Survival Strategy of Erwinia amylovora against Copper: Induction of the Viable-but-Nonculturable State

Copper compounds, widely used to control plant-pathogenic bacteria, have traditionally been employed against fire blight, caused by Erwinia amylovora. However, recent studies have shown that some phytopathogenic bacteria enter into the viable-but-nonculturable (VBNC) state in the presence of copper. To determine whether copper kills E. amylovora or induces the VBNC state, a mineral medium without copper or supplemented with 0.005, 0.01, or 0.05 mM Cu²⁺ was inoculated with 10⁷ CFU/ml of this bacterium and monitored over 9 months. Total and viable cell counts were determined by epifluorescence microscopy using the LIVE/DEAD kit and by flow cytometry with 5-cyano-2,3-ditolyl tetrazolium chloride and SYTO 13. Culturable cells were counted on King's B nonselective solid medium. Changes in the bacterial morphology in the presence of copper were observed by scanning electron microscopy. E. amylovora entered into the VBNC state at all three copper concentrations assayed, much faster when the copper concentration increased. The addition of different agents which complex copper allowed the resuscitation (restoration of culturability) of copper-induced VBNC cells. Finally, copper-induced VBNC cells were virulent only for the first 5 days, while resuscitated cells always regained their pathogenicity on immature fruits over 9 months. These results have shown, for the first time, the induction of the VBNC state in E. amylovora as a survival strategy against copper.     

Decomposition of overlapping protein complexes: A graph theoretical method for analyzing static and dynamic protein associations

Background  

Most cellular processes are carried out by multi-protein complexes, groups of proteins that bind together to perform a specific task. Some proteins form stable complexes, while other proteins form transient associations and are part of several complexes at different stages of a cellular process. A better understanding of this higher-order organization of proteins into overlapping complexes is an important step towards unveiling functional and evolutionary mechanisms behind biological networks.     

Pithecia pithecia's behavioral response to decreasing fruit abundance

In this article we describe the behavioral responses of a group of white-faced sakis' (Pithecia pithecia) to fruit and water scarcity. Six sakis were observed on Round Island in Guri Lake, Venezuela, between March and May 1996. These months are considered the dry season and the beginning of the wet season. Sakis specialize in eating seeds. During the present study only one tree species, Licania discolor (Chrysobalanaceae), fruited in substantial numbers. Licania seeds accounted for 88% of the time the sakis spent eating fruit in March, 87% in April, and 80% in May. We estimate that the sakis' intake of Licania seeds dropped from 2,573 seeds in the 15-day observation period in March to 956 seeds in the 16-day observation period in May. The sakis not only spent less time eating Licania, they ate the seeds at a much slower rate. The drop in the sakis' feeding rate was probably due to increased local search and inspection times. In response to the scarcity of fruit, the sakis ate more young leaves, insects, and flowers. Feeding bouts became more frequent but shorter. Mean distances between feeding bouts fell significantly and the sakis revisited trees less often.     

Mechanism of constitutive export from the golgi: bulk flow via the formation,protrusion, and en bloc cleavage of large trans-golgi network tubular domains

Transport of constitutive cargo proteins from the Golgi complex to the plasma membrane (PM) is known to be mediated by large tubular-saccular carriers moving along microtubules. However, the process by which these large structures emerge from the trans-Golgi network (TGN) remains unclear. Here, we address the question of the formation of Golgi-to-PM carriers (GPCs) by using a suitable cluster of morphological techniques, providing an integrated view of their dynamics and three-dimensional structure. Our results indicate that exit from the TGN of a constitutive traffic marker, the VSVG protein, occurs by bulk flow and is a three-step process. First, the formation of a tubular-reticular TGN domain (GPC precursor) that includes PM-directed proteins and excludes other cargo and Golgi-resident proteins. Notably, this step does not require membrane fusion. Second, the docking of this preformed domain on microtubules and its kinesin-mediated extrusion. Finally, the detachment of the extruded domain by membrane fission. The formation of GPCs does not involve cargo concentration and is not associated with the presence of known coat proteins on GPC precursors. In summary, export from the Golgi occurs via the formation, protrusion and en bloc cleavage of specialized TGN tubular-saccular domains.     

Large Number Discrimination in Newborn Fish

Quantitative abilities have been reported in a wide range of species, including fish. Recent studies have shown that adult guppies (Poecilia reticulata) can spontaneously select the larger number of conspecifics. In particular the evidence collected in literature suggest the existence of two distinct systems of number representation: a precise system up to 4 units, and an approximate system for larger numbers. Spontaneous numerical abilities, however, seem to be limited to 4 units at birth and it is currently unclear whether or not the large number system is absent during the first days of life. In the present study, we investigated whether newborn guppies can be trained to discriminate between large quantities. Subjects were required to discriminate between groups of dots with a 0.50 ratio (e.g., 7 vs. 14) in order to obtain a food reward. To dissociate the roles of number and continuous quantities that co-vary with numerical information (such as cumulative surface area, space and density), three different experiments were set up: in Exp. 1 number and continuous quantities were simultaneously available. In Exp. 2 we controlled for continuous quantities and only numerical information was available; in Exp. 3 numerical information was made irrelevant and only continuous quantities were available. Subjects successfully solved the tasks in Exp. 1 and 2, providing the first evidence of large number discrimination in newborn fish. No discrimination was found in experiment 3, meaning that number acuity is better than spatial acuity. A comparison with the onset of numerical abilities observed in shoal-choice tests suggests that training procedures can promote the development of numerical abilities in guppies.     

Nitric oxide and cell death in liver cancer cells

Nitric oxide (NO) is a lipophillic, highly diffusible, and short-lived physiological messenger which regulates a variety of physiopathological responses. NO may exert its cellular action through cGMP-dependent and cGMP-independent pathways which includes different postranslational modifications. The effect of NO in cancer depends on the activity and localization of NOS isoforms, concentration and duration of NO exposure, cellular sensitivity, and hypoxia/re-oxygenation process. NO regulates critical factors such as the hypoxia inducible factor-1 (HIF-1) and p53 generally leading to growth arrest, apoptosis or adaptation. NO sensitizes hepatoma cells to chemotherapeutic compounds probably through increased p53 and cell death receptor expressions.     

Shedding of the 67-kD laminin receptor by human cancer cells

The 67-kD laminin receptor (67LR) is a cell membrane-associated molecule exhibiting high affinity for the basement membrane glycoprotein, laminin. While export of the 67LR toward the extracellular matrix has been recently suggested by electron microscopy studies, there is to date no evidence of shedding of the 67LR from cells. Using two monoclonal antibodies directed against the 67LR, we developed a double-determinant radioimmunoassay that demonstrates that the 67LR is released from cancer cells into the culture medium. The shed molecule exhibited the same apparent molecular weight as that of the membrane-associated 67LR, suggesting that no proteolytic cleavage is involved in the process. Furthermore, we demonstrate that the 67LR is not anchored to the membrane through a glycolsyl-phosphatidylinositol bridge. However, the observation that lactose increased the release of 67LR suggests that a lectin-type interaction is involved in the cell membrane association of this laminin binding protein and the cell surface. Interestingly, the released 67LR recovered after HPLC gel filtration was found free as well as associated to high molecular weight complexes. The free 67LR retained its ability to bind to the cell surface. Our study is the first demonstration that the 67LR is effectively shed by cancer cells. The released free 67LR could play an important role in modulating interactions between cancer cells and laminin during tumor invasion and metastasis. © 1996 Wiley-Liss, Inc.