An Annotated List of Legume-Infecting Viruses in the Light of Metagenomics.

Legumes, one of the most important sources of human food and animal feed, are known to be susceptible to a plethora of plant viruses. Many of these viruses cause diseases which severely impact legume production worldwide. The causal agents of some important virus-like diseases remain unknown. In recent years, high-throughput sequencing technologies have enabled us to identify many new viruses in various crops, including legumes. This review aims to present an updated list of legume-infecting viruses. Until 2020, a total of 168 plant viruses belonging to 39 genera and 16 families, officially recognized by the International Committee on Taxonomy of Viruses (ICTV), were reported to naturally infect common bean, cowpea, chickpea, faba-bean, groundnut, lentil, peas, alfalfa, clovers, and/or annual medics. Several novel legume viruses are still pending approval by ICTV. The epidemiology of many of the legume viruses are of specific interest due to their seed-transmission and their dynamic spread by insect-vectors. In this review, major aspects of legume virus epidemiology and integrated control approaches are also summarized.

Virus Surveys in Olive Orchards in Greece Identify Olive Virus T, a Novel Member of the Genus Tepovirus.

Field surveys were conducted in Greek olive orchards from 2017 to 2020 to collect information on the sanitary status of the trees. Using a high-throughput sequencing approach, viral sequences were identified in total RNA extracts from several trees and assembled to reconstruct the complete genomes of two isolates of a new viral species of the genus Tepovirus (Betaflexiviridae), for which the name olive virus T (OlVT) is proposed. A reverse transcription-polymerase chain reaction assay was developed which detected OlVT in samples collected in olive growing regions in Central and Northern Greece, showing a virus prevalence of 4.4% in the olive trees screened. Sequences of amplified fragments from the movement-coat protein region of OlVT isolates varied from 75.64% to 99.35%. Three olive varieties (Koroneiki, Arbequina and Frantoio) were infected with OlVT via grafting to confirm a graft-transmissible agent, but virus infections remained latent. In addition, cucumber mosaic virus, olive leaf yellowing-associated virus and cherry leaf roll virus were identified.

Molecular analysis of Greek isolates of cucumber mosaic virus from vegetables shows a low prevalence of satellite RNAs and suggests the presence of host-associated virus strains.

Cucumber mosaic virus (CMV) is a generalist pathogen that infects many economically important crops in Greece. The present study was designed to evaluate the genetic variability of Greek CMV isolates in combination with their satellite RNAs (satRNAs). To achieve this goal, 77 CMV isolates were collected from symptomatic Greek vegetables, mainly tomatoes and cucurbits, alongside their neighboring crops, during a four-year period from 2015 to 2018. Phylogenetic analysis of a partial coat protein (CP) gene segment revealed that all of the isolates belong to CMV subgroups IA and IB and that they are closely related to previously reported Greek isolates. It should be noted, however, that the latter mainly included tomato isolates. Network analysis of the evolutionary relationships among the CP sequences of the Greek isolates in comparison to the corresponding sequences obtained from the GenBank database indicated two predominant common ancestors and at least three differentiated peripherals, and possibly host-associated (tomatoes, legumes, cucurbits) haplogroups (strain groups). More specifically, host-adaptive evolution can be postulated regarding the tomato isolates in subgroup IB. Necrogenic or non-necrogenic satRNAs were detected in four samples from tomato and melon, and this is the first report of non-necrogenic satRNAs in CMV in Greece.

Assessment of Genetic Variability of Citrus tristeza virus by SSCP and CE-SSCP.

Single-strand conformation polymorphism (SSCP) is a popular method used to study the genetic heterogeneity and population variability of Citrus tristeza virus (CTV) isolates. It is a simple, low-cost, and highly specific method for mutation detection of specific genes, mostly of the CTV major coat protein gene (p25). The technique is based on a comparison on polyacrylamide gel of electrophoretic profiles of single-stranded (ss) DNA sequences in terms of their spatial conformation. SSCP involves cDNA synthesis and amplification of the target gene, denaturation of single strands, and electrophoresis in non-denaturing conditions. The ssDNAs can be afterward visualized by staining the polyacrylamide gel. Alternatively, using fluorescently labeled primers, the procedure can be performed in automated sequencers equipped with an appropriate capillary (CE-SSCP), which increases the potential of high-throughput analysis, precision, and the reproducibility of results. CE-SSCP can be also directly applied to the virus particles obtained by elution from ELISA plates or tissue-print membranes.

The Westward Journey of Alfalfa Leaf Curl Virus.

Alfalfa leaf curl virus (ALCV), which causes severe disease symptoms in alfalfa (Medicago sativa L.) and is transmitted by the widespread aphid species, Aphis craccivora Koch, has been found throughout the Mediterranean basin as well as in Iran and Argentina. Here we reconstruct the evolutionary history of ALCV and attempt to determine whether the recent discovery and widespread detection of ALCV is attributable either to past diagnostic biases or to the emergence and global spread of the virus over the past few years. One hundred and twenty ALCV complete genome sequences recovered from ten countries were analyzed and four ALCV genotypes (ALCV-A, ALCV-B, ALCV-C, and ALCV-D) were clearly distinguished. We further confirm that ALCV isolates are highly recombinogenic and that recombination has been a major determinant in the origins of the various genotypes. Collectively, the sequence data support the hypothesis that, of all the analyzed locations, ALCV likely emerged and diversified in the Middle East before spreading to the western Mediterranean basin and Argentina.

Cucumber mosaic virus Isolates from Greek Legumes are Associated with Satellite RNAs that are Necrogenic for Tomato.

Worldwide, Cucumber mosaic virus (CMV) is the causal agent of many economically important diseases. Based on immunological or molecular analysis, three distinct subgroups of CMV isolates can be identified (IA, IB, and II). In addition, some CMV isolates are associated with satellite RNAs (satRNAs), a type of noncoding transcript that may alter the symptoms of CMV infections. This study presents an analysis of CMV isolates occurring in legumes in Greece in respect to their genetic diversity, and the presence and diversity of their satRNA. Phylogenetic analysis of the CMV coat protein sequence of 18 legume and 5 tomato CMV isolates collected throughout Greece classified them within subgroups IA and IB, with a limited genetic diversity. The CMV satRNAs found in nine field legumes exhibiting mild symptoms and in one tomato with a necrotic syndrome contained a functional necrogenic motif; therefore, they were grouped within the necrogenic group of CMV-satRNAs. The necrotic phenotype was expressed in all legume CMV isolates containing necrogenic satRNAs when mechanically inoculated onto tomato plants. To our knowledge, this is the first observation that legumes host necrogenic CMV-satRNAs. The possible role of legumes in the epidemiology of CMV and necrogenic satRNA complex is discussed.

Transmission of Moroccan watermelon mosaic virus (MWMV) by Aphids in Greece.

The aphid-transmitted Moroccan watermelon mosaic virus (MWMV; Potyvirus, Potyviridae) is an emerging pathogen in cucurbit crops in the Mediterranean basin but information on its transmitting vector species is limited. This study aimed to record the competence of 22 species of the Greek aphid fauna to vector MWMV. Timed-probe transmission experiments and arena tests were performed using laboratory colonies of aphid species abundant in field surveys; less common species were tested as apterous individuals collected directly from field plants in mass-inoculation (nonpersistent) tests. Depending on the test, aphids were tested in cohorts of 10 or 20 individuals on zucchini (Cucurbita pepo L.) plants and the frequency of transmission was calculated for a single aphid. Among 12 species tested in timed-probe transmission tests, Myzus persicae nicotianae (74.0%) appeared to be the most efficient vector, followed by M. persicae (48.0%), Aphis gossypii (11.8%), an unidentified Aphis sp. (11.8%), and A. spiraecola (11.0%). Alatae of nine species tested in arena tests transmitted the virus in rates varying from 0.7 to 53.6%; M. persicae was the most efficient species in virus spread. In mass-inoculation tests, the probability that apterae of 12 aphid species collected from field plants transmitted MWMV fluctuated from 0.3 to 5.3%. No transmission was obtained by Brevicoryne brassicae. The following species are reported as new vectors of MWMV: A. fabae, A. nerii, A. spiraephaga, A. umbrella, Capitophorus eleaegni, Dysaphis (Pomaphis) pyri, Macrosiphoniella sanborni, Macrosiphum rosae, Myzocallis castanicola, Myzus persicae nicotianae, M. cerasi, M. varians, Phorodon humuli, Ovatus crataegarius, Takecallis arundicolens (first report in Greece), Uroleucon sonchi, and U. (Uromelan) aeneum. These results contribute to a better understanding of MWMV epidemiology.

The Efficiency by Which Thrips tabaci Populations Transmit Tomato spotted wilt virus Depends on Their Host Preference and Reproductive Strategy.

ABSTRACT Arrhenotokous and thelytokous populations of Thrips tabaci from tobacco or leek plants were evaluated for their ability to transmit Tomato spotted wilt virus (TSWV) and for their host preference. Transmission efficiencies were comparatively studied using leaf disks of Petunia hybrida, Datura stramonium, and Nicotiana tabacum cv. Basmas. Adults of arrhenotokous populations collected on infected tobacco plants in the field were efficient transmitters (up to 48.5% transmission) and remained so when maintained on tobacco for several generations. Arrhenotokous T. tabacipopulations from leek plants were poor transmitters (up to 3.1% transmission), whereas no transmission was obtained with thelytokous populations from leek. All populations could infest leek, however none of the arrhenotokous and thelytokous populations from leek plants was able to infest tobacco. TSWV could be acquired by both first and second larval instars of a T. tabacipopulation from tobacco. However, the transmission by adults decreased with the age at which the virus was acquired by larvae. The highest efficiencies (61% of males and 51% of females transmitted) were obtained when newborn (0- to 24-h old) larvae acquired the virus. The majority of thrips started to transmit after becoming adult and rates were positively correlated with the temperature at which the thrips were kept. The median latent period values found for adults decreased with increasing temperature. The median acquisition access period (AAP50) of the population was 41 min, whereas the AAP(50) was 65 min for males and 35 min for females. The median inoculation access period of males was 246 and 365 min on tobacco and petunia, respectively, and 96 and 345 min for females. The results show that T. tabaci forms a complex in terms of host preference, reproductive strategy, and ability to transmit TSWV. The transmission parameters show that the thrips of arrhenotokous populations infesting tobacco are highly efficient vectors.