Ontario Food Safety Research Program
Compendium: 2004-2005
ISBN 0-7794-7817-7
Ontario
Food Safety Research Program Compendium 2004/05 (PDF)
Table of Contents
Preface
Today's investments in food safety research will result in tomorrow's
savings in health-care and industry costs and in an increased ability
for Ontario to compete on the world stage. The Ontario Ministry of Agriculture
and Food (OMAF) is dedicated to making Ontario's Food Safety System world-class
and Ontario's food supply among the safest in the world. For these reasons
OMAF is investing in state-of-the-art food safety research.
We are pleased to announce the funding of ten new research projects under
the Food Safety Research Program (FSRP) that will advance our knowledge
about the prevalence, prevention and detection methods of food-borne hazards.
With these new projects we are investing close to $0.7 million dollars
in research projects being performed at four research institutions across
Ontario: the University of Guelph, the University of Toronto, the National
Research Council and the Public Health Agency of Canada. This provincial
investment has leveraged an additional $2.3 million in financial and in-kind
support from research partners. Furthermore, the program has contributed
to the training of highly qualified personnel by funding a total of 25
Ph.D., M.Sc. and B.Sc. students.
This was the fifth year of the competitive Food Safety Research Program,
and it builds on the achievements from previous years, in which the FSRP
has invested $4 million for 34 research projects.
The FSRP has been successful not only in attracting excellent research
and in achieving its stated objectives, but also in fostering collaboration
in food safety research, and in disseminating results of research promptly
and widely.
For additional information on OMAF food safety research we encourage
you to visit our website at: www.omafra.gov.on.ca/english/research .
For further information on any specific projects listed in this compendium
you are encouraged to contact the lead researcher directly.
Finally we would like to recognize and thank the many researchers, universities,
federal and provincial government departments and industry organizations
that partner with OMAF to fund, perform and to communicate research results.
We are excited about the new opportunities that have been identified,
and we thank the many research institutions that have come together to
define and carry on excellent, multidisciplinary food safety research.
There are few things as important as the safety of our food and the research
projects supported by FSRP continues to help enhance its safety for the
benefit of Ontario citizens.
Maurice Bitran, Ph.D.
Director, Innovation and Risk Management Branch
Ontario Ministry of Agriculture and Food
Gwen Zellen, DVM
Director, Food Safety Policy Branch
Ontario Ministry of Agriculture and Food
Section One: Food Safety Research Program 2004/05
In this Compendium, we present the outcome of the Food Safety Research
Program (FSRP) 2004/05 grant application and funding process.
For your convenience this Compendium is divided into four sections to
allow greater search capabilities and ease of information dissemination.
Section One: Food Safety Research Program 2004/05
In this section you will find the program description, update, research
priorities, details about the latest program competition and interesting
statistical information.
Section Two: Abstracts
This section contains detailed abstracts of the projects categorized
into three areas: Detection/validation methodology (DM); Risk Assessment
(RA) - identification of emerging food safety hazards and contaminants;
Risk Management (RM) of food safety risks. These abstracts include contact
information for the lead researcher, name of collaborating researchers
and project duration. In addition, for each project you will find a brief
description of anticipated benefits to the food safety system in Ontario.
Section Three: Status of Previously Funded Projects (2000/01 - 2003/04)
This section provides a snapshot of all funded projects since the beginning
of the program with reference to the status of the project.
Section Four: Quick Search (In print and PDF only)
This section allows you to cross reference key words (e.g. Salmonella)
to supported projects.
Background
Ontario is recognized throughout the world for the quality and safety
of its agri-food products. To retain this position of leadership in food
safety, the province has initiated science-based, field-to-fork food safety
system improvements. In partnership with the Ministry of Health and Long-term
Care (MOHLTC) and the Ministry of Natural Resources (MNR), the Ontario
Ministry of Agriculture and Food (OMAF) led an Ontario food safety system
review. During the review process, OMAF recognized the need to update
its standards and requirements to keep pace with changes in scientific
information, technology, consumer behavior, consumer lifestyles and industry
practices. The review was designed to improve Ontario's food safety system
by increasing the government's capacity to maintain high standards of
food safety, protect public health and increase the marketability of Ontario
food products. The overall goal is to build a:
- modernized, science-based food safety system founded on the principles
of risk analysis/risk management;
- seamless system that covers the food chain from field to fork;
- market-friendly system consistent with Ontario's trade responsibilities
and industry needs.
Program description
The Ontario Food Safety Research Program is a competitive
research fund established in 2001 and funded on an annual basis. The mandate
of the program is to fund innovative food safety research projects that
enhance the safety of Ontario's food through:
- Development and/or validation of testing methods for the detection
of pathogens and chemicals for use in laboratory and field settings;
- Identification of emerging food hazards and contaminants;
- Risk analysis, risk assessment, risk management and control in food
safety.
Ultimate results of the program are new and/or enhanced technologies
and diagnostic tools that support the agri-food industry and government
regulatory and laboratory programs; new knowledge about emerging food
hazards and contaminants; new strategies to reduce, eliminate and manage
food safety risks. These results contribute to and support the implementation
of HACCP and quality assurance programs throughout the food chain.
A Research Requirements Document is issued on an annual basis to solicit
research proposals from academia, industry, and government or partnership
networks with demonstrated capability to perform quality research in their
area of expertise. For the 2004/05 competition the researchers were eligible
to apply for up to $100,000 per project that has to be completed within
two years. The program strongly encourages the applicants to demonstrate
extensive collaboration and secure matching funding if possible. OMAFRA
Staff and external peer reviewers review submitted proposals. Under the
2004/05 competition we received 40 Letters of Intent in response to our
June, 2004 Call for Proposals.
The full abstracts of these newly funded projects, as well as those from
previous FSRP funding cycles, can be found on our Web site:
www.omafra.gov.on.ca/english/research .
With these new projects we are investing almost $0.7 million dollars
for research being performed at four research institutions across Ontario,
including the University of Guelph, the University of Toronto, the National
Research Council, and the Public Health Agency of Canada.
It is important to highlight that from a total investment of $0.7 million,
90% goes directly to support research and only 10% are the administrative
costs.
To be successful in obtaining the program funds the researchers must
satisfy the following program criteria:
- Fit to the research priorities described in this document; the relevance
to current issues in food safety; the anticipated contribution to improving
the food safety system in Ontario
- Quality and clarity of experimental design and project work plan
- Research capabilities of the researcher and establishment
- Contribution from collaborators and the impact on the quality of research
attained
- Appropriateness of budget items - project costs must be reasonable
and detailed
- Effectiveness of the technology transfer and communications plan in
facilitating the adoption or commercialization of the research results
Overall the project proposals should:
- Foster innovative ideas (i.e., new detection methods, new strategies
to reduce, eliminate, manage food safety risks)
- Nurture collaboration and synergy between food safety scientists,
government agencies, policy makers and the industry
- Complement, build on, and/or feed into, but do not duplicate the research
programs of other funding agencies interested in food safety
- Encourage multi-disciplinary, collaborative participatory research
- Allow researchers to explore speculative 'high reward' opportunities
- Bring new researchers into food safety research
- Encourage special topics not well covered by other funding agencies
Statistical Summary:
Overall FSRP Funding in 2004/05
- Letters of Intent received = 40
- Letters of Intent invited to submit full proposal = 20
- Applications offered funding = 10
- Success rate = 25%
Applications and Awards by FSRP Priority Area:
|
Priority Area
|
# Applications
|
# Grants Awarded
|
|
Development and Validation of Testing Methods (DM)
|
15
|
4
|
|
Risk Assessment (RA)
|
17
|
2
|
|
Risk Management (RM)
|
8
|
4
|
FSRP 2004/05 Funding Highlights
- Four awards focus on innovative, exploratory, and high-risk/high reward
research projects in development of a novel approach in detection methodology
by focusing on development and/or validation of biosensors, microarrays
and PCR methodology.
- Two awards investigate risk assessment - one is focused on determining
a prevalence of food-borne pathogens on retail raw meat in an Ontario
county; the other is focused on finding a model to utilize existing
knowledge and to determine the best possible alternatives to manage
food safety hazards.
- Four awards deal with innovative approaches to eliminate, reduce and
manage food safety hazards, such as use of bacteriophages to destroy
E. coli O157:H7 in cattle, use of feed supplements, xylanase
to reduce Campylobacter jejuni colonization in poultry, use of
a combination of UV and Hydrogen Peroxide to decontaminate minimally
processed fruits and vegetables, and use of water acidifiers to reduce
the prevalence of Salmonella in pigs.
The FSRP wishes to acknowledge and thank the peer reviewers who participated
in the program review process for their service and dedication to the
Program.
SECTION TWO: Abstracts
Detection Methodology (DM)
DM1: Improvement and Validation of a Novel Protein Microarray Assay
for Salmonella Serotyping
Project Leader:
Shu Chen, Ph.D.
Laboratory Services Division
University of Guelph
Collaborating Researchers(s):
Hugh Cai, Animal Health Lab, University of Guelph; Anne Muckle, Laboratory
for Foodborne Zoonoses, Public Health Agency of Canada
Project Duration: Jan. 2005 - July. 2006
Contact Information:
Laboratory Services Division
University of Guelph
95 Stone Road West
Guelph, ON N1H 8J7
Tel: 519-767-6319
Email: schen@lsd.uoguelph.ca
Abstract:
A rapid, cost-effective and easy-to-use protein (antibody) microarray
assay has been successfully developed for Salmonella serotyping
by the Kauffmann-White scheme in our previous project. The assay involves
immobilization of Salmonella antibodies onto epoxy activated glass
slides, labelling of Salmonella cells with the fluorescent dye
Eosin Y, capturing Salmonella cells by the antibodies, and detection
of the fluorescent signal using a microarray scanner. A prototype antibody
array was constructed for identification of 20 commonly identified and
clinically important Salmonella serovars. The antibody array was
able to detect multiple Salmonella O antigens, and both phase 1
and 2 flagella antigens simultaneously, and thus allowed correct one-step
identification of the serovar. The assay was evaluated for Salmonella
serotyping using 117 target and 73 non-target Salmonella strains
belonging to 58 serovars. The microarray profiles allowed correct serovar
identification of 86 target strains, and correct identification of O and
most of the phase antigens for an additional 30 target strains. The assay
also allowed exclusion of the 73 non-target strains from the 20 target
serovars. Because of its speed, accuracy, low cost, and ability to identify
both flagellar antigens simultaneously, the antibody microarray-based
assay is a promising alternative to the current slide agglutination method
for Salmonella serotyping. However, the assay needs further optimization,
validation under applied conditions and standardized data analysis method/tool
for its implementation in diagnostic laboratories.
In this project, we will improve and validate the system to make it operational
by diagnostic labs. More specific antibodies and additional antibodies
will be evaluated and included on the microarray to minimize impact of
cross-reactions, and to allow exclusion of closely-related serovars. A
customized software program will be created to facilitate data analysis,
interpretation and reporting. Sufficient antibody arrays will be produced
and the system will be validated using at least 1000 Salmonella strains.
The validation will be conducted side-by-side with the standard slide
agglutination method under applied conditions in collaboration with the
OIE Reference Laboratory for Salmonellosis, Public Health Agency of Canada
(PHAC).
Expected Impact of Project Outcomes on Food Safety in Ontario:
Salmonella consists of over 2500 serovars. Serotyping is the most
important universal typing method for characterization of Salmonella
isolates, and involves more than 250 antisera for identification of
all serovars. The current Salmonella serotyping method only allows
for detection of a single antibody-antigen reaction at a time. A minimum
of three to six antibody-antigen reactions are needed for recognition
of a particular Salmonella serovar. The number of reactions required
can be many times greater if a less common serovar is tested. The assay
consumes high volumes of reagents, and takes three days to two weeks to
provide an answer.
Successful completion of our proposed project will result in an improved
and validated protein microarray-based test that can be used for rapid
and cost-effective serotyping of Salmonella in a single test. This
will allow testing laboratories and government inspection staff to conduct
risk assessment and outbreak studies in a cost-effective and timely manner
in support of monitoring and surveillance activities and enforcement of
regulatory programs. The method developed in this project can be extended
to serotyping of other food-borne pathogens such as Campylobacter.
DM2: Rapid Ultra-Concentration Technologies for Isolation of Pathogen
DNA and RNA Markers from Foodstuffs for On-line Detection and Screening
Project Leader:
Ulrich J. Krull, Ph.D.
University of Toronto at Mississauga
Collaboration Researchers:
Paul Piunno, and Xuezhu Liu, University of Toronto at Mississauga
Project Duration: Jan. 2005 - Dec. 2006
Contact Information:
Ulrich J. Krull
Chemical and Physical Sciences
University of Toronto at Mississauga
3359 Mississauga Road North
Mississauga, ON L5L 1 C6
Tel: 905-828-5437
Email: ukrull@utm.utoronto.ca
Abstract:
Two key challenges that must be overcome prior to the implementation
of practical biosensor and biochip technologies for testing of food and
water samples for nucleic acids indicative of pathogenic contaminants
are: (1) the provision of stable and reproducible sensing chemistries
to achieve the desired device performance and allow manufacture, and,
(2) to rapidly process on-line statistically representative samples (e.g.
litres of fluid or grams of tissue) and deliver isolated target molecules
to the sensing device in a small volume aliquot. We have developed a robust
and reliable optical sensor technology for nucleic acid analysis. In a
project funded by Genome Canada, we are currently developing a high sensitivity
detection system prototype that will be capable of measuring nucleic acid
targets directly from samples (i.e. without the need for enrichment of
the nucleic acid concentration by amplification methods such as, for example,
PCR). We are therefore well poised to address the second key challenge
of rapid sample preparation that will permit near real-time automated
analysis of pathogens in foodstuffs. On-line cell capture, ultrasonic
disruption and microfluidic methods will be developed for rapid isolation
and ultra-concentration of marker nucleic acids from food washes and surface
wipes, and water, which will permit automated analysis for critical pathogens
using our new nucleic acid biosensor technologies. By developing this
missing link in the overall diagnostic method, rapid quantitative testing
for multiple targets including bacteria, viruses and parasites at each
link in the food supply chain may be realised. The technology will provide
for inexpensive (per test) reliable screening that can be used in abattoirs,
food processing plants and in-field, as identified as goals of the HACCP
program.
Expected Impact of Project Outcomes on Food Safety in Ontario:
It is estimated that over 80 million food-borne illnesses occur in North
America each year, and that these can largely be attributed to E. coli,
Salmonella, Campylobacter and Listeria monocytogenes
(P.S. Mead et al., Emerg. Infect. Dis., 5: 607, 1999). Since Walkerton
there have been a number of E. coli outbreaks attributed to the
food supply. Public health officials have been anxious for a rapid monitoring
system to assess food and water supply systems for microbial contamination.
We have developed optical sensors that could rapidly detect interfacial
nucleic acid hybridization with high selectivity. Results showed that
genotyping hybridization assays could be done in minutes and that selective
binding was observed with discrimination of single-nucleotide polymorphisms.
Measurements were complete in 1-2 minutes, and the sensors have been demonstrated
to be sufficiently robust that they may be reused for over 500 cycles
of application (months of use) with good reproducibility (C.V. < 15
%) and no indication of degradation. The ultimate goal is approximately
1000 determinations per hour, at detection levels of 1000 molecules of
nucleic acid target, i.e. single cells or a few cells when screening RNA.
Our advances in detection technology have made it even more important
to shift focus towards creation of methods to decrease the amount of time
and effort necessary for sample collection and preparation. The problem
is to take a sample of some liters in volume or grams in tissue mass,
selectively collect organisms of interest while eliminating most of the
sample volume, and then to produce a measurable target and bring this
target in a concentrated small volume to a detector array. In partnership
with Genome Canada, and an industrial partner, SafeGuard Biosystems, we
intend to develop biosensor technologies that have the potential to provide
the speed, selectivity and sensitivity required for effective analysis
of food and water.
DM3: Mycobacterium avium subsp. paratuberculosis: Novel Tools
to Identify Risk for Contamination of Foods and Environments
Project Leader: Lucy Mutharia, Ph.D.
University of Guelph
Collaborating Researchers(s):
Joseph Odumeru, Ph.D. Laboratory Services Division, University of Guelph
Project Duration: Feb. 2005 - Jan. 2007
Contact Information:
Lucy M. Mutharia
Molecular and Cell Biology
University of Guelph
Guelph, ON N1G 2W1
Tel: 519-824-4120 x 56349
Email: lmuthari@uoguelph.ca
Abstract:
The goals of this research are to evaluate a three-step procedure to
identify the presence of live Mycobacterium avium subspecies paratuberculosis
(MAP) comprised of, i) affinity-based bacterial extraction, ii) broth
enrichment and iii) mRNA-based identification, for the potential to improve
sensitivity and reduce detection time of the contamination or infection.
Current procedures utilize extensive sample processing and decontamination
to separate the relatively few MAP from large numbers of microbial contaminants.
Although the sensitivity of testing by culture on agar is accepted as
poor, and takes 6-18 weeks for results, it is widely applied as the best
method available to detect live MAP. At present there is no recommended
standard laboratory protocol for live MAP tracking in milk or other sample
types. Given this lack of standards for sample preparation, decontamination,
and culture there is considerable variability in assay sensitivity, contamination
rates and time-to-detection among laboratories when processing identical
sample types.
Using various sample types, the study shall determine assay reliability,
specificity, detection range, and cost. The study will examine the procedures
for their application in clinical and food laboratories. Specifically,
the study will focus on the following 3 components of the assay:
a) Evaluate monoclonal antibody-based immunomagnetic capture for rapid
and specific isolation of MAP cells.
b) Evaluate the potential of broth enrichment to increase the sensitivity
of the assay.
c) Optimize an RNA-based protocol to detect MAP-specific transcripts,
hence viable MAP, directly in the captured MAP or after periods of broth
enrichment.
MAP is the cause of Johne's disease (JD), and affects approximately 37%
of Ontario dairy herds. Feces, milk, blood and lymph nodes from infected
animals can be contaminated with MAP bacteria. The infectious dose of
MAP is not known. Some MAP bacteria escape killing during pasteurization
and present an exposure risk to susceptible humans. Currently, it is not
known what factors affect MAP persistence in the environment, it's survival
in milk, or in meat. The potential for contamination of milk and meat
within the Ontario dairy industry are not known.
Expected Impact of Project Outcomes on Food Safety in Ontario:
Through collaboration with the University of Guelph Laboratory Services
Division the study will develop an assay for application in diagnostic
service to test both specimens for disease and food products for contamination.
The impetus for the development of a widely available, reliable, sensitive
and rapid assay is that it will enable the following:
a) Disease management: Reliable data are needed on the biological
and on-farm environmental sources of live MAP. Informed strategies can
then be implemented to reduce transmission within and among herds. There
are no effective treatments or vaccines. Identifying and removing sub-clinically
infected animals and other on-farm sources of infection will best achieve
control of JD. Wild ruminant and non-ruminant species infected with MAP
are a concern as they can act as reservoirs for the disease within a farm
and as vectors carrying the disease between farms and so interfere with
efforts to control the disease in livestock.
b) Human Food Safety: Pasteurization studied over a wide range
of temperatures and time combinations either report effective killing
of MAP or tailing with live bacteria. It is apparent that live MAP is
present in retail milk. Blood, lymph node tissue, feces and milk of animals
with subclinical disease can contain MAP organisms. Concerns regarding
the zoonotic potential of MAP indicate that the contamination of all products
designated for human consumption be accurately assessed, then reduced
or eliminated and monitored.
DM4: Reagentless Impedimetric Biosensors for Detection of Pathogens
in Greenhouse Operations and Pork Processing
Project Leader: Keith Warriner, Ph.D.
University of Guelph
Collaborating Researchers(s):
D. Thomas and C. Hall, University of Guelph
Project Duration: Jan. 2005 - Jun. 2006
Contact Information:
Keith Warriner
Department of Food Science
University of Guelph
Guelph, ON N1G 2W1
Tel: 519-824-4120 x 56072
Email: kwarrine@uoguelph.ca
Abstract:
The time between the introduction of a biohazard and subsequent detection
can strongly influence its impact (both in terms of health and economic).
Although rapid detection methods are available these are typically laboratory
based. Therefore, even if analysis can be performed within hours the time
to collect, deliver and process the sample can take days. In this respect
there is an identified need for sensors that can be used on-site that
act as an alarm for the presence of potential biohazards. In the following
project novel reagentless sensors for the detection of fecal indicators
and pathogens will be developed.
The sensors are based on conducting polymer nano-tubes formed within
microporous polycarbonate membranes onto which bioaffinity agents (host
cell or antibodies) are immobilized. Interaction of the immobilized bioaffinity
agent with analyte induces changes in the electrochemical properties of
the conducting polymer film that can be detected using impedance spectroscopy.
In the current project the target analytes selected will be F(+) coliphage
(bacteriophage commonly linked to fecal contamination) and Salmonella.
However, the generic sensing approach can be used to detect any biohazard
(bacterial pathogens, enteric viruses, protozoa, endospores, toxins) through
judicious choice of bioaffinity agent.
Initial work will fabricate the polymer electrodes and optimize immobilization
of bioaffinity agents within the conducting polymer nano-tubes. The kinetics,
sensitivity and selectivity of the sensors will be determined. Sensor
performance will be validated against current standard techniques. Finally
the ability to detect contamination within irrigation water or pork processing
facilities using the sensor devices will be evaluated. Matrix affects
on sensor performance will be evaluated and minimized through techniques
such as background subtraction.
The sensors to be developed will be reagentless with no sample volume
limitations or significant user input. The projected cost of the sensor
strips will be $2-5 and have a detection time of 5- 40 minutes. The limit
of detection for coliphage will be in the order of 1 pfu/ml. The lower
limit for Salmonella detection is projected to be 1-102 cfu/ml.
Expected Impact of Project Outcomes on Food Safety in Ontario:
OMAF has been pro-active at introducing HACCP and traceability schemes
throughout the food chain. This in turn has led to an increase in end-product
screening to reduce the risk of contaminated products reaching the marketplace.
However, a more efficient approach would be to screen for hazards at critical
points within the food chain thereby restricting the dissemination of
contamination.
The sensors will be fabricated from relatively inexpensive materials
and it is anticipated that each unit cost no more than $2-5. By using
plant derived antibodies (developed by Prof C A Hall) the cost can be
further reduced. Hand-held impedance analyzers are now commercially available
and can be customized to the appropriate measurement type required.
Availability of robust, reliable and cheap biohazard sensors is the foundation
of effective biosecurity, HACCP and traceability schemes. In this respect
the generic sensing approach to be developed will be of direct benefit
to food producers, processors, retailers and regulators.
Risk Assessment (RA)
RA1: A Novel Evidence-Based Tool in Support of Food Safety Policy Development
Project Leader/Co-Leader:
Scott McEwen, DVM, DVSc, Diplomate ACVP
University of Guelph
Andrijana Rajic, DVM, MSc
Public Health Agency of Canada
Collaborating Researchers:
Jan Sargent, McMaster University; Aamir Fazil, Susan Read, Public Health
Agency of Canada, Aboubakar Mounchili, University of Guelph, Mike Cassidy,
Ontario Ministry of Agriculture and Food
Project Duration: Jan. 2005 - Jan. 2007
Contact Information:
Andrijana Rajic
Public Health Agency of Canada
Laboratory for Foodborne Zoonoses
160 Research Lane, Unit 103
Guelph, ON N1G 5B2
Tel: 519-826-2255
Email: andrijana_rajic@phac-aspc.gc.ca
Abstract:
Policy makers are increasingly dealing with complex food safety and public
health issues such as BSE, antimicrobial resistance, avian influenza,
food-borne and other emerging pathogens and hazards. The recently published
'Justice Haines Report' has strongly recommended the use of a science-based
approach to food safety and more transparent food safety policy making
in Ontario. In a world where information is only a click away, food safety
professionals and policy makers need to identify information in a timely
way, and to appraise and synthesize the best evidence on targeted issues.
They also need to refine this evidence, to evaluate risks and to select
optimal mitigation strategies within the Ontario context. Therefore, this
research team, comprising researchers from two Canadian universities (Guelph,
McMaster), two governmental agencies (Public Health Agency of Canada and
OMAF) and collaborators from Iowa State University, proposes the development
and evaluation of a novel evidence-based tool as a potential standard
tool in support of food safety policy.
The tool will combine two existing knowledge synthesis and knowledge
transfer methodologies. These are systematic review and risk assessment.
Although they have been extensively used individually in other professional
sectors, this is the first attempt to combine and adapt them to food safety
policy making. The first methodology will enable trained research teams
to identify, evaluate, rank and summarize, qualitatively and quantitatively,
the best existing evidence on targeted issue(s). The second methodology
will utilize this evidence and synthesize it into a characterization of
the system so as to model risks associated with different scenarios/options.
The tool will be evaluated using one relevant food safety and trade issue,
namely 'Salmonella in pork'. This issue was selected because of recent
international and domestic trends that indicate an urgent need for policy
development. However, it will be applicable to any food safety issue and
agri-food sector in Ontario.
The project will be carried out in two phases. In phase 1, the research
team will develop and evaluate the protocol for conducting systematic
reviews on food safety effectiveness research. A rigorous and transparent
systematic review will be conducted to evaluate and rank the effectiveness
of potential on-farm interventions against Salmonella in swine
and their estimated impacts on Salmonella reduction. In phase 2,
quantitative risk assessment models will be developed to refine selected
interventions, to evaluate risks throughout the pork chain and to select
optimal mitigation strategies against Salmonella within the environment
of the Ontario pork production system.
A transparent, evidence-based summary and recommendations will be communicated
to Ontario's policy makers and pork industry. The needs, gaps and opportunities
for using systematic review and risk assessment as a potential standard
tool in support of evidence-based food safety policy making will be evaluated.
Expected Impact of Project Outcomes on Food Safety in Ontario:
- A novel evidence-based tool will be developed and evaluated as a potential
standard tool in support of food safety policy making in Ontario.
- The resulting tool will support timely and informed food safety policy
making in Ontario.
- Ontario's policy makers and agri-food sectors will negotiate existing
and forthcoming food safety issues based on powerful and sound evidence.
- Baseline and other research data generated through previous OMAF-funded
projects will be utilized in support of food safety policy making.
- The resulting information will allow the Ontario pork industry to
appropriately evaluate interventions against Salmonella in pork
and to make informed policy decisions regarding potential control options.
RA2: Prevalence and Enumeration of Foodborne Microbial Hazards in Retail
Raw Meat Products in an Ontario Community
Project Leader: Frank Pollari, Ph.D.
Public Health Agency of Canada (PHAC)
Collaborating Researcher(s):
Angela Cook, Katarina Pintar, Barbara Marshall, Andre Ravel, LFZ, PHAC
Project Duration: Jan. 2005 - Dec. 2006
Contact Information:
Frank Pollari
Public Health Agency of Canada
Laboratory for Foodborne Zoonoses
160 Research Lane, Unit 206
Guelph, ON N1G 5B2
Tel: 519-826-2184
Email: frank_pollari@phac-aspc.gc.ca
Abstract:
To address food-borne illness in Ontario, the proposed study will link
with a Public Health Agency of Canada/Agriculture and Agri-Food Canada
(PHAC/AAFC) surveillance initiative that is measuring the prevalence of
enteric pathogens in animals, retail food, water and humans. The study
objectives include linking with the C-EnterNet project to facilitate the
enumeration and additional sub-typing of retail food pathogen isolates
for one year. In addition, a summary of human health enteric illness data
will be developed, in conjunction with analysis of the retail food contamination
levels.
The collaborative nature of this project, and the development of retail-level
baseline enumeration data for the most important enteric food-borne pathogens,
will contribute valuable Ontario-specific data for risk assessment activities
and the development of Ontario's Food Safety Objectives. The study directly
addresses a number of recommendations from the Haines report. Collaborations
will involve the Region of Waterloo Public Health Unit, the Ontario Ministry
of Health and Long Term Care, the Public Health Agency of Canada (Laboratory
for Food-borne Zoonoses and Centre for Infectious Disease Prevention and
Control), and Agriculture and Agri-Food Canada. Discussions are currently
being pursued with the Centers for Disease Control (CDC) in the US to
collaborate on a component of the retail raw meat study (specifically
targeting Campylobacter levels on poultry products), and could provide
some international comparison data.
The study design is a cross-sectional survey of raw retail pork, beef
and poultry contamination (at the consumer level) for a 12-month period,
to generate valid and representative data about consumer-level exposure
to food-borne pathogens in meat, in one Ontario community.
The study area will be the Regional Municipality of Waterloo, a community
of 470,000 residents with urban/rural demographics similar to the Canadian
average.
Expected Impact of Project Outcomes on Food Safety in Ontario:
Science-based risk assessment along the food continuum is the modern
approach to food safety regulation and policy development and is strongly
endorsed by all levels of government. Unfortunately, there are limitations
when risk assessment principles are applied to food safety issues, including
the complexity of the food system, the lack of baseline data and inherent
assumptions made due to information gaps.
This proposal is a collaborative initiative with a five-year PHAC/AAFC
study, which will provide Ontario-specific baseline prevalence and enumeration
data for enteric pathogen contamination of raw meat products, in order
to provide relevant data that can be used in risk assessment.
This study will have many applications to current food issues. Pathogen
enumeration baseline data will support OMAF risk managers in the prioritization
of mitigation efforts, policy development, and the assessment of Ontario
quality assurance and HACCP programs. As well, a number of Justice Haines'
recommendations would be addressed.
C-EnterNet will integrate human, agri-food and water data, through enhanced
typing procedures, to provide a comprehensive understanding of the link
between food-borne pathogens and human health outcomes in Ontario. One
of the study objectives is to foster collaborative efforts between jurisdictions
and disciplines. One potential collaborative opportunity is a Campylobacter
enumeration study on retail chicken, which is in development by the CDC
in the US. These collaborations will provide an international perspective
to the issue of retail meat contamination, and an exponential return on
the investments of all of the collaborative partners.
Risk Management (RM)
RM1: Control of Salmonella and Other Pathogens of Public Health
Concern on Ontario Pig Farms
Project Leader: Robert M. Friendship, Ph.D.
University of Guelph
Collaborating Researcher(s):
Cate Dewey, Jeff Gray, University of Guelph
Project Duration: Jan. 2005 - Jan. 2007
Contact Information:
Robert M. Friendship
Department of Population Medicine
University of Guelph
Guelph, ON N1G 2W1
Tel: 519-824-4120 x 54022
Email: rfriends@ovc.uoguelph.ca
Abstract:
We propose to conduct field studies on Ontario pig farms in order to
establish effective intervention strategies to minimise the prevalence
of pathogens of public health significance. In particular, we hope to
demonstrate how Yersinia enterocolitica and Salmonella sp
can be controlled. These two disease agents appear to be the most important
food-borne pathogens associated with pork. If these organisms can be reduced
at the farm level then the risk through the rest of the food chain will
be also lessened.
Over the past 5 years, we have monitored approximately 80 finishing herds
on an annual basis and have identified herds with a high prevalence to
either Yersinia or Salmonella. In the case of Yersinia,
a major risk factor appears to be environmental contamination and we hypothesised
that strict hygiene and reduced mixing of pigs will greatly reduce the
prevalence of Yersinia enterocolitica. On the other hand,
Salmonella appears to be more closely associated with feeding techniques.
We have shown that farms using fermented liquid-feeding are much less
likely to have Salmonella-shedding market hogs compared to farms
using dry-feeding. It has been suggested that the main reason for this
apparent protection is the acidification of the diet. Whereas, liquid
feeding is an expensive technology that tends to be only available to
large farming operations, water acidification is easily applied to any
size of pig farm, making it a more acceptable approach.
We propose to select 10 farms that we have identified as having a high
prevalence of Salmonella and another 10 farms that have a high
prevalence of Yersinia. Initially, we will investigate the spread
of disease within each farm to determine when pigs appear to become infected
and therefore determine at which stage of production intervention would
be most appropriate. We will conduct field trials to evaluate intervention
strategies on 5 farms with Salmonella and 5 farms with Yersinia.
Intervention on five Salmonella farms will consist of water medication
with an acidifier. For five Yersinia farms, intervention will be
improved hygiene and control of mixing of pigs. Ten control and ten treatment
pens will be assigned and followed from entry in the grower-finisher barn
until market on each farm. Culture of faeces of market age pigs will be
used to determine the success of the treatment.
Expected Impact of Project Outcomes on Food Safety in Ontario:
Pork production is a major agricultural industry in Ontario and food
safety has become an important issue with regard to ensuring both domestic
and export markets. Countries, such as Denmark, have instituted on-farm
monitoring programs to measure the prevalence of Salmonella. There
will be pressure to institute similar programs in Ontario. The success
of a monitoring program depends on whether or not farms that are identified
with a high pathogen load can be instructed as to how they might be able
to reduce the prevalence of Salmonella or other pathogens. At present,
it is unlikely that some of the steps used in Denmark and elsewhere could
be easily implemented or would be successful.
The Ontario pork industry is very diverse with many different farm types
and management systems. In order to be able to implement a program to
minimize pathogens of public health concern, intervention strategies need
to be relatively inexpensive and practical for small farming operations
as well as large multi-site co-operatives. The benefit of this research
is to ensure that Ontario pig farmers are able to respond to demands and
pressures in the area of food safety. Our goals are to develop the monitoring
tools and practical intervention strategies to assist Ontario pig farmers
to meet food safety standards of the future.
RM2: Bacteriophage Therapy to Control E. coli O157:H7 in Cattle
Project Leader: Roger Johnson, M.V.Sc., Ph.D.
Public Health Agency of Canada
Collaborating Researchers(s):
Tim McAllister, Agriculture and Agri-Food Canada; Jack Gemmell and Paul
Averback, Nymox Pharmaceutical; Carlton Gyles and Scott McEwen, University
of Guelph
Project Duration: Apr. 2005 - Mar. 2007
Contact Information:
Roger Johnson
Public Health Agency of Canada
Laboratory for Foodborne Zoonoses
110 Stone Road West
Guelph, ON N1G 3W4
Tel: 519-822-3300 x 241
Email: roger_johnson@phac-aspc.gc.ca
Abstract:
Pre-harvest control of microbial food-borne and environmental hazards
is an essential component of an effective food production system. Our
earlier studies showed that bacteriophage (phage) therapy is potentially
an effective intervention to control E. coli O157:H7 in cattle,
which frequently carry this important human pathogen. The purpose of this
project is to study the safety and efficacy of this therapy. We will study
the safety of the phages in cattle and animals that share the same environments
as cattle (e.g. rodents and birds) and that may be potentially exposed
to the phages, and the efficacy of the therapy in cattle, firstly in controlled
experimental infection studies in calves, and secondly in naturally infected
cattle. This project will be conducted by researchers at the Laboratory
for Foodborne Zoonoses of the Public Health Agency of Canada, Lethbridge
Research Centre of Agriculture and Agri-Food Canada, and Nymox, a Canadian
company, all of whom have considerable experience and interest in phage
therapy. The outcomes of this project will provide information required
for field trials and eventual transfer of this technology to field use.
Expected Impact of Project Outcomes on Food Safety in Ontario:
Recognising the impact of E. coli O157:H7 on marketability of
cattle products domestically and internationally, cattle producers and
processors in Ontario and elsewhere have embraced the need for on-farm
control of this organism. Effective interventions at the farm level will
not only enhance the microbial safety of cattle food products, but will
reduce risks of contamination of water supplies, fresh produce, animals
and the environment. Although several on-farm interventions have been
investigated, most have met with little or only moderate success. However,
we have shown "proof of concept" that treatment of cattle with
E. coli O157 phages can potentially eliminate E. coli O157:H7
from cattle. This project addresses several remaining objectives of a
larger project designed to further develop and transfer this technology
to field use. If successful, this technology will benefit the Food Safety
System in Ontario and the agri-food sector in general, through safer beef
products, reduced contamination of the environment, water and other foods
with E. coli O157:H7, reduced risk of human exposure to E. coli
O157:H7, and enhanced market competitiveness in beef products domestically
and internationally.
RM3: Reduction of Campylobacter jejuni Colonization in Poultry
Project Leader: Christine Szymanski, Ph.D.
National Research Council (NRC)
Pathogen Genomics
Collaborating Researchers:
B. Allan, VIDO; C. Bihun, W. Sung, J.R. Brisson, J. Kelly, J. Nash, NRC
Project Duration: Jan. 2005 - Dec. 2006
Contact Information:
Christine Szymanski
National Research Council
Pathogen Genomics
100 Sussex Drive, Room 3149
Ottawa, ON K1A 0R6
Tel: 613-990-1569
Email: christine.szymanski@nrc-cnrc.gc.ca
Abstract:
Campylobacter jejuni is the leading cause of bacterial foodborne
illness in North America. Contaminated poultry are the primary risk factor
for C. jejuni infection in humans. Thus, reducing the levels of
C. jejuni colonization in poultry is a priority in the area of
risk management and control to increase food safety.
Supplementation of poultry feed with the high-efficiency feed supplement,
xylanase, was demonstrated to cause changes in mucin carbohydrates and
reduce mucin viscosity and C. jejuni colonization. In order to
further exploit this finding, we will investigate the role of chicken
mucin viscosity and carbohydrate content on C. jejuni virulence
and colonization potential. In addition, NRC-IBS has engineered a superior
xylanase supplement (http://ibs-isb.nrc-cnrc.gc.ca/ibs/ourstories/iogenstory_e.html)
that is able to resist extreme temperatures during animal feed pelleting
and remains active in the chicken gastrointestinal tract. Together we
will compare the effects of commercial and NRC-IBS modified xylanase on
C. jejuni colonization with the intent to develop a cost-effective
high efficiency animal feed supplement that will also improve Ontario
Food Safety by reducing the levels of C. jejuni colonization in
poultry. This objective will be achieved through the efforts of our group
with internationally recognized expertise in the analyses of colonization
factors (Szymanski/Allan), development of animal models (Allan/Bihun),
carbohydrate structure elucidation (Brisson/Kelly), gene expression profiling
(Nash) and xylanase engineering (Sung). These studies will involve using
well developed in vivo model systems, state-of-the art analytical equipment
available at NRC-IBS (http://ibs-isb.nrc-cnrc.gc.ca/ibs/immunochemistry/bioanalysis_e.html)
and in-house developed C. jejuni microarrays (http://ibs-isb.nrc-cnrc.gc.ca/ibs/immunochemistry/campychips_e.html).
Expected Impact of Project Outcomes on Food Safety in Ontario:
C. jejuni is the leading cause of gastroenteritis in Canada resulting
in a significant health burden to our economy. The primary risk factor
for campylobacter infection is contaminated poultry. Recently it
has been demonstrated that supplementation of chicken feed with xylanase
causes a decrease in C. jejuni colonization with a simultaneous
change in mucin viscosity and carbohydrate content . Other studies have
demonstrated that both the physical changes in viscosity and the presence
of mucin have affects on C. jejuni infectivity (2,3), Bourke, personal
communication). We will further extend these observations by using our
expertise in carbohydrates and understanding of C. jejuni regulatory
pathways, to examine the xylanase induced changes in mucin and how this
influences campylobacter colonization of poultry. In addition,
chicken feed will be supplemented with commercial xylanase and compared
with an NRC-IBS modified xylanase to determine reduction of campylobacter
colonization. With Iogen Corporation (Ottawa), the NRC-IBS enzyme has
been engineered to resist extreme temperatures during animal feed pelleting,
remains active in the chicken gastrointestinal tract, and facilitates
efficient feed conversion through better digestion and assimilation, leading
to enhanced meat and egg production. Thus, the modified xylanases, currently
approved for use in pulp bleaching with annual sales in the millions,
have further applications in food safety and the livestock industry.
RM4: Ultra Violet and Hydrogen Peroxide Combination for Decontaminating
Minimally Processed Fruits and Vegetables
Project Leader: Keith Warriner, Ph.D.
University of Guelph
Collaborating Researchers(s):
G.S. Mittal and D. Mercer, University of Guelph
Project Duration: Jan. 2005 - Dec. 2006
Contact Information:
Keith Warriner
Department of Food Science
University of Guelph
Guelph, ON N1G 2W1
Tel: 519-824-4120 x 56072
Email: kwarrine@uoguelph.ca
Abstract:
Minimally processed vegetables and fruits are become established as a
significant vehicle for foodborne pathogens. Due to the open nature of
the fresh produce chain, contamination of the product can occur at any
point during cultivation and post-harvest handling. Because bacteria (including
human pathogens) can become located within sub-surface structures of plants
(stomata, cut- edges) simply washing produce is inadequate even when chemical
sanitizers such as hypochlorite are used.
In this study the efficacy of applying a combination of Ultra Violet
(UV)/hydrogen peroxide (H2O2) in decontaminating
produce will be evaluated. When H2O2 (<2% v/v)
is illuminated with UV light highly anti-microbial, but short-lived, hydroxyl
radicals are formed. This provides a greater kill effect than when either
UV or hydrogen peroxide is applied alone. Although the combination of
UV and hydrogen peroxide have been used for over 20 years for sterilizing
carton packaging it has not been fully evaluated as a method for decontaminating
fruit or salad vegetables.
In the proposed project a treatment chamber will be constructed with
the capacity to decontaminate 3 kg batches of produce. Optimization of
operating conditions will be performed to maximize the generation of hydroxy
radicals from H2O2 (0.2-1%). The kinetics and mode
by which UV/ H2O2 inactivates a range of pathogenic
(E. coli O157, Salmonella, L. monocytogenes, Aeromonas
hydrophilia) and spoilage (Pseudomonas fluorescens and Erwinia
carotovara) bacteria, in addition to MS2 coliphage (enteric virus
surrogate), will be undertaken. The efficacy by which the treatment can
decontaminate a diverse range of product types (lettuce, red cabbage,
spring mix, spinach, red onion, carrot, tomatoes, cantaloupes, strawberries,
raspberries and apples) will be evaluated. A 5-log reduction in bacteria/bacteriophage
numbers without adversely affecting product quality will be used as the
criteria for success.
The project brings together Elopak (specialists in UV: H2O2
decontamination technology), Pride Pak Salads (major fresh cut producer
within Ontario) food microbiologists and engineers.
Expected Impact of Project Outcomes on Food Safety in Ontario:
The fresh-cut industry represents a significant sector of the Ontario
economy with an estimated Farm Gate Value of $224 M per annum. Although
foodborne outbreaks linked to fruit and vegetables are rare within Ontario
there is a need to take a pro-active approach with respect to ensuring
high food safety standards within the industry.
Currently there is no intervention step within the fresh cut chain that
can ensure the removal of field acquired contamination. It is widely acknowledged
that aqueous wash-based systems have limited ability to effectively penetrate
pathogens present in sub-surface locations. The proposed approach of using
UV/ H2O2 is specifically aimed at inactivating pathogens
located in such protective sites. The method is based on generating an
intense burst of anti-microbial hydroxyl radicals that rapidly inactivate
microbes located on the surface and sub-surface of produce. Because of
the transient nature of the radicals it is envisaged no bleaching of the
product will occur.
The project will provide a viable, cost effective, alternative to current
produce decontamination methods. If successful, the UV/ H2O2
based decontamination system will significantly enhance the safety of
minimally processed fruit and vegetables. The method will also find utility
in other sectors especially in relation to meat processing where virulent
pathogens such as E. coli O157 are an obvious concern.
SECTION THREE: Status of previously funded projects
(2000/01 - 2003/04)
Detection Methodology (DM)
|
Lead Researcher
|
Project Title
|
Status
|
Dr. Archambault Marie,
University of Guelph |
Johne's Disease - New Test Validation |
Completed |
| Dr. Bidawid Sabah, Health Canada, Ottawa |
Development of a Rapid Microarray Diagnostic Assay
for Detection of Norwalk-like Viruses in Food |
Ongoing |
| Dr. Brown Stephen, Queens University |
Development of a Rapid, Sensitive and Reliable Test
for the Detection and Quantification of Escherichia coli in
Foods
|
Completed |
Dr. Chen Shu,
University of Guelph |
Improvement and Validation of a DNA Microchip-based
Test for Rapid and Simultaneous Detection of Six Food-borne Pathogens
in Food Samples |
Completed |
Dr. Chen Shu,
University of Guelph |
Development of a Novel Protein Chip-based Test for Rapid
and Cost-effective Salmonella Serotyping |
Completed |
Dr. Chen Shu,
University of Guelph |
Development of a Robust DNA Preparation Method to Enhance
Simultaneous Detection of Multiple Pathogens in Foods by a Microarray-based
Assay |
Ongoing |
| Dr. Griffiths Mansel, University of Guelph |
New Technologies for Improving Real-time PCR Methods
for Detection of Food-borne Pathogens |
Completed |
| Dr. Griffiths Mansel, University of Guelph |
Rapid Phage-based Method for the Detection of Pathogens
in Food |
Ongoing |
| Dr. Gyles Carlton, University of Guelph |
Validation of a Method for Determining the Species of Origin of
Contaminant E. coli
|
Completed |
| Dr. Hall Christopher, University of Guelph |
Fluorescence Polarization Immunoassays (FPIA) for Food Safety:
a Rapid Detection System for Pathogens and Chemicals
|
Ongoing |
Dr. Ismail A.A.,
McGill University |
Rapid Whole-Organism Identification Methods Based on
Fourier Transform Infrared (FTIR) Spectroscopy |
Ongoing |
| Dr. McEwen Scott, University of Guelph |
Campylobacter Isolation Methodology and Molecular
Characterization |
Completed |
| Mitchell Mark, University of Guelph |
Development and Validation of a Screening Protocol for
Identifying Sulfamethazine-violative Swine Carcasses at Ontario Abattoris
Using the CHARM ROSA™ Sulfamethazine Test |
Completed |
| Dr. Odumeru Joseph, University of Guelph |
Application of the Impedance and Colorimetric Systems
for Rapid and Cost Effective Detection of Listeria Species
in Food and Environmental Samples |
Completed |
| Dr. Odumeru Joseph, University of Guelph |
Evaluation of Immunoassay Based Kits with High Sensitivity
and Specificity for Rapid Detection of E. coli O157:H7 in Foods
|
Completed |
| Dr. Schraft Heidi, Lakehead University |
Detection of Campylobacter jejuni by Fluorescent
in Situ Hybridization |
Completed |
| Spilsbury Louise, University of Guelph |
Development of an Analytical Method for the Confirmation
of Sulfonamides in Animal Tissues |
Completed |
Risk Assessment (RA)
|
Lead Researcher
|
Project Title
|
Status
|
| Dr. Allan Brenda, University of Saskatchewan |
Identification of Bacterial Components that Influence
Colonization of Poultry by Campylobacter jejuni |
Completed |
| Dr. De Lange, C. F. M., University of Guelph |
Liquid Feeding of Swine - Potential Positive and Negative
Impacts on Pork Safety |
Completed |
| Dr. Friendship Robert, University of Guelph |
Surveillance of Ontario Pig Farms for Diseases of Public
Health Significance |
Completed |
| Dr. Griffiths Mansel, University of Guelph |
Investigation of Routes for Transfer of Food and Water-borne
Pathogens to Produce |
Completed |
| Dr. Holley Richard, University of Manitoba |
Evaluation of Pesticide Solutions in the Transmission
of Pathogenic Bacteria to Horticultural Crops |
Completed |
| Dr. Kelton David, University of Guelph |
Assessing the Incidence of Antimcrobial Resistant E.
coli and Salmonella Bacterial Isolates in Cull Cows from
Ontario Free Stall Dairy Herds
|
Completed |
| Dr. McEwen Scott, University of Guelph |
Occurrence of Enteric Pathogens and Antimicrobial Resistance
Patterns in Selected Retail Poultry Products and Human Cases in a
Southwestern Ontario County |
Completed |
| Dr. Ribble Carl, University of Guelph |
Occurrence of Enteric Pathogens and Antimicrobial Resistance
Patterns in Selected Retail Turkey and Veal Products in Southwestern
Ontario |
Completed |
| Dr. Warriner Keith, University of Guelph |
Establishment of Critical Control Points for Enteric
Pathogens in Beef Production |
Ongoing |
Risk Management
|
Lead Researcher
|
Project Title
|
Status
|
| Dr. Abernathy Tom McMaster University |
A Community Trial to Determine an Effective
Intervention for the Delivery of HACCP to the Food Service Sector
|
Completed |
| Dr. Gong Joshua, Agriculture and Agri-Food Canada, Guelph
|
Evaluation of Essential Oils as an Alternative to Dietary
Antibiotics to Control Food-borne Pathogens in Livestock |
Ongoing |
| Dr. Henson Spencer, University of Guelph |
Understanding Barriers to the Effective Implementation
of HACCP in the Ontario Food Processing Sector |
Completed |
| Dr. McAllister T. A. , Agriculture and Agri-Food Canada,
Lethbridge |
Evaluation of the Ability of Seaweed Extract (Tasco-14)
to Reduce the Duration and Intensity of Fecal Shedding of Escherichia
coli O157:H7 and Total E. coli by Cattle |
Ongoing |
| Dr. Sharif Shayan, University of Guelph |
Antibiotic Replacement Therapy for Control of Food-borne
Pathogens in Poultry |
Ongoing |
| Skinner Alison, Ontario Beekeepers' Association |
Enhancing the Food Safety of Honey Bee Hive Products
through the Use of Organic Beekeeping Practices and Effective Monitoring
of Pest and Disease |
Completed |
| Dr. Warriner Keith, University of Guelph |
Elimination of Human Pathogens on Seeds Destined for
Sprout Production Using a Novel Sanitizer |
Completed |
| Dr. Zhou H., University of Guelph |
Practical Optimization of Ozonation Process for Enhancing
Microbial Safety and Food Quality |
Completed |
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