Genetic diversity of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> strains from China

A survey of bacterial wilt in China collected 286 strains of Ralstonia solanacearum from 17 plant species in 13 Chinese provinces to investigate genetic diversity using the biovar (bv.) and phylotype classification schemes. A phylotype-specific multiplex-PCR showed that 198 isolates belonged to phylotype I (bv. 3, 4 and 5) and 68 to phylotype II (bv. 2 and bv. 1). A phylogenetic analysis examined the partial sequence of the egl and hrpB gene of all strains and the genetic diversity of 95 representatives was reported, demonstrating that Chinese strains are partitioned into phylotype I (Asia) and II (Americas). Phylotype I strains (historically typed bv. 3, 4 and 5), had considerable phylogenetic diversity, including 10 different sequevars: seven previously described sequevars 12 to 18 and three new sequevars: 34, 44 and 48. Chinese strains Z1, Z2, Z3, Z7, Pe74 and Tm82 were not genetically distinguishable from the edible ginger reference strain ACH92 (r4-bv. 4) for sequevar 16. This is believed to be the first report of this ginger group in China. All Chinese bv. 2 strains falling into the genetically and phenotypically diverse phylotype II were placed into phylotype IIB sequevar 1 (historically the Andean race3-bv. 2 potato brown rot agent). In both the egl and hrpB sequence-based trees, strains isolated from mulberry were present in two distinct branches found in sequevars 12 and 48 (reference strains R292 and M2, respectively).     

Genetic diversity of Ralstonia solanacearum infecting solanaceous vegetables from India reveals the existence of unknown or newer sequevars of Phylotype I strains

Bacterial wilt is one of the important constraints in the cultivation of solanaceous vegetables in India. The disease is caused by Ralstonia solanacearum, a soil bacterium. We have collected 232 isolates of R. solanacearum infecting solanaceous vegetables (eggplant, tomato and chilli) and other crops from different parts of India. Pathogenicity of the isolates was tested on eggplant, tomato and chilli and the pathogen was confirmed by PCR. Multiplex PCR and biochemical tests indicated that all the isolates were phylotype I and biovar 3. Ninety-five representative isolates selected based on geographical region, host range and pathogenicity were subjected to further phylogenetic and diversity analysis. Sequence analysis of egl, pga and hrpB genes of 95 isolates and genetic diversity of 50 representative isolates was reported and discussed. Indian isolates within the Phylotype I did not group based on the host or geographical location, except clustering of isolates from the Andaman Islands. Indian isolates clustered into two sub groups based on egl and pga trees indicating the presence of two major population groups. Sub group 1 is the dominant group in the data set and consists of unknown/newer sequevars, and sub group 2 consist of mainly the isolates which are designated with sequevar numbers based on egl sequences. In the hrpB based tree, the sub group 2 is the dominant group in the data set and it is the same for the sub group 1 of the egl tree. Indian phylotpe I R. solanacearum strains are phenotypically diverse including the previously described sequevars 14, 17, 44, 47 and 48. Our studies indicated the existence of R. solanacearum isolates with unknown/newer sequevars; the diversity existing among the phylotype I isolates might be due to a continuous evolutionary process. To our knowledge this is the first detailed report on the diversity of phylotype I R. solanacearum strains infecting solanaceous vegetables and the existence of unknown/newer sequevars in India.     

Genetic diversity of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> strains causing bacterial wilt of solanaceous crops in Myanmar

In 2013 and 2014, an extensive survey of bacterial wilt in Myanmar was performed, and 70 strains of Ralstonia solanacearum (Rs) were collected from wilting plants of tomato, potato, chili and eggplant. Myanmar Rs strains were characterized by traditional and molecular methods. Polymerase chain reaction (PCR) test using Rs-specific primer set amplified one specific band (281-bp) from template DNA of all strains. Pathogenicity tests on the four solanaceous plants differentiated the strains into six pathogenic groups. Biovar determination tests showed that biovar 3 strains predominated (63%) among all Rs strains. Biovar 4 strains (7%) were obtained from both tomato and chili strains, whereas biovar 2 (30%) strains were isolated only from potato. Multiplex-PCR analysis indicated that tomato, eggplant and chili strains belonged to phylotype I, whereas potato strains comprised phylotype I and phylotype II. Strains in phylotype I, which was suggested to have originated from Asia, were the most prevalent in all surveyed areas. Phylogenetic analysis based on the endoglucanase (egl) gene sequences revealed that Myanmar strains partitioned into two major clusters that corresponded to phylotype I and II. Strains in phylotype I were further divided into seven subclusters, each corresponding to a distinct sequevar (15, 17, 46, 47, 48, unknown 1 or unknown 2). All strains in phylotype II belonged to sequevar 1. This is the first comprehensive report of the presence of diverse Rs strains in Myanmar.     

Potato bacterial wilt in India caused by strains of phylotype I, II and IV of Ralstonia solanacearum

Bacterial wilt or brown rot is one of the most devastating diseases of potato caused by a bacterium Ralstonia solanacearum (Smith 1986) Yabuuchi et al. (Microbiol Immunol 39:897–904 1995). Traditionally, R. solanacearum is classified into five races (r) on the basis of differences in host range and six biovars (bvs) on the basis of biochemical properties. Recently using molecular methods, R.?solanacearum has been classified into phylotypes based on the intergenic transcribed sequence of the ribosomal RNA genes 16S and 23S and into sequevars based on the endoglucanase gene (egl) sequence. In the present study, 75 bacterial strains, isolated from wilt infected potatoes from various potato growing regions of India, were classified by traditional and molecular methods. The identity of all the strains was confirmed as R. solanacearum as expected single 280-bp fragment resulted in all the strains following PCR amplification using R. solanacearum specific universal primer pair 759/760. Biovar (bv) analysis, based on utilization of disaccharide sugars and hexose alcohols, categorised the 75 strains into bv2 (78.7 %), 2 T (5.3 %), 3 (5.3 %) and 4 (10.7 %). The phylotype specific multiplex PCR assigned 78.7 % strains to phylotype II, 16.0 % to phylotype I and 5.3 % to phylotype IV. Phylogenetic analysis of egl gene sequences clustered all fifty nine phylotype II (bv2) strains with reference strain IPO1609 (IIB-1), all four phylotype IV (bv2T) strains with reference strain MAFF301558 (IV-8), three phylotype I (bv3) strains with reference strain MAFF211479 (I-30) and all eight phylotype I (bv4) and one phylotype I (bv3) strain with reference strain CIP365 (I-45). The study concluded that the Indian potato strains of R. solanacearum belong to three out of four phylotypes namely: the Asian phylotype I, the American phylotype II, and the Indonesian phylotype IV. This is the first study to address the diversity of R. solanacearum from potato in India using phylotype and sequevar scheme. We also report here for the first time the occurrence of phylotype IV sequevar 8 (bv2T) strain of R. solanacearum causing potato bacterial wilt in mid hills of Meghalaya in India.     

Phylotype and sequevar variability of Ralstonia solanacearum in Brazil,an ancient centre of diversity of the pathogen

The β‐proteobacterium Ralstonia solanacearum causes bacterial wilt of many plant species. Knowledge of phylotype and sequevar variability in populations of this microorganism is useful for implementing control measures, particularly host resistance. To this end, 301 isolates of R. solanacearum were collected from different geographic regions and hosts in Brazil. Their phylotype and sequevar characterization was used to determine the amount and distribution of phenetic and phylogenetic variability. Isolates were classified into phylotypes I (n = 48), clade 1; and phylotype II, clades 2–5. Phylotype II was divided into subclusters IIA (n = 112) and IIB (n = 141). Phylotype II was widely distributed, whereas phylotype I isolates were found in Central, Northern, and Northeastern regions of Brazil. There were 108 haplotypes identified among endoglucanase (egl) gene sequences from 301 isolates and 32 haplotypes among DNA repair (mutS) gene regions from 176 isolates. The egl and mutS sequence analyses identified eight known (1, 4, 7, 18, 27, 28, 41 and 50) and four new (54, 55, 56 and 57) sequevars. Phylotype IIB showed high diversity in sequevars and host range. Multiplex PCR, using primers specific to the Moko ecotype, characterized banana and long pepper isolates as sequevar 4 and 4/NPB, respectively. This constitutes the first report of the emergent ecotype IIB/4NPB in a new host, long pepper. The majority of sequevars were associated with geographic regions. This high variability of R. solanacearum in Brazil suggests use of host resistance to control bacterial wilt should be mainly focused by region.     

Diversity of Brazilian biovar 2 strains of Ralstonia solanacearum

Ralstonia solanacearum is responsible for bacterial wilt disease. Specific and accurate identification of this pathogen is essential for protection of susceptible crops as well as breeding resistant varieties. Historically, R. solanacearum has been classified into biovars based on the use of sugar and alcohol as carbon sources, into races based on its ability to infect different hosts, more recently into phylotypes based on the intergenic transcribed sequence of the ribosomal RNA genes 16S and 23S and into sequevars based on the endoglucanase gene (egl) sequence. Race 3 biovar 2 (R3Bv2) is widespread in South and Central America, and in Brazil it is present in all potato-producing regions as the most prevalent strain. In this study, we classified 53 Brazilian R. solanacearum biovar 2 (Bv2) strains by traditional and molecular methods. PCR with specific primers confirmed all 53 bacterial strains as belonging to the R. solanacearum species complex, and all were classified as biovar 2A or 2T based on acidification of sugars and alcohols. Multiplex phylotype PCR assigned all strains to phylotype II. Phylogenetic analysis of egl sequences showed that most Bv2 strains from Brazil analyzed in this study did not cluster with known sequevars and are less clonal than the R3Bv2 strains reported for other countries. This is the first study to address the diversity of a collection of Brazilian R. solanacearum strains using the phylotype and sequevar classification scheme.     

Application of matrix-assisted laser desorption ionization time-of-flight mass spectrometry for rapid and accurate identification of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> and <Emphasis Type="Italic">Ralstonia pseudosolanacearum</Emphasis>

Ralstonia solanacearum “species complex” (RSSC) represents soil-borne plant pathogenic bacteria, consisting of diverse and widespread strains that cause bacterial wilt on a wide range of host plants. A recent polyphasic taxonomic study has divided the RSSC into three bacterial species; Ralstonia pseudosolanacearum (phylotypes I and III), Ralstonia solanacearum (phylotype II) and Ralstonia syzygii (phylotype IV). Currently, standard identification of RSSC in plant health laboratories mainly relies on performance of two tests that are based on a different principle. However, these tests are inadequate to precisely discriminate among the three bacterial species in the RSSC. The accurate identification of each of the three bacterial species in the RSSC requires additional molecular tests, including a phylotype determination. These methodologies are labor-intensive, time consuming and rather impractical for routine identification purposes in a plant health laboratory. We explored the potential for an accurate identification of R. pseudosolanacearum (phylotypes I and III) and R. solanacearum (phylotype II) in RSSC, upon implementation of the MALDI-TOF MS tool, and after the creation and validation of an in-house database supplementing the commercial database and covering the entire known genetic diversity in RSSC. MALDI-TOF MS is an emerging approach for identification of bacterial plant pathogens and has been shown to be robust and reproducible. Additionally, when compared to the conventional microbial identification methods it is shown to be less laborious and less expensive. Validation data demonstrated that our in-house database (Mass Spectra Profiles, MSPs) was very specific resulting in the rapid and accurate identification of Ralstonia solanacearum (phylotype II), and Ralstonia pseudosolanacearum (phylotypes I and III). Additionally, no false positive results were obtained with our in-house database for other related Ralstonia sp., such as the R. picketii isolate PD 3286, or for the Pseudomonas syringae and Pseudomonas spp. isolates.     

Genetic diversity of Ralstonia solanacearum species complex strains obtained from Guangxi,China and their pathogenicity on plants in the Cucurbitaceae family and other botanical families

Bacterial wilt, caused by the Ralstonia solanacearum species complex (RSSC), is a destructive plant disease in Guangxi, China. However, the diversity of RSSC populations in the area is unknown. To this end, we performed an extensive bacterial wilt survey from 2015 to 2018. Using phylotype-specific multiplex PCR (Pmx-PCR) and an egl-based tree, 189 strains collected from 20 plant species were identified as R. pseudosolanacearum phylotype I, which included 14 sequevars (12, 13, 14, 15, 16, 17, 18, 30, 34, 44, 48, 54, 70, and 71); two strains isolated from potato plants belonged to R. solanacearum phylotype II, sequevar 1. Sequevars 13, 17, and 44 were prevalent in Guangxi, and sequevar 13 dominated the RSSC sequevars of four Cucurbitaceae plants. The susceptibility of different Cucurbitaceae species to bacterial wilt and the host range of 16 representative strains were further tested. Members of the Cucurbita, Momordica, and Luffa genera were susceptible to bacterial wilt, with wilt incidence ranging from 73% to 100%. Most strains were pathogenic to solanaceous plants, mulberry, and ginger plants but not to melon crops; however, the strains from kidney bean, pepper, and Cucurbitaceae plants were highly virulent to melon crops. This is the first comprehensive report on the genetic and host range diversity of the RSSC in Guangxi and the susceptibility of different Cucurbitaceae species to bacterial wilt, which can provide valuable information for the development of bacterial wilt control strategies.     

Bacterial wilt-resistant tomato rootstock suppresses migration of <Emphasis Type="Italic">ralstonia solanacearum</Emphasis> into soil

Ralstonia solanacearum, the causal agent of bacterial wilt of tomato, grows in infected plants and migrates from the roots into the soil. We investigated the effectiveness of bacterial wilt-resistant tomato rootstock in reducing the migration of R. solanacearum from susceptible scions into the soil. Rootstock stems were either 3–5 cm tall (low-grafted, LG) or ≥?10 cm tall (high-grafted, HG). After inoculation of scions of the susceptible cultivar (SC) with R. solanacearum below the first flower, there was no difference in disease progression among LG, HG, and ungrafted SC plants, and plants had wilted by 2 weeks. However, the rate of detection of R. solanacearum in the soil of wilted plants was reduced by grafting. The size of the R. solanacearum population in the soil of fully wilted plants increased in the order of HG?<?LG?<?SC. These results show that grafting onto resistant rootstock strongly suppressed the migration of R. solanacearum into the soil by the time of full wilting, and the effect was stronger with a longer rootstock. Migration of R. solanacearum into soil increased with increasing disease severity in SC, LG and HG. These facts suggest that early uprooting of slightly infected plants could control the spread of the bacteria into the soil.     

Phylogenetic relationships and population structure of Ralstonia solanacearum isolated from diverse origins in Taiwan

Bacterial wilt caused by Ralstonia solanacearum is a destructive disease for many crops. The aim of this study was to investigate the phylogenetic relationships and genetic structure of an R. solanacearum population from diverse origins in Taiwan. All 58 tested isolates belonged to phylotype I, except the two biovar 2 isolates from potato. These belonged to phylotype IIB sequevar 1 and were identical to known potato brown rot strains, which were probably introduced. Phylotype I isolates were grouped into 10 sequevars. Sequevar 15 was predominant (34 out of 56 isolates). Its distribution covered the whole island and it was largely associated with solanaceous crops such as tomato, and with tomato field soil. A total of 14 haplotypes were identified based on a partial endoglucanase gene sequence. Parsimony network analysis revealed that haplotype A was the oldest haplotype in the local population. It encompassed all but one of the sequevar 15 isolates. Large variation in virulence on tomato was observed among the 58 isolates, and seven pathotypes were identified. Significant genetic differentiation was detected among pathotypes. Moreover, genetic differentiation was detected between biovar 3 and biovar 4 subgroups and between the strains associated with solanaceous and non‐solanaceous species, but none was detected between strains from different geographic origins. The results suggest that the phylotype I population in Taiwan is homogeneous, while mutation and local adaptation to specific ecological niches keep shaping the population.     

A sensitive biosensor based on gold nanoparticles to detect <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> in soil

Ralstonia solanacearum, the devastating causal agent of potato bacterial wilt, is a soil-borne bacterium that can survive in the soil for a long time. The development of sensitive on-field detection methods for this pathogen is highly desirable due to its widespread host range and distribution. A novel nanobiosensor was thus developed to detect unamplified genomic DNA of R. solanacearum in farm soil. Gold nanoparticles functionalized with single-stranded oligonucleotides served as a probe to detect R. solanacearum genomic DNA. The advantages of this strategy include rapidity, facile usage and being a visual colorimetric method.     

Effect of simultaneous and sequential inoculations of <Emphasis Type="Italic">Meloidogyne incognita</Emphasis>, <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> and <Emphasis Type="Italic">Phomopsis vexans</Emphasis> on eggplant in sand mix and fly ash mix soil

Effects simultaneous and sequential inoculations of Meloidogyne incognita, Ralstonia solanacearum and Phomopsis vexans were studied on the growth, chlorophyll and carotenoid contents of eggplants grown in 25% fly ash and 25% sand mix soil. Plants grown in 25% fly ash mix soil had lesser plant growth than grown in 25% sand ash mix soil. Inoculation of M. incognita / R. solanacearum or P. vexans caused reduction in plant growth, chlorophyll and carotenoid contents in both types of soils but these pathogens in combination caused a greater reduction in than individual inoculation. Inoculation of M. incognita 20 days prior to R. solanacearum caused a greater reduction in plant growth than inoculation of M. incognita prior to P. vexans. Inoculation of P. vexans prior to R. solanacearum caused a lesser reduction in plant growth, chlorophyll and carotenoid contents than inoculation of P. vexans prior to M. incognita. Inoculation of R. solanacearum 20 days prior to M. incognita caused a greater reduction in plant growth, chlorophyll and carotenoid contents than inoculation of R. solanacearum prior to P. vexans. Galling and multiplication of M. incognita was higher in plants grown in 25% sand amended soil than with 25% fly ash soil. R. solanacearum and P. vexans had adverse effects on galling and nematode multiplication. Wilt and blight indices caused by R. solanacearum and P. vexans were 3 respectively. Wilt and blight indices were 4 when two pathogens were inoculated together.     

Suppression of bacterial wilt of tomato by soil amendment with mushroom compost containing <Emphasis Type="Italic">Bacillus amyloliquefaciens</Emphasis> IUMC7

Bacillus amyloliquefaciens IUMC7 and its culture supernatant, which we previously found to suppress Fusarium wilt, were found here to have antimicrobial activity against Ralstonia solanacearum. In in vivo experiments, mushroom compost inoculated with IUMC7 significantly reduced disease severity in tomato plants over that in control soils. The R. solanacearum population decreased in soil inoculated with IUMC7. A TLC–bioautography assay showed that one of the antimicrobial substances produced by IUMC7 is likely an iturin-like lipopeptide. These results suggest that these antimicrobial compounds are responsible for disease suppression and that mushroom compost containing IUMC7 has potential as a biocontrol product.     

<Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> upregulates marker genes of the salicylic acid and ethylene signaling pathways but not those of the jasmonic acid pathway in leaflets of <Emphasis Type="Italic">Solanum</Emphasis> lines during early stage of infection

Real-Time PCR assay was used to quantify the expression of marker genes of the salicylic acid, jasmonic acid and ethylene signaling pathways in seven Solanum lines after inoculation with a Ralstonia solanacearum phylotype I strain, R008. Four Solanum lycopersicum lines (CRA 66, Hawaii 7996, MST 32/1, Quatre carrées), one S. tuberosum line (Spunta), the wild Lycopersicon cerasiforme and Solanum commersonii were used for this investigation. Results revealed very little activation of the jasmonic acid pathway marker genes, lipoxygenase A (LoxA) and protease inhibitor II (Pin2), with no significant difference (p > 0.05) in fold change expression among the Solanum lines. In contrast the salicylic acid pathway marker genes, glucanase A (GluA) and PR-1a, and the ethylene pathway marker genes, osmotin-like (Osm) and PR-1b, were expressed at higher levels with a statistically significant difference (p < 0.05) in fold change expression among the Solanum lines. The resistant lines L. cerasiforme, CRA 66, Hawaii 7996 and S. commersonii showed stronger activation of the salicylic acid and ethylene marker genes than the moderately resistant cultivar (MST 32/1) and the susceptible lines (Quatre carrées and Spunta). The marker genes reached their highest expression levels earlier (4 h.p.i) in the resistant and moderately resistant lines than in the susceptible lines (48 h.p.i.). These results indicate that salicylic acid and ethylene signaling pathways have a significant role in defense against R. solanacearum. The timing and magnitude of the upregulation of gene expression may determine the plant ability to put up a defense response against the pathogen.     

Molecular typing of Japanese strains of Ralstonia solanacearum in relation to the ability to induce a hypersensitive reaction in tobacco

The genetic diversity of 120 Ralstonia solanacearum strains isolated from a variety of host plants across Japan was assessed on the basis of hypersensitive response (HR) in tobacco leaves and phylogenetic analyses of endoglucanase gene egl, hrpB, and gyrB. Phylogenetic analysis of egl revealed that only three strains belonged to phylotype IV, and 117 strains belonged to phylotype I. Partial sequences of HrpB were identical among phylotype I strains except for one strain. Analyses using the partial nucleotide sequences of the gyrB and egl gene fragments grouped phylotype I strains into 11 gyrB and 8 egl types, respectively, whereas analyses using the partial amino acid sequences of GyrB and Egl grouped phylotype I strains into 4 GyrB and 5 Egl types, respectively. Using multilocus sequence typing of GyrB and Egl, we identified 10 unique sequence types within the Japanese phylotype I strains. Strains belonging to the GyrB42 or GyrB66 type caused wilt in tobacco, and strains belonging to GyrB2 or GyrB9 type elicited HR, demonstrating that HR induction in tobacco is genetically differentiated in the Japanese strains of R. solanacearum.     

A chalcone synthase controls the verticillium disease resistance response in both <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> and cotton

Verticillium wilt is a devastating disease caused by the soil-borne fungus Verticillium dahliae that causes severe wilt symptoms in more than 400 plant species, including economically important cotton. However, the molecular mechanism of plant resistance to Verticillium remains unclear. In this study, we identified an Arabidopsis mutant, vsad1 (verticillium sensitive and anthocyanin deficient 1), which showed more serious disease symptoms such as discoloration and chlorosis than wild-type Arabidopsis. vsad1 is a previously identified allele of the transparent testa 4 gene (tt4), which encodes chalcone synthase (CHS), a key enzyme involved in the biosynthesis of flavonoids. Our results showed that VSAD1 expression was induced in response to Verticillium dahliae infection. Overexpression of VSAD1 partially recovered the anthocyanin accumulation phenotype of the vsad1–1 mutant. The concentration of V. dahliae increased and ROS accumulation decreased in the vsad1 mutant after infection with V. dahliae. Knockdown of the homologous gene GhCHS in cotton plants increased their susceptibility to V. dahliae infection. Thus, we conclude that VSAD1 is involved in the regulation of plant resistance to Verticillium wilt.     

我国长江流域和南方地区花生青枯菌遗传多样性分析

为明确不同青枯菌的遗传多样性和其在花生植株上的致病力差异,采用国际上新的青枯菌演化型分类模式,对从我国长江流域和南方地区9个花生种植区分离的95株花生青枯菌Ralstonia solanacearum菌株进行遗传多样性分析,基于内源葡聚糖酶基因egl对青枯菌进行系统发育研究,并对供试青枯菌的致病力进行测定。结果表明,所有95株菌株均属于青枯菌演化型I型,即亚洲分支类型。在序列变种分类上,所检测的9个花生种植区中有8个种植区的花生青枯菌菌株属于序列变种14,仅有1个种植区(广西壮族自治区贺州市)的花生青枯菌菌株属于序列变种48,表明我国长江流域和南方地区花生青枯菌群体遗传多样性水平较低。青枯菌致病力测定结果表明,来自赣州市的菌株GZ-1、贺州市的菌株HZ-2和宜昌市的菌株YC接种到花生植株14 d后,花生的病情指数分别为43.8、75.0和87.5,而来自其它6个花生种植区的菌株接种花生后,其病情指数均为100.0,表明菌株GZ-1和HZ-2的致病力较弱,而其它7个花生种植区代表性菌株的致病力均较强。     

Bacterial wilt symptoms are impacted by host age and involve net downward movement of <Emphasis Type="Italic">Erwinia tracheiphila</Emphasis> in muskmelon

Cucurbit bacterial wilt, caused by Erwinia tracheiphila, is a devastating disease of cucurbit crops in the Midwest and Northeast U.S. Current management of bacterial wilt relies primarily on insecticide applications to control striped and spotted cucumber beetles (Acalymma vittatum and Diabrotica undecimpunctata howardi, respectively), which vector E. tracheiphila. Development of alternative management strategies is constrained by a lack of understanding of bacterial wilt etiology. The impact of host age on rate on symptom development and extent of bacterial movement in the xylem of muskmelon (Cucumis melo cv. Athena) was evaluated following wound inoculation of 2- to 8-week-old plants in growth chamber experiments. Wilting occurred more rapidly in plants after inoculating E. tracheiphila into 2- or 4-week-old plants than 6- or 8-week-old plants. Recovery of viable cells from stem segments revealed that vascular spread of E. tracheiphila was more extensive below than above the inoculation point. These findings provide experimental evidence that host age impacts the rate of symptom development in cucurbit bacterial wilt and that movement of the xylem-inhabiting pathogen E. tracheiphila within muskmelon plants occurs primarily in the downward direction.     

<Emphasis Type="Italic">Fusarium oxysporum</Emphasis> and <Emphasis Type="Italic">Fusarium avenaceum</Emphasis> associated with yield-decline of pyrethrum in Australia

Fusarium wilt, one of the destructive diseases of cucumber can be effectively controlled by using biocontrol agents such as Trichoderma harzianum. However, the mechanisms controlling T. harzianum-induced enhanced resistance remain largely unknown in cucumber plants. Here we screened the potent T. harzianum isolate TH58 that could effectively control F. oxysporum (FO). Glasshouse efficacy trials also showed that TH58 decreased disease incidence by 69.7 %. FO induced ROS over accumulation, while TH58 inoculation suppressed ROS over accumulation and improved root cell viability under F. oxysporum infection. TH58 inoculation could reverse the FO-induced cell division block and regulate the proportional distribution of nuclear DNA content through inducing 2C fraction. Moreover, the expression levels of cell cycle-related genes such as CDKA, CDKB, CycA, CycB, CycD3;1 and CycD3;2 in TH58 - pre-inoculated seedlings were up-regulated compared with those infected with FO alone. Taken together, these results suggest that T. harzianum improved plant resistance against Fusarium wilt disease via alterations in nuclear DNA content and cell cycle-related genes expression that might maintain a lower ROS accumulation and higher root cell viability in cucumber seedlings.