Synthetic microbial consortia derived from rhizosphere soil protect wheat against a soilborne fungal pathogen.

Synthetic microbial communities (SynComs) could potentially enhance some functions of the plant microbiome and emerge as a promising inoculant for improving crop performance. Here, we characterized a collection of bacteria, previously isolated from the wheat rhizosphere, for their antifungal activity against soilborne fungal pathogens. Ten SynComs with different compositions from 14 bacterial strains were created. Seven SynComs protected wheat from Rhizoctonia solani AG8 infection, although SynComs were not more effective than single strains in reducing wheat root rot disease. Further, the mechanisms of interaction of the tested bacteria with each other and plants were explored. We found that nine bacteria and nine SynComs impacted the root growth of Arabidopsis. Nine bacteria and four SynComs significantly inhibited the growth of AG8 by producing volatiles. The cell-free supernatants from six bacteria inhibited the growth of AG8. Together, this study provided the potential for improving crop resilience by creating SynComs.

Occurrence of Mummy Berry Associated with Huckleberry (Vaccinium membranaceum) Caused by Monilinia spp. in Oregon.

In this Short Communication we describe the occurrence of mummy berry associated with huckleberry (Vaccinium membranaceum) caused by Monilinia spp. in Oregon. To our knowledge, this is the first report of a Monilinia spp. associated with mummy berry of huckleberry in Oregon. Sequence data from our specimens reveal the closest identity was Monilinia vaccinii-corymbosi, a pathogen of commercial blueberry (Vaccinium corymbosum). This may be a new species of Monilinia, not previously reported on huckleberry, and further investigation is needed. Of specific importance, the huckleberry holds cultural importance as a sacred First Food of the Confederated Tribes of the Umatilla Indian Reservation and other Pacific Northwest tribes. Although plant pathogen management in natural landscapes presents unique challenges, we will work with tribal authorities to determine whether cultural management techniques may mitigate yield loss due to Monilinia spp.

Responses of Soil Fungal Communities to Lime Application in Wheat Fields in the Pacific Northwest.

Liming is an effective agricultural practice and is broadly used to ameliorate soil acidification in agricultural ecosystems. Our understanding of the impacts of lime application on the soil fungal community is scarce. In this study, we explored the responses of fungal communities to liming at two locations with decreasing soil pH in Oregon in the Pacific Northwest using high-throughput sequencing (Illumina MiSeq). Our results revealed that the location and liming did not significantly affect soil fungal diversity and richness, and the impact of soil depth on fungal diversity varied among locations. In contrast, location and soil depth had a strong effect on the structure and composition of soil fungal communities, whereas the impact of liming was much smaller, and location- and depth-dependent. Interestingly, families Lasiosphaeriaceae, Piskurozymaceae, and Sordariaceae predominated in the surface soil (0-7.5 cm) and were positively correlated with soil OM and aluminum, and negatively correlated with pH. The family Kickxellaceae which predominated in deeper soil (15-22.5 cm), had an opposite response to soil OM. Furthermore, some taxa in Ascomycota, such as Hypocreales, Peziza and Penicillium, were increased by liming at one of the locations (Moro). In conclusion, these findings suggest that fungal community structure and composition rather than fungal diversity responded to location, soil depth and liming. Compared to liming, location and depth had a stronger effect on the soil fungal community, but some specific fungal taxa shifted with lime application.

Rhizosphere community selection reveals bacteria associated with reduced root disease.

BACKGROUND: Microbes benefit plants by increasing nutrient availability, producing plant growth hormones, and protecting against pathogens. However, it is largely unknown how plants change root microbial communities. RESULTS: In this study, we used a multi-cycle selection system and infection by the soilborne fungal pathogen Rhizoctonia solani AG8 (hereafter AG8) to examine how plants impact the rhizosphere bacterial community and recruit beneficial microorganisms to suppress soilborne fungal pathogens and promote plant growth. Successive plantings dramatically enhanced disease suppression on susceptible wheat cultivars to AG8 in the greenhouse. Accordingly, analysis of the rhizosphere soil microbial community using deep sequencing of 16S rRNA genes revealed distinct bacterial community profiles assembled over successive wheat plantings. Moreover, the cluster of bacterial communities formed from the AG8-infected rhizosphere was distinct from those without AG8 infection. Interestingly, the bacterial communities from the rhizosphere with the lowest wheat root disease gradually separated from those with the worst wheat root disease over planting cycles. Successive monocultures and application of AG8 increased the abundance of some bacterial genera which have potential antagonistic activities, such as Chitinophaga, Pseudomonas, Chryseobacterium, and Flavobacterium, and a group of plant growth-promoting (PGP) and nitrogen-fixing microbes, including Pedobacter, Variovorax, and Rhizobium. Furthermore, 47 bacteria isolates belong to 35 species were isolated. Among them, eleven and five exhibited antagonistic activities to AG8 and Rhizoctonia oryzae in vitro, respectively. Notably, Janthinobacterium displayed broad antagonism against the soilborne pathogens Pythium ultimum, AG8, and R. oryzae in vitro, and disease suppressive activity to AG8 in soil. CONCLUSIONS: Our results demonstrated that successive wheat plantings and pathogen infection can shape the rhizosphere microbial communities and specifically accumulate a group of beneficial microbes. Our findings suggest that soil community selection may offer the potential for addressing agronomic concerns associated with plant diseases and crop productivity. Video Abstract.

Baseline and Temporal Changes in Sensitivity of Zymoseptoria tritici Isolates to Benzovindiflupyr in Oregon, U.S.A., and Cross-Sensitivity to Other SDHI Fungicides.

Zymoseptoria tritici is the causal agent of Septoria tritici blotch (STB), a disease of wheat (Triticum aestivum) that results in significant yield loss worldwide. Z. tritici's life cycle, reproductive system, effective population size, and gene flow put it at high likelihood of developing fungicide resistance. Succinate dehydrogenase inhibitor (SDHI) fungicides (FRAC code 7) were not widely used to control STB in the Willamette Valley until 2016. Field isolates of Z. tritici collected in the Willamette Valley at dates spanning the introduction of SDHI (2015 to 2017) were screened for sensitivity to four SDHI active ingredients: benzovindiflupyr, penthiopyrad, fluxapyroxad, and fluindapyr. Fungicide sensitivity changes were determined by the fungicide concentration at which fungal growth is decreased by 50% (EC50) values. The benzovindiflupyr EC50 values increased significantly, indicating a reduction in sensitivity, following the adoption of SDHI fungicides in Oregon (P < 0.0001). Additionally, significant reduction in cross-sensitivity among SDHI active ingredients was also observed with a moderate and significant relationship between penthiopyrad and benzovindiflupyr (P = 0.0002) and a weak relationship between penthiopyrad and fluxapyroxad (P = 0.0482). No change in cross-sensitivity was observed with fluindapyr, which has not yet been labeled in the region. The results document a decrease in SDHI sensitivity in Z. tritici isolates following the introduction of the active ingredients to the Willamette Valley. The reduction in cross-sensitivity observed between SDHI active ingredients highlights the notion that careful consideration is required to manage fungicide resistance and suggests that within-group rotation is insufficient for resistance management.

Population Dynamics of Wheat Root Pathogens Under Different Tillage Systems in Northeast Oregon.

No-till or direct seeding can be described as seeding directly into the crop stubble from the previous season without use of tillage. A reduction in tillage can result in many benefits, including increased soil organic matter, increased water holding capacity, and reduced fuel costs. However, the effect of no-till and reduced tillage on crop root disease profiles is poorly understood. To study the effect of tillage on disease dynamics, soil samples were collected from commercial wheat fields representing a wide range of tillage strategies in fall 2016 and fall 2017. Because precipitation might affect soilborne diseases, wheat fields located across a diverse gradient of precipitation zones of the dryland Pacific Northwest were selected. Fusarium spp., Pythium spp., and Rhizoctonia spp. were quantified from soil samples using soil dilution plating and quantitative PCR (qPCR) assays. Results of dilution plating showed that the colony counts of Fusarium, Pythium, and Rhizoctonia at the genus level were negatively associated with tillage. However, the same patterns were not observed when specific causal agents of Fusarium, Pythium, and Rhizoctonia that are known to be pathogenic on wheat were quantified with qPCR. Furthermore, precipitation affected the population density of some fungal pathogens (F. culmorum, P. ultimum, and R. solani AG 8). Within the scope of inference of this study, results of this study indicate that the benefits of adopting reduced tillage likely outweigh potential risk for increased root disease.

Variable competitive effects of fungicide resistance in field experiments with a plant pathogenic fungus.

Classic evolutionary theory suggests that mutations associated with antimicrobial and pesticide resistance result in a fitness cost in the absence of the selective antimicrobial agent or pesticide. There is experimental evidence to support fitness costs associated with resistance to anti-microbial compounds and pesticides across many biological disciplines, including human pathology, entomology, plant sciences, and plant pathology. However, researchers have also found examples of neutral and increased fitness associated with resistance, where the effect of a given resistance mutation depends on environmental and biological factors. We used Zymoseptoria tritici, a model evolutionary plant pathogenic fungus, to compare the competitive ability of fungicide-resistant isolates to fungicide-sensitive isolates. We conducted four large-scale inoculated winter wheat experiments at Oregon State University agriculture experiment stations. We found a significant change in the frequency of fungicide resistance over time in all four experiments. The direction and magnitude of these changes, however, differed by experimental location, year of experiment, and inoculum resistance treatment (fungicide-resistant, resistant/sensitive mixture, and fungicide-sensitive). These results suggest that the competitive ability of resistant isolates relative to sensitive isolates varied depending upon environmental conditions, including the initial frequency of resistant individuals in the population.

Temporal Dynamics and Spatial Variation of Azoxystrobin and Propiconazole Resistance in Zymoseptoria tritici: A Hierarchical Survey of Commercial Winter Wheat Fields in the Willamette Valley, Oregon.

Fungicide resistance can cause disease control failure in agricultural systems, and is particularly concerning with Zymoseptoria tritici, the causal agent of Septoria tritici blotch of wheat. In North America, the first quinone outside inhibitor resistance in Z. tritici was discovered in the Willamette Valley of Oregon in 2012, which prompted this hierarchical survey of commercial winter wheat fields to monitor azoxystrobin- and propiconazole-resistant Z. tritici. Surveys were conducted in June 2014, January 2015, May 2015, and January 2016. The survey was organized in a hierarchical scheme: regions within the Willamette Valley, fields within the region, transects within the field, and samples within the transect. Overall, frequency of azoxystrobin-resistant isolates increased from 63 to 93% from June 2014 to January 2016. Resistance to azoxystrobin increased over time even within fields receiving no strobilurin applications. Propiconazole sensitivity varied over the course of the study but, overall, did not significantly change. Sensitivity to both fungicides showed no regional aggregation within the Willamette Valley. Greater than 80% of spatial variation in fungicide sensitivity was at the smallest hierarchical scale (within the transect) of the survey for both fungicides, and the resistance phenotypes were randomly distributed within sampled fields. Results suggest a need for a better understanding of the dynamics of fungicide resistance at the landscape level.

Reduced Virulence of Azoxystrobin-Resistant Zymoseptoria tritici Populations in Greenhouse Assays.

The development of resistance to multiple fungicide classes is currently limiting disease management options for many pathogens, while the discovery of new fungicide classes may become less frequent. In light of this, more research is needed to quantify virulence trade-offs of fungicide resistance in order to more fully understand the implications of fungicide resistance on pathogen fitness. The purpose of this study was to measure the virulence of azoxystrobin-resistant and -sensitive Zymoseptoria tritici populations collected from North and South Willamette Valley, Oregon, in 2012 and 2015. Inoculum mixtures of known fungicide-resistant phenotypes were used to simulate natural field conditions, where multiple genotypes exist and interact in close proximity. Six greenhouse inoculations were conducted over 2 years, and virulence of the isolate mixtures was evaluated in planta. We considered virulence to be "the degree of pathology caused by the organism" and visually estimated the percent area of leaf necrosis as a measure of virulence. In greenhouse conditions, a consistent association of reduced virulence with azoxystrobin-resistant Z. tritici isolate mixtures was observed. North Willamette Valley and South Willamette Valley populations did not differ in virulence.

An Improved Method for Measuring Quantitative Resistance to the Wheat Pathogen Zymoseptoria tritici Using High-Throughput Automated Image Analysis.

Zymoseptoria tritici causes Septoria tritici blotch (STB) on wheat. An improved method of quantifying STB symptoms was developed based on automated analysis of diseased leaf images made using a flatbed scanner. Naturally infected leaves (n = 949) sampled from fungicide-treated field plots comprising 39 wheat cultivars grown in Switzerland and 9 recombinant inbred lines (RIL) grown in Oregon were included in these analyses. Measures of quantitative resistance were percent leaf area covered by lesions, pycnidia size and gray value, and pycnidia density per leaf and lesion. These measures were obtained automatically with a batch-processing macro utilizing the image-processing software ImageJ. All phenotypes in both locations showed a continuous distribution, as expected for a quantitative trait. The trait distributions at both sites were largely overlapping even though the field and host environments were quite different. Cultivars and RILs could be assigned to two or more statistically different groups for each measured phenotype. Traditional visual assessments of field resistance were highly correlated with quantitative resistance measures based on image analysis for the Oregon RILs. These results show that automated image analysis provides a promising tool for assessing quantitative resistance to Z. tritici under field conditions.