Effect of medium composition, agitation and the presence of EDTA on the antimicrobial activity of cryptolepine

The antimicrobial activity of cryptolepine is influenced by the type of medium employed, agitation and the presence of non-inhibitory concentrations of EDTA. The use of Mueller–Hinton broth (MHB), iso-sensitest broth and tryptone soya broth (TSB) produced lower minimum inhibitory concentrations (MICs) for some of the test organisms compared with nutrient broth or yeast dextrose broth (YDB). For example, a fourfold drop in MIC was recorded for Saccharomyces cerevisiae in MHB compared with the same organism tested in YDB. Agitation of the broths during incubation nearly always produced lower MICs for the bacteria, an eightfold decrease in MIC being recorded for Escherichia coli cultured in nutrient broth with agitation compared with a statically maintained culture. A non-inhibitory concentration (10⁻³ mol l⁻¹) of disodium EDTA enhanced the antimicrobial activity of cryptolepine. Against E. coli NCTC 11560, an eightfold decrease in MIC and minimum bactericidal concentration (MBC) was recorded when tested in the presence of EDTA.     

Mutational analysis of slyD, an Escherichia coli gene encoding a protein of the FKBP immunophilin family

slyD encodes a 196 amino acid polypeptide that is a member of the FKBP family of cis–trans peptidyl–prolyl isomerases (PPIases). slyD mutations affect plaque formation by the phage φX174 by blocking the action of the phage lysis protein E. Here we describe the selection of a set of spontaneous slyD mutations conferring resistance to the expression of gene E from a plasmid. These mutations occur disproportionately in residues of SlyD that, based on the structure of the prototype mammalian FKBP12, make ligand contacts with immunosuppressing drug molecules or are conserved in other FKBP proteins. A wide variation in the plating efficiency of φX174 on these E  ^R strains is observed, relative to the parental, indicating that these alleles differ widely in residual SlyD activity. Moreover, it is found that slyD mutations cause significant growth rate defects in Escherichia coli B and C backgrounds. Finally, overexpression of slyD causes filamentation of the host. Thus, among the FKBP genes found in organisms across the evolutionary spectrum, slyD is unique in having three distinct drug-independent phenotypes.     

Isolation and characterization of a glycosylated form of human insulin-like growth factor I produced in Saccharomyces cerevisiae

Expression and secretion of human insulin-like growth factor-I (IGF-I) in Saccharomyces cerevisiae was achieved by linking an actin (ACT) promoter to an MF alpha 1 prepro leader peptide/IGF-I gene fusion. Purified human IGF-I from yeast culture media was found to contain, in addition to the native form, also a glycosylated variant. Structural studies showed that both IGF-I forms were processed identically, resulting in 70-amino-acid long polypeptides, with intact N-terminal and C-terminal residues of glycine and alanine, respectively. The glycosylation site was determined to threonine-29 (Thr29), by 1H NMR spectroscopy and protein sequence analysis of an isolated tryptic peptide(22-36). No other glycosylation sites were found. Only mannose was detected in the sugar analysis, with an estimated content of 4.5% w/w corresponding to 2 mannose residues per molecule of IGF-I. The carbohydrate structure, determined by 1H and 13C NMR spectroscopy, was found to be alpha-D-Manp(1----2)alpha-D-Manp(1----3)Thr corresponding to an O-linked glycoprotein structure. No other post-translational modifications could be identified in the glycosylated IGF-I form. Furthermore, this form was highly active, comparable to native IGF-I, exhibiting a specific activity of 20,500 units/mg, as determined by a radio-receptor assay.     

Field test of an automated radio‐telemetry system: tracking local space use of aerial insectivores

Documenting local space use of birds that move rapidly, but are too small to carry GPS tags, such as swallows and swifts, can be challenging. For these species, tracking methods such as manual radio‐telemetry and visual observation are either inadequate or labor‐ and time‐intensive. Another option is use of an automated telemetry system, but equipment for such systems can be costly when many receivers are used. Our objective, therefore, was to determine if an automated radio‐telemetry system, consisting of just two receivers, could provide an alternative to manual tracking for gathering data on local space use of six individuals of three species of aerial insectivores, including one Cliff Swallow (Petrochelidon pyrrhonota), one Eastern Phoebe (Sayornis phoebe), and four Barn Swallows (Hirundo rustica). We established automated radio‐telemetry systems at three sites near the city of Peterborough in eastern Ontario, Canada, from May to August 2015. We evaluated the location error of our two‐receiver system using data from moving and stationary test transmitters at known locations, and used telemetry data from the aerial insectivores as a test of the system's ability to track rapidly moving birds under field conditions. Median location error was ~250 m for automated telemetry test locations after filtering. More than 90% of estimated locations had large location errors and were removed from analysis, including all locations > 1 km from receiver stations. Our automated telemetry receivers recorded 17,634 detections of the six radio‐tagged birds. However, filtering removed an average of 89% of bird location estimates, leaving only the Cliff Swallow with enough locations for analysis of space use. Our results demonstrate that a minimal automated radio‐telemetry system can be used to assess local space use by small, highly mobile birds, but the resolution of the data collected using only two receiver stations was coarse and had a limited range. To improve both location accuracy and increase the percentage of usable location estimates collected, we suggest that, in future studies, investigators use receivers that simultaneously record signals detected by all antennas, and use of a minimum of three receiver stations with more antennas at each station.     

The Influence of Diabetic Peripheral Neuropathy on Local Postural Muscle and Central Sensory Feedback Balance Control

Poor balance control and increased fall risk have been reported in people with diabetic peripheral neuropathy (DPN). Traditional body sway measures are unable to describe underlying postural control mechanism. In the current study, we used stabilogram diffusion analysis to examine the mechanism under which balance is altered in DPN patients under local-control (postural muscle control) and central-control (postural control using sensory cueing). DPN patients and healthy age-matched adults over 55 years performed two 15-second Romberg balance trials. Center of gravity sway was measured using a motion tracker system based on wearable inertial sensors, and used to derive body sway and local/central control balance parameters. Eighteen DPN patients (age = 65.4±7.6 years; BMI = 29.3±5.3 kg/m²) and 18 age-matched healthy controls (age = 69.8±2.9; BMI = 27.0±4.1 kg/m²) with no major mobility disorder were recruited. The rate of sway within local-control was significantly higher in the DPN group by 49% (healthy local-control_slope = 1.23±1.06×10^-2 cm²/sec, P<0.01), which suggests a compromised local-control balance behavior in DPN patients. Unlike local-control, the rate of sway within central-control was 60% smaller in the DPN group (healthy central-control_slope-Log = 0.39±0.23, P<0.02), which suggests an adaptation mechanism to reduce the overall body sway in DPN patients. Interestingly, significant negative correlations were observed between central-control rate of sway with neuropathy severity (r _Pearson = 0.65-085, P<0.05) and the history of diabetes (r _Pearson = 0.58-071, P<0.05). Results suggest that in the lack of sensory feedback cueing, DPN participants were highly unstable compared to controls. However, as soon as they perceived the magnitude of sway using sensory feedback, they chose a high rigid postural control strategy, probably due to high concerns for fall, which may increase the energy cost during extended period of standing; the adaptation mechanism using sensory feedback depends on the level of neuropathy and the history of diabetes.     

Development of an Accelerometer-Linked Online Intervention System to Promote Physical Activity in Adolescents

Most adolescents do not achieve the recommended levels of moderate-to-vigorous physical activity (MVPA), placing them at increased risk for a diverse array of chronic diseases in adulthood. There is a great need for scalable and effective interventions that can increase MVPA in adolescents. Here we report the results of a measurement validation study and a preliminary proof-of-concept experiment testing the impact of Zamzee, an accelerometer-linked online intervention system that combines proximal performance feedback and incentive motivation features to promote MVPA. In a calibration study that parametrically varied levels of physical activity in 31 12-14 year-old children, the Zamzee activity meter was shown to provide a valid measure of MVPA (sensitivity in detecting MVPA = 85.9%, specificity = 97.5%, and r = .94 correspondence with the benchmark RT3 accelerometer system; all p < .0001). In a subsequent randomized controlled multi-site experiment involving 182 middle school-aged children assessed for MVPA over 6 wks, intent-to-treat analyses found that those who received access to the Zamzee intervention had average MVPA levels 54% greater than those of a passive control group (p < 0.0001) and 68% greater than those of an active control group that received access to a commercially available active videogame (p < .0001). Zamzee’s effects on MVPA did not diminish significantly over the course of the 6-wk study period, and were statistically significant in both females and males, and in normal- vs. high-BMI subgroups. These results provide promising initial indications that combining the Zamzee activity meter with online proximal performance feedback and incentive motivation features can positively impact MVPA levels in adolescents.