Progress 06/15/23 to 06/14/24
Outputs
Target Audience: The target audience for this project includes crop growers, agricultural personnel, environmental scientist, agricultural and industry stakeholders, and other researchers. The ability to improve the nutrient utilization by plants is beneficial to agriculture and the various stakeholders involved in research, development, and production. We will share our results and knowledge with the scientific community via peer-reviewed publications and scientific conferences, and through public engagement using extension articles and media. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project has provided the opportunity to train two undergraduate students, three graduate students and one postdoc scientists in interdisciplinary areas that integrate agricultural science, environmental engineering and computer and data science. The trainees learned new techniques in environmental microbiology, environmental engineering, soil chemistry, plant biology, soil science, and microbiology. The interdisciplinary training and a range of techniques gained in above areas will allow the students to advance in their careers. How have the results been disseminated to communities of interest?We have presented relevant preliminary results and shared our experimental approach and methods with scientific audience and communities with journal publications and at seminars, USDF PI meeting, workshops and conferences. In addition, we are currently working on developing five manuscripts based on the results generated: M Baldwin, Z Wang, P He, G Li, Y Yan, S Lee, J Han, H Yoon, Y Sang, ..and April Z Gu. Function-Targeted Single-Cell Raman Spectroscopy (SCRS)-enabled Phenotyping and Genotyping of Winter Wheat Rhizomicrobiome Reveals the Links of sub-surface Microbial Phosphorus Accumulation with Above-Ground Plant Health. AGU Fall Meeting Abstracts 2023, B44B-04 PeiSheng He, Nathan J. Sturdevant, Miguel A. Pineros and April Z. Gu "Assessment of Bioavailability of Polyphosphates and Impactions on Plant P Uptake and Phenotypes." Under revision Peisheng He 1, †, Yejin Son 2, †, Jennifer Berkowitz 2, Guangyu Li 1, Jangho Lee 1, IL Han 1, Eric Craft 3, Miguel Piñeros 2, 3, Jenny Kao-Kniffin 2, April Z. Gu 1, * Recycled Phosphorus Bio-Amendments from Wastewater Impact Rhizomicrobiome and Benefit Crop Growth: Sustainability Implications at Water-Food Nexus." Under revision Guangyu Li, Zijian Wang, Chieh Wu, David R. Kaeli, Jennifer G. Dy, April Z. Gu*"Towards High-Accuracy Bacterial Taxonomy Identification based on Phenotypic Single-Cell Raman Spectroscopy (SCRS) Data ." Under revision. What do you plan to do during the next reporting period to accomplish the goals?For the next reporting period, we plant to make progress towards all three objectives as described above. For objective 1, We will continue to finish the AI-enabled computation pipeline for SCRS-based phenotyping for agricultural applications and will work on generating a publication on this. We will also continue to establish the Raman-activated single cell (RASC) sorting coupled with downs-stream genomics analysis for soil samples applications. For Objective 2, we will finish all the data analysis for the collected a large amount of new rhizosphere soil samples with various crop genotypes, different growth stages and varying nutrient treatment conditions. We have developed outline based on these results for publications. For Objective 3, we expect to finish the remaining FACS, SCRS analysis of all the treatment samples. Specifically, we will compile all the data from all the objectives, preparing for generating products.
Impacts
What was accomplished under these goals?
Research Objective 1: Develop an integrated Single cell Raman microspectroscopy- FACS- genome sequencing plat form to discover PAOs in rhizosphere in relevant crops. For objective 1, for this period, we further refined and improved more automated SCRS raw data processing pipeline to enable more efficient SCRS data analysis. In addition, we improved the algorithm for downstream data analysis methods for rhizomicrobiome phenotyping. For this period, we have received the genomic sequencing data and have finished initial data analysis for rhizosphere soils samples from a variety of crops genotypes including wheat , sorghum, maize and other crops for our survey study. We subsequently applied this method in rhizosphere soils associated with a variety of crops in New York (NY) which revealed the unprecedented ubiquity of polyphosphate accumulating organisms (PAOs) in natural and agricultural niches. In this period, we finished most of the SCRS data acquisition and FACS-sorted cells 16s rRNA gene sequencing data, and the results are being organized into publication products. Single cell Raman spectroscopy (SCRS) for phenotyping PAOs in soil and rhizosphere samples were developed, and employed for a variety of crops. We developed algorithm to optimize the SCRS sampling size with the extended the use of operational phenotypic unit (OPU) clustering analysis, and more in-depth insights on rhizosphere microbial communities were revealed on the molecular, single-cell, and community level. In order to validate the capability and reproducibility of the SCRS platform, we are improving technological and computational modules for comparative biology correlation of genotypic and phenotypic signals, for the time-series detection of metabolic status of single cells and their network structures, for the visualization of molecular phenotyping cellular local structures via dimensional reductions, for the identification of molecular cellular heterogeneity within strains, and for the classification and prediction of intra-strain molecular cellular heterogeneity. For this period, we continued to develop and improve these computation modules. Research Objective 2: Investigate the activity of PAOs in the rhizosphere in relation to crop yield. We established the protocols and methods for monitor the PAOs in soil samples and we employed the methods for a variety of crops. For this period, we continued and finished the samples FACS, SCRS, genomics analysis for the NY crops survey study and the results are being summarized into publication products. Our findings revealed that PAOs were consistently present in the rhizosphere of all crop types, comprising approximately 1-10% of the total microorganisms, regardless of the specific crop or farming method. Our preliminary results showed that the composition of PAOs exhibited temporal changes at different crop growth stages. To further elucidate the activities and impacts of PAOs on crops, we also performed soil nutrients profiling, particularly P species including labile P, polyP, metal-bound P, humic-P, enzyme accessible P and proton accessible P etc. The novel results elucidated that below-ground P Species profiling were correlated with certain PAOs, highlighting the potential functional role of PAOs community in altering nutrient availability. We performed additional sampling of 10 rhizosphere soil samples associated with 10 different types of maize genotypes in New York state. We have finished samples pre-treatment, FACS soring, but the SCRS analysis and sequencing analysis are still in process. Research Objective 3: Explore the Application of PAOs Augmentation for Enhancing Bioavailable P and Plant P Utilization. With the improved SCRS data computation methods developed in this period, we re-analyzed the data from greenhouse experiment and the results was used to generate one journal manuscript that was submitted and under review. An environmentally controlled green-house experiment was developed in order further elucidate the impact of PAOs augmentation on enhancing bioavailable P and plant P utilization with sorghum seedlings in temperature-, light-, and moisture-controlled greenhouse. The data from FACS and SCRS analysis are still in the analysis. More insights will be obtained upon the completion of all the sample phenotyping analysis and 16s genomics analysis. The results will contribute to anticipated 1-2 publications.
Publications
|
Progress 06/15/22 to 06/14/23
Outputs
Target Audience:The target audience for this project includes crop growers, agricultural personnel, environmental scientist, agricultural and industry stakeholders, and other researchers. The ability to improve the nutrient utilization by plants is beneficial to agriculture and the various stakeholders involved in research, development, and production. We will share our results and knowledge with the scientific community via peer-reviewed publications and scientific conferences, and through public engagement using extension articles and media. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project has provided the opportunity to train two undergraduate students, fourgraduate students and two postdoc scientists in interdisciplinary areas that integrate agricultural science, environmental engineering and computer and data science. Two graduate students were trained to conduct machine learning based analytics. The trainees learned new techniques in environmental microbiology, environmental engineering, soil chemistry, plant biology, soil science, and microbiology. The interdisciplinary training and a range of techniques gained in above areas will allow the students to advance in their careers. How have the results been disseminated to communities of interest?We have presented relevant preliminary results and shared our experimental approach and methods with scientific audience and communities with journal publications and at seminars and conferences (see products section). In addition, we are currently working on developing five manuscripts based on the results generated. What do you plan to do during the next reporting period to accomplish the goals?For the next reporting period, we plant to make progress towards all three objectives as described above. For objective 1, we need to overcome some of the challenges we encountered regarding to the cell recovery rate from various soil samples. We will evaluate alternative approaches in soil sample pre-treatment and SCRS signal acquisition conditions with aim to improve cell recovery rate. We will continue to work on the AI-enabled computation pipeline for SCRS-based phenotyping for agricultural applications. We will also continue to establish the Raman-activated single cell (RASC) sorting coupled with downs-stream genomics analysis for soil samples applications. For Objective 2, we just collected a large amount of new rhizosphere soil samples with various crop genotypes, different growth stages and varying nutrient treatment conditions. We will finish the FACS and SCRS samples analysis for all the soil samples obtained from variety of crops. We also expect to complete the 16s genomics analysis of both original rhizosphere soil samples, and sorted samples with targeted phenotypic traits from FACS and RASC. For Objective 3, we expect to finish the remaining FACS, SCRS analysis of the polyp augmentation greenhouse experiments, as well as 16s analysis of all the treatment samples. Specifically, we will expand our phenotyping library for the rhizosphere related to key crops, we will identify and discover more unknown PAOs and deduce their potential roles in plant P uptake.
Impacts
What was accomplished under these goals?
Research Objective 1: Develop an integrated Single cell Raman microspectroscopy- FACS- genome sequencing plat form to discover PAOs in rhizosphere in relevant crops. For objective 1, for this period, we identified a technical issue related to cell FACS sorting recovery rate, which hinders its application for larger number of samples. We then further optimized the FACS protocols to significantly improve the cell recovery rate. We also added more automated SCRS raw data processing pipeline and improved the algorithm for data analysis methods to quantify and sort PAOs from soil samples for rapidly analysis of spatiotemporal dynamics of PAOs in rhizosphere. In this period, we collected more rhizosphere samples and finished partial SCRS data acquisition and FACS-sorted cells 16s rRNA gene sequencing data, and the data are still under analysis. We also employed this method for quantitatively measure and monitor the abundance and changes in the polyphosphate-accumulating organisms (PAOs) in crops during the various growth stages from seeding to mature over time. The samples are still being processed and analyzed. Single cell Raman spectroscopy (SCRS) for phenotyping PAOs in soil and rhizosphere samples were developed, and employed for a variety of crops. With Raman signatures being able to identify key intracellular metabolites such as PolyP, in this period, we proposed and developed a feature selection computation algorithm to cluster cells based on their SCRS signatures, and identify signature regions/peaks unique to each treatment or sample. We extended the use of operational phenotypic unit (OPU) clustering analysis, and more in-depth insights on rhizosphere microbial communities were revealed on the molecular, single-cell, and community level. Currently, we are working on the establishment and validation of the Raman-activated cell sorting coupled with single cell genomics platform. Our initial attempt in crop rhizosphere soil encountered several technical issues including too-low signal, low recovery rates. We are currently working on resolving these issues. Single-cell Raman Spectroscopy (SCRS) stands out as a particularly valuable tool for phenotypic and genotypic insights due to its non-intrusive and rapid capabilities. However, the limited availability of extensive SCRS datasets and inconsistent model selection hinder its standardized application across scientific disciplines. In this landmark study, we rigorously analyze and compare five dimensionality reduction (DR) methods with ten classifiers, using an expansive SCRS dataset meticulously labeled for taxonomy and cellular growth stages. In order to validate the capability and reproducibility of the SCRS platform, we are developing computational modules for comparative biology correlation of genotypic and phenotypic signals. Such AI-powered single-cell biotechnology platform offers novel complementary information to the current mainstream multi-omics techniques. For this period, we continued to develop and improve these computation modules. Research Objective 2: Investigate the activity of PAOs in the rhizosphere in relation to crop yield. We established the protocols and methods for monitor the PAOs in soil samples and we employed the methods for a variety of crops. For this period, we continued and finished the samples FACS, SCRS, genomics analysis for the NY crops survey study. Our findings revealed that PAOs were consistently present in the rhizosphere of all crop types, comprising approximately 1-10% of the total microorganisms, regardless of the specific crop or farming method. PAOs found in wheat, soybean, and corn grown in conventional farms had the least intracellular PolyP content, while PAOs inhabiting the rhizosphere of corn, pea, and alfalfa in organic farms, as well as conventionally grown triticale. We encountered some issues with the genomics sequencing due to problems at the Cornell Bio-Center that provided the sequencing service, and we had to re-send the samples to a different sequencing service center. This led to some delay in obtaining the sequencing data, which are currently under analysis. To further investigate the presence and activity of PAOs in relation to the crop yield, we sampled rhizosphere crop soils samples in a field testing during an entire growth period in collaboration with plant scientist at CALS in Cornell, where we conducted simultaneous monitoring and identification of PAOs and crops above-ground traits that were monitored via GIS remote sensing, such as chlorophyll content, leaf area index, global surface water index and normalized difference vegetative index etc. Our preliminary results showed that the composition of PAOs exhibited temporal changes at different crop growth stages. The key PAOs were also identified for the first time, including Sphingomonas and Delftia. Correlation analysis revealed the specific PAOs taxonomic groups that exhibited significant association with different crop yield and traits parameters derived from GIS. To further elucidate the activities and impacts of PAOs on crops, we also performed soil nutrients profiling, particularly P species including labile P, polyP, metal-bound P, humic-P, enzyme accessible P and proton accessible P etc. The novel results elucidated that below-ground P Species profiling were correlated with certain PAOs, highlighting the potential functional role of PAOs community in altering nutrient availability. To make up for the lost sampling reason in the Covid year, we performed additional sampling of 10 rhizosphere soil samples associated with 10 different types of maize genotypes in New York state. Upon receiving, the soil samples were subject to pre-treatment for the subsequent phenotypic analysis via SCRS and FACS as well as downstream genomic analysis. Pending the phenotypic analysis and time-sensitive genomic level analysis, samples were stored at 4°C or -80°C for other analyses later. We have finished samples pre-treatment, FACS soring, but the SCRS analysis and sequencing analysis are still in process. Research Objective 3: Explore the Application of PAOs Augmentation for Enhancing Bioavailable P and Plant P Utilization. With the improved SCRS data computation methods developed in this period, we re-analyzed the data from greenhouse experiment. Corn was cultivated under different fertilization regimes using biomass recycled from wastewater and Vermont green manures for a month before the plant growth and soil microbial communities were subsequently compared. Interestingly, the growth pattern of corn greatly differed depending on the fertilization methods employed. Further, distinct patterns were observed between the microbial community compositions present in the different soil treatments. Using 16S rRNA amplicon sequencing, we identified the development of different PAO communities between the soils of corn treated with synthetic EBPR sludge and those treated with Vermont manures. Our results revealed that Candidatus Accumulibacter spp. Is one of the most dominant species in activated sludge in the EBPR, can inhabit the rhizosphere of corn, take up its own niche, and proliferate throughout the soil. The results indicate that wastewater recycled biomass-originating PAOs represent a promising P biofertilizer that can potentially increase the growth and development of corn. An environmentally controlled green-house experiment was developed in order further elucidate the impact of PAOs augmentation on enhancing bioavailable P and plant P utilization with sorghum seedlings. Soil samples were taken at the beginning and the end of the growth period and subject to subsequent FACS and SCRS analyses. Portion of the FACS and SCRS analysis were completed. The student who was trained to perform FACS and SCRS analysis went on medical leave, and thus a delay in the analysis. More insights will be obtained upon the completion of all the sample phenotyping analysis and 16s genomics analysis.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
He, P., Son, Y., Berkowitz, J., Li, G., Baldwin, M.T., Wang, Z., Han, I., Lee, J., Kao-Kniffin, J., Gu, A.Z. Phenotypic and Genotypic Dynamics of Polyphosphate-Accumulating Organisms (PAOs) in Corn Rhizosphere in Response to Different Microbial Amendments and Phosphorus Sources. 2022 Phosphorus Week: Phosphorus Forum and Sustainable Phosphorus Summit, Raleigh, NC, November 1-4, 2022.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
Baldwin, M.T., He, P., Zhan, W., Han, I., Zhou, G., Yoon, H., Reid, M.C., Holeck, K., Rudstam, L., Gu, A.Z. Combined Tools to reveal the role of Polyphosphate-Accumulating Organisms in P Flux Dynamics in Oneida Lake Sediment and Impact of Dissolved Oxygen Level. 2022 Phosphorus Week: Phosphorus Forum and Sustainable Phosphorus Summit, Raleigh, NC, November 1-4, 2022.
- Type:
Journal Articles
Status:
Submitted
Year Published:
2023
Citation:
Guangyu Li, Zijian Wang, Chieh Wu, David R. Kaeli, Jennifer G. Dy, April Z. Gu* Towards High-Accuracy Bacterial Taxonomy Identification based on Phenotypic Single-Cell Raman Spectroscopy (SCRS) Data
- Type:
Journal Articles
Status:
Other
Year Published:
2023
Citation:
P. He, Z. Wang, Y. Son, G. Li, Il. Han, J. Kao-kniffin and A. Gu Enhancing Sustainable Agriculture: Exploring the Impacts of Phosphorus Recovery Bio-Amendments on Crop Growth and Rhizomicrobial Communities."
- Type:
Journal Articles
Status:
Other
Year Published:
2023
Citation:
PeiSheng He, Nathan J. Sturdevant, Miguel A. Pineros and April Z. Gu Assessment of Bioavailability of Polyphosphates and Impactions on Plant P Uptake and Phenotypes."
|
Progress 06/15/21 to 06/14/22
Outputs
Target Audience:The target audience for this project includes crop growers, agricultural personnel, environmental scientist, agricultural and industry stakeholders, and other researchers. The ability to improve the nutrient utilization by plants is beneficial to agriculture and the various stakeholders involved in research, development, and production. We will share our results and knowledge with the scientific community via peer-reviewed publications and scientific conferences, and through public engagement using extension articles and media. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project has provided the opportunity to train one undergraduate student, three graduate students and two postdoc scientists in interdisciplinary areas that integrate agricultural science, environmental engineering and computer and data science. The trainees learned new techniques in environmental microbiology, environmental engineering, soil chemistry, plant biology, soil science, and microbiology. The interdisciplinary training and a range of techniques gained in above areas will allow the students to advance in their careers. How have the results been disseminated to communities of interest?We have presented relevant preliminary results and shared our experimental approach and methods with scientific audience and communities with journal publications and at seminars and conferences. In addition, we are currently working on 2 manuscripts based on the results generated in this period. See details in the products section. What do you plan to do during the next reporting period to accomplish the goals?For the next reporting period, we plant to make progress towards all three objectives as described above. For objective 1, we will complete the AI-enabled computation pipeline for SCRS-based phenotyping for agricultural applications. We will also establish the Raman-activated single cell (RASC) sorting coupled with downs-stream genomics analysis for soil samples applications. For Objective 2, we will finish the FACS and SCRS samples analysis for all the soil samples obtained from variety of crops. We also expect to complete the 16s genomics analysis of both original rhizosphere soil samples, and sorted samples with targeted phenotypic traits from FACS and RASC. For Objective 3, we expect to finish the remaining FACS, SCRS analysis of the polyp augmentation greenhouse experiments, as well as 16s analysis of all the treatment samples. Specifically, we will expand our phenotyping library for the rhizosphere related to key crops, we will identify and discover more unknown PAOs and deduce their potential roles in plant P uptake.
Impacts
What was accomplished under these goals?
Research Objective 1: Develop an integrated Single cell Raman microspectroscopy- FACS- genome sequencing plat form to discover PAOs in rhizosphere in relevant crops. Year 3: 70% completion, estimated full completion in final year For objective 1, for this period, we further optimized the FACS protocols and data analysis methods to quantify and sort PAOs from soil samples for rapidly analysis of spatiotemporal dynamics of PAOs in rhizosphere. In order to quantify and identify polyphosphate accumulating organisms (PAOs), we developed and optimized two methods of phenotypic identification and sorting. The first of which centers around fluorescence-activated cell sorting (FACS) to quantify and sort microorganisms of interest from rhizosphere soils using fluorescence staining. In order to process rhizosphere samples, mechanical and chemical detachment of soil microorganisms from soil was develoed and optimized. Detached microbes were then separated and concentrated for fluorescent staining in FACS using a combination of SYBR Green II for DNA staining, Tetracycline for polyphosphate (PolyP) counter staining. We then coupled FACS with downstream genomic sequencing on sorted samples to further reveal the taxonomic and functional traits of microorganisms with desired phenotypes. Currently, the genomic sequencing data are still under analysis. We subsequently applied this method in rhizosphere soils associated with a variety of crops which revealed the unprecedented ubiquity of polyphosphate accumulating organisms (PAOs) in natural and agricultural niches. We also employed this method for quantitatively measure and monitor the abundance and changes in the polyphosphate-accumulating organisms (PAOs) in crops during the various growth stages from seeding to mature over time. Single cell Raman spectroscopy (SCRS) for phenotyping PAOs in soil and rhizosphere samples were developed, and employed for a variety of crops in New York (NY). SCRS reveals phenotypic and intrinsic biochemical fingerprints of soil microorganisms. With Raman signatures able to identify key intracellular metabolites such as PolyP and the use of operational phenotypic unit (OPU) clustering analysis, more in-depth insights on rhizosphere microbial communities were revealed on the molecular, single-cell, and community level. The high-resolution cellular metabolic state and phenotypic fingerprints was revealed by SCRS coupled with available cellular composition library . Since little is known of the identities of PAOs in soil, current mainstream genomic-based method can not detect unknown PAOs. SCRS provided cellular phenotypic profiles of both known and unknown unconventional PAOs and key microbial communities involved in P cycling. Currently, we are working on the establishment and validation of the Raman-activated cell sorting coupled with single cell genomics platform. In order to validate the capability and reproducibility of the SCRS platform, we established a pure-cultured soil-relevant bacterial database consisting of 36 soil agriculture-relevant strains growing at 4 growth stages belonging to 4 phyla, 15 families, and 16 genera with relevant functions on plant health and symbiosis. Five technological and computational modules were developed integrated into the RamanomeSpec software, including RamanTree for comparative biology correlation of genotypic and phenotypic signals, RamanMap for the time-series detection of metabolic status of single cells and their network structures, RamanVisual for the visualization of molecular phenotyping cellular local structures via dimensional reductions, RamanCluster for the identification of molecular cellular heterogeneity within strains, and RamanML for the classification and prediction of intra-strain molecular cellular heterogeneity. Such AI-powered single-cell biotechnology platform offers novel complementary information to the current mainstream multi-omics techniques . Research Objective 2: Investigate the activity of PAOs in the rhizosphere in relation to crop yield. Year 2: 65% completion, estimated full completion in final year We established the protocols and methods for monitor the PAOs in soil samples and we employed the methods for a variety of crops. 22 rhizosphere soil samples associated with 11 different types of crop plants were taken from 4 different farms (2 conventional and 2 organic) located close to Penn Yan, Cortland, and Sidney in New York state. Upon receiving, the soil samples were subject to phenotypic analysis via SCRS and FACS as well as downstream genomic analysis. Pending the phenotypic analysis and time-sensitive genomic level analysis, samples were stored at 4°C or -80°C for other analyses later. We have completed the FACS, SCRS analysis, and the data are being analyzed. 16s genomics analysis are under-going. In both SCRS and FACS analysis, PAOs were detected in all crops, with a relative abundance of 5-25% and 0.5-8% respectively. These novel results provided first-time evidence of the ubiquitous presence of PAOs in plant rhizosphere, suggesting its potential important role in plant growth. The abundance level varied among different crops with some have significant higher PAOs than others. Moving one layer of granularity deeper, from SCRS and FACS analyses, semi-quantified intracellular PolyP abundances ion crop rhizosphere microbial communities were found to be higher in organically-grown plant rhizosphere when compared to crops grown under conventionally growing practices across most paired. SCRS integrated with AI as described above enabled phenotypic profiling of the PAO communities in various crops. OPU clustering on all detected PAOs spectra showed that three dominant OPUs (OPU-1, -2, and -9) represented around 60% of PAOs detected via Raman phenotyping. Further, each sample contained at least one of these three OPUs (7 out of 22 contained two and 11 out of 22 contained all three). Research Objective 3: Explore the Application of PAOs Augmentation for Enhancing Bioavailable P and Plant P Utilization. Year 2: 70% completion, estimated full completion in final year An environmentally controlled green-house experiment was developed in order further elucidate the impact of PAOs augmentation on enhancing bioavailable P and plant P utilization. 60 replicated sorghum seedlings were grown in temperature-, light-, and moisture-controlled greenhouse. Chemical fertilizer, manure compost, and activated sludge from enhance biological phosphorus recovery were each applied to 20 sorghum as different treatments. Amounts applied were normalized to provide same amounts of total phosphorus to each sorghum seedlings. Further, to understand interaction with commonly occurring mycorrhizae, half of the sorghum seedlings in each treatment (10) were inoculated with AM fungal spores at the beginning of the total 6-week growth period. Soil samples were taken at the beginning and the end of the growth period and subject to subsequent FACS and SCRS analyses. Portion of the FACS and SCRS analysis were completed. NMR was also employed to detect polyp in soil samples. PAOs were detected in all treatment samples collected and the control group at the beginning of the growth period. In the control, about 0.7% PAOs were found, which could be seen as the background PAOs abundance from the soils used in this study. With the addition of manure compost and activated sludge, additional PAOs added to the soil at the same time contributing to around 0.3% and 1.1% respectively. PAOs were found to accumulate in sorghum rhizosphere during the growth period when treated with chemical fertilizer and manure compost but decreased when activate sludge was supplied. More insights will be obtained upon the completion of all the sample analysis and 16s genomics analysis.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2022
Citation:
Guangyu Li, Chieh Wu, Dongqi Wang, Varun Srinivasan, David R Kaeli, Jennifer G Dy, April Z Gu. Machine Leaning-based Determination of Sampling Depth for Complex Environmental Systems: Case Study with Single-Cell Raman Spectroscopy Data in EBPR Systems. Environmental Science & Technology, doi:https://doi.org/10.1101/2020.12.18.423496
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
Z. Wang, A. Gu, M. Reid, J. Kao-Kniffin, E. Craft, "Integrated biotechnology platform of single-cell Raman spectroscopy and advanced data analysis pipeline enables high-resolution phenotyping study of bacterial growth dynamics and cellular heterogeneity," in 2022 Association of Environmental Engineering and Science Professors (AEESP) Research and Education Conference, Jul. 2022.
- Type:
Other
Status:
Published
Year Published:
2021
Citation:
Invited Speaker, (2021), Function-targeted High-Resolution Phenotyping Platform to Deduce Genetics-Functions Relationships in Rhizomicrobiome for Promoting Plant Nutrients Utilization. Cornell Initiative of Digital Agriculture workshop, Ithaca, October 12th, 2021.
- Type:
Journal Articles
Status:
Other
Year Published:
2023
Citation:
P. He, Z. Wang, Y. Son, G. Li, Il. Han, J. Kao-kniffin and A. Gu. Ubiquitous and Significant Presence of Polyphosphate accumulating organism in rhizosphere of variety of crops in NY. In preparation.
- Type:
Journal Articles
Status:
Other
Year Published:
2023
Citation:
Yejin Son, Peisheng He, Jennifer Berkowitz, April Gu, Jenny Kao-Kniffin. Interaction between polyphosphate accumulating organism (PAO) and arbuscular mycorrhizal fungi (AMF) in sorghum bicolor. In preparation
|
Progress 06/15/20 to 06/14/21
Outputs
Target Audience:The target audience for this project includes crop growers, agricultural personnel, environmental scientist, agricultural and industry stakeholders, and other researchers. The ability to improve the nutrient utilization by plants is beneficial to agriculture and the various stakeholders involved in research, development, and business of agriculture and production. We will share our results and knowledge with the scientific community via peer-reviewed publications and scientific conferences, Cornell extension programs, and through public engagement using extension articles and media. We established collaborations with the USDA scientists. One of the USDA scientists become a visiting scholar in our lab. We provided laboratory experiences and instruction to two undergraduate students and2 graduate students. The two undergraduate students are continuing their participation on this project. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project has provided the opportunity to train one undergraduate student, two graduate students and two postdoc scientists in interdisciplinary areas that integrate agricultural science, environmental engineering and computer and data science. The trainees learned new techniques in environmental microbiology, environmental engineering, soil chemistry, plant biology, soil science, and microbiology. The interdisciplinary training and a range of techniques gained in above areas will allow the students to advance in their careers. How have the results been disseminated to communities of interest?We have presented relevant preliminary results and shared our experimental approach and methods with scientific audience and communities at seminars and workshops. In addition, we are published 2 manuscripts based on the results generated in this period, and working on 2 more manuscripts based on the results generated from this period. Due to COVID situation, we were not able to attend more conferences in person. And some conferences were canceled. What do you plan to do during the next reporting period to accomplish the goals?For the next reporting period, we plant to make progress towards all three objectives as described above. Specifically, we will expand our phenotyping library for the rhizosphere related to key crops. We plan to perform the field survey of various crops to identify and discover more unknown PAOsand other nutrient-relevant organisms. We will conduct more green house and field experiments with different types of cropsto examine the role and functions of PAOs in relevant to plant nutrient uptake and health. We will conduct lab and field testing of microbial in augmentation with different biosolids sources that contain PAOs such as wastes digestion product, manure digestion product, to explorehe Application of PAOs Augmentation for Enhancing Bioavailable P and Plant P Utilization.
Impacts
What was accomplished under these goals?
Due to Covid situation, the lab was shut-down for a period and the lab access was restricted. This has delayed some of your lab andfield experiments. The overall progress is a little behind the expected level. Research Objective 1: Develop an integrated Single cell Raman microspectroscopy- FACS- genome sequencing plat form to discover PAOs in rhizosphere in relevant crops. Year 2: 40% completion, estimated full completion in final year. For objective 1, we have cultured 36 rhizosphere microorganisms relevant to nutrient cycling and utilization and established the initial phenotyping library and database by performing single cell Raman spectroscopy. We further developed machine-learning based computation methods to achieve fast bacterial identification (4-20s per cell) with high accuracy (96.5%-99.1%) via their phenotypic traits . This enables further rhizomicrobiome phenotyping and fast identification of functionally relevant organisms (e.g., PAOs) under different growth stages. We also explored to establish the computational and statistical methodology to link the correlation of phenotypic fingerprints and phylogenetic genotypes, which holds the key to elucidating the functions and roles of PAOs in soil, and its relationship with plant growth. We have also established FACS protocols and methods to quantify and sort PAOs from soil and water samples for rapidly analysis of spatiotemporal dynamics of PAOs in rhizosphere and have employed the methods for real field soil samples. Research Objective 2: Investigate the activity of PAOs in the rhizosphere in relation to crop yield. Year 2: 40% completion, estimated full completion in final year We established the protocols and methods for monitor the PAOs in soil samples under different growth treatment conditions including variations in temperature, plant clipping, organic content and fertilizer types. We found that higher abundance of PAOs are associated with higher temperature, higher organic content and are impacted by the fertilizer types. In the initial testing with maize plants, the abundance of PAOs in the control groups remained the same, while it increased in manure treatment plants and decreased in bio-solids-amended treatments. Further investigation is needed to elucidate the fundamental factors that impact and govern the PAOs activities. Research Objective 3: Explore the Application of PAOs Augmentation for Enhancing Bioavailable P and Plant P Utilization. Year 2: 35% completion, estimated full completion in final year We have performed both greenhouse and field studies and expeirments to investigate the presence, abundance, characteristics and roles of PAOs in relevance to plant health.By employing single-cell Raman spectroscopy and 16S sequencing, PAOs are detected and quantified in agricultural media under different growth treatment conditions including sludge and composting. Phenotypic and genotypic dynamics of soil bacteria showed that some soil PAOs exhibited Raman phenotypic similarity to EBPR PAOs, but other unique PAO OPUs were also found in soil media. 16S sequencing results demonstrated the dominating roles of genusStreptomycesin rhizosphere with addition of sludge, whilePseudomonascontributed the most with the treatment of composting.In NCBI database and Silva Database, we retrieved 6, 159, 73 of fungi with PPK, PPX, PPN, and 197, 64 of archaea with PPK, PPX, and 4369, 10620 of bacteria with PPK, PPX. This enables us to identify more putative PAOs in rhizosphere and know the spatiotemporal dynamics of PAOs composition and structure in crop rhizosphere. In turn, this data mining process will guide us on further crops selection and application of PAOs based on the detected spatiotemporal PAO activities.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2021
Citation:
Status and Advances in Technologies for Phosphorus Species Detection and Characterization in Natural Environment-A Comprehensive Review
MM Alam, V Srinivasan, AV Mueller, AZ Gu
Talanta, 122458
- Type:
Journal Articles
Status:
Published
Year Published:
2021
Citation:
Unrevealed roles of polyphosphate?accumulating microorganisms
A Akbari, ZJ Wang, P He, D Wang, J Lee, IL Han, G Li, AZ Gu
Microbial Biotechnology 14 (1), 82-87
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Progress 06/15/19 to 06/14/20
Outputs
Target Audience:The target audience for this project includes crop growers, agricultural personnel, environmental scientist, agricultural and industry stakeholders, and other researchers. The ability to improve the nutrient utilization by plants is beneficial to agriculture and the various stakeholders involved in research, development, and business of agriculture and production. We will share our results and knowledge with the scientific community via peer-reviewed publications and scientific conferences, Cornell extension programs, and through public engagement using extension articles and media. We established collaborations with the USDA scientists. One of the USDA scientists become a visiting scholar in our lab. We provided laboratory experiences and instruction to two undergraduate students, 2 graduate students and 2 postdocs. The two undergraduate students are continuing their participation on this project. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project has provided the opportunity to train one undergraduate student, two graduate students and two postdoc scientists in interdisciplinary areas that integrate agricultural science, environmental engineering and computer and data science. The trainees learned new techniques in environmental microbiology, environmental engineering, soil chemistry, plant biology, soil science, and microbiology. The interdisciplinary training and a range of techniques gained in above areas will allow the students to advance in their careers. How have the results been disseminated to communities of interest?We have presented relevant preliminary results and shared our experimental approach and methods with scientific audience and communities at seminars and workshops. In addition, we are working on 1-2 manuscripts based on the results generated in this period: Invited Speaker, (2019), Function-targeted High-Resolution Phenotyping Platform to Deduce Genetics-Functions Relationships in Rhizomicrobiome for Promoting Plant Nutrients Utilization. Cornell Initiative of Digital Agriculture workshop, Ithaca, October 30th, 2019. Graduate student speaker (2019), Impact of Phosphorus Sources and Delivery Mechanisms on Plant P Uptake and Phenotypes in Hydroponic Culture". October 17th, 2019. Dongqi Wang, Peisheng He, Zijian Wang, Guangyu Li, Nehreen Majed, and April Z. Gu "Advances in Single Cell Raman Spectroscopy Technologies for Biological and Environmental Application", Current Opinion in Biotechnology, accepted, 2020 What do you plan to do during the next reporting period to accomplish the goals?For the next reporting period, we plant to make progress towards all three objectives as described above. Specifically, we will expand our phenotyping library for the rhizosphere related to key crops, we will identify and discover more unknown PAOs and other nutrient-relevant organisms. We will conduct more green house and field experiments with different types of crops to examine the role and functions of PAOs in relevant to plant nutrient uptake and health.
Impacts
What was accomplished under these goals?
Research Objective 1: Develop an integrated Single cell Raman microspectroscopy- FACs- genome sequencing plat form to discover PAOs in rhizosphere in relevant crops . Year 1: 30% completion, estimated full completion in final year For objective 1, we have developed and established the proposed novel tiered-sorting and phenotyping workflow to screen, identify PAOs in soil samples based on functional intracellular phenotypic signatures, followed by subsequent sequencing. We have optimized the key techniques for phenotyping whole microbial community and specific populations (i.e. PAOs, PHA-containing organisms) in soil samples. We have selected >40 rhizosphere microorganisms relevant to nutrient cycling and utilization and performed single cell Raman microspectroscpy analysis to establish the initial library and database for rhizosphere. This enables further rhizomicrobiome phenotyping and fast identification of functionally relevant organisms such as PAOs. Research Objective 2: Investigate the activity of PAOs in the rhizosphere in relation to crop yield. Year 1: 20% completion, estimated full completion in final year We established the protocols and methods for monitor the PAOs in soil samples under different growth treatment conditions including variations in temperature, plant clipping, organic content and fertilizer types. We found that higher abundance of PAOs are associated with higher temperature, higher organic content and are impacted by the fertilizer types. In the initial testing with maize plants, the abundance of PAOs in the control groups remained the same, while it increased in manure treatment plants and decreased in bio-solids-amended treatments. Further investigation is needed to elucidate the fundamental factors that impact the PAOs activities. Research objective 3: Explore the Application of PAOs Augmentation for Enhancing Bioavailable P and Plant P Utilization. Year 1: 30% completion, estimated full completion in final year We explored the augmentation of PAOs in maize to evaluate the impact of PAOs and polyphosphate on P utilization by the plant. A total of 30 maize plants were grown for 28 days receiving three different treatments. In control group, maize was grown in potting soil with no additional nutrient supply. In treatment group, maize was grown in potting soil supplied with solids containing PAOs from an lab-scale sequential batch reactor. Horticulture assays included measurement of plant height, root biomass, number of leaves, and stalk circumstance, to indicate plant nutrient uptake and health. Both short-term and long-term soil respiration tests confirmed that soil microbial respiration activities were increased in PAO-treated groups. Compared to controls, the PAO-treated plants had statistically higher plant height, more leaves and slightly larger stalk circumference.
Publications
- Type:
Journal Articles
Status:
Accepted
Year Published:
2020
Citation:
Dongqi Wang, Peisheng He, Zijian Wang, Guangyu Li, Nehreen Majed, and April Z. Gu Advances in Single Cell Raman Spectroscopy Technologies for Biological and Environmental Application , Current Opinion in Biotechnology, Invited article, accepted, 2020
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