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United States Department of Agriculture

Agricultural Research Service

Research Project: Development and Analysis of a Protocol to Assess Survival of Fecal Organisms in Agricultural Soils Amended with Raw Manure

Location: Environmental Microbial and Food Safety Laboratory

2012 Annual Report


1a.Objectives (from AD-416):
Conduct controlled environmental studies to.
1)compare survival of manure-borne pathogens with surrogates and.
2)develop a baseline reference of pathogen and surrogate survival in manure-amended soils for comparison to field survival, accounting for environmental effects.

Conduct field studies to evaluate survival of surrogates in raw manure-amended soils used for growing fresh produce.

Contribute to development of a systematized protocol to determine safe practices for use of raw manure as a soil amendment in the production of fresh fruits and vegetables in a manner compliant with safety standards of the Food Drug and Cosmetics Act, and mandates put forth in the Food Safety Modernization Act of 2011.


1b.Approach (from AD-416):
For controlled environmental studies, USDA-ARS and UMES will collect raw manure from local animal production facilities, will characterize (and supplement, as necessary) the microbial load of the manure, will amend with appropriate pathogenic microorganisms, and determine survival as under different environmental conditions.

For field studies, USDA-ARS and UMES will collect raw manure from local animal production facilities, will characterize (and supplement, as necessary) the microbial load of the manure, will apply manure to production fields consistent with current industry practices, and will monitor local weather and environmental impacts and the resultant survival and persistence of microorganisms of interest.

USDA-ARS and UMES, in consultation with FDA, will determine selected manure types/categories for study, appropriate field application methods, sampling times, analytical protocols and methods.

USDA-ARS and UMES will provide the FDA Project Officer, on at least a quarterly basis, a status report on the current state of the study, with information on data generated from completed milestones, status of current efforts, any technical difficulties encountered and plans to alleviate the issues while moving forward, and brief “next-step” discussions. No more than six months after the completion of field studies from all locations, USDA/ARS and UMES shall provide FDA/CFSAN with a final report on the project.


3.Progress Report:

Land application of raw animal manure to enhance soil productivity may pose a food safety risk from pathogenic microorganisms that survive and contact fresh produce grown on the soils. The presence of pathogens in fresh produce has resulted in several outbreaks of human gastroenteritis that have been linked to the consumption of contaminated fresh produce. Field, growth chamber, and greenhouse studies were established and conducted at two geographically separate locations in Maryland, one on the Eastern Shore (ES) of the coastal plain, in Princess Anne, and the other on the Western Shore Uplands (WSU) along the Fall Line of the Piedmont Plateau, in Beltsville. The purpose of these studies was to develop a model protocol for such studies and to analyze the survival and persistence of non-pathogenic generic E. coli (Ec) and non-pathogenic strains of E. coli O157:H7 (attO157) as influenced by application of broiler litter (BL) and dairy manure (DM). Field plots grouped by manure type and randomized for low and high-inoculum application rates were established at the two locations with manure-soil treatments as follows: poultry litter (ES and WSU), dairy manure liquids (WSU), dairy manure solids plus dairy manure liquids (WSU), dairy solids on organic (WSU) and on conventional soil (ES and WSU), and organic soil only (WSU). Three rifampicin-resistant (RifR) strains of Ec and two RifR strains of attO157 grown in dairy manure extract were sprayed onto plots as a single inoculation event at ~5x10^6 CFU/m^2 (low) and 5x10^8 CFU/m^2 (high). Soil samples collected at 0, 1, 2, 3, 7, 14 days, and monthly for 4 months post-inoculation were analyzed for the viable inoculated strains of E. coli. Results from the WSU location show that on day 0, average populations of both Ec and att0157 recovered from low treatments from WSU plots were 3.4 log10 CFU/g; whereas populations were 6.1 and 6.0 log10 CFU/g for Ec and attO157, respectively, in high inoculation rate treatments. After 14 days, Ec counts were 0.9 log10 CFU/g and 1.4 log10 CFU/g greater than attO157 counts, for low and high inoculum rate treatments, respectively. All E. coli survived at higher population densities in poultry litter and dairy manure liquids compared to other manure treatments. Rapid population decreases for attO157 occurred in organic soil as well as dairy solids treatments. Data from remaining days of the study up through 4-months post-inoculation are being processed and analyzed. In addition, data collected from growth chamber study using a multi-strain mixture of generic and pathogenic E. coli O157:H7 strains and the ES and WSU soils also are being analyzed. Companion studies in fields and greenhouses at the ES location are given in the progress report for Project 1265-32420-005-22S, titled: Survival of Fecal Organisms and Indicators in Agricultural Soils Amended with Raw Manure.


Last Modified: 9/10/2014
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