Studies on the Isolation, Identification and In Vitro Growth Rates of the Three Pathogenic Fungi from Panax notoginseng Cultivated in Wenshan Eparchy

Objective] The aim of this study was to simultaneously isolate and identify the main pathogenic fungi of the root rot, black spot and round spot from the Panax notoginseng plants cultivated in Wenshan Eparchy of Yunnan Province of China. [Method] The pathogenic fungi were isolated and purified by using potato dextrose agar （PDA） medium. The morphological identification was accomplished first according to the colony forms of the fungi when cultivated in vitro, then accord-ing to the symptom characteristics and colony forms of the re-isolated fungi in the reverse inoculation experiments. The molecular identification was performed accord-ing to the amplification and alignment of the internal transcribed space （ITS） se-quences of the fungi. The increases of the diameters and thickness of the colonies of the fungi cultivated in vitro were employed to indicate the growth rates of the fungi. [Results] The consistency of the colony forms and symptom characteristics and the 96%-99% similarities revealed in the ITS sequence alignments al proved that the main pathogenic fungi of the root rot, black spot and round spot of the P. notoginseng plants raised in Wenshan were Cylindrocarpon didymium, Alternaria panax and Mycocentrospora acerina, respectively. When cultivated in vitro in the same temperature, humidity and il umination, the increases of the colony diameters and thickness of C. didymium were the highest, fol owed by those of A. panax, then those of M. acerina. During different cultivation periods, the differences of the colony diameters and thickness of the three fungi al reached extremely significant level. However, at the same cultivation time, the differences of the diameters and thickness among the three fungi only reached significant level. [Conclusion] The main pathogenic fungi which result in the root rot, black spot and round spot of the P. notoginseng in Wenshan are C. didymium, A. panax and M. acerina, respec-tively. When these three diseases break out at the same time, the root rot wil spread fastest, fol owed orderly by the black spot and the round spot.

25347例白带常规检查结果分析

目的了解妇科阴道病的病原菌检出率、病原菌分布、混合感染及阴道清洁度情况,为临床预防和治疗妇科疾病提供参考。方法采用盐水涂片湿片法和直接涂片革兰染色法同时对25 347例白带检验结果进行分析。结果有9 009例检出了病原菌/体,总阳性率为35.5%。其中真菌5 976例占23.5%,滴虫1 510例占6.0%,线索细胞775例占3.0%,纤毛菌580例占2.3%,革兰阴性双球菌168例占0.6%。混合感染203例,二重感染176例,三重感染24例,四重感染3例。清洁度为Ⅰ、Ⅱ度(正常)的样本4 535例占17.9%,清洁度为Ⅲ、Ⅳ度(不正常)的样本20 812例占82.1%。结论妇科阴道病由多种病原菌感染引起,受检者阴道清洁度多数处于不正常状态。

AIDS-Associated Mycoplasmas Infections in Patients with Nongonococcal Urethritis and Mucopurulent Cervicitis

Background: The incidence of sexually transmittedHIV infections is rapidly increasing in China, and theprevalence of AIDS-associated mycoplasmas (includingMycoplasma genitalium, Mycoplasma penetrans, Myco-plasma pirum and Mycoplasma fermentans) infections isunknown in patients with sexually transmitted diseases.Objective: To investigate the prevalence of these 4species of AIDS-associated mycoplasmas infections inpatients with nongonococcal urethritis (NGU) andmucopurulent cervicitis (MPC).Methods: In 65 patients with NGU/MPC, detection ofM. genitalium, M. penetrans, M. pirum and M. fermentansin genital and pharyngeal specimens was performed byculture and nested polymerase chain reaction.Results: M. genitalium, M. penetrans, M. pirum and M.fermentans were identified in genital specimens from23.1% (15/65), 12.3% (8/65), 1.5% (1/65) and 0% ofpatients, respectively, and from pharyngeal samples in26.2% (17/65), 15.4% (10/65), 1.5% (1/65) and 0% ofpatients, respectively. M. genitalium was detected in bothgenital and pharyngeal samples in 10.8% (7/65) ofpatients, and M. penetrans in 4.6% (3/65) of patients. M.pirum was found in only 2 cases, and no M. fermentanswas discovered.Conclusions: This study suggests that M. genitaliumand M. penetrans infections are common in patients withNGU/MPC. M. genitalium and M. penetrans may be trans-mitted by genital-genital or oral-genital sex, and maycause urethritis and cervicitis.

This year's Nobel Prize to gastroenterology:Robin Warren and Barry Marshall awarded for their discovery of Helicobacter pylori as pathogen in the gastrointestinal tract

TO THE EDITORPeptic ulcer disease is a major health care concern in the society today, in view of personal suffering as well as economical health care costs.

OsWRKY03, a rice transcriptional activator that functions in defense signaling pathway upstream of OsNPR1

WRKY family proteins are a class of plant specific transcription factors that involve in many stress response pathways. It has been shown that one Arabidopsis WRKY protein, AtWRKY29/22, is activated by MAP kinase signaling cascade and confers resistance to both bacterial and fungal pathogens. However, little is known about the biological roles of WRKY proteins in rice. In this study, we investigated the expression patterns of rice AtWRKY29/22 homolog, OsWRKY03, under different conditions, and also its possible role involved in plant defense. Our results showed that OsWRKY03 was up-regulated by several defense signaling molecules or different treatments. Further analysis revealed that the expression of OsWRKY03 was light dependent. Transcriptional activation activity of OsWRKY03 was also demonstrated by yeast functional assay. Transient expression of OsWRKY03-GFP fusion protein in onion epidermis cells showed that OsWRKY03 was a nuclear localized protein. OsNPR1 as well as several other pathogenesis-related genes, such as OsPRlb, phenylalanine ammonia-lyase （ZB8） and peroxidase （POX22.3）, were induced in OsWRKYO3-overexpressing transgenic plants. These results indicated that OsWRKY03 is located upstream of OsNPR 1 as a transcriptional activator in salicylic acid （SA）-dependent or jasmonic acid （JA）-dependent defense signaling cascades.

Distinct immune response induced by peptidoglycan derived from Lactobacillus sp

AIM： To analyze the distinct immune responses induced by Lactobacillus peptidoglycan （PG）. METHODS： BALB/c mice were intraperitoneally injected with PG once a day for three consecutive days, Peritoneal macrophage and splenocyte mRNA was extracted and the gene expression profile was studied using high-density oligonudeotide microarrays. Inhibitory effects of Lactobacillus PG on colon tumor tissue were studied in vitro and in vivo, RESULTS： The gene expression profiles revealed that the TLR-NF-kB and Jak-STAT signaling pathways were highly activated. An inflammatory phenotype was induced when peritoneal macrophages were initially exposed to Lactobacillus PG and switched to a more complex phenotype when BALB/c mice were treated with three doses of Lactobacillus PG. A protective physiological inflammatory response was induced after three consecutive days of PG treatment. It was tending toward Thl dominant immune response. Lactobacillus PG also appeared to induce a significant in vivo anti-colon tumor effect. CONCLUSION： Lactobacillus PG is responsible for certain immune responses induced by Lactobacilli. Anti-tumor effects of Lactobacilliare likely to attribute to the activation of macrophages by PG expressed on the bacterial cell surface.

Initial Screening of 10 Plant Essential Oils in Vitro Antibacterial Activity Against Blueberry Pathogens

In order to develop natural antistaling agent for blueberries preservation,the antibacterial activities of 10 kinds of plant essential oils were analyzed by oxford-cup test and tube dilution method,and their inhibitory abilities to two kinds of representative Gram bacteria which are Escherichia coli and Staphylococcus aureus,and three kinds of harmful fungi in blueberries,involving Alternaria alternata,Penicillium digitatum and Botrytis cinerea,were tested.The results showed that 7 kinds of essential oils can inhibit the three fungi,among which cinnamon oil,clove oil,anise oil and thyme oil have strong fungi static efficacy.Cinnamon oil and thyme oil can effectively inhibit the Gram-positive bacterium(Staphylococcus aureus);cinnamon oil and masson pine oil are strongly inhibitory to Gram-negative bacterium(E.coli).Cinnamon oil has broad-spectrum ability to inhibit fungi,Gram-negative and positive bacteria,with a minimum inhibitory concentration(MIC)ranging from 0.0391 to 0.1560 L/mL.With strong inhibition effects on fungi,cinnamon oil can be used as a compound base oil and mixed with other essential oils to retain the freshness of blueberries,thus achieving better antibacterial and anti-corrosion effects.

Infection of the Anther-smut Microbotryum violaceum in Populations of Silene dioica: Variation in Floral Morphology, Patterns of Spore Deposition and Pathogen-Pollinator Mediated Selection

The anther-smut Microbotryum violaceum ( Brandenburger and Schvinn) G. Deml. and Oberw.) causes a systematic infection of its host Silene dioica ( L.) Clairv., resulting in sterility and production of teliospores (dispersal propagules) in flowers. These spores are transmitted to healthy plants mainly by flower visitors. The behavioral responses of flower visitors to a variation in floral characters are not only likely to affect rates of pollen export/import, but also the rate of spore deposition and probability of disease. In a transplantation experiment, using plants from four different populations, we tested for correlation between variation in female floral morphology and patterns of spore and pollen deposition, and a resulting risk of disease. The source populations in this experiment were located on four islands in Skeppsvik archipelago in northern Sweden, and represented a gradient of disease incidence from completely healthy ( Island 1), low incidence ( Island 2) to high incidences ( Islands 3 and 4) of disease. Fifty plants from each population were transplanted to the center, of the population on Island 4. There were significant differences among the transplants in floral characters, i.e. corolla size, style length and ovule number. Plants from the non-diseased population had larger flowers and longer styles than plants from the highly diseased populations. Numbers of pollen grains and spores deposited on flowers were strongly and positively correlated. We found that plants originating from the non-diseased population captured approximately 4 times more pollen and 9 times more spores, per flower than die individuals from the resident population (Island 4, population 4). The incidences of disease among plants, from the four populations differed significantly, and was 37%, 20%, 18% and 0 for populations 1, 2, 3 and 4 respectively. In a survey of ten populations we found a significant negative correlation between the mean style length ( positively correlated with corolla size and ovule number) among healthy plants and incidence of disease in these populations. The potentiality for pathogen-pollinator mediated selection oil floral characters; and consequences for gone flow between populations of Silene dioica are discussed.

Fungal genus Hypocrea/Trichoderma:from barcodes to biodiversity

Hypocrea/Trichoderma is a genus of soil-borne or wood-decaying fungi containing members important to mankind as producers of industrial enzymes and biocontrol agents against plant pathogens, but also as opportunistic pathogens of immuno- compromised humans and animals, while others can cause damage to cultivated mushroom. With the recent advent of a reliable, BarCode-aided identification system for all known taxa of Trichoderma and Hypocrea, it became now possible to study some of the biological fundamentals of the diversity in this fungal genus in more detail. In this article, we will therefore review recent progress in (1) the understanding of the geographic distribution of individual taxa; (2) mechanisms of speciation leading to development of mushroom diseases and facultative human mycoses; and (3) the possible correlation of specific traits of secondary metabolism and molecular phylogeny.

Association between cag-pathogenicity island in Helicobacter pylori isolates from peptic ulcer,gastric carcinoma,and non-ulcer dyspepsia subjects with histological changes

AIM： To investigate the presence of the cag-pathogenicity island and the associated histological damage caused by strains with complete cag-PAI and with partial deletions in correlation to the disease status. METHODS： We analyzed the complete cag-PAI of 174 representative Helicobacter pylori （H pylori ） clinical isolates obtained from patients with duodenal ulcer, gastric ulcer, gastric cancer, and non-ulcer dyspepsia using eight different oligonucleotide primers viz cagA1, cagA2, cagAP1, cagAP2, cagE, cagT, LEC-1, LEC-2 spanning five different loci of the whole cag-PAI by polymerase chain reaction （PCR）. RESULTS： The complete screening of the genes comprising the cag-PAI showed that larger proportions of subjects with gastric ulcer （97.8%） inhabited strains with complete cag-PAI, followed by gastric cancer （85.7%）, non-ulcer dyspepsia （7.1%）, and duodenal ulcer （6.9%）, significant differences were found in the percentage distribution of the genes in all the clinical groups studied. It was found that strains with complete cag-PAI were able to cause severe histological damage than with the partially deleted ones. CONCLUSION： The cag-PAI is a strong virulent marker in the disease pathogenesis as it is shown that a large number of those infected with strain with complete cag-PAI had one or the other of the irreversible gastric pathologies and interestingly 18.5% of them developed gastric carcinoma. The presence of an intact cag- PAI correlates with the development of more severe pathology, and such strains were found more frequently in patients with severe gastroduodenal disease. Partial deletions of the cag-PAI appear to be sufficient to render the organism less pathogenic.

Interaction of non-adherent suspended neutrophils to complement opsonized pathogens: a new assay using optical traps

Phagocytosis of opsonized pathogens by circulating non-adherent neutrophils is an essential step in host defense, which when overwhelmed contributes to sepsis. To investigate the role played by ligation of complement receptors CR3 and CR4 in non-adherent neutrophils, we designed a novel assay system utilizing dual optical traps, respectively, holding a suspended unactivated cell and presenting a specific ligand-coated bead to the cell surface. We chose anti-CD 18 as an example ligand, mimicking the bacterial opsonizing complement fragment iC3b. Presentation of anti-CD 18-coated beads elicited both pseudopodial protrusion and subsequent phagocytosis. This is in sharp contrast to previously reported responses of adherent neutrophils, which phagocytize opsonized particles without pseudopod formation. We used this same new assay to probe actomyosin pathways in the neutrophil＇s pseudopodial and phagocytic response. Disruption of actin or inhibition of myosin light-chain kinase dose-dependently reduced pseudopod formation and phagocytosis rates. In summary, i） the new dual trap assay can be used to study the responses of suspended neutrophils to a variety of ligands, and ii） in a first application of this technique, we found that local ligation of CR3/4 in unactivated neutrophils in suspension induces pseudopod formation and phagocytosis at that site, and that these events occur via an actomyosindependent pathway.

Diversity of Gram negative bacteria antagonistic against major pathogens of rice from rice seed in the tropic environment

With the use of a seed washing technique, more than 4000 Gram negative bacteria were isolated by two improved isolation methods from 446 batches of 1 kg rice seed samples obtained from 22 provinces in the Philippines. They were initially characterized on the basis of colony morphology and results of biochemical and pathogenicity tests. Six hundred and fifty two strains were further identified by Biolog, from which 133 were selected for fatty acid methyl ester (FAME) analysis together with 80 standard reference!strains. Sixteen species or types of Pseudomonas and 17 genera of non pseudomonads were identified, more than one third of which have not been recorded in rice. The most predominant species observed were P. putida and P. fulva. About 17% of the strains of Pseudomonas and 2% of the non pseudomonads were antagonistic to one or more fungal or bacterial pathogens of rice. Rice seed is an important source of biological control agents.

Efficacy of current guidelines for the treatment of spontaneous bacterial peritonitis in the clinical practice

AIM： To verify the validity of the International Ascites Club guidelines for treatment of spontaneous bacterial peritonitis （SBP） in clinical practice. METHODS： All SBP episodes occurring in a group of consecutive cirrhotics were managed accordingly and included in the study. SBP was diagnosed when the ascitic fluid polymorphonuclear （PIN） cell count was 〉 250 cells/mm^3, and empirically treated with cefotaxime. RESULTS： Thirty-eight SBP episodes occurred in 32 cirrhotics （22 men/20 women; mean age： 58.6 ＋ 22.2 years）. Prevalence of SBP, in our population, was 27%. Ascitic fluid culture was positive in nine （24%） cases only. Eleven episodes were nosocomial and 71% community-acquired. Treatment with cefotaxime was successful in 59% of cases, while 41% of episodes required a modification of the initial antibiotic therapy because of a less-than 25% decrease in ascitic PMN count at 48 h. Change of antibiotic therapy led to the resolution of infection in 87% of episodes. Among the cases with positive culture, the initial antibiotic therapy with cefotaxime failed at a percentage （44%） similar to that of the whole series. In these cases, the isolated organisms were either resistant or with an inherent insufficient susceptibility to cefotaxime. CONCLUSION： In clinical practice, ascitic PMN count is a valid tool for starting a prompt antibiotic treatment andevaluating its efficacy. The initial treatment with cefotaxime failed more frequently than expected. An increase in healthcare-related infections with antibiotic-resistant pathogens may explain this finding. A different first-line antibiotic treatment should be investigated.

Study on the Biology and Epidemiology of Uncinula necator---The Causal Agent of Grape Powdery Mildew Disease

This study was carried out during 2007-2009 in the vineyards of Ardebil province of lran to study the biology and the epidemiology of Uncinula necator--the causal agent of grape powdery mildew disease. The study concentrated on the survival and the initiation of primary inoculum of the fungal causal agent. Results of histopathological experiments indicated that U. necator survived as mycelium in the dormant buds of the grapes during winter season. Results of study on the effect of environmental factors on fungus biology showed that the pathogenic activity of the fungus began when the temperature was between 16-19 ℃ with a relative humidity more than 50%. It was also found that optimum temperature and relative humidity for the sporulation of U.necator was 20-25 ~C and 50%-100% respectively. According to the results, fungal conidia were trapped during formation of 5-6 true leaves and first disease symptoms were observed on the clusters on late June after fruit formation. Fungal cleistothecia were observed abundantly at the end of season on the leaves, petioles and twigs but they were not able to survive during winter. Formation of ascospores on young leaves was proved but their role as the primary inoculum was not supported by the results of this study. Results of this study and the new findings on the biology and epidemiology of U.necator may be of national and international interests for the management of powdery mildew disease which is one of the most destructive diseases around the world including Iran.

A bacterial pathogen infecting gametophytes of Saccharina japonica(Laminariales,Phaeophyceae)

A newly identified bacterial disease of kelp(Saccharina japonica) gametophytes was found in clone cultures.It is characterized by swollen gametophyte cells in the early period of infection followed by filamentous fading.An alginolytic marine bacterium referred to as A-1 was isolated from the diseased gametophytes.On the basis of 16S rDNA sequencing and morphological,physiological and biochemical characteristics,the bacterium was identified as a strain of the genus Alteromonas.By testing Koch's postulates,Alteromonas sp.A-1 was further confirmed as the pathogen.The infection process was also investigated using both scanning electron and light microscopy.

Survey of O-islands in Escherichia coli O157 and Other Enteric Pathogens——O-islands of E.coli O157∶H7

The genome of the enterohemorrhagic Escherichia coli O157∶H7 EDL933 contains 177 “O”-islands (OIs). To study their potential contribution to the O157-specific pathogenicity, we surveyed the distribution of 22 OIs by PCR and DNA hybridization in 17 isolates of Shiga toxin producing (Stx-positive) E.coli O157∶H7, and compared with their distribution in 21 isolates of Stx-negative E.coli O157 and 21 isolates of non-O157 enteric pathogens. Fourteen of 22 OIs were present in non-O157 entericpathogens analyzed. Eight of 22 OIs were found only in the 17 Shiga toxin- (Stx) positive E.coli O157∶H7 isolates, but they were absent from the 21 Stx-negative E.coli O157∶NM and O157︰Hund isolates tested. Among the 8 OIs, only OI43 or OI48 were exclusively detected in Stx-positive E.coli O157∶H7, absent from neither of Stx-negative E.coli O157 and non-O157 enteric pathogens, such as Salmonella, Shigella, Citrobacter, Vibrio cholera, enteropathogenic E.coli (EPEC), enteroadherent E.coli (EAEC), enteroinvasive E.coli (EIEC) and enterotoxingenic E.coli (ETEC). The OI43 and OI48 are 83 kb in size and identical in DNA sequences, which encode genes for urease, tellurite resistance and adherence. By analyzing their junction genes with PCR and DNA hybridization, we found that 21 Chinese isolates have OI48 only. However, for 7 Japanese patient isolates, 4 have OI43 and 3 have OI48; for American isolates, 2 have both of OI43 and OI48, 2 have OI48 only. These data confirmed the highly plasticity of the pathogenic E.coli genome. The unique presence of OI43/OI48 in Stx-positive E.coli O157∶H7 denotes its critical role in the pathogenicity specific to this pathogen.

In Vitro Activity of Lawsonia inermis （Henna） on Some Pathogenic Fungi

The present study was conducted to investigate antifungal activity ofLawsonia inermis （Henna plant）. Leaf samples of the plants were collected from Eastern Nile of Khartoum state, Sudan. Ethanol and petroleum ether extracts in various concentrations were obtained by maceration （cold method）. The extracts were bioassay in vitro to know their bioactivity to inhibit the growth of tested fungi. The cup plate agar diffusion method was adopted to assess the antifungal activity of the extracts against tested yeasts while agar incorporated method was used for other molds. Both extracts revealed anti fungal activity against all yeast strains except Pichiafabianii which was found resistant to both ethanol and ether extracts. The results displayed antifungal activity against tested fungi. Minimum mould concentration （MMC） of the extracts was determined. The obtained results revealed antifungal activity of henna leaves extracts which support the traditional use of the Henna in therapy of fungal infections. The possibility of therapeutic use of Sudanese henna as antifungal agents is recommended.

Alteration in Biochemical Response in Sesamum indicum upon Different Plant-Pathogen Interactions

Sesamum, an important oil yielding crop suffers a huge loss in its yield due to attack of large number of fungal pathogens. In semi-arid regions Sesamum is mainly affected by two major plant-pathogenic fungus viz. Macrophomina phaseolina and Fusarium oxysporum. The aim of the study was to analyze the metabolic alterations in Sesamum after infection with both pathogens. This accomplished by individually by （the word estimating is not quantitative） the levels of total phenolic compounds and the activities of phenylalanine ammonia lyase （PAL） of one week old plants. The PAL showed high activity in infected plants, revealing the active phase in the synthesis of secondary metabolites in the Sesamum plant after infection. As a consequence, in infected plants the contents of polyphenols along with salicylic acid （SA） considerably exceeded when compared to control plants. This in vivo study of M. phaseolina and F. oxysporum infection reveals the differences of resistance levels in sesame against these two pathogens. These results provide important information regarding the plant-pathogen interactions and also forfor Sesamum improvement programs seeking the adaptation to diverse range of fungal attack along with adverse environmental factors.

Clustering and Expression Analysis of Chitinases in Maize and Rice

Chitinases play an important role in regulating plant growth and development, especially defending themselves from fungal pathogens. It is important to do the biological analyses in crops. In this study, the result showed that the chitinases were distributed into the whole genome in rice, and nearly the whole genome in maize expect for Chromosome 9. The clustering results showed that one out of three chitinases from maize and rice belonged to new groups, which were separated from those in the conformed Classes I-VII. The identification of most amino acid sequences was very low among the chitinases from rice and/or maize. It was inferred that the chitinases with different functions were relatively stable during plant evolution. The relationship of chitinases expression between leaf blade and anther was positively significant in maize, but not significant in rice. Additionally, the ratio of chitinases with up- or down-regulated expression in sensitive maize under Fusarium moniliforme inoculation was different from that in sensitive rice under Magnaporthe grisea inoculation. It might be result from different tissues infected by different fungi. The number of chitinases from resistant maize was less than that from sensitive maize, which inferred that the resistant pathways on F. moniliforme should be not only chitin induced Pathogen-associated molecular PTI （patterns-triggered immunity） pathway, but also might include other PTI pathways that improve tolerance to F. moniliforme. The analysis of expression pattern of chitinases from maize and rice under fungi inoculation will be contributed into further research on the defense mechanism of fungi in crops.

Assessment of Seed-Borne Plant Pathogenic Fungi Associated with Rice Crop in South-Eastern Nigeria

This study was conducted to assess the incidence of seed-borne fungi on rice seeds sourced from the major rice growing areas of South-Eastern Nigeria. The rice seeds were collected during the dry seasons between 2009 and 2010. Four hundred rice seeds randomly collected and surface sterilized by washing in 3% sodium hypochlorite and rinsed two times with sterile distilled water were placed in three circular rings inside the Petri dishes. The incubation was done one after the other for each variety from all the locations. Ten fungi species from eight genera were isolated from the rice seeds in all the rice growing areas of South-Eastern Nigeria. The fungi pathogens vary from one locality to another, as well as the frequency of occurrence. The pathogens isolated include： Fusarium moniliforme, F. solani, F. oxysporium, Aspergillus spp., Botridiploidia spp., Helminthosporium spp., Penicillium spp., Trichoconis padwickii, Curvularia oryzae and Phoma oryzae. Though, there was the presence of some seed-borne fungi in the rice seeds from the areas studied, most of the cultivars experienced up to 90% germination. But some cultivars had very low germination. IR 1416 from Arochukwu and GB90 from Arondiuzogu had less than 1% germination. This study shows that seed-borne organisms are major constraint in rice production in the South-Eastern states of Nigeria as a result of low seed germination stemmed from infestation of fungi organisms in the seeds.