Between-year variation in flowering and fruit set in frost-prone and frost-sheltered populations of dioecious Rubus chamaemorus

Summary The flowering and fruiting patterns of the dioecious perennial herb Rubus chamaemorus L. were studied in frost-prone (open) and frost-sheltered (Shaded) habitats in northern Sweden over 6 years. The number of ramets with flower buds, the proportion of flower buds that opened, and fruit set varied markedly between years. In the frost-prone populations, the occurrence or absence of detrimental frosts during the development of flowers and fruit could explain much of the variation, both in the proportion of flower buds that developed into flowers, and in fruit set. In the frost-sheltered populations, most female flowers that did not develop into fruit aborted without any signs of physical damage and before any ovules had begun to enlarge. Flower mortality caused by herbivores feeding on reproductive parts was commonly low, but reached values higher than 10% in one of the shaded populations. Hand-pollination increased the proportion of ovules producing seeds in the mature fruits by about 20%, and in one year also increased fruit set significantly in one population. Fruit-producing female ramets had a higher mortality and a lower probability of flowering in the subsequent year than male ramets and non-fruiting female ramets. In R. chamaemorus, the conditions for fruit maturation are highly unpredictable at the time of flower initiation. It is suggested that the apparent over-initiation of flower buds is advantageous, as it allows the plant to attain a high reproductive success in years favourable for flowering and fruit development.     

Inhibited growth of common enteropathogenic bacteria in lactic-fermented cereal gruels

A natural lactic fermentation of mixtures of water and whole flour of either maize or high-tannin sorghum was obtained either before or after cooking to a weaning gruel: The preparations had a final pH of about 3.8 (range 3.67 to 4.00) and a ratio of lactic acid to acetic acid of 91 (w/w). The growth of added (about 10⁷ c.f.u./g gruel) Gram-negative intestinal pathogenic bacteria, enterotoxigenicEscherichia coli, Campylobacter jejuni, Shigella flexneri andSalmonella typhimurium, was strongly inhibited in the sour gruels, and the effect could primarily be explained by the low pH caused by the formation of lactic and acetic acids during the fermentation process. Of the added Gram-positive bacteria,Bacillus cereus andStaphylococcus aureus showed similar inhibited growth up to 7h after inoculation in the sour gruels. The strain ofStaphylococcus, however, showed only a continued reduction in growth in the fermented gruel samples, which had a viable lactic bacteria culture indicating the presence of a bacteriocin. This implies that a low pH (< 4.0) alone is not sufficient to sustain the inhibition of the growth ofStaphylococcus aureus. The survival studies were carried out at optimal temperatures for each respective enteropathogen.     

Genetic variation in relation to demography of peripheral pool frog populations (Rana lessonae)

Summary Genetic variation in an isolated northern metapopulation of the pool frog (Rana lessonae) in Sweden was compared to that of Central European populations using enzyme electrophoresis and literature data. Of the 31 loci scored, two (EST-2 andIDH-2) were polymorphic while no variation occurred in seven of the eight loci which are polymorphic in Central European populations.The heterozygosity level of the Swedish pool frogs is very low compared to that of other anuran populations, but their mean proportion of fertilized eggs within egg masses (97.5%) was not lower than in more heterozygous species, and their body size-specific fecundity did not differ from that of Polish conspecifics. The low genetic variability of the Swedish pool frogs is discussed in relation to features of the local populations such as size (N), calculated effective size (N _e ) reproductive success and probable history. It is concluded that long-term strong fluctuations in population size caused by reporductive failure in cold years have contributed more to the low genetic variability than could a single founder event due to a recent introduction by man.     

Inhibition of rat yolk sac tumour growth in vivo by a monoclonal antibody to the retroviral molecule P15E

Summary A monoclonal mouse IgG2b antibody 19F8, directed towards a determinant on the retroviral transmembranous molecule p15E, binds selectively to certain rat tumours, including all tested yolk sac tumours, one rat colon carcinoma, one spontaneous kidney carcinoma and an adenovirus-type-9-induced rat breast tumour, as tested by antibody-dependent cellular cytotoxicity (ADCC) and immunohistochemistry. Groups of rats receiving yolk sac tumour F56 isografts intraperitoneally (i.p.) or subperitoneally (s.p.) were treated with this monoclonal antibody (mAb), 19F8, inoculated twice a week in doses of 100 µg. Parallel control groups received analogous inoculations of an isotype-matched monoclonal antibody. A significant growth inhibitory effect was observed with 19F8. In 5/10 rats isografted i.p., tumour outgrowth was completely inhibited and in the other 5 rats the outgrowth was delayed compared to the 10 rats in the control group, which all developed tumours. All rats of the control group developed large volumes of ascites, whereas the 5 rats in the therapy group with eventual tumour outgrowth had little or no ascites. In two experiments with rats carrying subperitoneal isografts and treated with the 19F8 mAb, tumour grew out in 4/5 and 5/10 rats, though growth was delayed compared to the control groups, in which 5/5 and 9/9 rats developed tumours. The tumours grew significantly more slowly in the therapy groups compared to the controls. All rats that developed tumours in the therapy groups showed an anti-idiotypic response against mAb 19F8. The single tumour-free rat in the first experiment and 1/5 tumour-free rats in the other showed no such response. The draining lymph node cells from the tumour-free animals showed a specific proliferative response to yolk sac tumour F56 cells in a mixed lymphocyte tumour cell culture (MLTC), and the MLTC-induced cells were cytotoxic to F56 but not to the natural-killer-sensitive rat T cell lymphoma G1—Tc1. The cytotoxic cell population was more than 90% CD4⁺. It is concluded that the two test systems for identification of the epitope of p15E detected by mAb 19F8 correlated well in detection of the epitope in the cells (immunohistochemistry) and at the cell surface (ADCC). It is also concluded that mAb 19F8 has a growth-inhibitory effect on yolk sac tumour F56 and that, as a result of the treatment, T cells with specificity for F56 are appearing in draining lymph nodes of tumour-free animals.     

A sensitive method to introduce membrane-bound proteins into recipient cells based on affinity enrichment of lipid vesicles to the recipient cell prior to fusion

A sensitive analytical procedure for studying membrane-bound structures has been developed. Membrane glycoproteins inserted into liposomes were transferred to recipient cells by use of a lectin, concanavalin A, bound to the cells as a bridge to generate proximity between the recipient cell and the glycoprotein-containing liposome, prior to exposure to the fusing agent, poly(ethylene glycol). Partially purified histocompatibility antigen from rats was introduced into the membrane of human lymphocytes. After treating the cells with poly(ethylene glycol) under fusion conditions, some of the antigen present in the preparation could not be eluted with alpha-methyl mannoside and EDTA, indicating that incorporation in the cell membrane had taken place. This antigen remained exposed on the lymphocyte surface for approximately 1 h as demonstrated by sensitivity of the lymphocytes to the lytic effect of an antiserum to the histocompatibility antigen in the presence of complement. Some of the lectin molecules seemed to be internalized in the cells but no induction of cell mitosis was observed. The described method gives an opportunity to work with small amounts of membrane proteins inserted into liposomes, introducing them into recipient cells for analysis of their biological activities.     

Lactate dehydrogenase in developing rat oral epithelium

Freeze-dried sagittal, whole-body sections of 10-day-old rats were incubated for lactic dehydrogenase (LDH) using different media in the presence of the inhibitors urea and fluoropyruvate. Phenazine methosulfate (PMS) and menadione, which are regularly used in current histochemical media and are believed to promote the demonstration of LDH activity, were also added and shown to be insufficient for the demonstration of total LDH activity, and PMS even seemed to have an inhibitory effect on LDH activity in oral epithelium. However, cumulated data from the different incubations show that the oral epithelium of developing rats may contain two different types of LDH, one in the basal cells with possibly aerobic characteristics, and another in the spinosum/granulosum cells with anaerobic characteristics.     

Degradation of heparin in mouse mastocytoma tissue

1. Heparin was prepared from mouse mastocytoma tissue by mild procedures, including extraction of mast-cell granules with 2m-potassium chloride, precipitation of the extracted polysaccharide with cetylpyridinium chloride from 0.8m-potassium chloride and finally digestion of the isolated material with testicular hyaluronidase. The resulting product (fraction GE(H)) represented approx. 40% of the total heparin content of the tissue. 2. Fraction GE(H) was fractionated by gel chromatography on Sepharose 4B into three subfractions, with average molecular weights ( M(w)) of approx. 60000-70000 (highly polydisperse material), 26000 and 9000 respectively. Treatment of each of the subfractions with alkali or with papain did not affect their behaviour on gel chromatography. Amino acid and neutral sugar analyses indicated that the two low-molecular-weight fractions consisted largely of single polysaccharide chains lacking the carbohydrate-protein linkage region. It was suggested that these heparin molecules had been degraded by an endopolysaccharidase. 3. Pulse labelling in vivo of mastocytoma heparin with [(35)S]sulphate showed initial labelling of large molecules followed by a progressive shift of radioactivity toward fractions of lower molecular weight. Further, heparin-depolymerizing activity was demonstrated by incubating (35)S-labelled heparin in vitro with a mastocytoma 10000g-supernatant fraction. Appreciable degradation of the polysaccharide occurred, as demonstrated by gel chromatography. In contrast, no depolymerization was observed on subjecting (14)C-labelled chondroitin sulphate to the same procedure.     

Cytosolic free calcium elevation mediates the phagosome-lysosome fusion during phagocytosis in human neutrophils

Cytosolic free calcium ([Ca2+]i) and fusion of secondary granules with the phagosomal membrane (phagosome-lysosome fusion, P-L fusion) were assessed in single adherent human neutrophils during phagocytosis of C3bi-opsonized yeast particles. Neutrophils were loaded with the fluorescent dye fura2/AM and [Ca2+]i was assessed by dual excitation microfluorimetry. Discharge of lactoferrin, a secondary granule marker into the phagosome was verified by immunostaining using standard epifluorescence, confocal laser scanning and electron microscopy. In Ca2(+)-containing medium, upon contact with a yeast particle, a rapid rise in [Ca2+]i was observed, followed by one or more Ca2+ peaks (maximal value 1,586 nM and median duration 145 s): P-L fusion was detected in 80% of the cells after 5-10 min. In Ca2(+)-free medium the amplitude, frequency and duration of the [Ca2+]i transients were decreased (maximal value 368 nM, mostly one single Ca2+ peak and median duration 75 s): P-L fusion was decreased to 52%. Increasing the cytosolic Ca2+ buffering capacity by loading the cells with MAPT/AM led to a dose-dependent inhibition both of [Ca2+]i elevations and P-L fusion. Under conditions where basal [Ca2+]i was reduced to less than 20 nM and intracellular Ca2+ stores were depleted, P-L fusion was drastically inhibited while the cells ingested yeast particles normally. P-L fusion could be restored in Ca2(+)-buffered cells containing ingested particles by elevating [Ca2+]i with the Ca2(+)-ionophore ionomycin. The present findings directly indicate that although the ingestion step of phagocytosis is a Ca2(+)-independent event, [Ca2+]i transients triggered upon contact with opsonized particles are necessary to control the subsequent fusion of secondary granules with the phagosomal membrane.