Source: CORNELL UNIVERSITY submitted to NRP
METAGENOMIC DISCOVERY AND EXPRESSION OF CLASS IIA BACTERIOCINS FROM FERMENTED FOODS TO IMPROVE FOOD SAFETY AND QUALITY FROM FARM TO FORK
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1028144
Grant No.
2022-67011-36459
Cumulative Award Amt.
$120,000.00
Proposal No.
2021-09385
Multistate No.
(N/A)
Project Start Date
Jan 1, 2022
Project End Date
Dec 31, 2023
Grant Year
2022
Program Code
[A7101]- AFRI Predoctoral Fellowships
Recipient Organization
CORNELL UNIVERSITY
(N/A)
ITHACA,NY 14853
Performing Department
Food Science
Non Technical Summary
Humans have fermented foods for millennia to preserve plant and animal commodities, unknowing of their commensalism with microorganisms until recently. Bacteriocins are one of several molecules fermentation-relevant bacteria produce that protect agricultural commodities against the proliferation of human pathogens and spoilage microorganisms. The objectives of this project are to metagenomically identify and characterize known and unknown Class IIa bacteriocins encoded by microbial populations in fermented foods and then synthesize these bacteriocins to test for their potential as food preservatives. Use of metagenomic sequencing will accelerate the discovery of unknown Class IIa bacteriocins compared to traditional methods.Throughout the course of thispredoctoralfellowship, PD Jonathan Sogin will further develop technical, oral and written communication, mentoring, and project management skills. Jonathanwill expand his professional network by interacting with peers at conferences and through other networking opprotunities internal and externall to his institution, and will mentor other students within his lab and beyond. Dr. Randy Worobo will serve as Jonathan's primary mentor for the project, and will provide guidance and support when needed to achieve the objectives of this project. The successful completion of research and professional development activities associated with this project will aid in Jonathan's personal growth to become a leader in agriculture.
Animal Health Component
(N/A)
Research Effort Categories
Basic
80%
Applied
(N/A)
Developmental
20%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
71240101100100%
Knowledge Area
712 - Protect Food from Contamination by Pathogenic Microorganisms, Parasites, and Naturally Occurring Toxins;

Subject Of Investigation
4010 - Bacteria;

Field Of Science
1100 - Bacteriology;
Goals / Objectives
Goal:Contribute to sustainable agriculture practices by making the most efficient use of on-farm resources through the implementation of natural biological controls.Objective 1:Phylogenetic Reconstruction of Class IIa Bacteriocin Sequences from Publicly Available Whole Genome and Whole Metagenome Sequencing Data.Objective 2:Amplicon-Based Metagenomic Sequencing of Class IIa Bacteriocin-Encoding Genes from Fermented Foods.Objective 3: Purification and Analysis of Novel Class IIa Bacteriocins Expressed in E. coli.
Project Methods
Objective 1:Phylogenetic Reconstruction of Class IIa Bacteriocin Sequences from Publicly Available Whole Genome and Whole Metagenome Sequencing Data.-Search for known and unknown Class IIa Bacteriocins inwhole genome, whole metagenome, and protein sequences.-Compile a database of known and presumptiveClass IIa Bacteriocins found from the search.-Cluster known and presumptive Class IIa Bacteriocins into classes, describe the classes, and compare them to previously described classes.-Phylogenetically describe the distribution of Class IIa Bacteriocins in Bacteria.Objective 2:Amplicon-Based Metagenomic Sequencing of Class IIa Bacteriocin-Encoding Genes from Fermented Foods.-Design degenerate or multiplex primers to amplifynumerous classes of Class IIa Bacteriocins from environmental DNA.-Sequence Class IIa Bacteriocins from environmental DNA extracted from fermented foods and/or other sources via next-generation sequencing.-Identify known and presumptive Class IIa Bacteriocins from the selected sources to describe the distribution of Class IIa Bacteriocins in foods.Objective 3:Purification and Analysis of Novel Class IIa Bacteriocins Expressed in E. coli.-Identify presumptive Class IIa Bacteriocins of interestfor further study based on the results of Objectives 1 & 2.-Design gene blocks to express presumptive Class IIa Bacteriocins inE. coli; express novel Class IIa Bacteriocins.-Characterize novel Class IIa Bacteriocins within the context of food processing by conducting inhibitionassays ofspoialage and disease causing bacteria, and conducting pH, temperature, and protease stability tests.

Progress 01/01/22 to 12/31/23

Outputs
Target Audience:Research activity:General public and other scientists/peers, specifically those interested in food systems, food safety, food quality, and nutrition. Mentoring:In-training scientists, specifically those working with and for PD Sogin. Training:PD Sogin, in pursuit of his PhD from Cornnell University. Changes/Problems:PD Sogin graduatedsooner than expected in May 2023. The overall goals of training and workforce development weresuccessfully accomplished, but some of the objectives were not completed as originally written due to this time constraint. Furthermore, the computational aspect of Objective 1 was greater than anticipated, thus taking more time than originally expected, and affecting the completion of the other objectives. What opportunities for training and professional development has the project provided?During the course of this project PD Sogin participated in numerous professional development activities. Theseincluded training industry personnel in Juice HACCP principles, training and mentoring other students in his lab, substitute teaching a class a handful of times in his department and preparing materials to apply for jobs. The group NIFA Predoctoral Fellowship meeting that occurred was valuable for PD Sogin to hear from individuals at similar career stages. With the skills acquired with support from this grant, PD Sogin received his PhD in Food Science and Technology from Cornell University in May 2023. PD Sogin also secured employment as a Postdoctoral Associate where he now uses the skills he acquired with support from this grant to conduct and manage international food safety research. How have the results been disseminated to communities of interest?Research activity:Target audiences for research activitieswere the general public bypresentation of a seminar by PD Sogin at Cornell University; the general public byin-progressdissemination of peer-review journal article publication; other scientists through the presentation of research at PD Sogin's PhD defense; and the general public bypublication of PD Sogin's dissertation in fulfillment of PhD degree at Cornell University. Mentoring:Target audiences for mentoring were in-training scientists by dissemination of next-generation sequencing study design, library preparation, and data analysis; and peers by the dissemination of grant writing skills PD Sogin gained when applying to the NIFA predoctoral fellowship award. Training:PD Sogin gained professional networking experience via connections with industry partners made by supervisor Worobo; gained presentation experience through dissemination activities; gained grant writing experience by submitting this grant; gained job application and skills showcasing experience by applying for jobs towards the end of his PhD; and gained project management skills through the completion of activities planned throughout this grant. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Objective 1: Data was collected and is stillbeing analyzed and prepared for publication. Objective 2: This objective was reliant on data from Objective 1. Thus to accomplish the goal of learning to conduct next-generation sequencing analysis, and somewhat related to the original goal, the microbial communities of tempeh and various commercial kombucha products were analyzed and data from these studies are in review for publicationas academic journal articles. Objective 3:This objective was not be completed, as it was reliant on data from Objective 1, and because PD Sogin graduated in May 2023 before the termination of this award.

Publications

  • Type: Theses/Dissertations Status: Published Year Published: 2023 Citation: Sogin, J.H. 2023. Application of Rapid Diagnostics and Next-Generation Amplicon Sequencing to Address Product Quality in the Food Industry. Cornell University ProQuest Dissertations Publishing. 30425910.
  • Type: Journal Articles Status: Under Review Year Published: 2024 Citation: Sogin, J.H., and Worobo, R.W. 2024 High ethanol contents are not correlated to specific species in commercial kombucha products, but Lactobacillales are likely key contributors to lactic acid production and product differentiation. In Review.
  • Type: Journal Articles Status: Under Review Year Published: 2024 Citation: Sogin, J.H. and Worobo, R.W. 2024. Amplicon Metagenomic Analysis of Commercial Tempeh During Fermentation. In Review.


Progress 01/01/22 to 12/31/22

Outputs
Target Audience:Research activity: Research activity was provided to the general public through the presentation of a seminar by PD Soginat Cornell University. Other research activity is in the process of being prepared for publication. Mentoring: Mentoring other students was accomplished during this reporting period; PD Sogin trained other members of his lab how to do next-generation amplicon sequencing analysis. Training: PD Sogin interacted with private industry through his participation in training professionals juice HACCP with supervisor Worobo. PD Sogin assisted a private fruit business by helping them with environmental monitoring verification. Changes/Problems:PD Sogin will graduate sooner than expected in May 2023. The overall goal of training and workforce development will therefore have been successfully accomplished, but some of the objectives will not be completed as originally written. Objective 1: This objective is taking much longer than originally anticipated due tocomputation time and changing analytical methods, which are rapidly evolving in the field. The length of time to complete this objective is therefore longer than originally written, but will be completed as written otherwise. Objective 2: This objective was reliant on data from Objective 1. Thus to accomplish the goal of learning to conduct next-generation sequencing analysis, and somewhat related to the original goal, the microbial communities of tempeh and various commercial kombucha products were analyzed and data from these studies will be published as academic journal articles before the termination of this award. Objective 3: This objective will not be completed, as Objective 1 is taking much longer than expected, and PD Sogin is graduating sooner than originally expected. What opportunities for training and professional development has the project provided?During the course of this project PD Sogin participated in numerous professional development activities. These have included training industry personnel in Juice HACCP principles, training and mentoring other students in his lab, substitute teaching a class a handful of times in his department, and preparing materials to apply for jobs. How have the results been disseminated to communities of interest?Results from Objective 2 were presented at a research seminar by PD Sogin. Results are being prepared for dissemination to the public in the form of research articles published in academic journals. What do you plan to do during the next reporting period to accomplish the goals?PD Sogin will graduate sooner than anticipated in May 2023. Some of the objectives will not be completed because of this. Objective 1: Compile, analyze, and prepare data for publication. Publish research article in a scientific journal. Completed as written. Objective 2: Compile, analyze, and prepare data for publication. Publish research article in a scientific journal. This will be completed as adapted due to the unanticipated long time required to complete objective 1. Objective 3: N/A. This will not be completed.

Impacts
What was accomplished under these goals? Objective 1: Data for objective 1 was collected and is currently being analyzed and prepared for publication. Objective 2: Objective 1 took longer than expected and was required for objective 2. Therefore, to accomplish the next-generation sequencing training from objective 2, related albeit somewhat different projects were conducted. Data was collected, analyzed, and prepared for publication. Some of these manuscripts are currently in review. Objective 3: NA

Publications