Historical Contingencies Modulate the Adaptability of Rice Yellow Mottle Virus

The rymv1-2 and rymv1-3 alleles of the RYMV1 resistance to Rice yellow mottle virus (RYMV), coded by an eIF(iso)4G1 gene, occur in a few cultivars of the Asiatic (Oryza sativa) and African (O. glaberrima) rice species, respectively. The most salient feature of the resistance breaking (RB) process is the converse genetic barrier to rymv1-2 and rymv1-3 resistance breakdown. This specificity is modulated by the amino acid (glutamic acid vs. threonine) at codon 49 of the Viral Protein genome-linked (VPg), a position which is adjacent to the virulence codons 48 and 52. Isolates with a glutamic acid (E) do not overcome rymv1-3 whereas those with a threonine (T) rarely overcome rymv1-2. We found that isolates with T49 had a strong selective advantage over isolates with E49 in O. glaberrima susceptible cultivars. This explains the fixation of the mutation T49 during RYMV evolution and accounts for the diversifying selection estimated at codon 49. Better adapted to O. glaberrima, isolates with T49 are also more prone than isolates with E49 to fix rymv1-3 RB mutations at codon 52 in resistant O. glaberrima cultivars. However, subsequent genetic constraints impaired the ability of isolates with T49 to fix rymv1-2 RB mutations at codons 48 and 52 in resistant O. sativa cultivars. The origin and role of the amino acid at codon 49 of the VPg exemplifies the importance of historical contingencies in the ability of RYMV to overcome RYMV1 resistance.     

Involvement of the cylindrical inclusion (CI) protein in the overcoming of an eIF4E-mediated resistance against Lettuce mosaic potyvirus

The capacity of Lettuce mosaic virus to overcome the lettuce resistance conferred by the mo1¹ and mo1² alleles of the gene for eukaryotic translation initiation factor 4E (eIF4E) was analysed using reverse genetics. Mutations in the virus genome-linked protein (VPg) allowed mo1¹ only to be overcome, but mutations in the C-terminal portion of the cylindrical inclusion (CI) protein allowed both alleles to be overcome. Site-directed mutagenesis pinpointed a key role of the amino acid at position 621 in the virulence. This is the first example of the involvement of a potyviral CI protein in the breaking of an eIF4E-mediated resistance.     

Identification of a second major resistance gene to Rice yellow mottle virus, RYMV2, in the African cultivated rice species, O. glaberrima

Rice yellow mottle virus (RYMV) is the most damaging rice-infecting virus in Africa. However, few sources of high resistance and only a single major resistance gene, RYMV1, are known to date. We screened a large representative collection of African cultivated rice (Oryza glaberrima) for RYMV resistance. Whereas high resistance is known to be very rare in Asian cultivated rice (Oryza sativa), we identified 29 (8%) highly resistant accessions in O. glaberrima. The MIF4G domain of RYMV1 was sequenced in these accessions. Some accessions possessed the rymv1-3 or rymv1-4 recessive resistance alleles previously described in O. glaberrima Tog5681 and Tog5672, respectively, and a new allele, rymv1-5, was identified, thereby increasing the number of resistance alleles in O. glaberrima to three. In contrast, only a single allele has been reported in O. sativa. Markers specific to the different alleles of the RYMV1 gene were developed for marker-assisted selection of resistant genotypes for disease management. In addition, the presence of the dominant susceptibility allele (Rymv1-1) in 15 resistant accessions suggests that their resistance is under different genetic control. An allelism test involving one of those accessions revealed a second major resistance gene, i.e., RYMV2. The diversity of resistance genes against RYMV in O. glaberrima species is discussed in relation to the diversification of the virus in Africa.     

A BAHD acyltransferase is expressed in the tapetum of Arabidopsis anthers and is involved in the synthesis of hydroxycinnamoyl spermidines

BAHD acyltransferases catalyze the acylation of many plant secondary metabolites. We characterized the function of At2g19070 , a member of the BAHD gene family of Arabidopsis thaliana . The acyltransferase gene was shown to be specifically expressed in anther tapetum cells in the early stages of flower development. The impact of gene repression was studied in RNAi plants and in a knockout (KO) mutant line. Immunoblotting with a specific antiserum raised against the recombinant protein was used to evaluate the accumulation of At2g19070 gene product in flowers of various Arabidopsis genotypes including the KO and RNAi lines, the male sterile mutant ms1 and transformants overexpressing the acyltransferase gene. Metabolic profiling of flower bud tissues from these genetic backgrounds demonstrated a positive correlation between the accumulation of acyltransferase protein and the quantities of metabolites that were putatively identified by tandem mass spectrometry as N ¹, N ⁵, N ¹⁰-trihydroxyferuloyl spermidine and N ¹, N ⁵-dihydroxyferuloyl- N ¹⁰-sinapoyl spermidine. These products, deposited in pollen coat, can be readily extracted by pollen wash and were shown to be responsible for pollen autofluorescence. The activity of the recombinant enzyme produced in bacteria was assayed with various hydroxycinnamoyl-CoA esters and polyamines as donor and acceptor substrates, respectively. Feruloyl-CoA and spermidine proved the best substrates, and the enzyme has therefore been named spermidine hydroxycinnamoyl transferase (SHT). A methyltransferase gene ( At1g67990 ) which co-regulated with SHT during flower development, was shown to be involved in the O -methylation of spermidine conjugates by analyzing the consequences of its repression in RNAi plants and by characterizing the methylation activity of the recombinant enzyme.     

Growth and accumulation of N, K+, Ca2+ and Mg2+ in barley exposed to various nutrient regimes and root/shoot temperatures

Six cultivars of spring barley ( Hordeum vulgare L. cvs Salve, Nümberg II, Bomi, Risø 1508, Mona and Sv 73 608) were grown in water culture for three weeks with various combinations of mineral supply and differential roots/shoot temperatures during the growth period. Most important for growth and accumulation of N, K⁺, Ca²⁺ and Mg²⁺ was the mineral supply, followed by the root temperature and the choice of cultivar. Treatments with low mineral supply or low root temperature induced a uniform reduction in growth and accumulation of the ions studied. The effects of low mineral supply and low root temperature on growth and N accumulation was additive, which indicates that these factors exert their influence independently of each other.
Roots grown at 10°C were smaller and Rb⁺(⁸⁶Rb) influx was higher than in roots grown at 20°C. It is suggested that the control of Rb⁺(⁸⁶Rb) influx is affected by the root temperature and the age of the plants. The higher ⁸⁶Rb⁺ (⁸⁶Rb) influx into the low temperature roots could not compensate for the smaller root size. However, the lower total mineral accumulation made up for the needs of the smaller plants and cannot explain the reduction in growth.     

Salinity response of a freshwater charophyte, Chara vulgaris

Abstract. Chara vulgaris L. growing in an oligohaline lake was adapted to laboratory conditions and subjected to long-term salinity treatments ranging from 0 to 350 mol m ³ NaCl added to the lake water (40–680 mosmol kg ¹). Osmotic potential and concentration of the main osmotically active solutes (K⁺, Na⁺, Mg²⁺, Cl and sucrose) in the vacuolar sap of the central internodal cells were estimated. C. vulgaris did regulate turgor but incompletely. Turgor decreased from 335 mosmol kg ¹ under control conditions to 52–111 mosmol kg ¹ at 350 mol m ³ NaCl. The enhancement of πⁱ was achieved by increase in both ions and sucrose. Sterile and fertile plants differed in their response to osmotic stress. In sterile plants, the ions accounted for about 87% of the vacuolar osmotic potential. The increase of πⁱ under osmotic stress was exclusively due to an accumulation of Na⁺ and Cl^-. In fertile plants, sucrose accounted for about 35% of πⁱ and ions for about 51% Under osmotic stress, sucrose content increased together with the ionic content of Na⁺ and Cl^-.     

Growth analysis of wild-type and photomorphogenic-mutant tomato plants

A custom designed growth-measuring apparatus, controlled by a microcomputer has been used to study extension growth kinetics of wild-type and photomorphogenic-mutant tomato ( Lycopersicon esculentum Mill.) plants with and without end-of-day farred light (EODFR). The following photomorphogenic mutants were used. Far-red insensitive ( fri ^.1): deficient in phytochrome A (phyA); temporarily red light-insensitive ( tri ^.3): deficient in phytochrome Bl (phyB1), and their isogenic wild type (WT) cv. MoneyMaker. aurea (au) : deficient in phytochrome chromophore biosynthesis; high-pigment-1 ( hp-1 ): exhibiting exaggerated phytochrome responses, and their isogenic WT cv. Ailsa Craig. The stem elongation rate (SER) during a 24-h period of all the genotypes studied shows a similar pattern, having two dramatic transients, one shortly after the onset of the light period (a sharp decline in SER) and another shortly after the start of the dark period (a sharp increase in SER). These transients are probably associated with water relations as a consequence of opening and closure of the stomata. The fastest SER occurs during the dramatic oscillations early in the dark period. Between the genotypes there are large quantitative differences in SER. All the genotypes tested exhibited a strong EODFR response, resulting in a relative promotion of SER during the first period after the start of EODFR and in the subsequent light and dark periods. These results indicate that phyA, absent in the fri ^.1 mutant, does not play a major role in SER of light-grown tomato plants, whereas phyB 1, absent in the tri ^.3 mutant, is partly responsible for the compact stature of WT plants. An additional phytochrome other than phy A and phy B1 must therefore be capable of eliciting the EODFR response.     

Genetic control of the immune response in lines of chickens selected for graft-versus-host competences

The immune response of chickens to goat erythrocytes has been examined. The H line selected for high competence of graft-versus-host reaction (GVHR) showed higher immune responses than the L line selected for low GVHR competence. It appeared also that immune responses were controlled by the B blood group locus, which is the major histocompatibility locus in chickens. The relative immune responsiveness of B genotypes were B ¹¹/¹¹> B ⁹/ B ¹¹> B ⁹/B⁹.
Treatment of antiserum with 2-mercaptoethanol (2-ME) proved that the difference in immune responses between lines was due mainly to the 2-ME resistant antibody and that the difference between the B genotypes was due to the 2-ME sensitive antibody.     

Molecular Detection of Phytophthora cryptogea on Calendula officinalis and Gerbera jamesonii Artificially Inoculated with Zoospores

In this work, a protocol for zoospores production of Phytophthora cryptogea , an economically important plant pathogen was optimized. Five different concentrations of zoospores (5 × 10⁵, 5 × 10⁴, 5 × 10³, 5 × 10², 5 × 10¹ zoospores/ml) from four different isolates of P. cryptogea (Maria 1, Maria 2, S3 1-A, Amazzone) were used as inoculum on pot marigold ( Calendula officinalis ) and gerbera ( Gerbera jamesonii ) plants. Maria 1 was the most virulent isolate both on pot marigold and gerbera plants according to disease severity. A rapid and sensitive pathogen DNA extraction protocol suitable for large quantities of plant samples was adopted. Conventional polymerase chain reaction (PCR) was able to detect the pathogen in artificially inoculated symptomless pot marigold (collected day 12) and gerbera plants (day 8) after pathogen inoculation, with the suspension of 5 × 10⁵, 5 × 10⁴, 5 × 10³ P. cryptogea  zoospores/ml. Real-time PCR showed the possibility to detect the pathogen in artificially inoculated symptomless pot marigold (collected day 8) and gerbera plants (day 4) after pathogen inoculation, with the suspension of 5 × 10⁵, 5 × 10⁴ P. cryptogea  zoospores/ml. The first symptoms appeared on pot marigold plants 14 days after pathogen inoculation and on gerbera plants 10 days after inoculation. Real-time PCR showed the possibility to detect the pathogen 4 days before conventional PCR and 6 days before the appearance of disease symptoms both on pot marigold and gerbera plants.     

Random mutagenesis of Legionella pneumophila with mini-Tn10

Abstract The degradation of sheets of poly(3-hydroxybutyrate- co -3-hydroxyvalerate) (BIOPOL^®) by aerobic sewage sludge was analyzed. Degradation of the polymer was highly dependent on the pH of the culture medium and was maximal between pH 7 and pH 8.5. Below pH 6 and above pH 9 the degradation rate was very low. Agitation of the culture fluid had relatively little influence on the rates of degradation. 1.2×10⁵ aerobic polymer-degrading bacteria per ml sewage sludge were identified by halo formation on solid poly(3-hydroxybutyrate) (PHB)-containing media. The number of PHB-degrading bacteria in other ecosystems amounted to 3.8×10³ per ml sludge of a fresh-water lake, 9.2×10⁵ per g garden-soil, 1.3×10⁶ per g field-soil and 4.3×10⁶ per g compost.     

Diversity of Rice yellow mottle virus in Benin and Togo and Screening for Resistant Accessions

Surveys were conducted in rice fields in Benin, Cote d'Ivoire, Mali, Nigeria, Togo and Niger to assess the importance of Rice yellow mottle virus (RYMV). Diseased leaf samples were collected. In Togo, surveys were made mainly in the southern part of the country, and in Benin, all areas where rice is grown were covered. Leaf samples were serologically confirmed by ACP ELISA as bearing RYMV, propagated and all conserved. One hundred and forty‐eight (148) Beninese and 27 Togolese strains were serotyped. They were also phenotyped on three susceptible accessions including IR 64, four resistant lines with known alleles on RYMV 1 gene namely TOG 5681 (rymv 1‐3), TOG 5672 (rymv 1‐4 and RYMV 2), TOG 5674 (rymv 1.5) and Gigante (rymv 1‐2). RYMV spots with 9–100% incidences were identified. Serotyping by triple antibody sandwich (TAS) ELISA indicated that two main groups S1 and S2 coexisted in Benin with S1 being prevalent. In Togo, 26 strains were S1 and only one was S2. Phenotyping of the 148 Beninese and 27 Togolese indicated that they all attacked the three susceptible accessions, while TOG 5681 (rymv 1‐3), TOG 5672 (rymv 1‐4 and RYMV 2), TOG 5674 (rymv 1‐5) and Gigante (rymv 1‐2) remained symptomless. The use of the genes/alleles above in these countries against the disease is discussed. Three representative Beninese strains were selected to screen 48 accessions for disease resistance. Nine accessions were as susceptible as IR 64 to all strains while six (NERICA 9, NERICA 12, NERICA 13, TOG 7291, WAB56‐50, CG 14 and Moroberekan) were very resistant. Susceptibility of the six ARICA and resistance of some NERICA lines could be explained by the fact that one or both parental lines were, respectively, susceptible or resistant to the strains.     

Longevity of the freshwater anostracan Streptocephalus mackini (Crustacean: Anostraca) in relation to food (Chlorella vulgaris) concentration

SUMMARY 1. Temporary ponds are inhabited by a variety of invertebrates, of which anostracans are an important group. We studied the lifetables of male and female anostracan Streptocephalus mackini at 3 algal concentrations (0.5 × 10⁶, 1.0 × 10⁶ and 1.5 × 10⁶ cells mL⁻¹).
2. Regardless of sex, S. mackini showed better survivorship at lower food levels. The longest average lifespan observed was 85 ± 2 days for males fed Chlorella at 0.5 × 10⁶ cells mL⁻¹.
3. Both net reproductive rate and generation time decreased with increasing food level. The highest net reproductive rate was about 120 cysts per female. The longest generation time of about 40 days, observed at 0.5 × 10⁶ cells mL⁻¹, was more than three times that at 1.5 × 10⁶ cells mL⁻¹.
4. The rate of population increase ( r ) was nearly the same (0.31 ± 0.06) at high (1.5 × 10⁶ cells mL⁻¹) and intermediate (1.0 × 10⁶ cells mL⁻¹) food levels. The r -value at low food level (0.5 × 10⁶ cells mL⁻¹ of Chlorella ) was 0.20 ± 0.01 per day.     

Transformation of Aspergillus parasiticus using autonomously replicating plasmids from Aspergillus nidulans

Abstract A genetic transformation system for the aflatoxin-producing fungus Aspergillus parasiticus using two autonomously replicating plasmids from A. nidulans (ARp1 and pDHG25) is reported. Transformation frequencies using the plasmid pDHG25 were from 5 × 10² to 2.5 × 10⁴ transformants per 10⁶ viable protoplasts and μg DNA. The stability of the plasmids in the transformants was also studied. This transformation system offers a new opportunity to clone genes related to aflatoxin production using appropriate aflatoxin-defective mutants.     

Multiplication of Pseudomonas syringae pv. phaseolicola "in planta"

Multiplication of Pseudomonas phaseolicola was determined in 17 different bean cultivars ( Phaseolus vulgaris ) and 9 other plant species, and the effect of different inoculation methods and conditions was also studied.
In susceptible leaves, a generation time of 2.1 h was determined in the early phase (2 days after inoculation). Different multiplication rates in susceptible and resistant leaves were clearly observed 4 days after inoculation. At this time the first small water-soaked spots were visible in the susceptible cultivars. Bacteria multiplied up to the 7th day after inoculation with a maximum of 10⁹ cells per cm² leaf (equal to ca. 4 × 10¹⁰ bacterial cells/cm³). At the same time, the water-soaked spots had reached their maximum size in most cases. Thus, bacterial multiplication and development of water-soaked spots paralleled each other.
In resistant leaves, no water-soaked spots appeared, and the final bacterial concentration was 1/1000–1/100 of that in susceptible leaves. Gomparison of races 1 and 2 in several bean cultivars indicated the non-existence of a gene-for-gene relationship with this disease. Old leaves were less susceptible to infection. Some bacterial multiplication was also observed in non-host plants. There was a general correlation between bacterial multiplication in the non-host plants and their botanical relation to Phaseolus vulgaris .     

Characterization of Na+ exclusion mechanisms of salt-tolerant reed plants in comparison with salt-sensitive rice plants

Salt-tolerant reed plants ( Phragmites communis Trinius) and salt-sensitive rice plants ( Oryza sativa L. cv. Kinmaze) were grown in salinized nutrient solutions up to 50 m M NaCl, and growth, Na⁺ contents and kinetics of ²²Na⁺ uptake and translocation were compared between the species to characterize the salt tolerance mechanisms operating in reed plants. When both plants were grown under the same salinity, Na⁺ contents of the shoots were lower in reed plants, although those of the roots were quite similar. The shoot base region of both species accumulated Na⁺ more than the leaf blades did. Sodium-22 uptake and pulse-chase experiments suggested that the lower Na⁺ transport rate from root to shoot could limit excessive Na⁺ accumulation in the reed shoot. There was a possibility that the apparently lower ²²Na⁺ transport rate to the shoot of reed plants was due to net downward Na⁺ transport from shoot base to root.     

Effects of nitrate on influx, efflux and translocation of potassium in young sunflower plants

Influx, efflux and translocation of K⁺(⁸⁶Rb) were studied in the roots of sunflower seedlings ( Helianthus annuus L. cv. Uniflorus) treated with 0–4.0 m M NO₃⁻ during a 9 day growth period or a 24 h pretreatment period. Roots treated with high levels of NO₃⁻ absorbed and translocated more K⁺(⁸⁶Rb) than seedlings treated with low levels of NO₃⁻. The content of K⁺ in the shoots was, however, higher in seedlings treated with low levels of NO₃⁻, indicating a low rate of retranslocation of K⁺ in those plants. K⁺(⁸⁶Rb) efflux was highest into the low-NO₃⁻ solutions. All effects on K⁺(⁸⁶Rb)-fluxes were more obvious in high-K plants than in low-K plants. The results are discussed in relation to the Dijkshoorn-Ben Zioni hypothesis for K⁺+ NO₃⁻-uptake and translocation in plants.     

Biological activity of monomeric Sendai virus HN glycoprotein

Abstract We have developed a transformation system for Streptomyces wadayamensis , a cephamycin C producer. 1−5 × 10⁹ protoplasts can be obtained when late logarithmic phase cultures of this microorganisms are incubated with 10 mg/ml of lysozyme. Polyethylene glycol-Ca²⁺-mediated transformation of these protoplasts yielded 10⁶ transformants per μg of pIJ702 or pIJ365 DNA.     

Anion uptake in maize roots: Interactions between chlorate and nitrate

Effects of nitrate, chloride and chlorate ions upon nitrate and chlorate uptake by roots of maize ( Zea mays L., cv. B73) seedlings were examined. Net nitrate uptake, ³⁶ClO₃⁻ influx and ³⁶Cl⁻ influx (the latter two in a background of 0.5 m M KNO₃) displayed similar pH profiles with optima at pH 5.5 and below. External, non-labeled chloride had little effect on the accumulation of ³⁶ClO₃⁻ (both in 5 h and 20 min uptake assays), while nitrate and chlorate had almost identical, marked inhibitory effects. Nitrate pretreatment caused an apparent induction of both ³⁶ClO₃⁻ and ¹⁵NO₃⁻ uptake activities. After 5 h of treatment in nitrate, the uptake activities of chloride- and chlorate-pretreated plants increased to that of nitrate-pretreated plants. During 6 h exposure to chlorate, ³⁶ClO₃⁻ uptake activity of nitrate-pretreated plants decreased to that of chlorate- and chloride-pretreated plants. The results support the existence of a shared nitrate/chlorate transport system in maize roots which is not inhibited by external chloride, and which is induced by nitrate, but not by chlorate or chloride. The suggestion is made that selection of chlorate-resistant mutants of maize can identify nitrate uptake as well as nitrate reductase mutants.     

Detection of Yersinia enterocolitica in food by PCR amplification

A polymerase chain reaction (PCR) assay was developed for detection of pathogenic, virulent strains of Yersinia enterocolitica . By using both virulence loci virF and ail as markers for pathogenicity, detection of species with a virulence factor present was possible. DNA preparation in the presence of hexadecyl trimethy ammonium bromide (CTAB) was followed by two 44 cycle amplification reactions, one for each of the markers. As few as 10² Y. enterocolitica cells were detected in ground pork in the presence of 10⁵–10⁶ bacteria of other species. The described PCR assay provides a sensitive robust assay for the detection of virulent Y. enterocolitica in food.     

Efficacy of certain disinfectants against infectious pancreatic necrosis virus

The virucidal properties of iodophor, chlorine (sodium hypochlorite), formalin, thimerosal (organic mercurial compound), malachite green, and acriflavine were tested on infectious pancreatic necrosis virus (IPNV). Iodine and chlorine showed good activity, but efficacy depended on the concentration of virus, the presence of organic matter (calf serum), and water _pH. Water hardness (0-300 mg 1⁻¹ as CaCO₃) did not affect virucidal activity. In a 5 min exposure, 4 mg 1⁻¹ available iodine inactivated 10^3.9 TCID₅₀ m1⁻¹ IPNV but 16 mg 1⁻¹ iodine were needed for inactivation of 10^6.3TCID₅₀m1⁻¹. The addition of 0-5% calf serum significantly reduced the iodine concentration and the virucidal activity. In comparison, 4 mg 1⁻¹ chlorine were needed to inactivate 10⁴⁶ TCID₅₀ m1⁻¹ IPNV in 5 min. However, the addition of 0-07 % serum greatly reduced the chlorine concentration and extended the virucidal contact time to 30 min or more. IPNV at 10^6.3 TCID₆₀ m1⁻¹ was not inactivated by exposures for 60 min to 0-2% formalin, 10 min to 0-2% thimerosal, 60 min to 5 mg 1⁻¹ malachite green, or 20 min to 500 mg 1⁻¹ acriflavine. However, acriflavine at 0-5 mg 1⁻¹ in cell culture media prevented the development of cytopathology caused by IPNV and may be useful in the treatment of the disease.