Culture- and amplification-independent metagenomics approach for sensitive detection and identification of viruses isolated from produce
Description: Foodborne viruses are the leading cause of foodborne illnesses outpacing all other pathogens combined including bacteria. According to CDC, about 59% of total cases of foodborne illness are caused by viruses annually in the United States, and 58% of total cases by norovirus alone. The severity and duration of illnesses resulting from viral foodborne outbreaks emphasize the need for development of sensitive and accurate detection techniques for viruses from food samples. However, the genetic diversity of foodborne viruses, low quantities of viruses on contaminated food, the non-culturable nature of most of these viruses, and the globalization of food market make it difficult to quickly and (and no less important) accurately detect virus in contaminated food with current methods. The purpose of this study is to develop and optimize a metagenomics approach toward sensitive and accurate detection and identification of foodborne viruses from food, specifically the produce commodities in this study. We propose to utilize next generation sequencing (NGS) technology as the basis for detection of foodborne viruses at low copy number and/or in a mixture of species. The approach in this project involves artificially inoculating produce samples with either single or mixed viral species at various levels, and directly analyzing the recovered viruses from inoculated produce samples by NGS without pre-culture and pre-amplification. Through optimization of NGS data generation and analysis methods, we anticipate sensitive detection and/or identification of viruses responsible and/or associated with outbreaks of viral foodborne transmission and illness. This study will provide an initial investigation with NGS into the detection and identification of viruses isolated from food and will provide the Agency with a sensitive and accurate protocol to identify viruses for outbreak response and surveillance.
Development of a method for isolation and detection of Cronobacter species from fresh vegetables and dried plant-origin foods
Description: Cronobacter species, formerly known as Enterobacter sakazakii, are a group of opportunistic foodborne bacterial pathogens. The genus Cronobacter is comprised of seven species: C. sakazakii, C. malonaticus, C. turicensis, C. muytjensii, C. dublinensis, C. universalis, and C. condimenti. Of the seven species, the primary pathogen is C. sakazakii. The status of Cronobacter, as a pathogen, was elevated to an international public health concern when contaminated samples of powdered infant formula (PIF) or follow-up formula (FUF) were recognized by the food safety community which linked its presence to several neonatal meningitis outbreaks. It is well- defined now that contamination of reconstituted, temperature-abused PIF occurs both intrinsically and extrinsically; the main reservoir(s) and routes(s) of contamination have yet to be established, however. Furthermore, reports from numerous surveillance studies have shown that Cronobacter species are found in a variety of foods including dried foods (spices, herbs, flour, and cereals) and fresh ready-to-eat vegetables. This increasing body of evidence suggests that plants may serve as a reservoir. Moreover, linking the epidemiology of adult cases to consumption of PIF is difficult to explain, suggesting that there are still unknown sources, such as other foods which may be involved in causing adult infections. The current Cronobacter isolation procedure described in the Bacteriological Analytical Manual only deals with the isolation of Cronobacter from PIF and it uses a genus level real-time PCR assay. This research project is about the development of a new isolation method to obtain Cronobacter species which are contaminating ready to eat vegetables, and other foods of plant origin. A species-specific real-time PCR assay will also be developed.
Generation of Chloroplast Genomes for FDAâ€™s GenomeTrakr CP and Assay Development
Description: To support its public health mission, the FDA must verify labeling of products. This can be difficult when products are mixtures or heavily processed botanicals. In this internship, the student will learn how to extract and sequence plant DNA using next generation sequencing techniques. Chloroplast genomes from FDA species of interest will be targeted and the intern will also process the data bioinformatically. This will help continue to build a library of DNA sequences which will give FDA the ability to address the issue of processed botanicals by providing sequence data from which identification techniques can be developed. This database has grown steadily and will be useful to those in government, academia and industry. In addition, the intern will learn how to develop targeted assays of plants using the chloroplast genomes. The methods will include polymerase chain reactions (PCR) and/or real-time polymerase chain reaction (RT-PCR).
Fate of Listeria monocytogenes in frozen ready to eat foods
Description: Understanding the behavior of Listeria monocytogenes in various food matrices is critical to the understanding of the risk associated with L. monocytogenes contamination in these foods. In recent years, a number of novel food vehicles of L. monocytogenes have been identified. Specifically, frozen ready to eat (RTE) foods, such as ice cream, frozen vegetables and frozen fruits have been associated with outbreaks and recalls due to L. monocytogenes contamination, yet we do not know the behavior of L. monocytogenes on various frozen RTE products. Therefore, we have designed a project to study the behavior of L. monocytogenes in frozen RTE products.
Evaluation and optimization of the validated FDA method for detection of Cyclospora cayetanensis in prepared dishes.
Description: The research project will study the presence of the parasite Cyclospora cayetanensis in fresh produce. Outbreaks of cyclosporiasis have affected thousands of persons in the U.S occurring in a multi-state fashion in different years. The outbreaks have frequently been linked to fresh produce, which in many of these outbreaks, was consumed as a complex matrix such as salad. In 2018, of the two main outbreaks that have occurred, one was linked to pre-packaged vegetable trays containing broccoli, cauliflower, carrots, and dill dip, and in the second, FDA confirmed the presence of Cyclospora in an unused package of romaine lettuce and carrot mix, as part of salads purchased in a fast-chain restaurant. For FDA to fulfill its mission to protect consumers and improve food safety, the agency must ascertain the performance of the C. cayetanensis regulatory method (BAM Chapter 19B) for the detection of the parasite in complex matrices. This research will improve the ability of the FDA to detect C. cayetanensis in a variety of food samples that could be implicated in cyclosporiasis outbreaks. The objective of this project is to evaluate the performance of the C. cayetanensis BAM method in fresh produce and complex food matrices and adjust the techniques for optimal detection when applicable. This will allow the FDA to effectively apply the C. cayetanensis BAM method for the testing of foods in different scenarios, including leftovers that could be directly implicated as the source of the infection. CORE, ORA, and possibly the Office of Compliance will be impacted by this research. During past investigations, ORA laboratories have had problems analyzing complex matrix samples linked to cyclosporiasis outbreaks. The information/methods will be used to develop tools for outbreak investigations, to confirm sources of contamination, and may consequently support FDA regulatory actions. The information will allow the FDA to identify the specific source of infection and develop appropriate control measures to maintain the integrity of the U.S. food supply. The method will be adapted for analysis of complex matrices including: ready to eat fresh leafy salads, berry salads, and Mexican and Mediterranean-style side dishes. In addition, modified approaches, which may be necessary to optimize recovery and detection of oocysts from complex matrices, will be evaluated and adjusted to obtain optimal detection sensitivity for each group of complex matrices.
Assessment of Listeria monocytogenes serotype 4bV growth in soil compared to traditional disease-causing serotypes
Description: Recent outbreaks involving a subset of Listeria monocytogenes strains termed 4bV associated with fresh produce have raised questions about whether these strains have increased fitness in agricultural environments and/or food processing facilities. This project is aimed to fill that knowledge gap to determine whether special considerations are needed for these strains.
Detection of foodborne parasites in waste and surface water samples
Principle Investigator: da Silva, Alexandre
Location: CFSAN MOD-1, 8301 Muirkirk Road, Laurel, MD
Objective: 1- Optimize the DEUF for the detection of C. cayetanensis, on DNA extracted from filtrated samples with the same characteristics of WWTP. 2- Apply newly developed molecular methods in conjunction with the DEUF method to detect and characterize C. cayetanensis on water samples artificially contaminated with C. cayetanensis. 3-Mimic the contamination of agricultural water with feces containing C. cayetanensis oocysts and use DEUF and molecular methods to detect the parasite in such artificially contaminated samples.
Project Needs and Duration: The applicant should be familiar with general microbiology and molecular biology. The estimated duration of the internship project is one year. Time requirements include 8-10 h/week during the school year and 30 h/week during break sessions.
Description: Intestinal parasites such as Cyclospora cayetanensis is a common cause of multi-state diarrhea outbreaks in the US. The outbreaks caused by these pathogens have been associated with consumption of contaminated produce or water and have affected thousands of persons in the US and abroad. Especially in the case of C. cayetanensis outbreak, cases have been linked to consumption of imported produce. It is not very clear how water or produce is contaminated with these pathogens, but epidemiological data indicate that contaminated agricultural water could play a role in contamination of produce during irrigation or produce wash. The objective of this project is to analyze effluent samples from waste water treatment plants (WWTP) using the ultrafiltration method for detection of intestinal parasites and viruses. The ultrafiltration method to be used in this project uses hollow fiber filters (ultrafilters) which are designed to trap particles as small as 30 KDa. Therefore, samples concentrated using this method could be analyzed for several types of pathogens, especially those that are non-culturable and cannot be enriched biologically. The DEUF method is currently being evaluated for recovery of C. cayetanensis in agricultural water but it has been extensively used for recovery and detection of viruses in tap water and storm water. The samples processed by DEUF can also be used to study the microbiome of sewage or waste water with a goal to detect and ID the target potential produce contaminants. Such samples will be provided through ongoing studies led by North Carolina State University and Emory University. This project is part of OARSAâ€™s Produce Safety Research Consortium and has the potential to have a direct impact on FSMA policy decision-making.
Development and Validation of a Rapid Detection Method for Campylobacter in Leafy Green Produce
Description: The presence and persistence of the foodborne bacteria, Campylobacter, on fresh produce is not well known, and most likely underreported since there are currently no validated methods for detecting and isolating Campylobacter from fresh produce. High genetic diversity also makes it difficult to isolate and identify different pathogenic species of Campylobacter. The proposed research project intends to develop and validate a new method for BAM to detect Campylobacter species in leafy green produce. We will also compare various growth conditions (temperature, culture media, microaerophilic, spoilage) to understand the prevalence and persistence of Campylobacter in leafy green produce, which will provide data to assist in risk assessment analyses.
Field Portable Arsenic Speciation
Description: The purpose of this project is to adapt a commercial off-the-shelf water testing kit to measure inorganic arsenic in rice and infant rice cereals. This method will help satisfy the FDA proposed limit of 100 part per billion inorganic arsenic in infant rice cereal limit without requiring time- consuming methods and expensive instrumentation.â€ƒ
Advancing preventative food safety methods through the use of portable, near-infrared devices
Description: A priority for FDA has been on the development of improved, rapid screening methods to evaluate food content and authenticity. Recent advances in instrument technology have allowed a number of compact spectrometers to appear in commerce as low-cost devices. It is important for FDA to understand the analytical performance of these simplistic devices. Further, these devices have the potential to allow for portable, rapid, high-throughput methods that could be employed for FDA field use if the technology is accurate and sensitive for particular applications. This research will focus on evaluating a near infrared portable device in areas of FDA need (e.g., rapid screening of milk powders and olive oils) for potential future field applications and protection of public health.
Development of High Resolution Screening Procedures for Pesticide Screening in Foods
Description: One of the roles of the FDA's food safety program is to monitor for the presence of chemical residues and contaminants such as pesticides in foods. The FDA is responsible to enforce pesticide tolerances that are established by the U.S. Environmental Protection Agency. Since there are many pesticides and food types, improvement of current methods or development of methods to identify and quantify these potentially harmful chemicals presents many challenges. This research involves new technology, improvement of current procedures, expansion to new residues and chemicals and food matrices; and establishment of validated methods to be used for regulatory monitoring and surveillance. This research will involve the use of liquid chromatography-high resolution mass spectrometry, particularly quadrupole-orbitrap, to expand and improve current FDA procedures for the analysis of pesticides and other residues and contaminants (veterinary drugs, mycotoxins) in related foods and other related FDA regulated products. These applications will result in the improvement or complement existing procedures by providing rugged, rapid and robust multi-residue methods suitable for FDA and other laboratories using FDA methods and procedures.
Assessment of media perception/interpretation of communications messaging on CFSAN issues.
Description: Members of the OFVM Strategic Communications Outreach and Public Engagement (SCOPE) team in collaboration with CFSAN program office staff spend much of their time developing and clearing materials to communicate the Centerâ€™s policies and actions to media, consumers, and other interested parties through social media. The media serve as a conduit for the FDA to communicate to consumers, industry, Congress, state, local and international governments, and other stakeholders. Therefore, it is supremely important that our communications be effective. Currently, although we monitor news stories and capture them in a weekly media report, we donâ€™t have the resources to adequately evaluate the effectiveness of our efforts. Investing time to retrospectively evaluate the success of our communications strategies will help us better understand for which issues/topics we are communicating clearly and reaching the target audience. This helps inform decisions about future communications strategy and policy ventures, helping SCOPE and CFSAN staff work smarter not harder on the development and clearance of communications material.
Post Response Outbreak Data Analysis Support Project
Principle Investigator: Craig Cloyd, Tami
Location: CFSAN University Station, 4300 River Road, College Park, MD
Objective: â€¢ Perform analysis of FDA outbreak investigation data to provide summary statistics (e.g., number of outbreaks per year, number of illnesses, percentage of illnesses associated with specific pathogens or commodities, etc.) â€¢ Compile existing historical outbreak inspectional data associated with similar commodities, and analyze data for similar observations, deficiencies and corrective actions â€¢ Compile outbreak traceback data and assist with the analysis of the data to identify trends and repeat firms among outbreaks associated with specific commodities (e.g., leafy greens, sprouts, flour, etc.) â€¢ Perform a literature review and research regarding Cyclospora contributing to a peer-reviewed publication summarizing Cyclospora outbreak investigations and prevention activities â€¢ Review and verify CORE Post Response outbreak data with existing reference information and
Project Needs and Duration: â€¢ Applicable Major: Biology, Microbiology, Food Safety, Public Health, Food Science, Public Health (Epidemiology) â€¢ Knowledge of Excel, Microsoft Word and PowerPoint â€¢ Knowledge of PubMed, Web of Science, or ability to perform scientific literature searches â€¢ Ability to analyze data and summarize information. Attention to detail â€¢ Ability to work independently The estimated duration of the internship project is one year. Time requirements include 8-10 hours per week during the school year and 30 hours per week during break sessions.
Description: â€¢ Analysis of FDA outbreak data to contribute to 2019 public health conference presentations, and to support prioritization of FDA policy and any applicable CFSAN Workplan assignments â€¢ Compilation and analysis of inspectional data from outbreaks associated with similar commodities to support future outbreak investigations and regulatory decisions â€¢ Assistance with the analysis of outbreak traceback data to support future outbreak investigations and compliance activities â€¢ Literature review regarding Cyclospora outbreaks contributing to a peer-reviewed publication and FDA Cyclospora Taskforce objectives â€¢ Verification of CORE Post Response outbreak data to support improvement of future outbreak investigations and FDA regulatory activities
Content Analysis and Variation in State Retail Regulations and Retail Food Safety Guidance
Description: This project will involve examining and comparing state retail food regulations with the FDA Food Code and conducting content analysis of retail food safety interventions and laws enacted in all 50 states and the District of Columbia. CFSANâ€™s Retail Food Protection Staff (RFPS) is attempting to better understand and document: 1. The presence, similarities/differences of various Food Code related provisions and definitions in state Food Codes 2. The equivalency of various state Food Code provision with the FDA Food Code 3. Provisions that are present in state Food Codes but absent in the FDA Food Code 4. The types, similarities, differences, and characteristics of the retail food safety laws being enacted at the state level 5. How regulatory partners at the state, local, tribal, and territorial levels incorporate Food Code provisions into their regulatory documents and use the internet to deliver retail food safety information to their stakeholders. This information will be used to inform decisions regarding the technical assistance to be provided to regulatory retail food safety professionals by FDA to better understand policy diffusion, prioritize research, and evaluate the need to make potential changes to the FDA Food Code. The FDA publishes the Food Code, a model code that assists food control jurisdictions at all levels of government by providing them with a scientifically sound technical and legal basis for regulating the retail and food service segment of the industry (restaurants and grocery stores and institutions such as nursing homes). Local, state, tribal, and federal regulators use the FDA Food Code as a model to develop or update their own food safety rules and to be consistent with national food regulatory policy. To date, all 50 states, the District of Columbia, and 3 Territories have adopted some version of the FDA Food Code. RFPS manages the activities necessary to promote the adoption of the Food Code and to create an enhanced regulatory environment for retail food operations.