A single-source nosocomial outbreak of Aspergillus flavus uncovered by genotyping.

During construction work (2017-2019), an increase in Aspergillus flavus infections was noted among pediatric patients, the majority of whom were receiving amphotericin B prophylaxis. Microsatellite genotyping was used to characterize the outbreak. A total of 153 A. flavus isolates of clinical and environmental origin were included. Clinical isolates included 140 from 119 patients. Eight patients were outbreak-related patients, whereas 111 were outbreak-unrelated patients from Danish hospitals (1994-2023). We further included four control strains. Nine A. flavus isolates were from subsequent air sampling in the outbreak ward (2022-2023). Typing followed Rudramurthy et al.(S. M. Rudramurthy, H. A. de Valk, A. Chakrabarti, J. Meis, and C. H. W. Klaassen, PLoS One 6:e16086, 2011, https://doi.org/10.1371/journal.pone.0016086). Minimum spanning tree (MST) and discriminant analysis of principal components (DAPC) were used for cluster analysis. DAPC analysis placed all 153 isolates in five clusters. Microsatellite marker pattern was clearly distinct for one cluster compared to the others. The same cluster was observed in an MST. This cluster included all outbreak isolates, air-sample isolates, and additional patient isolates from the outbreak hospital, previously undisclosed as outbreak related. The highest air prevalence of A. flavus was found in two technical risers of the outbreak ward, which were then sealed. Follow-up air samples were negative for A. flavus. Microsatellite typing defined the outbreak as nosocomial and facilitated the identification of an in-hospital source. Six months of follow-up air sampling was without A. flavus. Outbreak-related/non-related isolates were easily distinguished with DAPC and MST, as the outbreak clone's distinct marker pattern was delineated in both statistical analyses. Thus, it could be a variant of A. flavus, with a niche ability to thrive in the outbreak-hospital environment. IMPORTANCE: Aspergillus flavus can cause severe infections and hospital outbreaks in immunocompromised individuals. Although lack of isogeneity does not preclude an outbreak, our study underlines the value of microsatellite genotyping in the setting of potential A. flavus outbreaks. Microsatellite genotyping documented an isogenic hospital outbreak with an internal source. This provided the "smoking gun" that prompted the rapid allocation of resources for thorough environmental sampling, the results of which guided immediate and relevant cleaning and source control measures. Consequently, we advise that vulnerable patients should be protected from exposure and that genotyping be included early in potential A. flavus outbreak investigations. Inspection and sampling are recommended at any site where airborne spores might disperse from. This includes rarely accessed areas where air communication to the hospital ward cannot be disregarded.

Resolution of eleven reported and five novel Podaxis species based on ITS phylogeny, phylogenomics, morphology, ecology, and geographic distribution.

The genus Podaxis was first described from India by Linnaeus in 1771, but several revisions of the genus have left the taxonomy unclear. Forty-four Podaxis species names and nine intraspecific varieties are currently accepted, but most fungarium specimens are labelled Podaxis pistillaris. Recent molecular analyses based on barcoding genes suggest that the genus comprises several species, but their status is largely unresolved. Here we obtained basidiospores and photographs from 166 fungarium specimens from around the world and generated a phylogeny based on rDNA internal transcribed spacer ITS1,5.8S and ITS2 (ITS), and a phylogenomic analysis of 3 839 BUSCO genes from low-coverage genomes for a subset of the specimens. Combining phylogenetics, phylogenomics, morphology, ecology, and geographical distribution, spanning 250 years of collections, we propose that the genus includes at least 16 unambiguous species. Based on 10 type specimens (holotype, paratype, and syntype), four recorded species were confirmed, P. carcinomalis, P. deflersii, P. emerici, and P. farlowii. Comparing phylogenetic analysis with described species, including morphology, ecology, and distribution, we resurrected P. termitophilus and designated neotypes, epitypes, or lectotypes for five previously described species, P. aegyptiacus, P. africana, P. beringamensis, P. calyptratus, and P. perraldieri. Lastly, based on phylogenies and morphology of type material, we synonymized three reported species, P. algericus, P. arabicus, and P. rugospora with P. pistillaris, and described five new species that we named P. desolatus, P. inyoensis, P. mareebaensis, P. namaquensis, and P. namibensis. Citation: Li GS, Leal-Dutra CA, Cuesta-Maté A, et al. 2023. Resolution of eleven reported and five novel Podaxis species based on ITS phylogeny, phylogenomics, morphology, ecology, and geographic distribution. Persoonia 51: 257-279. doi: 10.3767/persoonia.2023.51.07.

PATIENT-SPECIFIC DOSE ESTIMATES IN DYNAMIC COMPUTED TOMOGRAPHY MYOCARDIAL PERFUSION EXAMINATION.

The study aimed to implement realistic source models of a computed tomography (CT) scanner and Monte Carlo simulations to actual patient data and to calculate patient-specific organ and effective dose estimates for patients undergoing dynamic CT myocardial perfusion examinations. Source models including bowtie filter, tube output and x-ray spectra were determined for a dual-source Siemens Somatom Definition Flash scanner. Twenty CT angiography patient datasets were merged with a scaled International Commission on Radiological Protection (ICRP) 110 voxel phantom. Dose simulations were conducted with ImpactMC software. Effective dose estimates varied from 5.0 to 14.6 mSv for the 80 kV spectrum and from 8.9 to 24.7 mSv for the 100 kV spectrum. Significant differences in organ doses and effective doses between patients emphasise the need to use actual patient data merged with matched anthropomorphic anatomy in the dose simulations to achieve a reasonable level of accuracy in the dose estimation procedure.

CT beam dosimetric characterization procedure for personalized dosimetry.

Personalized dosimetry in computed tomography (CT) can be realized by a full Monte Carlo (MC) simulation of the scan procedure. Essential input data needed for the simulation are appropriate CT x-ray source models and a model of the patient's body which is based on the CT image. The purpose of this work is to develop comprehensive procedures for the determination of CT x-ray source models and their verification by comparison of calculated and measured dose distributions in physical phantoms. Mobile equipment together with customized software was developed and used for non-invasive determination of equivalent source models of CT scanners under clinical conditions. Standard and physical anthropomorphic CT dose phantoms equipped with real-time CT dose probes at five representative positions were scanned. The accumulated dose was measured during the scan at the five positions. ImpactMC, an MC-based CT dose software program, was used to simulate the scan. The necessary inputs were obtained from the scan parameters, from the equivalent source models and from the material-segmented CT images of the phantoms. 3D dose distributions in the phantoms were simulated and the dose values calculated at the five positions inside the phantom were compared to measured dose values. Initial results were obtained by means of a General Electric Optima CT 660 and a Toshiba (Canon) Aquilion ONE. In general, the measured and calculated dose values were within relative uncertainties that had been estimated to be less than 10%. The procedures developed were found to be viable and rapid. The procedures are applicable to any scanner type under clinical conditions without making use of the service mode with stationary x-ray tube position. Results show that the procedures are well suited for determining and verifying the equivalent source models needed for personalized CT dosimetry based on post-scan MC calculations.

Determination of the separation efficiencies of a single-stage cryogenic distillation setup to remove krypton out of xenon by using a (83m)Kr tracer method.

The separation of krypton and xenon is of particular importance for the field of direct dark matter search with liquid xenon detectors. The intrinsic contamination of the xenon with radioactive (85)Kr makes a significant background for these kinds of low count-rate experiments and has to be removed beforehand. This can be achieved by cryogenic distillation, a technique widely used in industry, using the different vapor pressures of krypton and xenon. In this paper, we present an investigation on the separation performance of a single stage distillation system using a radioactive (83m)Kr-tracer method. The separation characteristics under different operation conditions are determined for very low concentrations of krypton in xenon at the level of (83m)Kr/Xe = 1.9 ⋅ 10(-15), demonstrating, that cryogenic distillation in this regime is working. The observed separation is in agreement with the expectation from the different volatilities of krypton and xenon. This cryogenic distillation station is the first step on the way to a multi-stage cryogenic distillation column for the next generation of direct dark matter experiment XENON1T.

Identification of mycorrhizal fungi from single pelotons of Dactylorhiza majalis (Orchidaceae) using single-strand conformation polymorphism and mitochondrial ribosomal large subunit DNA sequences.

The mitochondrial ribosomal large subunit (Ls) DNA was used to identify the orchid mycorrhizal fungi found in roots of Dactylorhiza majalis. The gene was amplified using DNA extracted from single pelotons obtained from fresh and silica gel dried roots. Furthermore, sequencing a variety of well-characterized orchid isolates expanded the fungal database of the mitochondrial ribosomal LsDNA. Polymerase chain reaction product length variants present in D. majalis were sequenced and identified using the expanded database. These analyses revealed two different peloton-forming fungi in samples from D. majalis, which sometimes occurred together as a single two-taxa peloton within the same cortex cell. The first taxon belonged to the genus Tulasnella and the second taxon was distantly related to Laccaria.

Bacteria-induced release of white cell--and platelet-derived vascular endothelial growth factor in vitro.

BACKGROUND AND OBJECTIVES: Poor prognosis after resection of primary colorectal cancer may be related to the combination of perioperative blood transfusion and subsequent development of infectious complications. White blood cell--and platelet-derived cancer growth substances, including vascular endothelial growth factor (VEGF), may be involved in this process. Therefore, we studied the in vitro release of VEGF from white blood cells and platelets stimulated by bacterial antigens and supernatants from stored red cell components. MATERIALS AND METHODS: Eight units of whole blood (WB) and eight units of buffy-coat-depleted red cell (SAGM) blood were donated by healthy blood donors. Subsequently, half of every unit was leucocyte depleted by filtration, and all 32 half-units were stored under standard conditions for 35 days. Just after storage, and on days 7, 21 and 35 during storage, aliquots of the supernatants were removed from the units and frozen at -80 degrees C. WB from other healthy donors was stimulated for 2 h with sodium chloride (controls), with Escherichia coli lipopolysaccharide (LPS) alone, or with LPS plus supernatants from the non-filtered or prestorage leucofiltered WB units (diluted 1:10), or from non-filtered or prestorage leucofiltered SAGM blood units (diluted 1:20) stored for 0, 7, 21, or 35 days, respectively. Similar assays were performed using Staphylococcus aureus-derived protein A as a stimulatory antigen. The concentration of VEGF was determined in supernatants from stored blood and in assay supernatants by using enzyme-linked immunosorbent assay (ELISA). RESULTS: The concentration of VEGF increased significantly (P < 0.0001) in a storage time-dependent manner in non-filtered WB and SAGM blood, while the increase was abrogated by prestorage leucofiltration. The supernatant concentration of VEGF was significantly increased in LPS-stimulated (P = 0.002) and in protein A-stimulated (P < 0.0001) assays compared with controls. Addition of supernatants from stored, non-filtered WB or SAGM significantly increased the assay supernatant VEGF concentration storage-time dependently (P = 0.006) in LPS assays. In protein A assays, only supernatants from non-filtered WB significantly increased the assay supernatant VEGF concentration storage-time dependently (P = 0.022). This additional effect by supernatants from stored blood components was not observed with prestorage leucofiltered blood. CONCLUSIONS: Extracellular VEGF may accumulate in non-filtered red cell components, but this can be prevented by prestorage leucocyte depletion using filtration. In addition, bacterial antigens appear to induce release of VEGF from white blood cells and platelets. Addition of supernatants from stored, non-filtered WB or SAGM blood may increase the VEGF levels in a storage time-dependent manner, while prestorage leucofiltration may prevent further increase by supernatants.

Evolution of Fusarium oxysporum f. sp. vasinfectum Races Inferred from Multigene Genealogies.

ABSTRACT Fusarium wilt of cotton is a serious fungal disease responsible for significant yield losses throughout the world. Evolution of the causal organism Fusarium oxysporum f. sp. vasinfectum, including the eight races described for this specialized form, was studied using multigene genealogies. Partial sequences of translation elongation factor (EF-1alpha), nitrate reductase (NIR), phosphate permase (PHO), and the mitochondrial small subunit (mtSSU) rDNA were sequenced in 28 isolates of F. oxysporum f. sp. vasinfectum selected to represent the global genetic diversity of this forma specialis. Results of a Wilcoxon Signed-Ranks Templeton test indicated that sequences of the four genes could be combined. In addition, using combined data from EF-1alpha and mtSSU rDNA, the phylogenetic origin of F. oxysporum f. sp. vasinfectum within the F. oxysporum complex was evaluated by the Kishino-Hasegawa likelihood test. Results of this test indicated the eight races of F. oxysporum f. sp. vasinfectum appeared to be nonmonophyletic, having at least two independent, or polyphyletic, evolutionary origins. Races 3 and 5 formed a strongly supported clade separate from the other six races. The combined EF-1alpha, NIR, PHO, and mtSSU rDNA sequence data from the 28 isolates of F. oxysporum f. sp. vasinfectum recovered four lineages that correlated with differences in virulence and geographic origin: lineage I contained race 3, mostly from Egypt, and race 5 from Sudan; lineage II contained races 1, 2, and 6 from North and South America and Africa; lineage III contained race 8 from China; and lineage IV contained isolates of races 4 and 7 from India and China, respectively.

Relationships of the insect-pathogenic order Entomophthorales (Zygomycota, Fungi) based on phylogenetic analyses of nuclear small subunit ribosomal DNA sequences (SSU rDNA).

We sequenced the nuclear small subunit of ribosomal DNA (SSU rDNA) from seven species within the insect-pathogenic order Entomophthorales. These sequences were aligned with other published SSU rDNA sequences and phylogenies were inferred using phenetic and cladistic methods. Based on three different phylogenetic methods the Entomophthorales (excluding Basidiobolus ranarum) is monophyletic; B. ranarum was more closely related to chytrids from Chytridiales and Neocallimasticales than to Entomophthorales, as was proposed by Nagahama et al. (Mycologia 87: 203-209, 1995). Nuclear characters (large nuclei containing conspicuous condensed chromatin and lack of a prominent nucleolus) were of predictive value for the monophyly of the family Entomophthoraceae. Conidial characters separate the Entomophthoraceae, which only includes obligate pathogens, into at least two lineages: one lineage with uninucleate conidia and another with multinucleate conidia. The two species of Conidiobolus studied were paraphyletic in our analyses and only distantly related to each other. This information may prove to be important in the use of these fungi as biocontrol agents.

Enzymatic Activity of the Mycelium Compared with Oospore Development During Infection of Pea Roots by Aphanomyces euteiches.

ABSTRACT To describe the disease cycle of the root pathogen Aphanomyces euteiches, enzymatic activity in the mycelium was compared with the development of oospores in pea roots. Plants were inoculated with two zoospore concentrations to achieve different disease levels. Hyphae were stained for fungal alkaline phosphatase activity in the roots. Additionally, enzyme activity was measured after electrophoresis of an A. euteiches-specific glucose-6-phosphate isozyme. Development of oospores in the roots was measured after staining the oospores with trypan blue. In plants inoculated with the higher zoospore concentration, the enzymatic activity of the pathogen mycelium peaked 10 to 14 days after inoculation, when oospore formation was initiated. Oospore formation was associated with a gradual increase in disease symptoms. At the last harvest, plants inoculated with the higher zoospore concentration had died. In these plants, oospores were found in 90% of the root length, while the enzymatic activity of the mycelium was low. This suggests that the pathogen mycelium is only active on living plants and does not grow saprophytically on dead plant material.

Benomyl inhibits phosphorus transport but not fungal alkaline phosphatase activity in a Glomus-cucumber symbiosis.

Short-term effects of benomyl on the arbuscular mycorrhizal fungus Glomus caledonium (Nicol. & Gerd.) Trappe and Gerdeman associated with Cucumis sativus L. were studied by measuring effects on fungal P transport and on fungal alkaline phosphatase activity. Mycorrhizal plants were grown in three compartment systems where nylon mesh was used to separate n root-free hyphal compartment (HC) and a root + hyphal compartment(RHC) from The main root compartment (RC). Non-mycorrhizal control plants were grown in similar growth units. After 6 wk benomyl was applied to the plants in three ways: as soil drenches to RHC or HC, or as u spray to the leaves. Benomyl was added in three concentrations. Equal amounts of 32 P and 33 P were added to the HC and to the RHC respectively, immediately after the application of benomyl. Plants were harvested 4-6 d later. Hyphal transport of 32 P from the HC was inhibited when benomyl was applied to the HC at 10 µg g-1 soil, whereas the uptake of 32 P from RHC I roots + hyphae) was reduced only at the highest dose of application to the RHC (100 µ g g-1 soil). In contrast to the marked reduction of benomyl on fungal P transport, the activity of fungal alkaline phosphatase inside the roots was unaffected by benomyl.

Function after lower limb amputation.

Functional ability and social dependence were investigated by personal interview of 107 lower limb amputees surviving 1-5 years postoperatively. Among eight independent variables studied by multiple regression analysis, increased age was associated unfavorably with physical ability and social dependence. Independence from social provisions preoperatively showed favorable relationships with functional capacity and postoperative dependence. Above-knee or bilateral amputation and postoperative pain were associated with reduced functional ability, but not with social dependence. No significant association was found with cause of operation or sex of the amputees. The importance of proper prosthetic fitting and pain control is emphasized.

Fractures of the shaft of the tibia treated with AO-compression osteosynthesis.

Ninety-six displaced fractures of the shaft of the tibia in a series of 162 consecutive fractures were treated by AO internal fixation. Forty per cent were open fractures, of which 93 per cent received prophylactic treatment with antibiotics at the time of admission. The average time between the accident and the operation was 10 hours in closed fractures and 5 hours in open fractures. All cases were operated on by senior surgeons. The infection rate was 5.3 per cent in closed fractures, and 0 in open fractures. The average stay in hospital was 13 days. More than 90 per cent returned to work within 6 months after the accident. No case of pseudarthrosis or re-fracture was seen. The median time to final review was 36 months. Rigid internal fixation is advocated for all displaced fractures of the shaft of the tibia and is advocated as an urgent procedure especially in open fractures, and should be performed by experienced surgeons only. Rigid internal fixation appears to provide effective prophylaxis against secondary soft-tissue damage and limits the consequences of the initial soft-tissue damage.

Para-articular ossification following hip replacement. 70 arthroplasties ad modum Moore using McFarland's approach.

Seventy patients, representing all the survivors from a group of 117 consecutive Moore arthroplasties, were examined 2 years after the operation. A high incidence of para-articular ossification was observed. The ossification was associated with impaired function of the hip joint. The separation of periost and fascia from the greater trochanter in McFarland's approach may be responsible for the high incidence of bone formation. This theory is compatible with experimental evidence.