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This article was supported by the German Research Foundation (DFG) and the Open Access Publication Fund of Humboldt-Universität zu Berlin., After entry of a quarantine/regulated pathogen, infected plants shall be destroyed, and the cultivated area (e.g., greenhouse) shall be disinfected. Therefore, the selection of an effective disinfectant plays an important role. With the availability of different methods for virus quantification, we investigated the application of quantitative ELISA (qELISA), RT-qPCR (reverse transcription-quantitative polymerase chain reaction), and bioassays for the quantification of disinfectant efficacy. Therefore, we estimated the titer reduction in tomato brown rugose fruit virus (ToBRFV), a regulated pathogen, in plant sap and on germ carriers after treatment with MENNO Florades 4% for 16 h. The virus load before and after the treatment was measured with the mentioned methods. The RT-qPCR and qELISA methods showed very low efficacy in the presence of the disinfectant. Although bioassays are time-consuming, need purified particles for establishing the quantification models, and are less sensitive than RT-qPCR, they were able to quantify the differences in virus titer in the presence/absence of disinfectant. Interestingly, the bioassays reached at least the lower limit sensitivity of a qELISA. By being less sensitive to the presence of the disinfectant, bioassays proved to be the only technique for the determination of the disinfectant efficacy against ToBRFV on different germ carriers as well as on virus-infected plant sap., Menno Chemie-Vertriebs GmbH, Agro-Horti Testlabor, Deutsche Forschungsgemeinschaft, Open Access Publication Fund of Humboldt-Universität zu Berlin, Peer Reviewed

The tomato brown rugose fruit virus (ToBRFV) causes severe damage to tomato cultivars and has international economic importance. The harmful tobamovirus is easily mechanically transmissible and highly stable. An ongoing cultivation of infected tomato plants may lead to the spread of ToBRFV in and around the production area of the infested tomato farm. We conducted a study in which we collected a representative number of swab samples from various inanimate surfaces in greenhouses, packaging halls, and shared and private accommodations. In addition, numerous fabrics, such as outer clothing, bed linen, and items used by greenhouse workers, were tested. The infectivity of ToBRFV-contaminated surfaces was tested in bioassays using Nicotiana tabacum cv. Xanthi NN and confirmed using DAS-ELISA. The proportion of ToBRFV-contaminated surfaces varied among locations, from 48.7% in greenhouses to 0% in offices with limited access to staff. Samples from shared accommodation and private accommodation were 18.4% and 3.6% ToBRFV positive, respectively. Clothing and protective items were found to be highly contaminated with ToBRFV, and even around the sleeping area, infective ToBRFV was detected in a few apartments. This study provides evidence for the first time on how and where infectious ToBRFV can be spread by humans beyond the production area. To avoid further dissemination, strict hygiene protocols are required to interrupt transmission routes., German Federal Ministry of Food and Agriculture, Deutsche Forschungsgemeinschaft (DFG, German Research Foundation), Open Access Publication Fund of Humboldt-Universität zu Berlin, Peer Reviewed

Aspen mosaic-associated virus (AsMaV) is a newly identified Emaravirus, in the family Fimoviridae, Bunyavirales, associated with mosaic symptoms in aspen trees (Populus tremula). Aspen trees are widely distributed in Europe and understanding the population structure of AsMaV may aid in the development of better management strategies. The virus genome consists of five negative-sense single-stranded RNA (–ssRNA) molecules. To investigate the genetic diversity and population parameters of AsMaV, different regions of the genome were amplified and analyzed and full-length sequence of the divergent isolates were cloned and sequenced. The results show that RNA3 or nucleoprotein is a good representative for studying genetic diversity in AsMaV. Developed RT–PCR–RFLP was able to identify areas with a higher number of haplotypes and could be applied for screening the large number of samples. In general, AsMaV has a conserved genome and based on the phylogenetic studies, geographical structuring was observed in AsMaV isolates from Sweden and Finland, which could be attributed to founder effects. The genome of AsMaV is under purifying selection but not distributed uniformly on genomic RNAs. Distant AsMaV isolates displayed amino acid sequence variations compared to other isolates, and bioinformatic analysis predicted potential post-translational modification sites in some viral proteins., Einstein Foundation Berlin, COST action CA20132, DFG, German Research Foundation, Open Access Publication Fund of Humboldt-Universität zu Berlin, Peer Reviewed

This article was supported by the German Research Foundation (DFG) and the Open Access Publication Fund of Humboldt-Universität zu Berlin., Due to its global spread, easy mechanical transmissibility inside greenhouses, and detrimental effects on marketability, Tomato brown rugose fruit virus (ToBRFV) is considered to be one of the biggest threats to tomato production. Regarding such crop epidemics, it is essential to identify all conceivable transmission routes and to interrupt them with effective decontamination strategies. We analyzed the potential efficacy of reliable shoe sole cleaning in combination with a disinfection measure. For this purpose, first, a suspension test was undertaken that involved applying different disinfectants to an infected plant homogenate. This was followed by a simulated carrier test. Finally, shoe-sole decontamination was tested under practical conditions. The extent of decontamination was determined by bioassays of the infectivity of the initial load remaining after treatment. Thereby, necrotic local lesions on the susceptible indicator plant Nicotiana tabacum cv. Xanthi NN were counted. Recommendations for practical applications, based on suspension or simulated carrier tests, are limited in their applicability, since very short contact times between ToBRFV and disinfectants reduce efficacy. Under practical conditions, the approved disinfectant MENNO Florades was able to achieve complete inactivation of the virus in the disinfection mat following mechanical depletion from the shoe soles., Peer Reviewed

In order to fill in a decade-long information gap regarding the biological, serological and molecular data for plum tree viruses in Bosnia and Herzegovina, a three-phase study combining symptom evaluation, and serological and molecular assays with high-throughput sequencing (HTS) technology was conducted. The most frequently observed symptoms were discolorations in the form of ring patterns, bands and irregular shapes, as well as vein banding. Sharka-associated symptoms in the form of ring patterns and semicircles were prevalent. A total of 468 plum tree samples were tested by ELISA for the presence of PPV, ApMV, PDV, PNRSV, PBNSPaV, ACLSV and MLRSV. An overall infection incidence of 51.9% was detected, with PPV being the most prevalent (48.7%), followed by PDV (2.99%), PNRSV (0.21%) and mixed infections of PPV+PDV (1.71%). RT-PCR-assisted strain typing in 45 samples revealed PPV-D as the most common strain (22.22%), followed by PPV-REC (6.66%). Mixed infections of PPV-D+PPV-REC were detected (6.66%). HTS enabled the recovery of a 9743 nts long sequence of PPV-D (PPV_O7/80, MW412433), which shared the highest nucleotide and amino acid identities with isolates S13 (LC375131) from Serbia, SVN1 (LC375132) from Slovenia and N9 (LC375129) from Bulgaria. The phylogenetic analysis of the whole genome placed the isolate of the D strain in a distinctive group with the Slovenian isolate SVN1 (LC375132). In addition, the (Cter)NIb/(Nter)CP fragment of a PPV-REC isolate (MW412434) obtained in this survey formed a separate group with previously known isolates from Bosnia and Herzegovina (BOS64Pl and BOS257Pl)., European Cooperation in Science and Technology, Peer Reviewed

This article was supported by the German Research Foundation (DFG) and the Open Access Publication Fund of Humboldt-Universität zu Berlin., The highly infectious Tomato brown rugose fruit virus (ToBRFV) is a new viral threat to tomato production worldwide. In production, the very easy mechanical transmissibility combined with the high resistance in vitro is of great concern. We tested: (i) whether household cleaning products, commercial agricultural detergents, and an authorized plant protectant are suitable for cleaning contaminated clothing, and (ii) whether infectious viruses remain in the resulting cleaning water. The evaluation of the sanitation effect was performed using bioassays, by counting ToBRFV-associated necrotic local lesions on Nicotiana tabacum cv. Xanthi NN. For this purpose, leaves were mechanically inoculated with treated fabrics and cleaning solutions which would normally be discharged to the sewer system. The detergents Fadex H+ (FH) and Menno Hortisept Clean Plus, as well as the disinfectant Menno Florades (MF), led to an almost complete removal of ToBRFV from contaminated fabrics, corresponding to a reduction in local lesions by 99.94–99.96%. In contrast, common household cleaning products (Spee ActivGel (SAG), Vanish Oxi Action Gel (VO) did not effectively remove the pathogen from the fabric, where the reduction was 45.1% and 89.7%, respectively. In particular, cleaning solutions after the use of household cleaners were highly contaminated with ToBRFV. After a 16-h treatment with the disinfectant MF, infectious ToBRFV was no longer present in VO, FH, and MF cleaning solutions, as demonstrated by extensive bioassays., German Federal Ministry of Food and Agriculture, Deutsche Forschungsgemeinschaft (DFG), Open Access Publication Fund of Humboldt-Universität zu Berlin, Peer Reviewed

Emaravirus (Order Bunyavirales; Family Fimoviridae) is a genus comprising over 20 emerging plant viruses with a worldwide distribution and economic impact. Emaraviruses infect a variety of host plants and have especially become prevalent in important long-living woody plants. These viruses are enveloped, with a segmented, single-stranded, negative-sense RNA genome and are transmitted by eriophyid mites or mechanical transmission. Emaraviruses have four core genome segments encoding an RNA-dependent RNA polymerase, a glycoprotein precursor, a nucleocapsid protein, and a movement protein. They also have additional genome segments, whose number varies widely. We report here that the proteins encoded by these segments form three main homology groups: a homolog of the sadwavirus Glu2 Pro glutamic protease; a protein involved in pathogenicity, which we named “ABC”; and a protein of unknown function, which we named “P55”. The distribution of these proteins parallels the emaravirus phylogeny and suggests, with other analyses, that emaraviruses should be split into at least two genera. Reliable diagnosis systems are urgently needed to detect emaraviruses, assess their economic and ecological importance, and take appropriate measures to prevent their spread (such as routine testing, hygiene measures, and control of mite vectors). Additional research needs include understanding the function of emaravirus proteins, breeding resistant plants, and clarifying transmission modes., FAZIT-STIFTUNG Gemeinnützige Verlagsgesellschaft mbH, German Research Foundation (DFG), EINSTEIN Foundation, Agency of Renewable Resources (FNR), Hamburg Ministry for Economy and Innovation, European Cooperation in Science and Technology, Deutsche Forschungsgemeinschaft (DFG, German Research Foundation), Open Access Publication Fund of Humboldt-Universität zu Berlin, Humboldt-Universität zu Berlin, Peer Reviewed