The impact of γ-irradiation on the induction of bystander killing by genetically engineered ovarian tumor cells: implications for clinical use

Background Cellular based therapeutic approaches for cancer rely on careful consideration of finding the optimal cell to execute the cellular goal of cancer treatment. Cell lines and primary cell cultures have been used in some studies to compare the in vitro and in vivo efficacy of autologous vs allogeneic tumour cell vaccines. Methods This study examines the effect of γ-irradiation on a range of tumor cell lines in conjunction with suicide gene therapy of cancer. To determine the efficacy of this modality, a series of in vitro and in vivo experiments were conducted using genetically modified and unmodified tumor cell lines. Results Following co-culture of HSV-TK modified tumor cells and unmodified tumor cells both in vitro and in vivo we observed that the PA-STK ovarian tumor cells were sensitive to γ-irradiation, completely abolishing their ability to induce bystander killing of unmodified tumor cells. In contrast, TK-modified human and mouse mesothelioma cells were found to retain their in vitro and in vivo bystander killing effect after γ-irradiation. Morphological evidence was consistent with the death of PA-STK cells being by pyknosis after γ-irradiation. These results suggest that PA-STK cells are not suitable for clinical application of suicide gene therapy of cancer, as lethal γ-irradiation (100 Gy) interferes with their bystander killing activity. However, the human mesothelioma cell line CRL-5830-TK retained its bystander killing potential after exposure to similarly lethal γ-irradiation (100 Gy). CRL-5830 may therefore be a suitable vehicle for HSV-TK suicide gene therapy. Conclusions This study highlights the diversity among tumor cell lines and the careful considerations needed to find the optimal tumor cell line for this type of suicide gene therapy of cancer

Transcriptome sequencing, annotation and polymorphism detection in the hop bush, Dodonaea viscosa

The hop bush, Dodonaea viscosa, is a trans-oceanic species distributed oversix continents. It evolved in Australia where it is found over a wide range of habitat types and is an ecologically important species. Limited genomic resources are currently available for this species, thus our understanding of its evolutionary history and ecological adaptation is restricted. Here, we present a comprehensive transcriptome dataset for future genomic studies into this species.We performed Illumina sequencing of cDNA prepared from leaf tissue collected from seven populations of D. viscosa ssp. angustissima and spatulata distributed along an environmental gradient in South Australia. Sequenced reads were assembled to provide a transcriptome resource. Contiguous sequences (contigs) were annotated using BLAST searches against the NCBI non-redundant database and gene ontology definitions were assigned. Single nucleotide polymorphisms were detected for the establishment of a genetic marker set. A comparison between the two subspecies was also carried out.Illumina sequencing returned 268,672,818 sequence reads, which were de novoassembled into 105,125 contigs. Contigs with significant BLAST alignments (E value < 1e(-5))numbered at 44,191, with 38,311 of these having their most significant hits to sequences from land plant species. Gene Ontology terms were assigned to 28,440 contigs and KEGG analysis identified 146 pathways that the gene products from 5,070 contigs are potentially involved in. The subspecies comparison identified 8,494 fixed SNP differences across 3,979 contiguous sequences, indicating a level of genetic differentiation between them. Across all samples, 248,235 SNPs were detected.We have established a significant genomic data resource for D. viscosa,providing a comprehensive transcriptomic reference. Genetic differences among morphologically distinct subspecies were found. A wide range of putative gene regions were identified along with a large set of variable SNP markers, providing a basis for studies into the evolution and ecological adaptation of D. viscosa.Matthew J. Christmas, Ed Biffin and Andrew J. Low

Targeted capture to assess neutral genomic variation in the narrow-leaf hopbush across a continental biodiversity refugium

The Adelaide geosyncline, a mountainous region in central southern Australia, is purported to be an important continental refugium for Mediterranean and semi-arid Australian biota, yet few population genetic studies have been conducted to test this theory. Here, we focus on a plant species distributed widely throughout the region, the narrow-leaf hopbush, Dodonaea viscosa ssp. angustissima, and examine its genetic diversity and population structure. We used a hybrid-capture target enrichment technique to selectively sequence over 700 genes from 89 individuals across 17 sampling locations. We compared 815 single nucleotide polymorphisms among individuals and populations to investigate population genetic structure. Three distinct genetic clusters were identified; a Flinders/Gammon ranges cluster, an Eastern cluster, and a Kangaroo Island cluster. Higher genetic diversity was identified in the Flinders/Gammon Ranges cluster, indicating that this area is likely to have acted as a refugium during past climate oscillations. We discuss these findings and consider the historical range dynamics of these populations. We also provide methodological considerations for population genomics studies that aim to use novel genomic approaches (such as target capture methods) on non-model systems. The application of our findings to restoration of this species across the region are also considered.Matthew J. Christmas, Ed Biffin, Martin F. Breed & Andrew J. Low

THE B LYMPHOCYTE DIFFERENTIATION FACTOR (BAFF) IS EXPRESSED IN THE AIRWAYS OF CHILDREN WITH CF AND IN LUNGS OF MICE INFECTED WITH PSEUDOMONAS AERUGINOSA

Background Chronic lung infection with Pseudomonas aeruginosa remains a major cause of mortality and morbidity among individuals with CF. Expression of mediators promoting recruitment and differentiation of B cells, or supporting antibody production is poorly understood yet could be key to controlling infection. Methods BAFF was measured in BAL from children with CF, both with and without P. aeruginosa, and controls. Mice were intra-nasally infected with P. aeruginosa strain LESB65 for up to 7 days. Cellular infiltration and expression of B cell chemoattractants and B cell differentiation factor, BAFF were measured in lung tissue. Results BAFF expression was elevated in both P. aeruginosa negative and positive CF patients and in P. aeruginosa infected mice post infection. Expression of the B cell chemoattractants CXCL13, CCL19 and CCL21 increased progressively post infection. Conclusions In a mouse model, infection with P. aeruginosa was associated with elevated expression of BAFF and other B cell chemoattractants suggesting a role for airway B cell recruitment and differentiation in the local adaptive immune response to P. aeruginosa. The paediatric CF airway, irrespective of pseudomonal infection, was found to be associated with an elevated level of BAFF implying that BAFF expression is not specific to pseudomonas infection and may be a feature of the CF airway. Despite the observed presence of a potent B cell activator, chronic colonisation is common suggesting that this response is ineffective

Results of Archaeological Monitoring of the Spring Lake Section 206 Aquatic Ecosystem Restoration Project, Texas State University-San Marcos, Hays County, Texas

The Center for Archaeological Studies (CAS) at Texas State University-San Marcos conducted archaeological monitoring investigations in association with the Spring Lake Aquatic Ecosystem Restoration Project between October 2011 and July 2012. These archaeological monitoring investigations were the result of mitigation efforts proposed in the Historic Properties Treatment Plan drafted in accordance to the Memorandum of Agreement (MOA) signed and enacted between the U.S. Army Corps of Engineers, Texas State University-San Marcos, and the Texas Historical Commission. Archaeological monitoring investigations consisted of monitoring all demolition and ground-disturbing activities conducted during the course of the Spring Lake Aquatic Ecosystem Restoration Project. All cultural deposits or potential cultural deposits were identified, assessed, and documented during the project by archaeological monitors, and time-diagnostic artifacts were collected. Locations of deposits were recorded and uploaded to a GIS database of the Spring Lake area for future reference. No significant cultural remains were identified or impacted by demolition and ground-disturbing activities of the Spring Lake Aquatic Ecosystem Restoration Project

An update: improvements in imaging perfluorocarbon-mounted plant leaves with implications for studies of plant pathology, physiology, development and cell biology.

Plant leaves are optically complex, which makes them difficult to image by light microscopy. Careful sample preparation is therefore required to enable researchers to maximize the information gained from advances in fluorescent protein labeling, cell dyes and innovations in microscope technologies and techniques. We have previously shown that mounting leaves in the non-toxic, non-fluorescent perfluorocarbon (PFC), perfluorodecalin (PFD) enhances the optical properties of the leaf with minimal impact on physiology. Here, we assess the use of the PFCs, PFD, and perfluoroperhydrophenanthrene (PP11) for in vivo plant leaf imaging using four advanced modes of microscopy: laser scanning confocal microscopy (LSCM), two-photon fluorescence microscopy, second harmonic generation microscopy, and stimulated Raman scattering (SRS) microscopy. For every mode of imaging tested, we observed an improved signal when leaves were mounted in PFD or in PP11, compared to mounting the samples in water. Using an image analysis technique based on autocorrelation to quantitatively assess LSCM image deterioration with depth, we show that PP11 outperformed PFD as a mounting medium by enabling the acquisition of clearer images deeper into the tissue. In addition, we show that SRS microscopy can be used to image PFCs directly in the mesophyll and thereby easily delimit the "negative space" within a leaf, which may have important implications for studies of leaf development. Direct comparison of on and off resonance SRS micrographs show that PFCs do not to form intracellular aggregates in live plants. We conclude that the application of PFCs as mounting media substantially increases advanced microscopy image quality of living mesophyll and leaf vascular bundle cells

Annual Report to Texas State University, San Marcos, Hays County, Texas, for Texas Antiquities Permit No. 6775

The Center for Archaeological Studies (CAS) at Texas State University (University) conducted archaeological investigations for eight proposed undertakings on property owned by the University under Texas Antiquities Permit No. 6775 during the year 2014. Investigations were conducted to determine if intact cultural resources were present within the project areas and if they would be adversely affected by construction and development. Under a Memorandum of Agreement (MOA) between the Texas Historical Commission (THC) and the University, CAS is authorized to determine whether proposed undertakings have the potential to negatively impact cultural resources, and if so, to recommend to the University courses of future action that may avoid or offset that impact. As a result of archaeological monitoring, one previously unrecorded archaeological site, 41HY518, was documented. The boundaries of sites 41HY447 and 41HY37 have been revised to include associated deposits identified during archaeological investigations. No intact, significant cultural deposits were encountered or impacted during the University undertakings of 2014. Accordingly, CAS recommends that no additional investigations are warranted. CAS continues to recommend that the University avoid adversely impacting SALs 41HY160 and 41HY161, as well as sites 41HY37 and 41HY447. If avoidance of intact features and deposits of these sites is not possible, then additional work is recommended to offset the potential loss of information

The urgent need to develop emergency EYE care in the UK:the way forward?

For two decades prior to 2004, a steady state existed of ~14 million general Accident and Emergency (A&amp;E) annual attendances in England. This total has risen each year since, with 22.9 million attendances recorded in 2015/16 (https://www.england.nhs.uk/statistics/wp-content/uploads/sites/2/2016/06/Monthly-AE-Report-December-16.pdf). Resultant pressure on A&amp;E has received a great deal of public attention and extra resourcing with medical staff numbers rising 71% from 2002 to 2012

Competition and parasitism in the native White Clawed Crayfish Austropotamobius pallipes and the invasive Signal Crayfish Pacifastacus leniusculus in the UK

Many crayfish species have been introduced to novel habitats worldwide, often threatening extinction of native species. Here we investigate competitive interactions and parasite infections in the native Austropotamobius pallipes and the invasive Pacifastacus leniusculus from single and mixed species populations in theUK. We found A. pallipes individuals to be significantly smaller in mixed compared to single species populations; conversely P. leniusculus individuals were larger in mixed than in single species populations. Our data provide no support for reproductive interference as a mechanism of competitive displacement and instead suggest competitive exclusion of A. pallipes from refuges by P. leniusculus leading to differential predation. We screened 52 P. leniusculus and 12 A. pallipes for microsporidian infection using PCR. We present the first molecular confirmation of Thelohania contejeani in the native A. pallipes; in addition, we provide the first evidence for T. contejeani in the invasive P. leniusculus. Three novel parasite sequenceswere also isolated fromP. leniusculus with an overall prevalence of microsporidian infection of 38% within this species; we discuss the identity of and the similarity between these three novel sequences. We also screened a subset of fifteen P. leniusculus and three A. pallipes for Aphanomyces astaci, the causative agent of crayfish plague and for the protistan crayfish parasite Psorospermium haeckeli. We found no evidence for infection by either agent in any of the crayfish screened. The high prevalence of microsporidian parasites and occurrence of shared T. contejeani infection lead us to propose that future studies should consider the impact of these parasites on native and invasive host fitness and their potential effects upon the dynamics of native-invader systems

Research activity and the association with mortality.

INTRODUCTION: The aims of this study were to describe the key features of acute NHS Trusts with different levels of research activity and to investigate associations between research activity and clinical outcomes. METHODS: National Institute for Health Research (NIHR) Comprehensive Clinical Research Network (CCRN) funding and number of patients recruited to NIHR Clinical Research Network (CRN) portfolio studies for each NHS Trusts were used as markers of research activity. Patient-level data for adult non-elective admissions were extracted from the English Hospital Episode Statistics (2005-10). Risk-adjusted mortality associations between Trust structures, research activity and, clinical outcomes were investigated. RESULTS: Low mortality Trusts received greater levels of funding and recruited more patients adjusted for size of Trust (n = 35, 2,349 £/bed [95% CI 1,855-2,843], 5.9 patients/bed [2.7-9.0]) than Trusts with expected (n = 63, 1,110 £/bed, [864-1,357] p<0.0001, 2.6 patients/bed [1.7-3.5] p<0.0169) or, high (n = 42, 930 £/bed [683-1,177] p = 0.0001, 1.8 patients/bed [1.4-2.1] p<0.0005) mortality rates. The most research active Trusts were those with more doctors, nurses, critical care beds, operating theatres and, made greater use of radiology. Multifactorial analysis demonstrated better survival in the top funding and patient recruitment tertiles (lowest vs. highest (odds ratio & 95% CI: funding 1.050 [1.033-1.068] p<0.0001, recruitment 1.069 [1.052-1.086] p<0.0001), middle vs. highest (funding 1.040 [1.024-1.055] p<0.0001, recruitment 1.085 [1.070-1.100] p<0.0001). CONCLUSIONS: Research active Trusts appear to have key differences in composition than less research active Trusts. Research active Trusts had lower risk-adjusted mortality for acute admissions, which persisted after adjustment for staffing and other structural factors