First Report of bacterial speck caused by Pseudomonas syringae pv. tomato Race 1 affecting tomato in different Regions of Chile

In Chile, tomato is one of the most widely cultivated vegetables, with around 5,000 ha for fresh market and 8,000 ha for processing industry. During recent years, symptoms of bacterial speck caused by Pseudomonas syringae pv. tomato, have been observed more frequently in tomato plants in different regions of Chile. This pathogen was first identified in Chile in 1987 (Latorre & Lolas, 1988) and the presence of an apparent new variant was reported in 2004 (Besoain et al. 2004). To characterize the pathogen that was affecting this crop, samples of diseased tomato plants were taken in three regions of Chile. The samples were collected in 2016 in Northern Chile in Lluta Valley from the Arica y Parinacota Region, and in Central Chile, in 2014 in Limache from Valparaíso Region and in 2015 in Pichidegua from O´Higgins Region. Affected tomato plants exhibited dark brown to black lesions surrounded by yellow halos in the leaves, and dark brown to black lesions in the stems, pedicels, and peduncles. Plants tissues were macerated, and the suspension was spread on King’s B medium, resulting in fluorescent colonies visualized under 366 nm UV light. LOPAT tests results of three selected isolates from different Regions, were: levan production (+), oxidase reaction (-), potato soft rot (-), arginine dihydrolase production (-), and tobacco hypersensitivity (+) (Lelliot et al. 1966). Molecular identification was carried out by amplification and sequence analysis of housekeeping genes cts, encoding citrate synthase, gyrB, encoding DNA gyrase B, and rpoD, encoding sigma factor 70 (Hwang et al. 2005; Sarkar & Guttmann 2004) (GenBank Accessions No. OK001658-OK001666). BLAST analysis of cts and rpoD genes of the three isolates resulted in a match with a 100% identity (919 bp and 491 bp respectively) with Pseudomonas syringae pv. tomato strain B13-200 (GenBank: CP019871.1). BLAST analysis of gyrB gene of two isolates resulted in a match with a 100% identity (684 bp) and one isolate with 99.85% (683 bp) with Pseudomonas syringae pv. tomato strain B13-200. To identify the race 1, each strain was inoculated in five tomato plants cv. San Pedro, susceptible to both races of P. syringae pv. tomato, and cv. Rio Grande, resistant to race 0. The tomato plants were slightly wounded with a metal sponge and then sprayed with the bacterial suspension (108 CFU mL-1) of each isolate, including the reference strain DC3000 (race 0). Negative controls were sprayed with water. The plants inoculated with Chilean strains in both cv. San Pedro and cv. Rio Grande, showed symptoms of bacterial speck after 7 days. Plants inoculated with DC3000 strain showed symptoms only in cv. San Pedro, whereas control plants remained asymptomatic. Strains were re-isolated from symptomatic plants and identified by gene sequence analyses as Pseudomonas syryngae pv. tomato. This is the first report of Pseudomonas syryngae pv. tomato race 1 in Chile. Race 1 was previously reported in Canada (Lawton and MacNeill. 1986), in Italy (Buonaurio et al. 1996), in California (Arredondo and Davis 2000), in Portugal (Cruz et al. 2010), and in other states in the USA and countries in South America, Europe, Africa, and Australia, becoming the most commonly isolated race today (Cai et al 2011). These results will be the base for future studies of epidemiology, characterization, and virulence in order to explain the outbreak of this disease and the severity of symptoms observed.

A Green Nanostructured Pesticide to Control Tomato Bacterial Speck Disease

Bacterial speck disease, caused by Pseudomonas syringae pv. tomato (Pst), is one of the most pervasive biological adversities in tomato cultivation, in both industrial and in table varieties. In this work synthesis, biochemical and antibacterial properties of a novel organic nanostructured pesticide composed of chitosan hydrochloride (CH) as active ingredient, cellulose nanocrystals (CNC) as nanocarriers and starch as excipient were evaluated. In order to study the possibility of delivering CH, the effects of two different types of starches, extracted from a high amylose bread wheat (high amylose starch—HA Starch) and from a control genotype (standard starch—St Starch), were investigated. Nanostructured microparticles (NMP) were obtained through the spray-drying technique, revealing a CH loading capacity proximal to 50%, with a CH release of 30% for CH-CNC-St Starch NMP and 50% for CH-CNC-HA Starch NMP after 24 h. Both NMP were able to inhibit bacterial growth in vitro when used at 1% w/v. Moreover, no negative effects on vegetative growth were recorded when NMP were foliar applied on tomato plants. Proposed nanostructured pesticides showed the capability of diminishing Pst epiphytical survival during time, decreasing disease incidence and severity (from 45% to 49%), with results comparable to one of the most used cupric salt (hydroxide), pointing out the potential use of CH-CNC-Starch NMP as a sustainable and innovative ally in Pst control strategies.

Gene Based ­­­­Markers in Marker-Assisted Selection to Screen Tomato Genotypes Resistant to Fusarium Wilt, Late Blight, Verticillium Wilt, Leaf Mold, Bacterial Spot and Bacterial Speck

The authors have requested that this preprint be withdrawn due to author disagreement.

Gene Based ­­­­Markers in Marker-Assisted Selection to Screen Tomato Genotypes Resistant to Fusarium Wilt, Late Blight, Verticillium Wilt, Leaf Mold, Bacterial Spot and Bacterial Speck

The tomato crop is exposed to serious losses due to infection with several diseases and pests, which threaten tomato production in Egypt and worldwide. Therefore, selecting the tomato germplasm resistant or tolerant to a specific pathogen by molecular markers closely linked to resistance loci is a desirable goal of this study. In this work, seven co-dominant markers targeting six resistance genes (I-1, Ve, Ph3, Cf-9/Cf-4, Rx4, and Pto) for six main diseases [ fusarium wilt (Fusarium oxysporum f. sp. lycopersici), verticillium wilt (Verticillium dahliae and V. alboatrum), late blight (Phytophthora infestans), leaf mold (Cladosporium fulvum), bacterial spot (Xanthomonas campestris pv. vesicatoria) and bacterial speck (Pseudomonas syringae pv. tomato)], respectively were determined. Theses molecular markers differentiated among 19 tomato genotypes resistant (homozygote/heterozygote) and susceptible (homozygote) to the pathogens. Therefore, this study supplied us with novel tomato lines with resistance to multiple diseases, and their pyramiding inside domesticated tomato cultivars are suggested to apply in the tomato breeding programs of resistance against fungal and bacterial diseases.

Draft Genome Sequences of Pseudomonas syringae pv. tomato Strains J4 and J6, Isolated in Florida

ABSTRACT Pseudomonas syringae pv. tomato causes bacterial speck in tomato. We report the genome sequences of two P. syringae pv. tomato strains, J4 and J6, that are genetically closely related, with >99.9 average nucleotide identity (ANI), but vary in the presence of coronatine-associated genes.

The Effect of Pesticides on the Tomato Bacterial Speck Disease Pathogen Pseudomonas Syringae pv. Tomato

A significant part of the used pesticides does not reach the target organisms and, while remaining in the agrophytocenosis, influences all living organisms in it. Having a toxic and often mutagenic effect, pesticides induce morphological and physiological changes in the cells of microorganisms and are the cause of phenotypic heterogeneity of their populations. However, the effect of pesticides on phytopathogenic bacteria as non-target microorganisms remains out of the field of view for most researchers. However, the use of pesticides can lead to expansion of the diversity of existing phytopathogens and, as a consequence, complications of identification of the pathogens, loss of resistance by plants varieties, and increased harm from diseases caused by them. This study is focused on the effect of pesticides used in tomato plantations on the causative agent of bacterial speck of this crop—Pseudomonas syringae pv. tomato. The studies were carried out using the methods of classical microbiology. The mutagenic action of pesticides was recorded, taking into account the increase of the number of streptomycin resistance mutations in bacteria in the case of pesticide action. It is established that the fungicide aluminium phosethyl is characterised by a bacteriostatic effect on P. syringae pv. tomato. Deltamethrin insecticide does not affect the growth of P. syringae pv. tomato. However, there is an increase in the frequency of streptomycin resistance mutations in both studied strains of P. syringae pv. tomato after using deltamethrin. It is shown that the frequency of occurrence of R (rough colonies) forms of P. syringae pv. tomato IZ28 and IZ46 after using deltamethrin increased by 100 times when in comparison to the frequency of spontaneous morphological dissociation, or smooth-to-rough (S-R) mutation, of these bacteria. Therefore, aluminium phosethyl is characterised by moderate bacteriostatic action against P. syringae pv. tomato. Deltamethrin does not influence the growth of the pathogen of tomato speck but increases the frequency of formation of StrR mutants and R forms of phytopathogenic bacteria.