Erwinia spp. from pome fruit trees: similarities and differences among pathogenic and non-pathogenic species

The number of described pathogenic and non-pathogenic Erwinia species associated with pome fruit trees, especially pear trees, has increased in recent years, but updated comparative information about their similarities and differences is scarce. The causal agent of the fire blight disease of rosaceous plants, Erwinia amylovora, is the most studied species of this genus. Recently described species that are pathogenic to pear trees include Erwinia pyrifoliae in Korea and Japan, Erwinia spp. in Japan, and Erwinia piriflorinigrans in Spain. E. pyrifoliae causes symptoms that are indistinguishable from those of fire blight in Asian pear trees, Erwinia spp. from Japan cause black lesions on several cultivars of pear trees, and E. piriflorinigrans causes necrosis of only pear blossoms. All these novel species share some phenotypic and genetic characteristics with E. amylovora. Non-pathogenic Erwinia species are Erwinia billingiae and Erwinia tasmaniensis that have also been described on pome fruits; however, less information is available on these species. We present an updated review on the phenotypic and molecular characteristics, habitat, pathogenicity, and epidemiology of E. amylovora, E. pyrifoliae, Erwinia spp. from Japan, E. piriflorinigrans, E. billingiae, and E. tasmaniensis. In addition, the interaction of these species with pome fruit trees is discussed.     

New, simple medium for selective, differential recovery of Klebsiella spp

A highly selective, differential medium for the enumeration and isolation of Klebsiella spp. was developed. With pure cultures, 100% recovery of Klebsiella spp. was observed. Recovery of Klebsiella spp. on MacConkey-inositol-potassium tellurite (MCIK) agar was as good as or better than on MacConkey-inositol-carbenicillin agar either with pure cultures or environmental samples. Recovery and percent colony confirmation with MCIK agar were greater and easier to obtain than for other proposed Klebsiella selective media.     

A differential medium for the isolation and rapid identification of a plant soft rot pathogen, Erwinia chrysanthemi

A medium was developed for the isolation and differentiation of Erwinia chrysanthemi from other Erwinia spp. based on the production of blue-pigmented indigoidine. The medium, named NGM, consists of nutrient agar supplemented with 1% glycerol, that induces pigment production, and 2 mM MnCl2*4H2O, that further enhances color development. More than fifty E. chrysanthemi strains from six different plant hosts were tested. All tested strains of E. chrysanthemi grew well on the NGM medium, developing dark brownish to blue colonies easily distinguishable from other Erwinia spp. The results indicate that pigment production on the NGM medium is a very stable property and can be used as a phenotypic property to differentiate E. chrysanthemi from other Erwinia spp. In addition, a specific oligonucleotide primer set was designed for the detection of indC, which is involved in indigoidine biosynthesis. All E. chrysanthemi strains tested contained indC as determined by PCR amplification. No amplification was observed with other Erwinia spp. Thus, pigment production of E. chrysanthemi on the NGM medium is consistent with the existence of indC. The NGM medium was used to isolate and identify the causal agent of soft rot lesions of diseased Phalaenopsis orchids from three orchid cultivation areas in Taiwan. The causal agents of Phalaenopsis soft rot were all identified as E. chrysanthemi. The results indicate that the NGM medium is efficient in isolation and identification of E. chrysanthemi from plants with soft rot symptoms and can also be used for epidemiological studies.     

New, simple medium for selective, differential recovery of Klebsiella spp.

A highly selective, differential medium for the enumeration and isolation of Klebsiella spp. was developed. With pure cultures, 100% recovery of Klebsiella spp. was observed. Recovery of Klebsiella spp. on MacConkey-inositol-potassium tellurite (MCIK) agar was as good as or better than on MacConkey-inositol-carbenicillin agar either with pure cultures or environmental samples. Recovery and percent colony confirmation with MCIK agar were greater and easier to obtain than for other proposed Klebsiella selective media.     

Functions and origin of plasmids in Erwinia species that are pathogenic to or epiphytically associated with pome fruit trees

The genus Erwinia includes plant-associated pathogenic and non-pathogenic species. Among them, all species pathogenic to pome fruit trees (E. amylovora, E. pyrifoliae, E. piriflorinigrans, Erwinia sp. from Japan) cause similar symptoms, but differ in their degrees of aggressiveness, i.e. in symptoms, host range or both. The presence of plasmids of similar size, in the range of 30 kb, is a common characteristic that they possess. Besides, they share some genetic content with high homology in several genes associated with exopolysaccharide production and hence, with virulence, as well as in some other genes. Knowledge of the content of these plasmids and comparative genetic analyses may provide interesting new clues to understanding the origin and evolution of these pathogens and the level of symptoms they produce. Furthermore, genetic similarities observed among some of the plasmids (and genomes) from the above indicated pathogenic species and E. tasmaniensis or E. billingiae, which are epiphytic on the same hosts, may reveal associations that could expose the mechanisms of origin of pathogens. A summary of the current information on their plasmids and the relationships among them is presented here.     

核果和仁果褐腐病病原菌研究进展

核果和仁果类果品在我国果品生产和消费中占有重要地位.由链核盘菌属Monilinia几种真菌引起的褐腐病可导致枝枯和果实腐烂,造成严重的经济损失.其中部分种类还是一些国家或地区的检疫性病原物.近年来褐腐病病原菌的组成、名称和分布已有新的变化,对链核盘菌和褐腐病的研究也取得了许多新进展.本文概述了过去20年来国内外对核果和...     

Improved medium for recovery and enumeration of the farmer's lung organism, Saccharomonospora viridis

A new medium, which we propose to call R8, was developed for the isolation and enumeration of the thermophilic actinomycete, Saccharomonospora viridis. This organism has been implicated in a range of hypersensitivity pneumonitides, including farmer's lung, but is generally isolated in small numbers from contaminated environments. Recovery of S. viridis from moldy hay and mushroom compost on R8 medium was compared with recovery on conventional media. S. viridis was isolated from both substrates but in highest numbers and most consistently on the R8 medium. The selectivity of this medium was best observed when the sedimentation chamber method was used for hay samples. Here S. viridis accounted for up to 80% of the total number of actinomycetes recovered on R8 and could not be recovered on rifampin selective medium under the same conditions. R8 was also found to be an efficient recovery medium for a range of thermophilic actinomycetes from mushroom compost and for another allergenic species, Faenia rectivirgula, from moldy hay. Contamination of isolation plates by thermophilic bacilli was reduced on R8 compared with the activity on half-strength tryptone soy agar, supplemented with 0.2% casein hydrolysate, and this, together with specific improvements in S. viridis growth, accounts for the selective effect. It is possible that the occurrence of S. viridis and its role as a causative agent of hypersensitivity pnuemonitis have been underestimated by the use of suboptimal recovery protocols. It is hoped that use of R8 in conjunction with dilution plate techniques will generate information on the ecology of S. viridis and contribute to health risk assessment studies.     

Improved medium for recovery and enumeration of Pseudomonas aeruginosa from water using membrane filters.

A modified mPA medium, designated mPA-C, was shown to recover Pseudomonas aeruginosa from a variety of water sources with results comparable to those with mPA-B and within the confidence limits of a most-probable-number technique. Enumeration of P. aeruginosa on mPA-C was possible after only 24 h of incubation at 41.5 degrees C, compared with 72 h of incubation required for mPA-B and 96 h of incubation for a presumptive most probable number.     

Improved medium for recovery and enumeration of Pseudomonas aeruginosa from water using membrane filters.

A modified mPA medium, designated mPA-C, was shown to recover Pseudomonas aeruginosa from a variety of water sources with results comparable to those with mPA-B and within the confidence limits of a most-probable-number technique. Enumeration of P. aeruginosa on mPA-C was possible after only 24 h of incubation at 41.5 degrees C, compared with 72 h of incubation required for mPA-B and 96 h of incubation for a presumptive most probable number.     

Influence of the nutrient medium on the recovery of dividing cells from tobacco protoplasts

Systematic tests resulted in a nutrient solution containing the following, in milligrams per liter, for the culture of protoplasts isolated from Nicotiana tabacum L. callus cells: Murashige and Skoog salts (T. Murashige and F. Skoog, 1962. Physiol. Plant. 15: 473-497); sucrose, 15,000; mannitol, 110,000; α-naphthaleneacetic acid, 0.6; kinetin, 0-0.1; thiamine·HCl, 10; pyridoxine·HCl, 10; nicotinic acid, 5; myo-inositol, 100; and glycine, 2. In this medium, regeneration of cell wall has been observed in 85% and resumption of cell division among 35% of the protoplast isolates.     

A simple and rapid high recovery protocol for the purification of arginase

Summary A protocol of purification is devised that gave high recovery of homogeneous arginase from ox erythrocytes. The protocol involved haemolysis, heat treatment, CM- and DEAE-Sepharose chromatography, arginine AH-Sepharose chromatography and molecular sieving through Biogel P-150, all in the presence of 2-mercaptoethanol. It afforded (a) elution of arginase as a single peak, (b) almost 2900 fold purification, (c) 30% recovery and (d) electrophoretically homogeneous arginase. The purified arginase exhibited (a) M.W. of 115,000, (b) dissociation upon SDS treatment into homologous monomers of 30 kDa, (c) optimum pH and temperature 11.5 and 55° C, respectively and (d) Km value for L-arginine-HCl 2.2–2.3 mM, characteristic of the ureotelic type.     

Instability of short-sequence DNA repeats of pear pathogenic Erwinia strains from Japan and Erwinia amylovora fruit tree and raspberry strains

An array of short-sequence DNA repeats (SSRs) occurs in the plasmid pEA29 of the fire blight pathogen Erwinia amylovora. A large number of "fruit tree" strains, mainly from Central and Western Europe, were screened for their SSR numbers, and the analyses were extended to five raspberry strains from North America and six pear pathogenic Erwinia strains from Japan. The repeat ATTACAGA present in all E. amylovorastrains was found to be reiterated 3 to 15 times. The Japanese strains contained the major repeat sequence GGATTCTG, which was reiterated 16 to 24 times. ATTACAGG, which resembles the SSR of E. amylovora, was reiterated two or three times. In a novel approach, sequencing gels were used to visualize the rare occurrence of shorter arrays (down to three repeats) in E. amylovoraand the Japanese Erwinia strains. Changes in the repeat numbers in E. amylovora were observed repeatedly when the bacteria had been exposed to stress conditions. The repeat structures of homo- and heteroduplices of PCR-amplified repeats were also analyzed by cleavage of annealed molecules with the single-strand-specific endonuclease from bacteriophage T4. Not only heteroduplexes, but also homoduplexes showed non-matching regions in the SSRs, which could arise from transient formation of loops due to strand slippage during the assays.     

Improved medium for recovery and enumeration of the farmer's lung organism, Saccharomonospora viridis.

A new medium, which we propose to call R8, was developed for the isolation and enumeration of the thermophilic actinomycete, Saccharomonospora viridis. This organism has been implicated in a range of hypersensitivity pneumonitides, including farmer's lung, but is generally isolated in small numbers from contaminated environments. Recovery of S. viridis from moldy hay and mushroom compost on R8 medium was compared with recovery on conventional media. S. viridis was isolated from both substrates but in highest numbers and most consistently on the R8 medium. The selectivity of this medium was best observed when the sedimentation chamber method was used for hay samples. Here S. viridis accounted for up to 80% of the total number of actinomycetes recovered on R8 and could not be recovered on rifampin selective medium under the same conditions. R8 was also found to be an efficient recovery medium for a range of thermophilic actinomycetes from mushroom compost and for another allergenic species, Faenia rectivirgula, from moldy hay. Contamination of isolation plates by thermophilic bacilli was reduced on R8 compared with the activity on half-strength tryptone soy agar, supplemented with 0.2% casein hydrolysate, and this, together with specific improvements in S. viridis growth, accounts for the selective effect. It is possible that the occurrence of S. viridis and its role as a causative agent of hypersensitivity pnuemonitis have been underestimated by the use of suboptimal recovery protocols. It is hoped that use of R8 in conjunction with dilution plate techniques will generate information on the ecology of S. viridis and contribute to health risk assessment studies.