Quantification of hematozoa in blood smears

Ten thin blood smears from mourning doves (Zenaida macroura) infected with Haemoproteus maccallumi were examined by each of two observers using identical techniques and microscopy in an attempt to delineate the factors necessary to provide an accurate estimate of the number of parasites/n erythrocytes. The number of erythrocytes examined must be actually counted, not estimated from extrapolated partial counts or from the number of fields of view examined. Doubling the number of erythrocytes counted (1) decreased the overdispersed frequency distribution patterns in only 25% of the replicate counts for numbers of H. maccallumi/100 erythrocytes for a series of 2,000 versus 4,000 erythrocytes counted; and (2) did not significantly increase the accuracy for determining parasite intensities. Thus, the number of erythrocytes that must be counted to determine parasite intensities could be considerably reduced from the 10,000 or 20,000 estimated for most studies, and still provide an accurate determination of the number of parasites/n erythrocytes in datasets collected from hosts with moderate to high levels of parasitemia. This resulted in a decreased amount of time expended by the observer on each blood smear examined. With two equivalently trained individuals, differences between observers examining the same blood smears were minimal. This study suggests an approach by which a more standardized methodology for quantifying blood parasite intensities could be developed.     

Effect of photoperiod and temperature on the development of frost hardiness in three Alnus species

The influence of short day and low temperature on cold acclimation of A. crispa (Ait.) Pursh, A. glutinosa (L.) Gaertn. and A. rubra Bong, was investigated. Two clones of each species originating from in vitro propagation were exposed to three daylength/temperature treatments. Periodically plantlets were exposed to controlled freezing temperature in order to evaluate their level of frost hardiness.
Short day (SD) and cold temperature (CT) and long day (LD) and cold temperature (CT) were the most effective treatments for the development of frost hardiness in shoots and roots of the three species tested. Short day (SD) and warm temperature (WT) induced a significant increase in hardiness in shoots of all three species. However, this treatment did not trigger root hardening. A. crispa was found to be the hardiest species followed by A. glutinosa and A. rubra . Intraspecific variation was observed between the two A. glutinosa clones. A glutinosa clone AG8, a Russian provenance, showed a greater freezing resistance than A. glutinosa clone AG2, a German provenance.     

Distribution of activities of aspartate aminotransferase isoenzymes and malate dehydrogenase in guinea pig retinal layers

Distributions of activity of the cytosolic (cAAT) and mitochondrial (mAAT) isoenzymes of aspartate aminotransferase and of malate dehydrogenase (MDH) were determined in guinea pig retinal layers. The distribution of total AAT activity (tAAT = cAAT + mAAT) and of mAAT activity correlated well (r = 0.88-0.91) with the distribution of MDH activity. mAAT activity was highest in the inner segments of the photoreceptors; there was a greater than twelve-fold difference between activity in that layer and in the inner retinal layers. cAAT activity was also highest in the inner segments, but the difference between the activity in the inner segments and the other layers was not nearly as great as with mAAT. cAAT activity was also relatively high in the outer nuclear layer, outer plexiform layer, and part of the inner plexiform layer. The high activity of cAAT, mAAT, and MDH in the inner segments indicates that all of these enzymes are involved in metabolic reactions related to energy production and/or to photoreceptive processes in the outer segments and, therefore, that the enzymes are probably involved in energy-related metabolism at synapses. However, other functions, including those related to neurotransmission, are not excluded.     

Species specificity of bacterial palindromic units

We described previously a family of dispersed palindromic sequences highly repeated in Escherichia coli and Salmonella typhimurium genomes. These sequences, called PU (palindromic units), are located outside structural genes. We report here observations suggesting that PU may have a role in bacterial speciation.     

Comparison of sequences from the malB regions of Salmonella typhimurium and Enterobacter aerogenes with Escherichia coli K12: A potential new regulatory site in the interoperonic region

Summary The malE and malK genes from Salmonella typhimurium, and the MalEFG operon and a portion of malK from Enterobacter aerogenes were cloned and sequenced. Plasmid-borne malE genes from both species and the malF and malG genes from E. aerogenes were expressed normally in Escherichia coli, and their products function in maltose transport. This shows that the malB products from the three species are interchangeable, at least in the combinations tested. The general genetic organization of the malB region is conserved. Potential binding sites and distances between them are highly conserved in the regulatory intervals. An unexpected conserved region was detected, which we call the U box, and which could be another target for a regulatory protein. This hypothesis is supported by the presence of the U box in the regulatory, region of the pulA-malX operon in Klebsiella pneumoniae. The intergenic region between malE and malF from S. typhimurium and E. aerogenes, contains inverted repeats similar to the palindromic units (PU or REP) found at the same location in E. coli. The predicted amino acid sequence of the encoded proteins showed 90% or more identity in every pairwise comparison of species.     

Quantity and Kinetic Properties of Ribulose 1,5-Bisphosphate Carboxylase in C3, C4, and C3-C4 Intermediate Species of Flaveria (Asteraceae)

The kinetic properties of ribulose 1,5-bisphosphate carboxylase(RuBPC) appear to have been modified during evolution of photosynthesisto adjust to changes in substrate availability. C₄ plants areconsidered to have a higher concentration of CO₂ available toRuBPC than C₃plants. In this study, the K_m(CO₂ and catalyticcapacity (k_cat) of RuBPC and the ratio of RuBPC protein to totalsoluble protein from several Flaveria species, including C₃,C₃-C₄ intermediate, and C₄ species, were determined. The C₃and intermediate species had similar K_m(CO₂) values while theC₄ species on average had higher K_m(CO₂) values. The mean ratioof K_cat/K_m for species of each group was similar, supportingthe hypothesis that changes in K_m and K_cat, are linked. Theallocation of total soluble protein to RuBPC was lowest in theC₄ Flaveria species, intermediate in the C₃-C₄ species, andhighest in the C₃ species. The results suggest that during evolutionof C₄ photosynthesis adjustments may occur in the quantity ofRuBPC prior to changes in its kinetic properties. (Received January 4, 1989; Accepted April 11, 1989)     

Calcium-Independent Release of Acetylcholine from Electric Organ Synaptosomes and Its Changes by Depolarization and Cholinergic Drugs

Chemiluminescent detection was applied to measure the continuous spontaneous Ca2+-independent liberation of acetylcholine (ACh) from Torpedo electric organ synaptosomes. Differentiation between the release of ACh and choline was achieved by inhibiting cholinesterases with phospholine, and a way to quantify the continuous release was devised. The method permitted measurements during short time intervals from minute amounts of tissue and without an accumulation of ACh in the medium. Synaptosomes continuously liberated small amounts of ACh during incubations in the presence of 3 mM K+ and in the absence of Ca2+. The spontaneous liberation of ACh was similar both quantitatively and qualitatively at pH values of 8.6 and 7.8. It was unaltered by MgCl2 (10.4 mM), 2-(4-phenylpiperidino)cyclohexanol (10 microM), ouabain (104 microM), atropine (10 microM), and valinomycin (102 nM). Carbamoylcholine brought about a decrease, which could be partially reversed by atropine. The Ca2+-independent output of ACh was increased considerably when the concentration of K+ ions was raised (eightfold at 103 and 35-fold at 203 mM K+). Carbamoylcholine (104 microM) blocked the increase in ACh release produced by high K+; this effect of carbamoylcholine was not reversed by atropine (10 microM). When Ca2+ was added to synaptosomes depolarized by a high concentration of K+, the amount of ACh released during the first 1-3 min after the addition of Ca2+ was at least 20 times higher than in the absence of Ca2+, but the release returned rapidly to predepolarization values. Similarly high values of ACh release could be achieved by adding Ca2+ plus the ionophore A23187 and even higher values by adding Ca2+ plus gramicidin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production and Characterization of Monoclonal Antibodies to Wall-Localized Peroxidases from Corn Seedlings

A library of 22 hybridomas, which make antibodies to soluble wall antigens from the coleoptiles and primary leaves of etiolated corn (Zea mays L.) seedlings, was raised and cloned three times by limit dilution to assure monoclonal growth and stability. Two of these hybridomas made immunoglobulin G antibodies, designated mWP3 and mWP19, which both effectively immunoprecipitated peroxidase activity from crude and partially purified preparations of wall peroxidases. Direct peroxidase-binding assays revealed that both antibodies bound enzymes with peroxidase activity. As judged by immunoblot analyses, mWP3 recognized a M_r 98,000 wall peroxidase with an isoelectric point near 4.2, and mWP19 recognized a M_r 58,000 wall peroxidase. Immunogold localization studies showed both peroxidases are predominately in cell walls.     

Induction by antimycin A of cyanide-resistant respiration in heterotrophic Euglena gracilis: Effects of growth,respiration and protein biosynthesis

The addition of antimycin A during the logarithmic phase of growth of heterotrophic Euglena gracilis cultures (in lactate or glucose medium) was immediately followed by decreased respiration and a cessation of grwoth. Induced cyanideresistent respiration appeared 5 h after the addition of the inhibitor then the cells started to grow again and could be cultured in the presence of antimycin A. Thus the cells exhibited a cyanide-and antimycin-resistant respiration which was, in addition, sensitive to salicylhydroxamic acid and propylgallate. Antimycin-adapted Euglena and control cells were compared for their biomass production and protein synthesis. The difference in growth yield between control and antimycin-adapted cells was not as high as would be expected if only the first phosphorylation site of the normal respiratory chain was active in the presence of antimycin A. Furthermore, the ability to incorporate labelled valine into proteins, under resting-cell conditions, was not changed. Strong correlations were established between the effects of respiratory effectors on O₂ consumption and valine incorporation. These results suggest that sufficient energy for protein synthesis and growth is provided by the operation of the cyanide-resistant respiratory pathway in antimycin-adapted Euglena.Abbreviations DNP dinitrophenol - PG propylgallate - SHAM salicylhydroxamic acid     

Production of a fibronectin-associated lymphokine by cloned mouse T cells

Azobenzenearsonate-specific cloned mouse T cells able to transfer delayed hypersensitivity reactions in vivo produced macrophage agglutination factor (MaggF) after stimulation with mitogen or antigen in vitro. Mitogen (Con A) elicited MAggF production directly from T cells. Responses to Ag were Ag-specific, required syngeneic accessory cells in addition to T cells, and were independent of T cell fine specificity for azobenzenearsonate. Mouse MAggF shared a number of biochemical and immunochemical properties with the fibronectins (FN): 1) high Mr similar to that of plasma FN; 2) binding to gelatin, heparin, and polyclonal antibodies and mAb specific for cellular and plasma FN; 3) inhibition of activity in solution by monoclonal anti-human FN directed against plasma FN gelatin-binding domain; and 4) action on peritoneal exudate macrophages mediated through a FN-receptor cross reactive with one on human monocytes. MAggF production required active protein synthesis and was associated with significant increases in gelatin-binding immunoreactive FN (Mr 440 kDa on immunoblotting) in culture supernatants and T cell lysates. Metabolically labeled peptides could be precipitated by anti-FN from culture supernatants of activated T cells. Stimulated cultures contained significantly more cells with immunohistologically demonstrable cytoplasmic FN than unstimulated control cultures. We suggest that T cell FN is a distinct species of cellular FN which may play an important role in mediating delayed hypersensitivity inflammatory reactions in vivo.