Diversity of Serratia marcescens Strains Associated with Cucurbit Yellow Vine Disease in Georgia.

Cucurbit yellow vine disease (CYVD), caused by the squash bug (Anasa tristis)-transmitted bacterium Serratia marcescens, was first identified in Oklahoma and Texas in 1988 and in Georgia in 2012. S. marcescens is a highly diverse species found in many ecological niches. In previous studies, CYVD strains of S. marcescens formed a closely related group separate from non-CYVD strains based on biological and molecular characterization techniques. Multilocus sequence analysis (MLSA) of six housekeeping genes and repetitive elements-based polymerase chain reaction (rep-PCR) using the BOX and ERIC primers were used to assess the genetic diversity of CYVD strains of S. marcescens collected in Georgia together with a strain from Texas and seven non-CYVD strains of S. marcescens. rep-PCR results revealed genetic diversity among CYVD strains while MLSA results showed a 100% similarity across the six loci for all but one of the CYVD strains, which differed at the icd locus by five polymorphisms. For both methods, CYVD strains clustered separately from nonplant-pathogenic S. marcescens strains and were most similar to a rice endophyte strain. One CYVD strain isolated from a squash bug shared genetic similarities with non-CYVD strains, and may be the result of a recombination event between CYVD and non-CYVD strains.

Predictors of fuel poverty and the equity of local fuel poverty support: secondary analysis of data from Bradford, England.

AIMS: Addressing fuel poverty is a critical public health issue given its recent rise in prevalence across Europe. Although previous research identifies national risk markers of fuel poverty, evidence is lacking on whether these are consistent across local geographies, and the equity of local interventions. In the UK's current economic climate, it is more crucial than ever that services benefit households in greatest need. This study aimed to determine significant predictors of fuel poverty among households in Bradford, England, comparing them to national-level predictors, and evaluate if households possessing significant fuel poverty predictors were equitably referred to a local fuel poverty service (Warm Homes Healthy People, WHHP). METHODS: A multivariate logistic regression model determined significant fuel poverty predictors in Bradford using household-level data from the Energy Saving Trust and the Low Income High Costs fuel poverty definition. Statistical testing highlighted significant differences in predictors of fuel poverty between households referred to WHHP and all Bradford households. RESULTS: Significant (p < .05) predictors of fuel poverty included: living in an area with lower average household incomes and higher proportion of ethnic minority individuals, and living in a property with a lower energy efficiency rating. Households living in a detached or older property, and homeowners were more likely to be fuel poor. Differences in the direction of the relationship with fuel poverty were identified between some national and local predictors. Most predictors were significantly (p < .05) overrepresented among WHHP households, suggesting equitable service reach. Ethnic minorities, younger people, and multiperson households were underrepresented. CONCLUSIONS: Local fuel poverty predictors were similar to many national-level predictors, but identified differences in the direction of the relationship between some national and local predictors reaffirm the value of locally focused research. WHHP successfully targeted households possessing key predictors, but should ensure that ethnic minorities, younger people, and multiperson households are equitably referred.

Campylobacter spp. recovered from the Upper Oconee River Watershed, Georgia in a 4-year study.

Waterways should be considered in the migration routes of Campylobacter, and the genus has been isolated from several water sources. Inferences on migration routes can be made from tracking genetic types in populations found in specific habitats and testing how they are linked to other types. Water samples were taken over a 4-year period from waterways in the Upper Oconee River Watershed, Georgia, to recover isolates of thermophilic Campylobacter. The isolates were typed by multilocus sequence typing (MLST) and analyzed to determine the overall diversity of Campylobacter in that environment. Forty-seven independent isolates were recovered from 560 samples (8.4 %). Two (~4 %) isolates were Campylobacter coli, three (~6 %) isolates were putatively identified as Campylobacter lari, and the remaining 42 (~90 %) were Campylobacter jejuni. The C. jejuni and C. coli isolates were typed by the Oxford MLST scheme. Thirty sequence types (STs) were identified including 13 STs that were not found before in the MLST database, including 24 novel alleles. Of the 17 previously described STs, 10 have been isolated from humans, 6 from environmental water, and 6 from wild birds (five types from multiple sources). Seven sites had multiple positive samples, and on two occasions, the same ST was isolated at the same site. The most common type was STST61 with four isolates, and the most common clonal complex was CC179 with nine isolates. CC179 has been commonly associated with environmental water. Although some Campylobacter STs that were found in the Oconee River engage in widespread migration, most are tightly associated with or unique to environmental water sources.

Toxicity of copper to early-life stage Kootenai River white sturgeon, Columbia River white sturgeon, and rainbow trout.

White sturgeon (Acipenser transmontanus) populations throughout western North America are in decline, likely as a result of overharvest, operation of dams, and agricultural and mineral extraction activities in their watersheds. Recruitment failure may reflect the loss of early-life stage fish in spawning areas of the upper Columbia River, which are contaminated with metals from effluents associated with mineral-extraction activities. Early-life stage white sturgeon (A. transmontanus) from the Columbia River and Kootenai River populations were exposed to copper during 96-h flow-through toxicity tests to determine their sensitivity to the metal. Similar tests were conducted with rainbow trout (RBT [Oncorhynchus mykiss]) to assess the comparative sensitivity of this species as a surrogate for white sturgeon. Exposures were conducted with a water quality pH 8.1-8.3, hardness 81-119 mg/L as CaCO(2), and dissolved organic carbon 0.2-0.4 mg/L. At approximately 30 days posthatch (dph), sturgeon were highly sensitive to copper with median lethal concentration (LC(50)) values ranging from 4.1 to 6.8 µg/L compared with 36.5 µg/L for 30 dph RBT. White sturgeon at 123-167 dph were less sensitive to copper with LC(50) values ranging from 103.7 to 268.9 µg/L. RBT trout, however, remained more sensitive to copper at 160 dph with an LC(50) value of 30.9 µg/L. The results indicate that high sensitivity to copper in early-life stage white sturgeon may be a factor in recruitment failure occurring in the upper Columbia and Kootenai rivers. When site-specific water-quality criteria were estimated using the biotic ligand model (BLM), derived values were not protective of early-life stage fish, nor were estimates derived by water-hardness adjustment.

Determining divergence times of the major kingdoms of living organisms with a protein clock.

Amino acid sequence data from 57 different enzymes were used to determine the divergence times of the major biological groupings. Deuterostomes and protostomes split about 670 million years ago and plants, animals, and fungi last shared a common ancestor about a billion years ago. With regard to these protein sequences, plants are slightly more similar to animals than are the fungi. In contrast, phylogenetic analysis of the same sequences indicates that fungi and animals shared a common ancestor more recently than either did with plants, the greater difference resulting from the fungal lineage changing faster than the animal and plant lines over the last 965 million years. The major protist lineages have been changing at a somewhat faster rate than other eukaryotes and split off about 1230 million years ago. If the rate of change has been approximately constant, then prokaryotes and eukaryotes last shared a common ancestor about 2 billion years ago, archaebacterial sequences being measurably more similar to eukaryotic ones than are eubacterial ones.

Vibrio cholerae non-O1 infection in cirrhotics: case report and literature review.

Vibrio species are ubiquitous in the marine environment and can cause severe infections in cirrhotic patients. Patients with liver disease should be warned about the potential dangers of consuming raw or undercooked seafood, and avoiding exposure of wounds to seawater. We report a case of severe sepsis from Vibrio cholerae non-O1 in a patient with cirrhosis awaiting orthotopic liver transplant. This case is aimed to advise clinicians about the importance of V. cholerae subtypes, and non-cholera Vibrio species infections in cirrhotic patients, highlighting the need to educate these patients to stay away from undercooked seafood.

Determination of the mechanical properties of Bionate 80A and Bionate 75D for the stress analysis of cushion form bearings.

Two biomedical grade polyurethanes, currently under investigation as an alternative tibial bearing for a total knee replacement, are analysed. Extensive mechanical testing has been performed on these materials to determine the mechanical properties in terms of strain energy density functions for use in the finite element package MSC. Marc 2001 that do not rely on linear elasticity. A variation in Poisson's ratio in the fourth decimal place can lead to significant errors in the magnitude and distribution of stresses in a linear elastic analysis. As large variations in this parameter (up to the first decimal place) were determined experimentally, it is concluded that a linear elastic approach to the stress analysis of these materials is not valid.

Socio-Ecological Mechanisms Supporting High Densities of Aedes albopictus (Diptera: Culicidae) in Baltimore, MD.

Social, ecological, and climatic factors interact creating a heterogeneous matrix that determines the spatiotemporal distribution of mosquitoes and human risks of exposure to the diseases they transmit. We explore linkages between the social and institutional processes behind residential abandonment, urban ecology, and the interactions of socio-ecological processes with abiotic drivers of mosquito production. Specifically, we test the relative roles of infrastructure degradation and vegetation for explaining the presence of Aedes albopictus Skuse 1894 to better predict spatial heterogeneity in mosquito exposure risk within urban environments. We further examine how precipitation interacts with these socially underpinned biophysical variables. We use a hierarchical statistical modeling approach to assess how environmental and climatic conditions over 3 years influence mosquito ecology across a socioeconomic gradient in Baltimore, MD. We show that decaying infrastructure and vegetation are important determinants of Ae. albopictus infestation. We demonstrate that both precipitation and vegetation influence mosquito production in ways that are mediated by the level of infrastructural decay on a given block. Mosquitoes were more common on blocks with greater abandonment, but when precipitation was low, mosquitoes were more likely to be found in higher-income neighborhoods with managed container habitat. Likewise, although increased vegetation was a negative predictor of mosquito infestation, more vegetation on blocks with high abandonment was associated with the largest mosquito populations. These findings indicate that fine spatial scale modeling of mosquito habitat within urban areas is needed to more accurately target vector control.

Thermoelastic analysis of high-pressure angioplasty balloons.

Failures in angioplasty balloons are investigated using typical destructive techniques. The material properties of moulded balloons are derived from tensile tests and used to establish the reasons for failure of the balloons. Thermoelastic stress analysis is used to determine the stress distribution in the balloons, and a means of interpreting the data to derive actual stresses is described. The departure from linear elastic behaviour in the angioplasty balloons is identified using thermoelastic analysis. The results from the thermoelastic analysis are discussed and compared with those from the destructive tests, and the thermoelastic technique is shown to be a potential new means for non-destructive analysis of angioplasty balloons.

Degree of corneal anaesthesia after topical application of 0.4% oxybuprocaine hydrochloride and 0.5% proparacaine hydrochloride ophthalmic solution in clinically normal cattle.

OBJECTIVES: The use of corneal anaesthesia is necessary for a range of clinical purposes. Therefore, we assessed and compared the efficacy of corneal anaesthesia after application of 0.4% oxybuprocaine hydrochloride and 0.5% proparacaine hydrochloride ophthalmic solution in clinically normal cattle. METHODS: The 24 clinically normal cows were allocated into two groups. Cows in group 1 (n = 12) received 0.2 mL of 0.4% oxybuprocaine hydrochloride with fluorescein ophthalmic solution in one eye and 0.2 mL of sterile saline (0.9% NaCl) with fluorescein in the contralateral eye (control). Group 2 (n = 12) received 0.2 mL of 0.4% oxybuprocaine hydrochloride with fluorescein ophthalmic solution in one eye and 0.2 mL of 0.5% proparacaine hydrochloride with fluorescein in the contralateral eye (control). In each group, corneal touch threshold was determined by Cochet-Bonnet aesthesiometer for both eyes immediately prior to topical administration of solutions, at 1 min and 5 min after administration of topical solutions and every 5 min thereafter for a total of 75 min. RESULTS: Significant corneal anaesthesia was noted immediately following topical application of both oxybuprocaine and proparacaine as compared with controls, with maximal corneal anaesthesia noted 1 min after administration. Both oxybuprocaine and proparacaine produced significant corneal anaesthesia for the duration of the 75-min study. Neither oxybuprocaine hydrochloride nor proparacaine hydrochloride treatment resulted in visible adverse effects. CONCLUSION: There are limited data available demonstrating the efficacy and duration of corneal anaesthetic agents in cattle. Both oxybuprocaine hydrochloride and proparacaine hydrochloride should be considered practical options for providing corneal anaesthesia in cattle in a clinical setting.

The CaSm (LSm1) oncogene promotes transformation, chemoresistance and metastasis of pancreatic cancer cells.

The cancer-associated Sm-like (CaSm) oncogene is overexpressed in 87% of human pancreatic tumor samples and CaSm knockdown has demonstrated therapeutic efficacy in murine models of pancreatic cancer. Evidence indicates that CaSm modulates messenger RNA degradation; however, its target genes and the mechanisms by which CaSm promotes pancreatic cancer remain largely unknown. Here, we demonstrate that the CaSm overexpression alters several hallmarks of cancer-including transformation, proliferation, chemoresistance and metastasis. Doxycycline-induced CaSm expression enhanced proliferation and both anchorage-dependent and -independent growth of the human Panc-1 cells in vitro. CaSm induction decreased gemcitabine-induced cytotoxicity and altered the expression of apoptotic regulation genes, including Bad, E2F1 and Bcl-XL. CaSm-overexpressing Panc-1 cells were twofold more migratory and fourfold more invasive than the driver controls and demonstrated characteristics of epithelial-to-mesenchymal transition such as morphological changes and decreased E-cadherin expression. CaSm induction resulted in changes in RNA expression of metastasis-associated genes such as MMP1, SerpinB5, uPAR and Slug. Using a murine model of metastatic pancreatic cancer, injection of CaSm-induced Panc-1 cells resulted in a higher abundance of hepatic metastatic lesions. Overall, CaSm overexpression contributed to a more aggressive cancer phenotype in Panc-1 cells, further supporting the use of CaSm as a therapeutic target against pancreatic cancer.

Solution structure of the third immunoglobulin domain of the neural cell adhesion molecule N-CAM: can solution studies define the mechanism of homophilic binding?

Homophilic binding of the neural cell adhesion molecule (N-CAM) mediates the calcium-independent cell-cell adhesion that is involved in neuronal development. Two hypotheses have been advanced for the mechanism of homophilic binding. Cell-based experiments have implicated each of the five extracellular immunoglobulin (Ig) domains of N-CAM in the homophilic adhesion interaction, and have predicted that the third domain (Ig III) self-associates. The alternative hypothesis is based on solution observations, which implicate a specific antiparallel interaction between the first two Ig domains (Ig I and Ig II). In order to test these hypotheses, we have determined a high-resolution solution structure of recombinant Ig III (sequence derived from chicken N-CAM) and examined the aggregation behavior of isolated Ig domains in solution. The structure shows that Ig III adopts a canonical Ig fold, in which the beta strands ABED and A'GFCC' form two beta sheets that are linked by a disulfide bond. In contrast to the demonstrated aggregation of Ig III on solid supports, we were unable to demonstrate self-association of Ig III under any of a variety of solution conditions. The structure shows that the surface of Ig III is dominated by two large acidic patches, which may explain our failure to observe self-association in solution. To evaluate the involvement of the Ig I-Ig II interaction in cell-cell adhesion, we designed a point mutation in Ig I (F19S) that proved sufficient to abrogate the Ig I-Ig II interaction seen in solution. However, the introduction of this mutation into full-length N-CAM expressed in COS-7 cells failed to affect N-CAM-mediated cell-cell adhesion. The inability to observe Ig III self-association in solution, combined with the failure of the F19S mutation to affect N-CAM-mediated cell-cell adhesion, suggests that, although solution studies can give important insights into the structures of individual domains, the interactions observed in solution between the domains may not be representative of the interactions that occur on the cell surface.

PAX3 and ETS1 synergistically activate MET expression in melanoma cells.

Melanoma is a highly aggressive disease that is difficult to treat owing to rapid tumor growth, apoptotic resistance and high metastatic potential. The MET proto-oncogene (MET) tyrosine kinase receptor promotes many of these cellular processes, but while MET is often overexpressed in melanoma, the mechanism driving this overexpression is unknown. As the MET gene is rarely mutated or amplified in melanoma, MET overexpression may be driven to increased activation through promoter elements. In this report, we find that transcription factors PAX3 and ETS1 directly interact to synergistically activate MET expression. Inhibition of PAX3 and ETS1 expression in melanoma cells leads to a significant reduction of MET receptor levels. The 300-bp 5' proximal MET promoter contains a PAX3 response element and two ETS1 consensus motifs. Although ETS1 can moderately activate both of these sites without cofactors, robust MET promoter activation of the first site is PAX dependent and requires the presence of PAX3, whereas the second site is PAX independent. The induction of MET by ETS1 via this second site is enhanced by hepatocyte growth factor-dependent ETS1 activation, thereby MET indirectly promotes its own expression. We further find that expression of a dominant-negative ETS1 reduces the ability of melanoma cells to grow both in culture and in vivo. Thus, we discover a pathway where ETS1 advances melanoma through the expression of MET via PAX-dependent and -independent mechanisms.

Protection of neonatal kittens against feline immunodeficiency virus infection with passive maternal antiviral antibodies.

OBJECTIVE: Maternal antibodies from either vaccinated or feline immunodeficiency virus (FIV)-infected female cats (queens) were evaluated for their ability to protect kittens against homologous FIV infection. DESIGN: Kittens that received different levels of maternal antiviral antibodies from either vaccinated or infected queens were inoculated with homologous FIV at 1 week post-parturition and monitored for FIV infection. Maternal antiviral antibodies in the kittens were also measured and compared to the level of FIV infection. METHODS: Kittens at 1 week post-parturition were inoculated intraperitoneally with five median cat infectious doses of FIVPet. FIV infection was monitored by virus isolation for infectious FIV and by nested polymerase chain reaction for proviral DNA. Virus-neutralizing (VN) antibodies and antibodies to FIV transmembrane peptide and core protein were also monitored throughout the 25 weeks. RESULTS: Neonatal kittens that received high levels of antiviral antibodies from either vaccinated or infected queens were protected from FIV inoculation. Kittens that received low levels of maternal antiviral antibodies were not completely protected from similar FIV inoculation. Protection correlated more closely with the level of maternal VN antibodies than the anti-p25 antibodies transferred to the kittens. The unprotected kittens born to infected queens were not infected from vertical transmission because all littermates that were not FIV-inoculated remained free of FIV infection. CONCLUSIONS: Maternal antiviral antibodies, including VN antibodies, from either vaccinated or infected queens protected neonatal kittens from FIV inoculation. Thus, maternal antiviral antibodies play a key role in preventing or limiting infection in neonates and such antiviral immunity can be provided by vaccinated queens.

The glucose-regulated proteins (GRP78 and GRP94): functions, gene regulation, and applications.

The knowledge of GRPs as molecular chaperones is rapidly evolving. It is anticipated that the GRPs will make special contributions in the areas of basic cell biology, biotechnology, and cancer biology. In particular, they may play a role as the prototype of a class of genes that are regulated by signal transduction pathways originating in the ER and traveling to the nucleus. GRP78 and GRP94 function as molecular chaperones and can bind to malfolded proteins and unassembled complexes. They are induced in response to stress, but once the stress is removed the GRPs are posttranscriptionally modified into biologically inactive forms. The promoters of the grp genes are highly conserved, with several CCAAT-like motifs and GC-rich regions. The high level of redundancy that exists in the mammalian grp promoters may act to ensure that the expression of the genes, both of which are single copy, is unlikely to be significantly lowered in the event of mutation. These genes are thought to be controlled by several transcription factors whose complex interactions with the grp promoters allow variable patterns of grp induction. The promoters of the grp genes constitutively express their gene products, and their promoter activities can be further enhanced in cellular environments of low glucose or oxygen. The grp78 promoter is known to retain its strong activity in differentiated and undifferentiated tissues. These features make it an attractive alternative to viral promoters for use in gene therapy. Gene therapy may also be useful in treating cancer in some cases, especially solid tumors. In these instances, GRP levels are already likely to be quite high. These high levels of GRPs may inhibit the efficacy of several anti-cancer treatments. Suppression of GRP induction, perhaps by anti-sense or ribozyme technology, may prove to be useful in conjunction with anti-cancer drugs to treat tumors.

Synthesis, structure-activity relationships, and pharmacokinetic properties of dihydroorotate dehydrogenase inhibitors: 2-cyano-3-cyclopropyl-3-hydroxy-N-[3'-methyl-4'-(trifluoromethyl)phenyl ] propenamide and related compounds.

The active metabolite (2) of the novel immunosuppressive agent leflunomide (1) has been shown to inhibit the enzyme dihydroorotate dehydrogenase (DHODH). This enzyme catalyzes the fourth step in de novo pyrimidine biosynthesis. A series of analogues of the active metabolite 2 have been synthesized. Their in vivo biological activity determined in rat and mouse delayed type hypersensitivity has been found to correlate well with their in vitro DHODH potency. The most promising compound (3) has shown activity in rat and mouse collagen (II)-induced arthritis models (ED50 = 2 and 31 mg/kg, respectively) and has shown a shorter half-life in man when compared with leflunomide. Clinical studies in rheumatoid arthritis are in progress.

Induction of glucose-regulated protein (glucose-regulated protein 78/BiP and glucose-regulated protein 94) and heat shock protein 70 transcripts in the immature rat brain following status epilepticus.

Prior to 21 days of age, the immature rat brain is relatively resistant to excitotoxicity caused by the glutamate analogue, kainate. As stress-inducible proteins (GRP78, GRP94 and HSP70) have been proposed to possess molecular chaperone activity and protect cells from the deleterious effects of damaged proteins, we examined the pattern of expression of their respective messenger RNAs following systemic kainate at different postnatal ages. In untreated rats between seven and 21 days old, there was a higher basal level of grp78 and grp94 expression compared to hsp70. Unlike hsp70, which was inducible only in 21-day-old rats, kainate-mediated grp94 induction occurred in several regions of the brain as early as postnatal day 7. Grp78 messenger RNA expression was also increased by kainate treatment in 14-day-old rats, and the induction was most pronounced in the kainate-resistant dentate gyrus. With increasing age, longer lasting expression of both grp78 and grp94 messenger RNAs was observed in kainate-vulnerable regions, similar to observations in the adult rat brain. These results demonstrate non-overlapping expression patterns of glucose-regulated proteins and HSP70 in the immature central nervous system, suggesting that they serve different functions. While hsp70 induction could be a marker for potential cell injury and death, increased expression of grp78 and grp94 could play a neuroprotective role in the developing rat brain.

The design of a model for a three dimensional stress analysis of the cement layer beneath the medial plateau of a knee prosthesis.

Factors which have influenced the design of a large scale model for an analysis of the strain in three dimensions of the cement layer beneath the medial plateau of a knee prosthesis are discussed. Materials were selected to model the medial tibial plateau, underlying cement and bone for a typical prosthesis and a two dimensional finite element analysis was used to indicate where the strain gauges should be embedded in the model.

A protocol for repetitive somatic embryogenesis from mature peanut epicotyls.

The effects of 11 different auxins and one cytokinin-like compound were tested at four concentrations for their ability to induce primary and repetitive somatic embryos from mature, dry peanut (Arachis hypogaea L.) epicotyls of genotype AT120. Treatment with picloram and centrophenoxine at 83.0 and 124.4 µM resulted in the greatest number of embryos per explant and the highest percentage of explants responding. In a follow-up experiment, picloram, centrophenoxine, and dicamba were tested at 83.0 and 124.4 µM on four peanut genotypes (AT120, 59-4144, GK7, and VC1). Picloram and centrophenoxine induced similar numbers of globular-stage and total embryos from each genotype, while dicamba was less effective. Similar results were observed with percentage of responding axes. Genotypes AT120 and VC1 yielded more clusters of repetitive embryos than GK7 and 59-4144. After 5 months, embryos derived from repetitive embryogenic cultures were converted into mature plants.