae isolates from various sources in comparison with a repetitive sequence-based PCR (rep-PCR) and random amplification of polymorphic DNA (RAPD). Based on the higher discriminatory power of the microsatellite PCR with five separate primer pairs (Simpson's index of 1 vs 0 [RAPD] and 0 [rep-PCR]), the novel method was applied to eight additional isolates, including four well-characterised isolates from a cluster of infections in a next step. RESULTS: In total, microsatellite PCR identified 21 separate genotypes. A probable epidemiological association of the cluster isolates could be demonstrated by microsatellite genotyping. CONCLUSION: In conclusion, our findings demonstrate the value of microsatellite PCR in genotyping Cunninghamella bertholletiae and its potential for future applications with other species of the order Mucorales.

Multiplex PCR of bronchoalveolar lavage fluid in children enhances the rate of pathogen detection.

BACKGROUND: Culturing of bronchoalveolar lavage (BAL) fluid is a commonly used method for pathogen detection in pneumonia. However, the sensitivity is low, especially in patients pre-treated with anti-infective agents. The early detection of a pathogen is crucial for the outcome of respiratory tract infections. For bloodstream infections, a multiplex polymerase chain reaction (PCR) assay (SeptiFast®, SF) is available for improved pathogen detection from blood. OBJECTIVE: The aim of the present study was to determine whether the SF assay is applicable to the BAL of children with pulmonary infections and whether the frequency of pathogen detection is enhanced by the use of this multiplex PCR method. METHODS: We investigated 70 BAL samples of 70 children simultaneously by culture and multiplex PCR. The frequency of pathogen detection was compared. RESULTS: Pathogens were detected more frequently by SF than by culture (83% vs. 31%; p < 0.001). This advantage was shown for immunocompetent patients (p = 0.001) as well as for immunocompromised patients (p = 0.003). The majority (38/44; 86%) of the Gram positive cocci were only detected by SF. Fungal organisms were detected in 7/70 patients (10%) by SF and in 2/70 (3%) by culture (p = 0.125). CONCLUSION: Compared to conventional culture, the use of the SF assay on the BAL of children with pneumonia increases pathogen detection rates and therefore adds important information to guide anti-infective therapy.

Multiplex polymerase chain reaction to diagnose bloodstream infections in patients after cardiothoracic surgery.

BACKGROUND: Sepsis and other infectious complications are major causes of mortality and morbidity in patients after cardiac surgery. Whereas conventional blood culture (BC) suffers from low sensitivity as well as a reporting delay of approximately 48-72 h, real-time multiplex polymerase chain reaction (PCR) based technologies like "SeptiFast" (SF) might offer a fast and reliable alternative for detection of bloodstream infections (BSI). The aim of this study was to compare the performance of SF with BC testing in patients suspected of having BSI after cardiac surgery. METHODS: Two hundred seventy-nine blood samples from 169 individuals with suspected BSI were analyzed by SF and BC. After excluding results attributable to contaminants, a comparison between the two groups were carried out. Receiver operating characteristic (ROC) curves were generated to determine the accuracy of clinical and laboratory values for the prediction of positive SF results. RESULTS: 14.7% (n = 41) of blood samples were positive using SF and 17.2% (n = 49) using BC (n.s. [p > 0.05]). In six samples SF detected more than one pathogen. Among the 47 microorganisms identified by SF, only 11 (23.4%) could be confirmed by BC. SF identified a higher number of Gram-negative bacteria than BC did (28 vs. 12, χ2 = 7.97, p = 0.005). The combination of BC and SF increased the number of detected microorganisms, including fungi, compared to BC alone (86 vs. 49, χ2 = 13.51, p < 0.001). C-reactive protein (CRP) (21.7 ± 11.41 vs. 16.0 ± 16.9 mg/dl, p = 0.009), procalcitonin (28.7 ± 70.9 vs. 11.5 ± 30.4 ng/dl, p = 0.015), and interleukin 6 (IL 6) (932.3 ± 1306.7 vs. 313.3 ± 686.6 pg/ml, p = 0.010) plasma concentrations were higher in patients with a positive SF result. Using ROC analysis, IL-6 (AUC 0.836) and CRP (AUC 0.804) showed the best predictive values for positive SF results. CONCLUSION: The SF test represent a valuable method for rapid etiologic diagnosis of BSI in patients after cardiothoracic surgery. In particular this method applies for individuals with suspected Gram-negative blood stream. Due to the low performance in detecting Gram-positive pathogens and the inability to determine antibiotic susceptibility, it should be used in addition to BC only (Pilarczyk K, et al., Intensive Care Med Exp ,3(Suppl. 1):A884, 2015).

FunResDB-A web resource for genotypic susceptibility testing of Aspergillus fumigatus.

Therapy of invasive aspergillosis is becoming more difficult due to the emergence of azole resistance in Aspergillus fumigatus. A majority of resistant strains carries mutations in the CYP51A gene. Due to a lack of sensitivity of culture-based methods, molecular detection of A. fumigatus has become an important diagnostic tool. We set up the database FunResDB (www.nrz-myk.de/funresdb) to gather all available information about CYP51A-dependent azole resistance from published literature. In summary, the screening resulted in 79 CYP51A variants, which are linked to 59 nonsynonymous mutations. A tailor-made online sequence analysis tool allows for genotypic susceptibility testing of A. fumigatus.

Successful treatment of cerebral aspergillosis: case report of a patient with T-cell large granular lymphocytic leukemia (T-LGL).

BACKGROUND: Invasive aspergillosis involving patients with neutropenia or severe immunosuppression, such as patients with hematologic malignancies is associated with high mortality. Patients with T-cell large granular lymphocytic leukemia (T-LGL) on the other hand are considered to be less vulnerable for severe opportunistic fungal infection as their course of disease is chronic and marked by less violent cytopenia then in e.g. Aplastic Anemia. Only neutropenia is regarded as independent risk factor for severe opportunistic infection in T-LGL patients. CASE PRESENTATION: We report a case of a 53 year old patient with T-LGL, Immune-Thrombocytopenia (ITP) and combined antibody deficiency, who presented with fever and reduced general condition. The patient revealed a complicated infection involving the lungs and later the brain, with the presentation of vomiting and seizures. Broad microbiological testing of blood-, lung- and cerebrospinal fluid samples was inconclusive. In the absence of mycological proof, Aspergillus infection was confirmed by pathological examination of a brain specimen and finally successfully treated with liposomal amphotericin B and voriconazole, adopting a long-term treatment scheme. CONCLUSIONS: Beyond typical problems in the clinical practice involving fungal infections and hematologic malignancies, this case of invasive aspergillosis in a patient with T-LGL illustrates caveats in diagnosis, therapy and follow-up. Our data support careful ambulatory monitoring for patients with T-LGL, even in the absence of neutropenia. Especially those patients with combined hematologic malignancies and immune defects are at risk. Long-term treatment adhesion for 12 months with sufficient drug levels was necessary for sustained clearance from infection.