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Pseudomonas cannabina pv. alisalensis (Pcal) causes bacterial blight on cabbage. We previously conducted a screening for reduced virulence using Tn5 transposon mutants and identified one of the transcriptional factors, HexR, as a potential Pcal virulence factor. However, the role of HexR in plant pathogenic Pseudomonas virulence has not been investigated well. Here, we demonstrated that the Pcal hexR mutant showed reduced disease symptoms and bacterial populations on cabbage, indicating that HexR contributes to Pcal virulence. We used RNA-seq analysis to characterize the genes regulated by HexR. We found that several type three secretion system (T3SS)-related genes had lower expression of the Pcal hexR mutant. Five genes were related to T3SS machinery, two genes were related to type three helper proteins, and three genes encoded type three effectors (T3Es). We also confirmed that T3SS-related genes, including hrpL, avrPto, hopM1, and avrE1, were also down-regulated in the Pcal hexR mutant both in culture and in vivo by using RT-qPCR. T3SS functions to suppress plant defense in host plants and induce hypersensitive response (HR) cell death in non-host plants. Therefore, we investigated the expression profiles of cabbage defense-related genes, including PR1 and PR5, and found that the expressions of these genes were greater in the Pcal hexR mutant. We also demonstrated that the hexR mutant did not induce HR cell death in non-host plants, indicating that HexR contributes in causing HR in nonhost plants. Together, these results indicate that the mutation in hexR leads to a reduction in the T3SS-related gene expression and thus an impairment in plant defense suppression, reducing Pcal virulence.

The genus Pseudomonas includes some of the most problematic and studied foliar bacterial pathogens. Generally, in a successful disease cycle there is an initial epiphytic lifestyle on the leaf surface and a subsequent aggressive endophytic stage inside the leaf apoplast. Leaf-associated bacterial pathogens enter intercellular spaces and internal leaf tissues by natural surface opening sites, such as stomata. The stomatal crossing is complex and dynamic, and functional genomic studies have revealed several virulence factors required for plant entry. Currently, treatments with copper-containing compounds, where authorized and admitted, and antibiotics are commonly used against bacterial plant pathogens. However, strains resistant to these chemicals occur in the fields. Therefore, the demand for alternative control strategies has been increasing. This review summarizes efficient strategies to prevent bacterial entry. Virulence factors required for entering the leaf in plant-pathogenic Pseudomonas species are also discussed.

Bacterial canker of kiwifruit caused by Pseudomonas syringae pv. actinidiae (Psa) is a serious threat to kiwifruit production worldwide. Four biovars (Psa biovar 1; Psa1, Psa biovar 3; Psa3, Psa biovar 5; Psa5, and Psa biovar 6; Psa6) were reported in Japan, and virulent Psa3 strains spread rapidly to kiwifruit production areas worldwide. Therefore, there is an urgent need to develop critical management strategies for bacterial canker based on dissecting the dynamic interactions between Psa and kiwifruit. To investigate the molecular mechanism of Psa3 infection, we developed a rapid and reliable high-throughput flood-inoculation method using kiwifruit seedlings. Using this inoculation method, we screened 3000 Psa3 transposon insertion mutants and identified 91 reduced virulence mutants and characterized the transposon insertion sites in these mutants. We identified seven type III secretion system mutants, and four type III secretion effectors mutants including hopR1. Mature kiwifruit leaves spray-inoculated with the hopR1 mutant showed significantly reduced virulence compared to Psa3 wild-type, indicating that HopR1 has a critical role in Psa3 virulence. Deletion mutants of hopR1 in Psa1, Psa3, Psa5, and Psa6 revealed that the type III secretion effector HopR1 is a major virulence factor in these biovars. Moreover, hopR1 mutants of Psa3 failed to reopen stomata on kiwifruit leaves, suggesting that HopR1 facilitates Psa entry through stomata into plants. Furthermore, defense related genes were highly expressed in kiwifruit plants inoculated with hopR1 mutant compared to Psa wild-type, indicating that HopR1 suppresses defense-related genes of kiwifruit. These results suggest that HopR1 universally contributes to virulence in all Psa biovars by overcoming not only stomatal-based defense, but also apoplastic defense.

Bacteria are exposed to and tolerate diverse and potentially toxic compounds in the natural environment. While efflux transporters are generally thought to involve bacterial antibiotic resistance in vitro, their contributions to plant bacterial virulence have so far been poorly understood. Pseudomonas cannabina pv. alisalensis (Pcal) is a causal agent of bacterial blight of Brassicaceae. We here demonstrated that NU19, which is mutated in the resistance-nodulation-cell division (RND) transporter encoded gene, showed reduced virulence on cabbage compared to WT, indicating that the RND transporter contributes to Pcal virulence on cabbage. We also demonstrated that brassinin biosynthesis was induced after Pcal infection. Additionally, the RND transporter was involved in resistance to plant-derived antimicrobials and antibiotics, including the cabbage phytoalexin brassinin. These results suggest that the RND transporter extrudes plant-derived antimicrobials and contributes to Pcal virulence. We also found that the RND transporter contributes to Pcal virulence on Brassicaceae and tomato, but not on oat. These results suggest that the RND transporter contributes to Pcal virulence differentially depending on the host-plant species. Lastly, our expression-profile analysis indicated that the type-three secretion system (TTSS), which is essential for pathogenesis, is also involved in suppressing brassinin biosynthesis. Taken together, our results suggest that several Pcal virulence factors are involved in resistance to plant-derived antimicrobials and bacterial survival during infection.

The pathogen Colletotrichum orbiculare is causal fungus of cucurbit anthracnose. Multiple races have been identified in the United States, suggesting that it is necessary to cultivate suitable resistant cultivars and breed new cultivars with the most suitable resistance gene. This study examined the pathogenicity and virulence of 20 strains in Japan to clarify the existence of races and virulence differences. Based on the symptoms on inoculated cotyledons and true leaves of watermelon, we could evaluate the compatibility of each strain to each host cultivar. Our analysis based on the reaction to the host cultivar harboring the resistance gene Ar-1 (Cla001017) revealed the existence of three races in Japan. An alarming result was that a race that overcame Ar-1, which is a target gene in current watermelon breeding in Japan, is present in Japan. The cucumber and melon host cultivars showed diverse symptoms, whereas a squash cultivar was resistant to all strains. Three strains caused severe damage even to the most resistant cucumber cultivar 'Ban Kyuri' and resistant cultivars harboring Cssgr, a well-known gene conferring loss-of-susceptibility resistance. Screening genetic resources for novel resistance genes using strains with high virulence is of vital importance for watermelon, cucumber, and melon production. [ABSTRACT FROM AUTHOR]

The objective of this study was to investigate the growth and light-intercepting characteristics of tomatoes when movable benches are used in their cultivation. We cultivated tomatoes in a greenhouse (168 m2) during summer (9 July–9 September 2018) under different furrow distances (F1.0 = 1.0 m and F1.6 = 1.6 m) and movable benches (M indicates that the furrow distance = 0.4–0.8 m). Compared to the other treatments, when the movable bench was used to the change furrow distance depending on the plant growth stage (M treatment), the percentage of canopy light interception increased to ~90% at the early stage of plant growth (~20 days after transplanting). The percentage of canopy light interception for different treatments increased in the order of M > F1.0 > F1.6, and it increased towards the end of cultivation. In addition, the yield per unit area exhibited the same trend. Therefore, the solar radiation inside a greenhouse can be efficiently intercepted by plants when movable benches are used. This indicated that it was possible to increase plant yield per unit area using movable benches in plant cultivation.