Sequencing of diverse mandarin, pummelo and orange genomes reveals complex history of admixture during citrus domestication

Cultivated citrus are selections from, or hybrids of, wild progenitor species whose identities and contributions to citrus domestication remain controversial. Here we sequence and compare citrus genomes-a high-quality reference haploid clementine genome and mandarin, pummelo, sweet-orange and sour-orange genomes-and show that cultivated types derive from two progenitor species. Although cultivated pummelos represent selections from one progenitor species, Citrus maxima, cultivated mandarins are introgressions of C. maxima into the ancestral mandarin species Citrus reticulata. The most widely cultivated citrus, sweet orange, is the offspring of previously admixed individuals, but sour orange is an F1 hybrid of pure C. maxima and C. reticulata parents, thus implying that wild mandarins were part of the early breeding germplasm. A Chinese wild 'mandarin' diverges substantially from C. reticulata, thus suggesting the possibility of other unrecognized wild citrus species. Understanding citrus phylogeny through genome analysis clarifies taxonomic relationships and facilitates sequence-directed genetic improvement. (Résumé d'auteur

Towards the identification of flower-specific genes in Citrus spp

Citrus sinensis is a perennial woody species, for which genetic approaches to the study of reproductive development are not readily amenable. Here, the usefulness of the CitEST Expressed Sequence Tag (EST) database is demonstrated as a reliable new resource for identifying novel genes exclusively related to Citrus reproductive biology. We performed the analysis of an EST dataset of the CitEST Project containing 4,330 flower-derived cDNA sequences. Relying on bioinformatics tools, sequences exclusively present in this flower-derived sequence collection were selected and used for the identification of Citrus putative flower-specific genes. Our analysis revealed several Citrus sequences showing significant similarity to conserved genes known to have flower-specific expression and possessing functions related to flower metabolism and/or reproductive development in diverse plant species. Comparison of the Citrus flower-specific sequences with all available plant peptide sequences unraveled 247 unique transcripts not identified elsewhere within the plant kingdom. Additionally, 49 transcripts, for which no biological function could be attributed by means of sequence comparisons, were found to be conserved among plant species. These results allow further gene expression analysis and possibly novel approaches to the understanding of reproductive development in Citrus

Towards the identification of flower-specific genes in Citrus spp

Citrus sinensis is a perennial woody species, for which genetic approaches to the study of reproductive development are not readily amenable. Here, the usefulness of the CitEST Expressed Sequence Tag (EST) database is demonstrated as a reliable new resource for identifying novel genes exclusively related to Citrus reproductive biology. We performed the analysis of an EST dataset of the CitEST Project containing 4,330 flower-derived cDNA sequences. Relying on bioinformatics tools, sequences exclusively present in this flower-derived sequence collection were selected and used for the identification of Citrus putative flower-specific genes. Our analysis revealed several Citrus sequences showing significant similarity to conserved genes known to have flower-specific expression and possessing functions related to flower metabolism and/or reproductive development in diverse plant species. Comparison of the Citrus flower-specific sequences with all available plant peptide sequences unraveled 247 unique transcripts not identified elsewhere within the plant kingdom. Additionally, 49 transcripts, for which no biological function could be attributed by means of sequence comparisons, were found to be conserved among plant species. These results allow further gene expression analysis and possibly novel approaches to the understanding of reproductive development in Citrus.761768Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Exogenous Indole-3-Acetic Acid Production and Phosphate Solubilization by <i>Chlorella vulgaris</i> Beijerinck in Heterotrophic Conditions

The role of soil-borne microalgae in the edaphic ecosystem is barely known, especially concerning their plant-growth-promoting traits, although they are used as biofertilizers. In this study, a microalgal strain isolated from soil cultivated with maize was evaluated as an exogenous producer of indole-3-acetic acid (IAA) in potato dextrose broth—PD—and sugarcane molasses, with or without tryptophan, and phosphate solubilizer (in ‘National Botanical Research Institute’s phosphate’—NBRIP—medium with tricalcium phosphate)with plant-growth-promoting traits, under heterotrophic conditions. The species of microalga was identified, by sequencing the ITS region in the rDNA and the morphological characteristics, as Chlorella vulgaris. Its growth was significantly higher in the PD medium, with slower growth in molasses. The addition of tryptophan did not influence the growth of C. vulgaris in either medium, but it increased the production of IAA, mainly in the PD medium, to 265 µg mL−1. The microalga grew in the medium with insoluble phosphate, releasing phosphorus into the medium (30 mg L−1 after 72 h). This is the first study on the application of C. vulgaris as a phosphate solubilizer. More studies should be performed on C. vulgaris as a prospective plant-growth-promoting microorganism, besides its ability to produce exogenous IAA, and further investigations should be conducted on developing inexpensive culture media

Modified Monosaccharides Content of Xanthan Gum Impairs Citrus Canker Disease by Affecting the Epiphytic Lifestyle of Xanthomonas citri subsp. citri

Xanthomonas citri subsp. citri (X. citri) is a plant pathogenic bacterium causing citrus canker disease. The xanA gene encodes a phosphoglucomutase/phosphomannomutase protein that is a key enzyme required for the synthesis of lipopolysaccharides and exopolysaccharides in Xanthomonads. In this work, firstly we isolated a xanA transposon mutant (xanA::Tn5) and analyzed its phenotypes as biofilm formation, xanthan gum production, and pathogenesis on the sweet orange host. Moreover, to confirm the xanA role in the impaired phenotypes we further produced a non-polar deletion mutant (ΔxanA) and performed the complementation of both xanA mutants. In addition, we analyzed the percentages of the xanthan gum monosaccharides produced by X. citri wild-type and xanA mutant. The mutant strain had higher ratios of mannose, galactose, and xylose and lower ratios of rhamnose, glucuronic acid, and glucose than the wild-type strain. Such changes in the saccharide composition led to the reduction of xanthan yield in the xanA deficient strain, affecting also other important features in X. citri, such as biofilm formation and sliding motility. Moreover, we showed that xanA::Tn5 caused no symptoms on host leaves after spraying, a method that mimetics the natural infection condition. These results suggest that xanA plays an important role in the epiphytical stage on the leaves that is essential for the successful interaction with the host, including adaptive advantage for bacterial X. citri survival and host invasion, which culminates in pathogenicity

Severity assessment in the Nicotiana tabacum-Xylella fastidiosa subsp. pauca pathosystem: design and inter-laboratory validation of a standard area diagram

Nicotiana tabacum, a non-natural host, is an excellent plant model for biological and functional genomics studies on plant host-Xylella fastidiosa interactions. Symptoms by X. fastidiosa subsp. pauca on tobacco have been characterized but current severity assessment method is an ordinal scale developed for symptoms on citrus. We designed a standard area diagram (SAD) to aid in the visual estimation of percent area affected (% severity) and performed a multi-laboratory validation on tobacco cv. 'Havana inoculated with X. fastidiosa subsp. pauca. Inoculated plants were monitored over time and digital images of the symptoms were obtained. Three different softwares (APS Asses, ImageJ and Leaf Doctor) were used to segment the images and calculate % severity. Ten true-color images composed a 10-image SAD (0.5, 5, 10, 15, 25, 35, 45, 55, 65 and 75%). Fifty raters at four laboratories assigned % severity first without and then with the SAD on the same testing image set of 40 images following the same instructions provided by an examiner at each Laboratory. The means of % severity by all softwares were assumed to represent the actual % severity given the perfect agreement between them. The unaided estimates were less precise, biased towards overestimation (up to 50 percent points) and less concordant between raters. Accuracy and precision varied considerably among the raters and effect of the SAD on the overall concordance to the actual severity was dependent on the Laboratory; in all but one Laboratory, the group means of the agreement index statistically improved. The between-rater agreement improved in all Laboratories when using the SAD. The SAD may help to standardize and allow comparison across different laboratories, but care should be taken during instructions on how to discriminate symptoms that are not readily discernible as well as on how to correctly use of the SAD

Comparing submerged and solid-state fermentation of agro-industrial residues for the production and characterization of lipase by Trichoderma harzianum

Lipase production by Trichoderma harzianum was evaluated in submerged fermentation (SF) and solid-state fermentation (SSF) using a variety of agro-industrial residues. Cultures in SF showed the highest activity (1.4 U/mL) in medium containing 0.5 % (w/v) yeast extract, 1 % (v/v) olive oil and 2.5 C:N ratio. This paper is the first to report lipase production by T. harzianum in SSF. A 1:2 mixture of castor oil cake and sugarcane bagasse supplemented with 1 % (v/w) olive oil showed the best results among the cultures in SSF (4 U/g ds). Lipolytic activity was stable in a slightly acidic to neutral pH, maintaining 50 % activity after 30 min at 50 C. Eighty percent of the activity remained after 1 h in 25 % (v/v) methanol, ethanol, isopropanol or acetone. Activity was observed with vegetable oils (olive, soybean, corn and sunflower) and long-chain triacylglycerols (triolein), confirming the presence of a true lipase. The results of this study are promising because they demonstrate an enzyme with interesting properties for application in catalysis produced by fermentation at low cost. © 2012 Springer-Verlag and the University of Milan