Harlow H. Hall

Organic acids in the haemolymph of healthy and diseased Popillia japonica (Newman) larvae

Abstract Organic acid constituents in the haemolymph of healthy and diseased Popillia japonica (Newman) larvae were determined by partition chromatography, enzymatic analyses, and colorimetric procedures. Haemolymph from larvae infected with Bacillus popilliae Dutky, compared with that from healthy larvae, contained higher concentrations of malic, glycolic, tartaric pyruvic, and glyoxylic acids. No change in concentrations of butyric, propionic, acetic, formic, succinic, lactic, citric, and α-ketoglutaric acids occurred when larvae were infected. None of the acids decreased during the course of the milky disease. Gluconic and oxaloacetic acids were detected in the haemolymph but not determined. Organic acids account for most of the anion titre in the haemolymph.

Amino acids in the haemolymph of Popillia japonica (Newman) larvae

Abstract The proteins were separated from the acid-soluble components in haemolymph of Popillia japonica (Newman) larvae with 5% trichloroacetic acid, and the resulting materials analysed for amino acid content. Altogether twenty amino acids were identified in whole or hydrolysed haemolymph or in fractions. Analysis was done by paper chromatography and ion exchange chromatography. Glutamine, arginine, histidine, proline, glycine, alanine, and valine occur free in considerable amounts in the haemolymph. Lysine, aspartic acid, threonine, serine, glutamic acid, isoleucine, leucine, tyrosine, β-alanine, tryptophan, and asparagine were also detected. Sixteen amino acids were identified in the insoluble proteinaceous material. Phenylalanine and methionine were present in the insoluble fraction, but not to an appreciable extent in any other fractions. There were several peptides that disappeared on acid hydrolysis. There were at least two unidentified ninhydrinreacting substances that were stable to acid.

Hansenula Angusta, an Excellent Species for Demonstration of the Coexistence of Haploid and Diploid Cells in a Homothallic Yeast

Wickerham stated (1951) that the ratio of diploid to haploid cells is the dominant characteristic determining the evolutionary level attained by a species of Hansenula. This relationship holds whether the species is homothallic or heterothallic, and whether the yeast belongs to the phylogenetic lines which developed toward the free-living state or toward more dependence upon trees and insects. Other characteristics, such as increasing size of cells and transition from mucoid, to butyrous, to mat colonies, also reveal evolutionary succession in the various lines. Physiological characteristics, particularly the number of vitamins required from the environment, the number of sugars fermented, and the rate of fermentation, also indicate evolutionary position. The physiological reactions, however, became stronger in lines developing toward the freeliving state, and weaker in the lines developing toward greater dependence upon insects and trees. The genus Hansenula, as we now know it, apparently originated as a conifer-inhabiting species. The yeasts in the lines pointing toward independence from trees evolved successively as species dependent upon coniferous trees, then as species dependent upon deciduous trees, and finally as species living independently in soil and water. The completely dependent lines of Hansenula became limited in habitat to the coniferous

Hemolymph proteins of healthy and diseased larvae of the Japanese beetle, Popillia japonica

Abstract Proteins in the hemolymph of Popillia japonica larvae were separated and characterized by molecular-sieve chromatography, starch-gel and cellulose polyacetate zone electrophoresis, and ultracentrifugation. Hemolymph samples from normal larvae and from larvae infected with Bacillus popilliae , the milky disease organism, were compared to determine the effect of the disease on the proteins. During the course of milky disease, one protein disappeared and a smaller particle increased markedly in concentration. The protein present in highest concentration (major) exhibited the highest electrophoretic mobility and had an apparent isoelectric point of 5.85. This protein contained lipid as well as carbohydrate moieties. Hemolymph from diseased larvae showed a decrease in concentration of the major protein.

Infection of Popillia japonica larvae with heat-activated spores of Bacillus popilliae☆

Abstract Maximum outgrowth of spores of Bacillus popilliae resulted after they were heated at 50°C, but outgrowth diminished as the temperature was increased to 80°C. Although outgrowth was limited and often variable, infectivity was predictably influenced by heat activation of spores. The greatest number of Popillia japonica larvae (92.6%) were infected by injection of one million spores per larva heated at 50°C. By contrast, maximum infection from the same number of unheated spores was only 38.9%. Infectivity of spores heated at 60°C and at 70°C generally was intermediate between these values; heating at 80°C afforded minimal infectivity. Apparently infectivity depends on the efficacy of spore outgrowth which, in turn, is influenced by appropriate heat treatment.

Amino acids in the haemolymph of diseased Popillia japonica (Newman) larvae.

Abstract Free amino acid constituents in the haemolymph from diseased Popillia japonica (Newman) larvae were determined by ion-exchange chromatography, paper chromatography, enzymatic analysis, and amperometric titration. Haemolymph from larvae infected with Bacillus popilliae, compared with that from healthy larvae, contained higher concentrations of glutamic acid, β-alanine, aspartic acid, phenylalanine, threonine, serine, and lysine and lower concentrations of glycine, tyrosine, and histidine. Differences in the amino acid composition depend on the infective organism. The only amino acid to increase during infection with B. lentimorbus was glutamic acid. Amounts of histidine, proline, glycine, alanine, valine, isoleucine, tyrosine, and arginine decreased. The level of protein material in the haemolymph as well as the composition did not change markedly during infection with either B. popilliae or B. lentimorbus. Haemolymph contains little peptide of low molecular weight. Several additional amino acids were determined in haemolymph from healthy and diseased larvae. Milky disease apparently has no effect on the concentration of tryptophan, cystine, cysteine, glutamine, asparagine, and lanthionine.

Oxygen concentration in larval hemolymph of the Japanese beetle, Popillia japonica, infected with Bacillus popilliae

Abstract A polarographic method for measuring dissolved oxygen was used to follow changes during development of “milky disease” in larvae of the Japanese beetle, Popillia japonica . The dissolved oxygen of hemolymph of apparently healthy third-instar larvae in one group analyzed was equivalent to 33.4% to 42.0% saturation of 0.25 M KCl (0.078 to 0.100 μmoles/ml of hemolymph, 95% confidence limits). The concentration of oxygen was significantly lower early in the infection when only vegetative cells of Bacillus popilliae were present in the hemolymph. When both vegetative and spore forms were present, the oxygen level approached that of the uninfected group. Infection was initiated by feeding spores at the rate of 1–10 million spores/g of soil. At this level, 20% of the larvae became infected.

Identification of the Stimulatory Factors in Citrus Molasses for Carotenogenesis in Blakeslea trispora

CITRUS molasses and other citrus by-products stimulate carotenogenesis by mated cultures of Blakeslea trispora1. However, the substances in citrus by-products that stimulate carotene production in the fermentation process remain unknown. We attempted to determine the nature of the enhancing agent present in one of these by-products, citrus molasses.