Factors affecting the sporulation of Gloeosporium perennans and Gloeosporium fructigenum in vitro

An ability to spore predictably and plentifully when grown in darkness at a constant temperature, is a highly desirable attribute for any organism used to test antisporulant chemicals in vitro . The extreme variability of isolates, and their rapid loss of vigour in culture, resulted in erratic sporulation by Gloeosporium perennans : although sporulation could be temporarily improved by various means, it remained too variable for assessing the antisporulant activity of chemicals in vitro. G. fructigenum , on the other hand, grew vigorously throughout a long series of transfers, and spored reliably and plentifully when grown under the same conditions.     

THE ROTTING OF APPLES BY GLOEOSPORIUM PERENNANS ZELLER & CHILDS

The extent of rotting of commercially stored Cox's Orange Pippin apples by Gloeosporium species has been severe in recent years. G. perennans has been found to be mainly responsible.
Tree infections of this fungus have usually been found to be prevalent in the orchards on farms where storage losses of fruit have occurred, and conidia of G. perennans have been found on cankers at all seasons during the year. Winter as well as summer pruning cuts have been found to become infected. Detached leaves of Cox's Orange Pippin have been successfully inoculated with G. perennans in the laboratory and the fungus has also been found on dead detached leaves in the orchard after picking time. Considerable variations have been found in the susceptibility to rotting of samples of different varieties of apple and samples of Cox's Orange Pippin apples from different orchards. Periodic loading of fruit with spores after picking has shown that resistance declines with length of storage. Wide variation in the time at which rotting commences in commercially stored samples has been observed. A series of inoculations on picked and unpicked immature fruit has shown that the apple loses its resistance to attack on picking. Lenticels which are impermeable to gaseous exchange have been shown to be resistant to penetration by the fungus. Temperature affects the development of lenticel rotting by G. perennans , infection being most rapid at temperatures of about 20°. Rotting can occur at 0° C.     

OBSERVATIONS ON APPLE CANKER

Field observations on leaf-scar canker of apple confirm its importance in the southwest of England: on a heavily infected tree more than 5% of the current year's growth may be damaged. Leaf scars may become infected by spores of Nectria galligena which are sucked into the tracheids of a freshly exposed scar where they develop in a relatively protected environment. These freshly exposed scars become much less susceptible to infection after approximately one hour.
Infections established in leaf scars may develop immediately into cankers or they may be confined by a suberized or gum barrier. Mycelium from some of these confined lesions may spread into the surrounding healthy tissue through growth cracks when the buds develop. The lesions extend throughout the winter and spring; usually the scars earliest inoculated are the first to show cankers. It seems possible that infections of this type might be controlled by eradicant sprays applied just before leaf-fall.     

The control of apple canker (Nectria galligena) in a young orchard with established infections

In a factorial experiment on canker control the efficiency of phenylmercurie nitrate (PMN) applied at leaf fall, before bud burst or both was compared with the application of dodine, dithianon, triforine, thiabendazole, benomyl or carbendazim in May and June. Of the total number of cankers which developed in unsprayed trees 76–78% resulted from infections in April to August of each year of the experiment. Infection was reduced significantly by all of the fungicides applied in summer. Of these carbendazim was outstanding, controlling both summer and autumn infections. Sporulation throughout the summer was suppressed by carbendazim and to a lesser extent benomyl, but whereas the suppressant effect of carbendazim persisted until long after leaf fall that of benomyl was evident only until August. Dodine, dithianon, triforine and thiabendazole had no significant effect on spore production. Dithianon and dodine showed highest toxicity to the germination of Nectria galligena spores of all fungicides used in summer. All of the fungicides controlled apple scab (Venturia inaequalis) although thiabendazole was the least effective. Autumn applications of PMN reduced canker incidence but their value alone was not as great as carbendazim, dithianon, benomyl or dodine applied in summer. PMN applied in spring reduced the number of cankers in trees receiving no other fungicides but tended to increase the incidence of infections, particularly in autumn, when used in conjunction with fungicides applied in May and June. PMN applied before bud burst reduced sporulation of N. galligena for a few weeks, after which production resumed and in late summer and autumn actually exceeded the controls. None of the fungicides had any direct effect on leaf fall. Infection of the crotch and basal leaf scars was more common in summer than in autumn whereas infection of leaf scars above the basal region was more common in autumn than in summer. The rootstocks of many of the trees became infected via callus tissue associated with adventitious root development and this was controlled by the carbendazim treatment.     

Pollen germination, tube growth and morphology, and microtubule organization after exposure to benomyl

The inhibitory effects of benomyl on pollen tube growth have received little attention, particularly at the microscopic and immunohistochemical levels. Pollen germination and tube growth in the presence of benomyl were evaluated in Tradescantia virginiana to investigate the effects of this fungicide on pollen germination rate, tube growth and morphology, and microtubule (Mt) organization. Benomyl was incorporated in germination media at 0,480, 600 or 720 mg 1⁻¹. Inhibition of pollen germination, cytoplasmic streaming, and tube elongation were associated with benomyl treatments. Benomyl also induced abnormal pollen tube morphology and Mt organization. Compared to controls, Mts in the treated tubes were characteristically fewer in number, fragmented, sinuous and increasingly disorganized. At the two highest benomyl concentrations, Mts were considerably fewer or absent in apical/subapical regions of the pollen tubes. This work verifies that benomyl incorporated into germination media at concentrations lower than recommended field rates inhibit pollen germination and tube growth, and that the effects are associated with alterations of Mt organization.     

Field experiments in the control of Verticillium dahliae and Heterodera rostochiensis on potatoes

Potatoes were grown in soil infested with Verticillium dahliae and Heterodera rostochiensis and treated with methyl bromide, aldicarb or benomyl. Successive crops were grown in subsequent years on the same plots but without further treatment. Largest yields in the year of treatment were from plots fumigated with methyl bromide but in the second crop benomyl-treated plots yielded most. Neither methyl bromide nor aldicarb affected yields from trie third crop. After lifting the first crop, soil from plots treated with methyl bromide or benomyl contained less V. dahliae than that from plots receiving aldicarb or nothing. Soil was also less infective after the second crop following methyl bromide fumigation, but not the third. Plots treated with methyl bromide or aldicarb contained many fewer H. rostochiensis than untreated plots after the first crop, slightly less after the second and equal numbers after the third crop. H. rostochiensis were also fewer in benomyl-treated plots than in untreated after the first crop, but, surprisingly, were fewest after the second crop. The increased yield after applying benomyl to soil seems to depend more on its effect on H. rostochiensis than on V. dahliae. Fumigating with methyl bromide decreased common scab and black scurf on progeny tubers but increased infections by Oospora pustulans.     

An assessment of some new treatments for the control of rotting of stored apples

Benomyl (0.025% a.i.) and thiabendazole (0.05 % a.i.) applied in July, August and September to apple trees cvs. Sunset and Cox's Orange Pippin gave slightly better control of Gloeosporium spp. rots in stored fruit than captan (0.1% a.i.) applied at similar intervals. On Cox's Orange Pippin, treatment with captan at 0.1 % a.i. in July and at 0.2% a.i. in August was as effective as three sprays at 0.1 % a.i. in July, August and September in controlling both Gloeosporium spp. and Nectria galligena rots. Benomyl applied in the spring and early summer to control apple scab on orchard trees reduced the incidence of Gloeosporium spp. and Monilia fructigena in stored fruit. Pre-harvest sprays of benomyl and captan gave good control of rotting caused by N. galligena but thiabendazole was ineffective. Post-harvest dips of thiabendazole controlled rotting by Gloeosporium spp. butnot byN. galligena.     

Influence of Pesticide Use on the Natural Occurrence of Entomopathogenic Fungi in Arable Soils in the UK: Field and Laboratory Comparisons

The spectrum and abundance of entomopathogenic fungi in agricultural soil receiving different pesticide applications were evaluated . Seven small field plots within a barley crop were selected . Each plot had received a different pesticide treatment at slightly higher than the field rate each year for the previous 12-19 years . The field plots received either benomyl (fungicide) , triadimefon (fungicide) , aldicarb (insecticide) , chlorfenvinphos (insecticide) , glyphosate (herbicide) , all five of these pesticides or no pesticides at all (control) . Soil sampled from each plot was baited with Galleria mellonella larvae at either 18 or 26 C . Five species of entomopathogenic fungi infected these larvae . Beauveria bassiana was the dominant species , and the only species for which infection levels were high enough to be analyzed statistically . Significantly fewer G. mellonella larvae became infected with B. bassiana in soil treated with benomyl than in other treatments . This deleterious effect was confirmed in in vitro experiments where benomyl inhibited fungal growth significantly . Chlorfenvinphos also reduced the proportion of G. mellonella becoming infected with B. bassiana slightly (although not statistically significantly) . This effect was much greater in in vitro experiments in which chlorfenvinphos inhibited fungal growth significantly . B. bassiana- induced mortality was significantly greater in G. mellonella larvae placed in soil treated with triadimefon than in other treatments . In vitro studies , however , demonstrated that triadimefon inhibited rather than stimulated growth . A similar anomaly between field and laboratory observations was seen in the case of aldicarb . There was no significant difference in the number of G. mellonella larvae found infected with B. bassiana in soil treated with aldicarb than in the control soil . However , in the in vitro studies , aldicarb stimulated fungal growth at all concentrations except 10 times the field rate . In general , significantly fewer larvae became infected with B. bassiana at 26 C than at 18 C in all treatments . Pesticides may have a direct impact on the natural occurrence , infectivity and population dynamics of entomopathogenic fungi . They also affect other macro - and microorganisms in the soil which may interact with the ento mopathogenic fungi . In combination with climate and soil variables , these effects are difficult to separate . Although any suppression of these fungi may be detrimental to their capacity as natural enemies , this study has demonstrated that pesticides used under field conditions are unlikely either to kill all the entomopathogenic fungi present in the treated area or to limit their recolonization . It seems likely , therefore , that pesticides have the potential to be used in conjunction with mycoinsecticides in integrated pest management systems .     

Evaluation of the effects of chemical versus biological control on Botrytis cinerea agent of gray mould disease of strawberry

This study investigates on effects of four fungicide and six isolate from Trichoderma and Gliocladium on Botrytis cinerea agent grey mold of strawberry under library and greenhouse condition. The effect of four fungicides i.e. benomyl, dichlofluanid, captan and triadimenol on B. cinerea was studied in the laboratory condition by method mixed poison to culture medium. It was shown that the fungicide including benomyl, triadimenol, dichlofluanid and captan were able to inhibit mycelial growth of B. cinerea on PDA plate with EC50 of 0.16, 1.42, 3.40 and 7.73 ppm respectively. These fungicides delayed myceliogenic germination of sclerotia at 1000 ppm, while exhibiting no fungicidal effect. Moreover, the antagonistic effects of six fungi including Trichoderma koningii (T21), T. viride (T4), T. harzionum (T5), T. viride (T2), G. virens (G2), G. virens (G8) on B. cinerea were assessed. This assessment was done under library condition and its results as follows: The antagonistic mechanism occurred through branching at the end of B. cinerea hyphae, hyphal contact, coiling, vacuolization and lyses. Volatile metabolites of T. koningii (T21) and non-volatile metabolites of G. virens (G2 and G8) and T. koningii (T21) caused maximum inhibition of the fungal growth. Trichoderma spp and G. virens were able to colonize and sporulate on sclerotia and caused their lysis within 7-21 days. In greenhouse, a completely randomized design with 11 treatments (4 chemical and 6 biological and one untreated control) each replicated five times were used for the comparison. Greenhouse studies revealed that application of fungicides i.e. captan, dichlofluanid, triadimenol and benomyl reduces disease severity by 42, 45, 48 and 52% respectively. The fungal antagonists reduce the grey mold disease severity between 5-42%. All treatments caused a decline in post harvest disease, as the most effective treatment of chemical control was benomyl with 68.33% and for the biological treatment this was T. koningii (T21) with 56%.     

The effect of carbendazim-generating fungicides with and without a mineral oil additive on tomato stem lesions caused by carbendazim-sensitive and tolerant strains of Botrytis cinerea

Isolates of Botrytis cinerea were obtained from tomatoes in several localities in the West Scotland. Some isolates grew on agar containing 100 mg/1 benomyl (carbendazim-tolerant), while others did not (carbendazim-sensitive). Pot-grown tomato plants treated with benomyl and other carbendazim-generating fungicides, applied either as sprays or soil drenches, were inoculated on the leaf scars with some of these isolates. On treated plants the carbendazim-tolerant isolates formed lesions which were about as large as those on untreated plants. Sensitive isolates formed much smaller lesions on treated plants. There was evidence that the increase in lesion size during the period 7–14 days after inoculation with a carbendazim-sensitive isolate was less on plants sprayed with benomyl or carbendazim with added mineral oil (2% Actipron) than on plants to which the fungicides alone had been applied. No such effect was recorded with thio-phanate-methyl. There was also an indication that the addition of Actipron to a benomyl spray improved the effect of the fungicide against two tolerant isolates, though there was no effect on the relative increase in lesion size during the second week after inoculation. In two tests the addition of 2% and 4% Actipron to benomyl soil drenches did not improve the level of leaf scar lesion control.     

Effects of treating seed potatoes from commercial and stem cutting stocks with benomyl, thiabendazole and 2-aminobutane on yield and disease

Potato seed tubers of six cultivars from commercial stocks and from stocks derived from stem cuttings (healthier seed) were fumigated with 2-aminobutane 2 wk after lifting or treated with benomyl or thiabendazole in January. 2-aminobutane prevented skin spot and gangrene developing on treated tubers. Experiments were planted at Rothamsted (clay with flints soil) and at Woburn (sandy loam soil) in 1973–75. Healthier seed produced more stems/plant than commercial stocks and yielded on average 8% more at Rothamsted in 1973 and 1974 and respectively 5 and 10% more at Woburn in 1973 and 1975. Seed treatments did not consistently affect stem numbers or increase yield although all treatments tended to decrease tuber size. Infection of stem bases and tubers by Polyscytalum pustulans and Rhizoctonia solani was usually less from healthier than from commercial seed and was decreased by benomyl and thiabendazole in 1973 and 1974. Infection by Helminthosporium solani of the skin around tuber eyes was greater from healthier than from commercial seed but was decreased by benomyl and thiabendazole. 2-aminobutane sometimes decreased infection of tubers by P. pustulans and R. solani but neither of stem bases nor of tubers by H. solani. Gangrene on tubers uniformly wounded at lifting was not consistently affected by seed source or seed treatment. Treating seed with benomyl or thiabendazole in 1975 decreased skin spot and black scurf in tubers stored until March 1976. These treatments also decreased silver scurf on the produce of commercial seed at Rothamsted but gangrene was not consistently affected by seed treatments.     

The disappearance of benomyl from mushroom casing

The failure of Benlate (50% benomyl) to control Mycogone perniciosa on mushroom farms where isolates of the organism were benomyl sensitive was investigated. A comparison was made of carbendazim levels, the major fungitoxic breakdown product of benomyl, in unsterilised casing, autoclaved casing and autoclaved casing mixed with a small proportion (1.7%) of casing previously treated with benomyl but from which all detectable levels of carbendazim had disappeared. Added benomyl was effective in controlling M. perniciosa in autoclaved casing, less effective in unamended casing and ineffective in the amended casing mixture. The recovery of carbendazim from these treatments was directly related to disease occurrence. Studies on farms where benomyl or other carbendazim generating fungicides had been used indicated that a lack of control of M. perniciosa was associated with the disappearance of fungicide from the casing before cropping began. This was not so on a farm where carbendazim generating fungicides had not been used before these investigations. Thiabendazole, a fungicide closely related to the carbendazim generators showed only a slight decline in concentration in casing capable of degrading benomyl. Bacteria isolated from the casing were examined in vitro for their ability to degrade benomyl. Five groups of bacteria were compared and three bacteria, Pseudomonas putida, Pseudomonas aeruginosa and an unidentified member of the Entero-bacteriaceae were most able to degrade benomyl.     

Effect of the fungicide benomyl on spore germination and hyphal length of the arbuscular mycorrhizal fungus Glomus mosseae.

The fungicide benomyl inhibited spore germination and hyphal length of the arbuscular mycorrhizal fungus Glomus mosseae when applied at doses of 21.25 microg/ml (agronomic dose), 10.62 microg/ml and 10 microg/ml. G. mosseae was able to germinate in the presence of 2.12 microg/ml of benomyl, and the percentage of spore germination was unaffected by dosis of 0.1, 0.01 and 0.001 microg/ml of the fungicide. However, all doses of fungicide tested in this study decreased the hyphal length. When ungerminated G. mosseae spores previously exposed to benomyl were transferred to water-agar medium without benomyl, the maximum germination was 16%. Small spores of G. mosseae were more resistant to benomyl than the larger ones. Our results show some of the factors which can explain the variability of the effect of benomyl on arbuscular mycorrhizal fungi.     

Relationship of Endopolygalacturonase Inhibitor Activity to the Rate of Fungal Rot Development in Apple Fruits

An inhibitor extracted form the cell walls of apple fruits suppressed the activity of endopopygalacruronases (endo-PGs) produced in vivo and in vitro by Nectria galligena, Phomopsis mali, Fusarium Lateritium and Glomerella cingulata but not the endo-PGs produced by Penicillium expansum or Phytophtobora syringae. Of four apple cultivars tested Granny Smith tissue contained the highest levels of inhibitor and Cox's Orange Pippin contained the least. Linear rot expansion in the four apple cultivars inoculated with N. galligena was inversely related to inhibitor activity in the fruit tissue, rot development being slowest in Granny Smith fruits and most rapid in Cox's Orange Pippin fruits. Rot expansion in fruits inoculated with P. expansum bore no such relationship to inhibitor activity in the tissue Apple tissue maceration by the endo-PGs from N. galligena, P. mali. F. lateritium and G. cingulata was similarly related to inhibitory activity in the fruit. The properties of the partially purified inhibitor were consistent with it being proteinaceous but the relative slowness with which it was hear inactivated and the presence of a small percentage of carbohydrate might indicate that it was a glycoprotein.     

Induction of multiple germ tubes in Neurospora crassa by antitubulin agents

The antitubulin fungicide benomyl suppressed the linear growth of Neurospora crassa wild type strain St. Lawrence 74 at micromolar concentrations. The rate of germination of macroconidia was not affected. Macroconidia exposed to 1.7 microM benomyl for 5 h formed multiple germ tubes. When germlings incubated for 4 h were exposed to 1.7 microM benomyl for 3 h, their germ tube stopped growing, swelled and emitted several branches. Normal linear growth was restored after removal of the fungicide. Linear growth of N. crassa was resistant up to 16 microM nocodazole. This drug induced multipolar germination at 8 microM, and griseofulvin only at 140 microM. The microtubule (MT) cytoskeleton of N. crassa could be revealed by indirect immunofluorescence with the monoclonal antibody YOL 1/34 directed against yeast alpha-tubulin. We detected no striking effects of the benomyl treatments on MT organization. The MT-stabilizing agents deuterium oxide (D2O) and cAMP have no antagonistic effects on the benomyl-induced multipolar germination. The positioning of nuclei and mitochondria was determined from the DAPI and Rhodamine 123 fluorescence patterns, respectively. Benomyl inhibited nuclear migration into multiple germ tubes. Quantitative scanning cytophotometry revealed a peak in the intensity of the mitochondria-associated Rhodamine 123 fluorescence near the apex of untreated germlings. This peak disappeared in multiple germ tubes. Benomyl-resistant mutant bml 511 (r), mutated in its beta-tubulin gene, germinated normally in the presence of the fungicide. This strongly suggests that multiple germ tube formation was due to the effect of benomyl on beta-tubulin. Benomyl-resistant strain 74-3, constructed by reintroducing the cloned mutant N. crassa beta-tubulin gene into the cells by transformation, displayed a partial resistance to benomyl with respect to multipolar germination. Its rate of germination was slow (50% germination reached after 4 h at 37 degrees C as compared to 2.5 h for the wild type). In contrast to N. crassa, the other ascomycete Aspergillus nidulans is nocodazole-sensitive (linear growth suppressed at 1.6 microM). It did not respond to the MT inhibitors benomyl and nocodazole with respect to the pattern of germ tube emergence. Our results suggest that microtubule or membrane beta-tubulin is involved in the maintenance of developmental polarity during germ tube emergence and growth of N. crassa.     

Effects of soil treatments and cultivars on 'early dying' disease of potatoes caused by Globodera rostochiensis and Verticilliurn dahliae

Plots in a field infested with Globodera rostochiensis and Verticillium dahliae were treated in 1971 with methyl bromide, dazomet, aldicarb, benomyl or aldicarb + benomyl and potatoes (cv. Pentland Dell) were grown in four consecutive years. In 1971, all treatments delayed the development of haulm symptoms, decreased soil populations of G. rostochiensis and increased yields. In the second year yields were increased by all treatments except dazomet whereas in the third and fourth years only benomyl or aldicarb + benomyl increased yields. Soil populations of V. dahliae were decreased by methyl bromide in 1971 but not by other treatments or in later years. In 1976, Pentland Dell and Pentland Crown were grown on an infested field in plots fumigated with methyl bromide. Fumigation slightly delayed shoot emergence but increased plant height, ground cover and the size and persistence of the leaves; development of haulm symptoms was delayed and yield increased. Fumigating with methyl bromide at five times the normal rate was more beneficial to Pentland Dell than to Pentland Crown. In a comparison of 10 cultivars in 1975, symptoms developed sooner and yields were smaller on a field infested with G. rostochiensis and V. dahliae than on a non-infested field and on both sites Maris Peer plants died soonest and yielded least. Pentland Crown yielded most on the non-infested field and Maris Piper on the infested field.     

Observations on apple canker; the anatomy of the stem canker

The development of cankers caused by Nectria galligena on apple is described; the pathogen exploits all the tissues outside the xylem and will also penetrate the xylem to an appreciable depth, invading the xylem parenchyma, vessels and fibres.
Spread in the peripheral tissues is checked to some extent by successive barriers laid down by wound phellogens, but the barriers are eventually passed by the mycelium. There appear to be two distinct ways in which these barriers are passed: there may be direct penetration which appears to be associated with an aggregation of the mycelium into blocks, and symptoms suggestive of mechanical rupture; or there is an alternative route in which the pathogen grows in the lumen of the fibres and emerges behind the barrier.
In the xylem the spread of the pathogen is checked by tyloses and gumming in the vessels, and by gumming in the parenchymatous tissues, but in the fibres there appears to be no defence mechanism, and the spread beyond the gum barriers was usually recorded in the xylem fibres. It is suggested that the presence of the pathogen in the xylem fibres might provide the explanation of the formation of cankers on partially healed pruning cuts, and this type of canker is described in some detail.
The development of the canker depends on the balance between the development of the pathogen and the resistance of the host, and there is some discussion on the relation of this balance to the control of the disease.     

Comparison of paint and gel formulations for the treatment of Nectria cankers on apple trees

A gel formulation containing carbendazim and triadimefon completely eradicated infection from scraped or unscraped cankers and none became re-infected. Paint formulations containing mercuric oxide or octhilinone completely eradicated infection from scraped cankers but 30–50% of unscraped cankers receiving these treatments became re-infected. Scraping alone encouraged callus tissue formation but did not greatly reduce infection. Fungicide moved from gels into the wood beneath cankers and persisted there for up to 6 months; movement of fungicide from mercuric oxide or octhilinone paints was minimal. Carbendazim was more effective than thiophanate-methyl and xanthan gum than sodium alginate or alginate ester as fungicide and gel component respectively. Rainfall shortly after treatment probably aided movement of fungicide from gels into the wood but salt formation, by incorporation of acid, did little to improve fungicide performance. Unequivocal assessment of efficacy of canker treatment requires data on canker area changes, movement and persistence of active ingredient in the wood and extent of re-infection after 12 months.     

Induced Phytoextraction of Lead Through Chemical Manipulation of Switchgrass and Corn; Role of Iron Supplement

The effects of combined chemical application of benomyl, ethylenedianinetetraacetate (EDTA), and iron (Fe) (foliar and root) on lead (Pb) phytoextraction by switchgrass (Panicum virgatum) and corn (Zea mays) was examined. Switchgrass was grown in Pb-contaminated urban topsoil with the following treatments: (C) Control, (B) benomyl, (E) EDTA, (F) foliar-Fe, (BE) benomyl + EDTA, (BF) benomyl + foliar-Fe, (FE) foliar-Fe + EDTA, (BFE) benomyl + foliar-Fe + EDTA. Corn was grown in sand-culture supplemented with Pb (500 mg kg^?1) with the following treatments: (C) control, (B) benomyl, (E) EDTA, (F) root-Fe, (BE) benomyl + EDTA, (BF) benomyl + root-Fe, (FE) root-iron + EDTA, and, (BFE) benomyl + root-Fe + EDTA. All treatments were replicated three times and pots were arranged in a completely randomized design. Plants were analyzed for element concentration (Fe, Zn, P, and Pb) using either inductively coupled plasma (argon) atomic emission spectroscopy (ICP-AES) or graphite furnace atomic absorption spectrometer. Iron supplementation (foliar and root) affected Pb-translocation in plants. Foliar-Fe treatment increased translocation ratio of Pb (TF-Pb) significantly compared to other treatments with the exception of plants treated with benomyl and BF. Root-Fe treatment in combination with EDTA (FE) increased TF-Pb significantly compared to other treatments. Phytoextraction was improved by the combined chemical application; plants treated with BFE treatment increased Pb-total-phytoextraction by 424% compared to Control plants.     

Differential transfer and dissemination of hypovirus and nuclear and mitochondrial genomes of a hypovirus-infected Cryphonectria parasitica strain after introduction into a natural population

Biological control of plant diseases generally requires release of living organisms into the environment. Cryphonectria hypoviruses function as biological control agents for the chestnut blight fungus, Cryphonectria parasitica, and hypovirus-infected C. parasitica strains can be used to treat infected trees. We used naturally occurring molecular marker polymorphisms to examine the persistence and dissemination of the three genomes of a hypovirus-infected C. parasitica strain, namely, the double-stranded RNA genome of Cryphonectria hypovirus 1 (CHV1) and the nuclear and mitochondrial genomes of its fungal host. The hypovirus-infected strain was experimentally introduced into a blight-infested chestnut coppice forest by treating 73 of 246 chestnut blight cankers. Two years after introduction, the hypovirus had disseminated to 36% of the untreated cankers and to 35% of the newly established cankers. Spread of the hypovirus was more frequent within treated sprout clusters than between sprout clusters. Mitochondrial DNA of the introduced fungus also was transferred into the resident C. parasitica population. Concomitant transfer of both the introduced hypovirus and mitochondrial DNA was detected in almost one-half of the treated cankers analyzed. The introduced mitochondrial DNA haplotype also was found in three resident isolates from newly established cankers. The nuclear genome of the introduced strain persisted in the treated cankers but did not spread beyond them.