Two primers, specific for Phytophthora nicotianae (Pn6) and P. citrophthora (Pc2B), were modified to obtain Scorpion primers for real-time identification and detection of both pathogens in citrus nursery soils and roots. Multiplex PCR with dual-labelled fluorogenic probes allowed concurrent identification of both species of Phytophthora among 150 fungal isolates, including 14 species of Phytophthora. Using P. nicotianae specific primers a delayed and lower fluorescence increase was also obtained from P. cactorum DNA. However, in separate real-time amplifications, the aspecific increase of fluorescence from P. cactorum was avoided by increasing the annealing temperature. In multiplex PCR, with a series of 10-fold DNA dilutions, the detection limit was 10 pg ll )1 for P. nicotianae and 100 pg ll )1 for P. citrophthora, whereas in separate reaction DNA up to 1 pg ll )1 was detected for both pathogens.Simple and rapid procedures for direct DNA extraction from soil and roots were utilised to yield DNA whose purity and quality was suitable for PCR assays. By combining these protocols with a double amplification (nested Scorpion-PCR) using primers Ph2-ITS4 amplifying DNA from the main Phytophthora species (first round) and PnB5-Pn6 Scorpion and Pc2B Scorpion-Pc7 (second round), it was possible to achieve real-time detection of P. nicotianae and P. citrophthora from roots and soil. The degree of sensitivity was similar to that of traditional detection methods based on the use of selective media. The analyses of artificially and naturally infested soil showed a high and significant correlation between the concentration of pathogen propagules and the real-time PCR cycle threshold.

A study on seasonal abundance of Auchenorrhyncha species and their infectivity by Xylella fastidiosa in the Apulia region of Italy was conducted to identify ideal periods for monitoring and adoption of potential control measures against insect vectors. Adult populations of Auchenorrhyncha species were monitored monthly over a 2-yr period from five olive groves. A total of 15 species were captured, identified, and tested for presence of X. fastidiosa by polymerase chain reaction (PCR). For three species, Philaenus spumarius L., Neophilaenus campestris (Fallèn), and Euscelis lineolatus Brullé, positive reactions to X. fastidiosa were obtained, on average, in 16.3, 15.9 and 18.4% of adult insects, respectively. Philaneous spumarius was the dominant species (39.8% of total Auchenorrhyncha captured) with the highest adult abundance in summer months. Adult P. spumarius and N. campestris were first detected between March and May in both years, and all insects tested during these periods (year 1: n = 42, year 2: n = 132) gave negative reactions to X. fastidiosa by PCR. Similarly, first adults of E. lineolatus that appeared from October to November (year 1: n = 20, year 2: n = 15) tested negative for presence of X. fastidiosa Given the lack of transstadial and transovarial transmission of X. fastidiosa and considering that P. spumarius is univoltine, control measures against nymphal stages of P. spumarius should be investigated as means of population suppression to reduce spread of X. fastidiosa in olive groves.

Real-time loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) assays were developed targeting the internal transcribed spacer 2 region of the ribosomal DNA of Phytophthora infestans, the potato late blight causal agent. A rapid crude plant extract (CPE) preparation method from infected potato leaves was developed for on-site testing. The assay’s specificity was tested using several species of Phytophthora and other potato fungal and oomycete pathogens. Both LAMP and RPA assays showed specificity to P. infestans but also to the closely related species P. andina, P. mirabilis, P. phaseoli, and P. ipomoeae, although the latter are not reported as potato pathogen species. No cross-reaction occurred with P. capsici or with the potato pathogens tested, including P. nicotianae and P. erythroseptica. The sensitivity was determined using P. infestans pure genomic DNA added into healthy CPE samples. Both LAMP and RPA assays detected DNA at 50 fg/μl and were insensitive to CPE inhibition. The isothermal assays were tested with artificially inoculated and naturally infected potato plants using a Smart-DART platform. The LAMP assay effectively detected P. infestans in symptomless potato leaves as soon as 24 h postinoculation. A rapid and accurate on-site detection of P. infestans in plant material using the LAMP assay will contribute to improved late blight diagnosis and early detection of infections and facilitate prompt management decisions.

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Faleiro, J.R., M. Ferry, Th. Yaseen and S. Al-Dobai. 2019. Overview of the gaps, challenges and prospects of red palm weevil management. Arab Journal of Plant Protection, 37(2): 170-177. The Red Palm Weevil (RPW) Rhynchophorus ferrugineus Olivier has emerged as a key pest of palms in diverse agro-ecosystems worldwide. RPW has its home in South and SouthEast Asia where it has been a major pest of coconut. Ever since it was reported on date palm in the Middle East during the mid-1980s, it has spread rapidly mainly through infested planting material. Recent reports of RPW invasion suggest that the pest is gaining foot hold in the Caucasian region where it is detected from Sochi in Russia and Abkhazia in the republic of Georgia and also from East Africa in Djibouti. The current RPW IPM programmes, based on pheromone/bait trapping among other techniques have been implemented with limited success. Gaps and challenges in almost all the components of the strategy, particularly with regard to early detection of the pest, developing and implementing phytosanitary measures, lack of farmer participation in the programmes and scarcity of data on socioeconomic issues among several other factors have made RPW control and eradication extremely difficult. On the positive side, the pest has been eradicated in the Canary Islands and is approaching eradication in Mauritania. Eradication has also been obtained in various oasis in Oman but new introductions of infested palms have ruined these successes. The Food and Agriculture Organization of the UN during the Scientific and High Level Meeting on the Management of RPW in March, 2017 called for the urgent need to combat RPW by collaborative efforts and commitments at the country, regional and global levels to stop the spread of this devastating pest and formulated a framework strategy for eradication of RPW which aims to support efforts/programs of countries to stop its spread, to achieve a strong decline and if possible its eradication. This has led to the 'FAO Programme on RPW Eradication in the NENA Region' to intensify governance, monitoring, scientific research, capacity building and coordination. The program fosters the ongoing research on the applicable approaches of biological control and innovative detection and control methods. Furthermore, the 'FAO Global RPW management platform' aims mainly at monitoring the pest using mobile apps and GIS based techniques. This presentation highlights the gaps and challenges in the current RPW-IPM strategy with prospects for improving each component of the RPW-IPM program, based on a much better knowledge on the socioeconomic situation and the participation of the farmers and other stakeholders.

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The serious damages of Xylella fastidiosa (Xf) in Euro-Mediterranean countries (Italy, France, Spain) raise concerns for the Near East and North Africa (NENA). Therefore, a study was performed to: a) assess the risk of Xf entry, establishment and spread in target NENA countries (viz. Algeria, Egypt, Jordan, Lebanon, Libya, Morocco, Palestine, Syria, Tunisia); b) analyse risk perception and preparedness level among agri-food chain stakeholders; c) estimate potential socio-economic impacts for olives, grapes and citrus. Pest risk appraisal suggests that Morocco, Lebanon, Palestine and Syria are the most exposed to Xf risk; other target NENA countries, except Algeria, have intermediate risk. Risk perception analysis shows that governance efficacy and practices application can be improved by involving stakeholders and raising their awareness. Socio-economic impact assessment indicates declining yields, production, profitability, export, employment, and increasing import, with the highest impacts relating to olives, then citrus and grapes. The study suggests that the expected socioeconomic impacts are unacceptable and require urgent action against Xf at national and regional levels.