Ambient conditions of elevated temperature and CO2 levels are detrimental to the probabilities of transmission by insects of a Potato virus Y isolate and to its simulated prevalence in the environment.

Conditions of elevated temperature and CO2 levels [30 °C and 970 parts-per-million (ppm), respectively] reduced the systemic titers of a potato virus Y (PVY) isolate in Nicotiana benthamiana plants, relative to standard conditions (25 °C, ~405 ppm CO2). Under controlled conditions we studied how these growing environments affected the transmission of infection by aphids. Probabilities of transmission of infection by insects that fed on infected donor plants kept at either standard conditions, or at 30 °C and 970 ppm CO2 were both determined and found to positively correlate with titers in donor leaves, independently of the ambient conditions in which recipient plantlets would grow. With these data, viral prevalence was simulated under conditions of elevated temperature and CO2 levels and found that for it to remain comparable to that simulated under standard conditions, insect arrivals to recipient plants in the former scenario would have to increase several-fold in their frequency.

A cucumber mosaic virus (CMV) RNA 1 transgene mediates suppression of the homologous viral RNA 1 constitutively and prevents CMV entry into the phloem.

Resistance to Cucumber mosaic virus (CMV) in tobacco lines transformed with CMV RNA 1 is characterized by reduced virus accumulation in the inoculated leaf, with specific suppression of accumulation of the homologous viral RNA 1, and by the absence of systemic infection. We show that the suppression of viral RNA 1 occurs in protoplasts from resistant transgenic plants and therefore is not due to a host response activated by the cell-to-cell spread of virus. In contrast, suppression of Tobacco rattle virus vectors carrying CMV RNA 1 sequences did not occur in protoplasts from resistant plants. Furthermore, steady-state levels of transgene mRNA 1 were higher in resistant than in susceptible lines. Thus, the data indicate that sequence homology is not sufficient to induce suppression. Grafting experiments using transgenic resistant or susceptible rootstocks and scions demonstrated that the resistance mechanism exhibited an additional barrier to phloem entry, preventing CMV from moving a long distance in resistant plants. On the other hand, virus from susceptible rootstocks could systemically infect grafted resistant scions via the phloem. Analysis of viral RNA accumulation in the infected scions showed that the mechanism that suppresses the accumulation of viral RNA 1 at the single-cell level was overcome. The data indicate that this transgene-mediated systemic resistance probably is not based on a posttranscriptional gene-silencing mechanism.

[Anorexia nervosa: a multidisciplinary approach]. / Anorexia nerviosa. Abordaje multidisciplinar.

The childhood-adolescent psychiatrics field has, for various years, been confronted by a very significant increase in cases of nervous anorexia, a serious eating disorder characterized by a noticeable loss of weight. At the bottom of this situation lie complex biological, psychological and social-cultural problems, which demand an interdisciplinary approach to solve them. This article presents the predisposing factors, the initial factors, the factors which maintain this disorder...; what behaviors are considered to be normal; what the physical and psychological manifestations are; as well as what the medical evaluation carried out is ... to finalize with an explanation of the different functions to be performed by each member of a multidisciplinary team.

Tagging potato leafroll virus with the jellyfish green fluorescent protein gene.

A full-length cDNA corresponding to the RNA genome of Potato leafroll virus (PLRV) was modified by inserting cDNA that encoded the jellyfish green fluorescent protein (GFP) into the P5 gene near its 3' end. Nicotiana benthamiana protoplasts electroporated with plasmid DNA containing this cDNA behind the 35S RNA promoter of Cauliflower mosaic virus became infected with the recombinant virus (PLRV-GFP). Up to 5% of transfected protoplasts showed GFP-specific fluorescence. Progeny virus particles were morphologically indistinguishable from those of wild-type PLRV but, unlike PLRV particles, they bound to grids coated with antibodies to GFP. Aphids fed on extracts of these protoplasts transmitted PLRV-GFP to test plants, as shown by specific fluorescence in some vascular tissue and epidermal cells and subsequent systemic infection. In plants agroinfected with PLRV-GFP cDNA in pBIN19, some cells became fluorescent and systemic infections developed. However, after either type of inoculation, fluorescence was mostly restricted to single cells and the only PLRV genome detected in systemically infected tissues lacked some or all of the inserted GFP cDNA, apparently because of naturally occurring deletions. Thus, intact PLRV-GFP was unable to move from cell to cell. Nevertheless, PLRV-GFP has novel potential for exploring the initial stages of PLRV infection.

Characterisation of genetically modified cucumber mosaic virus expressing histidine-tagged 1a and 2a proteins.

Biological active cDNA clones of cucumber mosaic virus (CMV) RNAs 1 and 2 were modified by addition of sequences that encode hexahistidine (His-tag) at the amino- (N-) or carboxy- (C-) terminus of the 1a and 2a proteins. These proteins are essential components of the viral RNA-dependent RNA polymerase (RdRp). In all but one case, addition of the His-tag did not significantly affect the yields of the corresponding viruses and the His-tag-encoding sequences were maintained after mechanical passages. No differences were observed among the in vitro activities of the modified vs. wild-type viral RdRps. Subcellular fractionation showed that 2a protein was found both membrane-associated and in the 30,000 x g soluble fraction. Both termini of the native His-tag 2a protein could bind to a resin containing nickel-nitrilotriacetic acid (Ni(2+)-NTA). Detergent-treated RdRp containing C-terminal His-tagged 1a and 2a proteins was chromatographed on Ni(2+)-NTA resin. The activity of the eluted RdRp was template- dependent, in contrast to pre-chromatography fractions. However, only a small proportion of the viral RdRp as well as numerous host proteins bound to and eluted from the resin under non-denaturing conditions.

The hypersensitive response to cucumber mosaic virus in Chenopodium amaranticolor requires virus movement outside the initially infected cell.

Cucumber mosaic virus (CMV) expressing the green fluorescent protein (GFP), and lacking either the 3a movement protein or the coat protein (CP), failed to induce a hypersensitive response producing local lesions in inoculated leaves of Chenopodium amaranticolor. Cytological analysis showed that both viral-encoded proteins are required for cell-to-cell movement of the virus and the simultaneous appearance of cellular necrosis. In the absence of either or both proteins, infection was confined to single, non-necrotized, epidermal cells. CMV with a mutation in the 3a protein (M8 CMV) could infect tobacco systemically but did not induce necrotic lesions in C. amaranticolor. In this host, the mutated 3a protein was unable to promote viral movement out of the initially infected epidermal cell. Movement-deficient CMV expressing wild-type (WT) 3a protein as a fusion to the GFP, as well as WT CP, also failed to induce necrosis. Finally, single epidermal cells infected with a movement-deficient CMV expressing WT 3a protein, WT CP, and free GFP did not show necrosis. These data indicate that viral movement out of the initially infected epidermal cell, and not the simultaneous expression in this cell of the 3a protein and the CP, is required for the induction of cell death.

Transgenically expressed cucumber mosaic virus RNA 1 simultaneously complements replication of cucumber mosaic virus RNAs 2 and 3 and confers resistance to systemic infection.

Tobacco plants transformed with a cDNA copy of RNA 1 of the Fny strain of cucumber mosaic virus (CMV) promoted the asymptomatic accumulation of inoculated viral RNAs 2 and 3, which could be detected in noninoculated leaves, suggesting that the transgene also permitted viral long-distance movement. Typical symptoms of infection appeared later and correlated with the appearance of viral RNA 1 regenerated from the transgenic mRNA. Although all R0-generation plants were susceptible to Fny-CMV, one line displaying variable susceptibility to the virus in R1-and R2-generations led to selected R3-generation lines with systemic resistance to Fny-CMV. In the inoculated leaves of resistant plants, a dramatic decrease in the accumulation of viral RNA 1 was observed, relative to susceptible plants. No viral RNAs were detected in noninoculated leaves of the resistant plants, but such leaves were susceptible to infection. Furthermore, these leaves could sustain replication of inoculated CMV RNAs 2 and 3, indicating that a complete transgene-silencing had not been induced. Although a transgene-mediated, CMV RNA 1-suppression occurred in the inoculated leaf of resistant plants, the absence of a complete systemically acquired silencing suggests the existence of additional interferences with viral infection that prevented systemic infection by viral RNAs 2 and 3.

Characterization of cucumber mosaic virus. IV. Movement protein and coat protein are both essential for cell-to-cell movement of cucumber mosaic virus.

cDNA clones of cucumber mosaic virus (CMV) RNA 3 were modified to express the jellyfish green fluorescent protein (GFP) in place of the 3a movement protein (MP) or coat protein (CP), as fusions to the N (GFP-3a) or C (3a-GFP) terminus of the MP or from a separate open reading frame as part of tricistronic RNAs 3. CMV RNA transcripts containing the individual modified RNAs 3 were unable to infect either Nicotiana tabacum or Nicotiana benthamiana systemically. Infection, as measured by confocal microscopy of GFP fluorescence, generally was limited to one to three epidermal cells at each inoculation site. Limited cell-to-cell movement, but not systemic movement, could be detected by complementation involving expression of MP and CP from two different RNA 3 constructs, each also expressing GFP. Infection involving RNA 3 expressing the GFP-3a fusion showed bright granules of variable size distributed predominantly and nonuniformly throughout the cytoplasm and, to a lesser extent, associated with the cell wall in single fluorescent cells, while infections expressing the 3a-GFP fusion showed bright, punctate fluorescence associated only with the cell wall. Infected cells expressing either 3a-GFP or free GFP showed a halo of less bright, fluorescent, neighboring cells, indicating limited movement of GFP. The initially infected cells also allowed movement of 10-kDa fluorescent dextran to the neighboring halo cells, while infection did not spread, suggesting different requirements for movement of either MP or dextran versus RNA.

Prediction of depressive relapse in remitted bipolar patients using corticotrophin-releasing hormone challenge test.

Abnormalities in corticotrophin (ACTH) and cortisol levels before and after corticotrophin-releasing hormone (CRH) stimulation have been reported in depressed bipolar patients. The ACTH and free cortisol response to the injection of 100 micrograms of synthetic human CRH and plasma cortisol-binding globulin (CBG) levels were measured in 42 lithium-treated patients suffering from RDC bipolar-I disorder in remission, and in 21 age- and sex-matched control subjects. A 1-year follow-up was conducted in order to assess any possible relationship between outcome and the hormonal response. Bipolar patients showed higher baseline and peak ACTH concentrations than controls. A lower net area under the ACTH concentration curve after CRH stimulation predicted depressive relapse within 6 months by multiple regression analysis. The CRH challenge test could be a potentially good predictor of depressive relapse in remitted bipolar patients.

Transmission by aphids of a naturally non-transmissible plum pox virus isolate with the aid of potato virus Y helper component.

Two Spanish plum pox virus (PPV) isolates, 5.15 and 3.3, were used in transmission experiments involving the aphid vector Myzus persicae, with woody and herbaceous host plants. These isolates differ in the size of their coat protein (CP) and sequence analysis revealed that isolate 3.3 has a 15 amino acid deletion near the N terminus of the CP, affecting the same positions as in a previously reported non-aphid-transmissible PPV isolate from Germany. Aphid transmission experiments showed that isolate 5.15 was transmitted from infected plants whereas isolate 3.3 was not. In contrast, both isolates were readily aphid-transmitted when acquired through artificial membranes from purified virus preparations supplemented with purified helper component (HC) obtained from potato virus Y-infected plants. This indicates that non-transmissibility of isolate 3.3 may be due to a defect in the HC rather than in the CP.

Mini-pill in lactating women.

A non-comparative study of the progestogen-only oral contraceptive, norgestrel 0.075 mg, in breast-feeding women was conducted at the Centro de Investigaciones Regionales, Merida, Yucatan, Mexico. The study was designed to evaluate the overall acceptability and contraceptive efficacy of norgestrel in breast-feeding women. This report includes a survey of 200 women, all of whom were less than 26 weeks postpartum at admission; 113 were interval patients and 87 were postpartum. Follow-up visits were scheduled at 2, 6 and 12 months after admission. Overall, women experienced an increase in intermenstrual bleeding, amenorrhea, vaginal discharge and breast discomfort. The discontinuation rate at 12 months was 32.5 and the corresponding lost to follow-up rate was 22.5; this is a measure of acceptability. The 12-month life-table rate for pregnancy was 3.4 with a standard error of 2.0. Three women discontinued use of the mini-pill due to accidental pregnancy. One pregnancy was attributed to user failure and the woman conceived 9 months after entering into the study; the other two were attributed to method failure, one woman conceived 3 months after admission and the other conceived 6 months after admission.

Inhibition of adrenomedullary catecholamine release by propranolol isomers and clonidine involving mechanisms unrelated to adrenoceptors.

1 Transmural electrical stimulation (10 Hz, 40 V, 1 ms for 60s) increased total catecholamine secretion from perfused cat adrenal glands; this response was enhanced by neostigmine and inhibited by mecamylamine, suggesting that release of acetylcholine from splanchnic nerve terminals was stimulating nicotinic receptors and enhancing catecholamine secretion. 2 Isoprenaline, (-)-propranolol and (+)-propranolol (10(-7)-10(-5)M) inhibited the electrically-evoked secretory response by 40-70%; similar reductions were obtained with clonidine and yohimbine. Neither, (+)-propranolol nor (-)-propranolol inhibited K-evoked secretion from cat adrenals; in contrast, nimodipine potently inhibited it (IC50 = 24 nM). 3 Either, racemic propranolol or the (+)- or (-)-isomers (1-10 microM) equally inhibited [3H]-noradrenaline release evoked by nicotine or acetylcholine from cultured bovine adrenal chromaffin cells; clonidine (10 microM) inhibited secretion by 50% and yohimbine or isoprenaline did not affect it. 4 The results indicate that adrenomedullary catecholamine release evoked by splanchnic nerve stimulation is not modulated by alpha- or beta-adrenoceptors and suggest that propranolol may inhibit secretion by blocking ion fluxes through the acetylcholine receptor ionophore. Clonidine may inhibit secretion by this same mechanism, and/or by interfering with some intracellular event in the secretory mechanism.

Effect of oral contraceptive use on the erythrocytic glutathione reductase and aspartate aminotransferase activities in women with or without clinical signs of vitamin deficiency.

The effect of the chronic use of combined oral contraceptives (OCs) on the "activity coefficients" (alpha = coenzyme-stimulated activity/basal activity) of erythrocytic glutathione reductase and aspartate aminotransferase was studied in 2 groups of 90 female volunteers each; 1 of the groups, from the state of Yucatan in southeast Mexico, presented clinical lesions of vitamin deficiency, while the other group, from Mexico City, did not have any clinical evidence of vitamin deficiency. One half of the women (45) in each group were chronic OC users and the other half were not. The results were analyzed comparing OC users with non-users in each location. For both glutathione reductase and aspartate aminotransferase, the Mexico City OC users had significantly higher (p 0.001) alpha values than nonusers, while in the Yucatan women, the alpha values were similarly high independent of OC use.

Effects of lactation on hormonal levels in rural women.

Luteinizing hormone, follicle-stimulating hormone and prolactin levels were compared in ten rural and ten urban women, all of whom were experiencing lactational amenorrhea. The values of luteinizing hormone and follicle-stimulating hormone were similar in both groups. The prolactin levels were significantly elevated in the rural group.

PIP: Luteininzing hormone (LH), follicle-stimulating hormone (FSH) and prolactin levels were compared in 10 rural and 10 urban women in Mexico who were experiencing lactational amenorrhea in an effort to define any differences for these 2 groups. All the women had breast fed their babies without supplementation, and none of the women had menstruated or had taken oral contraceptives. Another 20 women who attended the Clinic for Family Planning and who did not take OCs served as controls. All the women had an extended postpartum period that varied from 9-12 months. The levels of LH in the urban group fluctuated between 1.0 and 12.0 Imu-ml, with an average of 5.1 Imu-ml. The FSH levels among the urban group ranged from 3.0 to 15.0 with an average of 7.3 Imu-ml, while the average prolactin level was 20.4 ng-ml. Duration of lactation varied from 7 to 16 months for the rural women. Their LH and FSH levels were similar to those of the urban group, but their prolactin levels were more elevated than those found in the urban group. The average prolactin level was 38.1 mg.ml, and this elevation was statistically significant. The LH values among the 20 controls fluctuated between 3.0 and 20.0 Imu-ml, with an average of 9.2 ImU-ml. The FSH levels for the control group ranged from 3.4 to 17.0 ImU-ml, with an average of 8.1 ImU-ml. Prolactin levels varied between 10.1 and 17.0 ng-ml, with an average of 14.5 ng-ml.