Final Report for GS09-084
Project Information
The objectives of this project included characterizing plant pathogen and general microbial population changes in soils under different sustainable management practices for strawberry production systems at two different locations in Arkansas. A brassica cover crop, mustard seed meal, solarization or a combination of the cover crop and solarization were compared to no soil treatment prior to establishing the strawberry crop. General microbial and suspect pathogen populations from soils were quantified by plate count methods. Additional soil samples were taken after cover crop incorporation to generate denatured gradient gel electrophoresis (DGGE) profiles for bacterial and fungal populations. Roots of strawberry plants, including the initial transplants, were also analyzed for the isolation frequency of plant pathogens.
Pythium populations were numerically higher in soils following the brassica cover crop compared to the control, but results were only significant at one site where Pythium was also increased when brassica cover crop was followed by solarization and in mustard seed meal amended soils compared to the control. Rhizoctonia was only isolated from soil at one location, where binucleate and multinucleate Rhizoctonia populations were higher in soils that received a brassica cover crop compared to no soil treatment. However, soil treatments usually did not significantly affect the frequency of Pythium, Rhizoctonia or Colletotrichum isolation from the roots of strawberry. The only exception was in plots that had been planted with a brassica cover crop and then solarized prior to strawberry planting. In these plots, 15.3% percent of strawberry plants were found to be colonized by Rhizoctonia solani compared to only 5.0% in control plots. Approximately 80% of strawberry transplants were colonized by Pythium, Rhizoctonia or Colletotrichum before they were planted into the test plots. Root ratings, which were used to monitor disease severity based on the percent discoloration of roots, did not differ among the treatments. Along with contamination of transplants, unusually high rainfall amounts prior to and shortly after strawberry transplanting likely contributed to a compromised root system for all plants.
Soil treatments did affect the level of bacterial, fungal and actinomycete populations in the soil at the time of brassica cover crop termination and at strawberry transplant. In all cases, there was a trend for higher bacterial, fungal and actinomycete populations in brassica, brassica plus solarization and mustard seed meal amended soils compared to solarized only and control soils, yet results were not always significant at both test locations. Total culturable, bacterial populations were significantly higher in soils that had been planted with a brassica cover crop followed by solarization and soils receiving mustard seed meal amendments at both locations at the time of strawberry transplanting. From soil samples taken at 7 and 25 days after the brassica cover crop was incorporated into the soil, denatured gradient gel electrophoresis (DGGE) produced unique profiles of bacteria and fungi compared to that of control soils. At the time of strawberry transplant, all bacterial DGGE profiles of soils from both locations receiving different treatments were still distinct and grouped separately in dendograms, yet fungal DGGE profiles were not as consistently distinct among treatments.
This project has successfully proven how including soil treatments such as a brassica cover crop, solarization or mustard seed meal application as a practice in annual strawberry production can enhance the soil microflora, especially the bacterial community. Since changes could be observed in both the bacterial and fungal communities throughout the sampling times, this system has the potential to produce a soil that is more diverse and possibly reduce populations or colonization of roots by soilborne pathogens. The impacts of these shifts in the soil microflora for soilborne diseases should be compared to chemical fumigants in soil with a history of strawberry production to examine their value in developing a sustainable strawberry production system.
Introduction
Soilborne pathogens are a limiting factor for strawberry production and historically have been managed through the use of fumigants, especially methyl bromide. Common soilborne diseases of strawberries are black root rot, crown and leather rot, red stele and Verticillium wilt (Guerena and Born, 2007; Maas, 1988). It has been shown that rotation with a brassica crop, broccoli, led to a reduction in Verticillium wilt of strawberry and microsclerotia in the soil compared to a non-brassica crop, lettuce (Subbarao et al., 2007). Additionally, in vitro studies have shown that macerated roots of brassica suppressed six different soilborne pathogens of strawberry, including Colletotrichum dematium, Cylindrocarpon destructans, Fusarium oxysporum, Pythium ultimum, Phytophthora cactorum and Rhizoctonia fragariae (Mattner et al., 2008). Another study has shown that a brassica-strawberry cropping sequence resulted in increased fruit yield compared to a non-treated control and showed no significant difference in yield from methyl bromide treated plots (Lazzeri et al., 2003).
Brassica spp. produce glucosinolates which decompose into chemicals that are inhibitory to a range of microorganisms including nematodes, fungi and bacteria. A proposed mechanism of disease suppression has been based on the release of these chemicals. However, these volatile chemicals of glucosinolate hydrolysis are detected in soil for only short periods of time, with reports ranging from 24 hours (Mazzola et al., 2007) to 12 days (Njoroge et al., 2008). When brassica seed meal was incorporated into soil four weeks prior to infestation with Rhizoctonia solani, there was a significant reduction in apple seedling infection, indicating other factors besides glucosinolate hydrolysis products play a role in suppressing disease (Mazzola et al., 2007). The authors of this study concluded that decreased root infection by Rhizoctonia was associated with increased populations of Streptomyces. Earlier research reported brassica seed meal amendments increased fluorescent Pseuodomonas, actinomycetes, and total bacterial populations (Mazzola et al., 2001), suggesting that benefits of using brassica cover crops could be associated with enriching the microbial population resulting in suppression of pathogens of the crop of interest.
This initial study investigated changes in the soil pathogen and general microbial populations over time after a brassica cover crop, mustard seed meal application, solarization and a combination of a brassica cover crop and solarization compared to control soils. Additionally, strawberry roots were examined for disease symptoms and colonization by potential pathogens. The goal of this research effort is to contribute to the development of a sustainable production system by suppression of soilborne strawberry diseases without the use of chemical soil fumigants.
This research is part of a larger project to develop a sustainable system for annual strawberry production using a brassica cover crop for soilborne disease suppression and will be done according to the following objectives:
1) Quantify changes in disease incidence and severity and soil pathogen populations in a strawberry cropping system that includes a brassica cover crop, mustard seed meal application, solarization or a combination of a brassica cover crop and solarization as preplant treatments
2) Quantify and characterize soil microbial changes in different strawberry production systems that include a brassica cover crop, mustard seed meal application, solarization or a combination of a brassica cover crop and solarization as preplant treatments
Monitoring changes in the soil microbial composition that result from the adoption of a brassica cover crop will help indicate whether this is a factor contributing to disease suppression. Strawberries are a high value crop identified as an attractive option for limited resource farmers or farmers with limited acreage. This research will aid in determining if brassica cover crops are a desirable option in strawberry disease protection for those farmers adverse to the high input, chemically intensive strawberry production system.
Cooperators
Research
Two field locations, the Vegetable Research Station at Kibler, AR, and the Southwest Research and Extension Center at Hope, AR, were established on sites not previously planted to strawberry. Soil treatments included a brassica cover crop, solarization, brassica cover crop followed by solarization (brassica plus solarization), mustard seed meal and a control that received no soil treatment prior to establishing the strawberry crop. The experiments were completely, randomized block designs with 4 replications per treatment. Fertilizer was applied throughout the experiment based on soil testing results and strawberry foliage nutrient analysis. The brassica cover crop, Seven Top Turnips, was planted on June 15 at Hope and on June 30 at Kibler. The cover crop was terminated by tilling it into the soil on August 21 at Hope and August 26 at Kibler. On these same dates, Oriental mustard seed meal (MPT Mustard Products & Technology, Saskatoon, SK) was applied at a rate of 1000 pounds per acre. Clear plastic was laid over the soil for brassica plus solarization and solarization treatments, and all other beds were covered with black plastic at this time. The clear plastic was painted black on October 5 at Hope and on November 6 at Kibler. Camerosa strawberries were transplanted into raised beds on October 12 at Hope, and Chandler strawberries were transplanted on October 19 at Kibler. The strawberry plants were transplanted into raised beds with 2 rows 12 inches apart per bed, and plants were spaced 14 inches apart within the rows. The plants were covered with frost protection cloth throughout the winter due to below freezing temperatures at both locations. ABOUND fungicide (active ingredient: azoxystrobin) was applied to strawberry plants on March 11 at both Hope and Kibler.
Soil samples were taken from the two field locations directly before cover crop termination and at strawberry transplant. Ten soil samples per plot were taken with a 2.5cm corer at a depth of 0 - 15cm. Pythium and Rhizoctonia populations were determined by dilution plating on P5ARP media (Jeffers and Martin, 1986) and by the multiple-pellet technique on Ko and Hora media (Ko and Hora, 1971), respectively.
Roots of the initial transplants and strawberry plants at mid-flowering (March 31 at Hope and April 5 at Kibler) and at the end of harvest (June 28 at Kibler only) were examined for pathogen colonization. Roots of 26 transplants and roots from 10 and 5 strawberry plants per plot at Kibler and Hope, respectively, were washed, disinfested in a 0.5% NaOCl solution for 90 seconds and blotted dry in paper towels. Segments of 4 separate roots per plant, giving preference to areas at the edge of lesions if possible, were cut into 2-3 cm segments and plated on 0.8% water agar supplemented with 10 mg/L rifampicin, 250mg/L ampicillin, and 0.5 ul/L of the miticide Danitol (WArad). Colonies emerging from roots 2-4 days after plating were transferred to Potato Dextrose Agar supplemented with10 mg/L rifampicin, 250mg/L ampicillin, and 0.5 ul/L of the miticide Danitol (PDArad) for identification.
In an attempt to isolate Phytophthora spp., 4 separate, washed roots were first cut into 2-3 cm segments, dipped into ethanol, blotted dry and plated on P5ARP media. P5ARP plates were stored at 17o C for 2 weeks. Colonies emerging from roots were transferred onto fresh P5ARP plates starting at 4 days after plating and up to 2 weeks. Cultures were then transferred to PDArad for observation of colony morphology. All isolates successfully grew on PDArad. Those isolates that possessed colony morphologies similar to Phytophthora were grouped, and 2 cultures from each group were sequenced using ITS1 and ITS4 primers (White et al., 1990) of the ITS region for tentative identification.
Roots were rated for disease symptoms based on the percent discoloration on a scale of 0-5, where 0 = 0-10%, 1 = 10-30%, 2 = 30-50%, 3 = 50-70%, 4 = 70-90% and 5 = 90-100% discoloration.
Soil samples taken directly before cover crop termination and at strawberry transplant were assayed for soil microbial populations including total bacteria, fungi and actinomycetes using dilution plating on Tryptic Soy Agar, Rose Bengal Agar supplemented with 30 mg/L streptomycin sulfate and Chitin media adjusted to pH 8.0-9.0, respectively.
Analyses for disease, isolation and population were conducted by GLM using SAS (SAS Institute, Cary, North Carolina). General soil populations were analyzed as Log10 populations. A protected LSD was used to compare treatments for variables having a significant P value (<0.05).
A subsample of the soil samples taken directly before cover crop termination and at strawberry transplant in addition to soil samples taken 7 and 25 days after the cover crop had been incorporated into soil were used to generate denatured gradient gel electrophoresis (DGGE) profiles for bacterial and fungal populations. As a result of heavy rainfall at Kibler, soil sampling at 25 days post cover crop termination was not possible, yet the respective sampling was carried out at the Hope location. Soil was frozen within 24 hours of collection and stored at -20o C until processed. DNA was isolated using the Powersoil DNA Isolation Kit (MoBio, Carlsbad, CA, USA). Variable regions of the 16S rRNA gene of bacteria were amplified using primers PRBA338F and PRUN518R (Nakatsu 2000) with the following cycling conditions: initial denaturing at 94o C for 2 min, a pause at 80o C when the polymerase was added, then touchdown PCR including 18 cycles of denaturing at 94o C for 1min, annealing at 67o C for 45 sec minus 0.5o C per cycle, extension at 72o C for 2 min, followed by 13 cycles of denaturing at 94o C for 1 min and annealing at 67o C for 45 sec. The final extension was set at 72o C for 10 min. Nested PCR was performed for amplification of the 18S rRNA gene of fungi using EF4 and Fung5 primers for the first round, followed by EF4 and NS2-GC primers for the second round (Zhang et al., 2007). Cycling conditions for the 18S rRNA gene included initial denaturing at 94o C for 3 min, a pause at 80o C when the polymerase was added, then 40 cycles of denaturing at 94o C for 1 min, annealing at 47o C for 1min and extension at 72o C for 3 min. The final extension step was set at 72o C for 10 min. Products from the first PCR were diluted 1:00 in water and used as the template for the second PCR. Cycling conditions for the second PCR were the same as the first. PCR products were loaded on 8% (w/v) acrylamide/bisacrylamide (37.5:1) gels and run for approximately 1000 volt hours at 60o C using the DCode system (Bio-Rad, Hercules, CA, USA). Gels were stained with SYBR green for 40 min followed by a 10 min destain, scanned with the Kodak EDAS 290 (ISC BioExpress, Kaysville, UT, USA) and then analyzed using Quantity One software (Bio-Rad). Lane background was subtracted using the rolling disk option. DNA bands were matched and compared among lanes, normalized (to account for loading error) and then assembled into a dendogram using an unweighted, UPGAMA cluster analysis.
Significant differences were found for Pythium populations among the various soil treatments. At the time of the cover crop termination, Pythium populations were numerically higher in soils planted with the brassica cover crop than control soils, yet the difference was not significant (P= 0.0603 at Kibler, and P= 0.0951 at Hope) (Table 1). At the time of strawberry transplanting, Pythium populations were significantly higher in soils that had only the brassica cover crop at both Hope and Kibler and also in soils with mustard seed meal and the brassica cover crop plus solarization at Kibler. Other studies have also observed increased Pythium populations after brassica amendments (Cohen and Mazzola, 2006; Njoroge et al., 2008). Cohen and Mazzola (2006) noted that, with Brassica napus seed meal, higher Pythium soil counts did not always correlate with an increase in Pythium infection of apple seedlings. The distribution of particular highly virulent versus less virulent Pythium spp. seemed to play a more important role in apple seedling infection instead of the population of Pythium in the soil (Cohen and Mazzola, 2006). In the current project, the distribution of Pythium spp. was not determined, yet higher amounts of Pythium in soils planted with a brassica cover crop or those amended with mustard seed meal did not result in significantly higher percentages of Pythium colonization in strawberry roots compared to control soils (Table 2). At flowering, the range of percent isolation of Pythium from strawberry roots was 57.5 – 64.2% at Kibler and 50.0 - 90.0% at Hope for all treatments, while the percent of plants with Pythium isolated from the roots at harvest ranged from 2.5 – 32.5% at Kibler.
Rhizoctonia populations were higher in those soils that received a brassica cover crop compared to the control at the Hope location. The increase in soil populations was significant for multinucleate Rhizoctonia solani and binucleate Rhizoctonia (BNR) isolates. Rhizoctonia was not detected in soil from any plots at Kibler (Table 1). Excessive rain at strawberry transplanting prevented the use of the multiple-pellet method to examine Rhizoctonia populations at Kibler and Hope.
Both BNR and R. solani are thought to be pathogens involved in the black root rot complex of strawberry (Mass, 1988; Sharon et al., 2007). Certain anastomosis groups (AG) of Rhizoctonia have been found to infect strawberry including BNR AG-A, AG-G, AG-I, AG-F and AG-K (Martin 2000; Sharon et al., 2007) and R. solani AG-4, AG-5, AG-6 (Sharon et al., 2007, Martin, 1998; Botha et al., 2003). It is yet to be determined the AG designation for the Rhizoctonia isolates found in this study. Although BNR was commonly isolated, no significant differences in colonization of strawberry roots by BNR was found among the treatments (Table 2). At flowering, BNR was isolated from the roots of 35.0 – 55.0% of plants at Kibler and 60.0 – 85.0% of plants at Hope for all treatments. At harvest, the range of BNR isolation from strawberry roots was 20.0 – 55.6% of plants at Kibler. Only the Kibler location exhibited a greater degree of MNR root colonization of strawberry planted in the brassica cover crop plus solarization soils, yet isolation percentages were only 15.3% in these soils compared to 5.0% and 2.5% for strawberry planted in control and solarized soils, respectively. With these results, it is still unclear if the given soil treatments consistently increase soil populations and colonization of strawberry by pathogenic Rhizoctonia, and, therefore, the AG and pathogenicity of the Rhizoctonia spp. isolates should be determined.
Colletotrichum was also commonly found from roots plated on WArad, yet soil treatments appeared to have no effect on the percent isolation of Colletotrichum (Table 2). Colletotrichum is a causative agent of crown rot of strawberry and can also cause a decline in plant vigor and wilting due to colonization of the roots and buds (Maas, 1988).
Despite attempts to isolate Phytophthora from the roots of strawberry, all suspect isolates that were sequenced from root isolations were not of this genus.
Approximately 80% of strawberry transplants proved to be colonized by Pythium, Rhizoctonia or Colletotrichum before they were ever planted into the test plots. The percentage of transplants colonized by Pythium, Rhizoctonia or Colletotrichum prior to planting was 42%, 35%, 42%, respectively. If transplants are being shipped already colonized or infested with these pathogens, it will be more difficult to achieve desirable results with preplant treatments such as those used in this project.
Root ratings, which were used to estimate disease severity based on the percent discoloration of roots, did not differ among the treatments (Table 3). At flowering, the mean root ratings for all treatments ranged from 3.4 – 3.9 at Kibler and 3.2 – 3.7 at Hope. At harvest, root ratings ranged from 2.9 – 3.7 for all treatments at Hope. Both test locations received record rainfall amounts causing saturated soil conditions prior to and shortly after strawberries were transplanted. Kibler received rainfall in excess of 6.66 and 12.26 inches of the average precipitation amounts for the months of September and October, respectively. Hope experienced rainfall that was 8.51 and 13.90 inches above normal in September and October, respectively. These weather conditions likely contributed to the compromised root system for all plants. Many plants at both locations did not survive. Yield was not recorded because fruit production was too low. By the end of harvest at Kibler, root discoloration ratings were still not significantly different according to treatment (Table 3). No samples were taken from Hope due to the high amount of plant death and very poor health of the remaining plants at this location.
Soil microbial changes were observed for the preplant treatments. Soil treatments affected the level of bacterial, fungal and actinomycete populations in the soil at the time of brassica cover crop termination and at strawberry transplant. In all cases, there was a trend for higher bacterial, fungal and actinomycete populations in brassica, brassica plus solarization and mustard seed meal amended soils compared to solarized only and control soils (Table 4). For example, bacterial populations were higher in brassica soils at the time of cover crop termination at Hope (P= 0.0436) and also at Kibler, yet the difference was not significant at Kibler (P= 0.0535). At the time of strawberry transplanting, the trend was for higher bacterial populations in all soils which had been planted with a brassica cover crop soils to control soils, yet results were only significant for brassica cover crop plus solarization and mustard seed meal amended soils at Hope and Kibler, as well as in the brassica cover crop soils at Kibler. Fungal populations were only significantly different at Kibler, where an increase in brassica planted soils compared to control soils was observed at the time of cover crop termination. Numerically higher fungal populations in brassica soils compared to control soils also were present at transplanting, but high coefficients of variation may have prevented differences from being significant. Finally, actinomycetes exhibited a trend for higher populations in brassica and brassica plus solarization soils, yet results were only significant at Kibler for the brassica soils at the time of cover crop termination and at Hope for the brassica and brassica plus solarization soils at the time of strawberry transplant. The increases of all mentioned microbial groups, including genera with pathogenic members, in brassica and mustard seed meal amended soils is likely due to the increase in soil organic matter associated with the amendments.
To detect changes in the composition, versus an increase or decrease, of microbial populations, DGGE proved beneficial. Unique microbial profiles were produced by DGGE representing bacterial and fungal communities of brassica versus control soils, and this effect was visible on those gels generated from sampling times at 7 days post cover crop incorporation (Figure 1) and on gels representing the fungal community at 25 days post cover crop incorporation into the soil (data not shown). Dendograms of the DGGE profiles of the 16S rRNA gene of bacteria at 7 days after brassica cover crop incorporation into soils showed that the brassica cover crop and control treatments were distinct in the effect on the bacterial community (Figure 2). The similarity among the represented bacterial community from brassica and control soils was 74% at Kibler and 77% at Hope. Furthermore, at 7 days post cover crop incorporation, the soils which had a brassica cover crop had more variability in the 16S rRNA gene profiles than control soils at both locations. From soils taken 25 days after the brassica cover crop was incorporated into soils, DGGE profiles representing the bacterial communities of cover crop and control soils were 85% similar (data not shown). Finally, samples taken at the time of strawberry transplant at both Kibler and Hope produced bacterial DGGE profiles that were still only 84% and 70% similar, respectively (Figure 3). When comparing brassica, brassica plus solarization, solarization and mustard seed meal treatments at both Kibler and Hope at the time of strawberry transplant, all treatments were unique in the effect on the soil bacterial DGGE profiles as can be seen by separation from each other and from the control in the dendograms (Figure 3).
The fungal community, as assessed by DGGE profiles of a region of the 18S rRNA gene, could be differentiated in brassica versus controls soils at 7 days post brassica cover crop incorporation into soils (Figure 4). Based on the fungal DGGE profiles at 7 days post cover crop incorporation, brassica and control soils were 84% and 76% similar at Hope and Kibler, respectively (Figure 5). At 25 days after the cover crop was terminated and incorporated, fungal DGGE profiles of brassica and control soils were only approximately 70% similar (data not shown). By the time of strawberry transplanting, only brassica soils at Kibler and solarized soils at Hope resulted in fungal profiles unique from all other treatments (Figure 6).
Amplification of the 16S rRNA gene of bacteria followed by DGGE resulted in detection of up to approximately 50 bands depending on the sampling time. However, band detection from amplification of the 18S rRNA fungal gene followed by DGGE was much less and reached approximately only 30 bands. Combing this data set with that derived from another set of PCR primers targeting conserved fungal genomic regions could provide more insight into the fungal community dynamics as affected by soil treatments used in this study. Nonetheless, the segments of the populations amplified and differentiated in this analysis was adequate to observe populations shifts for up to 25 days for fungal communities and up to 53 days for bacterial communities due to growing and incorporation of the brassica cover crop.
Changes in the soil microflora associated with a brassica cover crop, mustard seed meal application, solarization and a combination of a brassica cover crop and solarization could result in a more diverse soil microbial community with capabilities of suppressing pathogen populations and plant disease or promoting plant health. Annual strawberry production is a relevant system to incorporate these soil treatments due to the fallow period during the late summer months. In the case of this project, contamination of transplants and unusually high rainfall amounts prior to and shortly after strawberry transplanting likely contributed to poor root health and a difficulty in determining treatment effects on suppression of pathogen colonization of strawberry roots and disease symptoms. A further analysis of the pathogenicity of the Pythium and Rhizoctonia species associated with the strawberry roots before transplant and then later in the growing season could indicate the benefit of the mentioned soil treatments in preventing colonization by these pathogen groups. Also, testing on fields with a history of strawberry production could document the value of these treatments to disease suppression, productivity and sustainability for strawberry production compared to the current system which includes chemical soil fumigants.
- Table 1. Soil populations of select pathogenic genera for different preplant strawberry treatments
- Figure 5. Dendograms of 18S rRNA DGGE profiles from Kibler and Hope soils sampled 7 days post brassica cover crop incorporation. a) Kibler soils, b) Hope soils; C: control; B: brassica cover crop; 1,2,3 and 4: replication 1,2,3 and 4.
- Table 2. Isolation frequency of select pathogenic genera from strawberry roots at flowering and at the end of strawberry harvest.
- Table 3. Strawberry root discoloration ratings of plants for different preplant strawberry treatments.
- Table 4. Soil populations of select microbial groups for different preplant strawberry treatments.1
- Figure 1. DGGE profiles of bacterial 16S rRNA gene from Kibler and Hope soils sampled at 7 days post brassica cover crop incorporation. a) Kibler soils, b) Hope soils; C, control; B, brassica cover crop; 1,2,3 and 4: replication 1,2,3 and 4. Arrows indicate bands that display a change in relative abundance between treatments.
- Figure 6. Dendograms of bacterial 18S rRNA gene DGGE profiles from Kibler and Hope soils sampled at the time of strawberry transplant. a) Kibler soils, b) Hope soils; C, control; B, brassica cover crop; BS, brassica cover crop followed by solarization; S, solarization; M, mustard seed meal; 1,2,3 and 4: replication 1,2,3 and 4.
- Figure 2. Dendograms of 16S rRNA DGGE profiles from Kibler and Hope soils sampled 7 days post brassica cover crop incorporation. a) Kibler soils, b) Hope soils; C: control; B: brassica cover crop; 1,2,3 and 4: replication 1,2,3 and 4.
- Figure 3. Dendograms of bacterial 16S rRNA gene DGGE profiles from Kibler and Hope soils sampled at the time of strawberry transplant. a) Kibler soils, b) Hope soils; C, control; B, brassica cover crop; BS, brassica cover crop followed by solarization; S, solarization; M, mustard seed meal; 1,2,3 and 4: replication 1,2,3 and 4.
- Figure 4. DGGE profiles of the fungal 18S rRNA gene from Kibler and Hope soils sampled at 7 days post brassica cover crop incorporation. a) Kibler soils, b) Hope soils; C, control; B, brassica cover crop; 1,2,3 and 4: replication 1,2,3 and 4. Arrows indicate bands that display a change in relative abundance between treatments.
Educational & Outreach Activities
Participation Summary:
The results of this study will be published as part of a PhD dissertation, and we hope to publish the work completed in this project, in combination with another year of data with strawberry yield data included, in a peer-reviewed journal.
Project Outcomes
This project has successfully proven how including soil treatments such as a brassica cover crop, solarization or mustard seed meal application as a practice in annual strawberry production can enhance the soil microflora, especially the bacterial community. Since changes could be observed in both the bacterial and fungal communities throughout the sampling times, this system has the potential to produce a soil that is more diverse and possibly reduce populations or colonization of roots by soilborne pathogens. Annual strawberry production is a relevant system to incorporate the mentioned soil treatments due to the fallow period during the late summer months. The impacts of the shifts in the soil microflora on soilborne diseases should be compared to chemical fumigants in soil with a history of strawberry production to examine their value in developing a sustainable strawberry production system.
References:
Botha, A., Denman, S., Lamprecht, S. C., Mazzola, M., Crous, P. W., 2003. Characterization and pathogenicity of Rhizoctonia isolates associated with black root rot of strawberries in the Western Cape Province, South Africa. Australasian Plant Pathology 32 (2): 195-201.
Cohen, M. F., Mazzola, M., 2006. Resident bacteria, nitric oxide emission and particle size modulate the effect of Brassica napus seed meal on disease incited by Rhizoctonia solani and Pythium spp. Plant and Soil 286: 75-86.
Guerena, M., Born, M., 2007. Strawberries: Organic Production. National Sustainable Agriculture Information Service (ATTRA) and the National Center for Appropriate Technology (NCAT)
Jeffers, S. N., Martin, S. B., 1986. Comparison of two media selective for Phytophthora and Pythium species. Plant Disease 70: 1038-1043.
Ko, W. H., Hora, F. K., 1971. A selective medium for the quantitative determination of Rhizoctonia solani in soil. Phytopathology 61 (6): 707-710.
Lazzeri, L., Baruzzi, G., Malaguti, L., Antoniacci, L., 2003. Replacing methyl bromide in annual strawberry production with glucosinolate-containing green manure crops. Pest Management Science 59 (9): 983-990.
Maas, J. L., 1988. Compendium of strawberry diseases. APS Press, St. Paul, Minnesota, USA.
Martin, S. B., 1988. Identification, isolation and pathogenicity of anastomosis groups of binucleate Rhizoctonia spp. from strawberry roots. Phytopathology 78: 379–384.
Martin, F. N., 2000. Rhizoctonia spp. recovered from strawberry roots in central coastal California. Phytopathology 90 (4): 345-353.
Mattner, S. W., Porter, I. J., Gounder, R. K., Shanks, A. L., Wren, D. J., Allen, D., 2008. Factors that impact on the ability of biofumigants to suppress fungal pathogens and weeds of strawberry. Crop Protection 27 (8): 1165-1173.
Mazzola, M., Granatstein, D. M., Elfving, D. C., Mullinix, K., 2001. Suppression of specific apple root pathogens by Brassica napus seed meal amendment regardless of glucosinolate content. Phytopathology 91: 673-679.
Mazzola, M., Brown, J., Izzo, A., Cohen, M. F., 2007. Mechanism of action and efficacy of seed meal-induced suppression of pathogens inciting apple replant disease differ in a Brassicaceae species and time-dependent manner. Phytopathology 97: 454-460.
Nakatsu, C. H., Torsvik, V., Ovreas, L., 2000. Soil community analysis using DGGE of 16S rDNA polymerase chain reaction products. Soil Science Society of America 64 (4): 1382-1388.
Njoroge, S. M. C., Riley, M. B., Keinath, A. P., 2008. Effect of incorporation of Brassica
spp. residues on population densities of soilborne microorganisms and on damping-off and Fusarium wilt of watermelon. Plant Disease 92: 287-294.
Sharon, M., Freeman, S., Kuninaga, S., Sneh, B., 2007. Genetic diversity, anastomosis groups and virulence of Rhizoctonia spp. from strawberry. European Journal of Plant Pathology 117 (3): 247-265.
Subbarao, K. V., Kabir, Z., Martin, F. N., Koike, S. T., 2007. Management of soilborne diseases in strawberry using vegetable rotations. Plant Disease 91 (8): 964-972.
White, T. J., Bruns, T., Lee, S., Taylor, J., 1990. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. PCR Protocols: A guide to Methods and Applications (ed. M. A. Innis, D. H. Gelfand, J. J. Sninsky & T. J. White) Academic Press: San Diego, U.S.A, pp 315-322.
Zhang, W., Qiao, Z., Tang, Y., Hu, C., Sun, Q., Morimura, S., Kida, K., 2007. Analysis of the fungal community in Zaopei during the production of Chinese Luzhou-flavour Liquor. Journal of the Institute of Brewing and Distilling 113 (1): 21-27.
Farmer Adoption
At this stage in the research, the management strategies used in this project have not provided evidence of a benefit in regards to increased yield or disease suppression in an annual strawberry production system due to soil treatments such as a brassica cover crop, mustard seed meal application, solarization and a combination of a brassica cover crop and solarization. We believe that with more desirable weather conditions and a nursery source that can provide cleaner strawberry plants more conclusive data could be obtained. However, we are currently contacting strawberry farmers in the state to determine who could participate in repeating this research in whole or in part.
Areas needing additional study
Determining which species of bacteria, fungi and actinomycetes are enriched or suppressed by soil treatments and comparing these treatments with the conventionally used soil fumigants will help determine the unique contribution of a given treatment to strawberry disease suppression and yield. To better understand the soil treatment effects on soilborne pathogens of strawberry, it would be beneficial to monitor specific pathogenic species belonging to Pythium and Rhizoctonia genera in the soil and/or isolated from strawberry roots to determine if soil treatments influence a more or less virulent pathogen distribution.
The undesirable weather conditions during this project in conjunction with nursery-infested strawberry plants most likely contributed to a lower quality of plants at harvest. Therefore, this study will need to be repeated to determine more accurately the soil treatment effects on strawberry plant health and fruit production.