Cost-benefit analysis of inoculating blueberry bushes with ericoid mycorrhizae

2013 Annual Report for FNE12-770

Project Type: Farmer
Funds awarded in 2012: $8,890.00
Projected End Date: 12/31/2015
Region: Northeast
State: Vermont
Project Leader:
Ben Waterman
Watermans Berry Farm

Cost-benefit analysis of inoculating blueberry bushes with ericoid mycorrhizae


Project Year Two: Testing Differences in Leaf Tissue Nutrient Concentrations

While researchers have found Ericaceous plants colonized with Ericoid mycorrhizae (EM) can grow bigger and use nutrients more efficiently compared to the same plants without mycorrizae colonization, very little research has been done to quantify the economic impact of EM in commercial blueberry fields. Commercial EM inoculant products have recently become available, but we are curious whether or not the cost of buying and using these products is justified. Our research aims to verify that EM inoculation works, in the sense that roots are actually becoming colonized when inoculated, compared to roots that we do not inoculate. This is as much a study into the methods of using a microscope to observe the beneficial fungus as it is an investigation into the effectiveness of inoculating with a particular product. Our goal is to confirm EM observation methods that would be most practical for other growers with access to a microscope and a basic scientific background. 

Another goal for the project is to investigate whether inoculation with EM has an ultimate effect on fruit yield. Most work will take place during the third year of the project in 2014, when we will measure both differences in % root colonization and differences in yield, if any. This year, 2013, we tested if there were any differences in leaf tissue nutrient concentration between inoculated bushes and controls.

University of Vermont Plant and Soil Science department has been a collaborator, with Mark Starrett serving as the project adviser. Mark has studied Ericoid mycorrhizae, and found that many peat-based nursery media products contain Ericoid. His research implies that we might not find a significant difference in Ericoid colonization between inoculated bushes and controls, if Ericoid were to be present in the potting mix for all 1000 bushes (treatments and controls) that we outplanted. We do not test potting mixes in this project, but comparing colonization rates in the field should indicate whether or not there was EM colonization in pots before bushes were outplanted. We can also test whether EM is present in the native soil of our field by rooting cuttings in the soil as a media, and then testing for EM colonization.

In 2013, faculty and graduate staff from the University of Vermont Biology department became interested in the project. For future project ideas, Alison Brody and John Gonzales are intrigued to study the effect of EM colonization on blueberry nectar and pollinator activity. Another faculty, Gene Harris, is interested in studying any genetic alterations to the blueberry plant after colonization by EM. Brody and Gonzales have begun to do minor sampling of some of the project treatment and controls to gauge differences in EM colonization. We all plan to collaborate in 2014 to highlight the most practical methods of Ericoid observation for farmers. Gonzales has had some success with ink and vinegar staining, which we hope to develop into a feasible method for EM colonization observation for blueberry roots.

Attached are some photos taken by Brody and Gonzales of stained Ericoid mycorrhizae on Duke blueberry roots. 

2013 was a very quiet year, both in terms of this project and the overall work on our farm. Not much has changed with the blueberries, though we were able to confirm in 2013 that a living mulch strategy for the blueberries is not likely to work. In 2012 with the 1100-bush stand for this project, we seeded white dutch clover surrounding all bushes within rows. The theory was that the clover’s low and lateral spreading growth habit would make a good living much to surround the blueberry bushes and eliminate other weed pressure. We planted it densely around the bushes. While we had good, dense germination and spreading, the clover was no match for some quackgrass and other annuals that shot up out of the clover and outcompeted it. We will likely just dump on 5 inches of wood chips as a mulch surrounding the bushes next spring. We are not disappointed with the results, because we feel that the clover did provide competition for some weed pressure, and it also leaves a lot of good organic matter and nitrogen once weeded out around crowns or buried with other wood chip mulch.

Objectives/Performance Targets

No Significant Differences in Leaf Tissue Nutrient Concentrations

We continued to learn the methods for EM observation. It consists of a two-step process, clearing the roots of their natural pigment and then staining the fungus to highlight its presence. Unfortunately, one investigation got cut short because of personnel changes in UVM Plant and Soil Science. Our mini-experiment was to determine how long roots should soak in KOH at room temperature for adequate clearing before staining.  Samples are usually heated very quickly in an autoclave for clearing, but most growers don’t have access to an autoclave, so we wanted to test the long method where samples are left in KOH at room temperature. This is a simple experiment that we should be able to resume late this winter (Feb-March 2014).

Brody and Gonzales are investigating ink and vinegar staining at the time of filing this report (Dec-Jan 2013-2014).

The core project activity for 2013 was sampling leaf tissue to determine if there were any differences in leaf nutrient concentrations between treatments (inoculated with EM) and controls (not inoculated). We collected 40 composite samples. Each sample contained about 40-50 mature leaves from the top sections of healthy canes. To make that 40-50 leaf sample, we took 1-2 leaves from every bush in the 25-bush row, except for bushes at the ends. The 40 composite samples were sent to the UVM Extension lab for nutrient testing. Data for these samples is attached. Treatment and control data were analyzed using ANOVA statistics software provided by AGSTATS, available free online at : .

With the exception of Potassium, there were no significant differences between treatments and controls in all four blocks we tested for leaf tissue nutrient concentrations for all macro and micro nutrients. This is even when using a P-value of 20%, which would mean we would only need an 80% probability to declare there was a significant difference that was due to the specific effect of inoculation and not everyday field variations. Most data analysis for these types of projects use a 5% or 10% P-value to determine whether or not there is a significant difference. We cast a wide net at first, and planned to analyze for more certainty later, but all of the data sets (with the exception of Potassium) did not pass the test with P=20%.

So what’s going on with Potassium? In three out of the four blocks, leaf tissue concentrations of Potassium were higher for the controls than for the treated bushes (Each block contained 5 repetitions of treatment/control pairs). For the significant differences, the P-values were 16.6%, 8.22% and 10.21%, not a high degree of certainty, but still enough to warrant questioning. Why would Potassium concentrations in leaves be lower in inoculated bushes? There are various possible explanations. One is simply the degree of variation we observed is a normal phenomenon, in other words, Potassium levels in leaves fluctuate widely in the field. A statistical analysis would need to use a much higher degree of certainty to declare a real significant difference caused by the treatment. Another explanation is that the inoculation did effect the way Potassium is moving through the blueberry plant, and more Potassium is going to shoot or root growth and less to the leaves. A full comparison between treatments and controls of Potassium concentrations in roots and shoots would need to be conducted to verify this.

Another possibility explaining the lower leaf Potassium levels in inoculated bushes could be that nutrient use efficiency has changed. One of the most comprehensive studies of EM colonization effects on nutrient concentrations in blueberry bushes was conducted by USDA ARS researcher CF Scagel in 2005. (Available online at : ) The study found that for many of the cultivars grown in organic fertilizer systems, biomass nutrient concentrations either decreased or remained the same as controls, but biomass weights increased for bushes inoculated with EM. This is a very important concept to consider when thinking about leaf tissue nutrient concentrations. More is not necessarily better, and less is not necessarily worse. Nutrient use efficiency can change in cases where the plant can grow bigger using less of the nutrient in question. We will not be able to measure leaf, shoot or root biomass as part of this project, but our measurement of yields next year might indicate whether or not there are differences in nutrient use efficiency. If the bushes that showed lower levels of Potassium yield the same or more than controls, we might conclude that there is a difference in nutrient use efficiency between treatment and controls.

It should be noted that mean leaf tissue Potassium levels for both treatment and controls for all four blocks were well within the normal range. For Potassium, Cooperative Extension bulletins state there is a deficiency below 0.35% and the normal range is 0.4%-0.65%. In the three blocks mentioned above that had a significant difference between treatment and controls, mean levels for treatments were 0.55%, 0.49% and 0.50% compared to controls of 0.61%, 0.53% and 0.53% respectively. These levels don’t appear to be much different when the means alone are shown. The difference does not likely have practical significance, but it is something we will be keeping an eye on.

A final note about the leaf tissue nutrient level testing in this project: Many studies have found that mycorrhizae activity is lower when soil nutrient levels are inherently high or there is more-than-adequate fertilization by the grower. We can rule out this circumstance of high nutrient levels clouding out EM response. As an indicator that our field had low soil-available N, our field tested low leaf tissue Nitrogen levels for every sample tested. The threshold below which deficiency symptoms can adversely impact the crop for Nitrogen in blueberry leaves is about 1.7%. Our samples ranged from 1.25% to 1.69%. It is clear that the plants in our field in 2013 were under-fertilized in terms of one of the most critical nutrients for blueberry production, Nitrogen. EM inoculation did not seem to provide an observed difference to enable inoculated bushes to uptake more Nitrogen.

Was there no significant difference in leaf tissue concentrations of Nitrogen because there was no significant difference in colonization of EM on the roots? We look forward to doing the analysis of colonization levels in 2014. We also look forward to future comparisons of leaf tissue concentrations, as we are aware that plants are only their second year in the ground, still immature, and it might take a while for EM colonization or other biological systems below ground to develop to the point of representing the true character of the 30-50 year lifespan of the blueberry field.


Impacts and Contributions/Outcomes


Josh Roberts
Graduate Assistant
University of Vermont
Plant and Soil Science, Jeffords Hall, 63 Carrigan Drive
Burlington, VT 05405
Office Phone: 4842257878
Mark Starrett
Associate Professor
Plant and Soil Science, Jeffords Hall, 63 Carrigan Drive
Burlington, VT 05405
Office Phone: 8026560467