• Title/Summary/Keyword: Inoculation methods

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A Simple Method for the Assessment of Fusarium Head Blight Resistance in Korean Wheat Seedlings Inoculated with Fusarium graminearum

  • Shin, Sanghyun;Kim, Kyeong-Hoon;Kang, Chon-Sik;Cho, Kwang-Min;Park, Chul Soo;Okagaki, Ron;Park, Jong-Chul
    • The Plant Pathology Journal
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    • v.30 no.1
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    • pp.25-32
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    • 2014
  • Fusarium head blight (FHB; scab) caused mainly by Fusarium graminearum is a devastating disease of wheat and barley around the world. FHB causes yield reductions and contamination of grain with trichothecene mycotoxins such as deoxynivalenol (DON) which are a major health concern for humans and animals. The objective of this research was to develop an easy seed or seedling inoculation assay, and to compare these assays with whole plant resistance of twenty-nine Korean winter wheat cultivars to FHB. The clip-dipping assay consists of cutting off the coleoptiles apex, dipping the coleoptiles apex in conidial suspension, covering in plastic bag for 3 days, and measuring the lengths of lesions 7 days after inoculation. There were significant cultivar differences after inoculation with F. graminearum in seedling relative to the controls. Correlation coefficients between the lesion lengths of clip-dipping inoculation and FHB Type II resistance from adult plants were significant (r=0.45; P<0.05). Results from two other seedling inoculation methods, spraying and pin-point inoculation, were not correlated with adult FHB resistance. Single linear correlation was not significant between seed germination assays (soaking and soak-dry) and FHB resistance (Type I and Type II), respectively. These results showed that clip-dipping inoculation method using F. graminearum may offer a real possibility of simple, rapid, and reliable for the early screening of FHB resistance in wheat.

EVALUATION OF DISEASE RESISTANCE AND SUSCEPTIBILITY TO CHESTNUT BLIGHT FUNGUS, CRYPHONECTRIA PARASITTCA, OF CHESTNUT VARIETIES IN KOREA

  • Lee, Sang-Hyun;Hwang, Myung-Soo;Kim, Kyung-Hee;Lee, Jong-Kyu
    • Proceedings of the Korean Society of Plant Pathology Conference
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    • 2003.10a
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    • pp.69.2-70
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    • 2003
  • For the selection and breeding of chestnut varieties resistant to the chestnut blight fungus Cryphonectria parasitica, disease resistance and susceptibility of 28 varieties widely planted and growing in Korea were evaluated by artificial inoculation of a pathogenic fungus. For this experiment, a typical virulent strain (KCPC-19) was selected. Artificial inoculation was conducted into all varieties by using two different materials and methods, i.e., bark and wood tissue sections in the laboratory and living trees in the field. In the bark and wood tissue section method, the size of necrotic area and canker development on chestnut varieties were examined and compared 4 days after inoculation. There were wide variations of chestnut varieties in disease resistance and susceptibility against chestnut blight fungus, but 3 varieties, Daebo!, Ishizuchi, and Sandae, were shown to be relatively resistant to the disease with the necrotic area of 0.95-1.03 cm2, while Arima was the most susceptible with the size of 2.0 cm2. In the living tree inoculation examined 5 weeks after inoculation, 3 varieties, Daebo, Ishizuchi, and Riheiguri, showed the higher resistance, but Tono 2 did the highest susceptibility among tested varieties.

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Production of the Wild Entomopathogenic Fungi, Cordyceps militaris, in the Silkworm, Bombyx mori

  • Lee, Sang-Mong;Park, Nam-Sook;Cho, Sae-Yun;Hwang, Jae-Sam;Jin, Byung-Rae
    • International Journal of Industrial Entomology and Biomaterials
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    • v.3 no.1
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    • pp.105-108
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    • 2001
  • The wild entomopathogenic fungi, Cordyceps militaris, were collected at the Whawang mountain, Korea. The pupae of the silkworm, Bombyx mori, were used as infecting hosts for the production of the silkworm-mili-taris dongchunhacho, silkworm vegetable wasps and plant worms with C. militaris. Three inoculation methods in terms of injection, spray and immersion were tested against the silkworm pupae. The three inocu1ation methods revealed 100% infectivity to the silkworm pupae tested. Of the three inoculation methods, the injection method was highly effective in the reduction of the period required for the endosclerotium and the completion of fruiting body formation. These results indicate that the silkworm pupae are very effective host insects for the production of C. militaris.

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A novel method for high-frequency transgenic shoot regeneration via Agrobacterium tumefaciens in flax (Linum usitatissimum L.)

  • Beyaz, Ramazan;Darcin, E. Selcen;Aycan, Murat;Kayan, Mustafa;Yildiz, Mustafa
    • Journal of Plant Biotechnology
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    • v.43 no.2
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    • pp.240-247
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    • 2016
  • In this study, routinely used transformation method, which includes transferring explants onto co-cultivation medium after inoculating them with bacterial solution for a while, was compared with 3 different inoculation methods. In every 3 methods, hypocotyl explants excised from 7-day-old sterile flax seedlings having cotyledon leaves and no root system dried under air flow in sterile cabin for 35 min were inoculated with different volumes of bacterial solution at different inoculation periods. GV2260 line of Agrobacterium tumefaciens having 'pBIN 19' plasmid containing npt II (neomycin phosphotransferase II) gene and GUS reporter gene was used in transformation studies. After inoculation, hypocotyl segments of seedlings (0.5 cm in length) - were excised and left to co-cultivation for 2 days. Then, explants were transferred to regeneration medium supplemented with different antibiotics. The presence of npt-II and GUS genes in transformants was confirmed by PCR and GUS analysis. The highest results in all characters examined in all cultivars were obtained from the 2 inoculation method in which hypocotyls excised from seedlings inoculated with $500{\mu}l$ of bacterial solution after drying in sterile cabin for 35 min were used.

Artificial cultivation of Medicinal Mushroom, Phellinus linteus using Mulberry logs

  • Hong, In-Pyo;Sung, Gyoo-Byung;Chung, In-Mo;Lee, Won-Chu
    • Journal of Sericultural and Entomological Science
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    • v.44 no.2
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    • pp.74-81
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    • 2002
  • The optimal conditions for mycelial growth of P. linteus ASI 26011 were 25-30$^{\circ}C$ and pH 6.0, respectively. The mycelial growth of P. linteus was excellent on MCM medium. In case of carbon sources, the mycelial growth of P. linteus was best on the culture media that were contained with sucrose, mannose and glucose. Potassium nitrate and sodium nitrate were good for the mycelial growth of P. linteus as a nitrogen source. For comparison of the mycelial colonization of P. linteus on logs, several techniques of inoculation were tested; the sterilized short log inoculation, drilling inoculation and log-end sandwich inoculation. The mycelial colonization of P. linteus on logs was good in the treatment of sterilized short log inoculation, but poor in the traditional methods such as drilling inoculation and log-end sandwich. The initial mycelial growth and the full mycelial colonization of P. linteus were the best on 20 cm logs under the condition of 42% of moisture content in log. Also the initial mycelial growth of P. linteus was accelerated over 12 hours of sterilization. Burying method of logs after 5-6 months of incubation was the best for formation of basidiocarp of P. linteus. The formation of fruiting body of P. linteus was quite good in the cultivation house at the 31-35$^{\circ}C$ and over 96% of relative humidity.

Development of an Efficient Simple Mass-Screening Method for Resistant Melon to Fusarium oxysporum f. sp. melonis (덩굴쪼김병 저항성 멜론을 위한 효율적이고 간편한 대량 검정법 개발)

  • Lee, Won Jeong;Jang, Kyoung Soo;Choi, Yong Ho;Kim, Heung Tae;Kim, Jin-Cheol;Choi, Gyung Ja
    • Research in Plant Disease
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    • v.21 no.3
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    • pp.201-207
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    • 2015
  • This study was conducted to establish a simple mass-screening method for resistant melon to Fusarium wilt caused by Fusarium oxysporum f. sp. melonis (FOM). Root-dipping inoculation method has been used to investigate resistance of melon plants to Fusarium wilt. However, the inoculation method requires a lot of labor and time because of complicate procedure. To develop a simple screening method on melon Fusarium wilt, occurrence of Fusarium wilt on susceptible and resistant cultivars of melon according to inoculation method including root-dipping, soil-drenching, tip, and scalpel methods was investigated. Scalpel and tip methods showed more clear resistant and susceptible responses in the melon cultivars than root-dipping inoculation method, but tip method represented slightly variable disease severity. In contrast, in the case of soil-drenching inoculation method, disease severity of the susceptible cultivars was very low. Thus we selected scalpel method as inoculation method of a simple screening method for melon Fusarium wilt. By using the scalpel inoculation method, resistance degrees of the cultivars according to incubation temperature after inoculation (25 and $30^{\circ}C$) and inoculum concentration ($1{\times}10^6$ and $1{\times}10^7conidia/ml$) were measured. The resistance or susceptibility of the cultivars was hardly affected by all the tested conditions. To look into the effectiveness of scalpel inoculation methods, resistance of 22 commercial melon cultivars to FOM was compare with root-dipping inoculation method. When the melon cultivars were inoculated by scalpel method, resistance responses of all the tested cultivars were clearly distinguished as by root-dipping method. Taken together, we suggest that an efficient simple mass-screening method for resistant melon plant to Fusarium wilt is to sow the seeds of melon in a pot (70 ml of soil) and to grow the seedlings in a greenhouse ($25{\pm}5^{\circ}C$) for 7 days, to cut the root of seedlings with a scalpel and then pour a 10 ml-aliquot of the spore suspension of $1{\times}10^6conidia/ml$ on soil. The infected plants were cultivated in a growth room at 25 to $30^{\circ}C$ for about 3 weeks with 12-hr light a day.

Comparison of Three Different Methods for Campylobacter Isolation from Porcine Intestines

  • Shin, Eun-Ju;Lee, Yeon-Hee
    • Journal of Microbiology and Biotechnology
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    • v.19 no.7
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    • pp.647-650
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    • 2009
  • Using 200 porcine colon tissues, the efficiencies of three isolation methods of Campylobacter from porcine intestines were compared: Method 1, direct streaking of colon mucosa; Method 2, direct inoculation of intestinal contents with a swab; Method 3, inoculation of pre-enriched medium. A total of 460 Campylobacter isolates were obtained from 178 samples (89%) by direct streaking of colon mucosa, 142 samples (71%) by direct streaking of a swab, and 94 samples (47%) by pre-enrichment of intestinal contents in Preston broth. Direct streaking of colon mucosa was superior to the other two isolation methods, in terms of rapidity and higher efficiency. When isolates were identified with various biochemical tests and PCRs specific to 16s rRNA, mapA, and ceuE, C. coli was the predominant species (87%) in porcine, whereas the rest of the isolates were identified as C. lanienae.

An Alternative Method to Evaluate Resistance to Pear Scab (Venturia nashicola)

  • Kyungho Won;Eu Ddeum Choi;Keumsun Kim;Hae Won Jung;Il Sheob Shin;Seongsig Hong;Cecile Segonzac;Young Jin Kim
    • The Plant Pathology Journal
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    • v.39 no.2
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    • pp.228-233
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    • 2023
  • Two pear cultivars with different degrees of resistance to Venturia nashicola were evaluated on the basis of a disease severity rating for pear scab resistance under controlled environmental condition. Two inoculation techniques were tested: the procedure for inoculation by dropping conidia suspension of V. nashicola; the procedure by deposition of agar plug on the abaxial surface of pear leaves. All tested cultivars resulted in blight symptoms on the inoculated leaves and became spread to uninoculated region or other leaves. Although both methods provide satisfactory infection of V. nashicola on pear leaves, the mycelial plug method of inoculation was more reliable than the spray inoculation method for the evaluation of pear scab disease resistance. The incubation period of V. nashicola in the resistant pear cultivar, Greensis was longer than that in the susceptible cultivar, Hwasan.

The Effects of the Tumor Mass Size Inoculated in Immunologically Competent Balb/c Mice on Delayed-type Hypersensitivity Response (정상 면역 생쥐에 접종된 암세포주의 종괴 형성이 숙주의 지연성과민반응에 미치는 영향)

  • Lim, Hyun-Ja;Woo, A-Mi;Jung, Young-Ju;Kang, Jae-Seung;Shin, Dong-Hoon;Lee, Wang-Jae;Hwang, Young-Il
    • IMMUNE NETWORK
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    • v.6 no.4
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    • pp.185-191
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    • 2006
  • Background: Based on outstanding progresses in animal experiments, vaccines for some human tumors have been developed. However, clinical effects of these vaccines have been far below than expected. This discrepancy might come from differences between animal models and human patients with respect to immunocompetency. The immune status of mice after tumor inoculation has not been well studied, which make us cautious in interpreting and applying the results from mice to human. We evaluated cell-mediated immune responses in mice after tumor cell inoculation. Methods: Mice were inoculated with TA3Ha, CT26, or 4T1. Delayed-type hypersensitivity (DTH) responses were induced 2-4 weeks after inoculation using 2,4-dinitro-1-fluorobenzene as an antigen. The relationships between the severity of DTH responses and the duration of tumor inoculation or the size of tumor mass were analyzed. Results: In T A3Ha groups, DTH response was elevated 2 weeks after inoculation, but depressed after 4 weeks, compared to the control group. When analyzed based on the sizes of tumor masses elicited, DTH responses were inversely related to the mass size, especially in those greater than 10 mm in diameter. In CT26 groups, while the duration after inoculation did not affect the severity of DTH responses, those with large mass showed depressed responses regardless the duration of inoculation. 4T1 cells grew so slowly that the size of tumor mass was small even 4 weeks after inoculation, and this group showed much higher DTH responses compared to that of tumor-free group. Conclusion: At least in an experimental setting where tumor model was induced by inoculating tumor cell lines into immunologically competent mice, the host immune response was elevated in early stage, and then depressed in late stage when the mass grew over a critical size.

Screening Procedure of Tobacco Cultivars for Resistant to Bacterial Wilt Caused by Ralstonia solanacearum (담배세균성마름병[립고병(立枯病)]에 대한 담배품종의 저항성 검정법)

  • Jeon, Yong-Ho;Kang, Yue-Gyu
    • Journal of the Korean Society of Tobacco Science
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    • v.30 no.1
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    • pp.1-7
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    • 2008
  • Bacterial wilt caused by Ralstonia solanacearum has become a severe problem on tobacco in Korea. No effective single control measure is available at present time. One of the most potential way for controlling the bacterial wilt on tobacco is growing tobacco cultivars resistant to the bacterial wilt. In this study, optimal conditions for screening tobacco cultivars resistant to the bacterial wilt were examined to provide reproducible and efficient methods in growth chamber testing and field experiments for evaluating plant disease resistance. For this, already-known inoculation methods, inoculum densities, and incubation temperature, and plant growth stages at the time of inoculation were compared using tobacco cultivars resistant (Nicotiana tabacum cv, NC95), moderately resistant (N. tabacum cv. SPG70), and susceptible (N. tabacum BY4) to the bacterial disease. It was determined that root-dipping of tobacco seedlings at six true leaf stage into the bacterial suspension with inoculum level of $10^8$ colony-forming units (CFU)/ml for 20 min before transplanting was simple and most efficient in testing for resistance to the bacterial wilt of tobacco caused by R. solanacearum, for which disease incidences and severities were examined at 2 weeks of plant growth after inoculation at $20{\sim}25^{\circ}C$ in a growth chamber. These experimental conditions could discriminate one tobacco cultivar from the others by disease severity better than any other experimental conditions. In field testing, the optimum time for examining the disease occurrence was late June through early July. These results can be applied to establishing a technical manual for the screening of resistant tobacco cultivars against the bacterial wilt caused by R. solanacearum.