Resistance profiles of Botrytis cinerea populations to several fungicide classes on greenhouse tomato and strawberry in Lebanon

Tomato and strawberry are the most important protected crops in Lebanon and are seriously affected by grey mould disease, caused by Botrytis cinerea. In the present study, the fungicide sensitivity assays revealed medium to high frequencies of B. cinerea isolates resistant to benzimidazoles, dicarboximides, and anilinopyrimidines on tomato and strawberry. Fludioxonil- and boscalid-resistant mutants were uncommonly found at generally low frequency on both crops. Resistance to fenhexamid was detected in only one site on tomato but in most sites on strawberry with high frequencies, and the occurrence of resistance to QoI fungicides was ascertained on both crops. The majority of the tested isolates (>90%) exhibited multiple fungicide resistance, and isolates resistant to the seven antibotrydial fungicide classes were detected on strawberry in three locations. A high level of resistance was shown by B. cinerea mutants resistant to boscalid, fenhexamid, and QoI fungicides, while two levels of moderate and high resistance to anilinopyrimidines were identified. Genetic analysis revealed point mutations in the target genes commonly associated with resistance in B. cinerea isolates, with all mutants resistant to dicarboximides, fenhexamid, boscalid, and QoI fungicides carrying single-nucleotide polymorphims in BcOS1 (I365S/N, Q369P, and N373S), Erg27 (F412V/I), SdhB (H272R/Y), and cytb (G143A) genes, respectively. The general incorrect use of fungicides has caused the development and spread of fungicide resistance as a widespread phenomenon on protected tomato and strawberry in Lebanon. The implementation of appropriate antiresistance strategies is highly recommended

Identification and Characterization of the Helix-Destabilizing Activity of Rotavirus Nonstructural Protein NSP2

The rotavirus nonstructural protein NSP2 self-assembles into homomultimers, binds single-stranded RNA nonspecifically, possesses a Mg(2+)-dependent nucleoside triphosphatase (NTPase) activity, and is a component of replication intermediates. Because these properties are characteristics of known viral helicases, we examined the possibility that this was also an activity of NSP2 by using a strand displacement assay and purified bacterially expressed protein. The results revealed that, under saturating concentrations, NSP2 disrupted both DNA-RNA and RNA-RNA duplexes; hence, the protein possesses helix-destabilizing activity. However, unlike typical helicases, NSP2 required neither a divalent cation nor a nucleotide energy source for helix destabilization. Further characterization showed that NSP2 displayed no polarity in destabilizing a partial duplex. In addition, helix destabilization by NSP2 was found to proceed cooperatively and rapidly. The presence of Mg(2+) and other divalent cations inhibited by approximately one-half the activity of NSP2, probably due to the increased stability of the duplex substrate brought on by the cations. In contrast, under conditions where NSP2 functions as an NTPase, its helix-destabilizing activity was less sensitive to the presence of Mg(2+), suggesting that in the cellular environment the two activities associated with the protein, helix destabilization and NTPase, may function together. Although distinct from typical helicases, the helix-destabilizing activity of NSP2 is quite similar to that of the ςNS protein of reovirus and to the single-stranded DNA-binding proteins (SSBs) involved in double-stranded DNA replication. The presence of SSB-like nonstructural proteins in two members of the family Reoviridae suggests a common mechanism of unwinding viral mRNA prior to packaging and subsequent minus-strand RNA synthesis

Crystallographic Analysis Reveals Octamerization of Viroplasm Matrix Protein P9-1 of Rice Black Streaked Dwarf Virus

The P9-1 protein of Rice black streaked dwarf virus accumulates in viroplasm inclusions, which are structures that appear to play an important role in viral morphogenesis and are commonly found in viruses in the family Reoviridae. Crystallographic analysis of P9-1 revealed structural features that allow the protein to form dimers via hydrophobic interactions. Each dimer has carboxy-terminal regions, resembling arms, that extend to neighboring dimers, thereby uniting sets of four dimers via lateral hydrophobic interactions, to yield cylindrical octamers. The importance of these regions for the formation of viroplasm-like inclusions was confirmed by the absence of such inclusions when P9-1 was expressed without its carboxy-terminal arm. The octamers are vertically elongated cylinders resembling the structures formed by NSP2 of rotavirus, even though there are no significant similarities between the respective primary and secondary structures of the two proteins. Our results suggest that an octameric structure with an internal pore might be important for the functioning of the respective proteins in the events that occur in the viroplasm, which might include viral morphogenesis