Thiosulfinate Tolerance Gene Clusters Are Common Features of Burkholderia Onion Pathogens.

Burkholderia gladioli pv. alliicola, B. cepacia, and B. orbicola are common bacterial pathogens of onion. Onions produce organosulfur thiosulfinate defensive compounds after cellular decompartmentalization. Using whole-genome sequencing and in silico analysis, we identified putative thiosulfinate tolerance gene (TTG) clusters in multiple onion-associated Burkholderia species similar to those characterized in other Allium-associated bacterial endophytes and pathogens. Sequence analysis revealed the presence of three Burkholderia TTG cluster types, with both Type A and Type B being broadly distributed in B. gladioli, B. cepacia, and B. orbicola in both the chromosome and plasmids. Based on isolate natural variation and generation of isogenic strains, we determined the in vitro and in vivo contribution of TTG clusters in B. gladioli, B. cepacia, and B. orbicola. The Burkholderia TTG clusters contributed to enhanced allicin tolerance and improved growth in filtered onion extracts by all three species. TTG clusters also made clear contributions to B. gladioli foliar necrosis symptoms and bacterial populations. Surprisingly, the TTG cluster did not contribute to bacterial populations in onion bulb scales by these three species. Based on our findings, we hypothesize onion-associated Burkholderia may evade or inhibit the production of thiosulfinates in onion bulb tissues. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

A multi-provincial outbreak of Salmonella Newport infections associated with red onions: A report of the largest Salmonella outbreak in Canada in over 20 years.

An investigation into an outbreak of Salmonella Newport infections in Canada was initiated in July 2020. Cases were identified across several provinces through whole-genome sequencing (WGS). Exposure data were gathered through case interviews. Traceback investigations were conducted using receipts, invoices, import documentation, and menus. A total of 515 cases were identified in seven provinces, related by 0-6 whole-genome multi-locus sequence typing (wgMLST) allele differences. The median age of cases was 40 (range 1-100), 54% were female, 19% were hospitalized, and three deaths were reported. Forty-eight location-specific case sub-clusters were identified in restaurants, grocery stores, and congregate living facilities. Of the 414 cases with exposure information available, 71% (295) had reported eating onions the week prior to becoming ill, and 80% of those cases who reported eating onions, reported red onion specifically. The traceback investigation identified red onions from Grower A in California, USA, as the likely source of the outbreak, and the first of many food recall warnings was issued on 30 July 2020. Salmonella was not detected in any tested food or environmental samples. This paper summarizes the collaborative efforts undertaken to investigate and control the largest Salmonella outbreak in Canada in over 20 years.

Identification and functional characterization of the npc-2-like domain containing rust effector protein that suppresses cell death in plants.

The MD-2-related lipid-recognition (ML/Md-2) domain is a lipid/sterol-binding domain that are involved in sterol transfer and innate immunity in eukaryotes. Here we report a genome-wide survey of this family, identifying 84 genes in 30 fungi including plant pathogens. All the studied species were found to have varied ML numbers, and expansion of the family was observed in Rhizophagus irregularis (RI) with 33 genes. The molecular docking studies of these proteins with cholesterol derivatives indicate lipid-binding functional conservation across the animal and fungi kingdom. The phylogenetic studies among eukaryotic ML proteins showed that Puccinia ML members are more closely associated with animal (insect) npc2 proteins than other fungal ML members. One of the candidates from leaf rust fungus Puccinia triticina, Pt5643 was PCR amplified and further characterized using various studies such as qRT-PCR, subcellular localization studies, yeast functional complementation, signal peptide validation, and expression studies. The Pt5643 exhibits the highest expression on the 5th day post-infection (dpi). The confocal microscopy of Pt5643 in onion epidermal cells and N. benthamiana shows its location in the cytoplasm and nucleus. The functional complementation studies of Pt5643 in npc2 mutant yeast showed its functional similarity to the eukaryotic/yeast npc2 gene. Furthermore, the overexpression of Pt5643 also suppressed the BAX, NEP1, and H2O2-induced program cell death in Nicotiana species and yeast. Altogether the present study reports the novel function of ML domain proteins in plant fungal pathogens and their possible role as effector molecules in host defense manipulation.

Identifying and Controlling Anthracnose Caused by Colletotrichum Taxa of Welsh Onion in Sanxing, Taiwan.

Leaves of Welsh onion (Allium fistulosum) are subject to various fungal diseases such as anthracnose (Colletotrichum species) and Stemphylium leaf blight (Stemphylium vesicarium). These diseases are the main biotic limitations to Welsh onion production in northern Taiwan. From 2018 to 2020, anthracnose symptoms were observed throughout Welsh onion fields in northern Taiwan, mainly the Sanxing area. In total, 33 strains of Colletotrichum species were isolated from diseased leaves, and major causative agents were identified based on a multilocus phylogenetic analysis using four genomic regions (act, tub2, gapdh, and internal transcribed spacer). Based on this phylogeny, Colletotrichum species causing anthracnose of Welsh onion were identified as C. spaethianum (C. spaethianum species complex) and C. circinans (C. dematium species complex) in the Sanxing area, northern Taiwan. To determine and compare the pathogenicity of each species, representative fungal strains of each species were inoculated on the cultivar 'Siao-Lyu' by spraying spore suspension onto the leaf surface. Welsh onion plants were susceptible to both species, but disease incidence and severity were higher in C. spaethianum. In total, 31 fungicides were tested to determine their efficacy in reducing mycelial growth and conidial germination of representative strains of C. spaethianum and C. circinans under laboratory conditions. Five fungicides-fluazinam, metiram, mancozeb, thiram, and dithianon-effectively reduced mycelial growth and spore germination in both C. spaethianum and C. circinans. In contrast, difenoconazole and trifloxystrobin + tebuconazole, which are commonly used in Welsh onion production in northern Taiwan, mainly the Sanxing area, were ineffective. These results serve as valuable insights for growers, enabling them to identify and address the emergence of anthracnose caused by C. spaethianum and C. circinans of Welsh onion, employing fungicides with diverse modes of action. The findings of this study support sustainable management of anthracnose in Sanxing, northern Taiwan, although further field tests of the fungicides are warranted.

Rapid Detection of a Downy Mildew Pathogen, Peronospora destructor, in Infected Onion Tissues and Soils by Loop-Mediated Isothermal Amplification.

Downy mildew of onion caused by a soil-inhabiting water mold, Peronospora destructor, is one of the most devastating diseases that can destroy entire onion fields in a matter of days. In this study, we developed a loop-mediated isothermal amplification (LAMP) assay that allows for rapid detection of P. destructor by visual inspection. The internal transcribed spacer 2 region of P. destructor was used to design primer sets for LAMP reactions. The optimal temperature and incubation time were determined for the most efficient primer set. In the optimized condition, the LAMP assay exhibited at least 100 times more sensitivity than conventional PCR, detecting femtogram levels of P. destructor genomic DNA (gDNA). Detection of the pathogen from a small number of spores without gDNA extraction further confirmed the high sensitivity of the assay. For specificity, the LAMP assay was negative for gDNA of other fungal pathogens that cause various diseases on onion and oomycetes, whereas the assay was positive for gDNA extracted from onion tissues showing the typical downy mildew symptoms. Finally, we examined the efficacy of the LAMP assay in detection of P. destructor in soils. Soils collected from onion fields that had been contaminated with P. destructor were solarized for 60 days. Whereas the LAMP assay was negative for the solarized soils, we were able to detect P. destructor that oversummers in fields. The LAMP assay developed in this study enables rapid detection and diagnosis of downy mildew of onion in infected tissues and in soil.

Bosea rubneri sp. nov. Isolated from Organically Grown Allium cepa.

Strain ZW T0_25T was isolated from an onion sample (Allium cepa var. Hytech F1) within a storage trial and proofed to be a novel, aerobic, Gram-stain negative, rod-shaped bacterial strain. Analyses of the 16S rRNA gene sequence and of the whole draft genome sequences, i.e., digital DNA-DNA hybridization (dDDH), Average Nucleotide Identity (ANI) and Average Amino Acid Identity (AAI) showed that this strain represents a new species of the genus Bosea. The genome size of strain ZW T0_25T is 6.19 Mbp, and the GC content is 66.9%. As whole cell sugars, rhamnose, ribose and glucose were identified. Ubiquinone Q-10 is the major respiratory quinone with 97.8%. Polar lipids in strain ZW T0_25T are composed of one phosphatidylethanolamine, one phosphatidylglycerol, one aminophospholipid, two aminolipids, one glycolipid and two phospholipids whereas the fatty acid profile predominantly consists of C18:1 w7c (63.3%), C16:1 w7c (19.5%) and C16:0 (7.1%). Phenotypic traits were tested in the wet lab as well as predicted in silico from genome data. Therefore, according to this polyphasic approach, the new name Bosea rubneri sp. nov. with the type strain ZW T0_25T (= DSM 116094 T = LMG 33093 T) is proposed.

The Genetic Requirements for HiVir-Mediated Onion Necrosis by Pantoea ananatis, a Necrotrophic Plant Pathogen.

Pantoea ananatis is an unusual bacterial pathogen that lacks typical virulence determinants yet causes extensive necrosis in onion foliage and bulb tissues. The onion necrosis phenotype is dependent on the expression of the phosphonate toxin, pantaphos, which is synthesized by putative enzymes encoded by the HiVir (high virulence) gene cluster. The genetic contributions of individual hvr genes in HiVir-mediated onion necrosis remain largely unknown, except for the first gene, hvrA (phosphoenolpyruvate mutase, pepM), whose deletion resulted in the loss of onion pathogenicity. In this study, using gene-deletion mutation and complementation, we report that, of the ten remaining genes, hvrB to hvrF are also strictly required for the HiVir-mediated onion necrosis and in-planta bacterial growth, whereas hvrG to hvrJ partially contributed to these phenotypes. As the HiVir gene cluster is a common genetic feature shared among the onion-pathogenic P. ananatis strains that could serve as a useful diagnostic marker of onion pathogenicity, we sought to understand the genetic basis of HiVir-positive yet phenotypically deviant (non-pathogenic) strains. We identified and genetically characterized inactivating single nucleotide polymorphisms in the essential hvr genes of six phenotypically deviant P. ananatis strains. Finally, inoculation of cell-free spent medium of the isopropylthio-ß-galactoside (IPTG)-inducible promoter (Ptac)-driven HiVir strain caused P. ananatis-characteristic red onion scale necrosis as well as cell death symptoms in tobacco. Co-inoculation of the spent medium with essential hvr mutant strains restored in-planta populations of the strains to the wild-type level, suggesting that necrotic tissues are important for the proliferation of P. ananatis in onion. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

A Novel Biosynthetic Gene Cluster Across the Pantoea Species Complex Is Important for Pathogenicity in Onion.

Onion center rot is caused by at least four species of genus Pantoea (P. ananatis, P. agglomerans, P. allii, and P. stewartii subsp. indologenes). Critical onion pathogenicity determinants for P. ananatis were recently described, but whether those determinants are common among other onion-pathogenic Pantoea species remains unknown. In this work, we report onion pathogenicity determinants in P. stewartii subsp. indologenes and P. allii. We identified two distinct secondary metabolite biosynthetic gene clusters present separately in different strains of onion-pathogenic P. stewartii subsp. indologenes. One cluster is similar to the previously described HiVir phosphonate biosynthetic cluster identified in P. ananatis and another is a novel putative phosphonate biosynthetic gene cluster, which we named Halophos. The Halophos gene cluster was also identified in P. allii strains. Both clusters are predicted to be phosphonate biosynthetic clusters based on the presence of a characteristic phosphoenolpyruvate phosphomutase (pepM) gene. The deletion of the pepM gene from either HiVir or Halophos clusters in P. stewartii subsp. indologenes caused loss of necrosis on onion leaves and red onion scales and resulted in significantly lower bacterial populations compared with the corresponding wild-type and complemented strains. Seven (halB to halH) of 11 genes (halA to halK) in the Halophos gene cluster are required for onion necrosis phenotypes. The onion nonpathogenic strain PNA15-2 (P. stewartii subsp. indologenes) gained the capacity to cause foliar necrosis on onion via exogenous expression of a minimal seven-gene Halophos cluster (genes halB to halH). Furthermore, cell-free culture filtrates of PNA14-12 expressing the intact Halophos gene cluster caused necrosis on onion leaves consistent with the presence of a secreted toxin. Based on the similarity of proteins to those with experimentally determined functions, we are able to predict most of the steps in Halophos biosynthesis. Together, these observations indicate that production of the toxin phosphonate seems sufficient to account for virulence of a variety of different Pantoea strains, although strains differ in possessing a single but distinct phosphonate biosynthetic cluster. Overall, this is the first report of onion pathogenicity determinants in P. stewartii subsp. indologenes and P. allii. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Enabling Population Biology Studies of Stemphylium vesicarium from Onion with Microsatellites.

Stemphylium leaf blight (SLB), caused by the fungus Stemphylium vesicarium, is dominant within the foliar disease complex affecting onion production in New York (NY). The disease causes premature defoliation and significant reductions in bulb weight and quality. Foliar diseases of onion are usually managed by an intensive fungicide program, but SLB management is complicated by resistance to multiple single-site modes of action. The design of integrated disease management strategies is limited by incomplete knowledge surrounding the dominant sources of S. vesicarium inoculum. To facilitate genomic-based studies of S. vesicarium populations, nine microsatellite markers were developed. The markers were multiplexed into two PCR assays containing four and five fluorescently labeled microsatellite markers. Initial testing of the S. vesicarium isolates found the markers were highly polymorphic and reproducible with an average of 8.2 alleles per locus. The markers were used to characterize 54 S. vesicarium isolates from major NY onion production regions in 2016 (n = 27) and 2018 (n = 27). Fifty-two multilocus genotypes (MLGs) were identified between these populations. Genotypic and allelic diversities were high in both the 2016 and 2018 populations. A greater degree of genetic variation was observed within populations than between years. No distinct pattern of MLGs according to population was identified and some MLGs were closely related between 2016 and 2018. The lack of evidence for linkage among loci also was strongly suggestive of clonal populations with only minor differences between the two populations. These microsatellite markers will be a foundational resource for the testing of hypotheses surrounding the population biology of S. vesicarium and therefore informing disease management.

Isolation and Characterization of Bacteria Associated with Onion and First Report of Onion Diseases Caused by Five Bacterial Pathogens in Texas, U.S.A.

Bacterial diseases pose a severe challenge to growers and cause significant loss to the billion-dollar onion industry in the United States. Texas is the sixth largest onion producing state, yet the bacterial communities associated with short-day onion crops grown in Texas have not been studied. This study was conducted to identify, characterize, and understand the diversity of bacteria associated with onion production in Texas. In 2020, 190 foliar and 210 bulb samples were collected from onion crops in the Rio Grande Valley and Winter Garden regions of Texas. Sequencing of the 16s rRNA gene was used to identify each bacterial strains to a genus. The pathogenicity to onion of each bacterial strain was tested using three assays: a red onion scale assay, a yellow onion bulb assay, and a foliar assay. Whole genome sequencing was done to identify the onion-pathogenic strains to species. Collectively, isolates of 24 genera belonging to three phyla were detected, including 19 genera from foliar samples and nine genera from bulb samples. Isolates in the Phylum Proteobacteria, including 15 genera of Gram-negative bacteria, were the most abundant of the taxa, comprising 90.0% of the strains isolated. The diversity of foliar isolates was evenly distributed between Gram-positive and Gram-negative bacteria, while Gram-negative bacteria dominated the isolates from bulb samples. In total, 83.9% of the bacterial isolates were not pathogenic on onion, with only isolates of Pantoea, Pseudomonas, Burkholderia, Erwinia, Enterobacter, and Curtobacterium proving pathogenic. Strains of Burkholderia gladioli, Pseudomonas alliivorans, Pantoea agglomerans, P. ananatis, and P. allii are the first documented cases of these pathogens of onion in Texas. Identifying and characterizing the nature of onion microflora, including pathogens of onion, is vital to developing rapid disease detection techniques via pathogenomics and minimizing losses through the application of effective disease management measures.

Rahnella perminowiae sp. nov., Rahnella bonaserana sp. nov., Rahnella rivi sp. nov. and Rahnella ecdela sp. nov., isolated from diverse environmental sources, and emended description of the genus Rahnella.

Bacteria isolated from onion bulbs suffering from bacterial decay in the United States and Norway were previously shown to belong to the genus Rahnella based on partial housekeeping gene sequences and/or fatty acid analysis. However, many strains could not be assigned to any existing Rahnella species. Additionally, strains isolated from creek water and oak as well as a strain with bioremediation properties were assigned to Rahnella based on partial housekeeping gene sequences. The taxonomic status of these 21 strains was investigated using multilocus sequence analysis, whole genome analyses, phenotypic assays and fatty acid analysis. Phylogenetic and phylogenomic analyses separated the strains into five clusters, one of which corresponded to Rahnella aceris. The remaining four clusters could be differentiated both genotypically and phenotypically from each other and existing Rahnella species. Based on these results, we propose the description of four novel species: Rahnella perminowiae sp. nov. (type strain SL6T=LMG 32257T=DSM 112609T), Rahnella bonaserana sp. nov. (H11bT=LMG 32256T=DSM 112610T), Rahnella rivi sp. nov. (FC061912-KT=LMG 32259T=DSM 112611T) and Rahnella ecdela sp. nov. (FRB 231T=LMG 32255T=DSM 112612T).

Pseudomonas uvaldensis sp. nov., a bacterial pathogen causing onion bulb rot.

A Gram-stain-negative, aerobic and non-spore-forming bacterial strain, designated 20TX0172T, was isolated from a rotting onion bulb in Texas, USA. The results of phylogenetic analysis based on the 16S rRNA sequence indicated that the novel strain represented a member of the genus Pseudomonas and had the greatest sequence similarities with Pseudomonas kilonensis 520-20T (99.3 %), Pseudomonas corrugata CFBP 2431T (99.2 %), and Pseudomonas viciae 11K1T (99.2 %) but the 16S rRNA phylogenetic tree displayed a monophyletic clade with Pseudomonas mediterranea CFBP 5447T. In the phylogenetic trees based on sequences of four housekeeping genes (gap1, gltA, gyrB and rpoD), the novel strain formed a separate branch, indicating that the strain was distinct phylogenetically from known species of the genus Pseudomonas. The genome-sequence-derived average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between the novel isolate and P. mediterranea DSM 16733T were 86.7 and 32.7 %, respectively. These values were below the accepted species cutoff threshold of 96 % ANI and 70 % dDDH, affirming that the strain represented a novel species. The genome size of the novel species was 5.98 Mbp with a DNA G+C content of 60.8 mol%. On the basis of phenotypic and genotypic characteristics, strain 20TX0172T represents a novel species of the genus Pseudomonas. The name Pseudomonas uvaldensis sp. nov. is proposed. The type strain is 20TX0172T (=NCIMB 15426T=CIP 112022T).

Bi-national outbreak of Salmonella Newport infections linked to onions: the United States experience.

From 2016-2019, dry bulb onions were the suspected cause of three multistate outbreaks in the United States. We investigated a large multistate outbreak of Salmonella Newport infections that caused illnesses in both the United States and Canada in 2020. Epidemiologic, laboratory and traceback investigations were conducted to determine the source of the infections, and data were shared among U.S. and Canadian public health officials. We identified 1127 U.S. illnesses from 48 states with illness onset dates ranging from 19 June to 11 September 2020. Sixty-six per cent of ill people reported consuming red onions in the week before illness onset. Thirty-five illness sub-clusters were identified during the investigation and seventy-four per cent of sub-clusters served red onions to customers during the exposure period. Traceback for the source of onions in illness sub-clusters identified a common onion grower in Bakersfield, CA as the source of red onions, and onions were recalled at this time. Although other strains of Salmonella Newport were identified in environmental samples collected at the Bakersfield, CA grower, extensive environmental and product testing did not yield the outbreak strain. This was the third largest U.S. foodborne Salmonella outbreak in the last 30 years. It is the first U.S. multistate outbreak with a confirmed link to dry bulb onions, and it was nearly 10-fold larger than prior outbreaks with a suspected link to onions. This outbreak is notable for its size and scope, as well as the international data sharing that led to implication of red onions as the primary cause of the outbreak. Although an environmental assessment at the grower identified several factors that likely contributed to the outbreak, no main reason was identified. The expedient identification of the outbreak vehicle and response of multiple public health agencies allowed for recall and removal of product from the marketplace, and rapid messaging to both the public and industry on actions to protect consumers; these features contributed to a decrease in cases and expeditious conclusion of the outbreak.

Production of transgenic Allium cepa by nanoparticles to resist Aspergillus niger infection.

BACKGROUND: Transgenic plants are becoming a more powerful tool in modern biotechnology. Genetic engineering was used in biotech-derived products to create genetically modified (GM) plants resistant to diseases. The onion (Allium cepa, L.) is a common, important perennial vegetable crop grown in Egypt for food and economic value. Onions are susceptible to a variety of fungal infections and diseases. Aspergillus niger is a common onion phytopathogen that causes diseases such as black mould (or black rot), which is a major issue, particularly when exporting onions. A. niger grows between the bulb's outer (dead, flaky) skin and the first fleshy scales, which become water-soaked. Thionin genes produce thionin proteins, which have antimicrobial properties against a variety of phytopathogens, including A. niger. Chitosan nanoparticles act as a carrier for the thionin gene, which allows A. cepa to resist infection by A. niger. METHODS AND RESULTS: Thionin gene (Thio-60) was transformed into A. cepa to be resistance to fungal infection. The gene was loaded on chitosan nanoparticles to be transformed into plants. Transgenic A. cepa had a 27% weight inhibition compared to non-transgenic one, which had a 69% inhibition. The expressed thionin protein has a 52% inhibitory effect on A. niger spore germination. All these findings supported thionin protein's antifungal activity as an antimicrobial peptide. Furthermore, the data presented here demonstrated the efficacy of chitosan nanoparticles in gene transformation. CONCLUSION: The present study describes the benefits of producing transgenic onion resistance to black rot diseases via expression of thionin proteins.

Phylogeny and Identification of Pantoea Species Associated with Bulb Rot and Bacterial Leaf Blight of Onion Crops in Uruguay.

Onion is among the most consumed vegetables in Uruguay, grown in the northwestern and southern regions of the country. The onion supply presents interannual variations associated with significant postharvest losses, mainly caused by bacterial rots. Besides bulb rotting, onion leaf lesions as well as infections on seed-stalks during seed production may be devastating for some varieties under conducive conditions. This research aimed to identify the causal agents of bulb rots and leaf blight of onion crops in Uruguay. Symptomatic bulbs, seeds-stalks, and leaves were collected from commercial fields from 2015 to 2020. Bacterial colonies were isolated and identified at genera level using physiological tests and 16S rRNA gene sequence analysis. A collection of 59 Pantoea spp. isolates was obtained (11 from bulbs and 48 from leaves and seeds-stalks). Multilocus sequence analysis using four housekeeping genes (rpoB, gyrB, leuS, and fusA) allowed the assignment of the isolates to five Pantoea species: P. ananatis, P. agglomerans, P. allii, P. eucalypti, and P. vagans. The last two species were not previously reported as onion pathogens elsewhere. The ability to cause disease symptoms was tested by leaf inoculation and red onion scale assays. P. ananatis isolates showed the highest aggressiveness in both assays. Specific isolates from P. allii (MAI 6022), P. eucalypti (MAI 6036), P. vagans (MAI 6050), and Pantoea sp. (MAI 6049) ranked second in aggressiveness on onion leaves, whereas only three isolates belonging to P. eucalypti (MAI 6036 and MAI 6058) and P. agglomerans (MAI 6045) exhibited the same scale-clearing phenotype as P. ananatis. Leaf inoculation assays were also performed on a set of eight onion cultivars and breeding lines. Overall, P. ananatis MAI 6032 showed the highest aggressiveness in all tested cultivars, followed by P. eucalypti MAI 6036. The presence of new reported bacterial species leads to complex disease management and highlights the need for further studies on virulence factors and the epidemiology of these pathogens.

Preliminary Studies on Detection of Fusarium Basal Rot Infection in Onions and Shallots Using Electronic Nose.

The evaluation of crop health status and early disease detection are critical for implementing a fast response to a pathogen attack, managing crop infection, and minimizing the risk of disease spreading. Fusarium oxysporum f. sp. cepae, which causes fusarium basal rot disease, is considered one of the most harmful pathogens of onion and accounts for considerable crop losses annually. In this work, the capability of the PEN 3 electronic nose system to detect onion and shallot bulbs infected with F. oxysporum f. sp. cepae, to track the progression of fungal infection, and to discriminate between the varying proportions of infected onion bulbs was evaluated. To the best of our knowledge, this is a first report on successful application of an electronic nose to detect fungal infections in post-harvest onion and shallot bulbs. Sensor array responses combined with PCA provided a clear discrimination between non-infected and infected onion and shallot bulbs as well as differentiation between samples with varying proportions of infected bulbs. Classification models based on LDA, SVM, and k-NN algorithms successfully differentiate among various rates of infected bulbs in the samples with accuracy up to 96.9%. Therefore, the electronic nose was proved to be a potentially useful tool for rapid, non-destructive monitoring of the post-harvest crops.

Chemical or Genetic Alteration of Proton Motive Force Results in Loss of Virulence of Burkholderia glumae, the Cause of Rice Bacterial Panicle Blight.

Rice is an important source of food for more than half of the world's population. Bacterial panicle blight (BPB) is a disease of rice characterized by grain discoloration or sheath rot caused mainly by Burkholderia glumae. B. glumae synthesizes toxoflavin, an essential virulence factor that is required for symptoms of the disease. The products of the tox operons, ToxABCDE and ToxFGHI, are responsible for the synthesis and the proton motive force (PMF)-dependent secretion of toxoflavin, respectively. The DedA family is a highly conserved membrane protein family found in most bacterial genomes that likely function as membrane transporters. Our previous work has demonstrated that absence of certain DedA family members results in pleiotropic effects, impacting multiple pathways that are energized by PMF. We have demonstrated that a member of the DedA family from Burkholderia thailandensis, named DbcA, is required for the extreme polymyxin resistance observed in this organism. B. glumae encodes a homolog of DbcA with 73% amino acid identity to Burkholderia thailandensis DbcA. Here, we created and characterized a B. glumae ΔdbcA strain. In addition to polymyxin sensitivity, the B. glumae ΔdbcA strain is compromised for virulence in several BPB infection models and secretes only low amounts of toxoflavin (∼15% of wild-type levels). Changes in membrane potential in the B. glumae ΔdbcA strain were reproduced in the wild-type strain by the addition of subinhibitory concentrations of sodium bicarbonate, previously demonstrated to cause disruption of PMF. Sodium bicarbonate inhibited B. glumae virulence in rice, suggesting a possible non-toxic chemical intervention for bacterial panicle blight. IMPORTANCE Bacterial panicle blight (BPB) is a disease of rice characterized by grain discoloration or sheath rot caused mainly by Burkholderia glumae. The DedA family is a highly conserved membrane protein family found in most bacterial genomes that likely function as membrane transporters. Here, we constructed a B. glumae mutant with a deletion in a DedA family member named dbcA and report a loss of virulence in models of BPB. Physiological analysis of the mutant shows that the proton motive force is disrupted, leading to reduction of secretion of the essential virulence factor toxoflavin. The mutant phenotypes are reproduced in the virulent wild-type strain without an effect on growth using sodium bicarbonate, a nontoxic buffer that has been reported to disrupt the PMF. The results presented here suggest that bicarbonate may be an effective antivirulence agent capable of controlling BPB without imposing an undue burden on the environment.

Maximizing growth and productivity of onion (Allium cepa L.) by Spirulina platensis extract and nitrogen-fixing endophyte Pseudomonas stutzeri.

The study focuses on the impact of foliar spraying cyanobacterium Spirulina platensis extract and the inoculation with the endophyte N2-fixing Pseudomonas stutzeri, and their mixture in the presence of different nitrogen doses on growth and yield of onion under field conditions. Bioactive compounds of Spirulina and Pseudomonas were analyzed by GC-MC and amino acid production of Spirulina by the amino acid analyzer. Hydrogen cyanide (HCN), indole acetic acid (IAA), ammonia (NH3), pectinase activity, and N2-fixation of Pseudomonas were measured. Plant height (cm), leaf length (cm), number of green leaves, bulb diameter (cm), fresh and dry weight of plant (g), chlorophyll a, b of leaves, bulb weight (g), marketable bulb yield (t. ha-1), cull bulb weight (t. ha-1), total bulb yield (t. ha-1), bulb diameter (cm), total soluble solids (TSS%), dry matter content (DM%), evaluation of storage behavior, and economic feasibility were estimated. Spirulina extract has several bioactive compounds. Pseudomonas can produce HCN, NH3, IAA, pectinase, and nitrogen fixation. The application of mixture with recommended dose of nitrogen increases the onion plant parameters, marketable yield, total bulb yield, bulb weight, bulb diameter, TSS%, DM%, net return, benefit-cost ratio (B:C), lowest cumulative weight loss% of bulbs during storage, and reduce culls weight compared with other treatments in two seasons. Application of S. platensis extract and inoculation with endophyte nitrogen-fixing P. stutzeri enhance the growth and productivity of the onion under different doses of nitrogen fertilizer.

Pseudomonas allii sp. nov., a pathogen causing soft rot of onion in Japan.

Six phytopathogenic bacterial strains, MAFF 301512, MAFF 301513, MAFF 301514T, MAFF 301515, MAFF 301516 and MAFF 301517, were isolated from soft rot lesions of onion (Allium cepa L.) in Japan. The cells were Gram-reaction-negative, aerobic, non-spore-forming, motile with one or two polar flagella and rod-shaped. Analysis of their 16S rRNA gene sequences showed that they belong to the genus Pseudomonas, with the highest similarities to Pseudomonas poae DSM 14936T (99.86 %), Pseudomonas simiae OLiT (99.85 %), Pseudomonas trivialis DSM 14937T (99.79 %) and Pseudomonas extremorientalis KMM 3447T (99.79 %). Their genomic DNA G+C content was 60.9 mol% and the major fatty acids (>5 % of the total fatty acids) present were C16 : 0, summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c), summed feature 8 (C18 : 1 ω7c /C18 : 1 ω6c) and C17 : 0 cyclo. Phylogenetic and phylogenomic analyses based on the rpoD gene and whole genome sequences, respectively, demonstrated that the strains belong to the Pseudomonas fluorescens subgroup, but form a monophyletic and robust clade, with Pseudomonas azotoformans as their neighbour. Between the strains and P. azotoformans, the average nucleotide identity scores were 95.63-95.70 %, whereas the digital DNA-DNA hybridization scores of the strains against their closest relatives, including P. azotoformans, were 65.4 % or less, which are lower than the 70 % cut-off for prokaryotic species delineation. The strains were differentiated from their closest relatives by phenotypic characteristics, pathogenicity in onion and cellular fatty acid composition. The phenotypic, chemotaxonomic and genotypic data showed that the strains represent a novel Pseudomonas species, proposed to be named Pseudomonas allii sp. nov., with MAFF 301514T (=ICMP 23680T) being the type strain.

Evaluation of qualitative traits in modified Iranian Red Rey onion and comparison genetic resistance with primary mass selection and Red Azar-shahr cv to Fusarium oxysporum using laboratory and molecular markers.

BACKGROUND: The bulb onion (Allium cepa L.) is grown on all continents except Antarctica, and is prized by essentially all of the world's cultures for its flavor and health-enhancing attributes. Onion breeders focus primarily on bulb characteristics such as color, shape, soluble-solids content, pungency and flavor, storage ability, and health-enhancing attributes, as well as plant characters such as resistances to diseases. The use of breeding approaches, offers great promise for population improvement and hybrid development addressing changes in consumer preference and production environments. The aim of this study is to evaluate the storage and qualitative feature of modified Red Rey Iranian Onion. METHOD: Firstly, the modified population was obtained by the selection of superior bulbs, cultivation, its self-pollination and consequently the identification of the best families and implement open pollination between them. In next level, the Red Rey Iranian modified with basic population and Red Azar-shahr cultivar (comparative) was crossed. RESULTS: Our results showed that the selection procedure has leading to improvement in variety of traits in population. Also, the modified Red Rey is significantly superior to the base mass in qualitative traits such as: bulb stiffness, bulb dry matter, TSS, total sugar and glucose; So that the percentage of dry bulb content increased from 10.4% in the basal mass to 11.1% in the modified Red Rey; while spouring and rotting, minerals, and dry matter, vitamin C and fructose-reducing sugar was not affected by genotype. In the second step, resistances to Fusarium wilt disease (laboratory and molecular markers) were evaluated. Based on the results of phenotypic evaluation, the modified Red Rey had the lowest rate and level of infection and the highest score. According to the results of genotypic evaluation, there is a very high genetic affinity between resistant and susceptible cultivars.