An epidemic of almond witches'-broom has devastated almond production in Lebanon. Thousands of almond trees have died over the past 10 years due to the rapid spread of the disease. The symptoms, which include early flowering, stunted growth, leaf rosetting, dieback, off-season growth, proliferation of slender shoots, and witches'-brooms arising mainly from the main trunk and roots, resemble those caused by phytoplasmal infections. For the detection of the putative causal agent, nested polymerase chain reaction (PCR) was performed using universal primers (P1/P7, R16mF2/R16mR1, and R16F2n/R16R2) commonly used for the specific diagnosis of plant pathogenic phytoplasmas. Phytoplasmas were readily detected from infected trees with witches'-broom symptoms collected from three major almond growing regions in Lebanon. Restriction fragment length polymorphism (RFLP) analysis of PCR products amplified by the primer pair R16F2n/R16R2 revealed that the phytoplasma associated with infected almonds is similar to, but distinct from, members of the pigeon pea witches'-broom phytoplasma group (16SrIX). A new subgroup, 16SrIX-B, was designated. Sequencing of the amplified products of the phytoplasma 16S rRNA gene indicated that almond witches'-broom (AlmWB) phytoplasma is most closely related to members of the pigeon pea witches'-broom phytoplasma group (with sequence homology ranging from 98.4 to 99.0%). Phylogenetic analysis of 16S rDNA sequences from AlmWB phytoplasma and from representative phytoplasmas from GenBank showed that the AlmWB phytoplasma represents a distinct lineage within the pigeon pea witches'-broom subclade. The same phytoplasma appears also to infect peach and nectarine seedlings.

Petroleum ether (PE) and methanolic extracts of nine wild plant species were tested in vitro for their antimycotic activity against eight phytopathogenic fungi. The efficacy of PE extracts against all pathogens tested was higher than that of methanolic extracts. Wild marjoram (Origanum syriacum) PE extract showed the highest and widest range of activity. It resulted in complete inhibition of mycelial growth of six of eight fungi tested and also gave nearly complete inhibition of spore germination of the six fungi included in the assay, namely, Botrytis cinerea, Alternaria solani, Penicillium sp., Cladosporium sp., Fusarium oxysporum f. sp. melonis, and Verticillium dahlia. The other plant extracts showed differential activities in the spore germination test, but none was highly active against mycelial growth. Inula viscosa and Mentha longifolia were highly effective (>88%) in spore germination tests against five of six fungi tested, whereas Centaurea pallescens, Cichorium intybus, Eryngium creticum, Salvia fruticosa, and Melia azedarach showed >95% inhibition of spore germination in at least two fungi. Foeniculum vulgare showed the least antimycotic activity. Fractionation followed by autobiography on TLC plates using Cladosporium sp. as a test organism showed that O. syriacum PE extracts contained three inhibition zones, and those of Inula viscosa and Cichorium intybus, two, whereas the PE extracts of the remaining plants showed each one inhibition zone. Some of the major compounds present in these inhibition zones were identified by GC-MS. The possibility for using these extracts, or their mixtures, to control plant diseases is discussed.

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BackgroundIn the early 2000s, two cucurbit-infecting begomoviruses were introduced into the eastern Mediterranean basin: the Old World Squash leaf curl virus (SLCV) and the New World Watermelon chlorotic stunt virus (WmCSV). These viruses have been emerging in parallel over the last decade in Egypt, Israel, Jordan, Lebanon and Palestine.MethodsWe explored this unique situation by assessing the diversity and biogeography of the DNA-A component of SLCV and WmCSV in these five countries.ResultsThere was fairly low sequence variation in both begomovirus species (SLCV π = 0.0077; WmCSV π = 0.0066). Both viruses may have been introduced only once into the eastern Mediterranean basin, but once established, these viruses readily moved across country boundaries. SLCV has been introduced at least twice into each of all five countries based on the absence of monophyletic clades. Similarly, WmCSV has been introduced multiple times into Jordan, Israel and Palestine.ConclusionsWe predict that uncontrolled movement of whiteflies among countries in this region will continue to cause SLCV and WmCSV migration, preventing strong genetic differentiation of these viruses among these countries.

In the summer of 2011, whiteflies (Bemisia tabaci) were collected from cucumbers plants showing interveinal yellowing on the lower leaves growing in commercial greenhouses in Jiyye area, South Lebanon. About twenty whiteflies per seedling were placed on healthy cucumber (Cucumis sativus cv. Beit alpha) seedlings at the second-leaf stage and grown in insect-proof cages in the university glasshouse. After 4 days of feeding, the whiteflies were sprayed with an insecticide (Imidacloprid). Three weeks post inoculation, three out of eight inoculated plants developed typical yellowing symptoms on old leaves. Symptoms started with chlorotic spots that later coalesced into interveinal chlorosis similar to that induced by Cucurbit yellow stunting disorder virus (CYSDV), a widely occurring virus in Lebanon. Total RNA was extracted from the three symptomatic plants using TRI reagent (Sigma-Aldrich, St Louis, MO). The three symptomatic plants, however, tested negative for CYSDV by tissue blot immuno assay and reverse transcription (RT)-PCR (2). Another Crinivirus, Cucurbit chlorotic yellows virus (CCYV), was suspected (1). Analysis of the RNA extracts of the three symptomatic plants by RT-PCR using the CCYV heat shock protein (HSP70h) specific primer pair (CCYV-HSP-F1/CCYV-HSP-R1) (3) showed a band of the expected size (462 bp). A small scale survey from February to April 2012, covering the two major cucumber growing areas on the Lebanese coastal areas, Byblos (North) and Jiyye (South), was conducted. Eight out of 10 cucumber leaf samples collected from Jiyye and seven out of 38 from Byblos area tested positive for CCYV by RT-PCR. Therefore, RNA extracts from cucumber which were positive for CYSDV and stored at –80°C since spring 2011 were tested again for CCYV. Fourteen randomly selected RNA extracts out of 76 samples were all found to have mixed infections with CCYV. For partial molecular characterization of the Lebanese CCYV isolates collected during summer 2011, regions of the HSP70h and coat protein (CP) genes from CCYV were amplified using specific primers (CCYV-CPs/CCYV-CPas) (1, 3). Sequences of the purified amplicons of 462 and 725 bp respectively, were submitted to GenBank (Accession Nos. JX014262 and JX014263). The HSP70h and CP genes were highly homologous to the Japanese CCYV isolate (AB523789), sharing nucleotide sequence identities of 99.8% and 99.7%, respectively. The CCYV HSP70h gene of the Lebanese isolate is distantly related to other criniviruses sharing nucleotide sequence identities of 76.4, 75.8, and 74.7% with Lettuce chlorosis virus (FJ380119), Bean yellow disorder virus (EU191905), and CYSDV (AY242078), respectively, whereas the CCYV CP gene shares 77, 76, and 70% nucleotide sequence identities with the latter viruses, respectively. CCYV has been reported only in Japan, China, Taiwan, and Sudan (1). To our knowledge, this is the first report of CCYV in Lebanon and the Mediterranean basin. CCYV may have also spread to neighboring countries but has not been reported yet. The rapid outbreak of new whitefly-transmitted viruses poses a serious threat to cucurbit production in the Mediterranean region and Europe. References: (1) K. Hamed et al. Plant Dis. 95:1321, 2011. (2) H. Hourani and Y. Abou-Jawdah J. Plant Pathol. 85:1, 2003 (3) R. Zeng et al. Plant Dis. 95:354, 2011.

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Cucurbit yellow stunting disorder virus (CYSDV), genus Crinivirus and family Closteroviridae, has emerged as a serious whitefly-transmitted virus of cucurbit crops, causing between 30 and 50% yield losses. Development of resistant cultivars represents an economically and environmentally sound approach to management of this disease. In all, 124 cucumber accessions were evaluated for reaction to CYSDV under high inoculum pressure over three growing seasons. Seven accessions showed delayed expression of symptoms, milder final symptoms, and lower percentages of infected plants compared with susceptible cucumbers. Although none of these accessions were immune to CYSDV, virus concentrations in the middle leaves of the tolerant accessions were significantly lower than those of susceptible accessions.

BackgroundAlmond witches’-broom (AlmWB), a devastating disease of almond, peach and nectarine in Lebanon, is associated with ‘Candidatus Phytoplasma phoenicium’. In the present study, we generated a draft genome sequence of ‘Ca. P. phoenicium’ strain SA213, representative of phytoplasma strain populations from different host plants, and determined the genetic diversity among phytoplasma strain populations by phylogenetic analyses of 16S rRNA, groEL, tufB and inmp gene sequences.ResultsSequence-based typing and phylogenetic analysis of the gene inmp, coding an integral membrane protein, distinguished AlmWB-associated phytoplasma strains originating from diverse host plants, whereas their 16S rRNA, tufB and groEL genes shared 100 % sequence identity. Moreover, dN/dS analysis indicated positive selection acting on inmp gene. Additionally, the analysis of ‘Ca. P. phoenicium’ draft genome revealed the presence of integral membrane proteins and effector-like proteins and potential candidates for interaction with hosts. One of the integral membrane proteins was predicted as BI-1, an inhibitor of apoptosis-promoting Bax factor. Bioinformatics analyses revealed the presence of putative BI-1 in draft and complete genomes of other ‘Ca. Phytoplasma’ species.ConclusionThe genetic diversity within ‘Ca. P. phoenicium’ strain populations in Lebanon suggested that AlmWB disease could be associated with phytoplasma strains derived from the adaptation of an original strain to diverse hosts. Moreover, the identification of a putative inhibitor of apoptosis-promoting Bax factor (BI-1) in ‘Ca. P. phoenicium’ draft genome and within genomes of other ‘Ca. Phytoplasma’ species suggested its potential role as a phytoplasma fitness-increasing factor by modification of the host-defense response.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-015-0487-4) contains supplementary material, which is available to authorized users.