An analysis of the durability of resistance to plant viruses

ABSTRACT Genetic resistance often fails because a resistance-breaking (RB) pathogen genotype increases in frequency. On the basis of an analysis of cellular plant pathogens, it was recently proposed that the evolutionary potential of a pathogen is a major determinant of the durability of resistance. We test this hypothesis for plant viruses, which differ substantially from cellular pathogens in the nature, size, and expression of their genomes. Our analysis was based on 29 plant virus species that provide a good representation of the genetic and biological diversity of plant viruses. These 29 viruses were involved in 35 pathosystems, and 50 resistance factors deployed against them were analyzed. Resistance was found to be durable more often than not, in contrast with resistance to cellular plant pathogens. In a third of the analyzed pathosystems RB strains have not been reported, and in another third RB strains have been reported but have not become prevalent in the virus population. The evolutionary potential of the viruses in the 35 pathosystems was evaluated with a compound risk index based on three evolutionary factors: the population of the pathogen, the degree of recombination, and the amount of gene and genotype flow. Our analysis indicates that evolutionary potential may be an important determinant of the durability of resistance against plant viruses.     

The effects of repeated applications of the molluscicide metaldehyde and the biocontrol nematode Phasmarhabditis hermaphrodita on molluscs, earthworms, nematodes, acarids and collembolans: a two-year study in north-west Spain

Over two years, six consecutive field experiments were done in which the chemical molluscicide metaldehyde and the nematode biocontrol agent Phasmarhabditis hermaphrodita (Schneider) were applied at the standard field rates to replicated mini-plots successively planted with lettuce, Brussels sprouts, leaf beet and cabbage, to compare the effectiveness of different treatments in reducing slug damage to the crops. Soil samples from each plot were taken prior to the start of the experiments, and then monthly, to assess the populations of slugs, snails, earthworms, nematodes, acarids and collembolans. The experiments were done on the same site and each plot received the same treatment in the six experiments. The six treatments were: (1) untreated controls, (2) metaldehyde pellets, (3 and 4) nematodes applied to the planted area 3 days prior to planting without or with previous application of cow manure slurry, (5) nematodes applied to the area surrounding the planted area 3 days prior to planting, and (6) nematodes applied to the planted area once (only in the first of the six consecutive experiments). Only the metaldehyde treatment and the nematodes applied to the planted area at the beginning of each experiment without previous application of manure significantly reduced slug damage to the plants, and only metaldehyde reduced the number of slugs contaminating the harvested plants. The numbers of slugs, snails and earthworms in soil samples were compared among the six treatments tested: with respect to the untreated controls, the numbers of Deroceras reticulatum (Müller) were significantly affected only in the metaldehyde plots, and the numbers of Arion ater L only in the plots treated with nematodes applied to the planted area 3 days prior to planting without previous application of manure; numbers of snails (Ponentina ponentina (Morelet) and Oxychilus helveticus (Blum)) were not affected by the treatment. The total numbers of all earthworm species and of Lumbricus spp were unaffected by the treatment, but Dendrobaena spp increased significantly in the plots treated with manure. The numbers of nematodes, acarids and collembolans in soil samples were compared between the untreated controls and the treatments with nematodes applied 3 days prior to planting to the planted area or to the surrounding area, without previous application of manure: the treatment had a significant effect on the number of nematodes in soil samples, but acarids and collembolans were unaffected.     

Congenital skeletal abnormalities in a tawny owl chick (Strix aluco)

In this case report, we describe a tawny owl chick (Strix aluco) coming from a Wild Fauna Recovery Center with multiple congenital malformations in the limbs. The animal was unable to fly and showed marked malnutrition and poor general appearance. Physical, radiologic, and anatomic examinations showed osseous malformations including dislocation of radius and carpometacarpus with abnormal nonfunctional fixation of ligamentum propatagialis, absence of most parts of the bones of the manus in both wings, and twisted nonfused left tarsometatarsus with marked griphosis of digits. Routine toxicologic and pathologic examinations did not reveal a specific etiology.     

Effect of first-pass hepatic metabolism on the disposition of levamisole after intravenous administration in rabbits

OBJECTIVE: To evaluate the contribution of first-pass hepatic metabolism of levamisole on levamisole disposition in rabbits. ANIMALS: 30 male New Zealand White rabbits. PROCEDURES: Rabbits were randomly placed into 2 groups. Rabbits in the first group received levamisole via the marginal ear vein at the following 3 doses: 12.5, 16, and 20 mg/kg (5 rabbits for each dose). Rabbits of the second group received levamisole via the jejunal vein at the same doses (5 rabbits each). During the following 240-minute period, plasma samples were obtained and quantified for levamisole concentrations by reversed-phase high-performance liquid chromatography. RESULTS: No significant differences were found between pharmacokinetic parameters calculated by compartmental or noncompartmental analysis. Mean hepatic extraction ratio ranged from -0.044 to 0.017 and from 0.020 to 0.081 when area under the plasma concentration-time curve values were obtained after compartmental or noncompartmental analysis, respectively. After compartmental analysis, plasma concentration decreased bi-exponentially. Mean pharmacokinetic parameter values were as follows for each dose (12.5, 16, and 20 mg/kg, respectively): after levamisole administration via the marginal ear vein, volume of distribution at steady state (Vss) = 4.26, 4.33, and 3.20 L/kg; total body clearance (CI) = 49.04, 43.77, and 39.26 mL/kg x min; and half-life associated with beta-phase (t1/2beta) = 77.93, 85.39, and 69.79 minutes. After levamisole administration via the jejunal vein, Vss = 4.38, 2.85, and 2.97 L/kg; CI = 48.14, 42.40, and 39.69 mL/kg x min; and t1/2b = 101.9, 76.71, and 76.13 minutes. CONCLUSIONS: Levamisole has a low degree of hepatic extraction in rabbits.     

Two subpopulations of Colletotrichum acutatum are responsible for anthracnose in strawberry and leatherleaf fern in Costa Rica

Strawberry, Fragaria × ananassa, and leatherleaf fern, Rumohra adiantiformis, are two important crops in Costa Rica. One of the most severe diseases affecting these crops is anthracnose, caused by members of the fungal genus, Colletotrichum (teleomorph; Glomerella). Eighty single-spore isolates from strawberry and leatherleaf fern were identified as Colletotrichum acutatum by species-specific PCR, and were further characterised by Universally Primed PCR (UP-PCR) fingerprinting analysis, and sequence analysis of the ribosomal internal transcribed spacer (ITS) region. Morphological differences, genotypic variation revealed by UP-PCR fingerprinting analysis, and a single sequence polymorphism within the ITS2 region were found between the isolates from strawberry and leatherleaf fern, respectively. The UPGMA cluster analysis of the fingerprints clearly separated the isolates derived from strawberry and leatherleaf fern into two different clusters. Pathogenicity assays on detached strawberry fruits confirmed the apparent difference between the two groups of isolates. It is therefore suggested that the pathogens responsible for strawberry anthracnose fruit rot and leatherleaf fern anthracnose in Costa Rica, belong to two distinct subpopulations of C. acutatum.     

Hemocyanin lipid uptake in Polybetes pythagoricus is altered by fenitrothion

The spider very high density lipoprotein (VHDL), which contains hemocyanin as the major apoprotein, transports most of the circulating lipids. In this work, the effect of the pesticide fenitrothion (FS) on the ability of VHDL-apoproteins to uptake different lipids was investigated. For this, VHDL was delipidated using Triton X-100 and recombined with different radiolabeled lipids in the presence or the absence of FS. The oligomeric structural integrity was maintained after delipidation as shown by non-denaturating PAGE. In the presence of the insecticide, palmitic acid uptake decreased by 28.2 and 62.4% after treating the apolipoprotein with 10 and 20 ppm FS, respectively. Decreases in the uptake of cholesterol, triolein, and phosphatidylcholine caused by FS were 29, 23, and 31% using 10 ppm, and 40, 44, and 29% using 20 ppm FS, respectively. Fluorescence measurements with the hydrophobic probes diphenylhexatriene (DPH) and diphenylhexatrienyl-propionic acid (DPH-PA) indicate that FS induces a red shift, decreases the intensity and increases the anisotropy of the emission of these probes in the VHDL. These results indicate that insecticide binding to the lipoprotein enhances the environment polarity and restricts the mobility of these probes at their binding site. These changes at the hydrophobic VHDL binding sites could lead to the decreased affinity for lipids and hydrophobic ligands. It is inferred that FS could alter the normal lipid exchange between this lipoprotein and tissues.     

Evaluation of cell-surface IgE receptors on the canine mastocytoma cell line C2 maintained in continuous culture

OBJECTIVE: To assess expression and function of cell-surface IgE receptors on the canine mastocytoma cell line C2 maintained in continuous culture. SAMPLE POPULATION: C2 cells maintained in medium lacking IgE for up to 10 passages before being stored at -80 C. PROCEDURE: Cells were thawed, cultured in medium without IgE for 1 to 3 passages, sensitized for 7 days with IgE-rich serum from dogs naturally sensitized to Ascaris suum, and stimulated with antigen Asc S1 from A suum, goat polyclonal anti-canine IgE, or calcium ionophore and phorbol myristate acetate (PMA). Percentage of intracellular beta-hexosaminidase released and concentration of tumor necrosis factor-alpha (TNF-alpha) synthesized after stimulation were determined. Expression of cell-surface IgE receptors was assessed by use of a flow cytometry. RESULTS: Immunologic stimulation (antigen or anti-IgE) failed to induce release or synthesis of detectable amounts of beta-hexosaminidase or TNF-alpha. In contrast, nonimmunologic stimulation (calcium ionophore and PMA) led to release of beta-hexosaminidase (mean +/- SEM maximum release, 23.95+/-1.96%) and synthesis of TNF-alpha (maximum concentration, 34.34+/-2.34 pg/10(6) cells). As revealed by use of flow cytometry, C2 cells expressed surface IgE receptors that bound canine IgE in vitro. CONCLUSIONS: Continuous culture of the canine mastocytoma cell line C2 in medium without exogenous IgE or cytokines and other growth factors resulted in cell-surface expression of nonfunctional IgE receptors. However, C2 cells maintained in continuous culture may still be a useful tool for the evaluation of mast cell responses to nonimmunologic stimulation and IgE receptor differentiation and maturity.     

Phylogeography of New World isolates of Anaplasma marginale based on major surface protein sequences

Gene and protein sequences of major surface proteins (MSP) 1a and 4 of Anaplasma marginale (Rickettsiales: Anaplasmataceae) were used to infer phylogenetic relationships between New World isolates from Argentina, Brazil, Mexico and the United States. Seventeen isolates of A. marginale plus two outgroup taxa (A. centrale and A. ovis) were used for maximum-parsimony analysis of MSP4, while 20 isolates were used for phylogenetic analysis of MSP1a. msp4 analysis provided strong bootstrap support for a Latin American clade and, within this clade, support was detected for Mexican and South American clades. Isolates of A. marginale from the United States also grouped into two clades from the southern (isolates from Florida, Mississippi, and Virginia) and west-central (isolates from California, Idaho, Illinois, Oklahoma, and Texas) states. Although little phylogeographic resolution was detected within these higher clades, msp4 sequences appear to be a good genetic marker for inferring phylogeographic patterns of A. marginale isolates. In contrast to the phylogeographic resolution provided by msp4, MSP1a DNA and protein sequence were quite variable and did not provide phylogeographic resolution. Most variation in MSP1a sequences appeared unique to a given isolate and similar DNA sequence variation in msp1alpha was detected within isolates from Idaho and Florida and from Idaho and Argentina. The results of these studies demonstrated that msp4 provided phylogenetic information on the evolution of A. marginale isolates. In contrast MSP1a sequences appeared to be rapidly evolving and these sequences may provide phylogeographic information only when numerous isolate MSP1a sequences are analyzed from a geographic area.     

Risk factors for Aujeszky's disease in pig herds and detection of field virus antibodies in fattening pigs in the state of Yucatan, Mexico

Two epidemiological studies were conducted from August 1997 to May 1998: a case-control study to identify herd level risk factors for antibodies to Aujeszky's disease virus (ADV) in sows in the state of Yucatan, Mexico and a cross-sectional study to determine the prevalence of antibodies against ADV in fattening pigs. In the case-control study, data on herd management and biosecurity were obtained from all the 27 ADV known field-virus-seropositive farms (cases) and 62 randomly selected seronegative farms (controls) by questionnaire. Breeding animals of these seropositive farms had received a gE-deletion vaccine. In the cross-sectional study, 26 farrow-to-finish farms of the 27 seropositive farms were used and blood samples taken from 60 fattening pigs per herd (15 pigs for each stage of production). Serum samples were analyzed by the screening-ELISA and gE-ELISA tests. In the case-control study, three of the 15 risk factors were significant. Odds ratios for distance to the nearest farm (< or = 2.5km), not sampling for the detection of ADV and herds with origin of breeding animals within the state were 9.5, 18.1 and 8.7. In the cross-sectional study, 11 (42.3%) of the 26 sampled farms were seropositive to vaccine antibodies. None of the piglets were positive to antibodies against field virus risk--suggesting that the strategy of vaccinating only the breeding animals reduced the ADV infection of the piglets.     

Escherichia coli heat-stable enterotoxin b (STb) in vivo internalization within rat intestinal epithelial cells

Heat-stable enterotoxin b (STb) is a low molecular weight toxin known to bind sulfatide, its receptor. The fate of STb bound to rat intestinal epithelium cells was followed using an anti-toxin gold labeled assay and transmission electron microscopy. The data suggest that STb toxin and the fusion protein maltose binding protein (MBP)-STb were internalized whereas its mutant I41 E-M42R with reduced hydrophobicity did not show internalization. There was a significant difference in the mean of gold particles per field between rat intestine incubated with STb or the fusion protein MBP-STb and the negative control consisting of intestine incubated with PBS alone. No subcellular compartment seems to be particularly aimed by the toxin as gold particles were randomly distributed within the cell.