A new species concept for the clinically relevant Mucor circinelloides complex.

Mucor species are common soil fungi but also known as agents of human infections (mucormycosis) and used in food production and biotechnology. Mucor circinelloides is the Mucor species that is most frequently isolated from clinical sources. The taxonomy of Mucor circinelloides and its close relatives (Mucor circinelloides complex - MCC) is still based on morphology and mating behaviour. The aim of the present study was a revised taxonomy of the MCC using a polyphasic approach. Using a set of 100 strains molecular phylogenetic analysis of five markers (ITS, rpb1, tsr1, mcm7, and cfs, introduced here) were performed, combined with phenotypic studies, mating tests and the determination of the maximum growth temperatures. The multi-locus analyses revealed 16 phylogenetic species of which 14 showed distinct phenotypical traits and were recognised as discrete species. Five of these species are introduced as novel taxa: M. amethystinus sp. nov., M. atramentarius sp. nov., M. variicolumellatus sp. nov., M. pseudocircinelloides sp. nov., and M. pseudolusitanicus sp. nov. The former formae of M. circinelloides represent one or two separate species. In the MCC, the simple presence of well-shaped zygospores only indicates a close relation of both strains, but not necessarily conspecificity. Seven species of the MCC have been implemented in human infection: M. circinelloides, M. griseocyanus, M. janssenii, M. lusitanicus, M. ramosissimus, M. variicolumellatus, and M. velutinosus.

[The German keratomycosis registry : Initial results of a multicenter survey]. / Das Deutsche Pilz-Keratitis-Register : Erste Ergebnisse einer multizentrischen Erhebung.

BACKGROUND AND PURPOSE: Mycotic keratitis is a serious but relatively rare disease. No targeted data collection in Germany existed until the foundation of the German Pilz-Keratitis Register in 2015. PATIENTS AND METHODS: The inclusion of retrospective and prospective patients was carried out. INCLUSION CRITERIA: diagnosis confirmed by the polymerase chain reaction (PCR), culture, histology or confocal microscopy (IVCM). Collected parameters: date of symptom onset, date and method of diagnosis, risk factors, visual acuity and findings at admission and at follow-up, conservative and surgical treatment. RESULTS: By January 2018, a total of 102 eyes from the years 2000-2017 were reported from 16 centers (64.3% female, mean age 52 years, range 18-95 years). The initial diagnosis was made correctly in only 20.6% of cases. The mean time to correct diagnosis was 31.7 ±â€¯46.9 (0-296) days. The diagnosis was confirmed in cultures in 74.5%, histologically in 30.4%, by PCR in 38.2% and IVCM in 27.4%. Fungal species identified were: 36.7% Fusarium spp., 35.8% Candida spp., 6.4% Aspergillus spp. and 21.1% other. The most important risk factor was the use of contact lenses. The most commonly used antifungal agent was voriconazole (64.7%) followed by amphotericin B (37.2%). Penetrating keratoplasty was performed in 65.7% of the cases and 8.8% of the affected eyes had to be enucleated. The visual acuity of the entire study population increased from the initial 0.16 ±â€¯0.25 (0.001-1.0) decimal to 0.28 ±â€¯0.34 (0-1.0) decimal. CONCLUSION: The correct diagnosis of fungal keratitis is often significantly delayed. The treatment can be very difficult and keratoplasty is often necessary. In order to gain a better understanding of this disease, to recognize previously unknown risk factors and, if necessary, a change in the spectrum of pathogens and to identify approaches to treatment optimization, the fungal keratitis registry will be continued.

[Tintelnotia destructans: new enemy at the gates]. / Tintelnotia destructans: Ein neuer Feind vor dem Tore.

Tintelnotia destructans is a fungal species described for the first time in 2016, which can cause infections of the nails and of the cornea. We describe the second known case worldwide of Tintelnotia destructans-associated keratitis and its therapy. A good sensitivity for amphotericin B and voriconazole was demonstrated in the resistogram for the first time and the successful clinical course was confirmed. The present case study also shows the importance of intensive diagnostics in atypical microbial keratitis.

Caspofungin infusion solutions (50 mg/100 mL): chemical stability and antifungal activity against Candida spp.

BACKGROUND: Ready to use caspofungin infusion bags are centrally prepared in the Hospital Pharmacy, University Hospital of Heidelberg, for economic reasons and possibly occurring problems with drug shortages. The aim of this study was a quality control of the in-house preparation of caspofungin infusion bags and the preparation process. Caspofungin concentration with regard to chemical stability and antifungal activity of caspofungin preparations were defined as quality parameters. METHODS: Three caspofungin infusion bags (50 mg in 100 mL 0.9% sodium chloride) were examined every seven days for a total of four weeks. Chemical stability of caspofungin solutions was analyzed using a validated high performance liquid chromatography (HPLC) method. Antifungal activity was assessed by microdilution tests according to the EUCAST protocol. Additionally, concentration and sterility were determined in returned caspofungin infusion bags. RESULTS: The amount of caspofungin in the infusion solutions still exceeded 90% after four weeks (2-8 °C). Antifungal activity was consistent over 28 days with a MIC ≤2 mg/L for different Candida spp. In returned infusion bags, caspofungin concentration was found to be ≥90% in 12 out of 13 bags and sterility was given in all preparations. CONCLUSION: These results show that chemical stability of caspofungin infusion solutions (50 mg/100 mL) can be guaranteed for four weeks at 2-8 °C and are confirmed by corresponding results regarding sterility and antifungal activity.

DNA barcoding of clinically relevant Cunninghamella species.

Mucormycosis caused, in part, by representatives of the genus Cunninghamella is a severe infection with high mortality in patients with impaired immunity. Several species have been described in the literature as etiologic agents. A DNA barcoding study using ITS rDNA and tef-1α provided concordance of molecular data with conventional characters. The currently accepted Cunninghamella species were well supported in phylogenetic trees of both markers except for C. septata with ITS that clustered in the C. echinulata clade. Sequence variability was distinctly higher for the ITS than for tef-1α. Intraspecific ITS variability of some of the species exceeded that between some closely related species, but the marker remained applicable for species identification. The most variable species for both markers was C. echinulata. Cunninghamella bertholletiae is the main pathogenic species; infections by C. blakesleeana, C. echinulata, and C. elegans are highly exceptional.

Effects of three postexercice recovery treatments on femoral artery blood flow kinetics.

AIM: This study aimed to compare the kinetics of muscle leg blood flow during three recovery treatments following a prolonged exercise: contrast water therapy (CWT), compression stockings (CS) or passive recovery (PR). METHODS: Fifteen men came to the laboratory three times to perform a 45-min exercise followed 5 min after by a standardized 12-min recovery treatment in upright position, alternating between two vats every 2 min: CWT (cold: ~12 °C to warm: 36 °C), CS (~20 mmHg) or PR. The order of treatments was randomized. Blood flow was measured using Doppler ultrasound during the recovery treatments (i.e., min 3, 5, 7 and 9) in the superficial femoral artery distally to the common bifurcation (~3 cm) (above the water and stocking). RESULTS: Blood flow was significantly higher during CWT (P<0.01; +22.91%) and CS (P<0.05; +15.26%) than during PR. Although no statistical difference between CWT and CS was observed, effect sizes were larger during CWT (large) than during CS (moderate). No changes in blood flow occurred in the femoral artery between hot and cold transitions of CWT. CONCLUSION: During immediate recovery of a high intensity exercise, CWT and CS trigger higher femoral artery blood flow than PR. Moreover, effect sizes were greater during CWT than during CS.

Effect of antioxidant vitamin supplementation on endothelial function in type 2 diabetes mellitus: a systematic review and meta-analysis of randomized controlled trials.

Controversy exists among trials assessing whether prolonged antioxidant vitamin supplementation improves endothelial function in type 2 diabetes mellitus (T2DM) subjects. The aim of this study was to systematically review and quantify the effect of antioxidant vitamin supplementation on endothelial function in T2DM subjects. MEDLINE, Cochrane, Scopus and Web of Science were searched up to February 2013 for randomized controlled trials assessing the effect of antioxidant vitamin E and/or C supplementation on endothelial function in T2DM subjects. Ten randomized controlled trials comparing antioxidant vitamin-supplemented and control groups (overall n = 296) met the inclusion criteria. Post-intervention standardized mean difference (SMD) in endothelial function did not reach statistical significance between groups (0.35; 95% confidence interval = -0.17, 0.88; P = 0.18). In subgroup analysis, post-intervention endothelial function was significantly improved by antioxidant vitamin supplementation in T2DM subgroups with body mass index (BMI) ≤ 29.45 kg m(-2) (SMD = 1.02; P < 0.05), but not in T2DM subgroups with BMI > 29.45 kg m(-2) (SMD = -0.07; P = 0.70). In meta-regression, an inverse association was found between BMI and post-intervention SMD in endothelial function (B = -0.024, P = 0.02). Prolonged antioxidant vitamin E and/or C supplementation could be effective to improve endothelial function in non-obese T2DM subjects.

DNA barcoding in Mucorales: an inventory of biodiversity.

The order Mucorales comprises predominantly fast-growing saprotrophic fungi, some of which are used for the fermentation of foodstuffs but it also includes species known to cause infections in patients with severe immune or metabolic impairments. To inventory biodiversity in Mucorales ITS barcodes of 668 strains in 203 taxa were generated covering more than two thirds of the recognised species. Using the ITS sequences, Molecular Operational Taxonomic Units were defined by a similarity threshold of 99 %. An LSU sequence was generated for each unit as well. Analysis of the LSU sequences revealed that conventional phenotypic classifications of the Mucoraceae are highly artificial. The LSU- and ITS-based trees suggest that characters, such as rhizoids and sporangiola, traditionally used in mucoralean taxonomy are plesiomorphic traits. The ITS region turned out to be an appropriate barcoding marker in Mucorales. It could be sequenced directly in 82 % of the strains and its variability was sufficient to resolve most of the morphospecies. Molecular identification turned out to be problematic only for the species complexes of Mucor circinelloides, M. flavus, M. piriformis and Zygorhynchus moelleri. As many as 12 possibly undescribed species were detected. Intraspecific variability differed widely among mucorealean species ranging from 0 % in Backusella circina to 13.3 % in Cunninghamella echinulata. A high proportion of clinical strains was included for molecular identification. Clinical isolates of Cunninghamella elegans were identified molecularly for the first time. As a result of the phylogenetic analyses several taxonomic and nomenclatural changes became necessary. The genus Backusella was emended to include all species with transitorily recurved sporangiophores. Since this matched molecular data all Mucor species possessing this character were transferred to Backusella. The genus Zygorhynchus was shown to be polyphyletic based on ITS and LSU data. Consequently, Zygorhynchus was abandoned and all species were reclassified in Mucor. Our phylogenetic analyses showed, furthermore, that all non-thermophilic Rhizomucor species belong to Mucor. Accordingly, Rhizomucor endophyticus was transferred to Mucor and Rhizomucor chlamydosporus was synonymised with Mucor indicus. Lecto-, epi- or neotypes were designated for several taxa.

The family structure of the Mucorales: a synoptic revision based on comprehensive multigene-genealogies.

The Mucorales (Mucoromycotina) are one of the most ancient groups of fungi comprising ubiquitous, mostly saprotrophic organisms. The first comprehensive molecular studies 11 yr ago revealed the traditional classification scheme, mainly based on morphology, as highly artificial. Since then only single clades have been investigated in detail but a robust classification of the higher levels based on DNA data has not been published yet. Therefore we provide a classification based on a phylogenetic analysis of four molecular markers including the large and the small subunit of the ribosomal DNA, the partial actin gene and the partial gene for the translation elongation factor 1-alpha. The dataset comprises 201 isolates in 103 species and represents about one half of the currently accepted species in this order. Previous family concepts are reviewed and the family structure inferred from the multilocus phylogeny is introduced and discussed. Main differences between the current classification and preceding concepts affects the existing families Lichtheimiaceae and Cunninghamellaceae, as well as the genera Backusella and Lentamyces which recently obtained the status of families along with the Rhizopodaceae comprising Rhizopus, Sporodiniella and Syzygites. Compensatory base change analyses in the Lichtheimiaceae confirmed the lower level classification of Lichtheimia and Rhizomucor while genera such as Circinella or Syncephalastrum completely lacked compensatory base changes.

Endothelial dysfunction, inflammation, and oxidative stress in obese children and adolescents: markers and effect of lifestyle intervention.

With an increasing prevalence, pediatric obesity is often a prelude to adulthood obesity, and represents a major public health issue. Comorbidities are very common and severe in obese adults, justifying the search for earlier markers or risk factors for cardiovascular diseases in obese children. Endothelial dysfunction has been found to be present in the early stages of atherosclerosis, and can be non-invasively assessed with widely accepted and well-standardized techniques at the macrocirculation level. Endothelial dysfunction at the microcirculation level is less documented in obese children. Obesity in children has been repeatedly and independently correlated to endothelial dysfunction, inflammation and oxidative stress markers, although the relationship between these factors remains to be investigated. However, this would not only allow substantial improvements in risk stratification, but also provide essential data regarding the evolution of endothelial dysfunction in childhood obesity, especially during puberty when pro-inflammatory and pro-oxidative changes, with relative insulin resistance, occur. Therapeutic strategies such as lifestyle interventions in early childhood obesity appear all the more necessary, optimally including both exercise and diet because of their known effects on inflammatory and oxidative stress markers, potentially reversing endothelial dysfunction.

Vascular reactivity at rest and during exercise in middle-aged obese men: effects of short-term, low-intensity, exercise training.

OBJECTIVE: Although increased blood flow (BF) in exercising muscles is thought to be impaired in obese subjects and may contribute to physical inactivity, data are scarce in this regard and the involvement of endothelium dysfunction remains partly hypothetical. METHODS: A total of 16 middle-aged obese men (body mass index, BMI ≥ 30 kg m(-2)) and 16 normal-weight men (BMI<25 kg m(-2)), matched for age, were recruited. We used ultrasonography to compare intima-media thickness (IMT) and distensibility of the carotid artery, flow-mediated dilation (FMD), nitrate-dependent dilation (NDD) and peak BF during post-ischemic hyperemia in the brachial artery (a conduit artery), and leg BF during knee-extensor exercise (indicative of resistance vessel function) in obese and in normal-weight men. In addition, 10 obese men participated in an 8 week individualized low-intensity training program. RESULTS: Compared with normal-weight men, obese men had higher carotid IMT (0.50 ± 0.01 vs 0.62 ± 0.04 mm, P < 0.05) but lower carotid distensibility (0.26 ± 0.03 vs 0.11 ± 0.03 mm Hg(-1) 10(-2), P < 0.05), FMD (5.7 ± 0.4 vs 3.3 ± 0.5%, P < 0.05) and peak BF during post-ischemic hyperemia (398 ± 52 vs 229 ± 24%, P < 0.05), despite similar maximal shear rate, without NDD differences. Lower limb BF (ml min(-1) 100 g(-1)) increased significantly from rest to maximal exercise in both groups with lower values in obese men (at peak power, 36.9 ± 1.6 vs 31.5+2.2 ml min(-1) 100 g(-1), P < 0.05). Exercise training normalized carotid distensibility (0.14 ± 0.04 before vs 0.23 ± 0.03 mm Hg(-1) 10(-2) after training, P = 0.09) and FMD (2.7 ± 0.4 before vs 4.8 ± 0.5% after training, P < 0.05), but did not improve brachial post-ischemic peak BF or exercising leg BF. CONCLUSIONS: In obese men, conduit and resistance vessel reactivity is depressed, but a short-term low-intensity exercise training improves distensibility and endothelium dependent vasodilation in the large conduit artery, but not post ischemic or exercise muscle BF.

Flow-mediated dilation and exercise-induced hyperaemia in highly trained athletes: comparison of the upper and lower limb vasculature.

AIM: The main purpose of the present study was to assess whether similar vascular adaptive changes could be obtained by long-term intensive training involving predominantly either the lower or the upper limb musculature. METHODS: In 11 cyclists (C), 10 swimmers (S) and 10 sedentary controls (Sed), duplex Doppler ultrasonography was used to measure post-occlusion endothelium-dependent flow-mediated dilation (FMD), endothelium-independent, glycerine trinitrate-induced dilation (GTND) and exercise-induced blood flow changes in the arm (axillary artery) and leg (superficial femoral artery). Limb-specific exercise was achieved by one elbow-flexion or one leg-extension maximal exercise test, thereby allowing assessment of upper and lower limb muscle perfusion, vascular conductance and vasodilatory capacity of resistance vessels during effort. RESULTS: C and S exhibited vascular remodelling associated with improved FMD and GTND in the predominantly trained limbs compared to Sed. Both showed greater muscle perfusion and vascular conductance than Sed during isolated exercise involving the predominantly trained musculature. C showed also higher FMD in the brachial artery and greater peak muscle perfusion and conductance in the non-exercising muscles, whereas S presented only enhanced FMD in the superficial femoral artery. CONCLUSION: Therefore, in the upper as well as in the lower limb vasculature, repetitive exposure to increased shear stress over a long-term period results in improved FMD of large conduit arteries as well as greater vasodilatory capacity during isolated exercise in the predominantly trained muscles. Long-term training involving predominantly the lower limbs also results in enhanced vascular reactivity in upper limb conduit and resistance vessels.

Reproducibility of automated pulse wave velocity measurement during exercise. Running head: pulse wave velocity during exercise.

Pulse wave velocity measurement is used as an index of arterial stiffness. The purpose was to evaluate the reproducibility of pulse wave velocity measurement at rest, during exercise and recovery from exercise, using an automated device. Twelve healthy young adults (mean age 22.0 +/- 3.1 yrs) underwent an upright submaximal cycle test on two separate occasions, one week apart. Pulse wave velocity, systolic and diastolic blood pressures and heart rate were assessed at rest, during the last 2 min of exercise and 10 min later. Pulse wave velocity was measured on the upper limb and the forearm by the cross-correlation function of photoplethysmography and Doppler signals. Brachial artery pulse wave velocity was calculated from upper limb and forearm pulse wave velocities. No significant difference was found on duplicate measurements of heart rate, systolic and diastolic blood pressures at rest, during exercise and recovery, showing that pulse wave velocity was measured under similar conditions. Coefficient of variation for upper limb and forearm pulse wave velocities ranged from 2.9 to 5.9% at rest and during recovery, and were respectively 2.9% and 8.3% during exercise. However, coefficient of variation for brachial pulse wave velocity was 7.7 and 10.3% at rest, 15.7% during exercise, and 5.8% during recovery. During exercise, pulse wave velocity measurements were satisfying, but indirect assessment of brachial artery pulse wave velocity showed poor reproducibility. Thus, upper limb and forearm pulse wave velocities may be used during exercise to assess the effect of training or drugs on arterial wall mechanical properties.

Factors affecting outcome in penetrating oesophageal trauma.

BACKGROUND: Penetrating oesophageal trauma is rare and the risk factors affecting outcome have not been clearly identified. Delayed management has been cited as a factor contributing to the high rates of morbidity and mortality, but evidence for this is lacking. METHODS: A retrospective study was undertaken of patients with penetrating oesophageal trauma presenting to a level I trauma centre over 8 years. Outcome was assessed in terms of mortality, morbidity (oesophageal and non-oesophageal), and length of hospital and intensive care unit (ICU) stays. RESULTS: Fifty-two patients with oesophageal injury who reached the operating theatre were included. The overall mortality rate was 6 per cent. Fifteen patients (29 per cent) developed oesophageal injury-related complications. Time from injury to management was the only important risk factor for the development of oesophageal complications (P = 0.001), but did not affect the length of ICU (P = 0.560) or hospital (P = 0.329) stay, incidence of non-oesophageal injury-related complications (P = 0.963) or death (P = 0.937). Patients with gunshot injuries spent longer in the ICU (P = 0.007) and the duration of hospital stay was longer for those with higher-grade oesophageal injuries (P = 0.025). CONCLUSION: The risk of oesophageal injury-related complications was directly related to the interval between the trauma and definitive management of the oesophageal injury.

Excision of a giant hydatid cyst of the lung under thoracic epidural anaesthesia.

We present a patient with a large pulmonary hydatid cyst compressing underlying lung, with previous pulmonary tuberculosis, who presented in respiratory failure. After institution of thoracic epidural anaesthesia employing 0.25% bupivacaine, 1% lignocaine and fentanyl, the patient was placed in the sitting position and the hydatid cyst excised and drained after a limited rib resection. An air leak persisted until the 16th postoperative day. A marked improvement in symptoms as well as in spirometly and arterial blood gases occurred, and the patient was discharged on the 20th day. Thoracic epidural anaesthesia may be a safer method than general anaesthesia for removal of a hydatid cyst in a patient with severe respiratory compromise.

Thoracoscopy in undiagnosed pleural effusions.

OBJECTIVE: To review the indications and accuracy of diagnostic thoracoscopy for pleural effusions of unknown origin. DESIGN: Retrospective review of consecutive patients referred for diagnostic thoracoscopy over a 5-year period from 1 January 1989 to 31 December 1993. SETTING: Tertiary referral cardiothoracic unit. PATIENTS: Thirty-four patients referred from either medical or oncology services within a university-affiliated academic complex. INTERVENTIONS: All patients had diagnostic thoracoscopy performed under general anaesthesia. Retrospective data were collected in respect of presenting symptoms, gross findings, final pathological findings, amount of drainage, length of hospital stay and complications of the procedure. In 7 patients (21%), iodised talc was insufflated at the same time to create pleurodesis. MAIN RESULTS: Final diagnoses were: 17 (50%) malignant disease, 6 (18%) tuberculosis and 9 (26%) 'negative' pathology. In 2 (6%), further intervention was required to make a conclusive diagnosis. The diagnostic sensitivity for malignant disease was 89% and the specificity 100%. For pleural tuberculosis both the sensitivity and specificity were 100%. For 'negative' diagnoses the negative predictive value was 82%. A history of fever and sweats had a marked association (P = 0.002) with the final diagnosis of tuberculosis. No association could be identified between the gross observations at the time of thoracoscopy and the final diagnosis. The average length of hospital stay was 6.7 (range 1-25) days. There was 1 in-hospital death (3%), and 9 patients (26%) had major complications related to the procedure. CONCLUSIONS: Diagnostic thoracoscopy is a useful modality for obtaining a diagnosis in effusions of unknown origin where other methods have failed. The presence of symptoms such as fever and sweats is highly associated with a final diagnosis of tuberculosis.

[Cell therapy and its risks]. / Zelltherapie und ihre Gefahren.

Parenteral administration of fresh cells, frozen cells (snapfrozen cell suspensions) or lyophilized cells (sicca cells), is known as cellular or cell therapy. While the German Health Office (BGA) provisionally banned the use of dried cell preparations in 1987, injection of fresh cells is still allowed. There have been repeated reports of life-threatening, and even fatal, complications of this type of therapy. Since it involves the administration of heterologous biological material, most of the complications that have been observed have been of the allergic/hyperergic type resembling experimental allergic encephalomyelitis (EAE) and experimental allergic neuritis (EAN). When evaluating the risks of this form of alternative-medical treatment, the well-known risks of injection therapy must also be borne in mind. In the case of cell therapy, too, the hoped for effect must be weighed against the risks of the procedure, and our guiding principle must be: nil nocere.

WAL 2014--a muscarinic agonist with preferential neuron-stimulating properties.

The ability of WAL 2014 to elicit muscarinic responses was investigated in various in vitro and in vivo models. In CHO cells transfected with human m1- or m3- receptor genes, WAL 2014 was clearly more effective in stimulating the M1-mediated PI response. In isolated tissue preparations, WAL 2014 exhibited full agonist properties in the rabbit vas deferens (putative M1 receptor) and behaved like a partial agonist at M2 receptors in the atrium and M3 receptors in the ileum of guinea-pigs. In the pithed rat, in which the increase in blood pressure is mediated through a stimulation of M1 receptors in sympathetic ganglia, WAL 2014 produced a full dose response curve, whereas the reference compounds RS 86 and arecoline exhibited a bell-shaped behaviour. This is in accord with the view that WAL 2014 selectively activates M1 receptors in sympathetic ganglia, whereas conventional agonists in the same dose range stimulate both ganglionic M1 and vascular M3 receptors. The preferential neuron-stimulating properties were confirmed by EEG recording in the rabbit, in which muscarinic activation occurred at doses similar to those for ganglionic stimulation in the pithed rat. On the other hand, higher doses of WAL 2014 were needed to elicit muscarinic effects in peripheral effector organs, i.e. bradycardia, urinary bladder contraction and increase in airway resistance. It is concluded that WAL 2014 due to its preferential neuronal activity is a promising candidate for a cholinergic substitution therapy in Alzheimer's disease.