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Though information exists regarding the pathogenesis of the shot-hole disease (SH) in flowering cherry (FC), there has been a lack of research focusing on SH management. Therefore, here, we investigated the inhibitory activities of antagonistic bacteria against SH pathogens both in vitro and in vivo as well as their biochemical characteristics and bioactive compounds. Two biosurfactant-producing bacterial antagonists, identified as Bacillus velezensis strains JCK-1618 and JCK-1696, exhibited the best effects against the growth of both bacterial and fungal SH pathogens in vitro through their cell-free culture filtrates (CFCFs). These two strains also strongly inhibited the growth of the pathogens via the action of their antimicrobial diffusible compounds and antimicrobial volatile organic compounds (VOCs). Crude enzymes, solvent extracts, and biosurfactants of the two strains exhibited antimicrobial activities. Liquid chromatography/electrospray ionization time-of-flight mass spectrometric analysis of the partially purified active fractions revealed that the two antagonists produced three cyclic lipopeptides, including iturin A, fengycin A, and surfactin, and a polyketide, oxydifficidin. In a detached leaf assay, pre-treatment and co-treatment of FC leaves with the CFCFs led to a large reduction in the severity of the leaf spots caused by Epicoccum tobaicum and Bukholderia contaminans, respectively. In addition, the two antagonists produced indole-3-acetic acid, siderophore, and a series of hydrolytic enzymes, along with the formation of a substantial biofilm. To our knowledge, this is the first report of the antimicrobial activities of the diffusible compounds and VOCs of B. velezensis against the SH pathogens and their efficiency in the biocontrol of SH.

Passiflora mottle virus (PaMoV), an aphid-borne potyvirus, is the primary causal virus of devastating passionfruit woodiness disease in Vietnam. Here we generated a nonpathogenic, attenuated PaMoV strain for disease control by cross protection. A full-length genomic cDNA of PaMoV strain DN4 from Vietnam was constructed to generate an infectious clone. The green fluorescent protein was tagged at the N-terminal region of the coat protein gene to monitor in planta the severe PaMoV-DN4. Two amino acids within the conserved motifs of helper component protease (HC-Pro) of PaMoV-DN4 were mutated individually or in combination as K53E or/and Risi I. Mutants PaMoV-E53 and PaMoV-Ilgl induced local lesions in Chenopodiumquinoa plants, while PaMoV-EsjIisi caused infection without apparent symptoms. In passionfruit (Passiflora edulis) plants, PaMoV-F.53 elicited severe leaf mosaic and PaMoV-Lsi induced leaf mottling, while PaMoV-E53I181 caused transient mottling followed by symptomless recovery. PaMoV-E.53li8i was stable after six serial passages in yellow passionfruit (Passiflora edulis f. flavicarpa) plants. Its temporal accumulation levels were lower than those of the wild type, with a zigzag accumulation pattern, typical of a beneficial protective virus. An RNA silencing suppression (RSS) assay revealed that all three mutated HC-Pros are defective in RSS. Triplicated cross-protection experiments with a total of 45 plants showed that the attenuated mutant PaMoV-E53I181 provided a high protection rate (91%) against the homologous wild-type virus in passionfruit plants. This work revealed that PaMoV-Ejdm can be used as a protective virus to control PaMoV by cross protection. [ABSTRACT FROM AUTHOR]

Passionfruit plantings in Vietnam increased to 10,000 ha in 2019. However, outbreaks of passionfruit woodiness disease (PWD) have become a serious threat to production. In this study, five virus isolates (DN1, DN4, NA1, GL1, and GL2) were collected from different areas of Vietnam. Their causal roles in PWD were verified by back-inoculation to passionfruit. Analyses of coat protein (CP) and genomic sequences revealed that the GL1 isolate is closely related to East Asia Passiflora virus (EAPV) AO strain of Japan (polyprotein nt and aa identities of 98.1 and 98.2%, respectively), and the GL2 isolate is related to Telosma mosaic virus (TelMV) isolate PasFru, China (polyprotein nt and aa identities of 87.1 and 90.9%, respectively). CP comparison, host range, and cytological characterization indicated that DN1, DN4, and NA1 are potyviruses but are different from EAPV and TelMV. Phylogenic analyses of their CP and genome sequences indicated that these three isolates and the passionfruit severe mottle-associated virus Fujian isolate of China belong to a distinct clade, which does not meet the threshold (76% nt identity of polyprotein) to be regarded as any of potyviral species. Thus, a new species name, Passiflora mottle virus, (PaMoV), has been proposed by the International Committee on Taxonomy of Viruses. A rabbit antiserum was produced against the CP of DN1, and it can distinguish PaMoV from TelMV and EAPV in western blotting and enzyme-linked immunosorbent assay (ELISA) without cross-reactions. Field surveys of 240 samples by ELISA and reverse transcription PCR found that PWD in Vietnam is caused mainly by PaMoV, followed by EAPV, mixed infection of PaMoV and EAPV, and rare cases of TelMV. [ABSTRACT FROM AUTHOR]