Upgrading fungal gene expression on demand: improved systems for doxycycline-dependent silencing in Aspergillus fumigatus.

Conditional gene expression is key for functional studies in any given microorganism. To allow tight regulation in the pathogenic mold Aspergillus fumigatus, improved versions of the doxycycline-dependent Tet-On system were generated by replacing functional elements of the precursor module, thereby circumventing the former problem of leakiness due to intramolecular recombination.

Functional, Spiroergometric, and Subjective Comparisons Between Forearm Crutches and Hands-Free Single Crutches in a Crossover Study.

Background: Following below-knee surgery, the optimal medical mobility device remains controversial as adequate nonweightbearing of the operated extremity is critical to ensure successful healing. The use of forearm crutches (FACs) is well established but requires using both upper extremities. The hands-free single orthosis (HFSO) is an alternative that spares the upper extremities. This pilot study compared functional, spiroergometric, and subjective parameters between HFSO and FAC. Methods: Ten healthy (5 females, 5 males) participants were asked to use HFSOs and FACs in a randomized order. Five functional tests were performed: climbing stairs (CS), an L-shaped indoor course (IC), an outdoor course (OC), a 10-meter walk test (10MWT), and a 6-minute walk test (6MWT). Tripping events were counted while performing IC, OC, and 6MWT. Spiroergometric measurements consisted of a 2-step treadmill test with speeds of 1.5 and 2 km/h, each for 3 minutes. Lastly, a VAS questionnaire was completed to collect data regarding comfort, safety, pain, and recommendations. Results: Significant differences between both aids were observed in CS and IC (HFSO: 29.3 seconds; FAC: 26.1 seconds, P < .03; and HFSO: 33.2 seconds, FAC: 18 seconds, P < .001, respectively). The other functional tests showed no significant differences. The trip events were not significantly different between the use of the 2 aids. Spiroergometric tests showed significant differences regarding heart rate (HFSO: 131.1 bpm at 1.5 km/h and 131 bpm at 2 km/h; FAC: 148.1 bpm at 1.5 km/h and 161.8 bpm at 2 km/h) and oxygen consumption (HFSO: 15.4 mL/min/kg at 1.5 km/h and 16 mL/min/kg at 2 km/h; FAC: 18.3 mL/min/kg at 1.5 km/h and 21.9 mL/min/kg at 2 km/h) at both speeds (all P < .01). In addition, significantly different ratings regarding the items comfort, pain, and recommendation were recorded. Both aids were equally rated for safety. Conclusion: HFSOs may be an alternative to FACs, especially in activities that require physical stamina. Further prospective studies in patients with below-knee surgical intervention concerning everyday clinical use would be interesting. Level of Evidence: Level IV pilot-study.

A prospective pilot study using a low power piggy-back toric implantable Collamer lens to correct residual refractive error after multifocal IOL implantation.

PURPOSE: To assess whether residual refractive error after in-the-bag multifocal intraocular lens (mIOL) implantation can efficaciously and safely be corrected with a piggy-back low power Visian Toric Implantable Collamer Lens® (VTICL, STAAR Surgical) placed in the ciliary sulcus. PATIENTS AND METHODS: Twenty-four eyes of 23 patients (mean age: 57.5 years) with diminished uncorrected distance visual acuity (UDVA) of ≥2 lines due to residual refractive error after mIOL implantation were included in the study. VTICL size was calculated using the standard STAAR Visian ICL calculation software for phakic eyes. Postoperative study visits (1 day, 1 week, 3 months and 6 months after VTICL implantation) included UDVA, corrected distance visual acuity (CDVA), VTICL axis alignment, vault (space between mIOL and VTICL), IOP and documentation of adverse events. RESULTS: At 6 months, mean UDVA (logMAR) increased from 0.26 preoperatively to -0.01 (P<0.001) while mean CDVA remained unchanged. Mean VTICL misalignment from the preoperative target axis was 5.3° and mean vault was 1385 µm. In the initial phase of the study, 2 VTICL had to be exchanged due to oversizing. CONCLUSION: Piggy-back low power VTICL can efficaciously correct residual refractive error after mIOL implantation and significantly increase UDVA. Advantages of this novel surgical approach include: VTICL availability in small diopter steps, no significant surgical-induced astigmatism, atraumatic and reversible procedure.

Influence of blurred vision, accommodation, and target laser settings on eye movements during LASIK.

PURPOSE: To analyze the influence of blurred vision, accommodation, and target laser settings on eye movements during laser in situ keratomileusis (LASIK). SETTING: Department of Ophthalmology, Goethe University, Frankfurt, Germany. DESIGN: Prospective randomized study. METHODS: Participants had simulated LASIK treatment. They were instructed to focus on the fixation light; the treatment laser was blocked, and all other settings were applied according to standard LASIK treatments. To simulate blurred vision a 0.0 diopter (D) soft contact lens received a 5.0 D myopia laser treatment and was then applied to the participant's eye. To diminish accommodation, a second lens that had a refraction of the patient's spherical equivalent, plus 3.0 D to compensate for accommodation, was used. There were 4 treatment modalities as follows: (1) blurred lens with target laser on, (2) blurred lens with target laser turned off, (3) +3.0 D lens with target laser on, and (4) +3.0 D lens with target laser turned off. Lateral and torsional eye movements were recorded. Fourier analysis was used to obtain temporal power spectra from dynamic eye movements. The Fn criterion was set as the frequency below which n% of eye movements in the cohort occurred (n = 95%, 80%, and 50%). RESULTS: The study comprised 11 eyes of 11 participants. There was 1 significant difference between the eye movements based on measurement modalities. In 1 variable in the y-axis, there was movement that showed a significant difference in the F80 criterion. CONCLUSION: Surgical circumstances such as blurred vision, accommodation, and target light had little influence on eye movements during LASIK. FINANCIAL DISCLOSURE: Proprietary or commercial disclosures are listed after the references.

Mutant characterization and in vivo conditional repression identify aromatic amino acid biosynthesis to be essential for Aspergillus fumigatus virulence.

Pathogenicity of the saprobe Aspergillus fumigatus strictly depends on nutrient acquisition during infection, as fungal growth determines colonisation and invasion of a susceptible host. Primary metabolism has to be considered as a valid target for antimycotic therapy, based on the fact that several fungal anabolic pathways are not conserved in higher eukaryotes. To test whether fungal proliferation during invasive aspergillosis relies on endogenous biosynthesis of aromatic amino acids, defined auxotrophic mutants of A. fumigatus were generated and assessed for their infectious capacities in neutropenic mice and found to be strongly attenuated in virulence. Moreover, essentiality of the complete biosynthetic pathway could be demonstrated, corroborated by conditional gene expression in infected animals and inhibitor studies. This brief report not only validates the aromatic amino acid biosynthesis pathway of A. fumigatus to be a promising antifungal target but furthermore demonstrates feasibility of conditional gene expression in a murine infection model of aspergillosis.