Molecular epidemiology of <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">syringae</Emphasis> strains isolated from barley and wheat infected with bacterial black node

A repetitive sequence-based (rep)-polymerase chain reaction (PCR) and inter-simple sequence repeat (ISSR)-PCR were used to molecular type Pseudomonas syringae pv. syringae (PSS) strains isolated from barley and wheat plants with bacterial black node symptoms grown in 22 different locations and six different seed-production districts in Japan. Eighteen genomic fingerprinting (GF) genotypes were obtained from the combined results of BOX-, REP-, and GTG₅-PCR, indicating that the PSS population in Japan has high genetic diversity. The result based on logistic regression indicated that the population of GF genotype A was significantly related to a seed-producing district and that the epidemic of PSS strains in fields originated mainly from seed infection. This study will be applicable to future studies of the molecular epidemiology of bacterial plant diseases that have multiple infection routes.     

Spatiotemporal distribution of tomato plants naturally infected with leaf mold in commercial greenhouses

The distribution of tomato plants infected by Passalora fulva, causal agent of leaf mold, was analyzed by using Taylor’s model and Iwao’s model to assess the patterns of spatial distribution within the greenhouses. In Taylor’s model, the sample variance (s ²) of diseased plants newly recognized at each observation increased only slightly with mean density (m). In Iwao’s model, the mean crowding (m*) of newly recognized diseased plants at each observation increased with m of diseased plants. The statistical analysis in this study suggests that new infections in greenhouses observed during this investigation tended to cluster around a diseased plant and that secondary infections occurred as independent cluster points.     

A new pathovar of <Emphasis Type="Italic">Pseudomonas syringae</Emphasis>, pathovar <Emphasis Type="Italic">allii,</Emphasis> isolated from onion plants exhibiting symptoms of blight

Bacterial pathogens of onion (Allium cepa) plants and their undetected presence in seed can cause substantial losses to onion producers. In this study, 23 Pseudomonas syringae strains were isolated from five onion plants and 18 onion seeds. The symptoms on leaves and seed stalks were irregular lesions with necrotic centres and water soaked margins. The aim of the study was to characterize these P. syringae strains using Biolog GN III carbon source utilization, multilocus sequence typing (MLST) based on partial sequences of four housekeeping genes (cts, gapA, gyrB and rpoD), and to determine whether or not the strains were pathogenic on onion (cv. Granex 33), chive (Allium schoenoprasum cv. Grasiue), leek (Allium porrum cv. Giant Italian) and spring onion (Allium fistulosum cv. Salotte) plants. Both Biolog analysis and MLST analysis separated onion strains into two clusters, one supporting the existence of a new pathovar of P. syringae, and the other corresponding to P. syringae pv. porri. Pseudomonas syringae strains belonging to the new pathovar we pathogenic only on onion plants of the Allium spp. tested. The results of this study revealed that bacterial blight of onion in South Africa is caused by two pathovars of P. syringae sensu lato, namely, the newly described pathovar, allii, and P. syringae pv. porri. The symptoms caused by these two pathovars in the field were indistinguishable.     

<Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">alliifistulosi</Emphasis> pv. nov., the causal agent of bacterial leaf spot of onions

In 1972, bacterial leaf spot of onion (BLSO) was first recorded in Japan by Goto. The pathogen was considered as a pathovar of Pseudomonas syringae specifically causing disease on onion and Welsh onion, but it has not been taxonomically investigated in detail. In 2012 and 2014, a disease suspected as BLSO re-emerged on onion in Shizuoka and Hyogo Prefectures, Japan, respectively. A pathogenic bacterium isolated from the infected onions was thought to be the BLSO agent after preliminary examinations. Strains isolated from BLSO in 1969, 1986, 1987, 2012 and 2014 were characterized and compared with the causal agent of bacterial blight of leek (P. syringae pv. porri), which causes similar symptoms on Allium plants. The result of rep-PCR distinguished the BLSO agent from P. syringae pv. porri. Multilocus sequence analysis on housekeeping genes and hrp genes encoding the type-III secretion system revealed that the strains of the BLSO agent clustered independently of P. syringae pv. porri. The BLSO agent and P. syringae pv. porri also differed in utilization of erythritol, dl-homoserine, glutaric acid and other bacteriological characteristics and caused different reactions on onion, Welsh onions, chives, shallot, rakkyo, leek, garlic and Chinese chive. Thus, the BLSO agent clearly differs from P. syringae pv. porri and is considered to be a new pathovar of P. syringae. The name P. syringae pv. alliifistulosi is proposed with pathotype strain ICMP3414.     

<Emphasis Type="Italic">Alternaria alternata</Emphasis>: A new seed-transmitted disease of coriander in South Africa

This is the first report of Alternaria leaf spot disease on coriander (Coriandrum sativum L.) in South Africa. Using the agar plate method, Alternaria alternata was isolated from coriander seed lots together with four other fungal genera, which included Aspergillus, Fusarium, Penicillium and Rhizopus. Standard seed germination tests of coriander seed lots infected with seed-borne mycoflora showed a positive correlation with the number of diseased seedlings (r?=?0.239, p?<?0.01). Pathogenicity tests demonstrated that this seed-borne A. alternata was pathogenic on coriander and symptoms on leaves first appeared as small, dark brown to black, circular lesions (<5 mm diam.) that enlarged and coalesced to form dark brown blotches as time progressed. Leaf spot disease was most severe (64%) on wounded leaves inoculated with A. alternata. Re-isolation of A. alternata from diseased coriander plants satisfied the Koch’s postulates, thus confirming it as the causal agent of Alternaria leaf spot disease. Parsimony analysis based on rpb2 (GenBank Accession No. KT895947), gapdh (KT895949) and tef-1α (KT895945) sequences confirmed identity of the Alternaria isolate, which grouped within the A. alternata clade. Alternaria alternata was shown to be transmitted from infected coriander seed to the developing plants.     

AlgU contributes to the virulence of <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">tomato</Emphasis> DC3000 by regulating production of the phytotoxin coronatine

Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), which causes bacterial speck disease of tomato, has been used as a model pathogen to investigate the molecular basis of plant–pathogen interactions. The function of many potential virulence factors encoded in the Pst DC3000 genome and their modes of action are not fully understood. P. syringae is known to produce the exopolysaccharide alginate. Although AlgU, a sigma factor, is known to regulate the expression of genes such as algD related to alginate biosynthesis, the molecular mechanisms of AlgU in the virulence of Pst DC3000 is still unclear. To investigate the function of AlgU and alginate in plant–bacterial pathogen interactions, we generated ΔalgU and ΔalgD mutants. After inoculation with ΔalgU but not ΔalgD, host plants of Pst DC3000 including tomato and Arabidopsis had milder disease symptoms and reduced bacterial populations. Expression profiles of Pst DC3000 genes revealed that AlgU can regulate not only the expression of genes encoding alginate biosynthesis, but also the expression of genes related to type III effectors and the phytotoxin coronatine (COR). We also demonstrated that the ΔalgU mutant showed full virulence in the Arabidopsis fls2 efr1 double mutant, which is compromised in the recognition of PAMPs. Further, the application of COR was able to restore the phenotype of the ΔalgU mutant in the stomatal response. These results suggest that AlgU has an important role in the virulence of Pst DC3000 by regulating COR production.     

Black band of Jew’s marrow caused by <Emphasis Type="Italic">Lasiodiplodia theobromae</Emphasis>

Stem rot and wilt of Jew’s marrow (nalta jute, Corchorus olitorius) were found on Is. Okinawa, Okinawa Prefecture, Japan, in March 2000. An anamorphic fungus, Lasiodiplodia theobromae was isolated repeatedly from the diseased plants and demonstrated to cause the disease. We coined the Japanese name “kurogare-byô” of Jew’s marrow for the present disease because it was new to Japan, although it had already been reported in India and Bangladesh as black band of the plant.     

Alternative hosts of <Emphasis Type="Italic">Curtobacterium flaccumfaciens</Emphasis> pv. <Emphasis Type="Italic">flaccumfaciens</Emphasis>, causal agent of bean bacterial wilt

Alternative hosts are an important way of phytopathogenic bacteria survival between crop seasons, constituting a source of inoculum for the following crops. Bacterial wilt, caused by Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff), is one of the most important diseases for common bean, and little information is available about the host range of the bacterium. In this study, we assessed possible alternative hosts for Cff, especially those cultivated during winter, in rotation systems with common bean. Plants of barley, black oat, canola, radish, ryegrass, wheat and white oat, were assessed under field and greenhouse conditions. Cff colonized epiphytically all plant species and endophytically black oat, ryegrass, wheat and white oat plants assessed in the greenhouse assays. Under field conditions, Cff colonized all plant species by except radish. All bacterial strains re-isolated from the plants were pathogenic to common bean and identified as Cff by PCR with specific primers. Based on our results, the cultivation of bean crop in succession with barley, black oat, canola, ryegrass, wheat and white oat should not be recommended, mainly in areas with a history of bacterial wilt occurrence. In these cases, the better option for crop rotation during the winter is radish, a non-alternative host for Cff.     

<Emphasis Type="Italic">Clavibacter michiganensis</Emphasis> subsp. <Emphasis Type="Italic">michiganens</Emphasis>is strains virulence and genetic diversity. a first study in Argentina

Tomato leaves showing severe leaf spot symptoms have been observed and sampled in the central west and southwest Taiwan during 2015 and 2016. The symptoms were similar to those of bacterial leaf spot/late blight diseases, but only Stemphylium-like fungi were consistently isolated from the diseased tomato. Upon spray inoculation of tomato, Stemphylium-like isolates caused leaf spot symptoms identical to those of naturally infected plants, and the pathogenic isolates were successfully re-isolated from inoculated leaves. The tomato-pathogenic isolates were identified as S. lycopersici based on morphological characterization and molecular identification. S. lycopersici has been previously reported to cause gray leaf spot of tomato in the temperate regions, but the majority of S. lycopersici-caused lesions were black/dark brown rather than gray in our surveillance. Accordingly, it is suggested that S. lycopersici-caused disease of tomato is named Stemphylium leaf spot of tomato more appropriately than tomato gray leaf spot. Moreover, S. lycopersici-caused leaf spot disease on tomato has been distributed in major tomato production regions in Taiwan. The information provided by our study will be important for future breeding of tomato cultivars, especially for tomato producers in Taiwan.     

Evaluation of seedborne bacterial pathogens on common bean cultivars grown in central Anatolia region,Turkey

Bacterial diseases of bean cause economically important losses worldwide. The most important method for managing bacterial diseases on bean is the use of pathogen-free seed. In this study, 198 different dry bean seed samples of six different cultivars including Dermason, Cali, Sira, Battal, Bombay and Seker, were collected from 12 provinces of Central Anatolia Region of Turkey. All were tested to investigate the seedlots as primary inoculum sources of the major bacterial diseases. The data revealed that 22,72 %, 13,63 %, 11,11 %, 1,51 % and 0.5 % of seed samples tested were contaminated with five seedborne bacterial pathogens, Pseudomonas savastanoi pv. phaseolicola (Psp), Pseudomonas syringae pv. syringae (Pss), Xanthomonas axonopodis pv. phaseoli (Xap), X. axonopodis pv. phaseoli var. fuscans (Xapf) and Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff), respectively. All bacterial strains isolated were identified based on morphological, physiological, biochemical, molecular and pathogenicity tests. The results showed that Psp and Pss were found together on cv. Cali; Psp and Xap on cv. Dermason and cv. Sira; and Pss and Xap on cv. Seker, cv. Dermason, and cv. Cali. Therefore, the results in the present study suggested that evaluation and selection of pathogen-free seeds are very important for preventing the spread of pathogens and effective management of seed borne bacterial diseases prevalent in bean growing regions; in addition to implementation of integrated crop production strategies such as crop rotation, sanitation, seed treatment, tolerant/resistant cultivar selection and proper bactericide application.     

Phylogenetic relationships of fluorescent <Emphasis Type="Italic">Pseudomonad</Emphasis> isolates associated with bacterial canker of stone fruit trees in Kurdistan province,Iran

Bacterial canker is one of the most important diseases of stone fruit trees in various locations of Kurdistan province, Iran. Genetic diversity and evolutionary relationships among 20 fluorescent pseudomonads isolated from stone fruit trees with symptoms similar to bacterial canker were investigated using a polyphasic approach by means of phenotypic characterizations, repetitive PCR using the REP and ERIC primers and multilocus sequence typing (MLST) of four housekeeping genes (gapA, rpoD, gyrB and gltA). The pathogenicity of strains was carried out under greenhouse conditions. Twelve strains produced an expected amplified DNA fragment of about 752-bp which indicated the presence of the syrB gene. Based on MLST, these strains belonged to P. syringae species complex and included in the genomospecies 1, phylogroup 2b and 2d. Phylogenetic analysis of the other eight fluorescent pseudomonad strains by using gyrB and rpoD sequences allowed the identification of strains into P. fluorescens, P. putida and P. lutea groups. Unweighted pair group method analysis (UPGMA) of genomic fingerprints obtained by rep-PCR revealed 17 different patterns which grouped P. syringae strains into three clusters clearly separated from other fluorescent pseudomonads. MLST confirmed the genetic variability among strains obtained by rep-PCR. Grouping identified of P. syringae strains by both rep-PCR and MLST was related to geographic locations of strains.     

Identification of resistance to bacterial canker (<Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">syringae</Emphasis>) disease on apricot genotypes grown in Turkey

Bacterial canker caused by Pseudomonas syringae pv. syrinage (Pss) in apricot has widely spread in Turkey, especially in Malatya province, in recent years. The main objective of this study was to determine resistance of apricot cultivars to bacterial canker caused by Pss in apricot cultivars grown in Turkey. During the 2006–2007 growing period, bacterial isolations were taken from diseased apricot trees in Malatya and 53 Pseudomonas syringae isolates were obtained. Forty-two isolates were determined as Pseudomonas syringae pv. syringae and 11 isolates as pv. morsprunorum. In a pathogenicity test, leaves of cv. Hacihalilo?lu were used and five Pss isolates (K24, K25, K43, K47 and K51) were detected to be the most virulent and were used to test for cultivar resistance to Pss. Leaves of fifteen apricot cultivars (Alyanak, Çatalo?lu, Çölo?lu, Erken A?erik, Hacihalilo?lu, Hasanbey, ?smaila?a, Kabaa?i, Karacabey, Sakit 2, So?anci, ?am, ?ekerpare, Tokalo?lu (Erzincan) and Turfanda Eski Malatya) were tested for resistance to Pss. Green shoots were spray-inoculated with a concentration of 10⁸ cfu ml^?1 Pss mixed culture. Sprayed shoots were covered with moist plastic bags for 3 days and maintained in the growth chamber and monitored for symptom development. Hasanbey, Çölo?lu, So?anci and ?ekerpare apricot cultivars were resistant and ?am, Tokalo?lu (Erzincan) and Erken A?erik apricot cultivars were susceptible to Pss. This is the first report of a resistance source in apricot cultivars grown in Turkey against Pss.     

Diversity of rDNA-ITS sequences of <Emphasis Type="Italic">Polymyxa graminis</Emphasis> from wheat and barley in Japan

Polymyxa graminis is a soil-borne obligate organism that transmits bymoviruses and furoviruses to barley and wheat. We analyzed rDNA-ITS sequences of P. graminis from roots of wheat and barley in fields in Japan and obtained five kinds of sequences; two sequences were almost the same as known ribotype Ia and IIa, respectively, and three were close to ribotype Ib. When infection of P. graminis was examined using PCR, ribotype Ia was detected only in barley, but ribotypes Ib and IIa were detected in both wheat and barley. Our analysis suggested that Japanese ribotype Ib transmits furoviruses and bymoviruses.     

Improved control of black rot of broccoli caused by <Emphasis Type="Italic">Xanthomonas campestris</Emphasis> pv. <Emphasis Type="Italic">campestris</Emphasis> using a bacteriophage and a nonpathogenic <Emphasis Type="Italic">Xanthomonas</Emphasis> sp. strain

We examined the potential for biological control of black rot of broccoli, caused by Xanthomonas campestris pv. campestris (Xcc), using nonpathogenic Xanthomonas sp. strain 11-100-01 (npX) mixed with bacteriophage XcpSFC211 (pXS). Inoculation of intact broccoli plants in greenhouse trials with either npX or pXS did not control black rot. After injured plant inoculation, however, npX alone or npX with pXS significantly controlled black rot. When a mixed suspension of npX with pXS was placed on a membrane filter, then washed with distilled water and air-dried, a substantial amount of pXS adsorbed to the surface of npX. In a field trial, broccoli plants were sprayed with a suspension of npX with pXS, then inoculated with Xcc. A meta-analysis of the results from five field trials showed an integrated risk ratio (IRR, the ratio of disease incidence in inoculated broccoli plants to the incidence in control plants) of 0.69 after treatment with only npX and 0.59 with npX with pXS, indicating that black rot incidence was significantly reduced by each treatment. The difference between these two treatments was also significant. IRR was 1.24 when comparing suppression by npX with pXS and that by basic copper sulfate wettable powder; thus, their control was comparable. The combination of npX with pXS improved the preventive effect against black rot. This is the first report describing that a nonpathogenic Xanthomonas sp. strain mixed with a bacteriophage effectively controlled black rot of broccoli in field trials.     

Efficacy of some rhizospheric and endophytic bacteria <Emphasis Type="Italic">in vitro</Emphasis> and as seed coating for the control of <Emphasis Type="Italic">Fusarium culmorum</Emphasis> infecting durum wheat in Tunisia

Sixty two rhizospheric and endophytic bacterial strains were evaluated for their biocontrol effect on two aggressive Fusarium culmorum isolates (Fc2 and Fc3). We observed that 35 % and 23 % of the tested strains inhibited the in vitro growth of Fc2 and Fc3 respectively. The observed antagonism was due to inhibition by contact (13–19 % of the strains) or at distance (10–16 % of the strains) for both fungal isolates. Some of the antagonistic bacteria showed the ability to produce diffuse and/or volatile compounds that inhibit the growth, the sporulation and macroconidia germination of F. culmorum. None of the tested antagonistic bacteria showed chitinase activity on synthetic medium. The sequencing of the 16S rDNA genes of some antagonistic bacteria showed that they belong to the genera Bacillus, Pseudomonas and Microbacterium. The double inoculation of durum wheat seeds by the antagonistic bacterial strains (B13, B18, BSE1, BSE3 and B16E) and the two F. culmorum isolates showed that germination and seedling vigor were generally improved in vitro. The percentage of infected seeds was also reduced. In greenhouse trials, the biocontrol effectiveness of F. culmorum was dependant from the virulence of the fungal strain and the specificity of the antagonistic interaction between bacterial and fungal strains. The bacterial strains B18 and B16E reduced F. culmorum infection on durum wheat plants probably due to their antagonistic and plant growth promoting activities and they may be used in a mixture as seed biopriming inoculum for plant growth bio-promoting and Fusarium wheat diseases biocontrol.     

Antagonistic activity of <Emphasis Type="Italic">Trichoderma</Emphasis> spp. against <Emphasis Type="Italic">Scytalidium lignicola</Emphasis> CMM 1098 and antioxidant enzymatic activity in cassava

Trichoderma spp. are used as antagonists against different pathogens. Despite many possibilities of using Trichoderma as an antagonist, there are gaps in the knowledge of the interaction between Trichoderma, cassava and Scytalidium lignicola. This fungus causes cassava black root rot and is an inhabitant of the soil, so it is difficult to control. Antagonists may contribute to the possible induction of resistance of plants because, when exposed to such pathosystems, plants respond by producing antioxidative enzymes. The test for potential inhibition of growth of S. lignicola CMM 1098 in vitro was performed in potato-dextrose-agar with two Trichoderma strains T. harzianum URM3086 and T. aureoviride URM 5158. We evaluated the effect of the two selected Trichoderma to reduce the severity of cassava black root rot and shoots. Subsequently, the production of enzymes (ascorbate peroxidase, catalase, peroxidase and polyphenol oxidase) was evaluated in cassava plants. All two Trichoderma strains show an inhibition of the growth of S. lignicola CMM 1098. The most efficient was T. harzianum URM 3086, with 80.78% of mycelial growth inhibition. T. aureoviride URM 5158 was considered the best chitinase producer. All treatments were effective in reducing severity, especially treatments using Trichoderma. Cassava plants treated with T. aureoviride URM 5158 had the highest enzyme activity, especially peroxidase and ascorbate peroxidase. Trichoderma harzianum URM3086 and Trichoderma aureoviride URM 5158 were effective in reducing the severity of cassava black root rot caused by S. lignicola CMM 1098.     

Development of two strains of <Emphasis Type="Italic">Fusarium culmorum</Emphasis> with a different aggressiveness in the soil and on the roots of barley of two genotypes

Fusarium culmorum can affect plants including cereals or develop saprophytically in the soil. It is unknown whether its saprophytic ability is different in strains with a different aggressiveness. This knowledge could be used for effective breeding of resistant cultivars. Here, we aimed to study the development of two F. culmorum strains with a different aggressiveness in the soil under barley, to compare their saprophytic ability and to reveal the influence, if any, of the host plant on the development of strains in the soil and on the roots. Saprophytic development of the strains was studied on membranes inoculated with macroconidia and placed into non-sterile soil under barley of two genotypes. The fungus was identified on the membranes and on barley roots by indirect immunofluorescent method. Both strains could develop saprophytically. The more aggressive strain (Fc538) showed a lesser saprophytic fitness than the less aggressive strain (Fc885): its mycelial density was lower and the number of chlamydospores was greater. Barley genotypes influenced the development of the fungal strains. Interestingly, conditions for saprophytic development of both strains were more favourable in the soil under barley of the resistant genotype. The more aggressive strain colonized barley of the resistant genotype more actively. Rot symptoms appeared earlier in the barley of the resistant genotype, but the number of diseased plants was greater in the barley of the susceptible genotype. The presence of a saprophytic stage in life cycle should slow down the accumulation of aggressive races in field populations of F. culmorum. Possible interactions between F. culmorum strains and barley plants are discussed.     

<Emphasis Type="Italic">Dittrichia viscosa</Emphasis> and<Emphasis Type="Italic">Rubus ulmifolius</Emphasis> as reservoirs of aphid parasitoids (Hymenoptera: Braconidae: Aphidiinae) and the role of certain Coccinellid species

The role of the self-sown shrubsDittrichia viscosa (L.) W. Greuter andRubus ulmifolius Schott as reservoirs of aphid parasitoids was investigated. In the field studies conducted,D. viscosa grew adjacent to crops of durum wheat and barley andR. ulmifolius grew adjacent to cotton. The relative abundance of the parasitoids of(a) Capitophorus inulae (Passerini) onD. viscosa, (b) Rhopalosiphum padi (Linnaeus) on durum wheat and barley,(c) Aphis ruborum (Börner) onR. ulmifolius, and(d) Aphis gossypii Glover on cotton in various parts of Greece, was assessed during the years 1996–2000. In 2000, the fluctuation of parasitization of the above four aphid species was recorded and the action of the aphidophagous predators of the family Coccinellidae was studied. It was observed thatAphidius matricariae Haliday predominated onC. inulae andR. padi in all sampling cases. In contrast,Lysiphlebus fabarum (Marshall) was the dominant species parasitizingA. ruborum onR. ulmifolius andA. gossypii on cotton in Thessaly (central Greece) and Macedonia (northern Greece), whereasLysiphlebus confusus Tremblay et Eady andBinodoxys acalephae (Marshall) were the dominant parasitoid species in Thrace (northern Greece).Coccinella septempunctata Linnaeus was the most abundant coccinellid species on durum wheat, whereasAdonia variegata (Goeze) predominated on cotton. However, coccinellid individuals were scarce on bothD. viscosa andR. ulmifolius. The present study indicated that these two shrubs can be regarded as useful reservoirs of aphid parasitoids.