Effects on microbial activity by extraction of indigenous cells from soil slurries

Abstract: Possible effects on the physiological activity and culturability of soil microorganisms by different soil dispersion procedures, and effects on activity caused by extracting bacteria from soil, were investigated. There was no apparent difference in cfu's with dispersion of a silty loam soil and a loamy sand soil with pyrophosphate as compared to dispersion in NaCl. Substrate-induced respiration was reduced in the silty loam soil, and methanol oxidation was reduced in the loamy sand soil with dispersion in pyrophosphate, and the soil pH was irreversibly increased by the treatment. Extracted bacterial fractions had lower numbers of culturable cells as percentage of the total number of bacteria in each fraction, lower respiration rates and no methanol oxidation activity as compared to the soil slurry both before and after extraction. The physiological activity was apparently not affected by the number of cells extracted. This indicates that the increased extraction rate of indigenous soil bacteria obtained by effective disruption of aggregates and detachment of cells from surfaces, only results in increased extraction of cells that have been physiologically changed as a result of the extraction process.     

Species- and age-dependent sensitivity to ozone in young plants of pea,wheat and spinach: Effects on acyl lipid and pigment content and metabolism

Acyl lipids and pigments were analyzed in young plants of garden pea, spring wheat and spinach exposed to < 5 or 65 nl l^?1 ozone 12 h per day for 6 days. In one set of experiments, the plants were exposed to ¹⁴CO₂ for 2 h 3 days prior to ozone exposure. The plants responded differently to the moderately enhanced level of ozone used Spinach was not at all sensitive while in both pea and wheat, leaves of different ages differed in ozone sensitivity. In pea, ozone sensitivity increased with leaf age. In the second and third oldest leaves, the amounts of galactolipids per leaf area and the proportions of 18:3 of the total lipid extract and of phosphatidylglycerol decreased. In the second oldest leaf, ozone also caused a decreased proportion of 18:3 of monogalactosyldiacylglycerol. In the fourth oldest leaf, lipid composition and galactolipid unsaturation was unaffected, but ozone caused decreased leaf expansion resulting in increased acyl lipid content per leaf area. In both the first and second leaves of wheat, ozone fumigation caused a marked decrease in the content of monogalactosyldiacylglycerol and in the first leaf, the contents of phosphatidylcholine and phosphatidylethanolamine increased. The proportion of 18:3 in phosphatidylcholine was larger in ozone-fumigated than in control plants, while the reverse applied for phosphatidylglycerol. In the oldest sampled leaves of pea and wheat, ozone caused an increase in the radioactivity associated with β-carotene, indicating increased turnover. Thus, while spinach was unaffected, in both pea and wheat ozone caused a decrease in the proportion of chloroplast membrane lipids to non-chloroplast membrane lipids in older leaves while younger leaves were less sensitive.     

Tn5 insertion mutants of Pseudomonas sp. 267 defective in siderophore production and their effect on clover (Trifolium pratense) nodulated with Rhizobium leguminosarum bv. trifolii

Pseudomonas sp. strain 267 isolated from soil promoted growth of different plants under field conditions and enhanced symbiotic nitrogen fixation in clover under gnotobiotic conditions. This strain produced pyoverdine-like compound under low-iron conditions and secreted vitamins of the B group. The role of fluorescent siderophore production in the beneficial effect of strain 267 on nodulated clover plants was investigated. Several non-fluorescent (Pvd^-) Tn5 insertion mutants of Pseudomonas sp. strain 267 were isolated and characterized. The presence of Tn5 insertions was confirmed by Southern analysis of EcoRI digested genomic DNA of each derivative strain. The siderophore-negative mutants were compared to the parental strain with respect to their growth promotion of nodulated clover infected with Rhizobium leguminosarum bv. trifolii 24.1. We found that all isolated Pvd^- mutants stimulated growth of nodulated clover plants in a similar manner to the parental strain. No consistent differences were observed between strain 267 and Pvd^- derivatives strains with respect to their plant growth promotion activity under gnotobiotic conditions.Dr Deryto died in august 1994     

Somatic embryogenesis and organogenesis in Encephalartos dyerianus and E. natalensis

Callus cultures were initiated from zygotic embryos of Encephalartos dyerianus and E. natalensis. Callus of both species were transferred onto a modified B5 medium containing different combinations of 2,4-dichlorophenoxyacetic acid and kinetin. Somatic embryogenesis and shoot organogenesis occurred in both species. The embryos were dicotyledonary. To date none of the embryos have matured.Abbreviations ABA abscisic acid - 2,4-D 2,4-dichlorophenoxyacetic acid     

Overexpression of P-glycoprotein in heat-and/or drug-resistant hepatoma variants

We have earlier isolated a glucocorticoid-resistant, dedifferentiated rat hepatoma variant, the clone 2, which exhibited deficient stress activation of the major stress-inducible heat-shock protein hsp68.Multidrug-resistant variants were isolated from clone 2 cells using increasing concentrations of colchicine. The induction deficiency of hsp68 was maintained in the colchicine-resistant clone 2 cells grown for several months in the presence of 1 g/ml colchicine (termed ashighly multidrug-resistant variant) indicating that this heat-shock protein is not involved in the multidrug resistance. No alteration of the protein synthesis pattern was observed except the strong increase of the P-glycoprotein, which correlated with high level of corresponding mRNA. Stableheat-resistant variants of clone 2 were also isolated, which showed increaseddrug resistance to several drugs, i.e. they becamemoderately multidrug-resistant. This moderate multidrug resistance of the heat-resistant variants was further increased by stepwise selection with colchicine (highly multidrug-resistant heat-resistant variants). The levels of P-glycoprotein mRNA and protein were elevated both in the heat-resistant, non drug selected, moderately drug-resistant and in heatresistant, colchicine selected, highly drug-resistant variants. Decreased retention of antitumor drugs was observed in all multidrug-resistant variants indicating that P-glycoprotein was functional. Verapamil increased doxorubicin retention and cytotoxicity significantly. Our results showing that severely stressed hepatoma cells overexpressed the multidrug resistance gene(s) raise the possibility that the P-glycoprotein may participate in protection against enviromental stress such as heat.Abbreviations hsp heat-shock protein - MDR multidrug resistance - P-gp P-glycoprotein     

Isolation and characterization of extragenic mutations affecting the expression of the uroporphyrinogen decarboxylase gene (HEM12) in Sacharomyces cerevisiae

Uroporphyrinogen decarboxylase (Uro-d; EC 4.1.1.37), the fifth enzyme in the heme biosynthetic pathway, which catalyzes the sequential decarboxylation of uroporphyrinogen to coproporphyrinogen, is encoded by the HEM12 gene in Saccharomyces cerevisiae. The HEM12 gene is transcribed into a major short mRNA and a minor longer one, approximately 1.35 and 1.55 kb, respectively, in size, and that differ in the 5′ untranslated region. “Uroporphyric” mutants, which have no mutations in the HEM12 gene but accumulate uroporphyrinogen, a phenotype chracteristic of partial Uro-d deficiency, were investigated. Genetic analysis showed that the mutant phenotype depends on the combined action of two unlinked mutations, udt1 and either ipa1, ipa2, or ipa3. ipa1 is tightly linked to HEM12 The mutation udt1 apparently acts specifically on the HEM12 gene, and causes a six to tenfold decrease in the levels of the short HEM12 mRNA, in the β-galactosidase activity of a HEM12-lacZ fusion, in immunodetectable protein and enzyme activity. But heme synthesis is normal and porphyrin accumulation was modest. The mutations ipa1, ipa2, and ipa3 had no phenotype on their own, but they caused an increase in porphyrin accumulation in a udt1 background. This multiplicity of genetic factors leading to uroporphyric yeast cells closely resembles the situation in human porphyria cutanea tarda.     

Suppression of programmed neuronal death by a thapsigargin-induced Ca2+ influx

Rat sympathetic neurons undergo programmed cell death (PCD) in vitro and in vivo when they are deprived of nerve growth factor (NGF). Chronic depolarization of these neurons in cell culture with elevated concentrations of extracellular potassium ([K⁺]_o) prevents this death. The effect of prolonged depolarization on neuronal survival is thought to be mediated by a rise of intracellular calcium concentration ([Ca²⁺]_i) caused by Ca²⁺ influx through voltage-gated channels. In this report we investigate the effects of chronic treatment of rat sympathetic neurons with thapsigargin, an inhibitor of intracellular Ca²⁺ sequestration. In medium containing a normal concentration of extracellular Ca²⁺ ([Ca²⁺]_o), thapsigargin caused a sustained rise of intracellular Ca²⁺ concentration and partially blocked death of NGF-deprived cells. Elevating [Ca²⁺]_o in the presence of thapsigargin further increased [Ca²⁺]_i, suggesting that the sustained rise of [Ca²⁺]_i was caused by a thapsigargin-induced Ca²⁺ influx. This treatment potentiated the effect of thapsigargin on survival. The dihydropyridine Ca²⁺ channel antagonist, nifedipine, blocked both a sustained elevation of [Ca²⁺]_i and enhanced survival caused by depolarization with elevated [K⁺]_o, suggesting that these effects are mediated by Ca²⁺ influx through L-type channels. Nifedipine did not block the sustained rise of [Ca²⁺]_i or enhanced survival caused by thapsigargin treatment, indicating that these effects were not mediated by influx of Ca²⁺ through L-type channels. These results provide additional evidence that increased [Ca²⁺]_i can suppress neuronal PCD and identify a novel method for chronically raising neuronal [Ca²⁺]_i for investigation of this and other Ca²⁺-dependent phenomena. © 1995 John Wiley & Sons, Inc.