Testing of Old Reinforced Concrete Bridges, HR-390, 1997

The objective of this research project was to service load test a representative sample of old reinforced concrete bridges (some of them historic and some of them scheduled for demolition) with the results being used to create a database so the performance of similar bridges could be predicted. The types of bridges tested included two reinforced concrete open spandrel arches, two reinforced concrete filled spandrel arches, one reinforced concrete slab bridge, and one two span reinforced concrete stringer bridge. The testing of each bridge consisted of applying a static load at various locations on the bridges and monitoring strains and deflections in critical members. The load was applied by means of a tandem axle dump truck with varying magnitudes of load. At each load increment, the truck was stopped at predetermined transverse and longitudinal locations and strain and deflection data were obtained. The strain data obtained were then evaluated in relation to the strain values predicted by traditional analytical procedures and a carrying capacity of the bridges was determined based on the experimental data. The response of a majority of the bridges tested was considerably lower than that predicted by analysis. Thus, the safe load carrying capacities of the bridges were greater than those predicted by the analytical models, and in a few cases, the load carrying capacities were found to be three or four times greater than calculated values. However, the test results of one bridge were lower than those predicted by analysis and thus resulted in the analytical rating being reduced. The results of the testing verified that traditional analytical methods, in most instances, are conservative and that the safe load carrying capacities of a majority of the reinforced concrete bridges are considerably greater than what one would determine on the basis of analytical analysis alone. In extrapolating the results obtained from diagnostic load tests to levels greater than those placed on the bridge during the load test, care must be taken to ensure safe bridge performance at the higher load levels. To extrapolate the load test results from the bridges tested in this investigation, the method developed by Lichtenstein in NCHRP Project 12-28(13)A was used.

Residual antimalarial concentrations before treatment in patients with malaria from Cambodia: indication of drug pressure.

BACKGROUND: The Thai-Cambodian border has been known as the origin of antimalarial drug resistance for the past 30 years. There is a highly diverse market for antimalarials in this area, and improved knowledge of drug pressure would be useful to target interventions aimed at reducing inappropriate drug use. METHODS: Baseline samples from 125 patients with falciparum malaria recruited for 2 in vivo studies (in Preah Vihear and Pursat provinces) were analyzed for the presence of 14 antimalarials in a single run, by means of a liquid chromatography-tandem mass spectrometry assay. RESULTS: Half of the patients had residual drug concentrations above the lower limit of calibration for at least 1 antimalarial at admission. Among the drugs detected were the currently used first-line drugs mefloquine (25% and 35% of patients) and piperaquine (15% of patients); the first-line drug against vivax malaria, chloroquine (25% and 41% of patients); and the former first-line drug, quinine (5% and 34% patients). CONCLUSIONS: The findings demonstrate that there is high drug pressure and that many people still seek treatment in the private and informal sector, where appropriate treatment is not guaranteed. Promotion of comprehensive behavioral change, communication, community-based mobilization, and advocacy are vital to contain the emergence and spread of parasite resistance against new antimalarials.

Development of an Economic Dust Palliative for Limestone Surfaced Secondary Roads, HR-297, 1990

This research project was directed at laboratory and field evaluation of sodium montmorillonite clay (bentonite) as a dust palliative for limestone surfaced secondary roads. It had been postulated that the electrically charged surfaces of the clay particles could interact with the charged surfaces of the limestone and act as a bonding agent to agglomerate fine (-#200) particulates and also to band the fine particulates to larger (+#200) limestone particles. Laboratory testing using soda ash dispersed bentonite treatment of limestone fines indicated significant improvement of compressive strength and slaking characteristics. It was recommended that the project proceed to field trials and test roads were constructed in Dallas and Adair counties in Iowa. Soda ash dispersed bentonite solutions can be field mixed and applied with conventional spray distribution equipment. A maximum of 1.5% bentonite(by weight of aggregate)can be applied at one time. Higher applications would have to be staged allowing the excess moisture to evaporate between applications. Construction of higher application treatments can be accomplished by adding dry bentonite to the surfacing material and then by dry road mixing. The soda ash water solution can then be spray applied and the treated surfacing material wet mixed by motor graders to a consistency of 3 to 4 inch slump concrete. Two motor graders working in tandem can provide rapid mixing for both methods of construction. Calcium and magnesium chloride treatments are 2 to 3 times more effective in dust reduction in the short term (3-4 months) but are prone to washboarding and potholing due to maintenance restrictions. Bentonite treatment at the 2-3% level is estimated to provide a 30-40% dust reduction over the long term(18-24 months). Normal maintenance blading operations can be used on bentonite treated areas. Vehicle braking characteristics are not adversely affected up to the 3.0% treatment level. The bentonite appears to be functioning as a banding agent to bind small particulates to larger particles and is acting to agglomerate fine particles of limestone. This bonding capability appears recoverable from environmental effects of winter, and from alternating wet and dry periods. The bentonite appears to be able to interact with new applications of limestone maintenance material and maintains a dust reduction capability. Soda ash dispersed bentonite treatment is approximately 10 times more cost effective per percent dust reduction than conventional chloride treatments with respect to time. However,the disadvantage is that there is not the initial dramatic reduction in dust generation as with the chloride treatment. Although dust is reduced 30-40% after treatment there is still dust being generated and the traveling public or residents may not perceive the reduction.

Bentonite Treatment for Economical Dust Reduction on Limestone Surfaced Secondary Roads, HR-351, 1995

This research project was directed at laboratory and field evaluation of sodium montmorillonite clay (Bentonite) as a dust palliative for limestone surfaced secondary roads. It was postulated that the electrically charged surfaces (negative) of the clay particles could interact with the charged surfaces (positive) of the limestone and act as a bonding agent to agglomerate fine (-#200) particulates, and also to bond the fine particulates to larger (+#200) limestone particles. One mile test roads were constructed in Tama, Appanoose, and Hancock counties in Iowa using Bentonite treatment levels (by weight of aggregate) ranging from 3.0 to 12.0%. Construction was accomplished by adding dry Bentonite to the surfacing material and then dry road mixing. The soda ash/water solution (dispersing agent) was spray applied and the treated surfacing material wet mixed by motor graders to a consistency of 2 to 3 inch slump concrete. Two motor graders working in tandem provided rapid mixing. Following wet mixing the material was surface spread and compacted by local traffic. Quantitative and qualitative periodic evaluations and testing of the test roads was conducted with respect to dust generation, crust development, roughness, and braking characteristics. As the Bentonite treatment level increased dust generation decreased. From a cost/benefit standpoint, an optimum level of treatment is about 8% (by weight of aggregate). For roads with light traffic, one application at this treatment level resulted in a 60-70% average dust reduction in the first season, 40-50% in the second season, and 20-30% in the third season. Crust development was rated at two times better than untreated control sections. No discernible trend was evident with respect to roughness. There was no evident difference in any of the test sections with respect to braking distance and braking handling characteristics, under wet surface conditions compared to the control sections. Chloride treatments are more effective in dust reduction in the short term (3-4 months). Bentonite treatment is capable of dust reduction over the long term (2-3 seasons). Normal maintenance blading operations can be used on Bentonite treated areas. Soda ash dispersed Bentonite treatment is estimated to be more than twice as cost effective per percent dust reduction than conventional chloride treatments, with respect to time. However, the disadvantage is that there is not the initial dramatic reduction in dust generation as with the chloride treatment. Although dust is reduced significantly after treatment there is still dust being generated. Video evidence indicates that the dust cloud in the Bentonite treated sections does not rise as high, or spread as wide as the cloud in the untreated section. It also settles faster than the cloud in the untreated section. This is considered important for driving safety of following traffic, and for nuisance dust invasion of residences and residential areas. The Bentonite appears to be functioning as a bonding agent.

Gas-chromatography mass-spectrometry determination of phthalic acid in human urine as a biomarker of folpet exposure.

Agricultural workers are exposed to folpet, but biomonitoring data are limited. Phthalimide (PI), phthalamic acid (PAA), and phthalic acid (PA) are the ring metabolites of this fungicide according to animal studies, but they have not yet been measured in human urine as metabolites of folpet, only PA as a metabolite of phthalates. The objective of this study was thus to develop a reliable gas chromatography-tandem mass spectrometry (GC-MS) method to quantify the sum of PI, PAA, and PA ring-metabolites of folpet in human urine. Briefly, the method consisted of adding p-methylhippuric acid as an internal standard, performing an acid hydrolysis at 100 °C to convert ring-metabolites into PA, purifying samples by ethyl acetate extraction, and derivatizing with N,O-bis(trimethylsilyl)trifluoro acetamide prior to GC-MS analysis. The method had a detection limit of 60.2 nmol/L (10 ng/mL); it was found to be accurate (mean recovery, 97%), precise (inter- and intra-day percentage relative standard deviations <13%), and with a good linearity (R (2) > 0.98). Validation was conducted using unexposed peoples urine spiked at concentrations ranging from 4.0 to 16.1 μmol/L, along with urine samples of volunteers dosed with folpet, and of exposed workers. The method proved to be (1) suitable and accurate to determine the kinetic profile of PA equivalents in the urine of volunteers orally and dermally administered folpet and (2) relevant for the biomonitoring of exposure in workers.

Oxidation of proteins: Basic principles and perspectives for blood proteomics.

Protein oxidation mechanisms result in a wide array of modifications, from backbone cleavage or protein crosslinking to more subtle modifications such as side chain oxidations. Protein oxidation occurs as part of normal regulatory processes, as a defence mechanism against oxidative stress, or as a deleterious processes when antioxidant defences are overcome. Because blood is continually exposed to reactive oxygen and nitrogen species, blood proteomics should inherently adopt redox proteomic strategies. In this review, we recall the biochemical basis of protein oxidation, review the proteomic methodologies applied to analyse redox modifications, and highlight some physiological and in vitro responses to oxidative stress of various blood components.

Comparative analysis of amino acid repeats in rodents and humans

Amino acid tandem repeats, also called homopolymeric tracts, are extremely abundant in eukaryotic proteins. To gain insight into the genome-wide evolution of these regions in mammals, we analyzed the repeat content in a large data set of rat-mouse-human orthologs. Our results show that human proteins contain more amino acid repeats than rodent proteins and that trinucleotide repeats are also more abundant in human coding sequences. Using the human species as an outgroup, we were able to address differences in repeat loss and repeat gain in the rat and mouse lineages. In this data set, mouse proteins contain substantially more repeats than rat proteins, which can be at least partly attributed to a higher repeat loss in the rat lineage. The data are consistent with a role for trinucleotide slippage in the generation of novel amino acid repeats. We confirm the previously observed functional bias of proteins with repeats, with overrepresentation of transcription factors and DNA-binding proteins. We show that genes encoding amino acid repeats tend to have an unusually high GC content, and that differences in coding GC content among orthologs are directly related to the presence/absence of repeats. We propose that the different GC content isochore structure in rodents and humans may result in an increased amino acid repeat prevalence in the human lineage.

Dissecting the role of low-complexity regions in the evolution of vertebrate proteins

Low-complexity regions (LCRs) in proteins are tracts that are highly enriched in one or a few aminoacids. Given their high abundance, and their capacity to expand in relatively short periods of time through replication slippage, they can greatly contribute to increase protein sequence space and generate novel protein functions. However, little is known about the global impact of LCRs on protein evolution. We have traced back the evolutionary history of 2,802 LCRs from a large set of homologous protein families from H.sapiens, M.musculus, G.gallus, D.rerio and C.intestinalis. Transcriptional factors and other regulatory functions are overrepresented in proteins containing LCRs. We have found that the gain of novel LCRs is frequently associated with repeat expansion whereas the loss of LCRs is more often due to accumulation of amino acid substitutions as opposed to deletions. This dichotomy results in net protein sequence gain over time. We have detected a significant increase in the rate of accumulation of novel LCRs in the ancestral Amniota and mammalian branches, and a reduction in the chicken branch. Alanine and/or glycine-rich LCRs are overrepresented in recently emerged LCR sets from all branches, suggesting that their expansion is better tolerated than for other LCR types. LCRs enriched in positively charged amino acids show the contrary pattern, indicating an important effect of purifying selection in their maintenance. We have performed the first large-scale study on the evolutionary dynamics of LCRs in protein families. The study has shown that the composition of an LCR is an important determinant of its evolutionary pattern.

Complete nucleotide sequence of Birmingham IncP alpha plasmids. Compilation and comparative analysis.

The IncP alpha promiscuous plasmid (R18, R68, RK2, RP1 and RP4) comprises 60,099 bp of nucleotide sequence, encoding at least 74 genes. About 40 kb of the genome, designated the IncP core and including all essential replication and transfer functions, can be aligned with equivalent sequences in the IncP beta plasmid R751. The compiled IncP alpha sequence revealed several previously unidentified reading frames that are potential genes. IncP alpha plasmids carry genetic information very efficiently: the coding sequences of the genes are closely packed but rarely overlap, and occupy almost 86% of the genome's nucleotide sequence. All of the 74 genes should be expressed, although there is as yet experimental evidence for expression of only 60 of them. Six examples of tandem-in-frame initiation sites specifying two gene products each are known. Two overlapping gene arrangements occupy different reading frames of the same region. Intergenic regions include most of the 25 promoters; transcripts are usually polycistronic. Translation of most of the open reading frames seems to be initiated independently, each from its own ribosomal binding and initiation site, although, a few cases of coupled translation have been reported. The most frequently used initiation codon is AUG but translation for a few open reading frames begins at GUG or UUG. The most common stop-codon is UGA followed by UAA and then UAG. Regulatory circuits are complex and largely dependent on two components of the central control operon. KorA and KorB are transcriptional repressors controlling at least seven operons. KorA and KorB act synergistically in several cases by recognizing and binding to conserved nucleotide sequences. Twelve KorB binding sites were found around the IncP alpha sequence and these are conserved in R751 (IncP beta) with respect to both sequence and location. Replication of IncP alpha plasmids requires oriV and the plasmid-encoded initiator protein TrfA in combination with the host-encoded replication machinery. Conjugative plasmid transfer depends on two separate regions occupying about half of the genome. The primary segregational stability system designated Par/Mrs consists of a putative site-specific recombinase, a possible partitioning apparatus and a post-segregational lethality mechanism, all encoded in two divergent operons. Proteins related to the products of F sop and P1 par partitioning genes are separately encoded in the central control operon.

In vitro and in vivo evaluation of sunitinib eluting beads

Background: Chemoembolization is used to treat liver malignancies. However recurrence occurs frequently, possibly because of neoangiogenesis triggered by ischemia caused by the embolic agent. In this context, the combination of an embolic agent with an anti-angiogenic drug seems appealing. This study characterizes the in vitro loading and release profile of sunitinib eluting beads of different sizes and their pharmacokinetic profile in a rabbit model. Methods: 70-150 μm and 100-300 μm drug eluting beads (DC Bead, Biocompatibles UK) were loaded by incubation in a sunitinib hydrochloride solution. Drug was quantified by spectrophotometry at 430 nm. Drug release was measured over one-week periods and normalized using an internal standard in 30% ethanol in NaCl 0.9%. New-Zealand white rabbits were used. Eight animals received 0.2 ml of 100-300 μm DC Bead loaded with 6 mg of sunitinib in the hepatic artery (group 1) and 4 animals received 6 mg of sunitinib p.o. (group 2). Half of the animals were sacrificed after 6 hours and half after24 hours. Liver enzymes were measured at 0, 6 and 24 hours in both groups. Plasmatic sunitinib concentration was determined by tandem mass spectroscopy (LC MS/MS) at 0, 1, 2, 3, 4, 5, 6 and 24 hours. At sacrifice, the livers were harvested and sunitinib concentration in liver tissue was assessed by LC MS/MS. Results: High drug loading was obtained for both microsphere bead sizes. Particle shrinking was observed with adsorption of sunitinib. Almost complete release of sunitinib was detected under physiological conditions, with very similar release for 70-150 μm and 100-300 μm (t50%=1.2 h) DC Bead. Conclusions: Sunitinib eluting beads are well tolerated by rabbits when administered in the hepatic artery. No unexpected toxicity was observed. Very high drug concentration can be obtained at the site of embolization with minimal systemic passage.

The origins and impact of primate segmental duplications

Duplicated sequences are substrates for the emergence of new genes and are an important source of genetic instability associated with rare and common diseases. Analyses of primate genomes have shown an increase in the proportion of interspersed segmental duplications (SDs) within the genomes of humans and great apes. This contrasts with other mammalian genomes that seem to have their recently duplicated sequences organized in a tandem configuration. In this review, we focus on the mechanistic origin and impact of this difference with respect to evolution, genetic diversity and primate phenotype. Although many genomes will be sequenced in the future, resolution of this aspect of genomic architecture still requires high quality sequences and detailed analyses.

Association between bipolar disorder and monoamine oxidase A gene polymorphisms: results of a multicenter study

OBJECTIVE: Although genetic factors have been implicated in the etiology of bipolar disorder, no specific gene has been conclusively identified. Given the link between abnormalities in serotonergic neurotransmission and bipolar disorder, a candidate gene association approach was applied to study the involvement of the monoamine oxidase A (MAOA) gene, which codes for a catabolic enzyme of serotonin, in the susceptibility to bipolar disorder. METHOD: In France and Switzerland, 272 patients with bipolar disorder and 122 healthy subjects were typed for three polymorphic markers of the MAOA gene: the MAOA-CA repeat, the MAOA restriction fragment length polymorphism (RFLP), and a repeat directly adjacent to the variable number of tandem repeats (VNTR) locus. RESULTS: A significant difference in the distribution of the alleles for the MAOA-CA repeat was observed between the female bipolar patients and comparison group. CONCLUSIONS: The results obtained in the French and Swiss population confirm findings from two studies conducted in the United Kingdom.

Practical considerations for improving the productivity of mass spectrometry-based proteomics.

Mass spectrometry (MS) is currently the most sensitive and selective analytical technique for routine peptide and protein structure analysis. Top-down proteomics is based on tandem mass spectrometry (MS/ MS) of intact proteins, where multiply charged precursor ions are fragmented in the gas phase, typically by electron transfer or electron capture dissociation, to yield sequence-specific fragment ions. This approach is primarily used for the study of protein isoforms, including localization of post-translational modifications and identification of splice variants. Bottom-up proteomics is utilized for routine high-throughput protein identification and quantitation from complex biological samples. The proteins are first enzymatically digested into small (usually less than ca. 3 kDa) peptides, these are identified by MS or MS/MS, usually employing collisional activation techniques. To overcome the limitations of these approaches while combining their benefits, middle-down proteomics has recently emerged. Here, the proteins are digested into long (3-15 kDa) peptides via restricted proteolysis followed by the MS/MS analysis of the obtained digest. With advancements of high-resolution MS and allied techniques, routine implementation of the middle-down approach has been made possible. Herein, we present the liquid chromatography (LC)-MS/MS-based experimental design of our middle-down proteomic workflow coupled with post-LC supercharging.

A genomic island present along the bacterial chromosome of the Parachlamydiaceae UWE25, an obligate amoebal endosymbiont, encodes a potentially functional F-like conjugative DNA transfer system.

BACKGROUND: The genome of Protochlamydia amoebophila UWE25, a Parachlamydia-related endosymbiont of free-living amoebae, was recently published, providing the opportunity to search for genomic islands (GIs). RESULTS: On the residual cumulative G+C content curve, a G+C-rich 19-kb region was observed. This sequence is part of a 100-kb chromosome region, containing 100 highly co-oriented ORFs, flanked by two 17-bp direct repeats. Two identical gly-tRNA genes in tandem are present at the proximal end of this genetic element. Several mobility genes encoding transposases and bacteriophage-related proteins are located within this chromosome region. Thus, this region largely fulfills the criteria of GIs. The G+C content analysis shows that several modules compose this GI. Surprisingly, one of them encodes all genes essential for F-like conjugative DNA transfer (traF, traG, traH, traN, traU, traW, and trbC), involved in sex pilus retraction and mating pair stabilization, strongly suggesting that, similarly to the other F-like operons, the parachlamydial tra unit is devoted to DNA transfer. A close relatedness of this tra unit to F-like tra operons involved in conjugative transfer is confirmed by phylogenetic analyses performed on concatenated genes and gene order conservation. These analyses and that of gly-tRNA distribution in 140 GIs suggest a proteobacterial origin of the parachlamydial tra unit. CONCLUSIONS: A GI of the UWE25 chromosome encodes a potentially functional F-like DNA conjugative system. This is the first hint of a putative conjugative system in chlamydiae. Conjugation most probably occurs within free-living amoebae, that may contain hundreds of Parachlamydia bacteria tightly packed in vacuoles. Such a conjugative system might be involved in DNA transfer between internalized bacteria. Since this system is absent from the sequenced genomes of Chlamydiaceae, we hypothesize that it was acquired after the divergence between Parachlamydiaceae and Chlamydiaceae, when the Parachlamydia-related symbiont was an intracellular bacteria. It suggests that this heterologous DNA was acquired from a phylogenetically-distant bacteria sharing an amoebal vacuole. Since Parachlamydiaceae are emerging agents of pneumonia, this GI might be involved in pathogenicity. In future, conjugative systems might be developed as genetic tools for Chlamydiales.

The Flexural Fatigue Strength of Prestressed Steel I-Beams, HR-73, 1962

The purpose of this investigation was to study the flexural fatigue strength of two prestressed steel I-beams which had previously been fabricated in connection with a jointly sponsored project under the auspices of the Iowa State Highway Commission. The beams were prestressed by deflecting them under the action of a concentrated load at the center of a simple span, then welding unstressed high strength steel plates to the top and bottom flanges to retain a predetermined amount of prestress. The beams were rolled sections of A36 steel and the plates were USS "T-1" steel. Each of the two test specimens were subjected to an identical repeated loading until a fatigue failure occurred. The loading was designed to produce stresses equivalent to those which would have occurred in a simulated bridge and amounted to 84 percent of a standard H-15 live load including impact. One of the beams sustained 2,469,100 repetitions of load to failure and the other sustained 2,756,100 cycles. Following the fatigue tests, an experimental study was made to determine the state of stress that had been retained in the prestressed steel beams. This information, upon which the calculated stresses of the test could be superimposed, provided a method of correlating the fatigue strength of the beams with the fatigue information available on the two steels involved.