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1. This study used 4 wheat cultivars (Brigadier, Chaucer, Consort, Reaper) from three locations (Crossnacreevy, C; Downpatrick, D; Limavady, L), which had given rise to differences in wheat specific weight (SW), to examine the relationships between apparent metabolisable energy (AME) concentration, broiler performance and wheat SW. 2. The diets contained (g/kg): wheat 744, casein 142, blended vegetable fat 50, dicalcium phosphate 22, potassium bicarbonate 10.8, sodium bicarbonate 7.5, arginine 5, methionine 2, binder 8, trace minerals/vitamins 7.2, titanium dioxide 1.5. The diets were heat-treated (80 degrees C for 2 min) prior to pelleting (3 mm die). 3. SW ranged from 63 to 77 kg/hectolitre (hl), averaging 66, 69 and 76 kg/hl at D, C and L, respectively. In vitro viscosity of the wheat samples ranged from 5.2 to 17.5 cps and thousand grain weight (TGW) from 33.4 to 47.3 g. Mean TGW was similar at C and D (38.7, 37.0 g) but higher at L (43.1 g). In vitro viscosity was similar at C and L (11.2, 10.2 cps) but somewhat higher at D (14.4 cps). Crude protein (6.25 N) ranged from 116 to 147 g/kg and tended to be higher at D. Starch, which ranged from 612 to 656 g/kg, was least at D (617 g/kg) and greatest at L (641 g/kg). 4. Crude protein, crude fibre and total non-starch polysaccharide (NSP) were negatively correlated with SW, the R2 being respectively 0.38 (P

1. A 3x3x2 factorial experiment studied the interactions of fat source (tallow, soya, tallow:soya [2:1] blend), wheat level (700, 350, 0 g/kg) and enzyme inclusion (Avizyme 1300, absent, present) in diets for broilers fed ad libitum in individual cages from 7 to 35d. Bird performance, fat digestibility, viscosity of ileal contents and diet metabolisability (AME) were measured. 2. There were no significant effects of fat source on bird performance. However, there was a significant effect on fat digestibility, which was highest for soya and lowest for tallow. Diet AME content was also significantly affected by fat source and reflected differences in fat digestibility. 3. Dry matter (DM) intake, liveweight gain (LWG) and gain:food were all reduced at 700 g wheat/kg. Viscosity of ileal contents increased with increasing wheat inclusion. 4. There were no significant effects of enzyme on DM intake or LWG but gain:food was improved by 2%. Diet AME content was increased with enzyme addition, the effect being greatest (9%) with tallow at 700 g wheat/kg. 5. Viscosity of ileal contents was reduced and fat digestibility increased with enzyme addition and there were significant wheat enzyme interactions attributable to no differences with zero wheat but marked responses to enzyme at 700 g wheat/kg. 6. The results confirm important interactions between wheat content and fat composition in relation to fat digestibility, AME content and food efficiency.

1. The effect of diet form (mash, cold-pelleted, steam-conditioned/pelleted, wet mash, whole wheat with balancer pellet, restricted pellet) and enzyme inclusion (Avizyme 1300, absent, present) was studied in 2 trials using individually caged, male broilers from 14 to 42 d. Bird performance, viscosity of ileal contents and diet metabolisability (AME) were measured. 2. The performance of mash-fed birds was significantly poorer than for the other treatments in relation to dry matter intake, liveweight gain and gain:food. This was not due to reduced diet AME content. 3. There was no significant effect of heat treatment on any of the variables measured, although viscosity of ileal contents was increased by 30% as compared to the cold-pelleted diet. 4. Gain:food was improved with wet-mash feeding in comparison to the dry mash treatment but it was concluded that this was not due to any intrinsic improvement in diet quality, but rather to voluntary food restriction on introduction of the wet food. 5. Whole wheat feeding improved gain:food and diet AME content by 3% as compared to the complete diets and caused approximately a 50% increase in gizzard weight as compared with the pelleted diets. 6. Food enzyme inclusion did not improve performance although a significant improvement in diet AME content was observed with enzyme inclusion in trial 1.

Herpes simplex virus type 1 (HSV-1) mutants defective in immediate-early (IE) gene expression do not readily enter productive replication after infection of tissue culture cells. Instead, their genomes are retained in a quiescent, nonreplicating state in which the production of viral gene products cannot be detected. To investigate the block to virus replication, we used the HSV-1 triple mutant in1820K, which, under appropriate conditions, is effectively devoid of the transactivators VP16 (a virion protein), ICP0, and ICP4 (both IE proteins). Promoters for the HSV-1 IE ICP0 gene or the human cytomegalovirus (HCMV) major IE gene, cloned upstream of the Escherichia coli lacZ coding sequences, were introduced into the in1820K genome. The regulation of these promoters and of the endogenous HSV-1 IE promoters was investigated upon conversion of the virus to a quiescent state. Within 24 h of infection, the ICP0 promoter became much less sensitive to transactivation by VP16 whereas the same element, when used to transform Vero cells, retained its responsiveness. The HCMV IE promoter, which is not activated by VP16, also became less sensitive to the HCMV functional homolog of VP16. Both elements remained available for transactivation by HSV-1 IE proteins at 24 h postinfection, showing that the in1820K genome was not irreversibly inactivated. The promoters controlling the HSV-1 ICP4, ICP22, and ICP27 genes also became essentially unresponsive to transactivation by VP16. The ICP0 promoter was induced when hexamethylene bisacetamide was added to cultures at the time of infection, but the response to this agent was also lost by 24 h after infection. Therefore, promoter elements within the HSV-1 genome are actively repressed in the absence of IE gene expression, and repression is not restricted specifically to HSV-1 IE promoters.

Optical emission spectroscopy provides a rapid and precise method for determining 15N/14N ratios of 15N‐enriched plant and/or soil samples. The objective of this study was to determine the effect of an indicator added in the distillation solution on the success rate of tube lighting in the optical emission spectrometer over a large range of N concentration in 15N enriched plant samples. One‐hundred‐eighty plant samples with large ranges of N concentration (4 to 30 g.kg‐1 dry weight) and 15N atom enrichment (0.368 to 1.635%) were analyzed. Our data suggest that there was no difference in the success rate of tube lighting in emission spectroscopy and in 15N/14N ratio measured between samples prepared with and without addition of indicator in the distillation solution.

The thick folic (mainly upland forest) materials (> 40 cm of accumulated organic material) that occur in the Coastal Western Hemlock Biogeoclimatic Zone in British Columbia have not been described with regard to the spatial interrelationships of the soil constituents in context with the chemical composition of the different horizons. Micromorphological assessment and solid-state 13C NMR were used to characterize the accumulated folic materials from a Lignic Folisol (northern Vancouver Island), Histic Folisol (Prince Rupert, BC) and a Humic Folisol (Queen Charlotte Islands, BC). Micromorphology provided information on the spatial relationships of the soil constituents and 13C NMR provided data on the chemical components of the folic materials. Soil faunal activity, primarily from mites, was the dominant soil-forming process observed in the organic horizons of the Folisols, being especially prominent in the Lignic Folisol with the breakdown of woody materials. Solid state 13C CPMAS NMR spectra facilitated distinguishing three main types of horizons: (1) Horizons derived from accumulated residues (L, Fr, and Hr) showing higher carbohydrate-like C and O-alkyl C values and lower total aromatics; (2) Horizons with advanced decomposition (Hr2, Oh1, and Hd) which were higher in alkyl C; and (3) Horizons derived from ligneous material (Fw and Hdw) where carbohydrate-like C was less than total aromatic C. Implications for adequate nutrient content and forest growth were inferred from the observed micromorphology and chemical composition of the folic materials. Key words: Folisol, forest soils, micromorphology, 13C NMR, faunal activity

The assaying of nitrogen isotope abundance using emission spectroscopy commonly requires a plasma discharge in a sample of N2 gas at low pressure (∼0.5 kPa) in a sealed glass tube. Expensive high‐power (≳20 W) radio‐frequency or microwave amplifiers are often used for this purpose. However, with correct design, a simple and inexpensive, single transistor, 100 MHz, 1 W oscillator can be used to produce a bright plasma discharge in a low‐pressure sample of nitrogen in a 6 mm diameter by 10 cm tube. The device utilizes optimal impedance matching to minimize the power requirements. The principles and limitations of the matching are outlined, and a simple feedback circuit is also described which, with the aid of a phototransistor, enables the emission intensity to be controlled. Ignition of discharge by friction or with the help of a simple piezoelectric gas lighter is also described, and construction details are given.

International audience; The carbon (C) cycle in boreal regions is strongly influenced by fire, which converts biomass and detrital C mainly to gaseous forms (CO2 and smaller proportions of CO and CH4), and some 1?7% of mass to pyrogenic C (PyC). PyC is mainly produced as solid charred residues, including visually-defined charcoal, and a black carbon (BC) fraction chemically defined by its resistance to laboratory oxidation, plus much lower proportions of volatile soot and polycyclic aromatic hydrocarbons (PAHs). All PyC is characterized by fused aromatic rings, but varying in cluster sizes, and presence of other elements (N, O) and functional groups. There are several reasons for current interest in defining more precisely the role of PyC in the C cycle of boreal regions. First, PyC is resistant to decomposition, and therefore contributes to very stable C pools in soils and sediments. Second, it influences soil processes, mainly through its sorption properties and cation exchange capacity, and third, soot aerosols absorb solar radiation and may contribute to global warming. However, there are large gaps in the basic information needed to address these topics. While charcoal is commonly defined by visual criteria, analytical methods for BC are mainly based on various measures of oxidation resistance, or on yield of benzenepolycarboxylic acids. These methods are still being developed, and capture different fractions of the PyC "continuum". There are few quantitative reports of PyC production and stocks in boreal forests (essentially none for boreal peatlands), and results are difficult to compare due to varying experimental goals and methods, as well as inconsistent terminology. There are almost no direct field measurements of BC aerosol production from boreal wildfires, and little direct information on rates and mechanisms for PyC loss. Structural characterization of charred biomass and forest floor from wildfires generally indicates a low level of thermal alteration, with the bulk of the material having H/C ratios still >0.2, and small aromatic cluster sizes. For the more chemically-recalcitrant BC fraction, a variety of mainly circumstantial evidence suggests very slow decomposition, with turnover on a millennium timescale (5000?10 000 y), depending on environmental conditions and PyC properties, but the main limitation to PyC storage in soil is likely consumption by subsequent fires. Degraded, functionalized PyC is also incorporated into humified soil organic matter, and is transported to sediments in dissolved and particulate form. Boreal production is estimated as 7?17 Tg BC y?1 of solid residues and 2?2.5 Tg BC y?1 as aerosols. Primary research needs include basic field data on PyC production and stocks in boreal forests and peatlands, suitable to support C budget modeling, and development of standardized analytical methods and of improved approaches to assess the chemical recalcitrance of typical chars from boreal wildfires. To accomplish these goals effectively will require much greater emphasis on interdisciplinary cooperation.

Tannins influence ecosystem function by affecting decomposition rates, nutrient cycling, and herbivory. To determine the role of tannins in ecological processes, it is important to quantify their abundance and understand how structural properties affect reactivity. In this study, purified tannins from the foliage of nine species growing in the pygmy forest of the northern California coast were examined for chemical reactivity, protein precipitation capacity (PPC), and structural characteristics (13C NMR). Reactivity of purified tannins varied among species 1.5-fold for the Folin total phenol assay, and 7-fold and 3-fold, respectively, for the acid butanol and vanillin condensed tannin assays. There was about a 5-fold difference in PPC. Variation in chemical reactivity and PPC can be largely explained by differences in structural characteristics of the tannins determined by 13C NMR. In particular, the condensed versus hydrolyzable tannin content, as well as the hydroxylation pattern of the B-ring and stereochemistry at the C-2-C-3 position appear to influence reactivity. Due to the large differences in chemical reactivity among species, it is necessary to use a well-characterized purified tannin from the species of interest to convert assay values to concentrations. Our results suggest that structural characteristics of tannins play an important role in regulating their reactivity in ecological processes.

(1999). Effects of diet formulation and enzyme inclusion on apparent metabolisable energy (AME) concentration in wheat-based diets and on broiler performance. British Poultry Science: Vol. 40, No. sup001, pp. 37-38.

We constructed three recombinant vectors derived from the herpes simplex virus type 1 mutant ts K., each of which contained a different transgene under the control of the herpes simplex virus type 1 immediate early 3 promoter inserted into the thymidine kinase locus: the prokaryotic enzymes β-galactosidase and chloramphenicol acetyl transferase, and a fusion gene consisting of human tissue inhibitor of metalloproteinases linked to the last exon of Thy-1, which encodes for a glycosyl-phosphatidyl-inositol membrane anchor. Infection of postmitotic neocortical and hippocampal neurons in low-density primary cultures with these vectors, achieved reliable expression of all three foreign gene products in various neocortical cell types, e.g. pyramidal neurons, non-pyramidal neurons, and glial cells. The percentage of neurons expressing transgenes ranged from 1 to 46% depending on the multiplicity of infection ( highest assayed = 5 ); the percentage of glial cells expressing transgenes ranged from 0.5 to 98% ( highest multiplicity assayed = 3.4 ). Expression of transgenes could be detected for up to three days in approximately 20% of neurons infected at a multiplicity of infection of 1. Infection of neurons with tk K-derived recombinant vectors inhibited their protein synthesis by 40–50% at a multiplicity of infection of 10, but no effect was observed at a multiplicity of infection of 1. Infection of glial cells with the same vectors at a multiplicity of infection of 1 inhibited protein synthesis by more than 90%. Analysis of neuronal viability at different times post-infection indicated that more than 98% of neurons expressing transgenes 48 h post-infection were viable. Thus, low-density neuronal cultures can be used to assess the efficiency of herpes simplex virus type 1-derived gene transfer vectors and transgene expression in developing cortical postmitotic cells, before and after they establish polarity. In addition, we show that two cytoplasmic enzymes, β-galactosidase and chloramphenicol acetyl transferase, are able to diffuse freely in the cytoplasm reaching even growth cones in young neurons, while the chimeric protein tissue inhibitor of metalloproteinases/Thy-1 is correctly targeted to the plasma membrane via a glycosyl-phos-phatidylinositol anchor. This model system should be useful for investigation of cellular and molecular aspects of the development and establishment of neuronal polarity, as well as for analysis of signals involved in protein targeting in postmitotic neurons.

We have developed tissue culture systems which mimic certain aspects of herpes simplex virus (HSV) latency, in order to facilitate molecular studies of the phenomenon. The initial approach utilised incubation at 42 degrees C to suppress virus replication and establish stable latency after infection of human fibroblasts at low multiplicity with HSV-2. Latent virus was reactivated efficiently by superinfection of cultures with HSV-1 or human cytomegalovirus. Improvement of the system was achieved by use of the HSV-1 mutant in1814, which has a 12 base pair insertion in the coding sequences for the transactivating protein Vmw65 (VP16). Upon infection with in1814 at 42 degrees, or 37 degrees, it was possible to obtain cultures in which up to 21% of cells contained a viral genome. More recent modifications include the use of mutant in1820 (a derivative of in 1814) and pretreatment of cells with interferon alpha, which enable latency to be established in approximately 100% of cells. A system has therefore been developed that is suitable for the analysis of gene expression and genome structure during latency and reactivation.

Oogenesis within the hologonic ovary of the trichuroid nematode, Trichuris muris, was observed by light and electron microscopy. Early germinal stages in the form of oogonia and young primary oocytes were characterised by a high nuclear-cytoplasmic ratio, numerous ribosomes and several mitochondrial clusters. Previtellogenic primary oocytes contained a prominent nucleus with a nuclear envelope punctuated by pores. They also contained increased amounts of granular endoplasmic reticulum (GER), often arranged as annulate lamellae, several Golgi complexes and limited amounts of lipid. The appearance of three types of cytoplasmic inclusion, in the form of lipid, dense yolk granules and reticulate granules, indicated the onset of vitellogenesis. At this stage of oogenesis, all three types were distributed throughout the ooplasm. The possible role of the granules is discussed. During passage along the oviduct the oocyte was coated by an additional unit membrane and associated fibrillar layer external to the oole...

SummaryThe development of the egg-shell of the trichuroid nematode, Trichuris muris, was observed by light and electron microscopy. Initial stages of shell formation occurred immediately following sperm entry into the oocyte in the spermatheca of the female worm. The external vitelline layer separated from the oolemma, reticulate oocyte granules were discharged into the resultant perivitelline space and stored glycogen contributed to the formation of the chitinous shell. A lipid layer developed between the inner surface of the chitinous layer and the oolemma. The fully formed egg-shell thus comprised an external vitelline layer, a middle chitinous layer and an inner lipid layer. The outer vitelline layer resembled a unit membrane with strands of particulate material attached to its outer surface. The middle chitinous layer was composed of chitin microfibrils encased in a protein coat; the sheets of microfibrils were arranged as parallel lamellae forming a helicoidal architecture. The inner lipid layer contained a variable number of loosely arranged strands of electron-opaque and less-opaque material.

The RNA species synthesized in vitro by a transcribing nucleoprotein (TNP) complex of vesicular stomatitis virus (VSV) were translated with high efficiency in a fractionated cell-free system derived from reticulocytes. The use of TNP complexes isolated from VSV Indiana, VSV New Jersey, and Chandipura viruses showed that in each case the predominant polypeptides synthesized had electrophoretic mobilities identical to their virion N, NS, and M polypeptides in proportions reflecting those found in infected cells rather than purified virions. A minor polypeptide corresponding to unglycosylated polypeptide G was also observed, but the in vitro synthesis of polypeptide L was not detected. The addition of RNase inhibitor to transcription mixtures markedly increased the rate of RNA synthesis. Furthermore, the messenger activity of the RNA was significantly enhanced. The inclusion of S-adenosyl L-methionine during transcription substantially increased the messenger activity of the product RNA, suggesting a requirement for methylation. Fractionation by oligodeoxythymidylic acid-cellulose chromatography revealed that the RNA required a polyadnylic acid tract for messenger activity.

Two humic and one fulvic acid were hydrolyzed with hot 6M HCl and a combination of 12M and 0.5M H2SO4. Effects of acid hydrolysis on the chemical structures of the humic materials were assessed by comparing liquid-state13C NMR spectra of hydrolyzed with those of unhydrolyzed humic substances. Hydrolysis with 6M HCl was found to be more efficient for removing ptoteinaceous materials and carbohydrates than was hydrolysis with 12M-0.5M H2SO4. The latter appeared to better preserve the structural integrity of the humic materials, especially in the lower aliphatic region (0–50 ppm). Hydrolysis with the two acids increased the aromaticity of the humic materials, in part by removing proteinaceous components and carbohydrates. It also lowered the CO2H content of two out of three humic preparations, probably because of acid decarboxylation. The data presented demonstrate the great potential of13C NMR spectroscopy for structural research on humic materials.

mRNA's homologous to the herpes simplex virus type 1 DNA restriction endonuclease fragment BamHI p, which contains the thymidine kinase gene, have been identified and mapped by hybrid-arrested translation and mRNA selection. Such mRNA's, when translated in vitro, directed the synthesis of polypeptides of apparent molecular weights 43,000 (VI43) and 39,000 (VI39). mRNA for enzymatically active thymidine kinase was enriched by more than 20-fold after selection. Mapping was carried out with restriction endonuclease fragments of BamHI p, and locations of the 5' and 3' termini of VI43 mRNA were deduced. Analysis of nucleotide sequences around the 5' terminus revealed several consensus sequences commonly found at the start of eucaryotic mRNA's and which are presumably involved in initiation of transcription by RNA polymerase II. Translation of mRNA's for VI43, VI39, and the thymidine kinase enzyme was arrested only by a 1,170-base-pair region of BamHI p. Since this region is insufficient for adjacent genes, coding sequences for VI43 and VI39 must overlap; the possible relationship of these two polypeptides is discussed. A virus-induced product equivalent to VI39 was detected in infected cells.

Eighteen humic acids (HA's) were characterized by chemical analyses, infrared (IR) and electron spin resonance spectra, and by solution and solid-state cross polarization magic-angle spinning (CPMAS) C nuclear magnetic resonance (NMR). The main objectives of this investigation were: (i) to compare the solution and solid-state C NMR spectra of the HA's; (ii) to examine effects of different delay times (D1) on the solution spectra; and (iii) to find reasons for the failure of C NMR to detect the presence of significant concentrations of phenolic C in unmethylated HA's. Compared to solution spectra, CPMAS spectra consisted of relatively few, broad bands, indicative of considerable overlapping of individual signals, especially in the aliphatic regions. Proportions of aliphatic C determined by solution C NMR at D1 = 0.74 s were greater than those obtained at D1 = 1.90 s and by CPMAS C NMR (where D1 = relaxation delay). By contrast, proportions of aromatic C and aromaticities were greatest at D1 = 1.90 s. The highest values for CO2H groups were obtained by CPMAS C NMR and approached total acidities; values measured by solution C NMR at D1 = 1.90 s were intermediate, while those determined by solution C NMR with D1 = 0.74 s were the lowest and closest to CO2H concentrations determined chemically. Because the presence in HA's of significant concentrations of phenolic OH groups has been confirmed by several independent methods, it is possible that in C NMR spectra of HA's, signals arising from phenolic C are shifted downfield so that they overlap with resonances of C in CO2H groups. The net result is that CO2H groups are overestimated and phenolic OH groups underestimated. Additional Index Words: phenolic hydroxyl groups, carboxyl groups, aliphatic C, aromatic C, aromaticity. View complete article To view this complete article, insert Disc 4 then click button8

The cuticular permeability of anhydrobiotic 4th-stage juveniles of Ditylenchus dipsaci decreases during rehydration, as indicated by staining with osmium tetroxide, the measurement of permeability coefficients by the rate of uptake of tritiated water from a HOH/C-inulin bathing solution and by an increased ability to slow down the rate of water loss and to survive subsequent desiccation. The initial decrease in permeability coefficient is rapid, suggesting a physical effect due to the rehydration of the cuticle, followed by a slower reduction over the subsequent 1–4 h which is dependent upon metabolic activity. The reestablishment of the permeability barrier is sensitive to metabolic inhibitors which affect enzyme activity and post-transcriptional protein synthesis. Sodium iodoacetamide was the only inhibitor which affected maintenance of the permeability barrier. An Arrhenius plot of changes in cuticular permeability with temperature indicated a sharp increase in permeability at 40–50°C and brief exposure to diethyl ether resulted in loss of the permeability barrier, suggesting that a superficial layer, probably the epicuticle, is responsible for controlling cuticular permeability.

To assess effects of soil Cu on the yield and nutrition of a crop, onions (Allium cepa ’Autumn Spice’) were grown in field microplots at sites A (peat), B (muck) and C (mucky peat). The surface layers (0–20 cm) of the plots contained four levels of residual fertilizer Cu up to 1200 ppm (wt/wt) at sites A and B and up to 600 ppm at site C. The highest Cu treatment at sites A and B contained about 4–6 times the Cu required for mitigating the excessive decomposition and subsidence of such organic soils. Neither the Cu treatment level nor the total soil Cu concentration influenced crop yields at sites B or C. Crop yield responded positively to the two highest Cu treatment levels at site A where the background level of soil Cu was the lowest among the three sites. The Cu concentrations in the crop at all sites were below the level considered to be phytotoxic (20 ppm). Fe and Zn contents in plants were also not depressed by higher Cu concentrations except for foliar Fe on the highest Cu treatment plots at site A. These plots were intrinsically poorer in Fe than those treated with lesser Cu. Foliar Fe:Cu and Zn:Cu ratios were also lowest, but not below adverse levels, for the highest Cu treatment levels at sites A and B, respectively. Even the highest levels of soil Cu did not reduce N supply for, or N nutrition of, the onion crop, or alter the crop concentrations or uptakes of P, K, Ca, Mg and Mn. Key words: Copper, onions, subsidence, histosol, muck, peat

As a part of attempts at examining the suitability of using moderate amounts of Cu for mitigating the undesirably fast decomposition and subsidence of some organic soils, this investigation revealed that both total and DTPA-TEA-extractable Cu contents of 14 samples of cultivated organic soils correlated negatively with the total microbial population counted on soil extract-acitidione-gelatin agar plates and positively with the percentage of that population capable of extracellular proteolysis. On the other hand, the concentrations of both total and extractable Cu did not correlate with either the total number of proteolytic organisms counted, or the capacities of the soils for ammonification and nitrification. The results are, therefore, in accord with the belief that Cu curtails the decomposition of organic soils primarily through inactivation of accumulated soil enzymes, but not through direct toxic effects on the general soil microbial population.

Microinjection of herpes simplex virus (HSV)-infected cell mRNA into Xenopus laevis oocytes resulted in the production of a new exonuclease activity. This enzyme strongly resembled the HSV alkaline exonuclease in many biochemical properties, and hybrid-arrested translation studies showed that it was virus coded, mapping at 0.080 to 0.185 genome map units. Exonuclease mRNA had a size and genome location equivalent to the mRNA encoding V185 in reticulocyte lysates, suggesting that V185 is the exonuclease. The enzyme synthesized in oocytes was found to act as an exonuclease in vivo. Two plasmids containing HSV DNA fragments directed the synthesis of exonuclease when microinjected into oocyte nuclei, and this finding enabled the coding and control sequences for this gene to be localized to 0.155 to 0.185 genome map units.

The BglII N fragment of herpes simplex virus type 2 (HSV-2) DNA (approximately 0.58 to 0.63 map unit) was examined for encoded products. Using plasmid pGZ59, which consists of BglII-N cloned in pAT153, in conjunction with hybrid arrested translation, mRNA selection, and in vitro protein synthesis, we found that the major translated product of this region has an approximate molecular weight of 37,800. By further mapping, coding sequences for this polypeptide were located within the region of BglII-N representing approximately 0.58 to 0.61 genome map unit. To demonstrate immunological specificity, we used staphylococcal A protein immunoprecipitation with rabbit anti-HSV-1 or HSV-2 sera and antigens from HSV-1 or HSV-2 total mRNA translated in vitro and BglII-N-selected mRNA. The results show that the 37,800-dalton polypeptide has HSV-2 immunological specificity, as it is precipitated with anti-HSV-2 sera but not with anti-HSV-1 or control sera.

Copper concentrations should be maintained at 100 ppm (gg Cu g−1 dry soil) in the surface layers of undecomposed peat soils, and 400 ppm in humified muck soils. The Cu supports plant nutrition, and inhibits enzymes that degrade the organic soils (Histosols). The required or extravagant applications of Cu may threaten groundwater quality if the Cu, or elements displaced by the Cu, moves downwards in the soils. To test this, powdered CUSO4.5H2O was applied to the top 15 cm of replicated microplots of organic soils to increase their Cu concentrations by 0, 150, 500, and 1500 ppm at field sites A (peat) and B (muck) in May 1978 and by 0, 100, 300, and 800 ppm at field site C (mucky peat) in 1979. Duplicate cores of up to 50 cm depth were taken in the spring of 1981 from each of the 56 microplots and analyzed. At all sites, none of the Cu additions caused significant displacement and downward movement of Ca, Mg, K, Fe, Mn, or Zn. There was a leaching of small fractions of the applied Cu down to 40 cm depth only when 1500 ppm of Cu was added to the humus-poor, acidic peat at site A. Some of the Cu applied at the 500 ppm rate at site A and 1500 ppm rate at site B was found in the 20 to 30 cm zone. At site C, none of the applied Cu moved from the top 20 cm (plow layer).

Summary An investigation of the activity of nuclear RNA polymerase following infection of LS cells with HSV-1 shows a decline in both major activities. This effect is not entirely due to inhibition of cellular protein synthesis, and the effect of α-amanitin-sensitive RNA polymerase is mediated by a protein(s) synthesized in the infected cell. Changes in the properties of this RNA polymerase activity include a reduction in the relative UTP/GTP incorporation ratio and an increased sensitivity to inhibition by actinomycin D, indicating that RNA polymerase II is involved in virus transcription.

Previous studies (R. J. Watson and J. B. Clements, Virology 91:364--379, 1978; C. M. Preston, J. Virol. 29:275--284, 1979) have shown that the herpes simplex virus type 1 (HSV-1) mutant tsK has a temperature-sensitive lesion in an immediate early polypeptide whose function is to induce synthesis of new viral transcripts, including mRNA, for pyrimidine deoxyribonucleoside kinase. The studies presented here examine the properties of immediate early polypeptides in wild-type HSV-1 and tsK-infected cells at 31 and 38.5 degrees C. The overall pattern of immediate early protein synthesis was similar in wild-type HSV-1- and tsK-infected cells when radiolabeled with [35S]methionine or 14C-amino acid mixture. Further investigation, however, revealed two aberrant properties of the polypeptide Vmw 175 in tsK-infected cells at 38.5 degrees C. Upon cell fractionation, large amounts of this polypeptide were recovered in the cytoplasmic fraction, in contrast to tsK-infected cells at 31 degrees C or wild-type HSV-1-infected cells at either temperature. Furthermore, at 38.5 degrees C tsK-induced Vmw 175 was not processed normally to forms of lower electrophoretic mobility. Both of these defects were reversible upon downshift of tsK-infected cells, even in the absence of further protein synthesis, but were not observed in cells infected with a revertant of tsK. Coinfection of tsK-infected cells with wild-type HSV-1 did not alleviate these lesions, suggesting that they resulted from an abnormal Vmw 175 polypeptide rather than from a defective processing enzyme. Temperature upshift of tsK-infected cells caused reversion of Vmw 175 to the mutant form. The progression to synthesis of late polypeptides was also arrested; therefore, a functional lesion was also reversible upon temperature changes between 31 and 38.5 degrees C during the early stages of infection. The identification of a polypeptide with abnormal properties in tsK-infected cells and the demonstration that these properties, and the functional lesion, are reversible may provide an important system for investigation of HSV-1 transcriptional control.

The incubation of a cell-free protein-synthesizing system prepared from rabbit reticulocytes with cytoplasmic RNA from herpes simplex virus (HSV)-infected cells resulted in increased thymidine kinase activity. This enzyme activity was specifically inhibited by anti-HSV antiserum and was relatively unaffected by TTP, an inhibitor of cellular thymidine kinases. Induction of the new activity was prevented by addition of inhibitors of eucaryotic protein synthesis, and no new activity was detected when RNA from cells infected with pyrimidine deoxyribonucleoside kinase-deficient mutants, instead of wild-type HSV, was added. An increased deoxycytidine kinase activity with similar properties to the HSV-specified enzyme activity was also present in cell-free systems incubated with RNA from HSV-infected cells. Phosphorylation of thymidine and deoxycytidine at 30 degrees C continued for longer than 11 h. The findings are consistent with the accurate synthesis in vitro of enzymically active HSV-specified pyrimidine deoxyribonucleoside kinase.

A cell-free system active in translation of HSV mRNA was obtained by fractionation of reticulocyte lysates. Endogenous protein synthesis was further reduced by preincubation with micrococcal nuclease, although this treatment did not affect the translation of infected (or uninfected) BHK cell RNA. Polypeptides synthesized by the fractionated reticulocyte cell-free system were very similar to those produced by unfractionated reticulocyte lysates.

Polyadenylated immediate-early transcripts of herpes simplex virus type 1, made in BHK cells infected and maintained in the presence of cycloheximide, have been separated on denaturing agarose gels containing methyl mercuric hydroxide. Three virus-specific mRNA bands of estimated sizes 4.7, 3.0, and 2.0 kilobases (kb) were detected, and these mRNA's were mapped on the virus genome and also used to direct protein synthesis in vitro. The 4.7- and 3.0-kb mRNA's hybridized predominantly to certain DNA fragments which are located in the short and long repetitive regions of the genome, respectively, whereas the 2.0-kb mRNA's mapped to three discrete regions of the virus DNA. In vitro translation of these separated mRNA size classes indicated that the 3.0-kb mRNA specified the synthesis of virus polypeptide Vmw 110, whereas the 2.0-kb mRNA's specified Vmw 68, 63, and 12. The synthesis of small amounts of Vmw 175 was specified by the 4.7-kb mRNA. In contrast with the mRNA's which specify these other immediate-early polypeptides, that specifying Vmw 12 is much larger than required for its coding sequences.

This paper deals with control of mRNA levels, assayed by in vitro translation, in cells infected with herpes simplex virus type 1 (HSV-1). A particularly useful marker has been pyrimidine deoxyribonucleoside kinase (dPyK) mRNA, for which the enzymatically active product can be assayed quantitatively. Cells infected with the HSV-1 temperature-sensitive mutant tsK at the nonpermissive temperature (38.5 degrees C) or with wild-type HSV-1 in the continuous presence of cycloheximide contained no detectable dPyK mRNA. Upon temperature shift-down of tsK-infected cells to 31 degrees C, dPyK mRNA was produced, and this event was inhibited by actinomycin D but not cycloheximide. This result demonstrated that the defective polypeptide in tsK-infected cells was involved in transcription of the dPyK gene and could regain activity at 31 degrees C. Because tsK-infected cells synthesized mainly immediate early polypeptides at 38.5 degrees C, the involvement of this polypeptide class in synthesis of dPyK mRNA was investigated. Analysis of the kinetics of inductions of dPyK mRNA indicated that the temperature-sensitive lesion in tsK lies in an immediate early polypeptide which is directly responsible for activation of the dPyK gene at the transcriptional level.

The structure and development of the polar plugs of Trichuris muris eggs were observed by light and electron microscopy. Initial stages in plug formation commenced in the spermatheca of the adult female where two polar papillae became delimited from the remainder of the oocyte cytoplasm. These papillae exhibited a discrete PAS positive reaction and were regarded as prospective plug regions from which cytoplasmic granules were absent. Glycogen rosettes, initially concentrated in these prospective areas, were later transformed into an irregular fine network of chitin-protein microfibrils. This arrangement of microfibrils was in contrast to that of the surrounding collar region which displayed a distinct lamellate organization. The fully developed polar plug was of lower electron density than the shell. Each plug was covered externally by the vitelline layer and lined internally by the lipid layer of the eggshell.

We carried out a 13C NMR solution study on a humic and a fulvic acid from a mineral soil. T1 values were 0.2 to 2.O s, with maximum values for carboxyl and methoxyl carbons, and nOes were up to 1 (theoretical maximu 2). Qualitative spectra were effectively obtained with short acquisition tim