Kinetics of nitrite oxidation in two Nitrobacter species grown in nitrite-limited chemostats

The influence of growth rate, the presence of acetate and variation in the dissolved oxygen concentration on the kinetics of nitrite oxidation was studied in suspensions of intact cells of Nitrobacter winogradskyi and Nitrobacter hamburgensis. The cells were grown in nitrite-limited chemostats at different dilution rates under chemolithotrophic and mixotrophic conditions. Growth of N. hamburgensis in continuous culture was dependent on the presence of acetate. Acetate hardly affected the maximal nitrite oxidation rate per cell (V _max), but displayed a distinctly negative effect on the saturation constants for nitrite oxidation (K _m ) of both Nitrobacter species. This effect was reversible; when acetate was removed from the suspensions the K _m -values for nitrite oxidation returned to their original values. A reduction of the dissolved oxygen concentration from 100% to 18% air saturation slightly decreased the V _max of chemolithotrophically grown N. winogradskyi cells, whereas a 2.3 fold increase was observed with mixotrophically grown cells of N. hamburgensis. It is suggested that the large variation in K _m encountered in field samples could be due to this observed phenotypic variability. The V _max per cell is not a constant, but apparently is dependent on growth rate and environmental conditions. This implies that potential nitrite oxidation activity and numbers of cells are not necessarily related. Considering their kinetic characteristics, it is unlikely that N. hamburgensis is able to compete succesfully with N. winogradskyi for limiting amounts of nitrite under mixotrophic conditions. However, at reduced partial oxygen tensions, N. hamburgensis may become the better competitor.     

Comparative sequence analysis of rotavirus genomic segment 6--the gene specifying viral subgroups 1 and 2

Cloned DNA copies of rotavirus genomic segment 6 from simian 11 (subgroup 1) and human strain Wa (subgroup 2) rotaviruses have been used to determine the nucleotide sequences of the gene that determines viral subgroup specificity. Both genomic segments are 1,356 nucleotides in length and possess 5'- and 3'-terminal untranslated regions of 23 and 142 nucleotides, respectively. The inferred amino acid sequence reveals VP6 to be a polypeptide of 397 amino acids in which more than 90% of the amino acid sequence is conserved between the two viruses. There are 34 amino acid changes between the subgroup 1 and 2 polypeptides, most clustered in three regions of the molecule at residues 39 through 62, 80 through 122, and 281 through 315.     

Antigenic drift in influenza virus H3 hemagglutinin from 1968 to 1980: multiple evolutionary pathways and sequential amino acid changes at key antigenic sites

Surveys of the antigenic properties of a wide range of variants of the H3N2 (Hong Kong) influenza virus subtype have revealed complex patterns of variants cocirculating during each of the main epidemic eras of the subtype. We determined hemagglutinin (HA) gene sequences for 14 isolates chosen to give the wildest possible spread of variant types. The addition of these data to existing HA gene sequence information for other variants provides a comprehensive picture of HA gene evolution during antigenic drift among H3N2 subtype viruses. The data reveal the existence of multiple evolutionary pathways during at least one period of development of the subtype and strikingly demonstrate that amino acid changes are limited to a small number of locations on the HA molecule during antigenic drift. The occurrence of sequential amino acid changes at key positions within these variable regions suggests that the HA structure has remained constant during subtype evolution so that only limited possibilities remain for further antigenic drift among H3N2 viruses.     

Sequences in rotavirus glycoprotein VP7 that mediate delayed translocation and retention of the protein in the endoplasmic reticulum

Glycosylation and translocation of the simian rotavirus protein VP7, a resident ER protein, does not occur co-translationally in vivo. In pulse-chase experiments in COS cells, nonglycosylated VP7 was still detectable after a 25-min chase period, although the single glycosylation site was only 18 residues beyond the signal peptide cleavage site. After labeling, glycosylated and nonglycosylated VP7 was recovered in microsomes but the latter was sensitive to trypsin (i.e., the nascent protein became membrane associated) but most of it entered the ER posttranslationally because of a rate-limiting step early in translocation. In contrast with the simian protein, bovine VP7 was glycosylated and translocated rapidly. Thus, delayed translocation per se was not required for retention of VP7 in the ER. By constructing hybrid proteins, it was further shown that the signal peptide together with residues 64-111 of the simian protein caused delayed translocation. The same sequences were also necessary and sufficient for retention of simian VP7 in the ER. The data are consistent with the idea that certain proteins are inserted into the ER membrane in a loop configuration.     

The immunogenicity of VP7, a rotavirus antigen resident in the endoplasmic reticulum, is enhanced by cell surface expression.

The glycoprotein VP7, the major serotype antigen of rotaviruses, is localized to the endoplasmic reticulum (ER) of the cell, where it is retained as a membrane-associated protein before assembly into mature virus particles. Wild-type VP7 expressed by a recombinant vaccinia virus was also located internally and was poorly antigenic. Using recombinant techniques, a correctly processed, secreted form of VP7 (S.C. Stirzaker and G.W. Both, Cell 56:741-747, 1989) was modified by addition to its C terminus of the membrane anchor and cytoplasmic domains from the influenza virus hemagglutinin. The hybrid protein was directed to the surface of cells, where it was anchored in the plasma membrane. When expressed in mice and rabbits by a recombinant vaccinia virus, the surface-anchored antigen stimulated a level of rotavirus-specific antibodies that was greater than 100-fold above the level induced by wild-type VP7. T-cell responses to the novel antigen were also elevated in comparison with the wild-type, intracellular protein. Cell surface anchoring may provide a strategy to increase the immunogenicity of intracellular antigens from other parasites and viruses.     

The dynamics of the cassava green mite Mononychellus tanajoa in a seasonally dry area in Kenya

The population dynamics of the cassava green mite Mononychellus tanajoa was studied on cassava during 35 weeks (early March to first of November 1989) in an experimental field near Lake Victoria in Western Kenya. The mite population peaked at the onset of the long dry season with 1,100 mites/leaf, declined sharply to a level of about 300 individuals/leaf, not to increase again until the next rainy season commenced. An indigenous phytoseiid predator Iphiseius degenerans was abundant during the dry spell with a maximum about 9 predators/leaf.A nonlinear regression analysis revealed that food depletion in combination with I. degenerans predation limited the population growth of the mites, whereas rain intensity had no effect. The predator exhibited no aggregative response to high densities of M. tanajoa and stayed mainly in the lower part of the canopy while the spider mites preferred the top, indicating that I. degenerans is a generalist predator without capacity to control M. tanajoa alone. However, in combination with another density dependent factor, such as food depletion, the predator may have prevented the spider mites from causing complete defoliation during the dry season.     

The effect of the cassava green mite Mononychellus tanajoa on the growth and yield of cassava Manihot esculenta in a seasonally dry area in Kenya

The effect of the cassava green mite Mononychellus tanajoa on the growth and yield of cassava Manihot esculenta was studied over a 10-month period in two field trials near Lake Victoria in Kenya. One plot was maintained free of mites by means of acaricide, while the other was artificially infested.The highest population density of M. tanajoa occurred during the dry season. A maximum leaf area index (LAI) of about 2 was reached at the onset of the dry season. The total leaf area of mite infested plants was reduced compared with uninfested plants during the dry spell. During the following rainy season infested plants recovered and attained the same leaf area as uninfested plants. A multiple regression model predicting the leaf area showed that 58% of the seasonal variation could be explained by plant age, soil water, and leaf injury.The net growth rate of infested plants was lower than that of uninfested plants. Maximum values of 21 (infested plants) and 49 (uninfested plants) g m^-2 week^-1 were attained at the onset of the second rainy season. No difference was found between uninfested and infested plants with respect to net assimilation rates per unit leaf area during the dry season. The net assimilation rates reached a maximum almost at the same time as the growth rates, but the infested plants peaked slightly earlier and at a lower level than the uninfested plants. M. tanajoa did not affect the relative allocation of dry matter into stems and storage roots, but the absolute allocation of dry matter declined with increasing mite injury. Thus, after 10 months the dry matter of infested plants was reduced by 29% and 21% for storage roots and stems, respectively, compared with the uninfested plants.     

The genes encoding the peripheral cannabinoid receptor and alpha-L-fucosidase are located near a newly identified common virus integration site, Evi11.

A new common region of virus integration, Evi11, has been identified in two retrovirally induced murine myeloid leukemia cell lines, NFS107 and NFS78. By interspecific backcross analysis, it was shown that Evi11 is located at the distal end of mouse chromosome 4, in a region that shows homology with human 1p36. The genes encoding the peripheral cannabinoid receptor (Cnr2) and alpha-L-fucosidase (Fuca1) were identified near the integration site by using a novel exon trapping system. Cnr2 is suggested to be the target gene for viral interference in Evi11, since proviruses are integrated in the first intron of Cnr2 and retroviral integrations alter mRNA expression of Cnr2 in NFS107 and NFS78. In addition, proviral integrations were demonstrated within the 3' untranslated region of Cnr2 in five independent newly derived CasBrM-MuLV (mouse murine leukemia virus) tumors, CSL13, CSL14, CSL16, CSL27, and CSL97. The Cnr2 gene encodes a seven-transmembrane G-protein-coupled receptor which is normally expressed in hematopoietic tissues. Our data suggest that the peripheral cannabinoid receptor gene might be involved in leukemogenesis as a result of aberrant expression of Cnr2 due to retroviral integration in Evi11.     

Cholinergic Plasticity of Oscillating Neuronal Assemblies in Mouse Hippocampal Slices

The mammalian hippocampus expresses several types of network oscillations which entrain neurons into transiently stable assemblies. These groups of co-active neurons are believed to support the formation, consolidation and recall of context-dependent memories. Formation of new assemblies occurs during theta- and gamma-oscillations under conditions of high cholinergic activity. Memory consolidation is linked to sharp wave-ripple oscillations (SPW-R) during decreased cholinergic tone. We hypothesized that increased cholinergic tone supports plastic changes of assemblies while low cholinergic tone favors their stability. Coherent spatiotemporal network patterns were measured during SPW-R activity in mouse hippocampal slices. We compared neuronal activity within the oscillating assemblies before and after a transient phase of carbachol-induced gamma oscillations. Single units maintained their coupling to SPW-R throughout the experiment and could be re-identified after the transient phase of gamma oscillations. However, the frequency of SPW-R-related unit firing was enhanced after muscarinic stimulation. At the network level, these changes resulted in altered patterns of extracellularly recorded SPW-R waveforms. In contrast, recording of ongoing SPW-R activity without intermittent cholinergic stimulation revealed remarkably stable repetitive activation of assemblies. These results show that activation of cholinergic receptors induces plasticity at the level of oscillating hippocampal assemblies, in line with the different role of gamma- and SPW-R network activity for memory formation and –consolidation, respectively.