Improving power of association tests using multiple sets of imputed genotypes from distributed reference panels

The accuracy of genotype imputation depends upon two factors: the sample size of the reference panel and the genetic similarity between the reference panel and the target samples. When multiple reference panels are not consented to combine together, it is unclear how to combine the imputation results to optimize the power of genetic association studies. We compared the accuracy of 9,265 Norwegian genomes imputed from three reference panels—1000 Genomes phase 3 (1000G), Haplotype Reference Consortium (HRC), and a reference panel containing 2,201 Norwegian participants from the population‐based Nord Trøndelag Health Study (HUNT) from low‐pass genome sequencing. We observed that the population‐matched reference panel allowed for imputation of more population‐specific variants with lower frequency (minor allele frequency (MAF) between 0.05% and 0.5%). The overall imputation accuracy from the population‐specific panel was substantially higher than 1000G and was comparable with HRC, despite HRC being 15‐fold larger. These results recapitulate the value of population‐specific reference panels for genotype imputation. We also evaluated different strategies to utilize multiple sets of imputed genotypes to increase the power of association studies. We observed that testing association for all variants imputed from any panel results in higher power to detect association than the alternative strategy of including only one version of each genetic variant, selected for having the highest imputation quality metric. This was particularly true for lower frequency variants (MAF < 1%), even after adjusting for the additional multiple testing burden.     

EUCAST technical note on the EUCAST definitive document EDef 7.2: method for the determination of broth dilution minimum inhibitory concentrations of antifungal agents for yeasts EDef 7.2 (EUCAST-AFST)

The European Committee on Antimicrobial Susceptibility Testing-Subcommittee on Antifungal Susceptibility Testing (EUCAST-AFST) has revised the EDef 7.1 document on the method for the determination of broth dilution minimum inhibitory concentrations of antifungal agents for fermentative yeasts. Changes are: dimethylsulphoxide is now the recommended solvent for caspofungin, micafungin and fluconazole; the shelf-life of plates containing the echinocandins prepared from stock solutions in dimethylsulphoxide is extended to 6 months at ?80°C; testing of amphotericin and Cryptococcus has been incorporated; and minimum inhibitory concentration ranges for quality control strains and anidulafungin are included.     

Caspofungin Etest susceptibility testing of Candida species: risk of misclassification of susceptible isolates of C. glabrata and C. krusei when adopting the revised CLSI caspofungin breakpoints

The purpose of this study was to evaluate the performance of caspofungin Etest and the recently revised CLSI breakpoints. A total of 497 blood isolates, of which 496 were wild-type isolates, were included. A total of 65/496 susceptible isolates (13.1%) were misclassified as intermediate (I) or resistant (R). Such misclassifications were most commonly observed for Candida krusei (73.1%) and Candida glabrata (33.1%). The revised breakpoints cannot be safely adopted for these two species.     

Echinocandin susceptibility testing of Candida spp. Using EUCAST EDef 7.1 and CLSI M27-A3 standard procedures: analysis of the influence of bovine serum albumin supplementation, storage time, and drug lots

The MICs of echinocandins against Candida isolates with fks mutations are higher than those for wild-type (WT) isolates. However, the MIC ranges for susceptible and mutant populations overlap or are poorly separated. It was recently reported that a greater separation could be achieved in the presence of serum. To more fully explore this possibility, we compared the performances of the reference microdilution methods by using standard and bovine serum albumin (BSA)-supplemented growth medium. Anidulafungin, caspofungin, and micafungin MICs were determined according to EUCAST and CLSI methods and with 50% BSA in the medium for 93 clinical isolates, including Candida albicans (20/10 [number of isolates/number of mutants]), C. glabrata (19/10), C. dubliniensis (2/1), C. krusei (16/3), C. parapsilosis (19), and C. tropicalis (19/4) isolates. Stability of the plates was tested after storage at -80°C for 2 and 6 months, and the performance of two different lots of caspofungin was investigated. The addition of BSA to the medium resulted in higher MICs (1 to 9 2-fold dilution steps) for all isolates and compounds. The increases were greatest for anidulafungin and micafungin and, among WT isolates, for C. parapsilosis. The number of very major errors (VMEs) was reduced (24% [20/84 isolates] versus ≤ 7% [6/84 isolates]) using BSA-supplemented EUCAST medium but not using BSA-supplemented CLSI medium (6% versus 9%). MIC results were unchanged after 6 months of storage of test plates. The two lots of caspofungin yielded identical results. Addition of BSA to the EUCAST medium increases the ability to differentiate between WT isolates and isolates harboring resistance mutations.     

Superior fixation of pegged trabecular metal over screw-fixed pegged porous titanium fiber mesh: a randomized clinical RSA study on cementless tibial components

Background and purpose

Lasting stability of cementless implants depends on osseointegration into the implant surface, and long-term implant fixation can be predicted using radiostereometric analysis (RSA) with short-term follow-up. We hypothesized that there would be improved fixation of high-porosity trabecular metal (TM) tibial components compared to low-porosity titanium pegged porous fiber-metal (Ti) polyethylene metal backings.

Methods

In a prospective, parallel-group, randomized unblinded clinical trial, we compared cementless tibial components in patients aged 70 years and younger with osteoarthritis. The pre-study sample size calculation was 22 patients per group. 25 TM tibial components were fixed press-fit by 2 hexagonal pegs (TM group) and 25 Ti tibial components were fixed press-fit and by 4 supplemental screws (Ti group). Stereo radiographs for evaluation of absolute component migration (primary effect size) and single-direction absolute component migration (secondary effect size) were obtained within the first postoperative week and at 6 weeks, 6 months, 1 year, and 2 years. American Knee Society score was used for clinical assessment preoperatively, and at 1 and 2 years.

Results

There were no intraoperative complications, and no postoperative infections or revisions. All patients had improved function and regained full extension. All tibial components migrated initially. Most migration of the TM components (n = 24) occurred within the first 3 months after surgery whereas migration of the Ti components (n = 22) appeared to stabilize first after 1 year. The TM components migrated less than the Ti components at 1 year (p = 0.01) and 2 years (p = 0.004).

Interpretation

We conclude that the mechanical fixation of TM tibial components is superior to that of screw-fixed Ti tibial components. We expect long-term implant survival to be better with the TM tibial component.Tibial component loosening remains one of the major causes of failure of cementless total knee arthroplasty (TKA), and the early degree of knee implant migration detected by radiostereometric analysis (RSA) has been shown to predict the long-term survival of the implant (Ryd et al. 1995). With cementless knee arthroplasty, stability is achieved by biological fixation within the first weeks after surgery and the success relies on both correct component position and immediate macrofixation (Soballe et al. 1992). Porous implant surfaces support tissue ingrowth and are generally effective in supplementing bony integration of cementless implants (Bobyn et al. 1982). On the other hand, fibrous integration of tibial knee components leads to reduced strength of mechanical fixation, which is detectable under physiological loads, and to increased early migration measured with RSA, and it may indicate an increased risk of loosening at a later stage (Bellemans 1999). RSA is therefore particularly useful during the first postoperative years (Valstar et al. 2005).The pore size and structural geometry of coatings in cementless arthroplasty are important factors for early and safe bone ingrowth. Low-porosity coatings, i.e. fiber-metals and beads, may have inferior osseointegration compared to high-porosity coatings with regular interconnecting pores, i.e. trabecular metal (tantalum), which is a newer prosthetic material (Bobyn et al. 1982, Bobyn 1999).The prosthetic design also influences implant survival and function. A monobloc tibial design offers advantages compared to a modular design in terms of elimination of back-side wear problems and elimination of metallic debris produced by the polyethylene locking mechanism. A pegged tibial design without screw-holes provides an increased surface area for bony fixation and eliminates points where wear debris can directly enter the bone. On the other hand, a modular and screw-fixed design offers a consistent intraoperative macro-fixation (Sumner et al. 1994) with the option of isolated polyethylene liner revision later on (Ryd et al. 1993).It has been recommended that the fixation of new products for prosthetic surgery should be evaluated by RSA prior to general use (Valstar et al. 2005), and at the time of initiation of this study no clinical RSA data were available for the trabecular metal implant. The aim of this randomized clinical trial (RCT) was to compare the early clinical and migration results (absolute total migration and absolute single-direction migration) in younger osteoarthritic patients treated with two different cementless tibial implants: a new double-pegged trabecular metal tibial component and a well-documented porous, pegged screw-fixed titanium fiber-mesh tibial component.     

Astrocyte-mediated control of cerebral blood flow

Local increase in blood flow during neural activity forms the basis for functional brain imaging, but its mechanism remains poorly defined. Here we show that cortical astrocytes in vivo possess a powerful mechanism for rapid vasodilation. We imaged the activity of astrocytes labeled with the calcium (Ca(2+))-sensitive indicator rhod-2 in somatosensory cortex of adult mice. Photolysis of caged Ca(2+) in astrocytic endfeet ensheathing the vessel wall was associated with an 18% increase in arterial cross-section area that corresponded to a 37% increase in blood flow. Vasodilation occurred with a latency of only 1-2 s, and both indomethacin and the cyclooxygenase-1 inhibitor SC-560 blocked the photolysis-induced hyperemia. These observations implicate astrocytes in the control of local microcirculation and suggest that one of their physiological roles is to mediate vasodilation in response to increased neural activity.     

National surveillance of fungemia in Denmark (2004 to 2009)

A 6-year nationwide study of fungemia in Denmark was performed using data from an active fungemia surveillance program and from laboratory information systems in nonparticipating regions. A total of 2,820 episodes of fungemia were recorded. The incidence increased from 2004 to 2007 (7.7 to 9.6/100,000) and decreased slightly from 2008 to 2009 (8.7 to 8.6/100,000). The highest incidences were seen at the extremes of age (i.e., 11.3 and 37.1/100,000 for those <1 and 70 to 79 years old, respectively). The rate was higher for males than for females (10.1 versus 7.6/100,000, P = 0.003), with the largest difference observed for patients >50 years of age. The species distribution varied significantly by both age and gender. Candida species accounted for 98% of the pathogens, and C. albicans was predominant, although the proportion decreased (64.4% to 53.2%, P < 0.0001). C. glabrata ranked second, and the proportion increased (16.5% to 25.9%, P = 0.003). C. glabrata was more common in adults and females than in children and males, whereas C. tropicalis was more common in males (P = 0.020). C. krusei was a rare isolate (4.1%) except at one university hospital. Acquired resistance to amphotericin and echinocandins was rare. However, resistance to fluconazole (MIC of >4 μg/ml) occurred in C. albicans (7/1,183 [0.6%]), C. dubliniensis (2/65 [3.1%]), C. parapsilosis (5/83 [6.0%]), and C. tropicalis (7/104 [6.7%]). Overall, 70.8% of fungemia isolates were fully fluconazole susceptible, but the proportion decreased (79.7% to 68.9%, P = 0.02). The study confirmed an incidence rate of fungemia in Denmark three times higher than those in other Nordic countries and identified marked differences related to age and gender. Decreased susceptibility to fluconazole was frequent and increasing.     

Diagnostics of fungal infections in the Nordic countries: we still need to improve!

A Nordic External Quality Assessment programme in medical mycology was established in 2005. In order to monitor not 'best practice' but the level of routine diagnostics, specimens were designed to resemble clinical samples and laboratories were asked to handle the samples like routine samples. Five simulated clinical samples were distributed to 59 participating Nordic laboratories of clinical microbiology. The specimens contained the following microorganisms: 1) Candida glabrata and C. albicans in a ratio of 1:20; 2) Cryptococcus neoformans; 3) Aspergillus fumigatus, C. albicans and Enterobacter cloacae; 4) C. tropicalis, Klebsiella pneumonia and Enterococcus faecium; 5) None. 66% of the laboratories failed to detect the C. glabrata isolate in sample no. 1. 34% of the laboratories reporting susceptibility results incorrectly reported the Cryptococcus neoformans isolate as fluconazole susceptible. 24% of the laboratories failed to detect Aspergillus fumigatus in specimen no. 3 despite the accompanying clinical information notifying that it was a BAL sample from a neutropenic patient in an ICU. In conclusion, this distribution of simulated clinical samples illustrates that the traditional quality assessment programmes may give a false sense of satisfactory performance, that mycological diagnosis is difficult, and that there is a need of further improvement and attention.     

Rapid and accurate identification of Candida albicans isolates by use of PNA FISHFlow

We developed the simple, rapid (1 h), and accurate PNA FISH(Flow) method for the identification of Candida albicans. The method exploits unique in solution in situ hybridization conditions under which the cells are simultaneously fixed and hybridized. This method facilitates the accurate identification of clinical yeast isolates using two scoring techniques: flow cytometry and fluorescence microscopy.