Molecular Identification of a ‘Candidatus Phytoplasma ziziphi’‐related Strain Infecting Amaranth (Amaranthus retroflexus L.) in China

Amaranth (Amaranthus retroflexus L.) is a common weed that grows vigorously in orchards, roadside verges, fields, woods and scrubland in China. In 2009, phytoplasma disease surveys were made in orchards in Beijing, China, and stem/leaf tissues were collected from asymptomatic amaranths. Direct PCR using universal phytoplasma primers P1/P7 detected 16S rRNA gene sequences in every DNA sample extracted from the symptomless amaranths. Sequence alignment and phylogenetic analyses of the 16S rRNA gene determined that the amaranth phytoplasma strain was related to ‘Candidatus Phytoplasma ziziphi’. Furthermore, virtual RFLP pattern analysis showed that the amaranth phytoplasma belonged to the 16SrV‐B subgroup. This is the first report of symptomless plants containing a ‘Candidatus Phytoplasma ziziphi’‐related strain.     

Identification and Pathogenicity of Rhizoctonia spp. Causing Wirestem of Betula nigra in China

During July 2004, wirestem was frequently observed on the seedlings of Betula nigra at Dehong district in Yunnan Province, China. Isolates of Rhizoctonia spp. consistently obtained from their diseased leaves, roots and stems were identified as belonging to binucleate Rhizoctonia anastomosis groups (AG) AG‐P and AG‐R, and R. solani AG‐I IB and AG‐4 HG‐I, based on cultural characteristics, nuclear staining, anastomisis reaction and analysis of their ITS rDNA region. The percentage of recovery of AG‐P, AG‐1, AG‐R and AG‐4 was 48%, 39%, 8% and 3%, respectively. This is the first report of wirestem of red birch cause by binucleate Rhizoctonia AG‐P and AG‐R, and R. solani AG‐1 IB and AG‐4 HG‐I in China.     

Characterization and Sensitivity to Fungicides of Rhizoctonia spp. Recovered from Potato Plants in Bolu,Turkey

Isolates of Rhizoctonia spp. associated with stem canker and black scurf disease of potato were examined for their anastomosis group, sequence variations in the ITS‐5.8S rDNA region, pathogenicity and sensitivity to fungicides. A total of 92 isolates were obtained from diseased tuber, stolon and sprouts of the potato plants, collected from five districts of Bolu province, Turkey. Based on the anastomosis group and the similarity of the nucleotide sequence of the ITS‐5.8S rDNA, most of the isolates (81.5%) were identified as AG 3 PT. Other isolates belonged to AG 2‐1 (1.08%), AG 2‐2 IV (1.08%), AG 4 HG II (8.07%), AG 5 (2.17%), binucleate Rhizoctonia AG A (1.08%) and AG K (4.35%). Pathogenicity tests showed that isolates of AG 3 PT, AG 4 HG II and AG 5 caused similar degrees of disease severity on 45‐day‐old potato seedlings, whereas AG 2‐1 was moderately virulent. AG 2‐2 IV and binucleate Rhizoctonia spp. were weakly pathogenic or non‐pathogenic on potato seedlings. In this study, anastomosis groups of Rhizoctonia spp. isolates associated with potato in Turkey were characterized for the first time using molecular techniques and classified at the level of subgroups. Furthermore, the effect of selected fungicides was evaluated on disease development caused by soil‐borne inoculums of different anastomosis groups (AGs). Flutolanil and Bacillus subtilis QST 713 were found to be most effective against the Rhizoctonia isolates tested. These results revealed significant differences among the fungicides on disease development resulted from the different AGs.     

First Report of AG-A of Binucleate Rhizoctonia in China, Pathogenic to Soya Bean, Pea, Snap Bean and Pak Choy

Twenty binucleate Rhizoctonia (BNR) isolates were collected from roots of soya bean, pea, snap bean and pak choy with root rot symptoms in Yunnan Province, China. Chinese isolates anastomosed with the tester isolate of anastomosis group‐A (AG‐A; C‐517) with a high C2 fusion rate (>70%). Chinese isolates were pathogenic to soya bean, pea, snap bean and pak choy and had 97% similarity sequence of 5.8S rDNA‐internal transcribed spacer with AG‐A tester isolates SN‐2 and C‐662. When compared with other groups, AG‐Ba and AG‐Bb, Chinese isolates showed 77% sequence similarity. These results show that Chinese isolates belong to AG‐A of BNR. Growth rate, hyphal diameter, cultural characteristics and pathogenicity of the Chinese isolates differed significantly from the tester isolate of AG‐A. This is the first report on AG‐A in China.     

Increases in 1,5-Anhydroglucitol Levels in Germinating Amaranth Seeds and in Ripening Banana

To examine whether 1,5-anhydroglucitol (AG) is derived from starch degradation in plant tissues, we colorimetrically measured AG contents of germinating amaranth seeds and ripening banana pulp. In both cases, as starch degradation proceeded, AG levels were significantly increased, but were 1,700-5,000 times lower than those of total soluble carbohydrates. α-1,4-Glucan lyase activity, which is measured by the 1,5- anhydrofructose (AF) liberated from non-reducing glucose residues of starch or glycogen, was too low to be detected in amaranth or banana by the 3,5-dinitrosalicylic acid method. On the other hand, AF reductase, which reduces AF to AG, was detected in germinating amaranth seeds and banana pulp. Thus, the increases in AG levels are conceived to be derived from starch breakdown, although further investigation is needed to answer whether the starch degradation pathway via α-1,4-glucan lyase/AF reductase exists in plant tissues.     

Genetic Diversity of Thanatephorus cucumeris Infecting Tomato in Iran

The necrotrophic fungus Thanatephorus cucumeris (anamorph Rhizoctonia solani) is among the most important soil‐borne pathogens which causes tomato foot and root rot worldwide. We investigated virulence and genetic relationships among and within different taxonomic groups of R. solani from the tomato‐growing regions in the north‐east of Iran. Characterization of R. solani taxonomic groups revealed that, of 56 isolates, four were AG‐2‐1, 16 were AG‐3 PT, 21 were AG‐4 HG‐I and 15 were AG‐4 HG‐II. Because interprimer binding site (iPBS), which is based on amplification of retrotransposons, is known as novel and powerful DNA fingerprinting technology, we selected four iPBS primers, which can detect polymorphisms of tomato foot root and root rot pathogen, for investigating genotypic variability of the isolates. The iPBS analyses separated various taxonomic groups of R. solani and showed great diversity among the isolates, demonstrating that the R. solani isolates obtained from tomato were not a clonal population. Crop rotation strategies and geographic location seem to be important factors affecting genetic structure of the isolates. Pathogenicity tests on tomato cultivar ‘Mobil’ showed significant differences in the virulence of various isolates. The overall results indicated that isolates of AG‐3 and AG‐4 were more virulent than AG‐2‐1. There was no significant correlation between genetic diversity and virulence of the isolates. This is the first report of R. solani AG‐4 HG‐II, causing tomato foot and root rot. Also, our research is the first in assessment of genetic diversity in fungal populations using iPBS molecular markers.     

Identification and Molecular Characterizations of Neoscytalidium dimidiatum Causing Stem Canker of Red‐fleshed Dragon Fruit (Hylocereus polyrhizus) in Malaysia

During the year 2008 to 2009, a new disease of stem canker was noticed in most red‐fleshed dragon fruit (Hylocereus polyrhizus) plantations in Malaysia. The symptoms observed were small circular sunken orange spot, black pycnidia and rotted stem. This study was conducted to determine the occurrence of the stem canker on H. polyrhizus in Malaysia, subsequently to isolate, identify and characterize the fungal pathogen based on morphology and molecular characteristics and pathogenicity test. From the surveyed 20 plantations in Malaysia, stem canker was detected in all the plantations. A total of 40 isolates of Scytalidium‐like fungus were isolated and identified as Neoscytalidium dimidiatum based on morphological characteristics and ITS region sequences, which showed 99% similarity to N. dimidiatum (FJ648577). From the phylogenetic analysis using maximum‐likelihood tree, isolates of N. dimidiatum from stem canker of H. polyrhizus were grouped together and did not show any sequence variation. From pathogenicity test, all 40 isolates of N. dimidiatum were pathogenic causing stem canker on H. polyrhizus. To our knowledge, this is the first report of stem canker of H. polyrhizus caused by N. dimidiatum in Malaysia.     

Detection of the antimicrobial peptide gene in different Amaranthus species

Using primers to amplify the gene AMP2 in Amaranthus caudatus, we found the gene to be present in seven other species of the Amaranthus genus (A. albus, A. cruentus, A. blitum, A. hybridus, A. hypochondriacus, A. retroflexus and A. tricolor), in which it had not been described previously. The PCR products were sequenced and it was established that all the sequences were identical, except for two polymorphisms. These single nucleotide polymorphisms occurred at nucleotide positions 45 and 246. This exchange of one nucleotide for another was manifested in an amino acid change in both cases. Due to the fact that both polymorphisms lay outside the region encoding the chitin-binding peptide domain, which is crucial for antimicrobial peptide function, they will not likely affect the proper functioning of the peptide. With the exception of the above-mentioned polymorphisms, all sequences were identical to the sequence of the AMP2 gene that codes for the A. caudatus Ac-AMP2 (antimicrobial peptide isolated from Amaranthus caudatus seeds). The detection of sequences with high degree of sequence similarity to A. caudatus AMP2 gene leads us to the assumption that an antimicrobial peptide could also be produced by other amaranth species.     

Amaranth oil reduces accumulation of 4-hydroxynonenal-histidine adducts in gastric mucosa and improves heart rate variability in duodenal peptic ulcer patients undergoing Helicobacter pylori eradication

Helicobacter pylori-induced oxidative stress in gastric mucosa (GM) is a milieu for the development of chronic gastritis, duodenal peptic ulcer (DPU), gastric cancer, and a number of extragastric diseases. Because our previous study revealed the accumulation of the protein adducts of lipid peroxidation product 4-hydroxynonenal (HNE) in GM, which persists after eradication of H. pylori, the aim of the study was to test whether Amaranth oil supplementation in addition to standard anti-Helicobacter treatment could prevent such accumulation of HNE in GM in H. pylori-positive DPU patients. Seventy-five patients were randomly split into two groups: group 1 – standard treatment (n?=?39) and group 2 – standard treatment with additional supplementation of 1?ml of concentrated oil from amaranth seeds (Amaranthus cruenthus L., n?=?36). Clinical analysis, including endoscopy with biopsies from antrum and corpus of the stomach were performed before and after the treatment, as was heart rate variability (HRV) recorded, as parameter of systemic, extragastric pathophysiological alterations in DPU patients. Improvement of clinical, endoscopic and histologic manifestations, and successful ulcer healing were observed in both the groups. Moreover, supplementation of amaranth oil in addition to standard anti-H. pylori treatment significantly reduced accumulation of HNE-histidine adducts in GM and increased HRV in DPU patients (p?相似文献   

Domestication,phylogeny and taxonomic delimitation in underutilized grain Amaranthus (Amaranthaceae) – a status review

Many neglected, underutilized species are extremely important for food production especially in Low Income Food Deficient Countries (LIFDCs). Grain amaranth is one of such crops originated and domesticated in different parts of American continent. Lack of knowledge on taxonomy and phylogenetic relationship with other related crops, analysis of the extent and distribution of genetic diversity together with work on local and traditional knowledge, are the main constrains for genetic improvement of neglected, underutilized and crop related wild species. The phylogenetic relationship and taxonomic delimitation in genus Amaranthus are still not resolved with extreme clarity. But classification of the genus Amaranthus into three subgenera viz., Acnida, Amaranthus and Albersia, based on morphological parameters is quite acceptable. Phylogenetic analysis revealed clear separation of vegetable and grain amaranths. The derivation of grain and vegetable amaranths represent two lines of descent from weed progenitor. Amaranthus hybridus L. along with grain amaranths supposed to have formed a complex in which taxonomic problems are far from being clarified, especially because of apparent common hybridization and misapplication of nomenclature. (© 2014 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Characterization of microsatellite loci developed for <Emphasis Type="Italic">Amaranthus hypochondriacus</Emphasis> and their cross-amplifications in wild species

The present study reports isolation and characterization of 12 polymorphic microsatellite markers for Amaranthus hypochondriacus. A total of 92 alleles were detected across the 20 accessions, with an average of 7.7 alleles per locus. The observed (H _O ) and expected (H _E ) heterozygosity values ranged from 0 to 0.95 and from 0.49 to 0.92, respectively. At significance threshold (P < 0.05), nine loci deviated from Hardy-Weinberg equilibrium (HWE), whereas significant linkage disequilibria (LD) were observed between five pairs of loci. The 12 loci were successfully amplified in 18 other amaranth species representing cultivated grain and vegetable species, their putative progenitors and wild species. These results demonstrate wide potential applicability of these markers for the study of intra- and inter-specific genetic diversity as well as evolutionary relationships among cultivated and wild amaranths.     

Taxonomical observation on the grain amaranths and new varieties of Amaranthus cruentus (Amaranthaceae)

The grain amaranths (Amaranthus hypochondriacus, A. caudatus and A. cruentus) have a long history and a unique process of domestication. They are supposedly derived from a weed progenitor. The hypothesis proposing a monophyletic origin of the grain amaranths from the close relative A. hybridus is widely favoured. The present study applied morphological and biochemical tools, and revealed a close affinity between A. hypochondriacus and A. caudatus, and a distinct divergence of A. cruentus from the others, in accordance with previous observations. Accessions of A. cruentus showed a prominent variability, especially in seed surface features, which strongly proposed a segregation of the population into two groups; one having rusty brown seeds with reticulate spermoderm and another group having greyish white seeds with rugulate spermoderm: Amaranthus cruentus var. albus var. nov. and Amaranthus cruentus var. cruentus var. nov. This introduction was well supported by cluster analysis, showing a clear delimitation of brown and greyish white seeded accession of A. cruentus.     

First Report of Stem Canker on Phoenix Tree (Firmiana simplex) Caused by Fusarium oxysporum in China

A stem canker disease was observed on the phoenix trees located in the region of Dezhou, Shandong province. Symptomatic stems were collected and evaluated for the possible casual agent of the disease. A fungus resembling Fusarium sp. was consistently isolated from pieces of symptomatic tissues. The fungus formed abundant aerial mycelium on potato dextrose agar and produced the micro‐ and macro‐conidia on carnation leaf agar. The nucleotide sequences of the internal transcribed spacer of the rDNA from three representative isolates showed 100% identical to those of Fusarium oxysporum isolates deposited in the GenBank database. On the basis of morphological characteristics, pathogenicity test and molecular identification, the causal agent was identified as F. oxysporum. To our knowledge, this is the first report of stem canker on phoenix tree caused by F. oxysporum in China.     

An Investigation on Strains of Clavibacter michiganensis subsp. michiganensis in North and North West of Iran

Tomato bacterial canker disease was first reported from Urmiyeh in West Azerbaijan province in Iran. The disease causes lesion (canker), wilting and dryness of infected plants, leaf and fruit spots and the decline of the whole plant. Out of 102 isolates obtained from the fields in the major tomato producing areas of understudy regions, 98 were found Gram positive, yellow‐pigmented isolates, identified as Clavibacter michiganensis subsp. michiganensis based on the morphological and biochemical characteristics described in previous studies. Among these strains, 64 were virulent and 34 showed poor virulence. A strain of Cmm (NCPPB382) was used as a check (standard) in all steps of this study. DNA fingerprinting with repetitive‐sequence‐based PCR (rep‐PCR) (BOX primer) carried out among 11 representative strains (eight strains from West Azerbaijan, two from Golestan and one as standard). The most virulent strain was chosen as representative in each location. Dendrograms were prepared using NTSYS‐pc version 2/o2e software, unweighted pair group with arithmetic average method and simple matching similarity coefficient. According to the site of cut‐off line, three groups (clusters) with 82/5% similarity and six groups with 55% similarity were separated based on biochemical and SDS‐PAGE data, and rep‐PCR reactions respectively. Low similarity among groups (55%) can be explained as high genetic diversity among the strains. One strain of west Azerbaijan and the strains of Golestan, clustered in the same group suggesting that they may have been originated from a common source. Other strains of west Azerbaijan were clustered into different groups including II, III, IV, V and VI, suggesting the possibility of occurrence of different populations in a geographical region.     

Characterization and Pathogenicity of Rhizoctonia spp. from Onion in Amasya, Turkey

Forty‐two isolates of Rhizoctonia spp. were obtained from onion in Amasya, Turkey. Of these, 29% were Rhizoctonia solani (AG‐4), 69% were Waitea circinata var. zeae (Rhizoctonia zeae) and 2% were binucleate Rhizoctonia (AG‐B). Most of the isolates were recovered from rhizosphere soil. In pathogenicity tests on onion, R. solani AG‐4 caused the greatest disease severity, those of W. circinata var. zeae were moderately virulent but binucleate Rhizoctonia isolates were of low virulence. This is the first report of binucleate Rhizoctonia AG‐B and W. circinata var. zeae occurring on onion in Turkey.     

Fusarium spp. are Responsible for Shoot Canker of Kumquat in China

Shoot and branch canker and tree decline of kumquat (Fortunella margarita cv. Guban) were recorded in Yangshuo County, Guilin City, in the Guangxi Zhuang Autonomous Region of China during 2008–2011. Fusarium oxysporum and a new Fusarium species within the Gibberella fujikuroi complex (Fusarium sp. GLB1) were isolated repeatedly from the infected shoots and branches. Species identifications were verified by their high translation elongation factor 1‐alpha (TEF1) sequence similarity with those of the species epitypes. Koch's postulates were fulfilled on kumquat (cv. Guban) and mandarin establishing pathogenicity. To our knowledge, this is the first report of Fusarium shoot canker disease caused by F. oxysporum and Fusarium sp. on kumquat.     

Design,Synthesis, and Structure-Activity Relationship (SAR) Studies of Ketone-Isobenzofuranone Hybrid Herbicides

Thirty-five ketone-isobenzofuranone hybrids ( 1 – 35 ) were designed, synthesized, and evaluated for their herbicidal activity against Chinese amaranth (Amaranthus tricolor) and barnyard grass (Echinochloa crus-galli). The structure-activity relationship (SAR) results revealed that the position and type of substituent were crucial for activity. The o-substituted derivatives outperformed the m- and p-substituted derivatives. Compounds with strong electron-donating groups (OH, OMe) had low activity, while those with heterocycles (N-methylpyrrole, furan, and thiophene) had a moderate herbicidal effect. Compounds with a weak electron-donating group (Me) and weak, moderate, and strong electron-withdrawing groups (F, Cl, Br, and NO₂) showed promising herbicidal activity. Among these, the o-F substituted compound ( 20 ) was the most effective against Chinese amaranth, and the o-Cl substituted compound ( 23 ) was the most potent against barnyard grass. This is the first time the herbicidal potential of ketone-isobenzofuranone hybrids has been studied. The discovery of current chemical clues would be beneficial for the development of novel herbicides.     

Methicillin-resistant Staphylococcus pseudintermedius isolated from canine pyoderma in North China

Aims: To determine the prevalence of carriage of methicillin‐resistant Staphylococcus pseudintermedius (MRSP) among dogs with pyoderma from two small animal hospitals in North China during a 21‐month period and to characterize these isolates. Methods and Results: Swabs were taken from 260 dogs with pyoderma, and the staphylococcal species isolated and methicillin resistance were confirmed phenotypically and genotypically. The identified MRSP isolates were characterized by multilocus sequence typing (MLST), spa typing, staphylococcal cassette chromosome (SCC) mec typing, testing for susceptibility to nine antimicrobial agents and SmaI‐digested pulsed‐field gel electrophoresis. Thirty‐three (12·7%) dogs were positive for MRSP. The most prevalent genotypes detected among MRSP were ST71(MLST)‐t06(spa)‐II‐III(SCCmec) (n = 22, 66·7%), followed by ST5‐t19 (n = 8, 24·2%), ST126‐III(n = 2, 6·1%) and ST6‐t02‐V (3·0%). All MRSP isolates showed extended resistance to tested antimicrobial agents. Eight different SmaI patterns were observed in 21 typeable MRSP isolates. Conclusions: Clinical isolates of MSRP isolated from dogs in North China belonged to two major clonal lineages ST71 and ST5. Significance and Impact of the Study: This study is the first report on MRSP from canine pyoderma in China. Further surveillance study is needed to gain more detailed data concerning this major clinical challenge in veterinary medicine.     

Use of high resolution melting analysis to genotype the avirulence AvrLm4‐7 gene of Leptosphaeria maculans,a fungal pathogen of Brassica napus

The oilseed rape (Brassica napus) stem canker disease, due to the fungal pathogen Leptosphaeria maculans, is mainly controlled by host genetic resistance. Since 2004, the specific resistance gene Rlm7 is widely used in France. Specific resistance is effective when fungal populations are mainly composed of avirulent isolates. The development of molecular tools for the identification of virulent isolates towards Rlm7 was needed to undertake large‐scale surveys and to monitor the emergence of virulent populations in fields. Previous studies have described a large diversity of molecular events leading to virulence towards Rlm7, rendering conventional polymerase chain reaction (PCR) methods inapplicable to identify virulent isolates. Interestingly, a very limited nucleotide polymorphism was observed for avirulent, AvrLm7, alleles. Such characteristics were exploited here to develop a diagnostic method based on high resolution melting (HRM) analysis of the AvrLm4‐7 gene. High resolution melting analysis of a collection of 206 reference isolates revealed only four different profiles within 100 avirulent isolates and 87% of virulent isolates showed either no amplification or HRM curves distinct from those of avirulent isolates. The reliability of the method was confirmed using a second set of 119 unknown isolates, for which biological phenotyping and HRM genotyping were in agreement for 93% of the isolates. HRM combined with the PCR amplification of a larger fragment encompassing AvrLm4‐7 led to a correct diagnostic for 97.5% of the isolates.