Effect of NaCl, pH, temperature, and atmosphere on growth of Salmonella typhimurium in glucose-mineral salts medium

The interactions of pH (5.0, 6.0, and 7.0), temperature (19, 28, and 37 degrees C), and atmosphere (aerobic versus anaerobic) with NaCl (0, 1, 2, 3, 4, and 5%) on the growth of Salmonella typhimurium ATCC 14028 in defined glucose-mineral salts culture medium were evaluated. Response surface methodology was used to develop equations describing the response of S. typhimurium to environmental changes. The response to an increasing concentration of NaCl at any temperature tested was nonlinear. The maximum growth was predicted to occur at an NaCl concentration of 0.5%, a temperature of 19 degrees C, and an initial pH of 7.0 under aerobic growth conditions. The relative amounts of aerobic growth at 19 degrees C, pH 7.0, and NaCl concentrations of 0, 0.5, 1, 2, 3, 4, and 5% were predicted to be 99.2, 100.0, 98.8, 90.2, 73.5, 48.6, and 15.6%, respectively. Anaerobic growth conditions repressed the amount of growth relative to that under aerobic conditions, and the effects of NaCl and pH were additive at low salt concentrations; however, at higher salt levels anaerobiosis provided protection against the effects of NaCl.     

Fluorometric determination of polystyrene latex: application to the measurement of phagosomes and phagocytosis

Intrinsic fluorescence of polystyrene dissolved in organic solvents such as 1,2-dimethoxyethane was used to develop a sensitive method for the quantification of polystyrene latex beads. This method allows the assay of latex in the microgram range and is one order of magnitude more sensitive than the conventional spectrophotometric method. The fluorometric technique was used in the quantification of phagocytic latex particle uptake by macrophages and in the quantification of isolated phagosomal fractions.     

Topoisomerase II cleavage of herpes simplex virus type 1 DNA in vivo is replication dependent

The genome of herpes simplex virus type 1 contains a large number of recognition sites for eucaryotic DNA type II topoisomerase. Topoisomerase II sites were identified by means of the consensus sequence described previously (J.R. Spitzner and M.T. Muller, Nucleic Acids Res. 16:5553-5556, 1988) and then confirmed by sequencing DNA cleavages introduced by purified topoisomerase II. In vivo, host topoisomerase II also introduced double-stranded DNA breaks in the viral genome at sites predicted by the consensus sequence. Host topoisomerase II acted on all immediate-early genes as well as on genes from other temporal classes; however, cleavages were not detected until 4 to 5 h postinfection and were most intense at 10 h postinfection. Topoisomerase II cleavages were not detected when viral DNA replication was prevented with phosphonoacetic acid. These data indicate that, although progeny viral genomes are acted upon by host topoisomerase II, this enzyme either does not act on parental viral genomes before DNA replication or acts on them with such low efficiency that cleavages are beyond our limit of detection. The findings suggest that host topoisomerase II is involved in aspects of viral replication at late times in the infectious cycle.     

High-frequency shoot regeneration from leaves of the apple rootstock M26 (Malus pumila Mill.)

Regeneration of shoots was achieved from in vitro leaves of M26 at frequencies close to 100% on a medium based on MS salts and LS vitamins, containing 4.4 M BA and 0.5 M NAA. Dark and red light gave the best results in inducing shoot regeneration. White light at high intensity helped development of regenerated shoots. Inorganic nitrogen could be reduced by 75% without negative effect, and the presence of NH₄ ⁺ was necessary for regeneration. Leaves were able to regenerate after a 3 kR irradiation (gamma rays), not after 4 kR. Optimal dose should be between 1 and 2 kR.Publication No 261, Centro Studi Tecnica Frutticola-CNR. Research work supported by CNR, Italy special grant IPRA. Sub-project 1, paper No. 1496.     

Laboratory studies of host finding,acceptance and suitability of the dung-breeding fly,Haematobia thirouxi potans [Dipt.: Muscidae], byAleochara sp. [Col.: Staphylinidae]

Aleochara sp. from southern Africa is a potential candidate for introduction to Australia for control of the Australian buffalo fly,Haematobia irritans exigua De Meijere. Aspects of its host searching and acceptance behaviour were studied in South Africa usingHaematobia thirouxi potans (Bezzi), to assess, in part, its potential value as a biological control agent and to provide a basis for the development of a mass rearing technique. First instars ofAleochara sp. were found to parasitize most effectively those hosts that had buried themselves in the substrate, probably by orienting to a chemical cue and using the tunnels made by the post-feeding larvae. Lower rates of parasitization occurred in pupae placed on the surface of the substrate or buried by the experimenters. They would accept, parasitize and develop on all ages ofH. thirouxi potans pupae, but showed reduced survival in the very oldest ones. Although there was no difference in relative acceptability among hosts of different ages, a higher proportion of younger hosts were parasitized when these had buried themselves, probably due to a tendency of the parasitoids to follow fresher tunnels into the substrate. Survival was high (>80%) at relative humidities from 10–91% but was less than 20% at an R.H. of 100%. In rearing trials, the most economical ratio of parasitoid eggs: hosts was 1∶1.5, with 70% of the parasitoids developing successfully to adults.      

Accessibility and structural role of histone domains in chromatin. biophysical and immunochemical studies of progressive digestion with immobilized proteases

The accessibility and role of histone regions in chromatin fibres were investigated using limited proteolysis with enzymes covalently bound to collagen membranes. The changes in chromatin conformation and condensation monitored by various biophysical methods, were correlated to the degradation of the histone proteins revealed by antibodies specific for histones and histone peptides. Upon digestion with trypsin and subtilisin, chromatin undergoes successive structural transitions. The cleavage of the C-terminal domains of H1, H2A and H2B, and of the N-terminal tail of H3 led to a decondensation of chromatin fibres, indicated by increases in electric birefringence and orientational relaxation times. It corresponds to a 15% increase in linear dimensions. The degradation of the other terminal regions of histones H3, H2A and H2B resulted in the appearance of hinge points between nucleosomes without alteration of the overall orientation of polynucleosome chains. Despite the loss of all the basic domains of H1, H3, H2A and H2B, no significant change in DNA-protein interactions occurred, suggesting that most of these protease-accessible regions interact weakly, if at all, with DNA in chromatin. Further proteolysis led to H4 degradation and other additional cleavages of H1, H2B and H3. This caused the relaxation of no more than 8% of the total DNA but resulted in changes in the ability of chromatin to condense at high ionic strength. More extensive digestion resulted in a total unravelling of nucleosomal chains which acquired properties similar to those of H1-depleted chromatin, although the globular part of H1 was still present. The data suggest that histone-histone interactions between H1 and core histone domains play a central role in stabilizing the chromatin fibres, and cuts in H3, H2A and H2B as well as H1, seem necessary for chromatin expansion. On the contrary, H4 might be involved in the stabilization of nucleosomes only.