Cross-Reactivity of Neutralizing Antibodies among Malignant Catarrhal Fever Viruses

Some members of the gamma herpesvirus genus Macavirus are maintained in nature as subclinical infections in well-adapted ungulate hosts. Transmission of these viruses to poorly adapted hosts, such as American bison and cattle, can result in the frequently fatal disease malignant catarrhal fever (MCF). Based on phylogenetic analysis, the MCF viruses (MCFV) cluster into two subgroups corresponding to the reservoir hosts’ subfamilies: Alcelaphinae/Hippotraginae and Caprinae. Antibody cross-reactivity among MCFVs has been demonstrated using techniques such as enzyme linked immunosorbent and immunofluorescence assays. However, minimal information is available as to whether virus neutralizing antibodies generated against one MCFV cross react with other members of the genus. This study tested the neutralizing activity of serum and plasma from select MCFV-infected reservoir hosts against alcelaphine herpesvirus 1 (AlHV-1) and ovine herpesvirus 2 (OvHV-2). Neutralizing antibody activity against AlHV-1 was detected in samples from infected hosts in the Alcelaphinae and Hippotraginae subfamilies, but not from hosts in the Caprinae subfamily. OvHV-2 neutralizing activity was demonstrated in samples from goats (Caprinae) but not from wildebeest (Alcelaphinae). These results show that neutralizing antibody cross reactivity is present to MCFVs within a virus subgroup but not between subgroups. This information is important for diagnosing infection with MCFVs and in the development of vaccines against MCF.     

Development of efficient suicide mechanisms for biological containment of bacteria

To optimize plasmid containment, we have systematically investigated the factors that limit the killing efficiency of a suicide system based on the relF gene from Escherichia coli controlled by inducible lac promoters and placed on plasmids. In induction experiments with this suicide system, killing efficiency was unaffected by temperature and growth medium; there was no requirement for great promoter strength or high plasmid copy number. We could demonstrate that the factors limiting killing were the mutation rate of the suicide function and the reduced growth rate caused by a basal level of expression of the suicide gene during normal growth, which can give a selective growth advantage to cells with mutated suicide functions. The capacity of the plasmid-carried killing system to contain the plasmid was tested in transformation, transduction, and conjugational mobilization. The rate of plasmid transfer detected in these experiments seemed too high to provide adequate biological containment. As expected from the induction experiments, plasmids that escaped containment in these transfer experiments turned out to be mutated in the suicide function. With lac-induced suicide as a test, the efficiency of the system was improved by tightening the repression of the suicide gene, thereby preventing selection of cells mutated in the killing function. Reduction of the mutational inactivation rate of the suicide system by duplication of the suicide function augmented the efficiency of the suicide dramatically. These results permit the construction of extremely efficient biological containment systems.     

Adenosine and Glutamate Modulate Each Other's Release from Rat Hippocampal Synaptosomes

Abstract: In rat hippocampal synaptosomes, adenosine decreased the K⁺ (15 mM) or the kainate (1 mM) evoked release of glutamate and aspartate. An even more pronounced effect was observed in the presence of the stable adenosine analogue, R-phenylisopropyladenosine. All these effects were reversed by the selective adenosine A₁ receptor antagonist 8-cyclo-pentyltheophylline. In the same synaptosomal preparation, K⁺ (30 mM) strongly stimulated the release of the preloaded [³H]adenosine in a partially Ca²⁺-dependent and tetrodotoxin (TTX)-sensitive manner. Moreover, in the same experimental conditions, both l -glutamate and l -aspartate enhanced the release of [³H]adenosine derivatives ([³H]ADD). The gluta-mate-evoked release was dose dependent and appeared to be Ca²⁺ independent and tetrodotoxin insensitive. This effect was not due to metabolism because even the nonmetabolizable isomers d -glutamate and d -aspartate were able to stimulate [³H]ADD release. In contrast, the specific glutamate agonists N-methyl-d -aspartate, kainate, and quisqualate failed to stimulate [³H]ADD release, suggesting that glutamate and aspartate effects were not mediated by known excitatory amino acid receptors. Moreover, NMDA was also ineffective in the absence of Mg²⁺ and l -glutamate-evoked release was not inhibited by adding the specific antagonists 2-amino-5-phosphonovaleric acid or 6–7-dinitroquinoxaline-2, 3-dione. The stimulatory effect did not appear specific for only excitatory amino acids, as γ-anunobutyric acid stimulated [³H]ADD release in a dose-related manner. These results suggest that, at least in synaptosomal preparations from rat hippocampus, adenosine and glutamate modulate each other's release. The exact mechanism of such interplay, although still, unknown, could help in the understanding of excitatory amino acid neurotoxicity.     

Stimulation of ribose-5-phosphate and 5-phosphoribosyl-1-pyrophosphate generation by pyrroline-5-carboxylate in mouse liver in vivo: evidence for a regulatory role of ribose-5-phosphate availability in nucleotide synthesis.

Pyrroline-5-carboxylase (P5C), a physiological stimulator of hexose-monophosphate-pentose pathway activity, was found before to increase 5-phosphoribosyl-1-pyrophosphate (PRPP) generation and nucleotide synthesis in human erythrocytes and cultured fibroblasts. We now report the stimulation of PRPP generation by P5C also in mouse liver in vivo. In addition we demonstrated a simultaneous elevation in ribose-5-phosphate (R5P) concentration, which was relatively smaller and transient. The demonstrated effect of P5C on liver R5P and PRPP content in vivo provides strong evidence for the physiological role of R5P availability in the regulation of PRPP and purine production.     

A lung type prostaglandin F synthase is expressed in bovine liver: cDNA sequence and expression in E. coli.

Using the cDNA of bovine lung prostaglandin F synthase (EC 1.1.1.2) as a probe, we isolated a clone from a bovine liver cDNA library which differed in only eleven nucleotides from the probe. The corresponding protein contained three amino acid substitutions, including a leucine residue which is conserved throughout all aldo-keto reductases. We inserted the liver cDNA into expression vector pUC19 and expressed the recombinant liver enzyme in E.coli. The purified liver enzyme reduced prostaglandin H2 as well as prostaglandin D2 and various carbonyl compounds. The high relative activity against prostaglandin H2 in combination with a high Km value for prostaglandin D2 identified this liver enzyme as a lung type prostaglandin F synthase. However, the binding constant for NADPH of the liver enzyme was 3.5 fold higher than that of lung prostaglandin F synthase.     

Effects of some ionophore antibiotics and polyoxins on the growth of anaerobic rumen fungi

The growth of mixed rumen fungi in vitro was suppressed by both ionophore antibiotics (salinomycin, monensin and portmicin) and polyoxins (polyoxin B and D: inhibitors of chitin synthesis). The fungistatic effect of the ionophores on a Piromonas spp. was more pronounced than on a Neocallimastix spp. The polyoxins, however, were more potent fungistatically against the Neocallimastix spp. than the Piromonas spp. Higher concentrations of the polyoxins were required to elicit the same effect as that observed with the ionophores. Salinomycin administration decreased fungal count in the rumen of sheep, but fungal count increased after the cessation of the feeding of the antibiotic. Polyoxin D also suppressed the growth of fungi in vivo, but the effect was short-lived. Nevertheless, both bacterial and protozoal counts tended to increase during and after the administration of polyoxin D. Total volatile fatty acid concentrations in the rumen tended to increase during the period of polyoxin D administration. This increasing tendency was maintained for 10 d after the cessation of antibiotic administration. Offering polyoxin D to sheep increased production of propionate ( P < 0·05), while decreasing that of acetate. The results indicate that the rumen fungi are sensitive to chitin synthesis inhibitors as well as ionophores, and are essential members of microbes in the rumen ecosystem.     

Histophotometric characteristics of structural-metabolic heterogeneity of hepatocytes in acute hemorrhage and pulmonary artery thromboembolism]

Basing on the data obtained at early autopsies, we compared the structure and metabolism of liver acini in acute blood loss and pulmonary artery thromboembolism. Morpho- and pathogenetic impairment of hepatocytes of various acinic zones was correlated and studied histo-enzymatically.     

Seasonal and circadian fluctuations in blood biochemical indicators in mice in natural conditions and exposed to constant light]

The fluctuations of activity of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) and those of the levels of protein, glucose, cholesterol, bilirubin, creatinine, blood urea nitrogen, K+, Cl-, Na+ in blood plasma of mice in natural conditions (NC) and exposed to constant light (CL) were studied in different seasons of the year (in January, April, July, October) on days 18, 24, 6 (at 12 o'clock). Most indices both in NC and CL animals had seasonal rhythm similar for each of them. This proves a primary effect of environmental geoclimatic factors of formation of circadian periodicals as compared to desynchronization in constant light revealed by Kosinor analysis in winter (acrophase from 14.16 till 16.32 o'clock) and autumn (acrophase from 23.03 til 4.40 o'clock). During the same seasons one can observe the maximum desynchronization influences of constant light, which leads to abrupt falling (to the 10-fold and more) of the fluctuations amplitude and in some cases to stabilization of circadian rhythm.