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The Ewald method is applied to accelerate the evaluation of the Green's function (GF) of an infinite equispaced linear array of point sources with linear phasing. Only a few terms are needed to evaluate Ewald sums, which are cast in terms of error functions and exponential integrals, to high accuracy. It is shown analytically that the choice of the standard "optimal" Ewald splitting parameter E0 causes overflow errors at high frequencies (period large compared to the wavelength), and convergence rates are analyzed. A recipe for selecting the Ewald splitting parameter is provided. © 2006 Elsevier Inc. All rights reserved.

The Ewald method is applied to accelerate the evaluation of the Green's function of an infinite periodic phased array of line sources. The Ewald representation for a cylindrical wave is obtained from the known representation for the spherical wave, and a systematic general procedure is applied to extend previous results. Only a few terms are needed to evaluate Ewald sums, which are cast in terms of error functions and exponential integrals, to high accuracy. Singularities and convergence rates are analyzed, and a recipe for selecting the Ewald splitting parameter ε is given to handle both low and high frequency ranges. Indeed, it is shown analytically that the choice of the standard optimal splitting parameter ε0 will cause overflow errors at high frequencies. Numerical examples illustrate the results and the sensitivity of the Ewald representation to the splitting parameter ε. © 2005 IEEE.

An alternative direct procedure for applying the Ewald approach to obtain the Green's function for an array of line sources with 1D periodicity is presented. Furthermore, an algorithm for choosing the Ewald splitting parameter E that extends the efficiency of the method when the periodicity is somewhat larger than a wavelength is derived. In particular, the dyadic Green's function formalism is combined with the Ewald method.

Background A novel handheld point-of-care high-sensitivity cardiac troponin I analyser has recently been introduced to the market. Evaluating its diagnostic performance against laboratory standards is imperative, given the variations in cardiac troponin levels across populations. This study compared the diagnostic performance between the point-of-care high-sensitivity cardiac troponin I assay (Siemens Healthineers Atellica VTLi) and a laboratory high-sensitivity cardiac troponin I assay (Abbott ARCHITECT STAT High Sensitive Troponin-I) performed using blood samples from various populations (overall, male, female, younger and older) of Chinese patients with chest pain.Methods This cross-sectional study included 585 consecutive Chinese patients (age ≥18 year) who presented to an emergency department with chest pain (lasting >5 min) and were managed following the chest pain protocol between 1 August 2023 and 12 June 2024. For both assays, blood samples were collected at two time points (0 hour (initial) and 3 hour (subsequent)). The primary outcome was the diagnostic performance of the two assays, evaluated with their 99th percentile upper reference limits used as the cut-off values for diagnosing myocardial infarction. The gold standard for comparison was the final diagnoses made by attending physicians.Results The point-of-care and laboratory assays exhibited equivalent sensitivity and negative predictive values (both 100%) for blood samples collected at both time points. However, the point-of-care assay outperformed the laboratory assay in terms of specificity (initial: 90.5% to 96.3% vs 79.8% to 94.7%; subsequent: 87.8% to 94.8% vs 77.7% to 92.4%) and positive predictive value (initial: 24.4% to 30.8% vs 11.6% to 23.5%; subsequent: 12.5% to 25.0% vs 5.9% to 18.8%), particularly in older patients.Conclusion The point-of-care assay is recommended for rapid clinical decision-making. Future studies should explore the effects of its integration into clinical practice and the feasibility of using sex–race–age-specific 99th percentile upper reference limits to enhance its diagnostic performance.

Rice (Oryza sativa) is one of the most important cereal crops with a diverse set of pests and natural enemies. Rice fields often support a high diversity of arthropods which contribute significantly to productivity. This diversity is frequently threatened due to indiscriminate applications of pesticides. Our aim was to emphasize on the predator diversity in agrochemical exposed rice field as well as on the impact of surrounding vegetation on beneficial insect diversity. Natural enemies’ data were recorded from randomly selected 10 quadrates by visual observation from each treatment. A total of 5,590 individuals of predators were observed during the study period which included 27 species belonging to 16 families from five orders of arthropods during the kharif season of rice. Statistically, there were no significant differences between the population of general natural enemies such as Odonata, Coleoptera, Hymenoptera, and Araneae in plots with insecticide and control during the different growth stages of rice cultivation. Diversity indices were almost similar in fields where pesticide was sprayed and not sprayed. Our study concluded that natural enemies are conserved by ensuring crop heterogeneity, growing insect-friendly plants (with high levels of nectar and pollen) as border crops, and judicious application of granule insecticide like cartap hydrochloride in a rice agro-ecosystem.

A series of N-acetylated, methionine-containing dipeptides designated AcMet-Aa containing various C-terminal amino acids designated Aa are hydrolyzed in aqueous solution at 50 ºC and 0.95 < pD < 1.10 in the presence of three cis-[Pd(L)(H2O)2]2+ complexes, in which L are bidentate ligands en Me4en, and 3-OH-dtco. The reactions were monitored by 1H-NMR spectroscopy. The rate constant for hydrolytic cleavage of the Met-Aa bond decreases as the steric bulk of the amino acid Aa increases. Correlations to Taft’s Es values were made. The substituents on α-C and β-C atoms lower the rate constant most, those on the Ё-C atom lower it less, and those on the δ-C have no detectable effect. Partial selectivity for leaving amino acid Aa is attributed to differences in the volume of the side chain and to discrimination between leaving groups of similar volume but different branching patterns.

Abstract The recent rise in speed and efficiency of new sequencing technologies have facilitated high-throughput sequencing, assembly and analyses of genomes, advancing ongoing efforts to analyze genetic sequences across major vertebrate groups. Standardized procedures in acquiring high quality DNA and RNA and establishing cell lines from target species will facilitate these initiatives. We provide a legal and methodological guide according to four standards of acquiring and storing tissue for the Genome 10K Project and similar initiatives as follows: four-star (banked tissue/cell cultures, RNA from multiple types of tissue for transcriptomes, and sufficient flash-frozen tissue for 1 mg of DNA, all from a single individual); three-star (RNA as above and frozen tissue for 1 mg of DNA); two-star (frozen tissue for at least 700 μg of DNA); and one-star (ethanol-preserved tissue for 700 μg of DNA or less of mixed quality). At a minimum, all tissues collected for the Genome 10K and other genomic projects should consider each species’ natural history and follow institutional and legal requirements. Associated documentation should detail as much information as possible about provenance to ensure representative sampling and subsequent sequencing. Hopefully, the procedures outlined here will not only encourage success in the Genome 10K Project but also inspire the adaptation of standards by other genomic projects, including those involving other biota.

Abstract Background Matrix extracellular phosphoglycoprotein (MEPE) belongs to a family of small integrin-binding ligand N-linked glycoproteins (SIBLINGs) that play a key role in skeleton development, particularly in mineralization, phosphate regulation and osteogenesis. MEPE associated disorders cause various physiological effects, such as loss of bone mass, tumors and disruption of renal function (hypophosphatemia). The study of this developmental gene from an evolutionary perspective could provide valuable insights on the adaptive diversification of morphological phenotypes in vertebrates. Results Here we studied the adaptive evolution of the MEPE gene in 26 Eutherian mammals and three birds. The comparative genomic analyses revealed a high degree of evolutionary conservation of some coding and non-coding regions of the MEPE gene across mammals indicating a possible regulatory or functional role likely related with mineralization and/or phosphate regulation. However, the majority of the coding region had a fast evolutionary rate, particularly within the largest exon (1467 bp). Rodentia and Scandentia had distinct substitution rates with an increased accumulation of both synonymous and non-synonymous mutations compared with other mammalian lineages. Characteristics of the gene (e.g. biochemical, evolutionary rate, and intronic conservation) differed greatly among lineages of the eight mammalian orders. We identified 20 sites with significant positive selection signatures (codon and protein level) outside the main regulatory motifs (dentonin and ASARM) suggestive of an adaptive role. Conversely, we find three sites under selection in the signal peptide and one in the ASARM motif that were supported by at least one selection model. The MEPE protein tends to accumulate amino acids promoting disorder and potential phosphorylation targets. Conclusion MEPE shows a high number of selection signatures, revealing the crucial role of positive selection in the evolution of this SIBLING member. The selection signatures were found mainly outside the functional motifs, reinforcing the idea that other regions outside the dentonin and the ASARM might be crucial for the function of the protein and future studies should be undertaken to understand its importance.

Abstract Background Cytosolic glutathione transferases (cGST) are a large group of ubiquitous enzymes involved in detoxification and are well known for their undesired side effects during chemotherapy. In this work we have performed thorough phylogenetic analyses to understand the various aspects of the evolution and functional diversification of cGSTs. Furthermore, we assessed plausible correlations between gene duplication and substrate specificity of gene paralogs in humans and selected species, notably in mammalian enzymes and their natural substrates. Results We present a molecular phylogeny of cytosolic GSTs that shows that several classes of cGSTs are more ubiquitous and thus have an older ancestry than previously thought. Furthermore, we found that positive selection is implicated in the diversification of cGSTs. The number of duplicate genes per class is generally higher for groups of enzymes that metabolize products of oxidative damage. Conclusions 1) Protection against oxidative stress seems to be the major driver of positive selection in mammalian cGSTs, explaining the overall expansion pattern of this subfamily; 2) Given the functional redundancy of GSTs that metabolize xenobiotic chemicals, we would expect the loss of gene duplicates, but by contrast we observed a gene expansion of this family, which likely has been favored by: i) the diversification of endogenous substrates; ii) differential tissue expression; and iii) increased specificity for a particular molecule; 3) The increased availability of sequence data from diversified taxa is likely to continue to improve our understanding of the early origin of the different cGST classes.

Abstract Background The mitochondria produce up to 95% of a eukaryotic cell's energy through oxidative phosphorylation. The proteins involved in this vital process are under high functional constraints. However, metabolic requirements vary across species, potentially modifying selective pressures. We evaluate the adaptive evolution of 12 protein-coding mitochondrial genes in 41 placental mammalian species by assessing amino acid sequence variation and exploring the functional implications of observed variation in secondary and tertiary protein structures. Results Wide variation in the properties of amino acids were observed at functionally important regions of cytochrome b in species with more-specialized metabolic requirements (such as adaptation to low energy diet or large body size, such as in elephant, dugong, sloth, and pangolin, and adaptation to unusual oxygen requirements, for example diving in cetaceans, flying in bats, and living at high altitudes in alpacas). Signatures of adaptive variation in the NADH dehydrogenase complex were restricted to the loop regions of the transmembrane units which likely function as protons pumps. Evidence of adaptive variation in the cytochrome c oxidase complex was observed mostly at the interface between the mitochondrial and nuclear-encoded subunits, perhaps evidence of co-evolution. The ATP8 subunit, which has an important role in the assembly of F0, exhibited the highest signal of adaptive variation. ATP6, which has an essential role in rotor performance, showed a high adaptive variation in predicted loop areas. Conclusion Our study provides insight into the adaptive evolution of the mtDNA genome in mammals and its implications for the molecular mechanism of oxidative phosphorylation. We present a framework for future experimental characterization of the impact of specific mutations in the function, physiology, and interactions of the mtDNA encoded proteins involved in oxidative phosphorylation.

Abstract Background Insensitivity of advanced-stage prostate cancer to androgen ablation therapy is a serious problem in clinical practice because it is associated with aggressive progression and poor prognosis. Targeted therapeutic drug discovery efforts are thwarted by lack of adequate knowledge of gene(s) associated with prostate tumorigenesis. Therefore there is the need for studies to provide leads to targeted intervention measures. Here we propose that stable expression of U94, a tumor suppressor gene encoded by human herpesvirus 6A (HHV-6A), could alter gene expression and thereby inhibit the tumorigenicity of PC3 cell line. Microarray gene expression profiling on U94 recombinant PC3 cell line could reveal genes that would elucidate prostate cancer biology, and hopefully identify potential therapeutic targets. Results We have shown that stable expression of U94 gene in PC3 cell line inhibited its focus formation in culture, and tumorigenesis in nude mice. Moreover gene expression profiling revealed dramatic upregulation of FN 1 (fibronectin, 91 ± 16-fold), and profound downregulation of ANGPTL 4 (angiopoietin-like-4, 20 ± 4-fold) in U94 recombinant PC3 cell line. Quantitative real-time polymerase chain reaction (QRT-PCR) analysis showed that the pattern of expression of FN 1 and ANGPTL 4 mRNA were consistent with the microarray data. Based on previous reports, the findings in this study implicate upregulation of FN 1 and downregulation of ANGPTL 4 in the anti tumor activity of U94. Genes with cancer inhibitory activities that were also upregulated include SERPINE 2 (serine/cysteine protease inhibitor 2, 7 ± 1-fold increase) and ADAMTS 1 (a disintegrin-like and metalloprotease with thrombospondin type 1 motif, 7 ± 2-fold increase). Additionally, SPUVE 23 (serine protease 23) that is pro-tumorigenic was significantly downregulated (10 ± 1-fold). Conclusion The dramatic upregulation of FN 1 and downregulation of ANGPTL 4 genes in PC3 cell line stably expressing U94 implicate up-regulation of FN 1 and downregulation of ANGPTL 4 in anti tumor activity of U94. Further studies are necessary to determine functional roles of differentially expressed genes in U94 recombinant PC3 cell line, and hopefully provide leads to potential therapeutic targets in prostate cancer.

The introduction of genome engineering technology has transformed biomedical research, making it possible to make precise changes to genetic information. However, creating an efficient gene-editing system requires a deep understanding of CRISPR technology, and the complex experimental systems under investigation. While Large Language Models (LLMs) have shown promise in various tasks, they often lack specific knowledge and struggle to accurately solve biological design problems. In this work, we introduce CRISPR-GPT, an LLM agent augmented with domain knowledge and external tools to automate and enhance the design process of CRISPR-based gene-editing experiments. CRISPR-GPT leverages the reasoning ability of LLMs to facilitate the process of selecting CRISPR systems, designing guide RNAs, recommending cellular delivery methods, drafting protocols, and designing validation experiments to confirm editing outcomes. We showcase the potential of CRISPR-GPT for assisting non-expert researchers with gene-editing experiments from scratch and validate the agent's effectiveness in a real-world use case. Furthermore, we explore the ethical and regulatory considerations associated with automated gene-editing design, highlighting the need for responsible and transparent use of these tools. Our work aims to bridge the gap between biological researchers across various fields with CRISPR genome engineering technology and demonstrate the potential of LLM agents in facilitating complex biological discovery tasks.

Background: Vaccination remains a potent way to curb the present covid-19 global pandemic., Objectives: To assess knowledge, perception and willingness to receive covid-19 vaccines among tertiary students in Nigeria., Methods: In the descriptive cross-sectional study, a sample size of 750 respondents was randomly selected from a university, polytechnic and college of education (COE) in Osun State, Nigeria. Independent sample T and Pearson correlation tests were used to analyse the responses., Results: There was a significant increase in the percentage score of poor perception, relative to good perception among the university and polytechnic respondents. Among the COE respondents, significant increases in the percentage scores of poor knowledge, perception and willingness to receive covid-19 vaccines, relative to the good variables were observed. Weak positive correlations between knowledge and willingness & perception and willingness to receive covid-19 vaccines among all the respondents were noted. In addition, there was a significant increase in good perception to covid-19 vaccines among university and COE, relative to polytechnic respondents. Asides, a significant increase in good willingness to receive covid-19 vaccines was observed among the university, compared to COE respondents., Conclusion: There is poor knowledge, perception and willingness to receive covid-19 vaccines among tertiary students in Osun State, Nigeria., Competing Interests: None declared., (© 2023 Johnson WA et al.)

Highly dynamic geographical topology, two-direction mobility, and varying traffic density can lead to fairness issues in Vehicular Ad-hoc Networks (VANETs). The Medium Access Control (MAC) protocol plays a vital role in sharing the common wireless channel efficiently between vehicles in a VANET system. However, ensuring fairness between vehicles can be a challenge in designing MAC protocols for VANET systems. The existing protocol, IEEE 802.11 DCF, ensures that the packet transmission rate for a particular vehicle is directly proportional to the amount of time a vehicle spends within a service area, but it does not guarantee that faster vehicles will be able to send the minimum number of packets. Other existing MAC protocols based on IEEE 802.11 are able to provide a minimum amount of data transmission regardless of velocity, but are unable to provide an amount of data transmission that is more proportionate to the time a vehicle spends in the service area. To address the above limitations, we propose a Speed Aware Fairness Enabled MAC (SAFE-MAC) protocol that calculates the residence time of a vehicle in a service area by using mobility metrics such as position, direction, and speed to synthesize the transmission probability of each individual vehicle with respect to its residence time. This is achieved by dynamically altering the values of parameters such as minimum contention window, maximum backoff stage, and retransmission limit in the MAC protocol. We then develop an analytical model to compare the performance of our proposed protocol with contemporary MAC protocols. Numerical analysis results show that our proposed protocol significantly improves fairness among the speed-varying vehicles in VANET.

Signals from two different membrane proteins are combined to modulate how strongly sensory neurons respond to mechanical force.

Viral protein U (Vpu) is a type-III integral membrane protein encoded by Human Immunodeficiency Virus-1 (HIV- 1). It is expressed in infected host cells and plays several roles in viral progeny escape from infected cells, including down-regulation of CD4 receptors. But key structure/function questions remain regarding the mechanisms by which the Vpu protein contributes to HIV-1 pathogenesis. Here we describe expression of Vpu in bacteria, its purification and characterization. We report the successful expression of PelB-Vpu in Escherichia coli using the leader peptide pectate lyase B (PelB) from Erwinia carotovora. The protein was detergent extractable and could be isolated in a very pure form. We demonstrate that the PelB signal peptide successfully targets Vpu to the cell membranes and inserts it as a type I membrane protein. PelB-Vpu was biophysically characterized by circular dichroism and dynamic light scattering experiments and was shown to be an excellent candidate for elucidating structural models.

Root lesion nematodes (Pratylenchus spp.) hinder dryland wheat production worldwide. Montana, a leading U.S. wheat production region, has climatic conditions favorable for Pratylenchus spp. A 2006-2007 statewide soil survey revealed damaging populations of Pratylenchus neglectus, primarily in winter wheat production areas of Montana, whereas P. thornei was not found. Analyses of wheat yields in infested fields revealed negative correlations between yields and spring nematode populations (all P < 0.05 and all R 2 > 0.2). Statewide yield losses due to root lesion nematodes were an estimated 12 and 15% for winter wheat in 2006 and 2007, respectively. A subsequent study conducted in 2008 to 2009 revealed significant differences in reproductive success of P. neglectus among seven rotation treatments (P < 0.001). Nematode populations persisted from spring to fall under fallow, barley, pea, and camelina; increased under winter wheat and canola; and decreased under lentil. Populations were sustained through winter following winter wheat and barley but declined following canola, camelina, pea, lentil, and fallow. A screening of 19 barley lines for resistance to P. neglectus revealed significant variation in resistance among entries (P < 0.001), with 'Harrington' barley displaying promising levels of resistance. Development of resistant wheat cultivars remains a principal goal in managing this nematode.

Background: The complex Drosophila larval peripheral nervous system, capable of monitoring sensory input from the external environment, includes a family of multiple dendritic (md) neurons with extensive dendritic arbors tiling the inner surface of the larval body wall. The class IV multiple dendritic (mdIV) neurons are the most complex with dendritic nerve endings forming direct intimate contacts with epithelial cells of the larval body wall. Functioning as polymodal mechanonociceptors with the ability to respond to both noxious mechanical stimulation and noxious heat, the mdIV neurons are also activated by nanomolar levels of the endogenous reactive oxygen species (ROS), H2O2. Although often associated with tissue damage related to oxidative stress, endogenous ROS have also been shown to function as signaling molecules at lower concentrations. The overall role of ROS in sensory signaling is poorly understood but the acutely sensitive response of mdIV neurons to ROS-mediated activation is consistent with a routine role in the regulation of mdIV neuronal activity. Larvae respond to short wavelength ultraviolet (UVC) light with an immediate and visual system-independent writhing and twisting of the body previously described as a nociceptive response. Molecular and cellular mechanisms mediating this response and potential relationships with ROS generation are not well understood. We have used the UVC-induced writhing response as a model for investigation of the proposed link between endogenous ROS production and mdIV neuron function in the larval body wall., Results: Transgenic inactivation of mdIV neurons caused a strong suppression of UVC-induced writhing behavior consistent with a key role for the mdIV neurons as mediators of the behavioral response. Direct imaging of ROS-activated fluorescence showed that UVC irradiation caused a significant increase in endogenous ROS levels in the larval body wall and transgenic overexpression of antioxidant enzymes strongly suppressed the UVC-induced writhing response. Direct electrophysiological recordings demonstrated that UVC irradiation also increased neuronal activity of the mdIV neurons., Conclusions: Results obtained using UVC irradiation to induce ROS generation provide evidence that UVC-induced writhing behavior is mediated by endogenous production of ROS capable of activating mdIV mechanonociceptors in the larval body wall.

A subset of sensory neurons embedded within the Drosophila larval body wall have been characterized as high-threshold polymodal nociceptors capable of responding to noxious heat and noxious mechanical stimulation. They are also sensitized by UV-induced tissue damage leading to both thermal hyperalgesia and allodynia very similar to that observed in vertebrate nociceptors. We show that the class IV multiple-dendritic(mdIV) nociceptors are also required for a normal larval aversion to locomotion on to a dry surface environment. Drosophila melanogaster larvae are acutely susceptible to desiccation displaying a strong aversion to locomotion on dry surfaces severely limiting the distance of movement away from a moist food source. Transgenic inactivation of mdIV nociceptor neurons resulted in larvae moving inappropriately into regions of low humidity at the top of the vial reflected as an increased overall pupation height and larval desiccation. This larval lethal desiccation phenotype was not observed in wild-type controls and was completely suppressed by growth in conditions of high humidity. Transgenic hyperactivation of mdIV nociceptors caused a reciprocal hypersensitivity to dry surfaces resulting in drastically decreased pupation height but did not induce the writhing nocifensive response previously associated with mdIV nociceptor activation by noxious heat or harsh mechanical stimuli. Larvae carrying mutations in either the Drosophila TRP channel, Painless, or the degenerin/epithelial sodium channel subunit Pickpocket1(PPK1), both expressed in mdIV nociceptors, showed the same inappropriate increased pupation height and lethal desiccation observed with mdIV nociceptor inactivation. Larval aversion to dry surfaces appears to utilize the same or overlapping sensory transduction pathways activated by noxious heat and harsh mechanical stimulation but with strikingly different sensitivities and disparate physiological responses.

In volleyball, blocking is highly correlated with team success. The identification of specific techniques that produce a more successful block would be helpful knowledge for coaches and players. This study compared the traditional, swing, and "chicken wing" blocking techniques in combination with the running step footwork pattern in order to determine which technique enabled athletes to perform a more effective block. High-speed videography (7 cameras, Vicon Motion Analysis System) was used to capture the blocking movements of thirteen female NCAA Division I athletes (age = 19.4 ± 1.19 years, height = 1.82 ± 0.08 m, mass = 70.63 ± 7.96 kg, and years of participation at the collegiate level = 2.23 ± 1.17 years). Each player was familiar with each blocking technique. Reflective markers were placed on the players and in randomized order the players performed 3 blocking trials of each technique. The following dependent variables were assessed: The time it took the athletes to get off the ground and get their hands above (vertically) the net was calculated. The distance the hand reached over the net or hand penetration (displacement between the net and finger in the anterior and vertical planes) was also measured. Lastly, jump height was calculated. Repeated measures ANOVA and post-hoc comparisons were done (α = 0.05). There was no significant difference in the main effect for time to get off the ground (p > 0.05). There was a significant difference in the time to get the hands above the net (p < 0.05). The swing block was best for jump height (p <.001) and hand penetration (p < 0.05). These results can help coaches and players decide which blocking technique will benefit them most as a blocking team and as individual blockers. Key pointsThe swing blocking technique resulted in greater jump heights and increased hand penetration, relative to the traditional and chicken wing blocking techniques.The chicken wing blocking technique resulted in greater jump heights and increased hand penetration, relative to the traditional blocking technique.THE TRADITIONAL BLOCKING TECHNIQUE DOES NOT APPEAR TO PROVIDE ANY COMPETITIVE ADVANTAGE RELATED TO THE VARIABLES OBSERVED DURING THIS STUDY: (1) duration spent getting off of the ground and placing hands over the net, (2) jump height, and (3) hand penetration magnitude.

Controlled organismal growth to an appropriate adult size requires a regulated balance between nutrient resources, feeding behavior and growth rate. Defects can result in decreased survival and/or reproductive capability. Since Drosophila adults do not grow larger after eclosion, timing of feeding cessation during the third and final larval instar is critical to final size. We demonstrate that larval food exit is preceded by a period of increased larval surfacing behavior termed the Intermediate Surfacing Transition (IST) that correlates with the end of larval feeding. This behavioral transition occurred during the larval Terminal Growth Period (TGP), a period of constant feeding and exponential growth of the animal. IST behavior was dependent upon function of a subset of peripheral sensory neurons expressing the Degenerin/Epithelial sodium channel (DEG/ENaC) subunit, Pickpocket1(PPK1). PPK1 neuron inactivation or loss of PPK1 function caused an absence of IST behavior. Transgenic PPK1 neuron hyperactivation caused premature IST behavior with no significant change in timing of larval food exit resulting in decreased final adult size. These results suggest a peripheral sensory mechanism functioning to alter the relationship between the animal and its environment thereby contributing to the length of the larval TGP and determination of final adult size., ((c) 2010 Elsevier Inc. All rights reserved.)

The three Drosophila atypical soluble guanylyl cyclases, Gyc-89Da, Gyc-89Db, and Gyc-88E, have been proposed to act as oxygen detectors mediating behavioral responses to hypoxia. Drosophila larvae mutant in any of these subunits were defective in their hypoxia escape response-a rapid cessation of feeding and withdrawal from their food. This response required cGMP and the cyclic nucleotide-gated ion channel, cng, but did not appear to be dependent on either of the cGMP-dependent protein kinases, dg1 and dg2. Specific activation of the Gyc-89Da neurons using channel rhodopsin showed that activation of these neurons was sufficient to trigger the escape behavior. The hypoxia escape response was restored by reintroducing either Gyc-89Da or Gyc-89Db into either Gyc-89Da or Gyc-89Db neurons in either mutation. This suggests that neurons that co-express both Gyc-89Da and Gyc-89Db subunits are primarily responsible for activating this behavior. These include sensory neurons that innervate the terminal sensory cones. Although the roles of Gyc-89Da and Gyc-89Db in the hypoxia escape behavior appeared to be identical, we also showed that changes in larval crawling behavior in response to either hypoxia or hyperoxia differed in their requirements for these two atypical sGCs, with responses to 15% oxygen requiring Gyc-89Da and responses to 19 and 25% requiring Gyc-89Db. For this behavior, the identity of the neurons appeared to be critical in determining the ability to respond appropriately.

Growth of multicellular organisms proceeds through a series of precisely timed developmental events requiring coordination between gene expression, behavioral changes, and environmental conditions. In Drosophila melanogaster larvae, the essential midthird instar transition from foraging (feeding) to wandering (non-feeding) behavior occurs prior to pupariation and metamorphosis. The timing of this key transition is coordinated with larval growth and size, but physiological mechanisms regulating this process are poorly understood. Results presented here show that Drosophila larvae associate specific environmental conditions, such as temperature, with food in order to enact appropriate foraging strategies. The transition from foraging to wandering behavior is associated with a striking reversal in the behavioral responses to food-associated stimuli that begins early in the third instar, well before food exit. Genetic manipulations disrupting expression of the Degenerin/Epithelial Sodium Channel subunit, Pickpocket1(PPK1) or function of PPK1 peripheral sensory neurons caused defects in the timing of these behavioral transitions. Transient inactivation experiments demonstrated that sensory input from PPK1 neurons is required during a critical period early in the third instar to influence this developmental transition. Results demonstrate a key role for the PPK1 sensory neurons in regulation of important behavioral transitions associated with developmental progression of larvae from foraging to wandering stage.

Coordination of rhythmic locomotion depends upon a precisely balanced interplay between central and peripheral control mechanisms. Although poorly understood, peripheral proprioceptive mechanosensory input is thought to provide information about body position for moment-to-moment modifications of central mechanisms mediating rhythmic motor output. Pickpocket1 (PPK1) is a Drosophila subunit of the epithelial sodium channel (ENaC) family displaying limited expression in multiple dendritic (md) sensory neurons tiling the larval body wall and a small number of bipolar neurons in the upper brain. ppk1 null mutant larvae had normal external touch sensation and md neuron morphology but displayed striking alterations in crawling behavior. Loss of PPK1 function caused an increase in crawling speed and an unusual straight path with decreased stops and turns relative to wild-type. This enhanced locomotion resulted from sustained peristaltic contraction wave cycling at higher frequency with a significant decrease in pause period between contraction cycles. The mutant phenotype was rescued by a wild-type PPK1 transgene and duplicated by expressing a ppk1RNAi transgene or a dominant-negative PPK1 isoform. These results demonstrate that the PPK1 channel plays an essential role in controlling rhythmic locomotion and provide a powerful genetic model system for further analysis of central and peripheral control mechanisms and their role in movement disorders.

The ability to detect salt is critical for the survival of terrestrial animals. Based on amiloride-dependent inhibition, the receptors that detect salt have been postulated to be DEG/ENaC channels. We found the Drosophila DEG/ENaC genes Pickpocket11 (ppk11) and Pickpocket19 (ppk19) expressed in the larval taste-sensing terminal organ and in adults on the taste bristles of the labelum, the legs, and the wing margins. When we disrupted PPK11 or PPK19 function, larvae lost their ability to discriminate low concentrations of Na(+) or K(+) from water, and the electrophysiologic responses to low salt concentrations were attenuated. In both larvae and adults, disrupting PPK11 or PPK19 affected the behavioral response to high salt concentrations. In contrast, the response of larvae to sucrose, pH 3, and several odors remained intact. These results indicate that the DEG/ENaC channels PPK11 and PPK19 play a key role in detecting Na(+) and K(+) salts.

This article explores late-onset crack use among midlife and older adults as an emerging risk factor for HIV infection. Most research on illicit drug use and HIV infection/AIDS has focused on younger drug users, typically those who inject. The initiation into crack use during later adulthood challenges this narrow view. The analysis the authors present was drawn from qualitative and quantitative data collected as part of their ongoing research on illicit drug use. The subsample consisted of 27 men who began using crack at the age of 50 or older and 40 women who started using crack at the age of 35 or older. The findings suggest a typology of late-onset users with differing forms of HIV risk and prevention needs. The authors end the article with recommendations for effective HIV risk reduction programs for late-onset crack users.

The Drosophila tracheal system and mammalian airways are branching networks of tubular epithelia that deliver oxygen to the organism. In mammals, the epithelial Na(+) channel (ENaC) helps clear liquid from airways at the time of birth and removes liquid from the airspaces in adults. We tested the hypothesis that related Drosophila degenerin (DEG)/ENaC family members might play a similar role in the fly. Among 16 Drosophila DEG/ENaC genes, called pickpocket (PPK) genes, we found 9 expressed in the tracheal system. By in situ hybridization, expression appeared in late-stage embryos after tracheal tube formation, with individual PPK genes showing distinct temporal and spatial expression patterns as development progressed. Promoters for several PPK genes drove reporter gene expression in the larval and adult tracheal systems. Adding the DEG/ENaC channel blocker amiloride to the medium inhibited liquid clearance from the trachea of first instar larvae. Moreover, when RNA interference was used to silence PPK4 and PPK11, larvae failed to clear tracheal liquid. These data suggest substantial molecular diversity of DEG/ENaC channel expression in the Drosophila tracheal system where the PPK proteins likely play a role in Na(+) absorption. Extensive similarities between Drosophila and mammalian airways offer opportunities for genetic studies that may decipher further the structure and function of DEG/ENaC proteins and development of the airways.

Axonal selection of synaptic partners is generally believed to determine wiring specificity in the nervous system. However, we have recently found evidence for specific dendritic targeting in the olfactory system of Drosophila: second order olfactory neurons (Projection Neurons) from the anterodorsal (adPN) and lateral (lPN) lineages send their dendrites to stereotypical, intercalating but non-overlapping glomeruli. Here we show that POU domain transcription factors, Acj6 and Drifter, are expressed in adPNs and lPNs respectively, and are required for their dendritic targeting. Moreover, misexpression of Acj6 in lPNs, or Drifter in adPNs, results in dendritic targeting to glomeruli normally reserved for the other PN lineage. Thus, Acj6 and Drifter translate PN lineage information into distinct dendritic targeting specificity. Acj6 also controls stereotypical axon terminal arborization of PNs in a central target, suggesting that the connectivity of PN axons and dendrites in different brain centers is coordinately regulated.

Objectives: To assess HIV prevalence, incidence, and associated risk factors among IDUs in Chicago., Methods: Seven hundred ninety-four street-recruited IDUs ranging in age from 18 to 50 years, who were not in drug treatment at study enrollment, were interviewed and tested for HIV at baseline and at two follow-ups scheduled 6 and 12 months after baseline. Questionnaires assessed respondents' demographic characteristics, medical and drug treatment histories, drug use, and sexual practices., Results: HIV seroprevalence at baseline was 18%. Logistic regression identified the following determinants of prevalent HIV infection: Puerto Rican ethnicity, homosexual or bisexual self-identification, injecting for 4 or more years, and having smoked crack cocaine in the past 6 months. Follow-up data were collected from 584 (73.6%) participants. Mean duration of follow-up was 16.5 months, indicating that most subjects had follow-up intervals longer than the scheduled 6 and 12 months. Seven HIV seroconversions were observed in 632 person years of risk, yielding an incidence rate of 1.1 per 100 person years of risk. Injection for 3 or less years was positively associated with HIV seroconversion., Conclusions: The findings provide evidence of a decline in HIV incidence among IDUs, though newer injectors remain at elevated risk for infection.

The Drosophila Vestigial protein has been shown to play an essential role in the regulation of cell proliferation and differentiation within the developing wing imaginal disc. Cell-specific expression of vg is controlled by two separate transcriptional enhancers. The boundary enhancer controls expression in cells near the dorsoventral (DV) boundary and is regulated by the Notch signal transduction pathway, while the quadrant enhancer responds to the Decapentaplegic and Wingless morphogen gradients emanating from cells near the anteroposterior (AP) and DV boundaries, respectively. MAD-dependent activation of the vestigial quadrant enhancer results in broad expression throughout the wing pouch but is excluded from cells near the DV boundary. This has previously been thought to be due to direct repression by a signal from the DV boundary; however, we show that this exclusion of quadrant enhancer-dependent expression from the DV boundary is due to the absence of an additional essential activator in those cells. The Drosophila POU domain transcriptional regulator, Drifter, is expressed in all cells within the wing pouch expressing a vgQ-lacZ transgene and is also excluded from the DV boundary. Viable drifter hypomorphic mutations cause defects in cell proliferation and wing vein patterning correlated with decreased quadrant enhancer-dependent expression. Drifter misexpression at the DV boundary using the GAL4/UAS system causes ectopic outgrowths at the distal wing tip due to induction of aberrant Vestigial expression, while a dominant-negative Drifter isoform represses expression of vgQ-lacZ and causes severe notching of the adult wing. In addition, we have identified an essential evolutionarily conserved sequence element bound by the Drifter protein with high affinity and located adjacent to the MAD binding site within the quadrant enhancer. Our results demonstrate that Drifter functions along with MAD as a direct activator of Vestigial expression in the wing pouch.

During Drosophila embryogenesis the CNS midline cells have organizing activities that are required for proper elaboration of the axon scaffold and differentiation of neighboring neuroectodermal and mesodermal cells. CNS midline development is dependent on Single-minded (Sim), a basic-helix-loop-helix (bHLH)-PAS transcription factor. We show here that Fish-hook (Fish), a Sox HMG domain protein, and Drifter (Dfr), a POU domain protein, act in concert with Single-minded to control midline gene expression. single-minded, fish-hook, and drifter are all expressed in developing midline cells, and both loss- and gain-of-function assays revealed genetic interactions between these genes. The corresponding proteins bind to DNA sites present in a 1 kb midline enhancer from the slit gene and regulate the activity of this enhancer in cultured Drosophila Schneider line 2 cells. Fish-hook directly associates with the PAS domain of Single-minded and the POU domain of Drifter; the three proteins can together form a ternary complex in yeast. In addition, Fish can form homodimers and also associates with other bHLH-PAS and POU proteins. These results indicate that midline gene regulation involves the coordinate functions of three distinct types of transcription factors. Functional interactions between members of these protein families may be important for numerous developmental and physiological processes.

Mutations in the Drosophila class IV POU domain gene, abnormal chemosensory jump 6 (acj6), have previously been shown to cause physiological deficits in odor sensitivity. However, loss of Acj6 function also has a severe detrimental effect upon coordinated larval and adult movement that cannot be explained by the simple loss in odorant detection. In addition to olfactory sensory neurons, Acj6 is expressed in a distinct subset of postmitotic interneurons in the central nervous system from late embryonic to adult stages. In the larval and adult brain, Acj6 is highly expressed in central brain, optic and antennal lobe neurons. Loss of Acj6 function in larval optic lobe neurons results in disorganized retinal axon targeting and synapse selection. Furthermore, the lamina neurons themselves exhibit disorganized synaptic arbors in the medulla of acj6 mutant pupal brains, suggesting that Acj6 may play a role in regulating synaptic connections or structure. To further test this hypothesis, we misexpressed two Acj6 isoforms in motor neurons where they are not normally found. The two Acj6 isoforms are produced from alternatively spliced acj6 transcripts, resulting in significant structural differences in the amino-terminal POU IV box. Acj6 misexpression caused marked alterations at the neuromuscular junction, with contrasting effects upon nerve terminal branching and synapse formation associated with specific Acj6 isoforms. Our results suggest that the class IV POU domain factor, Acj6, may play an important role in regulating synaptic target selection by central neurons and that the amino-terminal POU IV box is important for regulation of Acj6 activity.

Little is known about how the odor specificities of olfactory neurons are generated, a process essential to olfactory coding. We have found that neuronal identity relies on the abnormal chemosensory jump 6 (acj6) gene, originally identified by a defect in olfactory behavior. Physiological analysis of individual olfactory neurons shows that in acj6 mutants, a subset of neurons acquires a different odorant response profile. Certain other neurons do not respond to any tested odors in acj6. Molecular analysis of acj6 shows that it encodes a POU-domain transcription factor expressed in olfactory neurons. Our data suggest that the odor response spectrum of an olfactory neuron, and perhaps the choice of receptor genes, is determined through a process requiring the action of Acj6.

Competitive PCR was used to monitor the survival of a 520-bp DNA target sequence from a recombinant plasmid, pVACMC1, after admixture of the plasmid with freshly sampled human saliva. The fraction of the target remaining amplifiable ranged from 40 to 65% after 10 min of exposure to saliva samples from five subjects and from 6 to 25% after 60 min of exposure. pVACMC1 plasmid DNA that had been exposed to degradation by fresh saliva was capable of transforming naturally competent Streptococcus gordonii DL1 to erythromycin resistance, although transforming activity decreased rapidly, with a half-life of approximately 50 s. S. gordonii DL1 transformants were obtained in the presence of filter-sterilized saliva and a 1-microg/ml final concentration of pVACMC1 DNA. Addition of filter-sterilized saliva instead of heat-inactivated horse serum to S. gordonii DL1 cells induced competence, although with slightly lower efficiency. These findings indicate that DNA released from bacteria or food sources within the mouth has the potential to transform naturally competent oral bacteria. However, further investigations are needed to establish whether transformation of oral bacteria can occur at significant frequencies in vivo.

Drosophila melanogaster has proven to be a good model for understanding the physiology of ion channels. We identified two novel Drosophila DEG/ ENaC proteins, Pickpocket (PPK) and Ripped Pocket (RPK). Both appear to be ion channel subunits. Expression of RPK generated multimeric Na+ channels that were dominantly activated by a mutation associated with neurodegeneration. Amiloride and gadolinium, which block mechanosensation in vivo, inhibited RPK channels. Although PPK did not form channels on its own, it associated with and reduced the current generated by a related human brain Na+ channel. RPK transcripts were abundant in early stage embryos, suggesting a role in development. In contrast, PPK was found in sensory dendrites of a subset of peripheral neurons in late stage embryos and early larvae. In insects, such multiple dendritic neurons play key roles in touch sensation and proprioception and their morphology resembles human mechanosensory free nerve endings. These results suggest that PPK may be a channel subunit involved in mechanosensation.

We have identified the Drosophila UDP-glucose dehydrogenase gene as being involved in wingless signaling. Mutations in this gene, called kiwi, generate a phenotype identical to that of wingless. UDP-glucose dehydrogenase is required for the biosynthesis of UDP-glucuronate, which in turn is utilized in the biosynthesis of glycosaminoglycans. By rescuing the kiwi phenotype with both UDP-glucuronate and the glycosaminoglycan heparan sulfate, we show that kiwi function in the embryo is crucial for the production of heparan sulfate in the extracellular matrix. Further, injection of heparin degrading enzyme, heparinase (and not chondroitin, dermatan or hyaluronic acid degrading enzyme) into wild-type embryos leads to the degradation of heparin-like glycosaminoglycans and a 'wingless-like' cuticular phenotype. Our study thus provides the first genetic evidence for the involvement of heparin-like glycosaminoglycans in signal transduction.

Organogenesis of the Drosophila tracheal system involves extensive directed cell migrations leading to a stereotypic series of interconnected tubules. Although numerous gene products have been shown to be essential for tracheal morphogenesis, direct functional relationships between participants have not been previously established. Both the breathless gene, encoding a Drosophila fibroblast growth factor receptor tyrosine kinase homologue, and the POU-domain transcription factor gene, drifter, are expressed in all tracheal cells and are essential for directed cell migrations. We demonstrate here that ubiquitously expressed Breathless protein under control of a heterologous heat-shock promoter is able to rescue the severely disrupted tracheal phenotype associated with drifter loss-of-function mutations. In the absence of Drifter function, breathless expression is initiated normally but transcript levels fall drastically to undetectable levels as tracheal differentiation proceeds. In addition, breathless regulatory DNA contains seven high affinity Drifter binding sites similar to previously identified Drifter recognition elements. These results suggest that the Drifter protein, which maintains its own expression through a tracheal-specific autoregulatory enhancer, is not necessary for initiation of breathless expression but functions as a direct transcriptional regulator necessary for maintenance of breathless transcripts at high levels during tracheal cell migration. This example of a mechanism for maintenance of a committed cell fate offers a model for understanding how essential gene activities can be maintained throughout organogenesis.

Transcriptional regulators utilizing the POU domain DNA-binding motif have been shown to form multi-protein complexes dependent on the POU domain itself and its flexible recognition of various octamer sequence elements. We have identified two variant POU domain recognition elements DFRE1 and DFRE2, which are found within a 514-bp autoregulatory enhancer of the Drosophila melanogaster POU domain gene drifter (dfr). Both elements are capable of binding bacterially produced full-length DFR protein with high affinity, although they differ in the 5'-to-3' orientation of POU-specific and POU homeodomain subelements. When placed in dfr loss-of-function genetic backgrounds, all expression of dfr-lacZ fusion genes under control of the autoregulatory enhancer is dependent on DFR activity levels. However, the complete enhancer sequence directs beta-galactosidase expression in only a subset of cells which normally express the endogenous DFR protein, including the middle pair of midline glias of the ventral nerve cord, the oenocyte clusters, and all tracheal cells. In addition, DFRE1 and DFRE2 exhibit separable tissue-specific functions when independently disrupted or deleted. Disruption of DFRE1 function specifically abolishes beta-galactosidase expression in the middle pair of midline glias. Deletion of DFRE causes a specific loss of tracheal expression, leaving oenocyte and midline glia expression intact. These results suggest that dfr cell-specific autoregulation is determined by the context of DFR POU domain binding within the enhancer, which is possibly mediated by the formation of recognition element-specific heteromultimeric complexes containing additional tissue-specific factors.

A fundamental question in the basic biology of aging is whether there is a universal aging process. If indeed such a process exists, one would expect that it develops at a higher rate in short- versus long-lived species. We have quantitated pentosidine, a marker of glycoxidative stress in skin collagen from eight mammalian species as a function of age. A curvilinear increase was modeled for all species, and the rate of increase correlated inversely with maximum life-span. Dietary restriction, a potent intervention associated with increased life-span, markedly inhibited glycoxidation rate in the rodent. On the assumption that collagen turnover rate is primarily influenced by the crosslinking due to glycoxidation, these results suggest that there is a progressive age-related deterioration of the process that controls the collagen glycoxidation rate. Thus, the ability to withstand damage due to glycoxidation and the Maillard reaction may be under genetic control.

The clinical diagnosis, treatment management, and epidemiology of a case of unilateral posterior ophthalmomyiasis in a university farm manager are reported. Subretinal tracking was prominent, leading to vision abnormalities, which initially prompted the patient to seek medical evaluation. Identification of the organism based on study of photographs was judged to be a first-instar larva of a Cuterebra (rodent bot fly), although identification of first-instar (stage) myiasis-producing fly larvae is impossible. Inasmuch as the patient was a herdsman, it is likely that the larva is of the genus Hypoderma (cattle grub) or possibly Gasterophilus (horse bot). Laser treatment was effectively used to destroy and immobilize the organism. Subsequently, the patient's health remains satisfactory, and his vision has improved. This case is reported to confirm increasing frequency of ophthalmomyiasis and to improve awareness of its features. Early recognition of this condition, when tumors and other conditions remain suspect, would preclude unnecessarily invasive surgical procedures, including enucleation.

The Drosophila drifter (dfr) gene, previously referred to as Cf1a, encodes a POU-domain DNA-binding protein implicated as a neuron-specific regulator in the developing central nervous system (CNS). We have isolated full-length dfr cDNA clones that encode a 46-kD protein containing the conserved POU-domain DNA-binding domain. The use of alternate polyadenylation sites produces two dfr mRNA transcripts that are first expressed in stage 10 embryos at 5- to 6-hr of development. A specific anti-dfr polyclonal antiserum generated against a dfr-glutathione S-transferase fusion protein recognizes a 46-kD protein on Western blots and has been used to analyze the cell-specific distribution of dfr protein during embryonic development. dfr protein is distributed in a complex expression pattern including the tracheal system, the middle pair of midline glia, and selected CNS neurons. We have carried out a genetic characterization of the dfr locus, previously localized to region 65D of the third chromosome, by generating a series of overlapping deficiencies between 65A and 65E1 that were used to isolate dfrE82, an EMS-induced lethal allele. Analysis of dfrE82 mutant embryos shows a disruption of the developing tracheal tree as well as commissural defects in the developing CNS. Based on an examination of a cell-specific marker for tracheal cells and midline glia, these defects appear to be caused by a failure of these cells to follow their characteristic routes of migration. The dfrE82 tracheal phenotype is rescued by a dfr minigene present as a P-element transposon expressing wild-type dfr protein in tracheal cells. These results suggest that the dfr protein plays a fundamental role in the differentiation of tracheal cells and midline glia possibly by regulating the expression of essential cell-surface proteins required for cell-cell interactions involved in directed cell migrations.

Several genes have been identified that affect the specification of unique neuronal identities during development of the Drosophila central nervous system. At least two of these genes, fushi tarazu (ftz) and even-skipped (eve) share a common structural motif, the homoeobox, which encodes a sequence-specific DNA-binding domain (homoeodomain). A family of related proteins has been recently characterized in mammals and nematodes that contain a diverged homoedomain as part of a structure referred to as a POU domain. Mammalian genes encoding POU domains have region-specific patterns of expression in the central nervous system, indicating a potential role for them in the regulation of neuronal development. The nematode POU-domain gene, unc-86, is involved in the determination of neuroblast lineages leading to serotonergic and dopaminergic neurons. We have now identified a Drosophila gene, Cfla, encoding a sequence-specific DNA-binding protein containing a highly conserved POU domain. The Cfla gene product binds to a DNA element required for expression of the dopa decarboxylase gene (Ddc) in selected dopaminergic neurons, implying that it functions as a neuron-specific transcription factor.

The dopa decarboxylase gene (Ddc) of Drosophila melanogaster is expressed in the hypoderm and the nervous system and promoter elements mediating differential expression in these tissues have been identified (Scholnick et al., 1986). Here we report an additional mode of regulation; the unique primary transcript of the Ddc gene is spliced to form mRNAs in these two tissues which differ by a single internal exon. In vitro mutagenesis and P-element-mediated transformation were employed to manipulate the tissue-specific expression of these RNAs. This approach demonstrated that regulated splicing rather than differential stability causes the tissue-specific expression of these RNAs and allowed the identification of Ddc enzyme isoforms encoded by each mRNA. The Ddc enzyme in the central nervous system differs from the hypodermal Ddc protein by the addition of 33-35 amino acids on the N terminus.

A study was conducted to determine the effects of an environmentally induced force molt treatment on concentrations and daily variations of selected serum biochemicals during early molt. Laying hens were placed into two environmental control chambers. In one chamber, control (Con) laying hens were maintained on long photoperiods of 19 hr of light daily to maintain egg production. In the other chamber, early molt (EM) hens were exposed to short photoperiods of 6 hr of light daily to decrease egg production. After 2 weeks in the chambers, feed was withdrawn from EM hens for 2 days to initiate molt. Three days after feed was restored, hens from both groups were blood sampled every 2 hr for 2 days for selected serum biochemical analyses. Serum concentrations of calcium (Ca), inorganic phosphate (P), glutamic pyruvic transaminase (GPT), and albumin (Alb) were depressed in EM hens. Serum concentrations of alkaline phosphatase (Alk P), lactic acid dehydrogenase (LDH), glutamic oxaloacetic transaminase (GOT), glucose (Glu), cholesterol (Chol), and globulin (Glob) were elevated in EM hens. Daily temporal rhythms of Ca, Glu, total protein (TP), and Alb were found in both hen groups. These temporal changes were independent of force molt treatment. Serum GOT was positively correlated with nuclei free liver corticosteroids (LCS) and negatively associated with plasma corticosterone in Con hens. Serum GOT was positively correlated with nuclei free LCS in EM hens.

Circulating levels of corticosterone were determined in chick embryos from 10 to 21 days of incubation using eggs from a Leghorn breeder flock. In Experiment 1, eggs were incubated from 10 to 20 days for daily embryonic blood collection. To verify stage of development with day of incubation, embryo right middle toe lengths were measured concurrent with blood sampling. Serum from three embryos was pooled into one sample and the corticosterone content of 10 samples per day of incubation was determined using a radioimmunoassay procedure. The levels of corticosterone from day 10 to 14 fluctuated slightly and then increased rapidly until 16 days of incubation. At this time serum corticosterone remained relatively constant through day 18 with an apparent increasing trend up to day 20. The use of toe lengths to assure no day-to-day overlap in embryonic development proved effective. In Experiment 2, newly hatched (day 21) chicks were sorted into four 3-hr periods ranging from early to late hatching. Blood samples were collected from five individual chicks during four 15-min sampling periods for each of the four hatch times. Serum corticosterone levels were not affected by sampling periods or hatch times.