Occurrence of Centrouropoda almerodai and Uroobovella marginata (Acari : Uropodina) phoretic on the Red Palm Weevil in Malta

The unwanted introduction of the Red Palm Weevil (RPW) coincides with the spread in Malta of two species of Uropodid mites associated with this weevil. Usually, adult RPW carry phoretic forms of C. almerodai which are attached to the underside of elytrae, and U. marginata that prefers exposed surfaces of sternum, pygidium, head and legs. These mites use adult RPW to abandon dead palms and to colonize newly infested host-plants. Their role as plant pests is however negligible. Even the plant pathogen conidia, Curvularia which are carried by the mites, seem unable to germinate in palms under laboratory conditions. Both Centrouropoda almerodai and Uroobovella marginata are established in the Maltese Islands.peer-reviewe

Characterization of Citrus tristeza virus isolates recovered in Syria and Apulia (southern Italy) using different molecular tools

Citrus tristeza virus (CTV) is the causal agent of the most important virus disease of citrus. CTV isolates differing in biological and molecular characteristics have been reported worldwide. Recently, CTV was detected in Syria in citrus groves from two Governorates (Lattakia and Tartous) and several CTV outbreaks have been reported in Apulia (southern Italy) since 2003. To molecularly characterize the CTV populations spreading in Syria and Italy, a number of isolates from each region was selected and examined by different molecular approaches including: Multiple Molecular Markers analysis (MMM), real time RT-(q)PCR, single strand conformation polymorphism (SSCP) of the major coat protein (CP) gene (P25), and sequence analysis of the CP (P25), P18, P20 and RdRp genes. SSCP analysis of CP25 yielded two distinct simple patterns among the Syrian isolates and three different patterns in the Italian isolates. Based on MMM analysis, all Syrian CTV isolates were categorized as VT-like genotype, whereas the Italian isolates reacted only with the markers specific for the T30 genotype. These findings were also confirmed by RT-qPCR and by sequencing analysis of four genomic regions. The Italian isolates had nucleotide identities which varied: from 99.5 to 99.8 for the CP gene; from 97.4% to 98.3% for the P18 gene; from 98.6% to 99.8% for the P20 and from 97.8% to 99.1% for the partial RdRp sequenced. High sequence identity was found for all genomic regions analyzed between the Syrian isolates (from 98.9% to 99.6%). These results show that the CTV populations spreading in Apulia and Syria are associated with different genotypes, indicating different potential impacts on the citrus trees in the field. Since in both areas the introduction of the virus is relatively recent, infected plants resulted to contain a single and common genotype, suggesting that CTV is spreading from the first outbreaks by aphids or local movement of autochthonous infected plant material

A real time loop-mediated isothermal amplification (RealAmp) assay for rapid detection of Pleurostoma richardsiae in declining olive plants

Pleurostoma richardsiae is associated with host trunk diseases, known to cause dieback, cankers and wilting of woody trees, and human infections. This fungus was isolated from wood tissues of declining olive trees and grapevines showing esca disease symptoms, in the Apulia region of Italy. Fungus detection has been based on morphological and molecular features, which are time-consuming to identify and require well-trained personnel. Improvement of Pl. richardsiae detection in olive was achieved through development of real time loop-mediated isothermal amplification targeting the intergenic spacer (IGS) region of the fungus. Specificity of the assay was confirmed using ten Pl. richardsiae strains and 36 other fungus strains of species usually isolated from declining olive trees. The achieved limit of detection was 7.5 × 10-2 ng μL-1 of Pl. richardsiae genomic DNA. A preliminary validation of RealAmp was also performed using material from infected olive plants artificially inoculated in a greenhouse

Phytophthora nicotianae is the predominant Phytophthora species in citrus nurseries in Egypt

Phytophthora root rot is considered to be the most destructive disease to citrus production in Egypt. Phytophthora species are generally present in citrus nurseries, where soil pots containing the survival propagules are considered responsible for their spread into new orchards. The goal of this study was to investigate the distribution and seasonal variation of Phytophthora species in soil and feeder roots in two Egyptian citrus nurseries, characterized by different management, and to identify Phytophthora species associated with root rot. Soil and root samples were collected at monthly intervals from Sour orange and Volkameriana lemon rootstocks during March-July period. The inoculum density of Phytophthora species, and the percentage of infected feeder roots, were estimated using the plate dilution method in conjunction with selective media. Phytophthora isolates were identified according to their morphological characteristics and on the basis of the ITS regions of the rDNA. Phytophthora nicotianae was the predominant isolated species, followed by P. citrophthora and P. palmivora. Phytophthora nicotianae was detected in both nurseries, while P. citrophthora and P. palmivora were recovered only in one nursery. Inoculum density of Phytophthora species fluctuated during spring and summer according to the environmental conditions, rootstock, and nursery management practices

Comparison of the performance of the main real-time and conventional PCR detection tests for ‘Candidatus Liberibacter’ spp., plant pathogenic bacteria causing the Huanglongbing disease in Citrus spp

23 Pág.Huanglongbing disease affects the Rutaceae family and is associated with three phloem-limited bacterium species: Candidatus Liberibacter asiaticus, africanus and americanus. These species are considered quarantine pathogens in the world, and pose major risks for citrus production and industry. Due to the low titer and the uneven distribution of the bacteria within its host plant, conventional PCR detection protocols can lead to false negative results, especially for early detection. Herein, three real-time PCR diagnostic methods recommended by the EPPO and FAO for asiaticus and africanus species detection were evaluated for their performance and compared with a conventional duplex PCR. Assessments were done as part of an international cooperative project under the EUPHRESCO guidance. Intra-laboratory assessment of the analytical specificity and analytical sensitivity was performed on 33 target or non-target DNA samples and seven target DNA samples were used to determine the sensitivity. Thereafter, repeatability, reproducibility, and concordance odds ratio were assessed on 20 target or non-target DNA samples through a collaborative test performance study organized among eight international laboratories. Results showed that the Li protocol proved to be the best method for asiaticus and africanus species detection, along with the conventional duplex PCR; whereas the Morgan protocol showed high performance only for asiaticus species. Interlaboratory reproducibility was high, suggesting that these real-time PCR methods can be readily transferred to diagnostic laboratories.This research was funded by Anses - Plant Health Laboratory (LSV).Peer reviewe

Structure analysis of the ribosomal intergenic spacer region of Phaeoacremonium italicum as a study model

Increasing recognition of novel Phaeoacremonium species, and their recent taxonomic reassignment through phylogeny based on the β-tubulin and actin genes, have highlighted the presence of paraphyly, intraspecific variation, and incongruence of some Phaeoacremonium species. This study investigated the intergenic spacer rDNA regions of a representative collection of 31 Phaeoacremonium italicum strains, and compared their structures with those of the closest related species, Phaeoacremonium alvesii and Phaeoacremonium rubrigenum. These intergenic spacer sequences had five categories of repeat elements that were organised into distinct patterns. Morphological analyses of the P. italicum strains provided a more detailed description of P. italicum. The phylogenetic tree constructed using the intergenic spacer sequences compared with that obtained by combined analysis of β-tubulin and actin sequences indicated that the intergenic spacer rDNA region distinguished intraspecific and interspecific variations. Further molecular studies are required to determine whether intergenic spacer sequences can improve precision in defining Phaeoacremonium phylogeny, and prevent misidentification and the introduction of vague species boundaries for the genus

Antagonism and Antimicrobial Capacity of Epiphytic and Endophytic Bacteria against the Phytopathogen Xylella fastidiosa

Olive quick decline syndrome (OQDS), which is caused by Xylella fastidiosa, poses a severe threat to the agriculture of Mediterranean countries and causes severe damage to the olive trees in Italy. Since no effective control measures are currently available, the objective of this study was the screening of antagonistic bacteria that are potentially deployable as biocontrol agents against X. fastidiosa. Therefore, two approaches were used, i.e., the evaluation of the antagonistic activity of (i) endophytic bacteria isolated from two different cultivars of olive trees (Leccino and Ogliarola salentina) and (ii) epiphytic bacteria isolated from the phyllospheres of different host plant species of X. fastidiosa. In vitro dual culture tests showed that 12 out of 200 isolates inhibited X. fastidiosa growth, with appearances of clear zones between 4.0 and 38.6 mm. 16S rRNA gene sequencing revealed different species of the genera Paenibacillus, Bacillus, Pantoea, Microbacterium, Stenotrophomonas, Delftia, and Pseudomonas. Furthermore, an investigation for antimicrobial activity identified 5 out of the 12 antagonistic bacteria, Paenibacillus rigui, Bacillus subtilis, Bacillus pumilus, Microbacterium oxydans, and Stenotrophomonas rhizophila, that were able to produce culture filtrates with inhibitory activities. Our results are promising for further investigation to develop an eco-sustainable strategy to control X. fastidiosa using biocontrol agents or their secreted metabolites

Effectiveness of Low Copper-Containing Chemicals against Olive Leaf Spot Disease Caused by Venturia oleaginea

The high level of copper (Cu) accumulation in the soil, the risk of surface water contamination, and the potential public health problems due to Cu entering the food chain have raised concerns on the use of Cu compounds in agriculture, including olive growing. As a consequence, there is worldwide regulatory pressure on agricultural systems to limit the use of Cu compounds. Hence, a field trial was carried out to evaluate the effectiveness of low copper containing chemicals to control olive leaf spot (OLS) disease caused by Venturia oleaginea. The trial was conducted in 2021 in an olive (cv. Nabali Baladi) grove in Palestine. Copper complexed with lignosulphonate and gluconate (Disper Cu Max®) and the self-defense inducer Disper Broton GS® were evaluated and compared to dodine and the traditionally and frequently used copper hydroxide. In addition, untreated trees were used as the control. Treatments were made in March, July, and August. In March 2021, leaves grown in 2020 were present and 100% infected. V. oleaginea infections caused defoliation in untreated and treated olive trees with varying degrees of intensity: the Control had the most defoliation, followed by copper hydroxide and Disper Cu Max®, whereas dodine and, in particular, Disper Broton GS® had the least. All treatments reduced symptomatic leaves but their efficacy varied significantly: copper hydroxide was the least effective, Disper Cu Max® was intermediate, dodine and, mainly, Disper Broton GS® were the most effective. Overall, the results are promising since Disper Cu Max® and Disper Broton GS® were able to significantly reduce OLS damage and the amount of copper used for treatments

Detection of Erwinia amylovora in pear leaves using a combined approach by hyperspectral reflectance and nuclear magnetic resonance spectroscopy

Erwinia amylovora infections on pear leaves were studied using nuclear magnetic resonance and hyperspectral reflectance spectroscopies. Inoculated pear plants under controlled conditions were used for comparing Erwinia amylovora infected leaves with those infected by Pseudomonas syringae pv. syringae or non-infected controls. Hyperspectral reflectance-NMR covariance analysis allowed the transfer of metabolome information obtained by NMR to hyperspectral reflectance bands through a knowledge transfer approach. At 20 d after Erwinia amylovora inoculation, correlation was found between the NMR signal at 1.16 ppm (attributed to the methyl group of a fucosyl-containing polysaccharide, identified as a specific metabolite from Erwinia amylovora) and a hyperspectral reflectance band centred at 1400 nm. At 50 d after inoculation the same marker metabolite was correlated to hyperspectral reflectance bands centred at 850 nm and 1050 nm. These methods allow maps to be developed which represent the specific infection status of pear plants, and could facilitate development of simple, fast and affordable hyperspectral reflectance-based devices for the detection of Erwinia amylovora infections on pear leaves