The Use of Cyanobacteria Biofertilizers to Increase Crop Productivity, Improve Soil Health, and Agricultural Sustainability in Florida

Progress report for LS21-354

Project Type: Research and Education
Funds awarded in 2021: $242,000.00
Projected End Date: 03/31/2024
Grant Recipient: Florida International University
Region: Southern
State: Florida
Principal Investigator:
Dr. Sanku Dattamudi
Florida International University
Co-Investigators:
Dr. Mahadev Bhat
Florida International University
Dr. Saoli Chanda
Florida International University
Dr. Krishnaswamy Jayachandran
Florida International University
Dr. Leonard Scinto
Florida International University
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Project Information

Abstract:

Biofertilizers are defined as organic substances containing various microorganisms that improve soil and crop productivity. Cyanobacteria have become an important addition to biofertilizers due to their enhancement of crop production and soil health. However, abundance of cyanobacteria in water can develop hypoxic conditions and cause harmful effects to aquatic organisms. Lake Okeechobee and St. Lucie River of Florida are frequently subjected to cyanobacterial ‘blooms’ that degrade water quality and incur significant environmental, ecological, economic, and societal impacts. Cyanobacterial blooms covered 60% (894 km2) of Lake Okeechobee during June-October 2018. Chemical treatments attempted to manage cyanobacteria population of the Lake have not been successful and created unintended water quality issues. Therefore, we propose to physically remove cyanobacteria from the Lake and utilize the resultant materials as biofertilizer for sustainable tomato and okra production.

Cyanobacteria biofertilizer provide macro and micro-nutrients in the soil and improve nutritional properties (antioxidant, anthocyanin, carotenoids, and chlorophyll contents) of vegetables. Cyanobacteria also increase soil organic carbon (SOC) accumulation, improve soil structure and biodiversity, enhance soil enzymatic activities, and protect plants from pathogens. We estimated that cyanobacteria biofertilizer would reduce 40% fertilizer cost than synthetic fertilizers for vegetable productions. Nutrients in cyanobacteria biofertilizer are in organic form and will be released slowly as the materials decompose. This process is similar to commercially available slow-release fertilizers, that have become standard practice in reducing nutrient leaching and water quality protection. Therefore, this approach is expected to increase return on investment (ROI) of agricultural growers while preserving ecosystem quality. To our knowledge, no field-based experiment in the US have evaluated the effects of cyanobacteria biofertilizer on crop productions and soil health.

This three-year study will evaluate (1) the efficacy of cyanobacteria biofertilizer in improving crop productivity and soil health, (2) the soil and water quality parameters from agricultural fields treated with cyanobacteria, and (3) the economic efficiency of replacing synthetic fertilizer with cyanobacteria biofertilizer.  

We are collaborating with the US Army Corps of Engineers (USACE) and AECOM company to collect cyanobacteria from Lake Okeechobee using micro-flotation techniques. Collected cyanobacterial mats will be processed and packed as biofertilizer at Florida International University. We will test the efficacy of cyanobacteria biofertilizer for vegetable production both at FIU greenhouse and cooperative farmer's field. Yield parameters and compositional analyses of tomato and okra will be determined. Physicochemical and microbiological parameters of soils will be measured to evaluate the effect of biofertilizer in agricultural fields.

Cyanobacteria samples (n = 8) collected from Lake Okeechobee showed 1.79% N (similar to other organic fertilizers), 19.6% total C, high Fe (2005 ppm), other micro-nutrient contents, and lower C:N ratios. A preliminary pot experiment growing okra using cyanobacteria biofertilizer showed (at 7-weeks) similar plant height (69 cm), stem diameter (11.2 mm), and leaf chlorophyll content (51) compared to synthetic fertilizer and higher values than unfertilized control pots.

We expect major outcomes of this project will be restoration of Lake Okeechobee ecosystem, higher crop production and revenue generation, soil health improvement, and educational benefits to growers and students on organic agriculture.

 

Project report summary - Annual report (04/01/2022)

This year (2021 to 2022) annual report includes:

  1. updated project objectives and specific goals
  2. a detail description of how cyanobacteria materials were collected from lake Jesup, Central Florida with the help of algae harvester made by AMECOM company
  3. processing of the materials at Florida International University (FIU) for biofertilizer preparation,
  4. set up research experiments at FIU greenhouse,
  5. results - some data collected as crops are still growing,
  6. issues during crop production, and
  7. brief extension activities.   
Project Objectives:

Overall, this project will follow a system-based research approach where optimum crop yield can be obtained without disrupting the harmony of multiple sustainability components (environmental, ecological, economic, and social). This project will address the key question: “Can we increase overall agricultural sustainability by using the waste product of one system as a viable resource for another?” Specific objectives of this project are:

 

1) To evaluate the chemical and nutritional properties of cyanobacteria collected from Lake Okeechobee and the St. Lucie River ecological areas of Florida

We propose to quantify pH, organic matter content, essential macro (N, P, and K), secondary nutrient (Ca, Mg, and S), and micro (Fe, Zn, Mn, B, Cu, Mo, Cl, and Ni) plant nutrients available in cyanobacteria samples. Cyanobacterial mats will be collected from various locations of the Lake and surrounding area for chemical analyses. We hypothesize that analyses of nutritional and physicochemical properties of cyanobacteria will assist in developing efficient fertilizer application planning in the greenhouse and field trials.

To ensure the safe use of cyanobacteria biofertilizer in the field, microcystin (a common cyanotoxin) concentrations of collected cyanobacterial mats will be analyzed in the laboratory at different stages of biofertilizer development. This experiment will be based on previous study conducted by Co-PI, Dr. Jayachandran and colleagues (Ramani et al., 2012) who found that more than 75% of the cyanotoxins were degraded within 3-weeks after cyanobacteria harvesting from Lake Okeechobee. Our team estimated that it will take about 45 to 50 days to process the fresh cyanobacteria mats into a dried biofertilizer powder (collection, transportation, drying, grounding, and packing). Therefore, the rate of cyanotoxin degradation over time will be calculated from day of collection (time zero; T0) to 50 days after collection (prior to crop application; T50). We will measure the cyanotoxins in cyanobacteria samples every 10 days to develop the ‘decay rate curve’ of the toxin in biofertilizer. We hypothesize that microcystin concentrations in the cyanobacteria biofertilizer will be degraded to less than 1 ppb (WHO standard guideline) and safe for field application.  

Additionally, we will test the shelf-life of the cyanobacteria biofertilizer. Some cyanobacteria biofertilizer samples produced in first year (2021-2022) will be kept in a cool, dry place and the nutritional properties of those samples will be analyzed in project years 2 and 3 to determine the shelf-life of the fertilizer.

 

2) To assess the efficacy of biofertilizer application for crop production and improving soil health

In first section of this objective, we will quantify yield and physiological parameters of tomato and okra at different plant growth stages both in a controlled environment (FIU greenhouse) and in field trials (collaborative farmer’s field). We hypothesize that the application of cyanobacteria biofertilizer will improve soil fertility and increase crop production. The second section of this objective will test the effect of cyanobacteria biofertilizer in improving soil health which is specifically important in South Florida where organic matter content is less than 1%. We hypothesize that extracellular polysaccharides released by cyanobacteria in the soil will improve overall soil health components (soil aggregate stability, soil bulk density, and water holding capacity). Measuring different fractions of soil carbon (total C, organic C, particulate C) will provide us the idea of different ecosystem services cyanobacteria biofertilizer can offer.

 

3) To develop the ‘mineralization rate curve’ of nutrients released from cyanobacteria biofertilizer

Nutrients in cyanobacteria biofertilizer are in organic form and will be released slowly as the organic materials decompose. This process is similar to commercially available slow-release fertilizers, the use of which have become standard practice in reducing nutrient leaching and water quality protection. Therefore, we will develop the ‘mineralization rate curve’ of nutrients in the soil after cyanobacteria biofertilizer application. We hypothesize that developing ‘mineralization rate curves’ of nutrients will help us evaluate the amount of nutrients solubilized each year from this biofertilizer application. Additionally, we will get a better idea about long-term residual effect of nutrients (how much remained in the field) and annual soil organic C accumulation from cyanobacteria application in agroclimatic conditions of South Florida.

 

 4) To monitor nutrient loss (soil and water quality) from the soils

This objective is to quantify the loss of nutrients (inorganic N and soluble reactive P) from cyanobacteria biofertilizer applied plots. As suggested by one of the reviewers that installing lysimeters in the field to conduct leaching experiments is beyond the scope of this study; therefore, the leaching experiments will be conducted in the FIU laboratory as column studies. Column study in leaching experiments for biofertilizer treatments were reported in previous studies (Zhao et al., 2009; Rashmi et al., 2020). Details of the column experiment is discussed in ‘approach and methods’ section. We hypothesize that improved soil structure and slow nutrient release in biofertilizer applied field plots will have lower amount of nutrient loss compared to the plots receiving synthetic fertilizers. 

 

5) To assess the economic benefits of biofertilizer application for vegetable productions in Florida        

We will run stochastic economic models to integrate inputs (seeds, fertilizer, labor etc.) and outputs (crop yield, price, market demand etc.) for assessing the economic benefits of cyanobacteria biofertilizer in tomato and okra productions. We realize that there are uncertainties with crop yields, costs, and market prices under both conventional and proposed organic farming approaches. The stochastic economic model will utilize a Monte Carlo approach to account for the above uncertainties and help assess incremental (additional) net profits and variability in returns. We hypothesize that organic farming will generate higher profits as well as lower economic variability (risks) for growers than conventional farming practices.

 

Update - Project Objectives for year 2022 (greenhouse experiment)

1) To evaluate the chemical and nutritional properties of cyanobacteria collected from lake Jesup

- Macro and micronutrient contents of dried and ground cyanobacteria biofertilizer

- Other chemical properties such as pH, OM content of the biofertilizer

2) To assess the efficacy of biofertilizer application for crop production in the greenhouse setting

- Evaluate the beneficial effects of cyanobacteria biofertilizer on different plants

  parameters (plant height, stem diameter, leaf chlorophyll contents, and crop biomass at

  harvesting) for both okra and tomato.

- Nutritional properties of leaf samples of okra and tomato at different plant growth

  stages

3) To evaluate the microcystin contents in cyanobacteria materials

- Microcystin concentration of cyanobacteria samples immediately after collection and at

  every 12 days interval till 60 days after collection  

Research

Materials and methods:

Figure 3 Figure 4 Figure 5

Support letter from AECOM

Approaches and Methods:

The USACE has an ongoing project (Harmful Algal Bloom Interception, Treatment, and Transformation Systems; HABITATS) in collaboration with AECOM (an International company; website: https://aecom.com/) to remove cyanobacterial blooms from Lake Okeechobee and adjacent canals. The USACE and AECOM agreed to assist in collecting cyanobacteria from the Lake and a support letter from Mr. Dan Levy (Vice president, AECOM) is attached along with this proposal. Cyanobacterial scums will be collected using ‘float it up and skim it off’(micro-flotation) technique. An overview of cyanobacteria collection, processing, storage, field application, and extension outreach is presented in Figure 3. Detailed description of collection and processing are discussed below:

 

1.Cyanobacteria collection (AECOM) and processing: The AECOM will deploy a custom-built, mobile algae harvesting unit (Figure 4) that separates algae biomass from water. Even a small-scale harvester can process between 100-175 gallons per minute and produces up to 1,000 gallons of algae biomass in 8 hours. Skimmers will capture thick mats of algae floating on the water’s surface. Collected algae biomass slurry will be dewatered to produce semi-solid mats. Composite cyanobacterial mats will be collected from various locations of the Lake and adjacent river area. Collected cyanobacteria mats will be transported to FIU greenhouse for drying and processing. Dried and ground (in powder form) cyanobacteria biofertilizer will be stored in colored plastic bags for field application. Storage of the dried cyanobacteria biofertilizer in colored plastic bags can increase its shelf life (Jha and Prasad, 2006). In this phase we will invite our agroecology students to experience cyanobacteria collection and processing for biofertilizer preparation.

 

A preliminary study was conducted by our research team where we (a) collected cyanobacteria samples (mats; n = 8) from Lake Okeechobee, (b) dried those samples at 450C for 72 hours, and (c) ground those dried samples using mortar and pestle (Figure 5A). Commercial grinding machine (Model# BL-230) will be used to process the biofertilizer required for greenhouse study and field trials.

We found the dry matter content of the cyanobacterial mat was 17 to 19%.   

UPDATE - Annual report (2022)

Cyanobacteria collection and processing

Figure 2022-1 Figure 2022-2 Figure 2022-3

As we mentioned in our statement of work (SOW), we communicated several times with the AECOM company, who helped us collecting the cyanobacteria from Lake Jesup (28043’N, 81013’W) located in Central Florida. We also visited the floating cyanobacteria collection station of AECOM company (Figure 2022-1) and learned more about the working principles of the unit. The custom-built, mobile algae harvesting unit of AECOM was separating algae biomass from the lake Jesup water. Generally, a small-scale harvester can process between 100-175 gallons per minute and produces up to 1,000 gallons of algae biomass in 8 hours. Cyanobacterial slurries were collected using ‘float it up and skim it off’(micro-flotation) technique and later coagulated in the unit. Cyanobacterial slurry was collected in five-gallon buckets (Figure 2022-2) for easy transporting and brought back to the FIU campus for further processing. One small glass jar of sample was brought in ice cooler and stored at -200C at FIU laboratory for microcystin analysis. This sample will give us the microcystin concentration of the cyanobacteria sample immediately after harvesting from the lake. At FIU campus, the rest of the slurry was then air dried (after spreading on large plastic trays) in the greenhouse for 3 to 4 weeks (Figure 2022-3) to obtain dried cyanobacteria materials. Slurry on the plastic trays were stirred up occasionally to expedite the drying process. Dried cyanobacteria then powdered using a mechanical grinder and stored in colored plastic bags for future applications.

 

2. Timeline development for safe use of the biofertilizer:

This project also aims to develop the timeline for safe use of the cyanobacteria biofertilizer produced in the field for crop production. As commented by one of the reviewers about the safety concern of the biofertilizer, we will measure the microcystin, a common cyanotoxin found in cyanobacteria during first project year.  

Microcystin content of cyanobacteria will be measured following the USEPA method 546 (ADDA- Enzyme-Linked Immunosorbent Assay technique) from collection time (T0) till 50 days after collection (T50) at 10 days interval to develop a decay rate curve (Birbeck et al., 2019). Krishnaswamy Jayachandran (Co-PI of this project) and colleagues conducted an experiment on cyanobacteria samples collected from Lake Okeechobee to measure microcystin degradation over time (Ramani et al., 2012). They found that more than 75% of the cyanobacteria microcystin degraded within 3-weeks after harvesting from the Lake. Cyanobacteria powder do not cause airborne problems or human health issues. However, we will take proper pre-cautions (gloves, masks) during biofertilizer processing and field application.

We will conduct safety training and knowledge dissemination workshops for growers and the farm workers for safe and better handling of the biofertilizer.

UPDATE - Annual report (2022)

This year we collected cyanobacteria slurry samples every 12 days interval till 60 days after collection from the lake. The first sample collected immediately after cyanobacteria harvesting from the lake is noted as T0. All the slurry samples (T0, T1, T2, T3, T4, and T5) for microcystin concentration were collected and stored in -20 deg C for analysis.

 

3.Objective # 1: Nutrient and chemical analyses of cyanobacteria biofertilizer

Subsamples of cyanobacteria biofertilizer will be analyzed at the FIU Soil-Plant Microbiology Laboratory for nutrient (macro and micro) contents and other chemical properties including OM content, pH, and cation exchange capacity (CEC). The pH of the biofertilizer will be measured in 2:1 (water: biofertilizer) slurry using ‘Denver Instrument’ pH meter. Cation exchange capacity (CEC) of the biofertilizer will be measured by sodium acetate extraction following method USEPA 9081. A subsample of cyanobacteria will be combusted in 6000C for 6 hours in Fisher Scientific isotemp muffle furnace to obtain organic matter content of the biofertilizer. Total C and N contents will be analyzed in a Thermo-Scientific Flash EA (Dumas combustion method). For multi-element analysis (macro and micro elements), pre-weighed cyanobacteria samples will be combusted at 550°C and then extracted using concentrated hydrochloric acid (HCl). The extracted samples will be analyzed in ICP-OES (Perkin Elmer Optima 8300). Total P will be extracted by combusting samples in the presence of MgSO4 at 550°C and solubilizing the ash with 0.6 N HCl to convert all P to ortho-P forms followed by colorimetric analysis (antimony-phospho-molybdate complex) in TechniconTM AutoanalyzerTM Sampler IV.  

Agroecology graduate students and student interns will be involved to gain experimental and experiential learning about chemical analyses of cyanobacteria biofertilizer in the laboratory.

 

Preliminary results (fertilizer grade): Physicochemical parameters of processed cyanobacteria biofertilizer samples were analyzed at the extension soil testing laboratory of the University of Florida (Table 1).

Table 1. Nutritional properties of cyanobacteria biofertilizer used for pilot experiment. Values are presented as mean ± SD of n = 8 samples.

Parameters

Unit

Value

Total C (TC)

%

19.56 ± 0.71

Total N (TN)

%

1.79 ± 0.07

Total P (TP)

%

0.02 ± 0.00

Total S (TS)

%

0.13 ± 0.01

Potassium (K)

%

0.06 ± 0.01

Calcium (Ca)

%

22.12 ± 0.20

Magnesium (Mg)

%

0.12 ± 0.01

Iron (Fe)

ppm

2005 ± 160

Manganese (Mn)

ppm

132.56 ± 13.35

Zinc (Zn)

ppm

53.34 ± 0.69

Copper (Cu)

ppm

29.51 ± 4.80

Boron (B)

ppm

95.37 ± 4.21

Molybdenum (Mo)

ppm

1.82 ± 1.01

Nickel (Ni)

ppm

4.34 ± 1.78

 

The C:N ratio (mol:mol) of test cyanobacteria biofertilizer averaged 12.7:1, lower than other organic nutrient sources. Therefore, we expect increased N mineralization in soil when cyanobacteria biofertilizer is applied. Another important component was Fe content (average 2005 ppm) in our biofertilizer mix. South Florida soils generally lack Fe because of low SOM content, high temperature, and high soil pH (Ozores-Hampton, 2013). Vegetable farmers often face problems due to Fe deficiency in these soils (Liu et al., 2012). Our collaborative growers informed us of the need to apply chelated Fe (an expensive option) to meet the crop requirements. Thus, cyanobacteria biofertilizer would serve as an excellent source of Fe and other micronutrients. Visual observation from preliminary experiments indicate that the interveinal chlorosis of young okra leaves in control plots possibly resulted from Fe deficiency (Figure 5B). Leaf samples were collected and submitted for micronutrient analyses (analysis in progress at the time of this proposal submission).

Typical N% for commonly used organic fertilizers in South Florida such as poultry manure is 1.5 to 2.3% (Hochmuth et al., 2009). Our test cyanobacterial biofertilizer falls within this range. However, cyanobacteria biofertilizer provides micronutrients and offer ecosystem services which makes it a high potential biofertilizer for vegetable production.  

 

4.Greenhouse experiment at FIU:

Greenhouse experiment will be conducted during first project year (2021-2022) to evaluate the effect of cyanobacteria biofertilizer on crop growth and soil properties (pH, EC, total P, total N, total C content, OM content, aggregate stability, and nutrient loss). Representative South Florida soils will be collected from LNB groves farm, Homestead, FL (courtesy of Mr. Marc Ellenby, our collaborative grower) for this greenhouse study. Seven-gallon and five-gallon pots will be used to grow tomato (Lycopersicum spp variety: BHN 975) and okra (Abelmoschus esculentus variety: Cajun Delight), respectively. These tomato and okra varieties perform well in Florida’s climate and are highly resistant to insect pests and were selected after discussing with Dr. Qingren Wang (extension specialist for commercial vegetables; our team member) and Mr. Marc Ellenby. Fertilizer application rates will be 224 kg N ha-1, 112 kg P2O5 ha-1, and 168 kg K2O ha-1 for tomato, and 135 kg N ha-1, 112 kg P2O5 ha-1, and 112 kg K2O ha-1 for okra. Biofertilizers will be applied along with synthetic fertilizers in different combinations. Seven treatments combinations [T0: control (no fertilizer applied); T1: 100% biofertilizer; T2: 75% biofertilizer + 25% synthetic; T3: 50% biofertilizer + 50% synthetic; T4: 25% biofertilizer + 75 % synthetic; T5: only 50% synthetic fertilizer and no biofertilizer; and T6: 100% synthetic fertilizer] will be assigned in a randomized complete block design (RCBD) with 20 replications (5 main plot x 4 within plot = 20) per treatment.

Because of the highly reactive nature of N fertilizers and the porous sandy soils of Florida, the N fertilizer will be applied in two splits. Urea (H2N-CO-NH2; 46-0-0) will be used as the primary source of inorganic N. Triple super phosphate [Ca(H2PO4)2. H2O; 0-45-0] and muriate of potash (KCl; 0-0-62) will be used as inorganic source of P and K, respectively. We will conduct demonstration training on shade house experiments to the local farmers and students during organic farming workshops and field days.

 

Preliminary results (pot experiment): We arranged the pot experiment in RCBD at FIU shade house using 2-gallon pots for each plant. Four treatments (with six replications for each treatment) used in this study including control (no fertilizer), full synthetic N fertilizer (urea; 46-0-0), full cyanobacteria biofertilizer, and 50% synthetic + 50% cyanobacteria biofertilizer. Soils were collected from LNB Grove Farm, Homestead, FL (website: https://www.lnbgrovestand.com/).

 

Table 2. Effect of cyanobacteria biofertilizer on plant height, stem diameter, and leaf chlorophyll content (SPAD readings) of okra. Like letters (lowercase) indicate not significant at α = 0.05 level (SAS 9.4 was used)

Treatment

Week 4

Week 7

Plant height

(cm)

Stem diameter

(mm)

SPAD*

Plant height

(cm)

Stem diameter

(mm)

SPAD

n =

6

6 x 3 =18

6 x 6 = 36

6

6 x 3 =18

6 x 6 = 36

Control

34 ± 6 a

6.1 ± 0.6 b

31 ± 6 a

48 ± 6 b

7.4 ± 0.7 b

34 ± 5 b

Full synthetic

47 ± 9 a

8.4 ± 0.9 a

43 ± 8 a

67 ± 9 a

10.8 ± 0.9 a

49 ± 8 a

50-50 Fertilizer

44 ± 6 a

8.7 ± 0.9 a

42 ± 6 a

64 ± 9 a

10.5 ± 1.1 a

49 ± 7 a

Full cyanobacteria

45 ± 5 a

9.2 ± 0.8 a

45 ± 7 a

69 ± 12 a

11.2 ± 1.1 a

51 ± 8 a

* SPAD: Soil Plant Analytical Development represents chlorophyll contents of the leaves (no unit)

Overall, plant height (69 cm) and SPAD (51) readings at week-seven were significantly (P<0.05) higher in full biofertilizer treatment than unfertilized control. Stem diameter (9.2 and 11.2 mm at week 4 and week 7, respectively) was always significantly higher in fertilizer treatments than control (Table 2). However, no significant differences were found between fertilizers, indicating that cyanobacteria biofertilizer can achieve same crop physiological parameters as synthetic fertilizers.

UPDATE - Annual Report (2022)

Experimental set up in the greenhouse

Okra (Abelmoschus esculentus Var: Cajun Delight) and tomato (Lycopersicum spp Var: BHN 975) crops are growing at three- and five-gallon plastic pots, respectively, at FIU greenhouse. Plastic pots were filled up with soils collected from Homestead area of South Florida. Physicochemical properties of the background soil are presented in Table 2022-1. Okra and tomato pots were organized in separate bench tops with each pot was spaced about 3 ft from each other. Walking rows are available between each bench tops for watering the plants, regular maintenance, and recording the experimental data. Biofertilizer was applied along with synthetic fertilizers in different combinations. Seven treatment combinations were a) T0: control (no fertilizer applied); b) T1: 100% biofertilizer; c) T2: 75% biofertilizer + 25% synthetic; d) T3: 50% biofertilizer + 50% synthetic; e) T4: 25% biofertilizer + 75 % synthetic; f) T5: only 50% synthetic fertilizer and no biofertilizer; and g) T6: 100% synthetic fertilizer. The experimental set up followed a randomized complete block design (RCBD) with five replications for each treatment. At this point fruit setting just started for both okra and tomato. Okra seeds were planted in January 2022. We grew tomato seedlings in nursery trays from December, 2021 to January, 2022. One month tomato seedlings were then transplanted in the pots in 3rd week of January, 2022. Both okra and tomato plants were watered in regular intervals in the greenhouse.  

Table 2022-1. Physicochemical properties of the soils collected from the field and used to fill up the pots in the greenhouse

Parameters

Unit

Value

pH (1:1 soil slurry)

 

7.06 ± 0.33

TN

%

0.74 + 0.08

TC

%

14.39 ± 0.98

Ca

%

1.51 ± 0.026

Cu

%

0.0003 ± 0.00001

Mg

%

0.033 ± 0.0008

P

%

0.002 ± 0.00006

K

%

0.014 ± 0.0004

Na

%

0.014 ± 0.0012

S

%

0.022 ± 0.00009

Zn

%

0.001 ± 0.00009

 

5.Objective # 2: Efficacy of cyanobacteria biofertilizers (crop yield and soil health) in field settings

Agricultural field plots from our collaborative growers (Mr. John Mills and Mr. Marc Ellenby) will be used to test the effect of cyanobacteria biofertilizer on tomato and okra production and soil health analysis. Physicochemical analysis of background soil will be analyzed before conducting the experiment. Yield parameters (shoot weight, root weight, plant height, and fruit yield) and compositional analyses (leaf chlorophyll content, plant height, plant shoot diameter, and nutrient uptake) of tomato and okra will be recorded during plant growth stages (early transplanting/seeding stage, flowering, early fruit set/pod-filling) and at harvesting. The yield parameters for our study were selected after discussing with Mr. Robert Barnum and Mr. Marc Ellenby. Routine physicochemical parameters (pH, EC, TP, TN, TC, POM, biomass C, N and P, Mehlich-3 extractable plant essential nutrients, ammonium, nitrate, nitrite, OM%, water filled pore space WFPS%, and aggregate stability) of soil samples will be analyzed in regular intervals throughout crop production. Soil pH will be measured in 1:1 soil to water slurry. Soil EC would be measured using Oakton conductivity meter. Analysis of TC, TN, and TP content will follow methods discussed earlier for soil samples. Particulate organic matter (POM) is a powerful indicator of soil health and quality (Chan, 2001; Wilson et al. 2001). Particulate organic matter (POM) will be measured in dry soil using a method described by Nciizah and Wakindiki, 2012 and will be calculated using the formula:

 

POM g/g dry soil= [(Dry soil weight - Combusted soil weight)/(Dry soil weight)] x 100

 

Soil aggregate stability will be measured using dry and wet sieving techniques (Kemper and Rosenau, 1986). Water soluble aggregates (WSA) is generally determined using:

 

WSA = {(Wac - Wc) / Wf}*100

 

Where, WSA (%) is the water-soluble aggregate in particular class or range of aggregate class, Wac (gm) id the dry weight of aggregates and coarse material; Wc is the dry weight of coarse material and Wf is the dry weight of the soil under each fraction of aggregate class.

 

Paid student (assistantships/scholarships) will assist in sample analysis where they will gain knowledge about sustainable agricultural research projects.

 

6.Objective # 3: Develop the ‘mineralization rate curve’ of nutrients

Mineralization rate of organic fertilizers is an important component as it determines the nutrient availability for crop production. Soil mineralization rate of N is commonly measured using buried bag technique (Lentz and Gary, 2012; Hanselman et al., 2004). Composite soil samples from fertilizer treated plots (and control) will be used to fill the bags. Total 5 bags will be prepared for each treatment and for each plot. The bags will be buried at 15 cm soil depth at the beginning of the experiment. One bag from each plot will be retrieved at different crop growth stages (1. early transplanting/seeding stage, 2. flowering, 3. early fruit set, and 4. maturity) for inorganic N content analysis. The last bag in each plot will be left to obtain the mineralization rate of N over two-years of the field study. N mineralization for a treatment will be calculated as:

 

%N mineralized = (N mineralized from treatments - N mineralized from control)/(Total N applied for treatments)

Treatments = T1 to T6 for this study

 

 7.Objective # 4: Monitoring nutrient loss from the soil

We will conduct a column study in the laboratory for leaching experiment using field soils following the method described by Zhao et al., (2009) and Rashmi et al. (2020). Composite soil samples (mixture of 12 cores) will be collected from each treatment and each plot to conduct the column study. The treatment for the column study will be the same as the field level fertilizer treatments (T0: control; T1: 100% biofertilizer; T2: 75% biofertilizer + 25% synthetic; T3: 50% biofertilizer + 50% synthetic; T4: 25% biofertilizer + 75 % synthetic; T5: only 50% synthetic fertilizer and no biofertilizer; and T6: 100% synthetic fertilizer). Each treatment will have 8 lab replications. Total dissolved P, soluble reactive P, inorganic N content will be measured in leachate samples. At the end of the column study; 100 days after the start date, the soil column will be sectioned to analyze TP, Mehlich-3 extractable P, and inorganic N content. The experiment was specifically selected for 100-day period because okra and tomato are generally harvested in 60-65 days and 80-85 days, respectively. Lysimeter is often installed in the field to conduct leaching experiments, however, one of the reviewers pointed out that using lysimeter for leachate analysis is beyond the scope of this study. Therefore, we will have column studies for leaching experiment in the laboratory (controlled environment).

 

8.Objective # 5: Economic assessment of biofertilizer application

We assume that if farmers were to adopt the new biofertilizer, they would experience the following types of changes and/or opportunity costs: (a) changes (increase or decrease) in the annual costs of cultivation; (b) changes in crop yields (increase or decrease) and products sold; and (c) change in product market price (due to product quality changes). Therefore, the “true” or “incremental” benefits of integrating the proposed biofertilizer will be equal to the difference in the net profits between with and without implementation. The following methodology captures the economic effects of all the above three factors on the incremental benefits of applying new fertilizer. Assume that the per acre cost of crop production is c ($/ac). Per acre crop yield is y (lb/ac) and market price is p ($/lb). Let π ($/ac) denote the net profit. Subscripts pre and pos denote before and after implementation.

 

The expected net profit (πpre) before the implementation of the new fertilizer is,

 

πpre = (ypre · ppre – cpre)

 

Similarly, the expected net profit after the implementation of a given N-I combination (πpro) is,

 

πpos = (ypos · ppos – cpos)

 

Note that both quantity and quality of the product could vary from the current to the new practice. The annual per acre incremental benefits of biofertilizer application (IB) is the difference in the net profit between without implementation and with implementation. Formally,

 

            IB = πpos - πpre

 

Crop yield, costs, and price parameters used in the above economic model are stochastic in nature and may have certain underlying probability distributions. Based on the field and lab experimental trials (sample mean and standard deviation), we will construct appropriate probability distributions (e.g., normal distribution) for yield parameters. Historical market data (USDA) on prices and costs will be used to develop appropriate uniform or other probability distribution reflecting market uncertainty. Then, the incremental benefits (IB) will be estimated by running Monte Carlo simulations, which accounts for uncertainty in model parameter values (yields, costs, etc.) by expressing them as probability distributions rather than fixed values.

We will first need to gather the data for the economic enterprise budgets for existing tomato and okra crops in Florida. Using these as starting points, we will then develop mixed-crop budgets for alternative systems that involve tomatoes and okra production and seasons.  The information on the output side of the budget (e.g., market price, season, product quality, value addition, etc.) will directly come from secondary sources such as USDA. Monte Carlo stochastic analysis will be conducted around each mixed-crop budget in order to capture the effects of multiple scenarios reflecting alternative market channels, degree of value addition, output and input price variances. The final recommendation will be based on alternative production scenarios of biofertilizers, crops, seasons, and others.

Research results and discussion:

RESULTS AND DISCUSSION

Figure 2022-4 Figure 2022-5 Figure 2022-6 Figure 2022-7

This year we are conducting the experiment in FIU greenhouse. Both okra and tomato plants are still growing and are in their fruiting stages at this point (Figure 2022-4, 2022-5). Basic physical properties during crop production such as plant height, stem diameter, leaf chlorophyll contents were collected in regular intervals. Plant height, stem diameter, and SPAD readings for tomato plants during flowering and fruit settings are presented in Table 2022-2. We will collect two more sets of plant physiological data before harvesting. Soil samples from each treatment (during fruit settings) were collected and processed (dried and ground) for future analysis. Additional soil sample from each treatment will be collected during harvesting. Those data will be added to the progress report once the analyses are done.

Our results (as the data collected so far) from tomato experiment clearly indicate that 100% cyanobacteria biofertilizer (T1) applied plots performed better than control plots. Average plant height (92.71 cm), stem diameter (8.47 mm), and SPAD readings (47.09) of T1 was 22%, 15%, and 20% higher, respectively, than control plots. When we compared the plant parameters of T1 with 100% synthetic fertilizer applied plots (T6), no significant differences were found. Therefore, it can be noted that 100% cyanobacteria fertilizer application is performing similar to 100% synthetic fertilizer application till this point of data collection. However, better comparative analysis can be made once we will have more plant parameters and yield data available after harvesting.  

Table 2022-2. Physiological parameters of tomato plants during different growth stages

Treatments

Plant height (cm)

Stem diameter (mm)

SPAD reading

Flowering Stage

T0

73.66

6.95

37.78

T1

83.82

7.95

43.16

T2

79.59

7.66

44.47

T3

80.43

7.38

42.75

T4

82.97

7.77

44.36

T5

75.35

6.87

40.14

T6

86.36

7.52

43.04

Fruit Setting

T0

78.55

7.79

40.55

T1

101.60

8.99

51.01

T2

96.52

8.42

49.03

T3

94.05

8.14

46.70

T4

99.91

8.59

46.20

T5

82.55

7.94

42.84

T6

107.02

8.38

48.99

Common issues during production

A major problem occurred during the greenhouse experiment was unexpected cooler temperature in South Florida during January, 2022. Growth of both okra and tomato plants were little slower because of the lower temperature. In general, both okra and tomato can be harvested at 65 to 70 days and 90 to 100 days, respectively. However, harvesting of our okra and tomato plants will take little longer than their usual harvesting schedule. Control plots for both okra and tomato plants showed possible Fe deficiency (interveinal chlorosis; Figure 2022-6). We also found some fungal disease (possibly Septoria leaf spots) in our tomato plants (Figure 2022-7). We applied copper fungicide to control fungal spots in tomato plants. Visually at least 60 to 70% of the leaf spots were controlled by using copper fungicide. We collected and processed (dried and ground) plants leaf samples during different growth stages (flowering and fruit settings), however, the laboratory analysis of nutritional properties of those samples is scheduled to be done.   

Project timeline for 2021 to 2022

 

Activities

Time frame

Job status

Additional comments

1. Project notification

February, 2021

Done

 

2. Project account set up

November, 2021

Done

 

3. Cyanobacteria collection and processing

 

 

About 2 to 3% dry matter was present in the slurry

       Trip to lake Jesup for cyano collection

November, 2021

Done

 

       Taking raw samples for microcystin

       analysis

November, 2021 to January, 2022

Done

Samples were preserved at -200C for further analysis

       Drying cyano samples in plastic trays

November, 2021 to January, 2022

Done

 

       Grind samples to make biofertilizer

January, 2022

Done

 

4. Setting up tomato and okra experiment

 

 

At FIU greenhouse

       Soil samples collected and pots were

       prepared for greenhouse study

January, 2022

Done

 

       Fertilizer application in the pots

January, 2022

Done

 

5. Data collection

 

 

 

      Plant height, stem diameter, and SPAD

February to April, 2022

Continued

More samples will be collected

      Soil samples collection during

      different growth stages

February to April, 2022

Continued

Few sets were collected, more will be collected

      Plant tissue sampling

      - at flowering stage

      - at fruit settings

      - at harvesting

 

February, 2022

March, 2022

April, 2022

 

Done

Done

Scheduled

 

5. Microcystin analysis

April, 2022

Scheduled

 

6. Harvesting

April, 2022

Scheduled

 

7. Laboratory analysis

From April, 2022

Scheduled

 

8. Project update at the SARE portal

May, 2022

Scheduled

 

 

Project outcomes (2021-2022)

Webinar presentation:

Sanku Dattamudi, Saoli Chanda, Krishnaswamy Jayachandran, Leonard J Scinto, and Mahadev Bhat. Application of cyanobacteria as biofertilizer to increase vegetable production and soil health improvement in South Florida. July 27, 2021, Webinar, National Algae Association (NAA)

Manuscript:

Sanku Dattamudi, Saoli Chanda, Krishnaswamy Jayachandran, Leonard J. Scinto, and Mahadev Bhat. 2022. Use of cyanobacteria biofertilizer for okra production and improving agricultural sustainability in Florida, USA. Target journal: Agronomy. (In preparation)

Extension activities:

1.  Undergraduate and graduate students of FIU Agroecology have been involved in this project. We demonstrated them biofertilizer preparation and its use for vegetable production. 

2. Member of AECOM company visited our FIU greenhouse and the pilot study at FIU garden - where we shoed them the current experiment and the applications of cyanobacteria biofertilizer

3. Pre-collegiate student visited our experiments to learn more about soil and plant sciences and agroecology.

4. We had couple of field visits to our collaborative growers where we described them about the project progress and scheduled timeline for next year field trial.

 

 

 

Participation Summary

Educational & Outreach Activities

1 Journal articles
1 Webinars / talks / presentations
1 Other educational activities: Experimental and experiential learning activities for pre-collegiate student during the experiment. The student visited the greenhouse settings several times and learned about soil, plant sample collection, effect of biofertilizers and laboratory analyses.

Participation Summary:

Education/outreach description:

Support_Letter_DiMare Support letter_Dr. Wang Support letter_Dr. Li

Outreach and dissemination 

Florida International University (FIU) Agroecology Program sponsors and participates in several outreach programs and events to disseminate newly acquired knowledge about sustainable agricultural practices. This year we have Dr. Qingren Wang, an extension specialist for commercial vegetables. He will be our team leader for extension and outreach activities for this project.

Outreach program

When

Audience

Activities

GrowFest

October every year

Farmers, Growers, Stakeholders, General public

Workshops on biofertilizers

South Florida Regional Science and Engineering Fair

January every year

Middle and high school students

Conduct roundtable discussion on biofertilizers and sustainable agriculture

Compost workshop

Fall and Spring per year

Agroecology Program students and other students, faculty, beginning farmers

Discussion on organic agriculture, biofertilizer, sustainable agriculture

Agroecology Annual Symposium

March every year

Undergraduate and graduate students from FIU, St. Thomas University, Miami Dade College, Faculty

Keynote address on agriculture sustainability, ecological, economic sustainability through cyanobacterial biofertilizers

CASE Take Our Kid to Work Day

April every year

Elementary, middle, high school students

Laboratory and shade house tour of biofertilizer experiments

Miami International Cattle and Agriculture Show

April every year

General public, farmers

Workshop on biofertilizers

Farm Life Field Day

November in every year

Agroecology Program students

Tour of collaborating farmers field to visit biofertilizer tomato and okra field experiments

Organic Gardens in Sustainable Food Systems Workshop

Fall and Spring in each year

Students and Farmers

Demonstrate biofertilizer production, nutrient analysis, sustainable agriculture

Other Outreach/Extension/Dissemination Activities

a) Local news, news articles and websites

Cyanobacteria blooms have been a major concern in Central and South Florida. Several news channels such as CBS local, The Weather Channel, NPR have been regularly reporting news about HABs and their consequences on human health and aquatic lives. Our collaboration with the international company AECOM and US Army Corps of Engineers (USACE) to physically remove the cyanobacteria from the Lake Okeechobee water will be covered by local news channels. Most of the people living around the Lake are concerned about water contamination from microcystin, foul smell of cyanobacteria, and economic decline value of their property. Their urgent need to find a better solution to remove and discard HABs have been featured in several local news channels and newspapers (such as TC Palm). To our knowledge, no field-based research on cyanobacteria biofertilizer has been conducted in the US. An innovative approach of using this nuisance cyanobacteria scum for sustainable farming and better community service is expected to draw attention of local people. We are planning to highlight our work in local new channels to educate public about recycling renewable resources, organic farming, soil health, carbon sequestration, and over all sustainable living. Agroecology Program website and Southern SARE website will be included in the news, so that interested people can direct their questions and concerns to us. Our Agroecology webpage ‘Biofertilizer: A sustainable nutrient source’ will provide information about this project to students, faculty, farmers, and general public. This project provides a unique approach to increase overall environmental sustainability on integrated farming system.

b) Workshops, field days, and training sessions

Florida International University (FIU) Agroecology Program organize several symposia and professional workshops throughout the year.  FIU is in the process of acquiring a 40-acre organic farm (called Possum Trot) through gift, which attracts a large number of farmers, residents and schoolchildren for eco-tour and experiential learning.  In addition, under a grant from the USDA Office of Advocacy and Outreach, we have been conducting bi-weekly technical and managerial workshops and four-month long farm apprenticeship for Beginning Farmers, Veterans and Socially Disadvantaged (BVSD) farmers on this and other private farms in South Florida.  More than 20-25 participants take part in each workshop from Southeast Florida. For the last twelve years, we have been conducting annual Agroecology Symposium under various USDA NIFA grants program.  More than 200 people including farmers and industry people attend the symposium. We also conduct annual International Agroecology Workshop. Under the proposed project, we will invite organic growers each year to participate in a special workshop as part of the annual Agroecology Symposium to discuss project results and gather dissemination ideas, feedbacks on our research activity and identify gaps for effective outreach activities. The workshop will also invite USDA, UF and FIU scientists. One or more technical workshops we conduct for the farmers will focus on biofertilizer and sustainable agriculture. As part of the annual farm tour to be conducted by Miami Dade Farm Bureau and county Extension Office, we will invite farmers and other interest groups to take the tour of the ‘Possum Trot’ farm which houses already more than 350 tropical plant species. We have developed extensive network with Homestead area and other South Florida farmers and nursery owners. An important outreach activity will be our involvement with DiMare company (one of the largest tomato growers in the US). Mr. Tony DiMare (Vice president of DiMare company) has agreed to collaborate with us during sustainable agriculture demonstration through various workshops, field days, and training sessions in his farms. A support letter from Mr. DiMare is attached with this proposal.   

c) Regular Meetings with local growers

We plan to conduct regular meetings with local farmers, growers, nursery owners, and stakeholders to discuss about the beneficial effects of low-cost biofertilizer and the economic benefits of replacing industrial synthetic fertilizer. Our collaborative growers team has decades of experiences in sustainable agricultural farming systems, and their valuable inputs during interactions with local growers will be a key factor to the proposed outreach/extension and dissemination activities. Our NGO partner Colonel John Mills (CEO of Redland Ahead Inc.) is the primary contact to reach out to veteran farmers and other small farmers. Colonel Mills operates a 20-acre community-based farm known as Verde Farm. Based on the successful execution of field pilot studies, biofertilizer application will be part of the farm operations in Verde Farm.  

d) Incorporation in classroom teaching

We have series agriculture sciences courses offered in classroom on regular basis. Some of the courses are agroecology, sustainable agriculture, soils and ecosystems, soil biology, modern crop production. We plan on incorporating cyanobacterial biofertilizer – production, nutrient analysis, beneficial role in soil and crop production. Students gain both experiential and experimental knowledge on biofertilizers.

e) Annual conferences and peer-reviewed articles

We expect to generate valuable data from cyanobacteria collection, drying, nutrient analysis, green house studies, and field studies. We will have several extension articles and peer-reviewed scholarly publications. We also plan to present in annual conferences such as tri-society meetings (ASA-CSSA-SSSA) and regional farmers’ meet and more. Project reports will be submitted to SSARE based on the requirements. We will submit a final project report at the end of the third year.

f) Data Management Plan

The project will leverage the extensive data management strategies and protocols of the FIU Institute of Environment (InE) https://environment.fiu.edu/ and its trained archival staff. The project will assign a qualified data manager within the InE who is certified in disclosure risk management to act as steward for the data while they are being collected, processed, and analyzed. All research data collected as part of this project is owned by the University.

 

- Types of data

 

Both digital and non-digital data will be generated from lab work, field work, surveys, etc.

Agroecosystem data: Types of data to be collected, stored, and shared include: (a) Location of farm, (b) general farm and environmental data including crop species/variety grown, numbers, growth features, productivity, soil type, irrigation and fertilizer application, specific agronomic/horticultural practices.

 

The data will be stored as MS excel spread sheet, MS word, PDF files, and images/pictures as JPEG/TIFF files. Each data set will have specific identifiers and required information to help understand, validate, and use the data.

 

- Data storage and preservation

 

We will process and manage data in a secure environment (e.g., lockable computer systems with passwords, firewall system in place, power surge protection, virus/malicious intruder protection) and by controlling access to digital files with encryption and/or password protection. We will share the results of the study with the research community via journal publication, conferences, and seminars. We will also dedicate a website for this project to share and publish these results (available at an fiu.edu location). Our project team have received training in human subject’s protection and will operate under the IRB approval for the project. Only project team members will have access to the confidential individually identifiable data, and they will aggregate or anonymize all data for publication or for data sharing purposes.

 

- Data sharing and public access

 

Project results and report are accessible to all public including university scientists, students, SSARE scientists and administrators and farm organization and farmers. Additionally, the key results and images and other useful information from the project will be uploaded onto FIU agroecology website section on Biofertilizer.

 

The research team will hold the intellectual property rights for the research data they generate.  The team own the raw data, the processed data, and the products derived from this project and will have the first use of the data. We will permit re-use and re-distribution of the aggregated and anonymized data and creation and publication of derivatives from such data for data sharing purposes as long as those seeking to do so receive prior permission from the research team, and the research team receives attribution or co-authorship for the work in the form of an acknowledgement (for data generation) in any publications or products derived using the data. The research team does not plan to permit others to use the data to develop commercial products or that in any way produce a financial benefit for those requesting the data. Since the data is generated using public funds, we expect that the products derived will be used exclusively for public benefit. We will announce the conditions for re-use and re-distribution of data on the project website within three months of the grant closing.

 

The FIU Agroecology will make the research data from this project available to the broader organic agriculture research community. Public-use data files, in which direct and indirect identifiers have been removed to minimize disclosure risk, may be accessed directly through the repository website. After agreeing to Terms of Use, users with a repository data account and an authorized IP address from a member institution may download the data, and non-members may purchase the files. Restricted-use data files: These files are distributed in those cases when removing potentially identifying information would significantly impair the analytic potential of the data. Users (and their institutions) must apply for these files, create data security plans, and agree to other access controls.

 

- Roles and responsibilities

 

The Principal Investigator and Co-PI’s of this project and the InE Information Manager will take responsibility for the collection, management, and sharing of the research data.

 

- Plans for Archiving and Preservation of Access

 

We will utilize the Repository of InE for data storage, preservation, and dissemination over a period of three years beyond the conclusion of the project.  In addition, we will establish a cloud-based data repository (Dropbox and Google Drive), along with an external hard drive for storing and maintaining the collected data. The principle investigators will hold the intellectual property rights to the data but will grant redistribution rights to other users for research and education purposes.

 

g) Student involvement

Florida International University (FIU) Agroecology programs has 20 graduate, 75 undergraduate students, and 25 student interns. We continue to recruit students every year to our programs. We will involve students from cyanobacteria collection, processing, analysis, green house and field experiments. We encourage and mentor students to develop few hypotheses and design small experiments on biofertilizers. We have extensive experience mentoring students in such activities and proved successful. Students graduate from our program with biofertilizer experiential and experimental learning experience will serve as extension agents to disseminate the outcome of the project.

 

Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture or SARE.