ReDivia - Repositorio Digital de l'Instit Valencià d'Investigacions Agràries
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mRNA from selected genes is useful for specific detection and quantification of viable Xanthomonas citri subsp citri
Get PDFThe purpose of this study was to assess the stability of mRNA and rRNA for evaluation of viability for Xanthomonas citri subsp. citri (Xcc). Total RNA from Xcc suspensions subjected to different stress treatments (high temperature or chemical treatment with sodium orthophenylphenate at different concentrations) was extracted at different time periods post-treatment (0, 3, 24 and 48 h) and analysed by quantitative real-time reverse transcription PCR (Q-RT-PCR). Primers were designed from selected fragments of rRNA and mRNA from genes involved in bacterial fitness, virulence or general metabolic mechanisms (gumD, rpfB, avrBs2 and gyrB). After stress treatment, only a 445-bp fragment from the gumD mRNA was detected in live Xcc cells specifically, whereas other RNA fragments, as well as DNA targets, were detected in both viable and nonviable cells. Statistical analyses demonstrated that the amount of some transcripts from genes involved in xanthan synthesis, pathogenicity factor regulation and DNA processing was significantly reduced after lethal treatments. The amplification of the 445-bp product from gumD mRNA was demonstrated to be useful for the detection of viable Xcc; the product was detected specifically from viable bacteria on leaf and citrus fruit surfaces and in citrus canker lesions. Instability of long RNA fragments can be used as a practical tool for the study of survival of citrus canker bacteria or for diagnostic purposes when the presence of viable bacteria needs to be confirmed
Relation of post-thaw sperm DNA damage and lipid peroxidation with freezability in boars
No full textFunctions and origin of plasmids in Erwinia species that are pathogenic to or epiphytically associated with pome fruit trees
Get PDFThe genus Erwinia includes plant-associated pathogenic and non-pathogenic species. Among them, all species pathogenic to pome fruit trees (E. amylovora, E. pyrifoliae, E. piriflorinigrans, Erwinia sp. from Japan) cause similar symptoms, but differ in their degrees of aggressiveness, i.e. in symptoms, host range or both. The presence of plasmids of similar size, in the range of 30 kb, is a common characteristic that they possess. Besides, they share some genetic content with high homology in several genes associated with exopolysaccharide production and hence, with virulence, as well as in some other genes. Knowledge of the content of these plasmids and comparative genetic analyses may provide interesting new clues to understanding the origin and evolution of these pathogens and the level of symptoms they produce. Furthermore, genetic similarities observed among some of the plasmids (and genomes) from the above indicated pathogenic species and E. tasmaniensis or E. billingiae, which are epiphytic on the same hosts, may reveal associations that could expose the mechanisms of origin of pathogens. A summary of the current information on their plasmids and the relationships among them is presented here
Selection of Ceratitis capitata (Diptera: Tephritidae) Specific Recombinant Monoclonal Phage Display Antibodies for Prey Detection Analysis
Get PDFSeveral recombinant antibodies against the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae), one of the most important pests in agriculture worldwide, were selected for the first time from a commercial phage display library of human scFv antibodies. The specificity and sensitivity of the selected recombinant antibodies were compared with that of a rabbit polyclonal serum raised in parallel using a wide range of arthropod species as controls. The selected recombinant monoclonal antibodies had a similar or greater specificity when compared with classical monoclonal antibodies. The selected recombinant antibodies were successfully used to detect the target antigen in the gut of predators and the scFv antibodies were sequenced and compared. These results demonstrate the potential for recombinant scFv antibodies to be used as an alternative to the classical monoclonal antibodies or even molecular probes in the post-mortem analysis studies of generalist predators. Citation: Monzo C, Urbaneja A, Ximenez-Embun M, Garcia-Fernandez J, Garcia JL, et al. (2012) Selection of Ceratitis capitata (Diptera: Tephritidae) Specific Recombinant Monoclonal Phage Display Antibodies for Prey Detection Analysis. PLoS ONE 7(12): e51440. doi:10.1371/journal.pone.005144
SNP mining in C. clementina BAC end sequences; transferability in the Citrus genus (Rutaceae), phylogenetic inferences and perspectives for genetic mapping
Get PDFBackground: With the increasing availability of EST databases and whole genome sequences, SNPs have become the most abundant and powerful polymorphic markers. However, SNP chip data generally suffers from ascertainment biases caused by the SNP discovery and selection process in which a small number of individuals are used as discovery panels. The ongoing International Citrus Genome Consortium sequencing project of the highly heterozygous Clementine and sweet orange genomes will soon result in the release of several hundred thousand SNPs. The primary goals of this study were: (i) to estimate the transferability within the genus Citrus of SNPs discovered from Clementine BACend sequencing (BES), (ii) to estimate bias associated with the very narrow discovery panel, and (iii) to evaluate the usefulness of the Clementine-derived SNP markers for diversity analysis and comparative mapping studies between the different cultivated Citrus species. Results: Fifty-four accessions covering the main Citrus species and 52 interspecific hybrids between pummelo and Clementine were genotyped on a GoldenGate array platform using 1,457 SNPs mined from Clementine BES and 37 SNPs identified between and within C. maxima, C. medica, C. reticulata and C. micrantha. Consistent results were obtained from 622 SNP loci. Of these markers, 116 displayed incomplete transferability primarily in C. medica, C. maxima and wild Citrus species. The two primary biases associated with the SNP mining in Clementine were an overestimation of the C. reticulata diversity and an underestimation of the interspecific differentiation. However, the genetic stratification of the gene pool was high, with very frequent significant linkage disequilibrium. Furthermore, the shared intraspecific polymorphism and accession heterozygosity were generally enough to perform interspecific comparative genetic mapping. Conclusions: A set of 622 SNP markers providing consistent results was selected. Of the markers mined from Clementine, 80.5% were successfully transferred to the whole Citrus gene pool. Despite the ascertainment biases in relation to the Clementine origin, the SNP data confirm the important stratification of the gene pools around C. maxima, C. medica and C. reticulata as well as previous hypothesis on the origin of secondary species. The implemented SNP marker set will be very useful for comparative genetic mapping in Citrus and genetic association in C. reticulata
Isolation and characterization of polymorphic microsatellite markers in Tetranychus urticae and cross amplification in other Tetranychidae and Phytoseiidae species of economic importance
No full textTetranychus urticae Koch is a cosmopolitan phytophagous mite considered as the most polyphagous species among spider mites. Population genetic studies using molecular markers such as microsatellites have proven to be extremely informative to address questions about population structure, phylogeography and host preferences. The aim of this study was to increase the available molecular tools to gain insight into the genetic structure of T. urticae populations of citrus orchards, which might help in their management. Five microsatellite DNA libraries were developed using probes with the motifs CT, CTT, GT and CAC following the FIASCO protocol. Positive clones, those that included the insert with the microsatellite, were detected using the PIMA-PCR technique. Combinations of primers were designed on 22 out of 32 new microsatellites loci and their polymorphism was tested in four populations sampled along the eastern coast of Spain. Eleven successful amplifications were obtained. Cross amplification was tested in the tetranychids Aphlonobia histricina, Eutetranychus banksi, E. orientalis, Oligonychus perseae, Panonychus citri, Tetranychus evansi, T. okinawanus and T. turkestani, and the phytoseiids Amblyseius swirskii, A. cucumeris, A. andersoni, Euseius stipulatus, Neoseiulus barkeri, N. californicus, Phytoseiulus persimilis and Typhlodromus phialatus. Eight successful cross amplifications were obtained
Acta Horticulturae
No full textEarly defoliation has been found to be an effective tool to reduce cluster compactness and improve fruit composition in different areas of northern Italy. Our objective was to test this technique under warmer conditions in ‘Tempranillo’ vines in two locations of Spain. In Requena (East-Spain), non-defoliated vines were compared with three defoliation treatments performed at different phenological phases (flowering or fruit set) and intensities. Sixteen vines per treatment were randomly selected in the vineyard. In the Badajoz (West-Spain) site, where vines are more vigorous, control vines were compared with defoliation carried out at fruit set. Treatments were carried out in 108 plants per replicate and four replicates per treatment. In the first season, as an average for all defoliation treatments, yield was reduced 22% by leaf pulling in East-Spain but not in West-Spain where yield was similar in all treatments. In East-Spain the yield reduction was due to lower berry fresh weight, while fruit set was not affected by defoliation. The effects of leaf pulling on grape acidity were different in the two locations, with defoliation decreasing must pH in West-Spain but with an increasing trend in East-Spain. Berry phenolic concentration increased with leaf removal in East-Spain but not in West-Spain where however cluster health was improved by defoliation. In East-Spain, the experiment was repeated during a second season, and in that year there was a heavier reduction in yield (-41%) due to lower bud fertility and fruit set produced by leaf pulling, suggesting important carry-over effects of the treatments. Overall, results indicate that even in warm terroirs early defoliation can improve berry composition but in different fashion depending on specific site characteristics. Growers should take into account that an important yield penalty due to defoliation might occur
Crop loss, aetiology, and epidemiology of citrus black spot in Ghana
No full textCitrus Black Spot (CBS), caused by Guignardia citricarpa, was detected for the first time in Ghana and in West Africa. The disease was first observed in the Eastern Region in 1999 with typical disease symptoms including hard spot, virulent spot and false melanose were observed on several citrus species. A survey revealed that the disease has reached epidemic levels in the citrus-producing areas of the Eastern and Ashanti regions and is spreading rapidly within these areas and to other regions of the country. Currently, CBS is the most important fruit disease of citrus in Ghana, causing about 22% crop loss. Although the disease does not cause postharvest decay and the internal quality of the fruit is not affected, significant amounts of blemished fruit are discarded at the markets. Disease incidence and severity was found to be higher on mature than on young citrus trees. Pycnidia were found on fruit with hard spot symptoms, and pycnidia and pseudothecia typical of Guignardia spp. were found on decomposing leaves. Two species, G. citricarpa and G. mangiferae, were isolated from 15% of the samples collected and identified using the Oatmeal Agar test and by PCR with species-specific DNA primers. Isolates of G. citricarpa produced CBS symptoms after 80 to 233 days on 75% of the artificially inoculated young fruit of Valencia Late sweet orange. The fungus was re-isolated from symptomatic, inoculated fruit completing Koch's postulates. Isolates of the endophyte G. mangiferae did not induce symptoms in the pathogenicity tests. In epidemiological studies, infections were detected from November to February for the minor cropping season and from May to November for the major season. Fruit of Valencia Late sweet orange were susceptible to G. citricarpa infection for up to 7 months after petal fall. Knowledge of the disease cycle in Ghana will improve methods for disease control