RESUMO
Since 2018, bacterial-like symptoms, such as leaf streaks were observed on wheat plants (Triticum aestivum L.) in Córdoba province in Argentina, with 1 to 5% of disease incidence. Samples of wheat stem and spike collected in a trial of varieties for summer/autumn sowing in the experimental field of the INTA Marcos Juárez were disinfected, washed and macerated in mortars with sterile distilled water and extracts were streaked on Luria-Bertani (LB) agar. After 48 h incubation at 28 °C, circular, mucoid, convex, and cream colonies were observed and pure cultures were transferred to LB medium for further identification tests. Biochemical tests corroborated the detection of a Gram-negative bacillus. Conventional PCR was performed using DNA isolate from pure cultures and general primers for various species of genera Xanthomonas (Maes 1993) and Pseudomonas (Mulet et al. 2010). An isolate (Arg-1), with cream colored colonies was positive using general primers for Xanthomonas sp (amplified fragment of 444 bp). A bacterial suspension containing 108 CFU mL-1 grown for 48 h on LB medium at 28 °C was injected into three-week-old leaves of wheat plants to fulfill Koch's postulates. After 5 days, plants showed symptoms of chlorosis, streaks and then necrosis on the leaves. The bacteria were re-isolated from the inoculated plants, showing same symptoms observed in the original plants. Negative control plants, inoculated with sterile water remained without symptoms. The amplified 444 bp fragment described above was sequenced by the Sanger method (GenBank accession OM972662), as well as another 757 bp fragment amplified with universal primers that amplify the partial 16S rDNA gene (GenBank accession OM972661). Analyses of these sequences, as well as the protein profile of the isolate obtained by matrix assisted laser desorption/ionization time of-flight mass spectrometry (MALDI-TOF MS) Bruker Biotyper, allowed to identify only the genus Xanthomonas. With the purpose of determine the species status, the complete genome of isolate Arg-1 was sequenced using Oxford Nanopore Technologies (ONT). Total gDNA was isolate from pure cultures using a commercial kit (Wizard Genomic DNA Purification Kit, Promega). gDNA library was constructed using Ligation Sequencing Kit (SQK-LSK109) and sequenced using ONT platform on a MinION 1kb device. Raw basecalled sequences were filtered using Filtlong and assembled using Trycycler. The genome was assembled in a single contig comprising 5.410.641 bp with 4740 predicted CDSs and 63.9% GC content. Genome sequence was deposited in GenBank under accession number CP094827 and SRA data SRX14635308. Whole-genome Average Nucleotide Identity (ANI) analysis showed values of ~ 97% against the reference genomes of Xanthomonas prunicola (PHKX01.1, PHKV01.1 and PHKW01.1) and 100% in complete 16S rRNA gene sequences (1547 bp). These findings suggest that a new wheat pathogen within the genus Xanthomonas is present in Argentina, as well as was reported in Uruguay and USA (Clavijo et al. 2021). To our knowledge, this is the first report of X. prunicola affecting wheat in Argentina and the first complete genome registered for this specie. Accurate and specific diagnostics are required for the detection of X. prunicola in wheat crops to implement correct prevention and control strategies to this disease, avoiding the dissemination in lots where it has not yet been found.
RESUMO
BACKGROUND: In breeding programs, the selection of cultivars with the highest yield potential consisted in the selection of the yield per se, which resulted in cultivars with higher grains per spike (GN) and occasionally increased grain weight (GW) (main numerical components of the yield). In this study, quantitative trait loci (QTL) for GW, GN and spike fertility traits related to GN determination were mapped using two doubled haploid (DH) populations (Baguette Premium 11 × BioINTA 2002 and Baguette 19 × BioINTA 2002). RESULTS: In total 305 QTL were identified for 14 traits, out of which 12 QTL were identified in more than three environments and explained more than 10% of the phenotypic variation in at least one environment. Eight hotspot regions were detected on chromosomes 1A, 2B, 3A, 5A, 5B, 7A and 7B in which at least two major and stable QTL sheared confidence intervals. QTL on two of these regions (R5A.1 and R5A.2) have previously been described, but the other six regions are novel. CONCLUSIONS: Based on the pleiotropic analysis within a robust physiological model we conclude that two hotspot genomic regions (R5A.1 and R5A.2) together with the QGW.perg-6B are of high relevance to be used in marker assisted selection in order to improve the spike yield potential. All the QTL identified for the spike related traits are the first step to search for their candidate genes, which will allow their better manipulation in the future.