Genetic and molecular mapping of the pma1 mutation conferring vanadate resistance to the plasma membrane ATPase from Saccharomyces cerevisiae

Summary In the yeast Saccharomyces cerevisiae, the pma1 mutations confers vanadate-resistance to H⁺-ATPase activity when measured in isolated plasma membranes. In vivo, the growth of pma1 mutants is resistant to Dio-9, ethidium bromide and guanidine derivatives. This phenotype was used to man the pma1 mutation adjacent to LEU1 gene on chromosome VII. From a cosmid library of a wild-type Saccharomyces cerevisiae genome, a large 30 kb DNA fragment was isolated by complementation of a leu1-pma1 double mutant. A 5 kb HindIII fragment was subcloned and it restored both Leu⁺ and Pma⁺ phenotypes after integrative transformation. The restriction map of the 5 kb HindIII fragment and Southern blot analysis reveal that the cloned fragment contains the entire structural gene for the plasma membrane ATPase and the 5 end of the adjacent LEU1 gene. The pma1 mutation conferring vanadate-resistance is thus located in the structural gene for the plasma membrane ATPase.Publication n^o 2456 from the Biology Directorate of the Commission of European Communities     

Plant-water relations in wheat as influenced by root pruning

Summary A greenhouse study in which 24, 54 and 71 per cent roots of wheat (Triticum aestivum L.) were pruned on the 73rd day from the date of planting (anthesis stage) showed that during a 7-day period following root pruning, total transpiration and leaf water potential were significantly lower (P=0.05) and the stomatal resistance was significantly higher (P=0.05) where 54 and 71 per cent roots were pruned, as compared to no root pruning or 24 per cent root pruning. The leaf relative water content, however, showed no significant differences. Thus about one-fourth root sytem could be reduced without adversely affecting the plant-water status.     

Isolation of tomato cell lines with altered response to Fusarium cell wall components

Summary To obtain Tomato cell lines with an altered capacity to respond to heat-released cell wall components (elicitor) of a tomato pathogen (Fusarium oxysporum f. sp. lycopersici), positive and negative selection experiments, using BUdR enrichment techniques, were carried out on suspension cultures of the susceptible, low phytoalexin producer cultivar Red River. Both high and low phytoalexin producing clones were isolated. Further tests demonstrated that not all phytoalexin-producing clones were more susceptible to the elicitor toxic effect, and that they were altered also in the speed of response to fungal cell wall components. Cells selected with Fusarium elicitor showed the same behaviour when challenged by Phytophthora infestans elicitor, thus suggesting in this case lack of specificity. The results are finally discussed with a view to using the technique both as a tool to study the genetics and physiology of hostparasite interactions and as a possible new method for the selection of pathogen resistant genotypes.Paper no. 1224 IPRA-CNR; research supported by an EEC-BAP contract     

Inheritance of resistance to potato y viruses in Phaseolus vulgaris L

Summary Resistance to watermelon mosaic virus-2 in Phaseolus vulgaris L. is conferred by two distinct dominant alleles at independent loci. Based on segregation data one locus is designated Wmv, the other, Hsw. The dominant allele Wmv from cv. Great Northern 1140 prevents systemic spread of the virus but viral replication occurs in inoculated tissue. In contrast, Hsw confers both local and systemic resistance to WMV-2 below 30C. At higher temperatures, plants that carry this allele in the absence of modifying or epistatic factors develop systemic veinal necrosis upon inoculation with the virus that results in rapid death. Patho-type specificity has not been demonstrated for either allele; both factors confer resistance to every isolate tested. A temperature-sensitive shift in epistasis is apparent between dominant alleles at these loci. Because Hsw is very tightly linked if not identical to the following genes for hypersensitivity to potyviruses I, (bean common mosaic virus), Bcm, (blackeye cowpea mosaic virus), Cam, (cowpea aphid-borne mosaic virus) and Hss (soybean mosaic virus), parental, reciprocal dihybrid F₁ populations, and selected F₃ families were inoculated with each of these viruses and held at 35 C. F₁ populations developed vascular necrosis completely or primarily limited to inoculated tissue, while F₃ families from WMV-2-susceptible segregates were uniformly susceptible to these viruses. The relationship between Hsw, Wmv and other genes for potyvirus resistance suggest patterns in the evolution of resistance and viral pathogenicity. Characterization of the resistance spectrum associated with each factor provides an additional criterion to distinguish genes for plant virus resistance.     

Succinate metabolism related to lipid synthesis in the housefly Musca domestica L

The metabolism of succinate was examined in the housefly Musca domestica L. The labeled carbons from [2,3-¹⁴C]succinate were readily incorporated into cuticular hydrocarbon and internal lipid, whereas radioactivity from [1,4-¹⁴C]succinate was not incorporated into either fraction. Examination of the incorporation of [2,3-¹⁴C]succinate, [1-¹⁴C]acetate, and [U-¹⁴C]proline into hydrocarbon by radio-gas-liquid chromatography showed that each substrate gave a similar labeling pattern, which suggested that succinate and proline were converted to acetyl-CoA prior to incorporation into hydrocarbons. Carbon-13 nuclear magnetic resonance showed that the labeled carbons from [2,3-¹³C]succinate enriched carbons 1, 2, and 3 of hydrocarbons with carbon-carbon coupling showing that carbons 2 and 3 of succinate were incorporated as an intact unit. Radio-high-performance liquid chromatographic analysis of [2,3-¹⁴C]succinate metabolism by mitochondrial preparations showed that in addition to labeling fumarate, malate, and citrate, considerable radioactivity was also present in the acetate fraction. The data show that succinate was not converted to methylmalonate and did not label hydrocarbon via a methylmalonyl derivative. Malic enzyme was assayed in sonicated mitochondria prepared from the abdomens and thoraces of 1- and 4-day-old insects; higher activity was obtained with NAD⁺ in mitochondria prepared from thoraces, whereas NADP⁺ gave higher activity with abdomen preparations. These data document the metabolism of succinate to acetyl-CoA and not to a methylmalonyl unit prior to incorporation into lipid in the housefly and establish the role of the malic enzyme in this process.     

Effects of starvation,chilling, and injury on endocrine gland function in Galleria mellonella

Starvation, chilling, and injury of last instar Galleria mellonella larvae typically elicit extra larval molts or a delay in pupation. The primary sites of action and the nature of the signals by which these treatments affect development are not known. However, since the connections of the brain to the nerve cord are crucial for the effects of starvation and chilling, these signals apparently affect the brain-centered program of developmental regulation via the nerve cord. Chilling, and occasionally starvation, cause extra larval molts in last instar larvae treated prior to the nervous inhibition of their corpora allata; release of a cerebral allatotropin, which stimulates the production of juvenile hormone, appears to be involved in this effect. After this time, a delay in pupation is the principal effect of starvation and chilling, and is apparently due to a temporal inhibition of the release of the prothoracicotropic hormone. Chilling also appears to inhibit unstimulated ecdysteroid production by the prothoracic glands. The effect of injury is not mediated by the nerve cord, but appears to involve an inhibitory humoral factor that affects either the brain or the prothoracic glands themselves. Injury also stimulates juvenile hormone production, an effect which is enhanced when the brain is separated from the nerve cord and which is evidenced by a delay of ecdysis and the occasional retention of some larval features in the ecdysed insects. None of the effects of these various treatments on the brain and the endocrine glands persist when the brains or glands are implanted into untreated hosts.     

Variation in feeding behaviour of Nilaparvata lugens on resistant and susceptible rice varieties

The feeding behaviour of Nilaparavata lugens was monitored on three rice varieties showing different levels of resistance in the Philippines, using a video-assisted observation method. N. lugens made more frequent, shorter probes on the moderately resistant IR46 and resistant IR62 rice varieties than on the susceptible IR22. Honeydew production was significantly lower on the resistant varieties though insect weight gains in 24 h were similar on IR46 and IR22, both being significantly greater than on the highly resistant variety.Population development, growth index and damage ratings were low on IR62 indicating antibiosis and/or non preference. When IR46 plants were infested as seedlings population increase, growth index and damage ratings were similar to those on the susceptible IR22. When infested at a later stage of plant growth the damage rating showed a moderate level of resistance though some population development was maintained, indicating antibiosis and tolerance. N. lugens started probing less frequently after surface exploration on both resistant varieties than on IR22 suggesting the presence of a resistance factor associated with the surface waxes of these varieties.

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Résumé Le comportement alimentaire de Nilaparvata lugens sur variétés de riz, sensible (IR22), partiellement résistante (IR46) et fortement résistante (IR62), a été contrôlé avec une méthode associant la vidéo à l'observation. N. lugens faisait des piqûres plus fréquentes et plus brèves sur IR46 et IR62, que sur la variété sensible. La production de miellat était significativement plus faible sur les variétés résistantes, bien que les gains de poids des insectes aient été les mêmes en 24 h sur IR46 et IR22, les deux étant significativement supérieurs à celui sur IR62.La croissance de la population, l'indice de croissance et le taux de dégâts étaient tous plus faibles sur IR62, ce qui révèle une antibiose et/ou une absence de préférence. Quand la contamination des IR46 a au lieu au stade semis, la croissance de population, l'indice de croissance et le taux de dégâts étaient semblables à ceux de la variété sensible IR22. Quand la contamination avait lieu à un stade ultérieur, le laux de dégâts révélait un niveau modéré de résistance bien qu'une certaine croissance de population se soit maintenue, ce qui révèle antibiose et tolérance.Après exploration de la surface des feuilles des deux variétés résistantes, N. lugens sondait moins fréquemment que sur IR22, ce qui laisse présumer un facteur de résistance associé aux cires superficielles de ces variétés.

     

Further investigation of the primary mechanism of benzimidazole resistance in Haemonchus contortus

The binding of the [³H] benzimidazole carbamates (BZCs)—albendazole (ABZ), oxibendazole (OBZ), parbendazole (PBZ), mebendazole (MBZ), fenbendazole (FBZ) and oxfendazole (OFZ)—to tubulin from three ecologically-related isolates of adult Haemonchus contortus has been examined. The extent of binding of each BZC was inversely proportional to the known resistance status of the isolate. Biochemically, the change in the formation of the BZC-tubulin complex was due to a reduction in the amount of drug bound to resistant tubulin, with no significant change in the association constant of the complex. The resistance factors derived from the binding data support the hypothesis that the complex is ligand-dependent, with the aryl-substituted BZCs—MBZ, OFZ and FBZ—demonstrating lower resistance factors than those of the alkyl-substituted BZCs—ABZ, OBZ and PBZ. Examination of the slope derived from plots of binding against protein concentration demonstrated that the failure of resistant or partially resistant isolates to bind was due to either a decrease in the number of binding sites or, more likely, to reduced stability of the BZC-tubulin complex rendering it unstable to charcoal extraction.     

Ribosomal protein S12 as a site for streptomycin resistance in Nicotiana chloroplasts

Summary A streptomycin resistant Nicotiana plastome mutant, X/str ^R6, was subjected to molecular analysis. In this mutant, a single nucleotide transition, C » T, in the chloroplast gene for ribosomal protein S12 alters codon 90 from proline to serine while the nucleotide sequence of the chloroplast 16 S rRNA gene is identical to that of the wild type. Mutant X/str ^R6 thus differs from several previously reported streptomycin resistant chloroplast mutants which are altered in the gene for 16 S rRNA.