Effect of contact and systemic insecticides on the sharpshooter Bucephalogonia xanthophis (Hemiptera: cicadellidae), a vector of Xylella fastidiosa in citrus

The knockdown and toxic effects of insecticides of different chemical groups and modes of action registered for citrus in Brazil were investigated for effective control of Bucephalogonia xanthophis, a sharpshooter vector of Xylella fastidiosa in citrus. The active ingredients dimethoate (1.2 mL/1.2L), imidacloprid (0.24 mL/1.2L) and lambda-cyhalothrin (0.24 mL/1.2L), as well as a control (water), were sprayed onto branches of potted-citrus nursery trees to evaluate the effect of residual contact. The insects were confined on sprayed branches by using sleeve cages, in groups of 10 per branch (5 branches/treatment). Lambdacyhalothrin showed a knockdown effect on B. xanthophis (>70% mortality within 2 h of exposure), and the residues were effective for approximately one wk. Imidacloprid, lambdacyhalothrin and dimethoate suppressed the vector populations for up to 3 wk after application, when the insects were exposed to sprayed plants for at least 24 h. In another experiment, 2 neonicotinoid insecticides (thiamethoxam and imidacloprid) were applied by soil drench to potted nursery trees, in order to study their systemic effect, i.e., mortality by ingestion on sharpshooter adults. Thiamethoxam and imidacloprid effectively controlled the vectors at all concentrations tested, when the insects were exposed to treated plants for 24 h (>80% mortality) or 48 h (near 100% mortality). The knockdown effect of thiamethoxam and lambda-cyhalothrin might be particularly important to prevent vector transmission of X. fastidiosa in citrus groves.

Endophytic Methylobacterium extorquens expresses a heterologous beta-1,4-endoglucanase A (EglA) in Catharanthus roseus seedlings, a model host plant for Xylella fastidiosa

Based on the premise of symbiotic control, we genetically modified the citrus endophytic bacterium Methylobacterium extorquens, strain AR1.6/2, and evaluated its capacity to colonize a model plant and its interaction with Xylella fastidiosa, the causative agent of Citrus Variegated Chlorosis (CVC). AR1.6/2 was genetically transformed to express heterologous GFP (Green Fluorescent Protein) and an endoglucanase A (EglA), generating the strains ARGFP and AREglA, respectively. By fluorescence microscopy, it was shown that ARGFP was able to colonize xylem vessels of the Catharanthus roseus seedlings. Using scanning electron microscopy, it was observed that AREglA and X. fastidiosa may co-inhabit the C. roseus vessels. M. extorquens was observed in the xylem with the phytopathogen X. fastidiosa, and appeared to cause a decrease in biofilm formation. AREglA stimulated the production of resistance protein, catalase, in the inoculated plants. This paper reports the successful transformation of AR1.6/2 to generate two different strains with a different gene each, and also indicates that AREglA and X. fastidiosa could interact inside the host plant, suggesting a possible strategy for the symbiotic control of CVC disease. Our results provide an enhanced understanding of the M. extorquens-X. fastidiosa interaction, suggesting the application of AR1.6/2 as an agent of symbiotic control.

Global gene expression under nitrogen starvation in Xylella fastidiosa: contribution of the σ54 regulon

Abstract Background Xylella fastidiosa, a Gram-negative fastidious bacterium, grows in the xylem of several plants causing diseases such as citrus variegated chlorosis. As the xylem sap contains low concentrations of amino acids and other compounds, X. fastidiosa needs to cope with nitrogen limitation in its natural habitat. Results In this work, we performed a whole-genome microarray analysis of the X. fastidiosa nitrogen starvation response. A time course experiment (2, 8 and 12 hours) of cultures grown in defined medium under nitrogen starvation revealed many differentially expressed genes, such as those related to transport, nitrogen assimilation, amino acid biosynthesis, transcriptional regulation, and many genes encoding hypothetical proteins. In addition, a decrease in the expression levels of many genes involved in carbon metabolism and energy generation pathways was also observed. Comparison of gene expression profiles between the wild type strain and the rpoN null mutant allowed the identification of genes directly or indirectly induced by nitrogen starvation in a σ54-dependent manner. A more complete picture of the σ54 regulon was achieved by combining the transcriptome data with an in silico search for potential σ54-dependent promoters, using a position weight matrix approach. One of these σ54-predicted binding sites, located upstream of the glnA gene (encoding glutamine synthetase), was validated by primer extension assays, confirming that this gene has a σ54-dependent promoter. Conclusions Together, these results show that nitrogen starvation causes intense changes in the X. fastidiosa transcriptome and some of these differentially expressed genes belong to the σ54 regulon.

Mutations in CEP57 cause mosaic variegated aneuploidy syndrome.

Using exome sequencing and a variant prioritization strategy that focuses on loss-of-function variants, we identified biallelic, loss-of-function CEP57 mutations as a cause of constitutional mosaic aneuploidies. CEP57 is a centrosomal protein and is involved in nucleating and stabilizing microtubules. Our findings indicate that these and/or additional functions of CEP57 are crucial for maintaining correct chromosomal number during cell division.

Danos causados pelo Zucchini lethal chlorosis virus (ZLCV) sobre a produção de frutos comerciais de abobrinha de moita 'Caserta'

O ZLCV é um tospovírus encontrado com freqüência causando severos danos em cucurbitáceas. Nesse trabalho avaliaram-se os danos causados pelo ZLCV em abobrinha de moita 'Caserta', em campo na ESALQ/USP, Piracicaba-SP, onde esse vírus é freqüente. Plantas obtidas pela semeadura direta foram monitoradas periodicamente quanto à infecção pelo ZLCV por meio dos sintomas e por PTA-ELISA. Monitorou-se ainda a contaminação com Papaya ringspot virus - type W e Zucchini yellow mosaic virus, desconsiderando a produção dessas plantas. As plantas foram agrupadas em função da época de aparecimento dos sintomas do ZLCV, avaliando a produção de frutos comerciais (FC) e não comerciais (FNC) de cada grupo e comparando com a de plantas que permaneceram sem sintomas até o final do experimento. As plantas que apresentaram sintomas até os 23 dias após a emergência (DAE) não produziram qualquer tipo de frutos. FC foram colhidos de plantas que apresentaram sintomas a partir dos 42 DAE. Mesmo assim, houve redução de 78,5 % na produção de FC. Plantas que mostraram sintomas por ocasião da última colheita (55 DAE) apresentaram redução na produção de FC de 9,6 %. A infecção com o ZLCV até o início da frutificação inviabiliza a produção de FC de abobrinha de moita 'Caserta'.

Tolerance to iron chlorosis in non-grafted quince seedlings and in pear grafted onto quince plants

Grafting is a technique that may affect plant tolerance to iron chlorosis in plants cultivated for their fruit. Therefore, the objective of this study was to evaluate the tolerance of non-grafted quince seedlings and pear grafted onto quince plants cultivated in pots with alkaline soil. The experiment was conducted in a greenhouse at the University of Cordoba, Spain, in pots (3 L) filled with alkaline soil, with one plant per pot. The treatments consisted of two genotypes, quince (Cydonia oblonga Mill) semi-woody rooted cuttings, cultivar BA29, and pear (Pyrus Communis L.), cultivar Ercolini, grafted onto quince cultivar BA29 (rootstock), and two nutrient solutions with and without iron (80 mu M Fe-EDDHA) arranged in a completely random design with eight repetitions. Each pot received 250 mL of the nutrient solution on June 3rd, 2010. Chlorophyll indirect measurements and the main stem length were evaluated for six weeks after the commencement of the treatments. During the last week, the main stem dry matter weight and the leaf total iron content were determined. It was found that grafting pear seedlings onto quince rootstock resulted in a higher tolerance to iron deficiency than when quince was not grafted. Non-grafted quince plants without iron in the nutrient solution, compared to the results with its application, showed low SPAD (Soil-Plant Analyses Development) values and resulted in plants with a lower leaf iron content and lower dry matter production; however, decreased seedling stem growth was observed only in the last week of cultivation.

Pulsing With Low Concentration Gibberellin Plus Benzyladenine or Commercial Floral Preservatives Affect Postharvest Longevity, Quality, and Leaf Chlorosis of Cut Lilies and Gladioli

Effects of pulsing with different concentrations of gibberellin plus benzyladenine (GA(4+7) + BA), a proprietary mixture of GA(4+7) plus BA in a commercial floral preservative (GA(4+7) + BA + preservative), or a propriety mixture of sugar plus acidifier developed for bulbous flowers (floral bulb preservative) were studied on postharvest performance and quality of cut lily (Lilium hybrids) and gladiolus ( Gladiolus hybrids) flowers. Pulsing of cut stems of lily with GA(4+7) + BA at 5 or 2 mL.L-1 GA(4+7) + BA + preservative for 20 hours at 3 +/- 1 degrees C extended the vase life and controlled leaf chlorosis of 'Cobra'oriental lily and 'Cappuccino'and Pot Corn'asiatic lily. Cut 'Orange Art'asiatic lily performed best when pulsed with GA(4+7) + BA at 10 mg.L-1. For cut gladiolus, pulsing with GA(4+7) + BA at 10 mg.L-1 extended the vase life of 'Alice', 'Mammoth', and 'Passion', while 'Scarlet'had the longest vase life when pulsed with 5 mg.L-1 GA(4+7) + BA. GA(4+7) + BA + preservative also extended the vase life and controlled leaf chlorosis, but the floral bulb preservative had no effect on vase life extension or preventing leaf chlorosis of lilies. Gladiolus cultivars had no or minor leaf chlorosis during vase period. Overall, overnight pulsing with GA(4+7) + BA + or GA(4+7) + BA + preservative extended the vase life and prevented leaf chlorosis

Indução de resistência a Tomato severe rugose virus e Tomato chlorosis virus em tomateiro determinado

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Variegated transgene expression in mouse mammary gland is determined by the transgene integration locus.

Mice carrying an ovine beta-lactoglobulin (BLG) transgene secrete BLG protein into their milk. To explore transgene expression stability, we studied expression levels in three BLG transgenic mouse lines. Unexpectedly, two lines exhibited variable levels of transgene expression. Copy number within lines appeared to be stable and there was no evidence of transgene rearrangement. In the most variable line, BLG production levels were stable within individual mice in two successive lactations. Backcrossing demonstrated that genetic background did not contribute significantly to variable expression. Tissue in situ hybridization revealed mosaicism of transgene expression within individual mammary glands from the two variable lines; in low expressors, discrete patches of cells expressing the transgene were observed. Transgene protein concentrations in milk reflected the proportion of epithelial cells expressing BLG mRNA. Furthermore, chromosomal in situ hybridization revealed that transgene arrays in both lines are situated close to the centromere. We propose that mosaicism of transgene expression is a consequence of the chromosomal location and/or the nature of the primary transgene integration event.

Overexploitation, malnutrition and stigma in a woman's illness: chlorosis in contemporary Spanish medicine (1877-1936)

The work was financed by the Project: 'Antecedentes históricos de la nutrición comunitaria en España: los primeros intentos de institucionalización, 1923-1947' (HUM2005-04961-C03-01).

First complete genome sequence of a capsicum chlorosis tospovirus isolate from Australia with an unusually large S RNA intergenic region

The first complete genome sequence of capsicum chlorosis virus (CaCV) from Australia was determined using a combination of Illumina HiSeq RNA and Sanger sequencing technologies. Australian CaCV had a tripartite genome structure like other CaCV isolates. The large (L) RNA was 8913 nucleotides (nt) in length and contained a single open reading frame (ORF) of 8634 nt encoding a predicted RNA-dependent RNA polymerase (RdRp) in the viral-complementary (vc) sense. The medium (M) and small (S) RNA segments were 4846 and 3944 nt in length, respectively, each containing two non-overlapping ORFs in ambisense orientation, separated by intergenic regions (IGR). The M segment contained ORFs encoding the predicted non-structural movement protein (NSm; 927 nt) and precursor of glycoproteins (GP; 3366 nt) in the viral sense (v) and vc strand, respectively, separated by a 449-nt IGR. The S segment coded for the predicted nucleocapsid (N) protein (828 nt) and non-structural suppressor of silencing protein (NSs; 1320 nt) in the vc and v strand, respectively. The S RNA contained an IGR of 1663 nt, being the largest IGR of all CaCV isolates sequenced so far. Comparison of the Australian CaCV genome with complete CaCV genome sequences from other geographic regions showed highest sequence identity with a Taiwanese isolate. Genome sequence comparisons and phylogeny of all available CaCV isolates provided evidence for at least two highly diverged groups of CaCV isolates that may warrant re-classification of AIT-Thailand and CP-China isolates as unique tospoviruses, separate from CaCV.

Transcriptome Analysis of Capsicum Chlorosis Virus-Induced Hypersensitive Resistance Response in Bell Capsicum

Background Capsicum chlorosis virus (CaCV) is an emerging pathogen of capsicum, tomato and peanut crops in Australia and South-East Asia. Commercial capsicum cultivars with CaCV resistance are not yet available, but CaCV resistance identified in Capsicum chinense is being introgressed into commercial Bell capsicum. However, our knowledge of the molecular mechanisms leading to the resistance response to CaCV infection is limited. Therefore, transcriptome and expression profiling data provide an important resource to better understand CaCV resistance mechanisms. Methodology/Principal Findings We assembled capsicum transcriptomes and analysed gene expression using Illumina HiSeq platform combined with a tag-based digital gene expression system. Total RNA extracted from CaCV/mock inoculated CaCV resistant (R) and susceptible (S) capsicum at the time point when R line showed a strong hypersensitive response to CaCV infection was used in transcriptome assembly. Gene expression profiles of R and S capsicum in CaCV- and buffer-inoculated conditions were compared. None of the genes were differentially expressed (DE) between R and S cultivars when mock-inoculated, while 2484 genes were DE when inoculated with CaCV. Functional classification revealed that the most highly up-regulated DE genes in R capsicum included pathogenesis-related genes, cell death-associated genes, genes associated with hormone-mediated signalling pathways and genes encoding enzymes involved in synthesis of defense-related secondary metabolites. We selected 15 genes to confirm DE expression levels by real-time quantitative PCR. Conclusion/Significance DE transcript profiling data provided comprehensive gene expression information to gain an understanding of the underlying CaCV resistance mechanisms. Further, we identified candidate CaCV resistance genes in the CaCV-resistant C. annuum x C. chinense breeding line. This knowledge will be useful in future for fine mapping of the CaCV resistance locus and potential genetic engineering of resistance into CaCV-susceptible crops.

Optical fiber laser induced fluorescence spectroscopy as a citrus canker diagnostic

Citrus canker is a serious disease caused by Xanthomonas citri subsp. citri bacteria, which infects citrus plants (Citrus spp.) leading to large economic losses in citrus production worldwide. In this work, laser induced fluorescence spectroscopy (LIF) was investigated as a diagnostic technique for citrus canker disease in citrus trees at an orchard using a portable optical fiber based spectrometer. For comparison we have applied LIF to leaves contaminated with citrus canker, citrus scab, citrus variegates chlorosis, and Huanglongbing (HLB, Greening). In order to reduce the noise in the data, we collected spectra from ten leaves with visual symptoms of diseases and from five healthy leaves per plant. This procedure is carried out in order to minimize the environmental effect on the spectrum (water and nutrient supply) of each plant. Our results show that this method presents a high sensitivity (similar to 90%), however it does present a low specificity (similar to 70%) for citrus canker diagnostic. We believe that such poor performance is due to the fact that the optical fiber collects light from only a small part of the leaf. Such results may be improved using the fluorescence imaging technique on the whole leaf. (C) 2010 Optical Society of America

Phytoplasma closely related to the pigeon pea witches`-broom phytoplasma (16Sr IX) is associated with citrus huanglongbing symptoms in the state of Sao Paulo, Brazil

In February 2007, sweet orange trees with characteristic symptoms of huanglongbing (HLB) were encountered in a region of Sao Paulo state (SPs) hitherto free of HLB. These trees tested negative for the three liberibacter species associated with HLB. A polymerase chain reaction (PCR) product from symptomatic fruit columella DNA amplifications with universal primers fDI/rPI was cloned and sequenced. The corresponding agent was found to have highest 16S rDNA sequence identity (99%) with the Pigeon pea witches`-broom phytoplasma of group 16Sr IX. Sequences of PCR products obtained with phytoplasma 16S rDNA primer pairs fU5/rU3, fU5/P7 confirm these result.,;. With two primers D7f2/D7r2 designed based oil the 16S rDNA Sequence of the cloned DNA fragment, positive amplifications were obtained from more than one hundred samples including symptomatic fruits and blotchy mottle leaves. Samples positive for phytoplasmas were negative for liberibacters, except for four samples, which were positive for both the phytoplasma and `Candidatus Liberibacter asiaticus`. The phytoplasma was detected by electron microscopy in the sieve tubes of midribs from symptomatic leaves. These results Show that a phytoplasma of group IX is associated with citrus HLB symptoms ill northern, central, and Southern SPs. This phytoplasma has very probably been transmitted to citrus from an external Source of inoculum, but the Putative insect vector is not yet known.