Identification of members of the Burkholderia cepacia complex by species-specific PCR

Definitive identification of the species in the Burkholderia cepacia complex by routine clinical microbiology methods is difficult. Phenotypic tests to identify B. multivorans and B. vietnamiensis have been established; more recent work indicates B. stabilis may also be identified by growth characteristics and biochemical tests. However, attempts to identify genomovars I and III have, thus far, proved unsuccessful. Previously, we demonstrated the utility of two primer pairs, directed to the rRNA operon, to specifically identify the B. cepacia complex in a PCR. One of these primer pairs, G1-G2, only amplified a DNA fragment from genomovars I and III and B. stabilis in a PCR with genomic DNA isolated from prototypical strains representing the five genomovars. Sequence analysis of the rRNA operon for all the genomovars indicated that this primer pair targeted a region shared by these isolates. Further analysis revealed a region of heterogeneity between genomovar III and B. stabilis internal to the amplified product of G1-G2. Primers designed to target this region were tested with prototypical strains following an initial amplification with the G1-G2 primer pair. New primers specific for the prototypical genomovar III and B. stabilis were designated SPR3 and SPR4, respectively. Analysis of 93 isolates representing 18 genomovar I, 13 B. multivorans, 36 genomovar III, 11 B. stabilis, and 15 B. vietnamiensis isolates was performed. DNA from all isolates of genomovars I and III and B. stabilis was amplified by G1-G2. Genomovar III isolates yielded a product with SPR3/G1 while B. stabilis amplified with SPR4-G1. Genomovar I isolates were amplified by either SPR3-G1 or SPR4-G1, but not both. B. multivorans yielded a product with SPR3-G1 but not G1-G2, and B. vietnamiensis isolates were negative in all PCRs. Thus using an algorithm with G1-G2, SPR3-G1, and SPR4-G1 primers in a PCR analysis, genomovar III isolates can be separated from B. stabilis and the identity of B. multivorans and B. vietnamiensis can be confirmed.     

Microglandular adenosis: a prime suspect in triple‐negative breast cancer development

Microglandular adenosis (MGA) and atypical MGA (AMGA) are unusual lesions of the breast. They were once regarded as benign proliferative lesions and innocent bystanders. Several lines of evidence suggested that they could be neoplastic, clonal lesions and a non‐obligate precursor for triple‐negative breast cancers (TNBC). Recent work published in The Journal of Pathology by Guerini‐Rocco and colleagues provided further evidence regarding the precursor–product relationship between MGA/AMGA and TNBC. Using a massively parallel sequencing approach, they demonstrated that MGA/AMGA, particularly those associated with TNBC, could be clonal neoplastic lesions showing clonal non‐synonymous mutations, but none in pure MGA. Importantly, those alterations were observed in the associated TNBC. They were also able to identify recurrent alterations in TP53 in those MGA/AMGA cases as well as their associated TNBC. The findings, in conjunction with others, underscore the significance for MGA in clinical diagnosis. The potential of a benign lesion to progress into an aggressive malignant tumour implies that modification of the current management approach may be necessary. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Measuring distortion of skeletal elements in Lodox Statscan‐generated images

Due to a scarcity of available skeletal material, anthropologists and other practitioners face difficulties with either the creation or validation of techniques used to estimate a biological profile in subadults. To address this problem, radiographic images of living individuals are often used in lieu of dry skeletal elements. However, radiographic images suffer from distortion. Some problems with metric analyses when using radiographic images may be addressed with the Lodox Statscan, an X‐ray machine that claims to produce minimal distortion along the scan‐axis due to a linear slot‐scanning design. The purpose of this research was to measure the distortion of skeletal elements in radiographic images generated from a Lodox Statscan. Skeletal elements subject to multiple imaging variables that affect distortion were radiographed, measured, and then compared to the dry bone measurements through multiple approaches. An 85% percent agreement within a ±1 mm range and a 97% agreement within a ±2 mm range was obtained. Percent difference results demonstrate that slot‐axis measurements incurred more distortion than scan‐axis measurements (11.8% and 2.7%, respectively). Inclusion of foam results in 4.5% more error than when foam is not included in the image. Angled scan‐axis measurements also incurred more distortion than either nonangled slot‐ and scan‐axis measurements. A Bland‐Altmanplot reveals an overall agreement between the radiographic and dry bonemeasurements, with most measurements falling within the upper and lower limits. Similar measurement error is found in Statscan radiographic and dry bone measurements; therefore, the Statscan offers a radiographic venue to collect metric data. Clin. Anat. 26:780–786, 2013. © 2013 Wiley Periodicals, Inc.     

Distribution of mutations in the PEX gene in families with X-linked hypophosphataemic rickets (HYP)

Mutations in the PEX gene at Xp22.1 (phosphate-regulating gene with homologies to endopeptidases, on the X-chromosome), are responsible for X-linked hypophosphataemic rickets (HYP). Homology of PEX to the M13 family of Zn2+ metallopeptidases which include neprilysin (NEP) as prototype, has raised important questions regarding PEX function at the molecular level. The aim of this study was to analyse 99 HYP families for PEX gene mutations, and to correlate predicted changes in the protein structure with Zn2+ metallopeptidase gene function. Primers flanking 22 characterised exons were used to amplify DNA by PCR, and SSCP was then used to screen for mutations. Deletions, insertions, nonsense mutations, stop codons and splice mutations occurred in 83% of families screened for in all 22 exons, and 51% of a separate set of families screened in 17 PEX gene exons. Missense mutations in four regions of the gene were informative regarding function, with one mutation in the Zn2+-binding site predicted to alter substrate enzyme interaction and catalysis. Computer analysis of the remaining mutations predicted changes in secondary structure, N-glycosylation, protein phosphorylation and catalytic site molecular structure. The wide range of mutations that align with regions required for protease activity in NEP suggests that PEX also functions as a protease, and may act by processing factor(s) involved in bone mineral metabolism.      

Alteration of cross-bridge kinetics by myosin light chain phosphorylation in rabbit skeletal muscle: implications for regulation of actin-myosin interaction.

Myosin light chain phosphorylation in permeable skeletal muscle fibers increases isometric force and the rate of force production at submaximal levels of calcium activation; myosin light chain phosphorylation may underlie the increased rate and extent of force production associated with isometric twitch potentiation in intact fibers. To understand the mechanism by which myosin light chain phosphorylation manifests these effects, we have measured isometric force, isometric stiffness, rate of isometric force redevelopment after isotonic shortening, and isometric ATPase activity in permeabilized rabbit psoas muscle fibers. These measurements were made in the presence and absence of myosin light chain phosphorylation over a range of calcium concentrations that caused various levels of activation. The results were analyzed with a two-state cross-bridge cycle model as suggested by Brenner [Brenner, B. (1988) Proc. Natl. Acad. Sci. USA 85, 3265-3269]. The results indicate that myosin light chain phosphorylation exerts its effect on force generation and the isometric rate of force redevelopment in striated muscle through a single mechanism, namely, by increasing the rate constant describing the transition from non-force-generating cross-bridges to force-generating states (fapp). gapp, the reverse rate constant, is unaffected by phosphorylation as are the number of cycling cross-bridges. Since both calcium and myosin light chain phosphorylation increase fapp, the possibility is considered that modulation of fapp may represent a general mechanism for regulating force in actin-myosin systems.     

Direct versus indirect transfer for traumatic brain injury to James Cook University Hospital: a retrospective study

IntroductionPatients with traumatic brain injury are referred to the neurosurgical unit at James Cook University Hospital, Middlesbrough, either from local accident and emergency departments (direct transfer from the scene) or from other hospitals (indirect transfer). This study looked at the outcome in both groups.Material and methodsThis was a retrospective observational study using trauma audit research network data for patients treated for traumatic brain injury at the neurosurgery department at the neurosurgical unit at James Cook University Hospital.ResultsA total of 356 patients with traumatic brain injury were admitted under the care of neurosurgeons; 143 (40%) of these patients had a neurosurgical procedure. Of the patients undergoing a neurological procedure, 111 patients were transferred directly while 32 were indirect transfers; 213 patients were managed conservatively. Of those managed conservatively, 165 were transferred directly while 48 were indirect transfers.We compared the length of hospital stay and Glasgow Outcome Scale score for the patients based on whether they were conservatively managed or required surgery in the direct and indirect transfer groups. The difference in the length of stay in the surgical and conservative groups following direct and indirect transfer was insignificant (p = 0.07). The time to the operation in direct and indirect transfer was also not statistically significant (p = 0.06).ConclusionPatients are as safe, if not safer, by reaching the nearest trauma unit with facilities for resuscitation and imaging.     

Speech-controlled generation of radiology reports

Voice entry has been successfully employed to generate radiology reports with a word recognizer with a 1,000-word lexicon capacity. About 50% of reports were able to be dictated with a single 900-word lexicon. This was split into five sections by anatomic or subspecialty application. Each was augmented to 900 words. By switching from one lexicon to another, it was possible to dictate more than 70% of reports. With exclusive use of three lexicons in subspecialty areas (gastrointestinal radiology, neuroradiology, and mammography), and with further modification of the respective vocabulary, it has been possible to employ the system 88% of the time. Twelve percent of cases included wording that was beyond the scope of the lexicon. Computer subsets that allow different translations of some words when used in different contexts have been used. Some of these are used as triggers that will print whole lines, sentences, or even complete reports. Dictation times with voice entry take about 20% longer. Recognition reliability has been greater than 95%.