Escherichia coli Contamination of Vegetables Grown in Soils Fertilized with Noncomposted Bovine Manure: Garden-Scale Studies

In this study we tested the validity of the National Organic Program (NOP) requirement for a ≥120-day interval between application of noncomposted manure and harvesting of vegetables grown in manure-fertilized soil. Noncomposted bovine manure was applied to 9.3-m² plots at three Wisconsin sites (loamy sand, silt loam, and silty clay loam) prior to spring and summer planting of carrots, radishes, and lettuce. Soil and washed (30 s under running tap water) vegetables were analyzed for indigenous Escherichia coli. Within 90 days, the level of E. coli in manure-fertilized soil generally decreased by about 3 log CFU/g from initial levels of 4.2 to 4.4 log CFU/g. Low levels of E. coli generally persisted in manure-fertilized soil for more than 100 days and were detected in enriched soil from all three sites 132 to 168 days after manure application. For carrots and lettuce, at least one enrichment-negative sample was obtained ≤100 days after manure application for 63 and 88% of the treatments, respectively. The current ≥120-day limit provided an even greater likelihood of not detecting E. coli on carrots (≥1 enrichment-negative result for 100% of the treatments). The rapid maturation of radishes prevented conclusive evaluation of a 100- or 120-day application-to-harvest interval. The absolute absence of E. coli from vegetables harvested from manure-fertilized Wisconsin soils may not be ensured solely by adherence to the NOP ≥120-day limit. Unless pathogens are far better at colonizing vegetables than indigenous E. coli strains are, it appears that the risk of contamination for vegetables grown in Wisconsin soils would be elevated only slightly by reducing the NOP requirement to ≥100 days.     

Salmonella enterica serovar Typhimurium and Escherichia coli contamination of root and leaf vegetables grown in soils with incorporated bovine manure

Bovine manure, with or without added Salmonella enterica serovar Typhimurium (three strains), was incorporated into silty clay loam (SCL) and loamy sand (LS) soil beds (53- by 114-cm surface area, 17.5 cm deep) and maintained in two controlled-environment chambers. The S. enterica serovar Typhimurium inoculum was 4 to 5 log CFU/g in manure-fertilized soil. The conditions in the two environmental chambers, each containing inoculated and uninoculated beds of manure-fertilized soil, simulated daily average Madison, Wis., weather conditions (hourly temperatures, rainfall, daylight, and humidity) for a 1 March or a 1 June manure application and subsequent vegetable growing seasons ending 9 August or 28 September, respectively. Core soil samples were taken biweekly from both inoculated and uninoculated soil beds in each chamber. Radishes, arugula, and carrots were planted in soil beds, thinned, and harvested. Soils, thinned vegetables, and harvested vegetables were analyzed for S. enterica serovar Typhimurium and Escherichia coli (indigenous in manure). After the 1 March manure application, S. enterica serovar Typhimurium was detected at low levels in both soils on 31 May, but not on vegetables planted 1 May and harvested 12 July from either soil. After the 1 June manure application, S. enterica serovar Typhimurium was detected in SCL soil on 7 September and on radishes and arugula planted in SCL soil on 15 August and harvested on 27 September. In LS soil, S. enterica serovar Typhimurium died at a similar rate (P >or= 0.05) after the 1 June manure application and was less often detected on arugula and radishes harvested from this soil compared to the SCL soil. Pathogen levels on vegetables were decreased by washing. Manure application in cool (daily average maximum temperature of <10 degrees C) spring conditions is recommended to ensure that harvested vegetables are not contaminated with S. enterica serovar Typhimurium. Manure application under warmer (daily average maximum temperature >20 degrees C) summer conditions is not recommended when vegetable planting is done between the time of manure application and late summer. A late fall manure application will not increase the risk of contaminating vegetables planted the next spring, since further experiments showed that repeated freeze-thaw cycles were detrimental to the survival of S. enterica serovar Typhimurium and E. coli in manure-fertilized soil. The number of indigenous E. coli in soil was never significantly lower (P < 0.05) than that of S. enterica serovar Typhimurium, suggesting its usefulness as an indicator organism for evaluating the risk of vegetable contamination with manure-borne S. enterica serovar Typhimurium.     

Salmonella enterica Serovar Typhimurium and Escherichia coli Contamination of Root and Leaf Vegetables Grown in Soils with Incorporated Bovine Manure

Bovine manure, with or without added Salmonella enterica serovar Typhimurium (three strains), was incorporated into silty clay loam (SCL) and loamy sand (LS) soil beds (53- by 114-cm surface area, 17.5 cm deep) and maintained in two controlled-environment chambers. The S. enterica serovar Typhimurium inoculum was 4 to 5 log CFU/g in manure-fertilized soil. The conditions in the two environmental chambers, each containing inoculated and uninoculated beds of manure-fertilized soil, simulated daily average Madison, Wis., weather conditions (hourly temperatures, rainfall, daylight, and humidity) for a 1 March or a 1 June manure application and subsequent vegetable growing seasons ending 9 August or 28 September, respectively. Core soil samples were taken biweekly from both inoculated and uninoculated soil beds in each chamber. Radishes, arugula, and carrots were planted in soil beds, thinned, and harvested. Soils, thinned vegetables, and harvested vegetables were analyzed for S. enterica serovar Typhimurium and Escherichia coli (indigenous in manure). After the 1 March manure application, S. enterica serovar Typhimurium was detected at low levels in both soils on 31 May, but not on vegetables planted 1 May and harvested 12 July from either soil. After the 1 June manure application, S. enterica serovar Typhimurium was detected in SCL soil on 7 September and on radishes and arugula planted in SCL soil on 15 August and harvested on 27 September. In LS soil, S. enterica serovar Typhimurium died at a similar rate (P ≥ 0.05) after the 1 June manure application and was less often detected on arugula and radishes harvested from this soil compared to the SCL soil. Pathogen levels on vegetables were decreased by washing. Manure application in cool (daily average maximum temperature of <10°C) spring conditions is recommended to ensure that harvested vegetables are not contaminated with S. enterica serovar Typhimurium. Manure application under warmer (daily average maximum temperature >20°C) summer conditions is not recommended when vegetable planting is done between the time of manure application and late summer. A late fall manure application will not increase the risk of contaminating vegetables planted the next spring, since further experiments showed that repeated freeze-thaw cycles were detrimental to the survival of S. enterica serovar Typhimurium and E. coli in manure-fertilized soil. The number of indigenous E. coli in soil was never significantly lower (P < 0.05) than that of S. enterica serovar Typhimurium, suggesting its usefulness as an indicator organism for evaluating the risk of vegetable contamination with manure-borne S. enterica serovar Typhimurium.     

Fate of Salmonella enterica serovar Typhimurium on carrots and radishes grown in fields treated with contaminated manure composts or irrigation water

Three different types of compost, PM-5 (poultry manure compost), 338 (dairy cattle manure compost), and NVIRO-4 (alkaline-pH-stabilized dairy cattle manure compost), and irrigation water were inoculated with an avirulent strain of Salmonella enterica serovar Typhimurium at 10(7) CFU g(-1) and 10(5) CFU ml(-1), respectively, to determine the persistence of salmonellae in soils containing these composts, in irrigation water, and also on carrots and radishes grown in these contaminated soils. A split-plot block design plan was used for each crop, with five treatments (one without compost, three with each of the three composts, and one without compost but with contaminated water applied) and five replicates for a total of 25 plots for each crop, with each plot measuring 1.8 x 4.6 m. Salmonellae persisted for an extended period of time, with the bacteria surviving in soil samples for 203 to 231 days, and were detected after seeds were sown for 84 and 203 days on radishes and carrots, respectively. Salmonella survival was greatest in soil amended with poultry compost and least in soil containing alkaline-pH-stabilized dairy cattle manure compost. Survival profiles of Salmonella on vegetables and soil samples contaminated by irrigation water were similar to those observed when contamination occurred through compost. Hence, both contaminated manure compost and irrigation water can play an important role in contaminating soil and root vegetables with salmonellae for several months.     

Fate of Salmonella enterica Serovar Typhimurium on Carrots and Radishes Grown in Fields Treated with Contaminated Manure Composts or Irrigation Water

Three different types of compost, PM-5 (poultry manure compost), 338 (dairy cattle manure compost), and NVIRO-4 (alkaline-pH-stabilized dairy cattle manure compost), and irrigation water were inoculated with an avirulent strain of Salmonella enterica serovar Typhimurium at 10⁷ CFU g⁻¹ and 10⁵ CFU ml⁻¹, respectively, to determine the persistence of salmonellae in soils containing these composts, in irrigation water, and also on carrots and radishes grown in these contaminated soils. A split-plot block design plan was used for each crop, with five treatments (one without compost, three with each of the three composts, and one without compost but with contaminated water applied) and five replicates for a total of 25 plots for each crop, with each plot measuring 1.8 × 4.6 m. Salmonellae persisted for an extended period of time, with the bacteria surviving in soil samples for 203 to 231 days, and were detected after seeds were sown for 84 and 203 days on radishes and carrots, respectively. Salmonella survival was greatest in soil amended with poultry compost and least in soil containing alkaline-pH-stabilized dairy cattle manure compost. Survival profiles of Salmonella on vegetables and soil samples contaminated by irrigation water were similar to those observed when contamination occurred through compost. Hence, both contaminated manure compost and irrigation water can play an important role in contaminating soil and root vegetables with salmonellae for several months.     

Survival of faecal indicator bacteria in bovine manure incorporated into soil

AIMS: Survival of Escherichia coli and enterococci was evaluated in bovine manure incorporated into two Wisconsin soils. METHODS AND RESULTS: Silty clay loam (SCL) and loamy sand (LS) were mixed with fresh bovine manure, exposed daily to 10 h at 22 degrees C/14 h at 9 degrees C, and watered weekly for 12 weeks. Escherichia coli numbers increased 1-2 log cfu g(-1), then decreased < 1 and about 2 log cfu g(-1) in SCL and LS, respectively. Enterococci numbers rose less and then declined faster than those of E. coli. Watering intervals of 3, 7 and 14 days were evaluated in weeks 13-19, but did not affect the slow decline in numbers of E. coli or enterococci. CONCLUSION: Escherichia coli and enterococci may survive at least 19 weeks at 9-21 degrees C in bovine manure/soil, with E. coli surviving better. SIGNIFICANCE AND IMPACT OF THE STUDY: Quantification of E. coli or enterococci in late spring/early summer soil may be useful in indicating recent application of bovine manure.     

Impact of Manure Fertilization on the Abundance of Antibiotic-Resistant Bacteria and Frequency of Detection of Antibiotic Resistance Genes in Soil and on Vegetables at Harvest

Consumption of vegetables represents a route of direct human exposure to bacteria found in soil. The present study evaluated the complement of bacteria resistant to various antibiotics on vegetables often eaten raw (tomato, cucumber, pepper, carrot, radish, lettuce) and how this might vary with growth in soil fertilized inorganically or with dairy or swine manure. Vegetables were sown into field plots immediately following fertilization and harvested when of marketable quality. Vegetable and soil samples were evaluated for viable antibiotic-resistant bacteria by plate count on Chromocult medium supplemented with antibiotics at clinical breakpoint concentrations. DNA was extracted from soil and vegetables and evaluated by PCR for the presence of 46 gene targets associated with plasmid incompatibility groups, integrons, or antibiotic resistance genes. Soil receiving manure was enriched in antibiotic-resistant bacteria and various antibiotic resistance determinants. There was no coherent corresponding increase in the abundance of antibiotic-resistant bacteria enumerated from any vegetable grown in manure-fertilized soil. Numerous antibiotic resistance determinants were detected in DNA extracted from vegetables grown in unmanured soil. A smaller number of determinants were additionally detected on vegetables grown only in manured and not in unmanured soil. Overall, consumption of raw vegetables represents a route of human exposure to antibiotic-resistant bacteria and resistance determinants naturally present in soil. However, the detection of some determinants on vegetables grown only in freshly manured soil reinforces the advisability of pretreating manure through composting or other stabilization processes or mandating offset times between manuring and harvesting vegetables for human consumption.     

Listeria spp. found on fresh market produce.

From October 1987 to August 1988, 1,000 tests were conducted on 10 types of fresh produce from two Minneapolis area supermarkets to detect Listeria spp. The produce included broccoli, cabbage, carrots, cauliflower, cucumbers, lettuce, mushrooms, potatoes, radishes, and tomatoes. The vegetables were tested by the Food and Drug Administration method for isolation of Listeria spp., with the addition of LiCl-phenylethanol-moxalactam agar in the last 280 tests; 8.6 and 11.4% of these tests were positive by modified McBride and LiCl-phenylethanol-moxalactam agars, respectively. Listeria monocytogenes was isolated from cabbage, cucumbers, potatoes, and radishes; L. innocua was isolated from cucumbers, lettuce, mushrooms, potatoes, and radishes; L. seeligeri was isolated from cabbage and radishes; and L. welshimeri was isolated from cucumbers, potatoes, and radishes. The isolates were of various serotypes; however, the L. monocytogenes isolates were predominantly serotype 1 (82%). Only potatoes (25.8% positive) and radishes (30.3% positive) showed significant amounts of L. monocytogenes contamination.     

Listeria spp. found on fresh market produce

From October 1987 to August 1988, 1,000 tests were conducted on 10 types of fresh produce from two Minneapolis area supermarkets to detect Listeria spp. The produce included broccoli, cabbage, carrots, cauliflower, cucumbers, lettuce, mushrooms, potatoes, radishes, and tomatoes. The vegetables were tested by the Food and Drug Administration method for isolation of Listeria spp., with the addition of LiCl-phenylethanol-moxalactam agar in the last 280 tests; 8.6 and 11.4% of these tests were positive by modified McBride and LiCl-phenylethanol-moxalactam agars, respectively. Listeria monocytogenes was isolated from cabbage, cucumbers, potatoes, and radishes; L. innocua was isolated from cucumbers, lettuce, mushrooms, potatoes, and radishes; L. seeligeri was isolated from cabbage and radishes; and L. welshimeri was isolated from cucumbers, potatoes, and radishes. The isolates were of various serotypes; however, the L. monocytogenes isolates were predominantly serotype 1 (82%). Only potatoes (25.8% positive) and radishes (30.3% positive) showed significant amounts of L. monocytogenes contamination.     

Fate of Escherichia coli O157:H7 in manure-amended soil

Escherichia coli O157:H7 cells survived for up to 77, >226, and 231 days in manure-amended autoclaved soil held at 5, 15, and 21 degrees C, respectively. Pathogen populations declined more rapidly in manure-amended unautoclaved soil under the same conditions, likely due to antagonistic interactions with indigenous soil microorganisms. E. coli O157:H7 cells were inactivated more rapidly in both autoclaved and unautoclaved soils amended with manure at a ratio of 1 part manure to 10 parts soil at 15 and 21 degrees C than in soil samples containing dilute amounts of manure. The manure-to-soil ratio, soil temperature, and indigenous microorganisms of the soil appear to be contributory factors to the pathogen's survival in manure-amended soil.     

Survival and growth of Escherichia coli O157:H7 on salad vegetables.

The influence of modified-atmosphere packaging, storage temperature, and time on survival and growth of Escherichia coli O157:H7 inoculated onto shredded lettuce, sliced cucumber, and shredded carrot was determined. Growth of psychotrophic and mesophilic microorganisms and changes in pH and sensory qualities of vegetables, as judged by subjective evaluation, were also monitored. Packaging under an atmosphere containing 3% oxygen and 97% nitrogen had no apparent effect on populations of E. coli O157:H7, psychotrophs, or mesophiles. Populations of viable E. coli O157:H7 declined on vegetables stored at 5 degrees C and increased on vegetables stored at 12 and 21 degrees C for up to 14 days. The most rapid increases in populations of E. coli O157:H7 occurred on lettuce and cucumbers stored at 21 degrees C. These results suggest that an unknown factor(s) associated with carrots may inhibit the growth of E. coli O157:H7. The reduction in pH of vegetables was correlated with initial increases in populations of E. coli O157:H7 and naturally occurring microfloras. Eventual decreases in E. coli O157:H7 in some samples, e.g., those stored at 21 degrees C, are attributed to the toxic effect of accumulated acids. Changes in visual appearance of vegetables were not influenced substantially by growth of E. coli O157:H7. The ability of E. coli O157:H7 to growth on raw salad vegetables subjected to processing and storage conditions simulating those routinely used in commercial practice has been demonstrated.     

Leaching of Escherichia coli O157:H7 in diverse soils under various agricultural management practices

Application of animal manures to soil as crop fertilizers is an important means for recycling the nitrogen and phosphorus which the manures contain. Animal manures also contain bacteria, including many types of pathogens. Manure pathogen levels depend on the source animal, the animal's state of health, and how the manure was stored or treated before use. Rainfall may result in pathogen spread into soil by runoff from stored or unincorporated manure or by leaching through the soil profile. Steady rainfall consisting of 16.5 mm h(-1) was applied to 100-mm disturbed soil cores that were treated with manure and inoculated with Escherichia coli O157:H7 strain B6914. The level of B6914 in leachate was near the inoculum level each hour for 8 h, as was the level of B6914 at several soil depths after 24 h, indicating that there was a high rate of growth. Bacterial movement through three different types of soil was then compared by using disturbed (tilled) and intact (no-till) soil cores and less intense rainfall consisting of 25.4 mm on 4 consecutive days and then four more times over a 17-day period. Total B6914 levels exceeded the inoculum levels for all treatments except intact clay loam cores. B6914 levels in daily leachate samples decreased sharply with time, although the levels were more constant when intact sandy loam cores were used. The presence of manure often increased total B6914 leachate and soil levels in intact cores but had the opposite effect on disturbed soil cores. Ammonia and nitrate levels correlated with B6914 and total coliform levels in leachate. We concluded that tillage practice, soil type, and method of pathogen delivery affect but do not prevent vertical E. coli O157:H7 and coliform transport in soil and that soluble nitrogen may enhance transport.     

Potential Uptake of Escherichia coli O157:H7 from Organic Manure into Crisphead Lettuce

To investigate the potential transfer of Escherichia coli O157:H7 from contaminated manure to fresh produce, lettuce seedlings were transplanted into soil fertilized with bovine manure which had been inoculated with approximately 10⁴ CFU g⁻¹ E. coli O157:H7. The lettuce was grown for approximately 50 days in beds in climate-controlled rooms in a greenhouse. As the bacterium was not detected in the edible parts of the lettuce, the outer leaves of the lettuce, or the lettuce roots at harvest it was concluded that transmission of E. coli O157:H7 from contaminated soil to lettuce did not occur. The pathogen persisted in the soil for at least 8 weeks after fertilizing but was not detected after 12 weeks. Indigenous E. coli was detected only sporadically on the lettuce at harvest, and enterococci were not detected at all. The numbers of enterococci declined more rapidly than those of E. coli in the soil. Pseudomonas fluorescens, which inhibited growth of E. coli O157:H7 in vitro, was isolated from the rhizosphere.     

Persistence of Escherichia coli O157:H7 in soil and on plant roots

Soil microcosms were inoculated with Escherichia coli O157:H7 to test persistence in fallow soil, on roots of cover crops and in presence of manure. In fallow soils, E. coli O157:H7 persisted for 25-41 days, on rye roots for 47-96 days and on alfalfa roots, in a silt loam soil, for 92 days whereas on other legumes persistence ranged from 25-40 days, similar to fallow soil. Manure did not seem to affect the persistence of E. coli O157:H7 in these soils. Indigenous and manure-applied coliform populations often decreased faster when E. coli O157:H7 was applied, indicating possible competition between microflora. Coliform populations in microcosms not inoculated with E. coli O157:H7 decreased more slowly or increased. Microbial community analyses showed little effect for E. coli O157:H7 inoculation or addition of manure. Microbial community metabolic activity was enhanced from rye roots after 14 days and by 63 days from alfalfa roots. Microbial community lactose utilization increased over time on rye roots in all soils and on alfalfa roots in a silt loam soil when E. coli O157:H7 was inoculated. Lactose utilization also increased for uninoculated rye roots, soil around rye roots and in some fallow soils. Our data suggest that clay increases persistence and activity of E. coli O157:H7 and other coliforms. In frozen soil stored for over 500 days, E. coli O157:H7 was viable in 37% of tested samples. In summary, E. coli O157:H7 persisted longer and activity was enhanced with some cover crops in these soils due to plant roots, the presence of clay and freezing.     

Effects of cattle feeding regimen and soil management type on the fate of Escherichia coli O157:H7 and salmonella enterica serovar typhimurium in manure, manure-amended soil, and lettuce

Survival of the green fluorescent protein-transformed human pathogens Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium was studied in a laboratory-simulated lettuce production chain. Dairy cows were fed three different roughage types: high-digestible grass silage plus maize silage (6:4), low-digestible grass silage, and straw. Each was adjusted with supplemental concentrates to high and low crude protein levels. The pathogens were added to manure, which was subsequently mixed (after 56 and 28 days for E. coli O157:H7 and Salmonella serovar Typhimurium, respectively) with two pairs of organically and conventionally managed loamy and sandy soil. After another 14 days, iceberg lettuce seedlings were planted and then checked for pathogens after 21 days of growth. Survival data were fitted to a logistic decline function (exponential for E. coli O157:H7 in soil). Roughage type significantly influenced the rate of decline of E. coli O157:H7 in manure, with the fastest decline in manure from the pure straw diet and the slowest in manure from the diet of grass silage plus maize silage. Roughage type showed no effect on the rate of decline of Salmonella serovar Typhimurium, although decline was significantly faster in the manure derived from straw than in the manure from the diet of grass silage plus maize silage. The pH and fiber content of the manure were significant explanatory factors and were positively correlated with the rate of decline. With E. coli O157:H7 there was a trend of faster decline in organic than in conventional soils. No pathogens were detected in the edible lettuce parts. The results indicate that cattle diet and soil management are important factors with respect to the survival of human pathogens in the environment.     

Influence of soil type, moisture content and biosolids application on the fate of Escherichia coli in agricultural soil under controlled laboratory conditions

AIMS: To determine the fate of the enteric indicator organism, Escherichia coli, in sewage sludge (biosolids)-amended agricultural soil in relation to soil type and moisture status under controlled conditions. METHODS AND RESULTS: We enumerated Escherichia coli in soil by membrane filtration and most probable number techniques. The background concentration of E. coli was higher in sandy loam than in silty clay soil. E. coli numbers increased in soil following addition of dewatered, mesophilic anaerobically digested sludge. Escherichia coli declined to a small extent with time in both moist and air-dried unamended control soils, although decay was only highly significant (P < 0.001) in moist sandy loam (T(90) = 100 days). Removal rates were high in sludge-treated moist soil (T(90) = 20 days), but were significantly reduced in amended air-dried soil. CONCLUSIONS: Slow removal of E. coli in air-dried soil as against their rapid decay in moist soil after sludge application indicated that the soil biota are involved in pathogen reduction processes in sludge-amended soil. SIGNIFICANCE AND IMPACT OF THE STUDY: Soil ecological mechanisms are implicated as having a critical role in the fate of enteric organisms introduced into temperate agricultural soil in sewage sludge.     

Manure Enhances Plasmid Mobilization and Survival of Pseudomonas putida Introduced into Field Soil

We report a field study on plasmid mobilization in an agricultural soil. The influence of pig manure on the mobilization of the IncQ plasmid pIE723 by indigenous plasmids or by the IncP(alpha) plasmid pGP527 into the recipient Pseudomonas putida UWC1 (Rif(supr) Nal(supr)) was studied in field soil. Six plots were prepared in duplicate, three of which were treated with manure prior to inoculation of the donor and recipient strains. As a donor strain, either Escherichia coli J53(pIE723) or E. coli 600(pIE723, pGP527) was used. Putative transconjugants obtained on a selective medium were confirmed by DNA hybridization and PCR. Plasmid mobilization by indigenous mobilizing plasmids was observed on two occasions in manured soil. Manuring of soil significantly enhanced the frequency of pIE723 mobilization by pGP527, since mobilization frequencies into P. putida UWC1 were at least 10-fold higher in manured soil than in nonmanured soil. Enhanced numbers of P. putida UWC1 transconjugant and recipient colonies could be observed in manured soil throughout the 79-day field test. Transfer of pIE723 or pG527 into indigenous soil or rhizosphere bacteria could not be detected when indigenous bacteria isolated by selective cultivation were screened for the presence of these plasmids by DNA hybridization. Furthermore, the presence of IncN-, IncP-, or IncQ-specific sequences was confirmed in total community DNA extracted directly from the manured or nonmanured soil by PCR. IncW plasmids were detectable only in manured soil, indicating entry of these plasmids into soil via manure.     

Reduction of indicator and pathogenic microorganisms by psychrophilic anaerobic digestion in swine slurries

The objective of this study was to evaluate the efficiency of a low temperature anaerobic treatment to reduce viable populations of indicator microorganisms (total coliforms, Escherichia coli) and the presence of selected pathogens (Salmonella, Yersinia enterocolitica, Cryptosporidium and Giardia) in swine slurries from different sources. Experiments were carried out in 40 l Sequencing Batch Reactors (SBRs). Experimental results indicated that anaerobic digestion of swine manure slurry at 20 degrees C for 20 days in an intermittently fed SBR: (1) reduced indigenous populations of total coliforms by 97.94-100%; (2) reduced indigenous populations of E. coli by 99.67-100%; (3) resulted in undetectable levels of indigenous strains of Salmonella, Cryptosporidium, and Giardia. It can be considered as a promising method for reducing indigenous indicator and pathogenic microorganisms populations in liquid swine manure slurries.     

Leaching of Escherichia coli O157:H7 in Diverse Soils under Various Agricultural Management Practices

Application of animal manures to soil as crop fertilizers is an important means for recycling the nitrogen and phosphorus which the manures contain. Animal manures also contain bacteria, including many types of pathogens. Manure pathogen levels depend on the source animal, the animal's state of health, and how the manure was stored or treated before use. Rainfall may result in pathogen spread into soil by runoff from stored or unincorporated manure or by leaching through the soil profile. Steady rainfall consisting of 16.5 mm h⁻¹ was applied to 100-mm disturbed soil cores that were treated with manure and inoculated with Escherichia coli O157:H7 strain B6914. The level of B6914 in leachate was near the inoculum level each hour for 8 h, as was the level of B6914 at several soil depths after 24 h, indicating that there was a high rate of growth. Bacterial movement through three different types of soil was then compared by using disturbed (tilled) and intact (no-till) soil cores and less intense rainfall consisting of 25.4 mm on 4 consecutive days and then four more times over a 17-day period. Total B6914 levels exceeded the inoculum levels for all treatments except intact clay loam cores. B6914 levels in daily leachate samples decreased sharply with time, although the levels were more constant when intact sandy loam cores were used. The presence of manure often increased total B6914 leachate and soil levels in intact cores but had the opposite effect on disturbed soil cores. Ammonia and nitrate levels correlated with B6914 and total coliform levels in leachate. We concluded that tillage practice, soil type, and method of pathogen delivery affect but do not prevent vertical E. coli O157:H7 and coliform transport in soil and that soluble nitrogen may enhance transport.     

Potential role of Diploscapter sp. strain LKC25, a bacterivorous nematode from soil, as a vector of food-borne pathogenic bacteria to preharvest fruits and vegetables

Diploscapter, a thermotolerant, free-living soil bacterial-feeding nematode commonly found in compost, sewage, and agricultural soil in the United States, was studied to determine its potential role as a vehicle of Salmonella enterica serotype Poona, enterohemorrhagic Escherichia coli O157:H7, and Listeria monocytogenes in contaminating preharvest fruits and vegetables. The ability of Diploscapter sp. strain LKC25 to survive on agar media, in cow manure, and in composted turkey manure and to be attracted to, ingest, and disperse food-borne pathogens inoculated into soil or a mixture of soil and composted turkey manure was investigated. Diploscapter sp. strain LKC25 survived and reproduced in lawns of S. enterica serotype Poona, E. coli O157:H7, and L. monocytogenes on agar media and in cow manure and composted turkey manure. Attraction of Diploscapter sp. strain LKC25 to colonies of pathogenic bacteria on tryptic soy agar within 10, 20, 30, and 60 min and 24 h was determined. At least 85% of the worms initially placed 0.5 to 1 cm away from bacterial colonies migrated to the colonies within 1 h. Within 24 h, > or =90% of the worms were embedded in colonies. The potential of Diploscapter sp. strain LKC25 to shed pathogenic bacteria after exposure to bacteria inoculated into soil or a mixture of soil and composted turkey manure was investigated. Results indicate that Diploscapter sp. strain LKC25 can shed pathogenic bacteria after exposure to pathogens in these milieus. They also demonstrate its potential to serve as a vector of food-borne pathogenic bacteria in soil, with or without amendment with compost, to the surface of preharvest fruits and vegetables in contact with soil.