Progress 04/15/17 to 04/30/21
Outputs
Target Audience:In the final year of the project, the target audiences included: Microbiologists working on microbial interactions, microbial ecology of the phyllosphere, microbiology of fermented foods, and other aspects of microbiome assembly Fermented food producers (including sauerkraut, kimci, and other fermented vegetables) and home fermented food producers Farmers of cabbages and other fermented vegetables Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?One graduate student, two post-docs, and four undergraduate students were trained during this project. They each learned skills in microbiome sequencing, plant tissue culture, and synthetic model microbiome development. They also learned new skills in project management, data visualization, and science outreach development. The graduate student is now working in an industry position in microbiome research. One post-doc is now an assitant professor of biology and the other is working as a genomic scientist for a genomics biotech. Three of the undergraduates are working in microbiology industry positions and one is applying for medical schools. How have the results been disseminated to communities of interest?We have disseminated results from our work through direct contact with stakeholders, at scientific conferences, through journal articles, and via science outreach portals. We have been working with the farmers and gardeners at sites where we sampled, and have communicated our findings via email to them and during in person visits. Results of this work have been presented via a poster presentations and talks by the graduate student at 8 national meetings ( including ASM Microbe,AgMicrobiomes Workshop, and the Cold Spring Harbor Microbiome conference)and at 18 talks by the PI at various university departments or conferences. As noted earlier, results of the work have been disseminated in four manuscripts (three published and one submitted for review). We have also disseminated our findings to the general public at outreach events through our "Microbiome Discovery Center." We have also conveyed findings from our results and related research on fermented vegetableson our fermented food outreach website MicrobialFoods.org. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
In the early phase of the project, we focused on identifying how large- and small-scale dispersal processes shape the cabbage phyllosphere microbiome. Our focus was on the distribution and diversity of the lactic acid bacteria as they are the key microbial group that ferments cabbage into products such as sauerkraut and other fermented vegetable products. We first created a network of 51 farms and community gardens to sample soil and leaves. Over the summer of 2017, we sampled each of these sites to to identify the potential sources of lactic acid bacteria in farm systems. Using culture-based and metagenomic amplicon sequencing approaches, we determined that lactic acid bacteria are relatively rare compared to other phyllosphere bacteria. Using our gnotobiotic cabbage system as well as isolates of phyllosphere bacteria from these sites, we experimentally identified why LAB are so rare in the cabbage phyllosphere. We have also used whole-genome sequencing of LAB from across the various sites to understand large-scale dispersal of these bacteria across farming systems. To move to field trials to test our observations from the gnotobiotic cabbage system, we built out an experimental garden where we will grow cabbages in the summer and fall of 2018. Using raised beds, we improved an existing experimental garden site on the Tufts University property that has not been used for the past several years. This results of this comprehensive research are described in our 2019 Applied and Environmental Microbiology publication (https://pubmed.ncbi.nlm.nih.gov/31003989). In addition to using the gnotobiotic cabbage system for the work described above, we test the utility of the system for other types of experiments including microbial interaction and growth experiments. We provided a detailed description of how this gnotobiotic plant system could be used by other researchers for other crops in our 2020 Journal of Visualized Experiments publication (https://pubmed.ncbi.nlm.nih.gov/32568237). Later phases of the project focused on microbiome diversity and assembly in fermented vegetable products. Fermented vegetables are increasing in popularity in North America, but little is known about the diversity of their microbial communities. Shotgun metagenomic sequencing was used to determine the bacterial and fungal diversity of 17 different fermented vegetable products (sauerkraut and kimchi) made in the United States and Canada. We found an average of 10.8 species of lactic acid bacteria (LAB) per product using metagenomic sequencing along with low levels (<1% relative abundance of Kazachstania) of yeast in some of the products. The most common LAB species was Enterococcus faecium, which was found in high abundance in all products. Other LAB detected in products were Leuconostoc mesenteroides, Lactobacillus (Lb.) plantarum, Lb. sakei, and Lb. brevis which are commonly isolated from most fermented vegetable products. Using culture-based quantification, we were surprised to find yeast in over half of the sampled products, in some instances at a greater cell count than LAB. Further experiments using in vitro fermentation and a six-member synthetic LAB and yeast community show that the presence of yeast is predominantly explained by the temperature of the fermentation, although the salt concentration and type of cabbage used in the fermentation can also influence yeast abundance. These results provide the first comprehensive survey of commercially available North American fermented vegetable products and demonstrate how temperature, salt, and ingredients can drive the community structure of bacterial and yeast communities in these products.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2019
Citation:
Miller ER, Kearns PJ, Niccum BA, O'Mara Schwartz J, Ornstein A, Wolfe BE. Establishment Limitation Constrains the Abundance of Lactic Acid Bacteria in the Napa Cabbage Phyllosphere. Appl Environ Microbiol. 2019 Jun 17;85(13):e00269-19. doi: 10.1128/AEM.00269-19. PMID: 31003989; PMCID: PMC6581170.
- Type:
Journal Articles
Status:
Published
Year Published:
2020
Citation:
Miller ER, O'Mara Schwartz J, Cox G, Wolfe BE. A Gnotobiotic System for Studying Microbiome Assembly in the Phyllosphere and in Vegetable Fermentation. J Vis Exp. 2020 Jun 3;(160). doi: 10.3791/61149. PMID: 32568237.
- Type:
Journal Articles
Status:
Awaiting Publication
Year Published:
2021
Citation:
Miller, ER, Dinh C, Anderson B, Edwards CB, and BE Wolfe. Diversity of yeast and lactic acid bacteria in fermented vegetable products sold in the United States. Submitted
|
Progress 04/15/17 to 04/14/18
Outputs
Target Audience:Over the past year, our research project has interfaced with over 50 small farms and community gardens throughout the Northeast. These farms range from the Hudson Valley of New York, all of Massachusetts, Connecticut, and Rhode Island, and up into the southern part of New Hampshire. Across all of these sites, we have been assessing the abundance and diversity of lactic acid bacteria at individual farms or gardens as part of Aim 1 of the research project. These are the primary bacteria that control the quality of vegetable fermentations, but their ecology from farm to ferment is poorly understood. We've communicated with this group directly through site visits, via emails, and through followup communications about results from samples/data collected at their sites. When sampling these sites, we have been able to describe our research and teach farmers and gardeners basic concepts in phyllosphere microbiology, microbial ecology, soil ecology, and food microbiology. We have also shared our findings with other phyllosphere microbiome scientists. This happens informally via email and phone conversations, but also at scientific meetings. The PI is also on the steering committeeof the NSF Ag Microbiomes Research Coordination Network and will be sharing results at a workshop at the International Plant Pathology Conference in Boston in summer of 2018. We have also interacted with the general public through outreach efforts that are part of ourMicrobiome Discovery Center. This is a portal microbiome outreach setup that we will be using at farmers' markets throughout New England in the next two years of the project. We piloted it last year at the Berkshire Fermentation Festival. Over 500 people visited the booth and learned about phyllosphere microbiomes through interactive exhibits and a live microscopy demonstration. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?One graduate student, one post-doc, and one undergraduate student were trained during the first year of this project. They each learned skills in microbiome sequencing, plant tissue culture, and synthetic model microbiome development. They also learned new skills in project management, data visualization, and science outreach development. How have the results been disseminated to communities of interest?We have been disseminating results from our first year of work through direct contact with stakeholders, at scientific conferences, through journal articles, and via science outreach portals. We have been working with the farmers and gardeners at sites where we sampled, and have communicated our preliminary findings via email to them and during in person visits. Results of this work have been presented via a poster by a graduate student atthe Boston Bacterial Meeting and at 8 talks by the PI at various university departments. The work will be presented at several scientific meetings during the summer of 2018.We are also writing up our first major manuscript which describe our observational and experimental datasets demonstrating dispersal limitation in the cabbage phyllosphere. This will be in review in June 2018.Another perspective piece that included insights gained from the gnotobiotic cabbage system was published in mSystems in early 2018. A description of our preliminary findings will also be posted on our fermented food outreach website MicrobialFoods.org. What do you plan to do during the next reporting period to accomplish the goals?During the next reporting period, we will transition to Aim 2, where we will be looking at how microbe-microbe interactions shape the diversity ofthe cabbage phyllosphere. Using a series of lab and field experiments, we will use synthetic microbial communities isolated in the past year to determine how biotic interactions shape the distributions of some of the major microbial taxa in the phyllosphere. We have built out an experimental garden on the grounds of Tufts University where we are carrying out field trials to confirm our lab findings. Another set of experiments over the next year will link our results from the phyllosphere to fermentation outcomes. These findings will have direct implications for food safety and quality. We will also further develop our Microbiome Discovery Center and deploy it at more farmers' markets throughout Massachusetts.
Impacts
What was accomplished under these goals?
Over the past year, we have focused on identifying how large- and small-scale dispersal processes shape the cabbage phyllosphere microbiome. Our focus has been on the distribution and diversityof the lactic acid bacteria as they are the key microbial group that ferments cabbage into products such as sauerkraut and other fermented vegetable products. We first created a network of 51 farms and community gardens to sample soil and leaves. Over the summer of 2017, wesampled each of these sites toto identify the potential sources of lactic acid bacteria in farm systems. Using culture-based and metagenomic amplicon sequencing approaches, we determinedthat lactic acid bacteria are relatively rare compared to other phyllosphere bacteria. Using our gnotobiotic cabbage system as well as isolates of phyllosphere bacteria from these sites, we have experimentally identified why LAB are so rare in the cabbage phyllosphere. We have also used whole-genome sequencing of LAB from across the various sites to understand large-scale dispersal of these bacteria across farming systems. To move to field trials to test our observations from thegnotobiotic cabbage system, we built out an experimental garden where we will grow cabbages in the summer and fall of 2018. Using raised beds, we improved an existing experimental garden site on the Tufts University property that has not been used for the past several years. We also developed and tested our Microbiome Discovery Center to begin the outreach component of this grant. We designed and implemented a farmers' market booth with a tent, posters, interactive exhibits, and live microscopy. We used this setup at one event in September 2018 to test our design. We received helpful feedback that we will use to improve the booth for the 2018 season.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2018
Citation:
Wolfe, B. E. (2018). Using Cultivated Microbial Communities To Dissect Microbiome Assembly: Challenges, Limitations, and the Path Ahead. mSystems, 3(2), e00161-17.
|
|