Towards dissecting the structural determinant of Peach latent mosaic viroid inducing mosaic symptoms

Most isolates of Peach latent mosaic viroid (PLMVd) do not incite foliar symptoms, but a few number of isolates cause peach mosaic (PM) or peach calico (PC), an extreme albino phenotype. The PC determinant has been previously mapped at an insertion of 12-13 nt folding into a hairpin capped by a U-rich loop but the PM determinant, which is not associated with a specific insertion, remains unidentified and could reside in one or more domains of the branched conformation proposed for PLMVd. To tackle this problem we have selected for further dissection one variant (GDS6), recovered from a typical PM isolate (GDS), which is very infectious and elicits consistently a characteristic PM. We have initially focused on G337, a position that appears associated with PM in multiple alignments that include GDS6 and other PM-inducing and latent variants. To determine the role of G337 in infectivity and symptoms, GF-305 peach seedlings were inoculated with in vitro transcripts of recombinant plasmids containing dimeric tandem inserts of PLMVd-cDNAs with all possible changes at this position introduced by site-directed mutagenesis. Deletion of G337 abolished infectivity, while substitutions by A, C or U incited, in most inoculated plants, PM symptoms. Cloning and sequencing showed that the A substitution at position 337 was preserved in the progeny or reverted to G, while C or U substitutions at this position were not stable and reverted to A or G in the progenies. Extending this approach to additional nucleotides of loop A, or of other PLMVd domains, may provide hints in identifying the determinant of PM. Keywords: Viroids, Pathogenesis, Peach diseas

Pome fruit viruses in Bosnia and Herzegovina

During autumn 2005 and summer 2006, field surveys were carried out to assess the sanitary status of pome fruit trees in Bosnia and Herzegovina. Inspections were done in the main pome fruit growing areas including 10 orchards, 2 nurseries and one varietal collection. A total of 65 apple and 50 pear cultivars were tested by biological indexing for the presence of Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV) and Apple mosaic virus (ApMV). The average infection level was 81%. Both species showed a similar infection rate (83% for apple and 78% for pear). The most frequent viruses of apple were ACLSV (72%) and ASPV (69%), and of pear ASGV (69%) and ACLSV (64%). The same samples were also tested by ELISA, with a lower virus detection rate compared to the biological indexing. Multiplex RT-PCR results of 20 randomly selected apple cultivars were in line with biological indexing. Results of our surveys report for the first time th e presence of ACLSV, ASPV, ASGV and ApMV on pome fruits in Bosnia and Herzegovina.Keywords: Malus, biological indexing, ELISA, multiplex RT-PCR, sanitary statu

Identification and characterization of citrus concave gum-associated virus infecting citrus and apple trees by serological, molecular and high-throughput sequencing approaches

Citrus concave gum-associated virus (CCGaV) is a negative-stranded RNA virus, first reported a few years ago in citrus trees from Italy. It has been reported in apple trees in the USA and in Brazil, suggesting a wider host range and geographic distribution. Here, an anti-CCGaV polyclonal antiserum to specifically detect the virus has been developed and used in a standard double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) that has been validated as a sensitive and reliable method to detect this virus both in citrus and apple trees. In contrast, when the same antiserum was used in direct tissue-blot immunoassay, CCGaV was efficiently detected in citrus but not in apple. Using this antiserum, the first apple trees infected by CCGaV were identified in Italy and the presence of CCGaV in several apple cultivars in southern Italy was confirmed by field surveys. High-throughput sequencing (HTS) allowed for the assembling of the complete genome of one CCGaV Italian apple isolate (CE-c3). Phylogenetic analysis of Italian CCGaV isolates from apple and citrus and those available in the database showed close relationships between the isolates from the same genus (Citrus or Malus), regardless their geographical origin. This finding was further confirmed by the identification of amino acid signatures specific of isolates infecting citrus or apple hosts. Analysis of HTS reads also revealed that the CE-c3 Italian apple tree, besides CCGaV, was simultaneously infected by several viruses and one viroid, including apple rubbery wood virus 2 which is reported for the first time in Italy. The complete or almost complete genomic sequences of the coinfecting agents were determined

Identification, full-length genome sequencing, and field survey of citrus vein enation virus in Italy

Citrus vein enation virus (CVEV) was described in Spain and then it has been reported in several citrus growing areas of Asia, America and Australia. Here, the occurrence of CVEV in Italy has been documented for the first time. The full genome sequence of a CVEV Italian isolate (14Q) was determined by high-throughput sequencing and the presence of the virus was confirmed by RT-PCR and graft-transmission to indicator plants, from which the virus was recovered six-months post-inoculation. Phylogenetic analysis based on the full-length genome of CVEV isolates from different countries showed that they are phylogenetically related to each other based on their geographic origin, rather than on their host and that the Italian isolate is more closely related to the Spanish isolate than to the other ones. A field survey revealed the presence of CVEV in some areas of Campania region (southern Italy), prevalently infecting lemon trees. In the frame of this survey, kumquat was identified for the first time as a host of CVEV. No symptoms were observed in the field so far. The infection of asymptomatic hosts and the transmission by aphid species present in Italy increase the risk that the virus could further spread

Root internalization of Salmonella Typhimurium in basil plants

Abstract Background Foodborne pathogens present in soil or irrigation water have the potential to internalize via root into edible parts of the plants and survive, representing a serious hazard for consumer's health (1). The present work assesses the ability of Salmonella Typhimurium (ST) to enter basil plants (Ocimum basilicum L., cultivar "Napoletano") from roots. This situation may represent a risk in food safety since that vegetable is largely used raw/undercooked, in the Mediterranean diet. Methods Sixteen basil plants were cultivated in hydroponics, from the seeds. Roots from 14 plants at 4 leaves-stage were individually immersed in liquid media contaminated with a suspension of ST isolated from Aterno river in Abruzzo region, Italy (9 log10 cfu/mL final concentration). Two plants were used as negative controls (no ST challenge); surface swabs were taken from leaves and stems to exclude external contaminations. After 24 hours, plants were tested for detection (UNI EN ISO 6579-1:2017) and enumeration (in-house method) of ST in leaves and stems. Results ST detection gave positive results in 14/14 plants; in particular, ST was present in 14/14 plants in the leaves and for 9/14 plants also in the stems. The internalized ST strains were also enumerated in 10/14 plants. In particular, 3/14 plants gave positive results both in the leaves and stems, 2/14 only in the stems and other 5/14 only in the leaves. The average ST counts were of 4 log10 cfu/g in leaves and stems. All negative ctr/surface swabs were negative to ST. Conclusions Even though antimicrobial properties of basil plants have been described (2), this study demonstrated the potential of ST to internalize, survive and spread to edible parts. The findings highlight the risk of human infections by ST in plant following root uptake, suggesting the importance of applying appropriate preventive pre-harvest strategies. Biblio EFSA. E. Jour. (2014);12(3):3600 Tenore et al. (2017). Natural product research, 31(17), 2067-2071 Key messages The ability of Salmonella Typhimurium to internalize, survive and spread to edible parts of basil plants. Pathogenic bacteria root uptake represents a potential risk for human health

Close similarities between Cherry chlorotic rusty spot disease from Italy and Cherry leaf scorch from Spain

Cherry chlorotic rusty spot (CCRS), a disease affecting sweet and sour cherry in Southern Italy was regularly found associated with an unidentified fungus and with a complex pattern of viral-like double-stranded RNAs as well as with two small circular RNAs (cherry small circular RNAs, cscRNAs). Further studies revealed that i) the ds-RNAs correspond to the genome of different mycoviruses belonging to the genera Chrysovirus, Partitivirus and Totivirus and ii) the two viroid-like RNAs consist of two groups of variants with similar sequences but differing in size (394–415 and 372–377 nt for cscRNA1 and cscRNA2, respectively). Here we report that the dsRNAs of Chrysovirus and Partitivirus have been detected by RT-PCR analysis with CCRS specific primers in nucleic acid preparations from cherry leaves affected by cherry leaf scorch (CLS) in Spain, a disease whose etiological agent is the ascomycetes Apiognomonia erythrostoma, order Diaporthales. Moreover, Northern-blot hybridization assays showed that a viroid-like RNA comigrating and sharing high sequence similarity with the cscRNA1 previously reported in Italy, accumulate in leaves from CLS affected trees in Spain. These data, together with other evidence showing similar symptoms, disease cycle and fungal fructifications in CCRS and CLS affected trees, suggest a close relationship between the two cherry disorders.Keywords: dsRNAs, cscRNAs, Apiognomonia erythrostoma, Diaporthale

A Feedforward Neural Network Approach for the Detection of Optically Thin Cirrus From IASI-NG

The identification of optically thin cirrus is crucial for their accurate parameterization in climate and Earth's system models. This study exploits the characteristics of the infrared atmospheric sounding interferometer-new generation (IASI-NG) to develop an algorithm for the detection of optically thin cirrus. IASI-NG has been designed for the European Organization for the Exploitation of Meteorological Satellites (EUMETSAT) polar system second-generation program to continue the service of its predecessor IASI from 2024 onward. A thin-cirrus detection algorithm (TCDA) is presented here, as developed for IASI-NG, but also in parallel for IASI to evaluate its performance on currently available real observations. TCDA uses a feedforward neural network (NN) approach to detect thin cirrus eventually misidentified as clear sky by a previously applied cloud detection algorithm. TCDA also estimates the uncertainty of "clear-sky" or "thin-cirrus" detection. NN is trained and tested on a dataset of IASI-NG (or IASI) simulations obtained by processing ECMWF 5-generation reanalysis (ERA5) data with the s-IASI radiative transfer model. TCDA validation against an independent simulated dataset provides a quantitative statistical assessment of the improvements brought by IASI-NG with respect to IASI. In fact, IASI-NG TCDA outperforms IASI TCDA by 3% in probability of detection (POD), 1% in bias, and 2% in accuracy, and the false alarm ratio (FAR) passes from 0.02 to 0.01. Moreover, IASI TCDA validation against state-of-the-art cloud products from Cloudsat/CPR and CALIPSO/Cloud-Aerosol Lidar with Orthogonal Polarization (CALIOP) real observations reveals a tendency for IASI TCDA to underestimate the presence of thin cirrus (POD = 0.47) but with a low FAR (0.07), which drops to 0.0 for very thin cirrus