Pectobacterium carotovorum subsp. brasiliense is a broad host range bacterial pathogen, which causes blackleg of potatoes and bacterial soft rot of vegetables worldwide. Production of plant cell wall degrading enzymes is usually critical for Pectobacterium infection. However, other virulence factors and the mechanisms of genetic adaptation still need to be studied in detail. In this study, the complete genome of P. carotovorum subsp. brasiliense strain SX309 isolated from cucumber was compared with eight other pathogenic bacteria belonging to the Pectobacterium genus, which were isolated from various host plants. Genome comparison revealed that most virulence genes are highly conserved in the Pectobacterium strains, especially for the key virulence determinants involved in the biosynthesis of extracellular enzymes and others including the type II and III secretion systems, quorum sensing system, flagellar and chemotactic genes. Nevertheless, some variable regions of the T6SS and the CRISP-Cas immune system are unique for P. carotovorum subsp. brasiliense. The extensive comparative genomics analysis revealed highly conserved virulence genes in the Pectobacterium strains. However, several variable regions of type VI secretion system and two subtype Cas mechanism-Cas immune systems possibly contribute to the process of Pectobacterium infection and adaptive immunity.

Pseudomonas syringae pv. tomato is a seedborne pathogen that causes bacterial speck disease in tomato. P. syringae pv. tomato is typically detected in tomato seed using quantitative real-time PCR (qPCR) but the inability of qPCR to distinguish between viable and nonviable cells might lead to an overestimation of viable P. syringae pv. tomato cells. In the present study, a strategy involving a propidium monoazide (PMA) pretreatment followed by a qPCR (PMA-qPCR) assay was developed for quantifying viable P. syringae pv. tomato cells in contaminated tomato seed. PMA could selectively bind to the chromosomal DNA of dead bacterial cells and, therefore, block DNA amplification of qPCR. The primer pair Pst3F/Pst3R was designed based on gene hrpZ to specifically amplify and quantify P. syringae pv. tomato by qPCR. The PMA pretreatment protocol was optimized for selectively detecting viable P. syringae pv. tomato cells, and the optimal PMA concentration and light exposure time were 10 μmol liter⁻¹ and 10 min, respectively. In the sensitivity test, the detection limit of PMA-qPCR for detecting viable cells in bacterial suspension and artificially contaminated tomato seed was 10² CFU ml⁻¹ and 11.86 CFU g⁻¹, respectively. For naturally contaminated tomato seed, viable P. syringae pv. tomato cells were quantified in 6 of the 19 samples, with infestation levels of approximately 10² to 10⁴ CFU g⁻¹. The results indicated that the PMA-qPCR assay is a suitable tool for quantifying viable P. syringae pv. tomato cells in tomato seed, which could be useful for avoiding the potential risks of primary inoculum sources from contaminated seed.

Cucumber target leaf spot, caused by Corynespora cassiicola, is an emerging disease with a high incidence that causes severe damage to cucumbers on a global scale. Therefore, efforts need to be undertaken to limit the spread and infection of this pathogen, preferably by using environmentally friendly methods. In this study, the effects of temperature and moisture on the sporulation of C. cassiicola were investigated in vitro and in vivo. The novelty of our study refers to the observation of spore production and size as well as the revelation of a correlation between spore size and virulence. On potato dextrose agar (PDA) and cucumber−leaf extract agar (CEA), temperature played a critical role in spore production, which was strongly influenced by both temperature and moisture on detached leaves and cucumber seedlings. Maximum spore production was found at 30 °C on PDA and 25 °C on CEA, cucumber detached leaves and living plants. Lower spore productions were observed with a stepwise change of 5 °C. In addition, the largest spore production was found at 100% relative humidity (RH) in comparison to the other tested moisture. Moreover, moisture was found to be the most important factor affecting spore size, accounting for 83.09–84.86% of the total variance in length and 44.72–73.10% of the total variance in width. The longest−narrowest spores were formed at 100% RH, and the shortest−widest spores were formed at 75% RH. Furthermore, the result showed that larger spores of C. cassiicola were more virulent and small spores were avirulent. Our findings will contribute to the development of new strategies for the effective alleviation and control of cucumber target leaf spot.

Clubroot (Plasmodiophora brassicae) is an important soilborne disease that causes severe damage to cruciferous crops in China. This study aims to compare the differences in chemical properties and microbiomes between healthy and clubroot-diseased soils. To reveal the difference, we measured soil chemical properties and microbial communities by sequencing 18S and 16S rRNA amplicons. The available potassium in the diseased soils was higher than in the healthy soils. The fungal diversity in the healthy soils was significantly higher than in the diseased soils. Ascomycota and Proteobacteria were the most dominant fungal phylum and bacteria phylum in all soil samples, respectively. Plant-beneficial microorganisms, such as Chaetomium and Sphingomonas, were more abundant in the healthy soils than in the diseased soils. Co-occurrence network analysis found that the healthy soil networks were more complex and stable than the diseased soils. The link number, network density, and clustering coefficient of the healthy soil networks were higher than those of the diseased soil networks. Our results indicate that the microbial community diversity and network structure of the clubroot-diseased soils were different from those of the healthy soils. This study is of great significance in exploring the biological control strategies of clubroot disease.

Fusarium is a soil-borne pathogen that causes root rot disease in cucumber. To date, quantitative real-time PCR (qPCR) is a common tool to detect the content of Fusarium in soil. However, qPCR cannot distinguish between viable and nonviable cells. The aim of this study was to develop a detection technique to pretreat tissue fluid with propidium monoazide (PMA) followed by extract DNA, and then to quantify viable Fusarium cells in contaminated soil. In this work, the specific primer pair F8-1/F8-2 was designed based on the translation elongation factor (EF) gene and a PMA-qPCR assay was established to amplify and quantify soils of viable Fusarium cells. The PMA pretreatment test was optimized, which indicated that the optimal PMA concentration and light exposure time were 50 mmol L⁻¹ and 15 min, respectively. The lowest limit of viable cells in suspension detected and soil by PMA-qPCR were 82 spore mL⁻¹ and 91.24 spore g⁻¹, respectively. For naturally contaminated soil, viable Fusarium cells were detected in eight of the 18 samples, and the Fusarium amount ranged from 10⁴ to 10⁶ spore g⁻¹. In conclusion, the PMA-qPCR method has the characteristics of high sensitivity, efficiency, and time saving, which could support nursery plants to avoid Fusarium infection and agro-industry losses.