Exploring the quantitative resistance to <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">phaseolicola</Emphasis> in common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L.)

Pseudomonas syringae pv. phaseolicola is an important disease that causes halo blight in common bean. The genetic mechanisms underlying quantitative halo blight resistance are poorly understood in this species, as most disease studies have focused on qualitative resistance. The present work examines the genetic basis of quantitative resistance to the nine halo blight races in different organs (primary and trifoliate leaf, stem and pod) of an Andean recombinant inbred line (RIL) progeny. Using a multi-environment quantitative trait locus (QTL) mapping approach, 76 and 101 main-effect and epistatic QTLs were identified, respectively. Most of the epistatic interactions detected were due to loci without detectable QTL additive main effects. Main and epistatic QTLs detected were mainly consistent across the environment conditions. The homologous genomic regions corresponding to 26 of the 76 main-effect detected QTLs were positive for the presence of resistance-associated gene cluster encoding nucleotide-binding and leucine-rich repeat (NL) proteins and known defence genes. Main-effect QTLs for resistance to races 3, 4 and 5 in leaf, stem and pod were located on chromosome 2 within a 3.01-Mb region, where a cluster of nine NL genes was detected. The NL gene Phvul.002G323300 is located in this region, which can be considered an important putative candidate gene for the non-organ-specific QTL identified here. The present research provides essential information not only for the better understanding of the plant-pathogen interaction but also for the application of genomic assisted breeding for halo blight resistance in common bean.     

ANTIFUNGAL ACTION OF STREPTOMYCIN-COPPER CHELATE AGAINST PHYTOPHTHORA INFESTANS ON TOMATO (LYCOPERSICON ESCULENTUM)

The antifungal protection afforded by streptomycin-copper chelate in vivo in greenhouse tests against Phytophthora infestans on tomato was found to be of the order of six times greater than that of streptomycin sulphate. The copper chelate also showed much greater resistance to simulated rainfall. Similar tests showed that streptomycin-copper chelate (in terms of streptomycin content) was from 2 1/2 to 5 times more effective as an antifungal prophylactic spray than zinc ethylene-bis(dithiocarbamate) (zineb), and some fifty times more active than copper oxychloride formulated as a commercial 50% wettable powder. These three materials showed roughly similar resistance to weathering by 'rainfall' and it was assumed therefore that 250 i.u./ml. streptomycin as streptomycin-copper chelate might be expected to give the same degree of protection against Phytophthora infestans in the field as would the usual field rates of usage of zineb and copper oxychloride.     

Improvement of the efficacy of linear undecapeptides against plant-pathogenic bacteria by incorporation of D-amino acids

A set of 31 undecapeptides, incorporating 1 to 11 d-amino acids and derived from the antimicrobial peptide BP100 (KKLFKKILKYL-NH(2)), was designed and synthesized. This set was evaluated for inhibition of growth of the plant-pathogenic bacteria Erwinia amylovora, Pseudomonas syringae pv. syringae, and Xanthomonas axonopodis pv. vesicatoria, hemolysis, and protease degradation. Two derivatives were as active as BP100, and 10 peptides displayed improved activity, with the all-d isomer being the most active. Twenty-six peptides were less hemolytic than BP100, and all peptides were more stable against protease degradation. Plant extracts inhibited the activity of BP100 as well as that of the d-isomers. Ten derivatives incorporating one d-amino acid each were tested in an infectivity inhibition assay with the three plant-pathogenic bacteria by using detached pear and pepper leaves and pear fruits. All 10 peptides studied were active against E. amylovora, 6 displayed activity against P. syringae pv. syringae, and 2 displayed activity against X. axonopodis pv. vesicatoria. Peptides BP143 (KKLFKKILKYL-NH(2)) and BP145 (KKLFKKILKYL-NH(2)), containing one d-amino acid at positions 4 and 2 (underlined), respectively, were evaluated in whole-plant assays for the control of bacterial blight of pepper and pear and fire blight of pear. Peptide BP143 was as effective as streptomycin in the three pathosystems, was more effective than BP100 against bacterial blight of pepper and pear, and equally effective against fire blight of pear.     

Activity of some aminoglycoside antibiotics against true fungi, Phytophthora and Pythium species

AIMS: To investigate the in vitro antifungal and antioomycete activities of some aminoglycosides against true fungi and Phytophthora and Pythium species and to evaluate the potential of the antibiotics against Phytophthora late blight on plants. METHODS AND RESULTS: Antifungal and antioomycete activities of aminoglycoside antibiotics (neomycin, paromomycin, ribostamycin and streptomycin) and a paromomycin-producing strain (Streptomyces sp. AMG-P1) against Phytophthora and Pythium species and 10 common fungi were measured in potato dextrose broth (PDB) and on seedlings in pots. Paromomycin was the most active against Phytophthora and Pythium species with a minimal inhibitory concentration of 1-10 microg ml(-1) in PDB, but displayed low to moderate activities towards other common fungi at the same concentration. Paromomycin also showed potent in vivo activity against red pepper and tomato late blight diseases with 80 and 99% control value, respectively, at 100 microg ml(-1). In addition, culture broth of Streptomyces sp. AMG-P1 as a paromomycin producer exhibited high in vivo activity against late blight at 500 microg freeze-dried weight per millilitre. CONCLUSIONS: Among tested aminoglycoside antibiotics, paromomycin was the most active against oomycetes both in vitro and in vivo. SIGNIFICANCE AND IMPACT OF THE STUDY: Data from this study show that aminoglycoside antibiotics have in vitro and in vivo activities against oomycetes, suggesting that Streptomyces sp. AMG-P1 may be used as a biocontrol agent against oomycete diseases.     

THE EFFECTS OF STREPTOMYCIN AND TERRAMYCIN, SINGLY AND IN COMBINATION, ON THE LEAF BLIGHT DISEASE OF MAIZE CAUSED BY BACTERIUM CAROTOVORUUM F. ZEAE SABET

The growth in vitro of Bacterium carotovorum f. zeae Sabet, the cause of the root and stalk rot and leaf blight disease of maize, was inhibited by dihydrostreptiomycin sulphate and by Terramycin (oxytetracycline). Joint applications showed a synergistic action, and the bacterium was most susceptible to a 1:1 mixture of the two antibiotics. This mixture is effective in checking the development of resistant strains which develop rapidly after treatment with streptomycin alone.
Roots of maize seedlings in water cultures, and leaves of older plants readily absorb both antibiotics. High water content of soil favoured absorption; sunlight had no effect. Absorption from leaf applications first increased with higher concentration and frequency of spraying, but after eight daily applications of streptomycin toxic symptoms appeared and there was evidence that the antibiotic was inactivated.
Antibiotic treatment of maize leaves shortly before inoculation with B. carotovorum f. zeae reduced the incidence and severity of leaf blight. The therapeutic effect extends at first both downward and upward but later only upward. Treatment of the leaves did not induce resistance to infection in the stems and roots.     

The action of streptomycin in a mutant of Escherichia coli with increased sensitivity to the antibiotic

A mutant of Escherichia coli with increased sensitivity to streptomycin has been studied. This strain differed from a normal str(s) strain in that streptomycin produced inhibition of protein synthesis and loss of viability with almost no lag period. Chloramphenicol protected a normal str(s) strain but not the mutant against the bactericidal action of streptomycin. The results obtained support the idea that access of streptomycin to its site of action in a normal cell is restricted, and that this restriction, which is much less effective in the mutant, probably involves a permeability barrier. Comparison of the inhibition of protein synthesis by streptomycin with concomitant changes in the distribution of polyribosomes in both strains suggested that the antibiotic can directly inhibit the translation of mRNA.     

Antibiotic treatment of ulcerative enteritis of bobwhite quail

Penicillin-streptomycin, bacitracin methylene disalicylate, zinc bacitracinm, soluble bacitracin methylene disalicylate, streptomycin (25%) and spectinomycin were used as prophylactic and therapeutic treatments for ulcerative enteritis in bobwhite quail (Colinus virginianus). Penicillin-streptomycin was the most effective prophylactic and streptomycin (25%) was the most effective therapeutic agent.     

Streptomycin Application Has No Detectable Effect on Bacterial Community Structure in Apple Orchard Soil

Streptomycin is commonly used to control fire blight disease on apple trees. Although the practice has incited controversy, little is known about its nontarget effects in the environment. We investigated the impact of aerial application of streptomycin on nontarget bacterial communities in soil beneath streptomycin-treated and untreated trees in a commercial apple orchard. Soil samples were collected in two consecutive years at 4 or 10 days before spraying streptomycin and 8 or 9 days after the final spray. Three sources of microbial DNA were profiled using tag-pyrosequencing of 16S rRNA genes: uncultured bacteria from the soil (culture independent) and bacteria cultured on unamended or streptomycin-amended (15 μg/ml) media. Multivariate tests for differences in community structure, Shannon diversity, and Pielou''s evenness test results showed no evidence of community response to streptomycin. The results indicate that use of streptomycin for disease management has minimal, if any, immediate effect on apple orchard soil bacterial communities. This study contributes to the profile of an agroecosystem in which antibiotic use for disease prevention appears to have minimal consequences for nontarget bacteria.     

THE SYSTEMIC INSECTICIDAL ACTION OF SODIUM FLUOROACETATE AND OF THREE PHOSPHORUS COMPOUNDS ON THE EGGS AND LARVAE OF PIERIS BRASSICAE L.

Four compounds, bis dimethylaminophosphonous anhydride ( anhydride ), bis (dimethylamino)fluorophosphine oxide ( oxide ), diethyl paranitrophenyl phosphate ( E 600) and sodium fluoroacetate ( acetate ), previously shown to have systemic insecticidal activity against aphids, have now been tested against the eggs and larvae of Pieris brassicae L.
The anhydride proved to have very little toxic action on Pieris , but the other three compounds showed both contact toxicity and systemic insecticidal action when taken up by the roots of cabbage plants from solution and from soil. The acetate, but more especially the E600, also showed systemic action following application to the leaves.
In all cases the order of decreasing toxicity was E600 > acetate > oxide > anhydride. E600 is the only compound which is outstandingly toxic to Pieris eggs and larvae. It has the added interest that when watered on to the roots of cabbage plants it kills larvae in egg batches on the leaves. Death takes place at the stage when the larvae are biting through the shells. The oxide and acetate proved to be surprisingly innocuous.
Unless it is felt to be too poisonous E600 is worthy of consideration as an insecticide against Pieris eggs and larvae, since it is highly effective as a contact insecticide and also has some systemic action.     

Chromogenic plate assay distinguishing bacteriolytic from bacteriostatic activity of an antibiotic agent

A solid agar plate assay was devised to discriminate bacteriolytic from bacteriostatic activity for a given antibacterial agent. The assay uses a bacterial culture harboring β-galactosidase enzyme as reporter of cellular lysis. When a drop of bacteriolytic compound is placed on the agar, β-galactosidase is released from the bacteria to the external solid medium where it hydrolyzes X-Gal substrate analogue, developing a blue halo at the edge of the inhibition growth zone. The assay was successfully evaluated against several antibiotics with well-known mechanism of action. It was found that bacteriostatic compounds consistently did not display blue halo at the inhibition zone.     

Phenotypic and genetic diversity of Erwinia amylovora: the causal agent of fire blight

Erwinia amylovora is a polyphagous bacterium causing fire blight on apple, pear and over 130 other plant species belonging mainly to the Rosaceae family. Although E. amylovora is regarded as a very homogenous species, the particular strains can differ in pathogenic ability as far as their host range is concerned (e.g. those originating from Rubus or Maloidae plants) as well as by the extent of the disease they cause. It was found that strains originating from North America are generally more genetically heterogeneous than those from Europe. Diversity of E. amylovora is also related to streptomycin resistance as a result of its application to control of fire blight. The level of genetic heterogeneity of E. amylovora is so low (comparative genome analysis revealed a similarity of over 99% for the two genomes tested) that standard DNA-based techniques fail in detection of intra-species variability. Amplified fragment length polymorphism was found to be most useful for differentiation of strains of fire blight causal agent as well as techniques ensuing release of pan-genome sequences of two E. amylovora strains: multi-locus variable number of tandem repeats analysis and clustered regularly interspaced short palindrome repeats.     

Antibacterial Activity of Tea and Coffee Wastes Against Some Plant Pathogenic Pseudomonas syringae Strains

In vitro and greenhouse trials were conducted to elucidate the potential use of extracts of tea and coffee wastes to control plant diseases caused by the bacterial pathogens, Pseudomonas syringae pv. pisi (race 1 and 2) and P. s. pv. phaseolicola (race 1). The antibacterial activity was measured as the diameter of the inhibition zone in agar and also by periodical viable cell counts in laboratory tests. The effect on the hypersensitive reaction and the potential for disease control after leaf infiltration and seed treatment were studied on bean plants in the greenhouse. Results showed that both the tea and coffee extracts possessed antibacterial activity against the three pathogens, but that the effects varied depending on the strain and the test method. Strong reduction of halo blight disease and improvement in plant growth was obtained in presence of the coffee extract. For the halo blight pathogen, P. s. pv. phaseolicola, there was a good correlation between the results from the viable cell count method and the greenhouse tests, but the results from the in vitro studies did not agree with those from greenhouse as regards the P. s. pv. pisi strains. It is suggested that the component(s) in tea and coffee responsible for controlling the bean pathogen may not be the same as that for the pea pathogens.     

The prophylactic administration of interleukin-2 increases the survival of mice with an acute intra-abdominal infection caused by Staphylococcus aureus]

The effect of RIL-2 on the survival of mice with acute Staphylococcus aureus strain 5/2 intra-abdominal [correction of intraperitoneal] infection was studied. RIL-2 was ineffective when administered simultaneously with the LD100 dose of bacteria. Antibiotics (gentamycin or combination of penicillin and streptomycin) administered in the same fashion cured 100% of animals. However, RIL-2 proved to be effective when administered simultaneously with LD70 dose of bacteria. The prophylactic course of RIL-2 consisting of repeated injections on days 3, 2 and 1 before the challenge with LD100 dose of bacteria also resulted in the marked increase of the survival of mice. The hypothetical mechanisms of action and the prospects of RIL-2 application are discussed.     

Biocontrol potential of phylloplane bacterium Ochrobactrum anthropi BMO‐111 against blister blight disease of tea

Aims

The present study was carried out to screen the phylloplane bacteria from tea for antagonism against grey blight caused by Pestalotiopsis theae and blister bight caused by Exobasidium vexans and to further evaluate the efficient isolates for disease control potential under field condition.

Methods and Results

A total of 316 morphologically different phylloplane bacteria were isolated. Among the antagonists, the isolates designated as BMO‐075, BMO‐111 and BMO‐147 exhibited maximum inhibitory activity against both the pathogens under in vitro conditions and hence were selected for further evaluation under microplot field trial. Foliar application of 36‐h‐old culture of BMO‐111 (1 × 10⁸ colony‐forming units ml^?1) significantly reduced the blister blight disease incidence than the other isolates. The culture of BMO‐111 as well as its culture filtrate effectively inhibited the mycelial growth of various fungal plant pathogens. The isolate BMO‐111 was identified as Ochrobactrum anthropi based on the morphological and 16S rDNA sequence analyses.

Conclusions

It could be concluded that the biocontrol agent O. anthropi BMO‐111 was effective against blister blight disease of tea.

Significance and Impact of the Study

Further study is required to demonstrate the mechanism of its action and formulation for the biocontrol potential against blister blight disease of tea.     

Involvement of phaseolotoxin in halo blight of beans: transport and conversion to functional toxin

Phaseolotoxin ([N^δ-phosphosulfamyl]ornithylalanylhomoarginine) is produced by Pseudomonas phaseolicola (Burkh.) Dows. in liquid culture. When phaseolotoxin was applied to leaves of bean (Phaseolus vulgaris L.) at 0.1 to 1 nmoles/g fresh weight of leaf by a prick-assay procedure, the characteristic “halo” symptom of bean halo blight disease developed after 24 to 48 hours. At higher concentrations (10-100 nmoles/g fresh weight) the systemic symptoms, which are commonly a feature of diseased plants, also developed after 24 to 48 hours.     

Prevention of the disturbance of protein synthesis is a possible mechanism of the interferon protective activity.

Interferon and interferon inducer (Newcastle disease virus), were shown to enhance the cell resistance against the damaging effects of streptomycin. The preliminary intravenous injection of NDV to mice resulted in a diminishing of both qualitative and quantitative changes in activity of the enzyme myeloperoxydase caused by streptomycin in leucocytes from peritoneal exudate. Protective effect was conferred also by immediate pretreatment of mouse leucocytes with homologous interferon. The data presented suggest that protective action of interferon involves the control of normal protein synthesis in the cell.     

A pathogen-responsive cDNA from potato encodes a protein with homology to a phosphate starvation-induced phosphatase

Infiltration of potato leaves with the phytopathogenic bacteria Pseudomonas syringae pv. maculicola induces local and systemic defense gene expression as well as increased resistance against subsequent pathogen attacks. By cDNA-AFLP a gene was identified that is activated locally in potato leaves in response to bacterial infiltration and after infection with Phytophthora infestans, the causal agent of late blight disease. The encoded protein has high homology to a phosphate starvation-induced acid phosphatase from tomato. Possibly, decreased phosphate availability after pathogen infection acts as a signal for the activation of the potato phosphatase gene.     

Effectiveness of acibenzolar-S-methyl, fungicides and antibiotics for the control of brown eye spot, bacterial blight, brown leaf spot and coffee rust in coffee

This study was carried out to evaluate the potential of acibenzolar‐S‐methyl (ASM), combined or not combined with fungicides and antibiotics for the control of brown eye spot (Cercospora coffeicola) and bacterial blight (Pseudomonas syringae pv. garcae) in coffee seedlings, and ASM combined with conventional fungicide application schedules for the control of coffee rust (Hemileia vastatrix) and brown leaf spot (Phoma costarricencis) under field conditions in two coffee crops in the State of São Paulo, Brazil. ASM protected coffee seedlings against C. coffeicola when applied at the rates of 2.5 and 5 g of active ingredient per hectolitre of water (g a.i. hL^?1), providing 34–55% of disease control, and against bacterial blight, when applied at the rates of 2.5, 10 and 20 g a.i. hL^?1, with 38–57% of disease control. Tebuconazole (100 g a.i. hL^?1) and azoxystrobin (10 g a.i. hL^?1) showed the best results for brown eye spot control. Oxytetracycline + streptomycin, kasugamycin hydrochloride, oxytetracycline + metallic copper, copper oxychloride and mancozeb + copper oxychloride also controlled bacterial blight in levels similar to those shown by ASM. In the field experiments, all fungicide application schedules tested, cyproconazole (December, February, April), epoxiconazole (December, March), tetraconazole (December, February, April), cyproconazole (December, February) and azoxystrobin (January, March) were effective for coffee rust control and provided partial control of brown leaf spot. The results also showed that for all experiments, there was no synergistic effect of the combination of ASM with azoxystrobin, cyproconazole or cupric fungicides.     

Pathovar-specific requirement for the Pseudomonas syringae lemA gene in disease lesion formation.

The lemA gene is conserved among strains and pathovars of Pseudomonas syringae. In P. syringae pv. syringae B728a, a causal agent of bacterial brown spot disese of bean, the lemA gene is required for lesion formation on leaves and pods. Using lemA-containing DNA as a probe, we determined that 80 P. syringae pv. syringae strains isolated from bean leaves could be grouped into seven classes based on restriction fragment length polymorphism. Marker exchange mutagenesis showed that the lemA gene was required for lesion formation by representative strains from each restriction fragment length polymorphism class. Hybridization to the lemA locus was detected within six different P. syringae pathovars and within Pseudomonas aeruginosa. Interestingly, a lemA homolog was present and functional within the nonpathogenic strain P. syringae Cit7. We cloned a lemA homolog from a genomic library of P. syringae pv. phaseolicola NPS3121, a causal agent of halo blight of bean, that restored lesion formation to a P. syringae pv. syringae lemA mutant. However, a lemA mutant P. syringae pv. phaseolicola strain retained the ability to produce halo blight disease symptoms on bean plants. Therefore, the lemA gene played an essential role in disease lesion formation by P. syringae pv. syringae isolates, but was not required for pathogenicity of a P. syringae pv. phaseolicola strain.     

Biological control of common blight of bean (Phaseolus vulgaris) causedby Xanthomonas axonopodis pv. phaseoli by using the bacteriumRahnella aquatilis

The aim of this study was to evaluate the bacterium Rahnella aquatilis (Ra) for protection of bean plants against common blight disease caused by Xanthomonas axonopodis pv. phaseoli (Xap). Xap isolates were isolated from a naturally blighted leaves of bean plants grown in Assiut governorate. The blight symptoms were produced by all three isolates, but the isolates differed in their degree of the pathogenicity. Xap1 was the most virulence one against bean plants. The effect of Ra against common blight of bean plant was tested. In vitro studies, Ra exhibited inhibitor effect against the pathogen. Under greenhouse and field conditions, beanvariety “Giza 6” treated by Ra resulted in marked disease suppression. Ahigh decrease of the disease was correlated with a reduction of the bacterial multiplication. In physiological studies, bean plants treated by Ra exhibited higher phenolic compounds contents and higher activity of peroxidase (PO) enzyme than untreated plants. In conclusion, application of Ra was effective and could be recommended for controlling the bean common blight disease.