Pepper Pathogen Research Articles

Antifungal activity of cow urine distillates of local botanicals against major pathogens of bell pepper

Cow urine distillates of botanicals viz, Ranunculus muricatus, Melia azedarach, Vitex negundo, Eupatorium sp. and Murraya koenigii and cow urine alone were evaluated at 0.5, 2.0, 4.0, 6.0, 8.0 and 10.0% concentrations under in vitro conditions against bell pepper pathogens namely Rhizoctonia solani, Sclerotinia sclerotiorum, Sclerotium rolfsii, Phytophthora nicotianae, Fusarium oxysporum f.sp. capsici, Fusarium solani and Colletotrichum capsici. M. koenigii and V. negundo provided the complete inhibition in mycelial growth of C. capsici, Rhizoctonia solani, Phytophthora nicotianae, Sclerotinia sclerotiorum, Fusarium solani and S. rolfsii at 10% concentration. R. muricatus and Murraya azedarach caused 100% inhibition of R. solani, S. sclerotiorum and S. rolfsii whereas more than 98% inhibition in mycelial growth of F. solani, P. nicotianae, R. solani and C. capsici was observed at 10% concentration. Eupatorium sp. showed 63.05 to 99.69% inhibition against all the test pathogens. Inhibition in rest of the concentrations ranged between 85.06 to 98.97%. Cow urine distillate exhibited more than 98% inhibition in mycelial growth of P. nicotianae and S. sclerotiorum and 93% inhibition of C. capsici, R. solani, F. solani, F. oxysporum f.sp. capsici and S. rolfsii at 10% concentration and inhibition in rest of concentrations was observed to the tune of 59.70 to 96.77%. Cow urine distillates of botanicals were found more inhibitory to pathogens than cow urine distillate alone. Key words: Antifungal activity, cow urine distillate, Ranunculus muricatus, Melia azedarach, Vitex negundo, Eupatorium sp., Murraya koenigii.

Inactivation of Escherichia coli O157:H7 and Salmonella Typhimurium in black pepper and red pepper by gamma irradiation

This study evaluated the efficacy of gamma irradiation to inactivate foodborne pathogens in black pepper (Piper nigrum) and red pepper (dried Capsicum annuum). Black pepper and red pepper inoculated with Escherichia coli O157:H7 and Salmonella Typhimurium were subjected to gamma irradiation in the range of 0, 1, 2, 3 and 5kGy, and color change was evaluated after treatment. Pathogen populations decreased with increasing treatment doses. A gamma irradiation dose of 5kGy decreased E. coli O157:H7 and S. Typhimurium populations >4.4 to >5.2logCFU/g in black pepper without causing color change. Similarly, 5kGy of gamma irradiation yielded reduction of 3.8 to >5.2logCFU/g for E. coli O157:H7 and S. Typhimurium in red pepper. During gamma irradiation treatment, L⁎, a⁎ and b⁎ values of red pepper were not significantly changed except for 297μm to 420μm size red pepper treated with 5kGy of gamma irradiation. Based on the D-value of pathogens in black pepper and red pepper, S. Typhimurium showed more resistant to gamma irradiation than did E. coli O157:H7. These results show that gamma irradiation has potential as a non-thermal process for inactivating foodborne pathogens in spices with minimal color changes.

Development of a novel codominant molecular marker for chili veinal mottle virus resistance in Capsicum annuum L.

Chili veinal mottle virus (ChiVMV) is one of the most destructive pepper pathogens in Asia. Development of ChiVMV-resistant cultivars is necessary to control ChiVMV infection on pepper farms. However, sources of variation for ChiVMV resistance have not been identified and only a recessive resistance gene has been identified. We initially screened 30 pepper lines from several countries using inoculation tests to further examine inheritance of ChiVMV resistance, to establish a relevant breeding program, and to develop a new resistant line. Here, we report a new genetically dominant source of resistance to ChiVMV in pepper. Secondly, we found two amplified fragment length polymorphisms linked to dominant resistance and converted them into high-resolution melting markers, which were located on chromosome 6. Furthermore, we obtained a cleaved amplified polymorphic sequence marker that was closer to the ChiVMV resistance locus using comparative mapping. The newly discovered marker, related to a single dominant gene, will help researchers develop a new ChiVMV-resistant pepper cultivar.

Systemic resistance and growth promotion of chili pepper induced by an antibiotic producing Bacillus vallismortis strain BS07

In the present study, the ability of plant growth-promoting rhizobacterium (PGPR), Bacillus vallismortis strain BS07 (BS07) was evaluated as one of the potential biocontrol agents against multiple pathogens in chili pepper. Under greenhouse conditions, chili pepper seedlings treated with BS07 or chemical control, benzothiadiazole (BTH) by soil drench suppressed the soft rot incidence significantly by reducing the percent disease lesion area per leaf disk compared to negative control. The treatment with BS07 or BTH induced plant resistance against phytophthora blight caused by Phytophthora capsici upon pathogen challenge. There was a greater reduction of anthracnose infection caused by Colletotrichum acutatum on matured fruits in BS07 or BTH treated plants when compared to untreated control. Under field conditions, percent of soft rot incidence and disease severity of phytophthora blight were significantly reduced in BS07 and BTH treated plants when compared to untreated control. In the case of anthracnose disease, the significant reduction of disease was observed during the two years on matured fruits, when compared to untreated control. The study also clearly showed that there was a significant increase of fruit yield in BS07 treated plants, and it has also been observed that BS07 treated plants showed higher chlorophyll content in the fresh leaves compared to untreated control. Reverse transcriptase (RT)-PCR analyses of Arabidopsis thaliana leaves revealed up-regulation of PR1a gene expression in Col-0, while there was no expression in Nag-G plants treated with BS07 suspensions. This suggests that the systemic resistance might be evolved by the mechanism of salicylic acid (SA) dependent pathway.

Solution Structure and Dynamics of a Plant Pathogen Effector

Plant diseases account for at least $220 billion of crop losses worldwide every year, and have a significant impact on global food security and bio-fuel availability. Bacterial, oomycetes and fungi pathogens secrete protein effectors into plant cells, where they perturb cellular processes, presumably to the benefit of the pathogen. In oomycetes, an N-terminal protein motif (RXLR) is important for targeting plant pathogen effectors into host cells (Whisson et al. 2007, Nature450, 115 and Oliva et al. 2010, Cell. Microbiol.12, 1015). However, little is known about the molecular mechanisms by which RXLR effectors manipulate host cell pathways. We have been studying the RXLR effector AVR3a11 (from Phytophthora capsici, a pathogen of pepper), which shares sequence similarity with the well-studied AVR3a effector from the Irish potato famine pathogen Phytophthora infestans (the causative agent of late blight in potato and tomato). Using a combination of 2D and 3D nuclear magnetic resonance (NMR) spectroscopy experiments, 75% of the AVR3a11 backbone has been assigned. With additional 13C-HSQC-NOESY and 15N-HSQC-NOESY experiments, a structural ensemble model has been generated which is in good agreement with the X-ray structure of AVR3a11 (PDB code 3ZR8, Boutemy et al. 2011, J. Biol. Chem.286, 35834). Backbone amide T1, T2 and heteronuclear NOE relaxation experiments at 800 MHz, show that AVR3a11 behaves as expected for a well ordered protein. However, the measured transverse relaxation is faster than theoretically expected for a protein this size, suggesting that conformational exchange may be occurring. Additional information on conformational exchange was obtained from hydrogen/deuterium exchange experiments.

Molecular Re-identification of Strains of the Colletotrichum acutatum Species Complex Deposited in the NIAS Genebank and Morphological Characteristics of its Member Species

Recently, nine phylogenetic groups were proposed for a plurivorous fungus, Colletotrichum acutatum by some researchers. Some of them were named as C. simmondsii, C. fioriniae, C. acutatum sensu stricto and C. carthami based on morphology and phylogenetic analyses of β-tubulin-2 gene partial sequences. We re-identified 170 strains deposited as C. acutatum in the NIAS Genebank, Japan based on phylogenetic analysis with the β-tubulin-2 gene sequences. Eighty strains with reddish colony colors and relatively uniform conidia belonged to C. fioriniae, 37 with a grayish to yellowish color and short conidia to C. simmondsii, 25 with a blackish to cream color and short conidia to C. carthami, 12 with a cream to pale orange color and narrow conidia to ‘C. acutatum group A2-P’ newly designated in this paper, 5 with a brownish to orange color and larger conidia to C. acutatum group A4, and 11 to other species of Colletotrichum. No strain of C. acutatum s. str. was found. A dichotomous key to the member species and groups was designated based on reverse colony color and conidial morphology. Because pathogenic strains of 18 host plants were re-identified, updating of pathogen names was proposed. Ten of them should be changed to C. fioriniae, and the pathogens of each five host plants such as strawberry and apple should be replaced with 2 to 3 species or the group. Sweet pepper pathogen was tentatively classified into C. acutatum group A2-P.

Bacillus sp. NAAS-1을 이용한 고추 탄저병 생물학적 방제

고추포장의 토양에서 분리한 Bacillus sp. NAAS-1 Colletotrichum acutatum에 대한 생물학적 방제 활성을 검정 하였다 이 분리균주를 일종의 작물보호제인 카밴다짐 가스가마이신 수화제와 비교하여 검정한 결과, 기준사용량 농도(50 <TEX>${\mu}L/mL$</TEX>)와 비슷한 항진균 활성을 나타냈다. 또한 생물 검정 결과, 공시 작물보호제의 기준사용량 농도(50 <TEX>${\mu}L/mL$</TEX>)에서 보다 고추 탄저병균의 발병억제효과가 높게 나타나내었다. Bacillus sp. NAAS-1 isolated from the field of Chili pepper was tested for biocontrol activity against anthracnose pathogen of Chili pepper caused by Colletotrichum acutatum. The antifungal activity of Bacillus sp. NAAS-1 culture broth was compared with synthetic fungicide containing carbendazim (40%) and kasugamycin (3.45%). Bacillus sp. NAAS-1 showed a similar fungicidal activity against the anthracnose pathogen at the concentration of 50 <TEX>${\mu}L/mL$</TEX> in comparison to the fungicide containing carbendazim (40%) and kasugamycin (3.45%) using a cup method. Bacillus sp. NAAS-1 also exhibited its potent fungicidal activity against the anthracnose in vivo test at the concentration of 50 <TEX>${\mu}L/mL$</TEX> when compared to the fungicide containing carbendazim (40%) and kasugamycin (3.45%).

Advances in Research on Phytophthora capsici on Vegetable Crops in The United States.

Since L. H. Leonian's first description of Phytophthora capsici as a pathogen of chile pepper in 1922, we have made many advances in our understanding of this pathogen's biology, host range, dissemination, and management. P. capsici causes foliar blighting, damping-off, wilting, and root, stem, and fruit rot of susceptible hosts, and economic losses are experienced annually in vegetable crops including cucurbits and peppers. Symptoms of P. capsici infection may manifest as stunting, girdling, or cankers for some cultivars or crops that are less susceptible. P. capsici continues to be a constraint on production, and implementation of an aggressive integrated management scheme can still result in insufficient control when weather is favorable for disease. Management of diseases caused by P. capsici is currently limited by the long-term survival of the pathogen as oospores in the soil, a wide host range, long-distance movement of the pathogen in surface water used for irrigation, the presence of fungicide-resistant pathogen populations, and a lack of commercially acceptable resistant host varieties. P. capsici can infect a wide range of hosts under laboratory and greenhouse conditions including cultivated crops, ornamentals, and native plants belonging to diverse plant families. As our understanding of P. capsici continues to grow, future research should focus on developing novel and effective solutions to manage this pathogen and prevent economic losses due to the diseases it causes.

PEPPER DISEASES IN BALKAN REGION

One of the most important problems affecting pepper production in the Balkan peninsula is the occurrence of common pathogens causing severe diseases and epidemics resulting in reduced and compromised yield. Phytophthora capsici, Botrytis cinerea, Sclerotinia sclerotiorum, Verticillium dahliae, Fusarium spp., Pseudomonas syringae pv. syringae, Xanthomonas vesicatoria, Pepper mild mottle virus (PMMV), Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), Tomato spotted wilt virus (TSWV), Alfalfa mosaic virus (AMV), Potato virus Y (PVY) etc. are among the most devastating and widely distributed pathogens. This region is also characterized with endemic, emerging or newly introduced pathogens threatening pepper production. In the last years new pepper fungal pathogens (Phomopsis capsici and several Colletotrichum spp.) were found in Bulgaria with increasingly frequency. In 2010 a SEE-ERA.NET project started combining the research efforts of seven scientific institutions from five Balkan countries (Albania, Bulgaria, Greece, Macedonia and Serbia). The strategic objectives of the project are: i) to explore Balkan biodiversity of Capsicum spp. in order to extract biotic stress resistant germplasm; ii) to update knowledge about the most economically important and emerging pathogens on Capsicum in the Balkan region and to form pathogen collection; iii) to develop database, concerning the pepper pathogen occurrence in the mentioned regions; iv) to identify areas at differing pathogen risk in the involved Balkan countries and to define risks related to introduction of new pathogen biotypes by trading. Adequate knowledge for pathogens is essential for the management of the diseases, caused by them and for solving problems in sustainable and conventional agriculture. First joint collecting expeditions have been carried out. Pathogens have been isolated, identified and characterized mainly at species level. Collections have been created and properly preserved in related institutes for further investigations concerning the race and strain specificity of the saved pathogens and host-pathogen interaction.

First Report of a Leaf Spot on Pepper Caused by Cladosporium oxysporum in China

Pepper (Capsicum annuum L.) is an important vegetable crop grown in Liaoning Province of China. In June 2009, leaf spotting was observed on hot pepper cv. 37-74 in Wafangdian County of Liaoning Province. By August 2011, the disease had spread to Pulandian and Donggang Counties. Symptoms initially appeared on both sides of leaves as pinpoint chlorotic spots that enlarged and developed into irregular, brown lesions, 1 to 8 mm in diameter. To identify the pathogen, leaf pieces (3 to 5 mm) taken at the edge of lesions with both infected and healthy portions were surface disinfected by placing them in 75% ethanol for 5 s, then transferred to a 0.1% aqueous mercuric chloride solution for 30 s and rinsed with sterilized water three times. The sections were placed on potato dextrose agar (PDA) at 25°C in the dark. Ten pure fungal cultures were obtained from single spores. For growth rate determination and morphological description of colonies, single conidial isolates were inoculated on PDA, malt extract agar (MEA), and oatmeal agar (OA), and incubated at 25°C for 14 days in darkness. For studies of microscopic morphology, isolates were grown on synthetic nutrient agar (SNA) in slide cultures. Colonies grew up to 77 to 80 mm in diameter on PDA, 62 to 66 mm on MEA, and 58 to 60 mm on OA after 14 days. Conidiophores were straight to slightly flexuous. Conidia ranged in shape from subglobose or ovoid to subcylindrical. Macronematous conidiophores measured 40 to 670 × 3 to 5.5 μm with swellings, micronematous conidiophores 15 to 137 × 1.5 to 3.5 μm, terminal conidia 2 to 4 × 2 to 3 μm, and intercalary conidia 4 to 10 × 2.5 to 4 μm. Ramoconidia were rarely observed. Secondary ramoconidia were zero- to one-septate and measured 5.2 to 14.8 × 2.4 to 4 μm. On the basis of these characteristics, the isolates were identified as Cladosporium oxysporum (1). The internal transcribed spacer (ITS) region from isolate W10-02 was amplified using primers ITS1 and ITS4 and sequenced (GenBank Accession No. JQ775499). The 525-bp amplicons had 99% identity to C. oxysporum (GenBank Accession No. EF029816). On the basis of morphological characteristics and nucleotide homology, the isolate was identified as C. oxysporum. Koch's postulates were fulfilled in the laboratory on pepper leaves inoculated with C. oxysporum conidial suspensions (107 conidia ml-1). Eight inoculated 4-week-old seedlings were incubated under moist conditions for 8 to 10 days at 25°C. All leaf spots that developed on inoculated leaves were similar in appearance to those observed on diseased pepper in the field. C. oxysporum was reisolated from lesions and its identity was confirmed by morphological characteristics. C. oxysporum was first reported as a pathogen of pepper in the Sultanate of Oman (2). To our knowledge, this is the first report of C. oxysporum causing leaf spot on pepper in China. The outbreak and spread of this disease may decrease the yield of pepper in the northern regions of China. More studies are needed on the management strategy including the resistance of pepper cultivars against C. oxysporum.

Benzothiadiazole-elicited defense priming and systemic acquired resistance against bacterial and viral pathogens of pepper under field conditions

Like the innate immunity in mammals, plants have developed an induced resistance, referred to as systemic acquired resistance (SAR). Recently, defense priming that is not related to the direct activation of defenses, but instead elicits more rapid induction of resistance mechanisms following trigger application, has been proposed to explain the long-lasting effect of SAR. However, the majority of previous studies have focused on understanding the molecular mechanism underlying priming under in vitro and laboratory conditions. This study examined whether defense priming occurred and was detectable with SAR marker genes by a chemical elicitor, benzothiadiazole (BTH), under field conditions. Pepper seedling application of 0.5 mM BTH was sufficient to prime the CaPR4 gene for 20 days as well as to induce SAR against bacterial spot caused by Xanthomonas axonopodis. Transcriptome analysis revealed to prime defense hormonal signaling and antimicrobial compound production genes. At the end of the season, when bacterial spot and Cucumber mosaic virus disease outbreaks naturally occurred, BTH-treated plants demonstrated less disease symptoms. Our results indicate that the priming of SAR genes plays a critical role in plant protection against pathogens under natural conditions.

병저항성 육종을 위한 고추 유전자원의 저항성 평가

For initiation of resistance breeding program of the red pepper, 21 PR ( resistance) cultivars, 36 cultivars collected from USA and 'Supermanitta' which is a susceptible cultivar against phytophthora blight were assayed against phytophthora blight, powdery mildew, and anthracnose. For seedling assay of phytophthora blight, three different mating type strains of were used (A1, A2, Sterile). The result showed that most of the pepper of PR cultivars were resistance or moderately resistance at each mating type. 'Yeokganghongjanggun' was resistant to all three strains and 'PR-Datta' and 'PR-Manitta' were resistant or moderately resistant at each type. In case of the collected cultivars, 'NuMex J.E.Parker', 'Omni Color', and 'SCM334' were resistant to all the three types and some cultivars including 'Sweet Banana' and 'Tabasco' were moderately resistant to each type fungi. 'Orange Habanero' and 'Black Cuban' were resistant to powdery mildew and 'Supermanitta' and 'PR Keumdong' were moderately resistant, while 'Santa Fe Grande', 'NuMex Pinata' and 'Puya' were very susceptible. In the case of anthracnose, 'Aji Limon' and 'Capsicum baccatum var. pendulum 3-4' were resistant and 'Pobalno', 'Omni Color', 'Negro', 'Mesilla', 'Mulato', 'Bhut Jolokia', 'Big Dipper', 'Black Cuban', 'NuMex Pinata', and 'NuMex Big Jim' were moderately resistant. The most PR cultivars except 'Taesan' were susceptible or very susceptible. These resistant individuals identified through this experiment can be used as sources of resistance to pepper pathogens in the future breeding programs.

&lt;i&gt;In Vitro&lt;/i&gt; Evaluation of Some Fungicides Alternatives Against &lt;i&gt;Fusarium Oxysporum&lt;/i&gt; the Causal of Wilt Disease of Pepper (&lt;i&gt;Capsicum &lt;/i&gt;&lt;i&gt;a&lt;/i&gt;&lt;i&gt;nnum &lt;/i&gt;L.)

The inhibitory effect of the antagonistic bioagents, chemical plant resistance inducers and some essential oils against the linear growth of two isolates of F. oxysporum the wilt pathogen of pepper (Capsicum annum L.) was evaluated in vitro. The antagonistic microorganisms, Trichoderma harzianum, T. viride, T. aureiviride, Bacillus subtilis and Pseudo- monas fluorescens were tested. Also, the tested chemical inducers were Sodium benzoate, Potassium bicarbonate, Potassium sorbate and Chitosan. Meanwhile, the tested essential oils were Cinnamon, Clove, Thyme, Lemon grass, Lemon, Mint, Pepper mint and Mustard. The obtained results indicate that the antagonistic bioagents, T. viride, B. subtilis, P. fluorescens showed superior inhibitory effect against the growth of pathogenic fungi compared with T. harzianum and T. aureiviride. The fungal mycelial growth reduced gradually by increasing of tested concentrations to reach complete reduction (100%) at the concentrations of 4% for Potassium bicarbonate and Sodium benzoate and at 6% for Potassium sorbate. Data also revealed that the fungicide Topsin-M had superior inhibitor effect on the fungal linear growth than that of tested salts. It cause com- plete growth reduction at concentration of 300ppm. Chitosan was found to affect the linear growth of the two isolates of F. oxysporum that the complete reduction in fungal growth (100%) was observed at concentration of 4.5 g/L. Results also showed that Thyme, Lemon grass, Peppermint, Clove and Mint oils had higher inhibitor effect on fungal mycelial growth than Limon, Cinnamon and Mustard oils. Fungal mycelial growth decreased significantly as the concentrations of essential oils were increased, to reach the fungal growth's minimum at the highest concentration used. Complete reduction (100%) in mycelial growth of two fungal isolates was recorded at concentration of 6% of all tested essential oils. The obtained results in the present study showed the possibility of usage antagonistic bioagents, various plant inducers and essential oils to control plant pathogenic fungi.

Distribution and Antifungal Activity of Endophytic Fungi in Different Growth Stages of Chili Pepper (Capsicum annuum L.) in Korea

This study investigated the distribution of endophytic fungi obtained from the surface sterilized tissues of leaves, stems and roots of chili pepper (<TEX>$Capsicum$</TEX> <TEX>$annuum$</TEX> L.) plants in seedling, flowering and fruiting stages in Korea and their antifungal activity. A total of 481 isolates were recovered and were identified using molecular techniques. Based on rDNA ITS gene sequence and phylogenetic analysis, 21 fungal genera were characterized, belonging to 16 Ascomycota and 5 Basidiomycota. <TEX>$Penicillium$</TEX> in seedling stage, <TEX>$Fusarium$</TEX> in flowering stage, <TEX>$Colletotrichum$</TEX> followed by <TEX>$Fusarium$</TEX>, <TEX>$Alternaria$</TEX> and <TEX>$Xylaria$</TEX> in fruiting stage was predominant and <TEX>$Alternaria$</TEX>, <TEX>$Cladosporium$</TEX> and <TEX>$Fusarium$</TEX> were common in all growth stages. Among 481 endophytes 90 phenotypes were evaluated for the antimicrobial activity against three major pathogens (<TEX>$Phytophthora$</TEX> <TEX>$capsici$</TEX>, <TEX>$Colletotrichum$</TEX> <TEX>$acutatum$</TEX> and <TEX>$Fusarium$</TEX> <TEX>$oxysporum$</TEX>) of chili pepper. Among them 16 isolates inhibited the growth of at least one test microorganisms. Three strains showed a broad spectrum antifungal activity and displayed strong inhibition against chili pepper pathogenic fungi.

Avirulence Proteins AvrBs7 from Xanthomonas gardneri and AvrBs1.1 from Xanthomonas euvesicatoria Contribute to a Novel Gene-for-Gene Interaction in Pepper

A novel hypersensitive resistance (HR) in Capsicum baccatum var. pendulum against the bacterial spot of pepper pathogen, Xanthomonas gardneri, was introgressed into C. annuum cv. Early Calwonder (ECW) to create the near-isogenic line designated as ECW-70R. A corresponding avirulence gene avrBs7, in X. gardneri elicited a strong HR in ECW-70R. A homolog of avrBs7, avrBs1.1, was found in X. euvesicatoria 85-10, which showed delayed HR on ECW-70R leaves. Genetic analysis confirmed the presence of a single dominant resistance gene, Bs7, corresponding to the two avr genes. Both AvrBs7 and AvrBs1.1 share a consensus protein tyrosine phosphatase (PTP) active site domain and can dephosphorylate para-nitrophenyl phosphate. Mutation of Cys(265) to Ser in the PTP domain and subsequent loss of enzymatic activity and HR activity indicated the importance of the PTP domain in the recognition of the Avr protein by the Bs7 gene transcripts. Superpositioning of AvrBs7 and AvrBs1.1 homology models indicated variation in the geometry of the loops adjacent to the active sites. These predicted structural differences might be responsible for the differences in HR timing due to differential activation of the resistance gene. Mutating the PTP domain of AvrBs1.1 to match that of AvrBs7 failed to activate HR on ECW-70R, indicating the possibility of differential substrate specificities between AvrBs1.1 and AvrBs7.

Antifungal Bioefficacy of Organic Inputs Against Fungal Pathogens of Bell Pepper

Five organic inputs namely panchgavya, vermiwash, biosol, cow urine and butter milk were evaluated against major pathogens of bell pepper namely Colletotrichum capsici, Phytophthora nicotianae, Sclerotium rolfsii, Fusarium solani, Fusarium oxysporum f.sp. capsici, Sclerotinia sclerotiorum and Rhizoctonia solani under in vitro and in vivo conditions. Under in vitro conditions, all inputs were tested at five concentrations ranging from 2.0 to 10.0 percent. Among these inputs, fermented cow urine showed maximum 100 and 99 percent inhibition in the mycelial growth of the test pathogens at 10 percent concentration followed by panchgavya exhibited complete mycelia inhibition of S. sclerotiorum and 99.0 percent inhibition in mycelia growth of S. rolfsii, F. solani and P. nicotianae. Similarly, vermiwash and biosol showed moe than 99.0 percent mycelia inhibition of all tested pathogens. Whereas, comparatively less inhibition was observed in case of fermented butter milk. Under in vivo conditions, all organic inputs were evaluated individually and in combinations against foliar pathogens and soilborne pathogens of bell pepper separately. Among all treatments, a combination of panchgavya (5%) + fermented cow urine (5%) caused maximum 87.02 percent control of phytophthora blight followed by anthracnose (85.22%) when applied as foliar spray at 15 days interval. Whereas, biosol (5%) + fermented cow urine (5%) showed 83.91 percent disease control of anthracnose. In case of soilborne diseases, drench with panchgavya (5%) + fermented cow urine (5%) exhibited maximum 88.04, 84.98 and 84.25 percent disease control of fusarium wilt, stem rot and root rot respectively.

고추와 토마토 종자에서 종자전염 세균 및 바이러스의 동시 검출을 위한 One-step Multiplex RT-PCR 방법

우리나라에서 주로 재배되는 가지과작물의 종자 전염 병원균 중에서 세균성 병원균 Xanthomonns campestris pv. vesicatoria(Xcv), Clavibacter michiganensis subsp. michiganensis (Cmm), Erwinia carotovora subsp. carotovora (Ecc)와 바이러스 병원균 Pepper mild mottle virus (PMMoV) and Tobacco mild green mosaic virus (TMGMV)를 종자로부터 동시검출하기 위한 One-step Multiplex RT-PCR을 개발하였다. 각각의 병원균을 특이적으로 증폭시키는 병원균 검출용 프라이머 15세트를 primer-blast 프로그램을 이용하여 제작하였고 각각의 병원균을 특이적으로 검출할 수 있는 5세트의 프라이머 세트(Cmm-F/R, Ecc-F/R, Xcv-F/R, PMMoV-F/R, TMGMVF/R)를 선발하였다. 이들 프라이머 세트는 프라이머간의 또는 병원균 cDNA간의 간섭없이 타겟 병원균만을 검출하였다. 가지과 작물 중에서 유통 중인 고추종자 20품종과 토마토종자 10품종에 대한 종자 감염 병원균 검출을 위한 PCR을 수행한 결과, 2개 품종의 토마토종자에서 Ecc가 검출되었고 4개 품종의 고추종자에서도 Ecc가 검출되었다. 특히 고추 종자에선 Ecc가 검출된 4개 품종 외에 1개 품종에서 PMMoV, TMGMV가 동시 검출되었다. 이로 인해 개발된 One-step multiplex RT-PCR은 서로 간섭 현상 없이 병원균을 효율적으로 검출할 수 있음을 보여 주었다. The aim of this study was to develop specific and sensitive PCR-based procedures for simultaneous detection of economically important plant seed infection pathogenic bacteria and virus, Xanthomonns campestris pv. vesicatoria (Xcv), Clavibacter michiganensis subsp. michiganensis (Cmm), Erwinia carotovora subsp. carotovora (Ecc), Pepper mild mottle virus (PMMoV) and Tobacco mild green mosaic virus (TMGMV) in pepper and tomato seeds. Most of pepper and tomato bacterial and virus diseases are responsible for germination and growth obstruction. PCR with arbitral primers: selection of specific primers, performance of PCR with specific primers and determination of the threshold level for pathogens detection. To detect simultaneously the Xcv, Cmm, Ecc, PMMoV and TMGMV in pepper and tomato seeds, five pairs (Cmm-F/R, Ecc-F/R, Xcv-F/R, PMMoV-F/R, TMGMV-F/R) of specific primer were synthesized by primer-blast program. The multiplex PCR for the five pathogens in pepper and tomato seeds could detect specially without interference among primers and/or cDNA of plant seeds and other plant pathogens. The PCR result for pathogen detection using 20 commercial pepper and 10 tomato seed samples, Ecc was detected from 4 pepper and 2 tomato seed samples, PMMoV was detected from 1 pepper seed sample, and PMMoV and TMGMV were simultaneously detected from 1 pepper seed sample.

English

Pepper leaf curl virus (PepLCV) is the most destructive pathogen of pepper and causes substantial economic losses of chilli production worldwide. Curling and puckering of leaves and stunted growth of the plants are typical symptoms of the viral infection. For a reliable detection of PepLCV, coat protein specific primer pairs were designed and after polymerase chain reaction amplification (PCR), -1 kb amplification product specific to coat protein gene was amplified. Cloning of this amplification product in an expression vector resulted in high-level expression of a 28 kD protein. Coat protein gene product is important for vector specificity and is also responsible for viral capsid formation and whiteflymediated transmission. The purified protein can be used for production of coat protein specific antisera leading to development of an efficient detection system for this very important viral pathogen of pepper.

Characterization ofGeminivirusResistance in an Accession ofCapsicum chinenseJacq.

Pepper golden mosaic virus (PepGMV) and Pepper huasteco yellow vein virus (PHYVV), members of the Geminiviridae family, are important pathogens of pepper (Capsicum annuum L.) and other solanaceous crops. Accession BG-3821 of C. chinense Jacq. was reported earlier as resistant to mixed infection with PepGMV and PHYVV. In this work, we characterized the Geminivirus resistance trait present in BG-3821. Segregation analysis suggested that resistance depends on two genes. Our data showed that PepGMV replication in protoplast of resistant plants is approximately 70% lower when compared with the levels observed in protoplasts from susceptible plants. Additionally, viral movement is less efficient in resistant plants. We also evaluated several characteristics commonly associated with systemic acquired resistance (SAR), which is a conserved defensive mechanism. The concentration of salicylic acid was higher in resistant plants inoculated with PepGMV than in susceptible plants. Marker genes for SAR were induced after inoculation with PepGMV in resistant leaves. Similarly, we found a higher accumulation of reactive oxygen species on resistant leaves compared with susceptible ones. A model for the mechanism acting in the Geminivirus resistance detected in BG-3821 is proposed. Finally, the importance of BG-3821 in Geminivirus resistance breeding programs is discussed.

Characterization of Xanthomonas euvesicatoria strains pathogens of pepper in Serbia

During spring and summer of 2008, 101 bacterial strains was isolated from the diseased pepper leaves collected from different pepper growing areas in the Republic of Serbia. The aim of this research was to characterize the isolated strains and determine their taxonomic position according to the most recent nomenclature. Pathogenic, biochemical and physiological characteristics of isolated bacteria were tested using standard bacteriological tests. The pathogen races were determined according to the reaction of differential varieties of Early Calwonder (ECW), their isogenic lines (ECW-10R, ECW-20R, ECW-30R) and Capsicum pubescens. The sensitivity of strains to bactericides was studied in vitro by culturing bacteria on sucrose pepton agar (SPA) plates, amended with filter-sterilized aqueous solution of streptomycin and kasugamycin (50, 100, 200 ppm) or copper-sulphate (100, 200 ppm). Based on pathogenic, biochemical and physiological characteristics, the investigated strains belonged to Xanthomonas euvesicatoria. The reaction of pepper differential varieties indicated that these strains belonged to pepper races P1, P3, P7, P8. Streptomycin resistant strains were not detected, but 6 strains were resistant to kasugamycin (50 ppm) and 13 strains to copper-sulphate (200 ppm), indicating bacterial resistance development.