Improving the Comprehension of Pathogenicity and Phylogeny in 'Candidatus Phytoplasma meliae' through Genome Characterization.

'Candidatus Phytoplasma meliae' is a pathogen associated with chinaberry yellowing disease, which has become a major phytosanitary problem for chinaberry forestry production in Argentina. Despite its economic impact, no genome information of this phytoplasma has been published, which has hindered its characterization at the genomic level. In this study, we used a metagenomics approach to analyze the draft genome of the 'Ca. P. meliae' strain ChTYXIII. The draft assembly consisted of twenty-one contigs with a total length of 751.949 bp, and annotation revealed 669 CDSs, 34 tRNAs, and 1 set of rRNA operons. The metabolic pathways analysis showed that ChTYXIII contains the complete core genes for glycolysis and a functional Sec system for protein translocation. Our phylogenomic analysis based on 133 single-copy genes and genome-to-genome metrics supports the classification as unique 'Ca. P. species' within the MPV clade. We also identified 31 putative effectors, including a homolog to SAP11 and others that have only been described in this pathogen. Our ortholog analysis revealed 37 PMU core genes in the genome of 'Ca. P. meliae' ChTYXIII, leading to the identification of 2 intact PMUs. Our work provides important genomic information for 'Ca. P. meliae' and others phytoplasmas for the 16SrXIII (MPV) group.

First report of Xanthomonas prunicola causing bacterial leaf streaks on wheat in Argentina.

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.

Draft Genome Sequence of "Candidatus Phytoplasma pruni" (X-Disease Group, Subgroup 16SrIII-B) Strain ChTDIII from Argentina.

Herein, we report the draft genome sequence of "Candidatus Phytoplasma pruni" strain ChTDIII (subgroup 16SrIII-B). The final assembly consists of 790,517 nucleotides organized in 67 contigs (minimal size, 1 kb), with a G+C content of 29.4% and encoding 672 proteins.

Description of 'Candidatus Phytoplasma meliae', a phytoplasma associated with Chinaberry (Melia azedarach L.) yellowing in South America.

China tree yellows (ChTY) phytoplasma is associated with the yellowing disease of the China tree (Melia azedarach) in Argentina. According to partial 16S rRNA gene analysis, ChTY phytoplasma belongs to the 16Sr XIII group, subgroup G. Strains of species of ChTY have 98-99 % 16S rDNA gene sequence similarity with 16SrXIII-group phytoplasmas, and less than 97.5 % when compared to all 'CandidatusPhytoplasma' described so far, except for the novel 'CandidatusPhytoplasma hispanicum'. However, strains of species of ChTY are differentiated from the latter due to having additional molecular and biological attributes. The presence of unique features in the 16S rDNA sequence distinguishes ChTY from all species of 'CandidatusPhytoplasma' currently described. The in silico RFLP profile of 16S rDNA (1.2 kb) and rpLV-rpsC (1.3 kb) genes distinguished ChTY, as in the 16SrXIII-G subgroup within the 16SrXIIII group. The phylogenetic analyses, based on 16S rDNA, rpLV-rpsC and secA gene sequences, in addition to the restricted host range, characteristic symptoms and geographical distribution, confirm that the collective strains of the species ChTY represent a distinct lineage within the phytoplasma clade and support the description of a novel species of 'CandidatusPhytoplasma meliae' with the reference strain being ChTY-Mo3 (Montecarlo, Argentina).

Viabilidade da antecipação do tratamento de sementes de arroz com inseticidas em relação à data de semeadura no controle de Oryzophagus oryzae (Coleoptera: Curculionidae)

Oryzophagus oryzae (Costa Lima, 1936) is one of the most important pests of irrigated rice, whose control is made traditionally with granulated insecticides on the irrigation water. However, the use of insecticides on the seeds has been considered a promising method. Although there are adequate implements to apply insecticides on the seeds, its hard to make this treatment with quality and in a big quantity, few days or hours before sowing. In this context, the viability of treat the rice seeds in an advance of sowing date has been evaluated. Two experiments were performed at Embrapa Clima Temperado, Capão do Leão, RS, Brazil in 1997/98 and 2004/05. The experimental design was in blocks at random with four repetitions, being evaluated the insecticides carbosulfan and fipronil applied manually to the seeds of rice in different times that preceded the sowing. Germination power and vigour, emerged plants, larval population and grains productivity were considered. The rice seeds treatment with insecticides in advance of sowing date trying to achieve the control of O. oryzae, demonstrate viability. The insecticides carbosulfan and fipronil are adequate to advance seeds treatment, if applied until 75 days before sowing.

Oryzophagus oryzae (Costa Lima, 1936) é uma das pragas mais prejudiciais da cultura do arroz irrigado, sendo seu controle efetuado tradicionalmente com inseticidas granulados aplicados na água de irrigação. No entanto, o uso de inseticidas aplicados às sementes tem sido considerado promissor. Embora existam máquinas adequadas para aplicação de inseticidas às sementes, há dificuldade de assegurar a qualidade das sementes tratadas, em poucos dias ou mesmo horas antes da semeadura. Assim, objetivou-se avaliar a viabilidade do tratamento de sementes de arroz com inseticidas aplicados com antecedência à data de semeadura para o controle de O. oryzae. Dois experimentos foram instalados na Estação Experimental de Terras Baixas da Embrapa Clima Temperado, localizada no município de Capão do Leão, Rio Grande do Sul (RS), em duas épocas (1997/1998 e 2004/2005). O delineamento experimental foi em blocos ao acaso com quatro repetições. Foram avaliados os inseticidas carbosulfano e fipronil aplicados manualmente às sementes de arroz em diferentes épocas que antecederam a semeadura e foram avaliadas as variáveis poder germinativo e vigor das sementes, plantas emergidas, população larval e produtividade de grãos. O tratamento das sementes de arroz com inseticidas, antecipadamente à data de semeadura, visando ao controle de O. oryzae, demonstrou ser viável. Os inseticidas carbosulfano e fipronil são adequados ao tratamento antecipado das sementes, se aplicados até 75 dias antes da semeadura.

Viabilidade da antecipação do tratamento de sementes de arroz com inseticidas em relação à data de semeadura no controle de Oryzophagus oryzae (Coleoptera: Curculionidae)

Oryzophagus oryzae (Costa Lima, 1936) is one of the most important pests of irrigated rice, whose control is made traditionally with granulated insecticides on the irrigation water. However, the use of insecticides on the seeds has been considered a promising method. Although there are adequate implements to apply insecticides on the seeds, it’s hard to make this treatment with quality and in a big quantity, few days or hours before sowing. In this context, the viability of treat the rice seeds in an advance of sowing date has been evaluated. Two experiments were performed at Embrapa Clima Temperado, Capão do Leão, RS, Brazil in 1997/98 and 2004/05. The experimental design was in blocks at random with four repetitions, being evaluated the insecticides carbosulfan and fipronil applied manually to the seeds of rice in different times that preceded the sowing. Germination power and vigour, emerged plants, larval population and grains productivity were considered. The rice seeds treatment with insecticides in advance of sowing date trying to achieve the control of O. oryzae, demonstrate viability. The insecticides carbosulfan and fipronil are adequate to advance seeds treatment, if applied until 75 days before sowing.

Oryzophagus oryzae (Costa Lima, 1936) é uma das pragas mais prejudiciais da cultura do arroz irrigado, sendo seu controle efetuado tradicionalmente com inseticidas granulados aplicados na água de irrigação. No entanto, o uso de inseticidas aplicados às sementes tem sido considerado promissor. Embora existam máquinas adequadas para aplicação de inseticidas às sementes, há dificuldade de assegurar a qualidade das sementes tratadas, em poucos dias ou mesmo horas antes da semeadura. Assim, objetivou-se avaliar a viabilidade do tratamento de sementes de arroz com inseticidas aplicados com antecedência à data de semeadura para o controle de O. oryzae. Dois experimentos foram instalados na Estação Experimental de Terras Baixas da Embrapa Clima Temperado, localizada no município de Capão do Leão, Rio Grande do Sul (RS), em duas épocas (1997/1998 e 2004/2005). O delineamento experimental foi em blocos ao acaso com quatro repetições. Foram avaliados os inseticidas carbosulfano e fipronil aplicados manualmente às sementes de arroz em diferentes épocas que antecederam a semeadura e foram avaliadas as variáveis poder germinativo e vigor das sementes, plantas emergidas, população larval e produtividade de grãos. O tratamento das sementes de arroz com inseticidas, antecipadamente à data de semeadura, visando ao controle de O. oryzae, demonstrou ser viável. Os inseticidas carbosulfano e fipronil são adequados ao tratamento antecipado das sementes, se aplicados até 75 dias antes da semeadura.

Plataformas de colheita e colheita manual com trilha mecânica sobre a qualidade de sementes de arroz ( Oryza sativa, L. )

During irrigated rice harvesting occur losses and physical and phisiological seed damage. Late 80's, appeared the strippers headers that strip the grain, instead of cutting the spike. However, little is know about physical and phisiological seed damage by harvest header. The objective of this work was to evaluate the mechanical damage caused to BR-IRGA 409 and BR-IRGA 410 rice cultivars by three harvesting methods: (a) manual harvesting and mechanical strip; (b) cutterbar harvesting and; (c) stripper header harvesting. Samples were collected directly in the grain tank when the harvest was mechanical. The experimental design was randomized blocks with six replications. Results demonstrated that the rice seeds of the studied variety didn't showed significant differences in physical and phisiological in seed quality when harvested by cutterbar or stripper header. These two harvesting forms, however, showed significantly seed damages when compared to manual harvesting and mechanical strip.

Durante a colheita do arroz irrigado ocorrem perdas e danos físicos e fisiológicos às sementes. No final da década de oitenta, surgiram as plataformas recolhedoras, que retiram ou arrancam o grão ao invés de cortar a panícula, porém, pouco se conhece a respeito dos danos físicos e fisiológicos que este sistema de plataforma pode causar às sementes. Este trabalho teve por objetivo avaliar os danos mecânicos causados às sementes dos cultivares de arroz BR-IRGA 409 e BR-IRGA 410, por três formas de colheita: (a) colheita manual e trilha mecânica; (b) colheita com plataforma de corte; (c) colheita com plataforma recolhedora. Quando a colheita foi mecânica, realizou-se a coleta das amostras diretamente no graneleiro. O delineamento experimental foi blocos ao acaso, com seis repetições. Os resultados demonstraram que as sementes de arroz irrigado dos cultivares estudados não apresentaram diferenças significativas em suas qualidades físicas e fisiológicas, quando colhidas com plataforma de corte e com a plataforma recolhedora. Estes dois métodos de colheita, porém, apresentaram danos significativamente maiores quando comparados à colheita manual e trilha mecânica.

Plataformas de colheita e colheita manual com trilha mecânica sobre a qualidade de sementes de arroz ( Oryza sativa, L. )

During irrigated rice harvesting occur losses and physical and phisiological seed damage. Late 80's, appeared the strippers headers that strip the grain, instead of cutting the spike. However, little is know about physical and phisiological seed damage by harvest header. The objective of this work was to evaluate the mechanical damage caused to BR-IRGA 409 and BR-IRGA 410 rice cultivars by three harvesting methods: (a) manual harvesting and mechanical strip; (b) cutterbar harvesting and; (c) stripper header harvesting. Samples were collected directly in the grain tank when the harvest was mechanical. The experimental design was randomized blocks with six replications. Results demonstrated that the rice seeds of the studied variety didn't showed significant differences in physical and phisiological in seed quality when harvested by cutterbar or stripper header. These two harvesting forms, however, showed significantly seed damages when compared to manual harvesting and mechanical strip.

Durante a colheita do arroz irrigado ocorrem perdas e danos físicos e fisiológicos às sementes. No final da década de oitenta, surgiram as plataformas recolhedoras, que retiram ou arrancam o grão ao invés de cortar a panícula, porém, pouco se conhece a respeito dos danos físicos e fisiológicos que este sistema de plataforma pode causar às sementes. Este trabalho teve por objetivo avaliar os danos mecânicos causados às sementes dos cultivares de arroz BR-IRGA 409 e BR-IRGA 410, por três formas de colheita: (a) colheita manual e trilha mecânica; (b) colheita com plataforma de corte; (c) colheita com plataforma recolhedora. Quando a colheita foi mecânica, realizou-se a coleta das amostras diretamente no graneleiro. O delineamento experimental foi blocos ao acaso, com seis repetições. Os resultados demonstraram que as sementes de arroz irrigado dos cultivares estudados não apresentaram diferenças significativas em suas qualidades físicas e fisiológicas, quando colhidas com plataforma de corte e com a plataforma recolhedora. Estes dois métodos de colheita, porém, apresentaram danos significativamente maiores quando comparados à colheita manual e trilha mecânica.