Characterization of the Subunit Structure of the Ribonucleic Acid Genome of Influenza Virus

Ribonucleic acid extracted from influenza virus was labeled at the 3' termini with (3)H and analyzed by polyacrylamide gel electrophoresis. Influenza virus was found to contain a minimum of seven and possibly as many as 10 polynucleotide chains, most of which appear to terminate at the 3' end in uridine.     

Internalization and processing of Bacillus anthracis lethal toxin by toxin-sensitive and -resistant cells

Anthrax lethal toxin consists of two separate proteins, protective antigen and lethal factor (LF). Certain macrophages and a mouse macrophage-like cell line, J774A.1, are lysed by low concentrations of lethal toxin. In contrast, another macrophage cell line, IC-21, and all other cell types tested were resistant to this toxin. To discover the basis for this difference, each step in the intoxication process was examined. No differences between sensitive and resistant cells were found in receptor binding or proteolytic activation of protective antigen, steps that are required prior to LF binding. To determine whether resistance results from a defect in translocation to the cytosol, we introduced LF into J774A.1 and IC-21 cells and a nonmacrophage cell line (L6 myoblast) by osmotic lysis of pinocytic vesicles. Only J774A.1 cells were lysed; no effect was observed in IC-21 and L6 cells. These results suggest that resistant cells either lack the intracellular target of LF or fail to process LF to an active form. The relatively low potency of LF introduced into J774A.1 cells by osmotic lysis suggests that protective antigen may also be required at a stage subsequent to endocytosis.     

The role of parasitoid and predator production in technology transfer of field crop biological control

The immediate goals for improving natural enemy production are to reduce costs, increase efficacy and provide additional species for pest management. This paper describes expanding markets for natural enemies that are or could be produced commercially, gives operational and experimental examples of parasitoid and predator production for use in field crop biological control, defines some of the obstacles and makes recommendations for producing and using natural enemies. Additionally, it provides recent published guidance for implementing biological control in Integrated Pest Management.     

Diaminopimelic Acid Synthesis in Cultures of an Escherichia coli Double Auxotroph: Effects of Cultural Conditions

The metabolic response to L-lysine of Escherichia coli ATCC 13002, a lysine-histidine double auxotroph, has been examined in a synthetic medium containing sucrose. In shaken cultures largest amounts of extracellular DAP were produced with an initial lysine concentration of 7·5 mg/1 and in static cultures of 2·5 mg/1. Considerably smaller amounts of DAP accumulated under stationary conditions. In cultures shaken for 20 and 43 h there was an overall decrease in the yields of DAP, expressed in terms of cell biomass and of sucrose consumed, as the initial concentration of lysine was increased from 0·75 mg/1 in steps up to 25 mg/1. The regulatory effect of lysine on DAP production was also observed when lysine was supplied to cultures at a constant rate employing diffusion capsules.     

Functional characterization of protease-treated Bacillus anthracis protective antigen.

Characterization of the functional domains of Bacillus anthracis protective antigen (PA, 83-kDa), the common cellular binding molecule for both anthrax edema toxin and anthrax lethal toxin, is important for understanding the mechanism of entry and action of the anthrax toxins. In this study, we generated both biologically active (facilitates killing of J774A.1 cells in combination with lethal factor, LF) and inactive preparations of PA by protease treatment. Limited proteolytic digestion of PA in vitro with trypsin generated a 20-kDa fragment and a biologically active 63-kDa fragment. In contrast, limited digestion of PA with chymotrypsin yielded a preparation containing 37- and 47-kDa fragments defective for biological activity. Treatment with both chymotrypsin and trypsin generated three major fragments, 20, "17," and 47 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This PA preparation was also biologically inactive. To investigate the nature of the defect resulting from chymotrypsin treatment, we assayed PA preparations for the ability to bind to the cellular receptor and to bind and internalize 125I-LF. All radiolabeled PA preparations bound with specificity to J774A.1 cells and exhibited affinities similar to native 83-kDa PA. Once bound to the cell surface receptor, both trypsin-treated PA and chymotrypsin/trypsin-treated PA specifically bound 125I-LF with high affinity. Finally, these PA preparations delivered 125I-LF to a Pronase-resistant cellular compartment in a time- and temperature-dependent fashion. Thus, the biological defect exhibited by chymotrypsin-treated PA is not at the level of cell binding or internalization but at a step later, such as toxin routing or processing by J774A.1 cells. These protease-treated preparations of PA should prove useful in both elucidating the intracellular processing of anthrax lethal toxin and determining the structure-function relationship of PA and LF.     

Receptor-mediated internalization and degradation of diphtheria toxin by monkey kidney cells.

The receptor-mediated internalization and degradation of radiolabeled diphtheria toxin by cultured monkey kidney cells was studied. The ability of a number of enzymes and chemicals to remove cell surface-bound toxin was tested; the combination of pronase and inositol hexaphosphate (PIHP) proved most effective. Using PIHP, the kinetics of toxin-cell association at 37 degrees C was resolved into two compounds: surface binding and internalization. The PIHP assay also allowed estimation of the half-time of toxin internalization (about 25 min). An assay involving precipitation of culture supernatants with trichloroacetic acid was developed and used to measure the rate of degradation and excretion of cell-associated toxin. Agents which markedly inhibited toxin internalization similarly prevented degradation, implying an intracellular location for the degradative process. The primary radioactive product excreted by Vero cells was monoiodotyrosine. The extent and rate of toxin degradation indicated lysosomal involvement. Finally, agents which blocked internalization or degradation, or both, (e.g. antibody and concanavalin A), protected cells from the cytotoxin action of diphtheria toxin, suggesting that these processes are necessary for expression of biological effect.     

Carbon dioxide output and mating in adult cabbage looper moths exposed to discrete light régimes

Carbon dioxide discharge and the number of spermatophores transferred were recorded from populations of adult cabbage loopers, Trichoplusia ni, maintained under various wavelengths and intensities of light. Nocturnal CO₂ output was enhanced by retaining 1 per cent of the diurnal intensity of near-u.v. and blue lights during the scotophase. However, addition of < 10 per cent of the diurnal intensities of u.v., blue, and white lights suppressed CO₂ production. Exposure to gold light resulted in the lowest 24 hr CO₂ accumulations, and red was intermediate. Similarly, maximum levels of mating occurred when cabbage loopers were exposed to nocturnal intensities of 1 per cent of the diurnal u.v., blue, or white lights, even though higher intensities were inhibitory. Considering all nocturnal intensities ranging from 0 to 100 per cent of diurnal levels, u.v. and red yielded the highest mating frequencies, blue and gold were intermediate, and white was lowest. Thus, relatively unique action spectra resulted from each régime.     

Phagocytosis of degenerating myelin in transected feline optic nerve: an immunohistochemical study

The phagocytic activity of neuroglial cells in adult feline degenerating optic nerve was investigated by immunocytochemistry at both light and electron microscopy levels. Degeneration was initiated by unilateral eye enucleation and the segment distal to the transection showing true Wallerian degeneration was examined. Following enucleation, twelve adult domestic cats were examined over a period of seven to 215 days. All cases showed slow clearance of myelin debris and absence of proliferating monocytes throughout the post-enucleation period. All phagocytic cells present were neuroglial cells, and many of these cells expressed oligodendroglial antigens. These findings demonstrate the persistence of an active population of oligodendrocytes that might play an additional functional role during Wallerian degeneration of feline optic nerve.