Rhizosphere competent <Emphasis Type="Italic">Pantoea agglomerans</Emphasis> enhances maize (<Emphasis Type="Italic">Zea mays</Emphasis>) and chickpea (<Emphasis Type="Italic">Cicer arietinum</Emphasis> L.) growth,without altering the rhizosphere functional diversity

Plant growth promoting Pantoea agglomerans NBRISRM (NBRISRM) was able to produce 60.4 μg/ml indole acetic acid and solubilize 77.5 μg/ml tri-calcium phosphate under in vitro conditions. Addition of 2% NaCl (w/v) in the media induced the IAA production and phosphate solubilization by 11% and 7%, respectively. For evaluating the plant growth promotory effect of NBRISRM inoculation a micro plot trial was conducted using maize and chickpea as host plants. The results revealed significant increase in all growth parameters tested in NBRISRM inoculated maize and chickpea plants, which were further confirmed by higher macronutrients (N, P and K) accumulation as compared to un-inoculated controls. Throughout the growing season of maize and chickpea, rhizosphere population of NBRISRM were in the range 10⁷–10⁸ CFU/g soil and competing with 10⁷–10⁹ CFU/g soil with heterogeneous bacterial population. Functional richness, diversity, and evenness were found significantly higher in maize rhizosphere as compared to chickpea, whereas NBRISRM inoculation were not able to change it, in both crops as compared to their un-inoculated control. To the best of our knowledge this is first report where we demonstrated the effect of P. agglomerans strain for improving maize and chickpea growth without altering the functional diversity.     

Distribution of Coliphages Against Four <Emphasis Type="Italic">E. Coli</Emphasis> Virotypes in Hospital Originated Sewage Sample and a Sewage Treatment Plant in Bangladesh

The distribution of coliphages infecting different Escherichia coli virotypes (EHEC, EIEC, EPEC, ETEC) and an avirulent strain (K-12) in sewage system of a hospital and a sewage treatment plant (STP) was investigated by culture-based agar overlay methods. Coliphages were found in all the samples except stool dumping site in the sewage system of the hospital and lagoon of the STP. Bacteriophage count (pfu/ml) infecting E. coli strains showed the following ascending pattern (EHEC < EIEC < EPEC < ETEC < E coli K-12) in all the collected samples except one. Phages capable of infecting avirulent E. coli K-12 strains were present in the highest number among all the examined locations. Phages specific for E. coli K-12 presented high diversity in plaque size on the bacterial lawn. Virulent E. coli specific coliphages rarely produced plaques with diameter of 1–2 mm or over. Conventional agar overlay method was found to be not satisfactory for phage community analysis from primary stool dumping site of the hospital, probably due to the presence of high concentration of antimicrobial substances. The gradual decrease seen in the five groups of coliphage quantity with the ongoing treatment process and then the absolute absence of coliphages in the outlet of the examined treatment plant is indicative of the usefulness of the treatment processes practiced there.     

Chemotaxis of free-living nitrogen-fixing bacteria towards maize mucilage

Summary A method for collecting sterile mucilage from maize root tips growing in sterile conditions has been devised.Enterobacter andAzospirillum strains were isolated from the rhizosphere of maize and rice using the spermosphere model method. To evaluate chemotaxis of these strains, a modification of Adler's microcapillary method was used. Under these conditions, the number of attracted bacteria was proportional to the concentration of mucilage. When comparing the chemotaxis ofA. lipoferum andE. cloacae from the rhizosphere of maize and from the rhizosphere of rice, it appeared that the strains isolated from maize were strongly attracted by maize mucilage whereas strains isolated from rice were not more attracted than the control (E. coli K12). Thus, bacteria of the same species are not equivalent in their chemotactic behaviour. This could imply that some degree of specificity exists in the establishment of plant-bacteria associations.     

Growth promotion of maize and lettuce by phosphate-solubilizing Rhizobium leguminosarum biovar. phaseoli

Rhizobium leguminosarum bv. phaseoli strains P31 and R1, Serratia sp. strain 22b, Pseudomonas sp. strain 24 and Rhizopus sp. strain 68 were examined for their plant growth-promoting potential on lettuce and forage maize. All these phosphate solubilizing microorganisms (PSM) were isolated from Québec soils. The plants were grown in field conditions in three sites having high to low amounts of available P. In site 1 (very fertile soil), strains R1 and 22b tended to increase the dry matter yield of lettuce shoots (p≤0.10). Lettuce inoculated with rhizobia R1 had a 6% higher P concentration (p≤0.10) than the uninoculated control. In site 2 (poorly fertile soil), the dry matter of lettuce shoots was significantly increased (p≤0.05) by inoculation with strain P31 and 24 plus 35 kg ha^-1 P-superphosphate, or with strain 68 plus 70 kg ha^-1 P-superphosphate. In site 3 (moderately fertile soil), the dry matter of maize shoots was significantly increased (p≤0.05) by inoculation with strain 24 plus 17.5 kg ha^-1 P-superphosphate, or with strain P31 plus 35 kg ha^-1 P-superphosphate. Inoculation with PSM did not affect lettuce P uptake in the less fertile soil in site 2. In site 3 with the moderately fertile soil, maize plants inoculated with strain R1 had 8% higher P concentration than the uninoculated control (p≤0.01), and 6% with strains P31 and 68 (p≤0.05). The results clearly demonstrate that rhizobia specifically selected for P solubilization function as plant growth promoting rhizobacteria with the nonlegumes lettuce and maize. The P solubilization effect seems to be the most important mechanism of plant growth promotion in moderately fertile and very fertile soils when P uptake was increased with rhizobia and other PSM.     

Soil Type and Maize Cultivar Affect the Genetic Diversity of Maize Root–Associated Burkholderia cepacia Populations

Abstract Burkholderia cepacia populations associated with the Zea mays root system were investigated to assess the influence of soil type, maize cultivar, and root localization on the degree of their genetic diversity. A total of 180 B. cepacia isolates were identified by restriction analysis of the amplified 16S rDNA (ARDRA technique). The genetic diversity among B. cepacia isolates was analyzed by the random amplified polymorphic DNA (RAPD) technique, using the 10-mer primer AP5. The analysis of molecular variance (AMOVA) method was applied to estimate the variance components for the RAPD patterns. The results indicated that, among the factors studied, the soil was clearly the dominant one in affecting the genetic diversity of maize root–associated B. cepacia populations. In fact, the percentage of variation among populations was significantly higher between B. cepacia populations recovered from maize planted in different soils than between B. cepacia populations isolated from different maize cultivars and from distinct root compartments such as rhizoplane and rhizosphere. The analysis of the genetic relationships among B. cepacia isolates resulted in dendrograms showing bacterial populations with frequent recombinations and a nonclonal genetic structure. The dendrograms were also in agreement with the AMOVA results. We were able to group strains obtained from distinct soils on the basis of their origin, confirming that soil type had the major effect on the degree of genetic diversity of the maize root–associated B. cepacia populations analyzed. On the other hand, strains isolated from distinct root compartments exhibited a random distribution which confirmed that the rhizosphere and rhizoplane populations analyzed did not significantly differ in their genetic structure. Received: 22 January 1999; Accepted: 7 April 1999     

Survival of lux-marked bacteria introduced into soil and the rhizosphere of bean (Phaseolus vulgaris L.)

The survival of lux-marked recombinants of Escherichia coli and Bacillus subtilis was studied in the rhizosphere of bean (Phaseolus vulgaris L.) and in bulk soil. The number of E. coli (pSB343) containing a complete lux operon did not differ significantly according to whether they were introduced into soil separately or together with a non-luminescent mutant Pseudomonas fluorescens R2fN. When genetically altered strains of E. coli and B. subtilis bearing a complete or an incomplete lux-reporter system were introduced into soil, the numbers of surviving cells were the same both in the rhizosphere and bulk soil. The insertion of lux genes into bacterial strains therefore does not affect their competitiveness and survival in the rhizosphere and bulk soil.The author is with the Department of Microbiology, University of Silesia, Jagielloska 28, 40-032 Katowice, Poland     

Evaluation of Petrifilm™ EC method for enumeration of E. coli from soil

Aims: To evaluate the suitability of commercially available Petrifilm? EC plates for enumeration of Escherichia coli from soil. Methods and Results: A confirmed E. coli strain isolated from liquid swine manure was inoculated into sterilized sandy clay loam and loam soils at the concentrations of 10², 10³, 10⁵ CFU g^?1 of soil. The efficiency of recovery on Petrifilm? EC plates for soils spiked with E. coli was compared with standard membrane filtration techniques on m‐FC basal medium supplemented with 3‐bromo‐4‐chloro‐5‐indoyl‐β‐d ‐glucopyranoside (BCIG) and most probable numbers (MPN) techniques in E. coli medium with 4‐methylumbelliferyl‐β‐d ‐glucuronide (EC‐MUG) broth. Petrifilm? EC and m‐FC (BCIG) methods were then assessed for the ability to recover E. coli from field soils applied with swine manure. No significant differences (P > 0·05) were observed between Petrifilm? EC, m‐FC (BCIG) and MPN methods for the recovery of E. coli from spiked samples, irrespective of soil type. However, recovery of E. coli from manure‐applied field soil samples showed a significant difference (P < 0·05) between the Petrifilm? EC method and the m‐FC method in enumerating E. coli possibly as a result of false positives on m‐FC. Conclusion: The Petrifilm? EC method is suitable for the enumeration of E. coli from soil with a detection limit of 10 CFU g^?1 soil. Significance and Impact of the Study: The commercially available Petrifilm? EC method is comparatively low cost, easy to use method for the enumeration of E. coli from soil without the need for further confirmation tests.     

Effect of iclR and arcA deletions on physiology and metabolic fluxes in Escherichia coli BL21 (DE3)

Deletion of both iclR and arcA in E. coli profoundly alters the central metabolic fluxes and decreases acetate excretion by 70%. In this study we investigate the metabolic consequences of both deletions in E. coli BL21 (DE3). No significant differences in biomass yields, acetate yields, CO₂ yields and metabolic fluxes could be observed between the wild type strain E. coli BL21 (DE3) and the double-knockout strain E. coli BL21 (DE3) ΔarcAΔiclR. This proves that arcA and iclR are poorly active in the BL21 wild type strain. Noteworthy, both strains co-assimilate glucose and acetate at high glucose concentrations (10–15 g l⁻¹), while this was never observed in K12 strains. This implies that catabolite repression is less intense in BL21 strains compared to in E. coli K12.     

Microbial Inoculants with Multifaceted Traits Suppress Rhizoctonia Populations and Promote Plant Growth in Cotton

The suppressive effects of microbial inoculants on cotton seedling mortality were assessed in Rhizoctonia solani‐infested soil. Per cent mortality ranged from 16 to 32 (60–120 days after sowing, DAS) and significant differences were recorded at 120 DAS, especially after drenching with compost tea of Azotobacter sp. and Anabaena torulosa—Trichoderma viride‐biofilmed formulations. The activity of hydrolytic enzymes was reduced in diseased root tissues due to a majority of the microbially inoculated treatments, compared with healthy root tissues. Per cent changes in the amounts of glomalin‐related soil proteins (GRSPs) were 2 to 85% greater than those of the uninoculated experimental controls. These microbial inoculants altered the rhizosphere bacterial communities as evident from the Denaturing gradient gel electrophoresis (DGGE) banding patterns and, also reduced the population of R. solani. While the copy numbers of the internal transcribed spacer (ITS) gene of R. solani in the uninoculated (infested soil) were approximately 1.47 × 10¹¹ per g soil, they were 1.34–1.42 × 10⁵ per g soil after the application of A. torulosa, Anabaena laxa and A. torulosa–Bacillus sp. Increases in yield (ranging from 3 to 23%) due to various microbial inoculants relative to uninoculated controls illustrated their promise as plant growth‐promoting and disease‐suppressing agents. This study illustrates the modulation of rhizosphere ecology through microbial inoculants as a mechanism of disease suppression and sustaining plant growth.     

Biodiversity of diazotrophic bacteria within the soil, root and stem of field-grown maize

Recent studies suggest a high diversity of diazotrophic bacteria in maize. However, none of these works have been based on a sufficient number of samples to provide reasonable quantitative estimates of diazotrophic bacterial diversity. Here we present the use of molecular tools and statistical inference to assess diazotrophic bacterial diversity within rhizosphere soils, roots and stems of field grown maize. DNA was isolated from the latter collected from six maize growing regions within the southern most state in Brazil, Rio Grande do Sul. Using conserved primers, nifH Cluster I gene fragments were amplified from each of the three zones, and the products cloned and sequenced. The majority of the sequences were classified within the Proteobacteria with the α-proteobacteria and β-proteobacteria being the most abundant in the rhizosphere soil and stem samples. The γ-proteobacteria were most abundant in rhizosphere soils, less so in roots, and least in the stem samples. According to three different diversity measures, the rhizosphere soil samples possessed greater diazotrophic bacterial diversity than the roots and stems of the maize plants. Only two genera, Azospirillum and Azotobacter, were found in virtually all samples at an abundance of over 1% of the total nifH sequences obtained. Other genera were largely restricted to soil (Methylocystis, Beijerinckia, Geobacter, Rhodovulum, Methylobacterium, Gluconacetobacter, Methylocella, and Delftia), roots (Dechloromonas), or stems (Methylosinus, Raoultella, and Rhizobium). Three genera, Herbaspirillum, Ideonella, and Klebsiella, appeared to dominate in the interior of the plant but were much rarer in soil.     

The legacy of bacterial invasions on soil native communities

Soil microbial communities are often not resistant to the impact caused by microbial invasions, both in terms of structure and functionality, but it remains unclear whether these changes persist over time. Here, we used three strains of Escherichia coli O157:H7 (E. coli O157:H7), a species used for modelling bacterial invasions, to evaluate the resilience of the bacterial communities from four Chinese soils to invasion. The impact of E. coli O157:H7 strains on soil native communities was tracked for 120 days by analysing bacterial community composition as well as their metabolic potential. We showed that soil native communities were not resistant to invasion, as demonstrated by a decline in bacterial diversity and shifts in bacterial composition in all treatments. The resilience of native bacterial communities (diversity and composition) was inversely correlated with invader's persistence in soils (R² = 0.487, p < 0.001). Microbial invasions also impacted the functionality of the soil communities (niche breadth and community niche), the degree of resilience being dependent on soil or native community diversity. Collectively, our results indicate that bacteria invasions can potentially leave a footprint in the structure and functionality of soil communities, indicating the need of assessing the legacy of introducing exotic species in soil environments.     

Manganese reduction in the rhizosphere of mycorrhizal and nonmycorrhizal maize

The influence of rhizosphere microorganisms and vesicular-arbuscular (VA) mycorrhiza on manganese (Mn) uptake in maize (Zea mays L. cv. Tau) plants was studied in pot experiments under controlled environmental conditions. The plants were grown for 7 weeks in sterilized calcareous soil in pots having separate compartments for growth of roots and of VA mycorrhizal fungal hyphae. The soil was left either uninoculated (control) or prior to planting was inoculated with rhizosphere microorganisms only (MO-VA) or with rhizosphere microorganisms together with a VA mycorrhizal fungus [Glomus mosseae (Nicol and Gerd.) Gerdemann and Trappe] (MO+VA). Mycorrhiza treatment did not affect shoot dry weight, but root dry weight was slightly inhibited in the MO+VA and MO-VA treatments compared with the uninoculated control. Concentrations of Mn in shoots decreased in the order MO-VA > MO+VA > control. In the rhizosphere soil, the total microbial population was higher in mycorrhizal (MO+VA) than nonmycorrhizal (MO-VA) treatments, but the proportion of Mn-reducing microbial populations was fivefold higher in the nonmycorrhizal treatment, suggesting substantial qualitative changes in rhizosphere microbial populations upon root infection with the mycorrhizal fungi. The most important microbial group taking part in the reduction of Mn was fluorescent Pseudomonas. Mycorrhizal treatment decreased not only the number of Mn reducers but also the release of Mn-solubilizing root exudates, which were collected by percolation from maize plants cultivated in plastic tubes filled with gravel quartz sand. Compared with mycorrhizal plants, the root exudates of nonmycorrhizal plants had two fold higher capacity for reduction of Mn. Therefore, changes in both rhizosphere microbial population and root exudation are probably responsible for the lower acquisition of Mn in mycorrhizal plants.     

Molecular and phenotypic characterization of potential plant growth-promoting <Emphasis Type="Italic">Pseudomonas</Emphasis> from rice and maize rhizospheres

In this study, Pseudomonas species were isolated from the rhizospheres of two plant hosts: rice (Oryza sativa cultivar Pathum Thani 1) and maize (Zea mays cultivar DK888). The genotypic diversity of isolates was determined on basis of amplified rDNA restriction analysis (ARDRA). This analysis showed that both plant varieties selected for two distinct populations of Pseudomonas. The actual biocontrol and plant promotion abilities of these strains was confirmed by bioassays on fungal (Verticillum sp., Rhizoctonia solani and Fusarium sp.) and bacterial (Ralstonia solanacearum and Bacillus subtilis) plant pathogens, as well as indole-3-acetic acid (IAA) production and carbon source utilization. There was a significant difference between isolates from rice and maize rhizosphere in terms of biological control against R.  solanacearum and B.  subtilis. Interestingly, none of the pseudomonads isolated from maize rhizosphere showed antagonistic activity against R.  solanacearum. This study indicated that the percentage of pseudomonad isolates obtained from rice rhizosphere which showed the ability to produce fluorescent pigments was almost threefold higher than pseudomonad isolates obtained from maize rhizosphere. Furthermore, the biocontrol assay results indicated that pseudomonad isolated from rice showed a higher ability to control bacterial and fungal root pathogens than pseudomonad isolates obtained from maize. This work clearly identified a number of isolates with potential for use as plant growth-promoting and biocontrol agents on rice and maize.     

Use of Peracetic Acid as a Disinfectant in a Water-Treatment Plant: Effect on the Plasmid Contents of Escherichia coli Strains

Total thermotolerant coliforms (TTC) and Escherichia coli strains were isolated from sewage from a treatment plant before and after peracetic acid (PAA) disinfection. The plasmid profiles of 120 E. coli strains were analyzed. Although PAA disinfection effectively reduced the number of TTC and E. coli strains, the percentage of E. coli strains containing plasmids was not statistically different among water samples. The sizes of the plasmids found ranged from <3 kb to >56 kb, but plasmids of between 3 and 5 kb were encountered most frequently.     

Population variation of invasive <Emphasis Type="Italic">Spartina</Emphasis> <Emphasis Type="Italic">alterniflora</Emphasis> can differentiate bacterial diversity in its rhizosphere

While several studies have documented that invasive plants can change the microbial communities, little is known about how soil microbial communities respond to population variation of invasive plants. Here, nine populations of Spartina alterniflora were selected from the east coast of China along latitudinal gradient to compare bacterial diversity of rhizospheres among these populations. The bacterial diversity in S. alterniflora rhizospheres was valued by denaturing gradient gel electrophoresis (DGGE) analysis. Shannon–Weaver diversity index (H′) and number of DGGE bands showed that rhizosphere bacterial diversity of S. alterniflora populations increased along a latitudinal gradient when all the populations were grown in a common garden. These findings suggest that population variation of S. alterniflora can differentiate the rhizosphere bacterial diversity, and the latitudinal gradient can shape the specific plant–bacterial diversity relationship. Our results adding to the recent literature suggest that invasive plant–soil biota interactions would have clinal variation with environmental gradients and improve our understanding of the mechanisms and processes of plant invasions.     

Effects of rhizosphere remediation and bioaugmentation on carbofuran removal from soil

Rhizosphere soil contains important sources of nutrients for microorganisms resulting in high number of microorganisms capable of degrading various types of chemicals in the soil. Thus, this study investigated a carbofuran dissipation in rhizosphere soils of 6 weeds namely, umbrella sedge (Cyperus iria L.), fuzzy flatsedge (C. pilosus V.), small flower umbrella plant (C. difformis L.), tall-fringe-rush hoorah grass (Fimbristylis miliacea V.), cover fern (Marsilea crenata P.), and water primrose (Jussiaea linifolia V.). Rhizosphere soil of fuzzy flatsedge showed the shortest half-life (t_1/2) of carbofuran (15 days) among other soils. So, it was selected to be used in the bioaugmentation experiment using carbofuran degrader namely Burkholderia cepacia, PCL3, as inoculum in order to examine whether they would improve carbofuran degradation in soil. The results showed that the addition of PCL3 into rhizosphere soil did not improve carbofuran degradation suggesting that microorganisms in rhizosphere soil might be capable enough to remove carbofuran from soil. The number of carbofuran degraders in the rhizosphere soils was greater than in bulk soil 10–100 times which might be responsible to a rapid degradation of carbofuran in rhizosphere soils without the addition of PCL3. The ability of PCL3 to degrade carbofuran was evident in bulk soil (t_1/2 of 12 days) and autoclaved soils (t_1/2 13–14 days) when compared to soils without an inoculation (t_1/2 of 58 days) indicated that the addition of a degrader was useful in improving carbofuran degradation in soil.     

3-Ketoglycoside-mediated metabolism of sucrose in E. coli as conferred by genes from Agrobacterium tumefaciens

Escherichia coli strains that did not have the ability to use sucrose as a sole carbon source gained this ability after receiving a cloned fragment of DNA from Agrobacterium tumefaciens. No invertase was detected in the sucrose-metabolizing E. coli, but evidence for the activity of certain enzymes, known to be produced by biotype 1 strains of Agrobacterium, were found. Evidence was found for the presence of d-glucoside 3-dehydrogenase (G3DH) and α-3-ketoglucosidase. The activity of enzyme extracts on 3-ketosucrose also indicated that 3-ketoglucose reductase, or some enzyme that acts on 3-ketoglucose, was present in the Suc⁺ E. coli as well. The fragment was found to complement a G3DH mutant of A. tumefaciens and was also found to confer chemotaxis towards sucrose in E. coli. Received: 13 September 1996 / Received revision: 15 January 1997 / Accepted: 24 January 1997     

Maize rhizosphere modulates the microbiome diversity and community structure to enhance plant health

Metagenomic has been explored in investigating microbiome diversity. However, there is limited available information on its application towards securing plant health. Hence, this study adopts the metagenomic approach to unravel the microbiome diversity associated with healthy (LI and MA) and Northern corn leaf blight (NCLB) infected (LID and MAD) maize rhizosphere in the maize growing field at Lichtenburg and Mafikeng, North-West province of South Africa. The extraction of whole DNA from the respective healthy and diseased rhizosphere soils was conducted and sequenced using shotgun metagenomics. A total of 12 bacteria, 4 archaea and 2 fungal phyla were found as predominant across the fields with the use of the SEED subsystem database. The most predominant bacteria phyla included Proteobacteria, Dienococcus-Thermus, Gemmatimonadetes, Chlorobi, Cyanobacteria, Planctomycetes, Verrucomicrobia, Acidobacteria, Firmicutes, Chloroflexi and Bacteroidetes. Archaea consisted of Euryarchaeota, Thaumarchaeota, Crenarchaeota and Korachaeota, while Ascomycota and Basidiomycota were the dominant fungal phyla. Microbial abundance and diversity were higher in the rhizosphere of healthy maize (LI and MA) rhizosphere as compared to the NCLB diseased (LID and MAD), in the order LI > MA > LID > MAD. At phylum and genus level, alpha diversity index showed no significant (p > 0.05) difference in the abundance of the microbial community of healthy and NCLB infected maize rhizosphere, while beta analysis produced a significant (p = 0.01) difference in the microbial diversity in the soil. Taken together, the study revealed that the abundance of microbial diversity in the maize rhizosphere influences the efficacy of the rhizosphere microbiome to modulate microbial functions towards managing and sustaining plant health.     

Persistence of enterohaemorrhagic and nonpathogenic E. coli on spinach leaves and in rhizosphere soil*

Aims: Survival of Escherichia coli O157:H7 and nonpathogenic E. coli on spinach leaves and in organic soil while growing spinach in a growth chamber was investigated. Methods and Results: Spinach plants were maintained in the growth chamber at 20°C (14 h) and 18°C (10 h) settings at 60% relative humidity. Five separate inocula, each containing one strain of E. coli O157:H7 and one nonpathogenic E. coli isolate were applied to individual 4‐week‐old spinach plants (cultivar ‘Whale’) grown in sandy soil. Leaf and soil inocula consisted of 100 μl, in 5 μl droplets, on the upper side of leaves resulting in 6·5 log CFU plant^?1 and 1 ml in soil, resulting in 6·5 log CFU 200 g^?1 soil per plant. Four replicates of each plant shoot and soil sample per inoculum were analysed on day 1 and every 7 days for 28 days for E. coli O157:H7 and nonpathogenic E. coli (by MPN) and for heterotrophic plate counts (HPC). Escherichia coli O157:H7 was not detected on plant shoots after 7 days but did survive in soil for up to 28 days. Nonpathogenic E. coli survived up to 14 days on shoots and was detected at low concentrations for up to 28 days. In contrast, there were no significant differences in HPC from days 0 to 28 on plants, except one treatment on day 7. Conclusions: Escherichia coli O157:H7 persisted in soil for at least 28 days. Escherichia coli O157:H7 on spinach leaves survived for less than 14 days when co‐inoculated with nonpathogenic E. coli. There was no correlation between HPC and E. coli O157:H7 or nonpathogenic E. coli. Significance and Impact of the Study: The persistence of nonpathogenic E. coli isolates makes them possible candidates as surrogates for E. coli O157:H7 on spinach leaves in field trials.     

Clonal Diversity of Escherichia Coli Isolates from Marketed Beef in East Malaysia

Summary Escherichia coli, including Shiga-like toxin producing E. coli (STEC), serogroup O157:H7 and E. coli O157, were isolated from raw beef marketed in Sarawak and Sabah, East Malaysia. Molecular subtyping by pulsed-field gel electrophoresis (PFGE) was performed on 51 confirmed E. coli isolates. Of the 51 isolates, five were E. coli O157:H7, four E. coli O157, two non-O157 STEC and 40 other E. coli isolates (non-STEC). Digestion of chromosomal DNA from these E. coli isolates with restriction endonuclease XbaI (5′-TCTAGA-3′), followed by PFGE, produced 45 restriction endonuclease digestion profiles (REDPs) of 10–18 bands. E. coli O157:H7 isolates from one beef sample were found to have identical PFGE profiles. In contrast, E. coli serogroup O157 from different beef samples displayed considerable differences in their PFGE profiles. These suggested that E. coli isolates of both serogroups were not closely related. A large variety of PFGE patterns among non-STEC isolates were observed, demonstrating a high clonal diversity of E. coli in the beef marketed in East Malaysia. The distance matrix values (D), calculated showed that none of the pathogenic E. coli strains displayed close genetic relationship with the non-STEC strains. Based on the PFGE profiles, a dendrogram was generated and the isolates were grouped into five PFGE clusters (A–E). From the dendrogram, the most related isolates were E. coli O157:H7, grouped within cluster B. The STEC O157:H7 beef isolates were more closely related to the clinical E. coli O157:H7 isolate than the E. coli O157:H7 reference culture, EDL933. Cluster A, comprising many of other E. coli isolates was shown to be the most heterogeneous. PFGE was shown to possess high discriminatory power in typing pathogenic and non-pathogenic E. coli strains, and useful in studying possible clonal relationship among strains.