Nasopharyngeal Pneumococcal Colonization Density Is Associated With Severe Pneumonia in Young Children in the Lao People's Democratic Republic.

BACKGROUND: No studies have explored the association between pneumococcal nasopharyngeal density and severe pneumonia using the World Health Organization (WHO) 2013 definition. In Lao People's Democratic Republic (Lao PDR), we determine the association between nasopharyngeal pneumococcal density and severe pneumonia in children. METHODS: A prospective observational study was undertaken at Mahosot Hospital, Vientiane, from 2014 to mid-2018. Children <5 years admitted with acute respiratory infections (ARIs) were included. Clinical and demographic data were collected alongside nasopharyngeal swabs for pneumococcal quantification by lytA real-time quantitative polymerase chain reaction. Severe pneumonia was defined using the 2013 WHO definition. For pneumococcal carriers, a logistic regression model examined the association between pneumococcal density and severe pneumonia, after adjusting for potential confounders including demographic and household factors, 13-valent pneumococcal conjugate vaccine status, respiratory syncytial virus co-detection, and preadmission antibiotics. RESULTS: Of 1268 participants with ARI, 32.3% (n = 410) had severe pneumonia and 36.9% (n = 468) had pneumococcal carriage. For pneumococcal carriers, pneumococcal density was positively associated with severe pneumonia (adjusted odds ratio, 1.4 [95% confidence interval, 1.1-1.8]; P = .020). CONCLUSIONS: Among children with ARIs and pneumococcal carriage, pneumococcal carriage density was positively associated with severe pneumonia in Lao PDR. Further studies may determine if pneumococcal density is a useful marker for pneumococcal conjugate vaccine impact on childhood pneumonia.

Actin-binding compounds, previously discovered by FRET-based high-throughput screening, differentially affect skeletal and cardiac muscle.

Actin's interactions with myosin and other actin-binding proteins are essential for cellular viability in numerous cell types, including muscle. In a previous high-throughput time-resolved FRET (TR-FRET) screen, we identified a class of compounds that bind to actin and affect actomyosin structure and function. For clinical utility, it is highly desirable to identify compounds that affect skeletal and cardiac muscle differently. Because actin is more highly conserved than myosin and most other muscle proteins, most such efforts have not targeted actin. Nevertheless, in the current study, we tested the specificity of the previously discovered actin-binding compounds for effects on skeletal and cardiac α-actins as well as on skeletal and cardiac myofibrils. We found that a majority of these compounds affected the transition of monomeric G-actin to filamentous F-actin, and that several of these effects were different for skeletal and cardiac actin isoforms. We also found that several of these compounds affected ATPase activity differently in skeletal and cardiac myofibrils. We conclude that these structural and biochemical assays can be used to identify actin-binding compounds that differentially affect skeletal and cardiac muscles. The results of this study set the stage for screening of large chemical libraries for discovery of novel compounds that act therapeutically and specifically on cardiac or skeletal muscle.

Super-relaxed state of myosin in human skeletal muscle is fiber-type dependent.

The myosin super-relaxed state (SRX) in skeletal muscle is hypothesized to play an important role in regulating muscle contractility and thermogenesis in humans but has only been examined in model organisms. Here we report the first human skeletal muscle SRX measurements, using quantitative epifluorescence microscopy of fluorescent 2'/3'-O-(N-methylanthraniloyl) ATP (mantATP) single-nucleotide turnover. Myosin heavy chain (MHC) isoform expression was determined using gel electrophoresis for each permeabilized vastus lateralis fiber, to allow for novel comparisons of SRX between fiber types. We find that the fraction of myosin in SRX is less in MHC IIA fibers than in MHC I and IIAX fibers (P = 0.008). ATP turnover of SRX is faster in MHC IIAX fibers compared with MHC I and IIA fibers (P = 0.001). We conclude that SRX biochemistry is measurable in human skeletal muscle, and our data indicate that SRX depends on fiber type as classified by MHC isoform. Extension from this preliminary work would provide further understanding regarding the role of SRX in human muscle physiology.

Adherence to 24-hour movement guidelines and academic performance in adolescents.

OBJECTIVE: The aim of the study was to examine the association between meeting combinations of the Canadian 24-h movement guidelines and academic performance in adolescents. STUDY DESIGN: This cross-sectional study used the 2017 Ontario Student Drug Use and Health Survey, a survey representative of Ontario students in grades 7-12 attending publicly funded schools. A total of 10,160 students were included in the analysis. METHODS: Moderate to vigorous physical activity (MVPA), screen time, sleep duration, and academic performance were self-reported. A multiple linear regression model was used to examine differences in academic performance between adolescents meeting and those not meeting the combinations of movement guidelines (≥60 min/day of MVPA; ≤2 h/day of screen time; 9-11 h/night of sleep for ages 11-13 years, 8-10 h/night for ages 14-17 years, and 7-9 h/night for ages 18 years or older). Covariates included age, sex, ethnicity, subjective socio-economic status, body mass index z-score, and substance use. RESULTS: We found that 5.1% of students met all three movement guidelines, whereas 39.0% did not meet any. Middle school students who met all three guidelines or either the screen time or sleep guideline displayed better academic performance than those who met none of the guidelines. High school students who met the screen time and sleep guidelines displayed better academic performance than those who did not meet any guidelines. CONCLUSIONS: Adhering to screen time and sleep duration recommendations is associated with better academic performance among adolescents.

A minor conformation of a lanthanide tag on adenylate kinase characterized by paramagnetic relaxation dispersion NMR spectroscopy.

NMR relaxation dispersion techniques provide a powerful method to study protein dynamics by characterizing lowly populated conformations that are in dynamic exchange with the major state. Paramagnetic NMR is a versatile tool for investigating the structures and dynamics of proteins. These two techniques were combined here to measure accurate and precise pseudocontact shifts of a lowly populated conformation. This method delivers valuable long-range structural restraints for higher energy conformations of macromolecules in solution. Another advantage of combining pseudocontact shifts with relaxation dispersion is the increase in the amplitude of dispersion profiles. Lowly populated states are often involved in functional processes, such as enzyme catalysis, signaling, and protein/protein interactions. The presented results also unveil a critical problem with the lanthanide tag used to generate paramagnetic relaxation dispersion effects in proteins, namely that the motions of the tag can interfere severely with the observation of protein dynamics. The two-point attached CLaNP-5 lanthanide tag was linked to adenylate kinase. From the paramagnetic relaxation dispersion only motion of the tag is observed. The data can be described accurately by a two-state model in which the protein-attached tag undergoes a 23° tilting motion on a timescale of milliseconds. The work demonstrates the large potential of paramagnetic relaxation dispersion and the challenge to improve current tags to minimize relaxation dispersion from tag movements.

Evaluation of capillary and myofiber density in the pectoralis major muscles of rapidly growing, high-yield broiler chickens during increased heat stress.

Skeletal muscle development proceeds from early embryogenesis through marketing age in broiler chickens. While myofiber formation is essentially complete at hatching, myofiber hypertrophy can increase after hatch by assimilation of satellite cell nuclei into myofibers. As the diameter of the myofibers increases, capillary density peripheral to the myofiber is marginalized, limiting oxygen supply and subsequent diffusion into the myofiber, inducing microischemia. The superficial and deep pectoralis muscles constitute 25% of the total body weight in a market-age bird; thus compromise of those muscle groups can have profound economic impact on broiler production. We hypothesized that marginal capillary support relative to the hypertrophic myofibers increases the incidence of microischemia, especially in contemporary high-yield broilers under stressing conditions such as high environmental temperatures. We evaluated the following parameters in four different broiler strains at 39 and 53 days of age when reared under thermoneutral (20 to 25 C) versus hot (30 to 35 C) environmental conditions: capillary density, myofiber density and diameter, and degree of myodegeneration. Our data demonstrate that myofiber diameter significantly increased with age (P > or = 0.0001), while the absolute numbers of capillaries, blood vessels, and myofibers visible in five 400 x microscopic fields decreased (P > or = 0.0001). This is concomitant with marginalization of vascular support in rapidly growing myofibers. The myofiber diameter was significantly lower with hot environmental temperatures (P > or = 0.001); therefore, the absolute number of myofibers visible in five 400X microscopic fields was significantly higher. The incidence and subjective degree of myodegeneration characterized by loss of cross-striations, myocyte hyperrefractility, sarcoplasmic vacuolation, and nuclear pyknosis or loss also increased in hot conditions. Differences among strains were not observed.

Characteristics of traditional Chinese medicine use in children with asthma: a nationwide population-based study.

Asthma is a chronic inflammatory airway disease accounting for severe morbidity and mortality in children. To determine the characteristics of traditional Chinese medicine (TCM) used to treat pediatric asthma, we conducted a nationwide population-based study by analyzing a cohort of one million randomly sampled patients from the beneficiaries of the National Health Insurance Program in Taiwan from 2002 to 2010. Children under 18 years of age with newly diagnosed asthma (ICD-9-CM code: 493, N = 45 833) were enrolled, and 57.95% (N = 26 585) of them had used TCM. The number of TCM users was significantly more than that of non-TCM users in school-age children. The most commonly prescribed TCM formula is Ding-chuan-tang, or Xing-ren (Semen Armeniacae Amarum) for the single herb. Our study is the first to reveal characteristics and prescription patterns of the use of TCM in children with asthma. Further research is needed to elucidate the efficacy and safety of these Chinese herbal products.

Heterogeneity in Blood Biomarker Trajectories After Mild TBI Revealed by Unsupervised Learning.

Concussions, also known as mild traumatic brain injury (mTBI), are a growing health challenge. Approximately four million concussions are diagnosed annually in the United States. Concussion is a heterogeneous disorder in causation, symptoms, and outcome making precision medicine approaches to this disorder important. Persistent disabling symptoms sometimes delay recovery in a difficult to predict subset of mTBI patients. Despite abundant data, clinicians need better tools to assess and predict recovery. Data-driven decision support holds promise for accurate clinical prediction tools for mTBI due to its ability to identify hidden correlations in complex datasets. We apply a Locality-Sensitive Hashing model enhanced by varied statistical methods to cluster blood biomarker level trajectories acquired over multiple time points. Additional features derived from demographics, injury context, neurocognitive assessment, and postural stability assessment are extracted using an autoencoder to augment the model. The data, obtained from FITBIR, consisted of 301 concussed subjects (athletes and cadets). Clustering identified 11 different biomarker trajectories. Two of the trajectories (rising GFAP and rising NF-L) were associated with a greater risk of loss of consciousness or post-traumatic amnesia at onset. The ability to cluster blood biomarker trajectories enhances the possibilities for precision medicine approaches to mTBI.

Exploratory Analysis of Concussion Recovery Trajectories using Multi-modal Assessments and Serum Biomarkers.

Clinicians need better tools to assess severity, prognosis, and recovery from mild Traumatic Brain Injury (mTBI), which can cause long term impairment. To enable better mTBI outcome prediction, an initial step is to analyze the trajectory of recovery metrics over time. This study provides an assessment of recovery trajectories of mTBI while incorporating heterogeneity of individual responses. We analyze the trajectories over multiple discrete time points from baseline to 6 months post injury using a combination of neurocognitive and postural stability assessments and serum biomarkers. The data, obtained from FITBIR, consists of concussed subjects and a matched control group, to allow for comparison in prognostic assessment. Outcomes derived from this exploratory analysis will aid future studies in developing a mTBI recovery timeline model.Clinical relevance- This study further informs clinicians as to the recovery trajectory of clinical measures and biomarkers after mTBI to support return to play decisions. GFAP biomarker and measures related to balance, memory, orientation, and concentration were significantly different than controls early after mTBI.

Drug use and other risk factors related to lower body mass index among HIV-infected individuals.

Malnutrition is associated with morbidity and mortality in HIV-infected individuals. Little research has been conducted to identify the roles that clinical, illicit drug use and socioeconomic characteristics play in the nutritional status of HIV-infected patients. This cross-sectional analysis included 562 HIV-infected participants enrolled in the Nutrition for Healthy Living study conducted in Boston, MA and Providence, RI. The relationship between body mass index (BMI) and several covariates (type of drug use, demographic, and clinical characteristics) were examined using linear regression. Overall, drug users had a lower BMI than non-drug users. The BMI of cocaine users was 1.4 kg/m(2) less than that of patients who did not use any drugs, after adjusting for other covariates (p=0.02). The BMI of participants who were over the age of 55 years was 2.0 kg/m(2) less than that of patients under the age of 35, and BMI increased by 0.3 kg/m(2) with each 100 cells/mm(3) increase in CD4 count. HAART use, adherence to HAART, energy intake, AIDS status, hepatitis B and hepatitis C co-infections, cigarette smoking and depression were not associated with BMI in the final model. In conclusion, BMI was lower in drug users than non-drug users, and was lowest in cocaine users. BMI was also directly associated with CD4 count and inversely related to age more than 55 years old. HIV-infected cocaine users may be at higher risk of developing malnutrition, suggesting the need for anticipatory nutritional support.

Age affects myosin relaxation states in skeletal muscle fibers of female but not male mice.

The recent discovery that myosin has two distinct states in relaxed muscle-disordered relaxed (DRX) and super-relaxed (SRX)-provides another factor to consider in our fundamental understanding of the aging mechanism in skeletal muscle, since myosin is thought to be a potential contributor to dynapenia (age-associated loss of muscle strength independent of atrophy). The primary goal of this study was to determine the effects of age on DRX and SRX states and to examine their sex specificity. We have used quantitative fluorescence microscopy of the fluorescent nucleotide analog 2'/3'-O-(N-methylanthraniloyl) ATP (mantATP) to measure single-nucleotide turnover kinetics of myosin in skinned skeletal muscle fibers under relaxing conditions. We examined changes in DRX and SRX in response to the natural aging process by measuring the turnover of mantATP in skinned fibers isolated from psoas muscle of adult young (3-4 months old) and aged (26-28 months old) C57BL/6 female and male mice. Fluorescence decays were fitted to a multi-exponential decay function to determine both the time constants and mole fractions of fast and slow turnover populations, and significance was analyzed by a t-test. We found that in females, both the DRX and SRX lifetimes of myosin ATP turnover at steady state were shorter in aged muscle fibers compared to young muscle fibers (p ≤ 0.033). However, there was no significant difference in relaxation lifetime of either DRX (p = 0.202) or SRX (p = 0.804) between young and aged male mice. No significant effects were measured on the mole fractions (populations) of these states, as a function of sex or age (females, p = 0.100; males, p = 0.929). The effect of age on the order of myosin heads at rest and their ATPase function is sex specific, affecting only females. These findings provide new insight into the molecular factors and mechanisms that contribute to aging muscle dysfunction in a sex-specific manner.

Swift and NuSTAR observations of GW170817: Detection of a blue kilonova.

With the first direct detection of merging black holes in 2015, the era of gravitational wave (GW) astrophysics began. A complete picture of compact object mergers, however, requires the detection of an electromagnetic (EM) counterpart. We report ultraviolet (UV) and x-ray observations by Swift and the Nuclear Spectroscopic Telescope Array of the EM counterpart of the binary neutron star merger GW170817. The bright, rapidly fading UV emission indicates a high mass (≈0.03 solar masses) wind-driven outflow with moderate electron fraction (Ye ≈ 0.27). Combined with the x-ray limits, we favor an observer viewing angle of ≈30° away from the orbital rotation axis, which avoids both obscuration from the heaviest elements in the orbital plane and a direct view of any ultrarelativistic, highly collimated ejecta (a γ-ray burst afterglow).

Structure-Performance Investigation of Thioxanthone Derivatives for Developing Color Tunable Highly Efficient Thermally Activated Delayed Fluorescence Emitters.

Thioxanthone derivatives consisting of undecorated carbazole as an electron donor and thioxanthone (TXO) or 9H-thioxanthen-9-one-S,S-dioxide (SOXO) as an electron acceptor in a donor-acceptor (D-A) or donor-acceptor-donor (D-A-D) structure were developed as thermally activated delayed fluorescence emitters to fabricate highly efficient fluorescent organic light emitting diodes. Their emission color was successfully tuned from blue to yellow by changing the sulfur atom valence state of the thioxanthone unit to tune intramolecular charge transfer effect. Their thermal, electrochemical, photophysical, and electroluminescent properties, and theoretical calculations were systematically investigated to illustrate the molecular structure and property relationships. Maximum external quantum efficiency (EQE) of 13.6% with Commission Internationale de L'Eclairage coordinates of (0.37, 0.57) was achieved for green light emission CzSOXO consisting of SOXO and carbazole in a D-A structure. Blue light emission CzTXO and DCzTXO consisting of TXO and carbazole in a D-A and D-A-D structure could also give EQE values exceeding 11%. Their efficiency roll-off with increasing current density was simulated by adopting triplet-triplet annihilation model, indicating that the TXO derivatives suffer more severe efficiency roll-off because of their relatively long delayed fluorescence lifetime (τ(D)).

Morphologic changes in tubular cells from in situ kidneys following experimental hypothermia and rewarming.

Although renal failure may occur following rewarming from deep accidental hypothermia, this subject has received little attention in experimental hypothermia and clinical case reports. In order to explore the integrity of hypothermic and posthypothermic renal morphology we used an experimental animal model of accidental hypothermia where the heart supports the circulation throughout cooling and rewarming without accompanying cardioplegia or ischemia. Ultrastructural changes in renal tubular cells from three groups of pentobarbital anesthetized Wistar rats: 1) controls (n=6) maintained at 37 degrees C for 4 h, 2) hypothermic rats (n=6) core-cooled and maintained at 15-13 degrees C for 4 h, and 3) rewarmed rats (n=10), were studied as a sensitive indicator of renal damage. Electron micrographs (EM) from hypothermic kidneys showed rounded up mitochondria with loss of contrast. These changes were observed in several though not all of the biopsies, but they were found in all kidneys. Areas exhibiting focal tubular necrosis were seen on most EM from three of these kidneys. EM from rewarmed kidneys showed alterations of mitochondrial ultrastructure with similarities to those observed after hypothermia, but in general the changes were more prominent. Extracellular edema, intracellular edema, swelling of mitochondria, margination of chromatin, necrosis of single tubular cells, and disrupting necrotic debris into tubular lumen could be found in micrographs from 7 of the 10 kidneys examined. Rewarming from experimental hypothermia, without episodes of ischemia or hypoxia, thus induces ultrastructural changes in renal tubular cells similar to changes observed in acute tubular necrosis, associated with renal failure.

The energy landscape of adenylate kinase during catalysis.

Kinases perform phosphoryl-transfer reactions in milliseconds; without enzymes, these reactions would take about 8,000 years under physiological conditions. Despite extensive studies, a comprehensive understanding of kinase energy landscapes, including both chemical and conformational steps, is lacking. Here we scrutinize the microscopic steps in the catalytic cycle of adenylate kinase, through a combination of NMR measurements during catalysis, pre-steady-state kinetics, molecular-dynamics simulations and crystallography of active complexes. We find that the Mg(2+) cofactor activates two distinct molecular events: phosphoryl transfer (>10(5)-fold) and lid opening (10(3)-fold). In contrast, mutation of an essential active site arginine decelerates phosphoryl transfer 10(3)-fold without substantially affecting lid opening. Our results highlight the importance of the entire energy landscape in catalysis and suggest that adenylate kinases have evolved to activate key processes simultaneously by precise placement of a single, charged and very abundant cofactor in a preorganized active site.

The laminin binding protein p40 is involved in inducing limb abnormality of mouse fetuses as the effects of methoxyacetic acid treatment.

This study is intended to characterize a protein that is linked with mouse limb teratogenicity as the effects of methoxyacetic acid (MAA) treatment. A single dose of MAA (10 mmol/kg body weight) was given by gavage on gestation day (GD) 11, whereas the control group were administered vehicle only. The pregnant mice were killed at 4 h after MAA treatment, and forelimb buds were isolated from both the control and treated group embryos. Proteins from forelimb buds GD 11 + 4 h, which were precipitated out using 40-60% ammonium sulfate, then were analyzed by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (2-D SDS-PAGE) technique. The 2-D gels reveal one protein with 41.6 kDa and pI 6.4, which expression was downregulated after MAA treatment. Tentative protein identification via peptide mass database search and definitive protein identification via a primary sequence database search indicate that the protein matches exactly to 34/67 kDa laminin binding protein (LBP; P14206, SwissProt), which is encoded by p40 gene (MGI:105381). The identity was further verified by Western blotting with an antibody against the 67 kDa LBP. The results suggest that MAA treatment to pregnant mice downregulates the LBP-p40 in the forelimb buds.

Markers of atherosclerosis and inflammation and mortality in patients with HIV infection.

OBJECTIVE: HIV-infected patients are at increased risk for cardiovascular disease, which may be mediated in part by inflammation. Surrogate marker studies suggest an increased prevalence of vascular abnormalities in HIV infection. We examined the association of all-cause mortality in HIV-infected patients with carotid artery intima-media thickness (cIMT) and high-sensitivity C-reactive protein (hsCRP). DESIGN AND METHODS: Baseline risk factors, cIMT and hsCRP were prospectively measured in 327 HIV-infected participants. Follow-up time with median of 3.1 years was calculated from baseline to death or censored dated 7/31/07. Cox Proportional Hazards models were used to study risk factors associated with mortality. RESULTS: Thirty-eight (11.6%) of participants have died since study enrollment. cIMT was significantly higher in those who died and decedents were significantly more likely to have cIMT above the 75th percentile. Those who died had higher hsCRP than those alive and more had hsCRP values above 3mg/L. CD4 count was lower and log(10) viral load was higher in decedents, but antiretroviral regimens were similar in both groups. cIMT and hsCRP levels were significantly associated with mortality (HR = 2.74, 95% CI 1.26-5.97, p = 0.01; HR = 2.38, 95% CI 1.15-4.9, p = 0.02). CONCLUSIONS: Our study demonstrated a strong association of carotid IMT and hsCRP with all-cause death in this HIV-infected population despite being similar with respect to exposure to antiretroviral medications. Together these surrogate markers may be indices of chronic inflammation and unfavorable outcomes in HIV-positive patients.

A teratoproteomics analysis: heat shock protein 70 is upregulated in mouse forelimb bud by methoxyacetic acid treatment.

BACKGROUND: Methoxyacetic acid (MAA) causes fetal limb abnormalities when the substance is administrated on gestation day (GD) 11 in mice. Limb abnormalities are caused mainly by extensive cell death in the mesoderm of the limb plate. This investigation focused on identifying a protein that is linked with mouse limb teratogenicity. METHODS: A single dose of MAA at 10 mmol/kg body weight was administered by gavage on GD 11; controls were administered vehicle only. Dams were killed by cervical dislocation 4 hr after treatment and forelimb buds were isolated from both the control and treated embryos. Proteins in forelimb buds GD 11 + 4 hr were precipitated out using 40-60% ammonium sulfate and were then analyzed by 2D SDS-PAGE. Excised protein spots were identified by mass spectrometry and amino acid internal sequence analysis. Identified protein was further confirmed by Western blotting. RESULTS: Two-dimensional gel analysis indicated that 1 protein spot of 81.7 kDa/pI 7.3 was overexpressed, and the protein matched heat shock protein 70 (HSP70; accession no. P08109, SwissProt). CONCLUSIONS: The results suggest that MAA, when administered to pregnant mice, upregulates HSP70 in the forelimb buds.

The induction of the primordial germ cells in the urodeles.

Xenoplastic recombinates of animal ectodermal caps with the ventral vegetative yolk mass of blastulae of stage 81/2-83/4 ofA. mexicanum, T. alpestris, T. cristatus carnifex andP. waltlii have demonstrated unambiguously that in the urodeles the primordial germ cells-together with other ventro-caudal mesodermal structures-develop from the animal ectodermal moiety of the blastula under an inductive influence emanating from the ventral vegetative yolk mass. Similar recombinates of3H-labeled and unlabeled ectodermal and endodermal components fully support this conclusion.Recombinates of the ventral vegetative yolk mass with different regions of the animal ectodermal hemisphere show that primordial germ cells can be formed by any region of the animal ectodermal hemisphere, including those regions which in normal development will never form them. The number of primordial germ cells formed differs significantly among the various regions, that of the ventral peripheral region being the highest and that of the central, animal region the lowest. The capacity for primordial germ cell formation shows two increasing gradients, one animal-vegetative and the other dorse-ventral (in the peripheral zone). Although accurate measurements could not be made, there seems to be a relation between the number of primordial germ cells formed and the amount of ventro-caudal mesoderm induced.The experiments, moreover, show that notochord differentiation largely or entirely suppresses primordial germ cell formation. Notochord differentiation shows a similar animalvegetative, but an opposite ventro-dorsal increase in frequency (in the peripheral zone) as compared with the capacity for primordial germ cell formation. The notochord-forming gradient in the peripheral regions is mainly due to the inductive action already exerted by the dorsal vegetative yolk mass in the intact blastula prior to isolation and recombination (see control explants). The ventro-dorsal decline in primordial germ cell formation in the peripheral regions is very probably due only to the inhibition of primordial germ cell formation by notochord differentiation (as an expression of dorsal mesoderm induction). Therefore, in the animal ectodermal moiety of the blastula there exists only an animal-vegetative gradient in mesodermal competence.These results make it very likely that in urodeles the primordial germ cells do not arise from predetermined elements such as those demonstrated in anurans, but develop from common, totipotent animal ectodermal cells. The discrepancy in the mode of origin of the primordial germ cells between anurans and urodeles could be due only to pronounced differences in the time of appearance of the germinal cytoplasm (in anurans during oogenesis, in urodeles possibly during determination of the primordial germ cells within the ventro-caudal mesoderm).The differences in site and mode of origin of the primordial germ cells between urodeles and anurans favor a dual phylogenetic origin of the two groups.

Detection of serum hepatitis C virus RNA in HCV antibody-seropositive volunteer blood donors.

Approximately 90% of subjects with chronic hepatitis resulting from hepatitis C virus infection have hepatitis C virus RNA in serum. However, the prevalence of hepatitis C virus RNA in serum from subjects with hepatitis C virus antibody associated with persistent normal liver biochemical values is unclear. Do these subjects have resolved or continuing infection with hepatitis C virus? The aim of this study was to examine whether subjects with hepatitis C virus antibody but normal ALT levels had evidence of ongoing infection. Our study population was divided into four groups. Groups 1, 2 and 3 comprised hepatitis C virus antibody-positive volunteer blood donors. Group 1 was made up of subjects found to be hepatitis C virus antibody-positive on enzyme-linked immunosorbent assay with persistent abnormal ALT levels (59 donors: 53 positive on recombinant immunoblot assay and 6 indeterminate). Group 2 members were hepatitis C virus antibody positive, with persistent normal ALT levels (50 donors: 39 positive on recombinant immunoblot assay and 11 indeterminate). Group 3 members were hepatitis C virus seropositive but negative on second-generation recombinant immunoblot assay (n = 48). Twenty patients (not blood donors) with chronic liver disease who were anti-hepatitis C virus seronegative were used as controls (group 4). Serum samples from all four groups were assayed for hepatitis C virus RNA on reverse transcription and a 40-cycle polymerase chain reaction with a combination of primers from the highly conserved 5'-noncoding and less-conserved third and fourth nonstructural regions. All assays were confirmed on hybridization with an internal probe.(ABSTRACT TRUNCATED AT 250 WORDS)