Nanopore Sequencing with GraphMap for Comprehensive Pathogen Detection in Potato Field Soil.

Early detection of causal pathogens is important to prevent crop loss from diseases. However, some diseases, such as soilborne diseases, are difficult to diagnose due to the absence of visible or characteristic symptoms. In the present study, the use of the Oxford Nanopore MinION sequencer as a molecular diagnostic tool was assessed due to its long-read sequencing capabilities and portability. Nucleotide samples (DNA or RNA) from potato field soils were sequenced and analyzed using a locally curated pathogen database, followed by identification via sequence mapping. We performed computational speed tests of three commonly used mapping/annotation tools (BLAST, BWA-BLAST, and BWA-GraphMap) and found BWA-GraphMap to be the fastest tool for local searching against our curated pathogen database. The data collected demonstrate the high potential of Nanopore sequencing as a minimally biased diagnostic tool for comprehensive pathogen detection in soil from potato fields. Our GraphMap-based MinION sequencing method could be useful as a predictive approach for disease management by identifying pathogens present in field soil prior to planting. Although this method still needs further experimentation with a larger sample size for practical use, the data analysis pipeline presented can be applied to other cropping systems and diagnostics for detecting multiple pathogens.

Sorghum Sheath Blight Caused by Fusarium spp. in Sinaloa, Mexico.

Sorghum (Sorghum bicolor) leaf sheath blight was observed for the first time in Sinaloa, Mexico in the summer of 2020. Fungal isolates were obtained from symptomatic tissue in potato dextrose agar. Fusarium spp. were associated with symptomatic plants in 10 sampling sites under field conditions. No root and stalk rot was observed during the sampling period. Analysis of fragments of the translation elongation factor alpha and RNA polymerase II second largest subunit genes indicated that all isolates belong to the Fusarium fujikuroi species complex (FFSC). Five groups were delineated from this complex: F. thapsinum, F. verticillioides, Fusarium sp. (four isolates), Fusarium sp. 4 (Fus4), and Fusarium sp. (Fus16), which is closely related to Fusarium madaense. The morphological characteristics (colony color and morphometry of conidia) of isolates with sequence similarities to those of F. thapsinum and F. verticillioides were in the expected range for these species. The morphology of isolates Fus7a, Fus7b, Fus11, and Fus17, as well as Fus4 and Fus16, were similar to those of the FFSC, specially to F. andiyazi and F. madaense. Inoculations of sorghum with representative isolates of F. thapsinum, F. verticillioides and the unidentified Fusarium species resulted in reddish brown lesions similar to those observed under field conditions; the original isolates inoculated were reisolated fulfilling the Koch's postulates. Although leaf sheaths on sorghum plants were heavily damaged, root and stalk rot were not observed in the greenhouse inoculations or under field conditions. Future research should focus on determining the identity of the unknown Fusarium spp. to design control measures for the disease. This is the first report of Fusarium spp. causing sorghum leaf sheath blight in Mexico.

PvLOX2 silencing in common bean roots impairs arbuscular mycorrhiza-induced resistance without affecting symbiosis establishment.

The arbuscular mycorrhizal (AM) symbiosis is an intimate association between specific soil-borne fungi and the roots of most land plants. AM colonisation elicits an enhanced defence resistance against pathogens, known as mycorrhizal-induced resistance (MIR). This mechanism locally and systemically sensitises plant tissues to boost their basal defence response. Although a role for oxylipins in MIR has been proposed, it has not yet been experimentally confirmed. In this study, when the common bean (Phaseolus vulgaris L.) lipoxygenase PvLOX2 was silenced in roots of composite plants, leaves of silenced plants lost their capacity to exhibit MIR against the foliar pathogen Sclerotinia sclerotiorum, even though they were colonised normally. PvLOX6, a LOX gene family member, is involved in JA biosynthesis in the common bean. Downregulation of PvLOX2 and PvLOX6 in leaves of PvLOX2 root-silenced plants coincides with the loss of MIR, suggesting that these genes could be involved in the onset and spreading of the mycorrhiza-induced defence response.