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백묵병 감염유충으로부터 병원균의 분리 및 형태학적 특성
남성희(Sung Hee Nam),이명렬(Myeong Lyeol Lee),이만영(Man Young Lee),김영수(Young Soo Kim),이병일(Ri Bing Li),장승종(Seung Jong Chang),윤병수(Byoung Su Yoon) 한국양봉학회 2004 韓國養蜂學會誌 Vol.19 No.1
Chalkbrood-infected larvae of honeybee were obtained from 8 apiaries in Korea. Total 548 samples varied in the rate of black and white mummies of honey bee. Among them, 306 black mummies were examined to identify species. The whole larvae were proved to be infected with Ascosphaera apis(Maasen ex Claussen) Olive & Spiltoir. In the early stage of chalkbrood diseases, the white/grey mass was formed on the surface of larvae. The larvae were soon shrunken to mummy, getting hard, and forming hard mummies. In the late stage of chalkbrood diseases, the mycelium grew densely and covered the larvae to the extent that it filled the whole cell. When the larvae were infected with one sexual type of A. apis, they became white mummies while the grey-black mummies were developea by the invasion of both sexual types. A. apis (Aaj623) was heterothallic and mostly grown 57.6 ㎜ in diameter in 7 days on potato dextrose agar(PDA). Spore cysts were globose, nearly smooth on outer surface and evenly verrucate on inner surface, 45~95 ㎛ in diameter(average 74 ㎛ diameter). Spore balls were globose, 9~17 ㎛ in diameter(average 12.9 ㎛ in diameter) and lacking a conspicuous granular coating. Ascospores were hyaline, ellipsoid and smooth, 1 cell, 2.3~3.0 × 1.0~1.5 ㎛(average 2.49 × 1.35 ㎛ in diameter).
Bo Xu,Yingzhe WANG,Shixin ZHU,Haizhu ZHOU,Changlong GOU,Wenlong DONG,Yu Wang,Yunhang GAO,Hongxia MA 한국곤충학회 2019 Entomological Research Vol.49 No.1
Chalkbrood, which results from Ascosphaera apis infection, is one of the bee diseases that causes serious damage to the bee colony. Understanding the molecular bases underlying immune response to chalkbrood disease would facilitate the genetic breeding of bees by selecting races with superior chalkbrood resistance. In this study, transcriptome sequencing was performed to identify genes and pathways involved in the immune response to As. apis infection in A. mellifera larvae. In total, 2,890 differentially expressed genes (DEGs) (FDR < 0.001) were identified between the healthy and As. apis infected bee larvae, including 2,214 up‐regulated and 676 down‐regulated unigenes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway classification of the DEGs revealed association with development, energy metabolism, signal transduction, disease, and immune response. Among the immune‐related genes, p38, toll‐like receptors (TLRs), integrin, and antimicrobial peptides were up‐regulated under As. apis infection. This study provides valuable gene transcriptional information towards the investigation of molecular mechanisms related to chalkbrood immune response and host pathogenesis in A. mellifera.
Control of Chalkbrood by using Actinobacterial Culture
Hojae Lee,Jaisoo Kim 한국양봉학회 2014 韓國養蜂學會誌 Vol.29 No.1
Chalkbrood is a major blight damage to bees, caused by a fungus Ascosphaera apis (A. apis) through infection to honeybee larvae. To control chalkbrood disease of honey bees, this study tested many actinobacterial isolates to find the antifungal activity against A. apis. As a result, we found 7 Streptomyces spp. Those are S. ferralitis, S. resistomycificus, S. sporoverrucosus, S. caniferus, S. racemochromogenes, S. anandii, and S. blastmyceticus. Among them, S. sporoverrucosus, S. racemochromogenes, and S. anandii showed the most effective antifungal activities with the diameters of inhibition zone, around 20 mm.
Real-Time PCR을 이용한 백묵병 원인균 Ascosphera apis의 신속 검출
이혜민(Hye-Min Lee),이도부(Do-Bu Lee),한상훈(Sang-Hoon Han),남성희(Seoung-Hee Nam),임윤규(Yoon-Kyu Lim),윤병수(Byoung-Su Yoon) 한국양봉학회 2005 韓國養蜂學會誌 Vol.20 No.2
Chalkbrood is a well known fungal disease of honeybees, that was caused by a fungus, Ascosphaera apis. Because there is no available control method for disease, and because co-infection with other pathogens could be induced a rapid detection system will be important in a hive management. In this study, Real-time PCR method for the rapid detection of chalkbrood was developed. A specific primer-pair was designed based on 185 rRNA gene of Ascosphaera apis. This PCR could easily detect the existence of the pathogen from larvae samples. In addition, the minimum concentration of template DNA was about 6 copies. From this result, shortening the detection time in Chalkbrood disease could induce a strong advance in this field.
항진균제 ALBIO K와 Pure Wild Organo Oil을 사용한 꿀벌 백묵병의 예방
이혜민(Hye-Min Lee),하정순(Jung-Soon Ha),조용호(Yong-Ho Jo),김동수(Dong-Soo Kim),윤병수(Byoung-Su Yoon) 한국양봉학회 2005 韓國養蜂學會誌 Vol.20 No.1
Chalkbrood is a highly contagious disease of the honeybee apis mellifera caused by the heterothallic fungus Ascosphaera apis. We tested the anti-fungal activity of ALBIO K and Pure Wild Organa Oil that were extracted from natural source against honeybee's Chalkbrood disease. As the results of using different experiments, it was found that the growth of A. apis was definitely inhibited by ALBIO K or Pure Wild Organo Oil in the early stage of germination. Especially, the anti-fungal effects of ALBIO K and Pure Wild Organo Oil were estimated in the disc-diffusion experiments via liquid or via air. Both ways of drug delivery will be offer more useful usage of these agents in the point of prevention against Chalkbrood and in the point of food safety.
남성희(Sung-Hee Nam),최지영(Ji Young Choi),이명렬(Myeong Lyeol Lee),홍인표(In Pyo Hong),성규병(Gyoo Byung Sung),이광길(Kwang Gill Lee),여주홍(Joo Hong Yeo) 한국양봉학회 2009 韓國養蜂學會誌 Vol.24 No.2
To investigate the factors of occurrence of A. apis, Chalkbrood disease was investigated at two farms in Chungbuk province. The lowest temperature within the hives was 32℃, and the highest was 37℃ from May to September. The humidity was 74~86% and it wasn’t shown significant difference compared with healthy hives. Ascosphaera apis grow faster under the 25℃ and was extinct completely at 40℃. Ascomata and ascospores developed early when it was cultured under the 10% CO₂ incubator. To select compounds for prevent chalkbrood disease, Ascosphaera apis was treated with 8 chemicals which was Ceder oil, Thymol, Oxalic acid dihydrate, Dodycine, Sodium hypochlorite, Ethanol, Anti-fungi paint, Morpholinium compound terpinen-4-ol. The honey bees were treated with 8 chemicals and avoidanced odor extremely on Sodium hypochlorite and Oxalic acid and wasn’t effected with Dodycine and Ethanol. Prevent effect of dodycine and sodium hypochorite were 98.5% and 100% on chalkbrood pathogen and they didn’t harm honey bees. As these results, Dodycine and Sodium hypochorite were proper disinfection chemical for hive and hive tools respectively.
백묵병 원인균 Ascosphaera apis의 Quick Real-time PCR 검출법
이혜민(Hye-Min Lee),유미선(Mi-Sun Yoo),김을환(Eul-Hwan Kim),이동우(Dong-Woo Lee),한상훈(Sang-Hoon Han),윤병수(Byoung-Su Yoon) 한국양봉학회 2006 韓國養蜂學會誌 Vol.21 No.2
Chalkbrood of honeybees is caused by a species of fungus, Ascosphaera apis, which could affect larvae of honeybee. For the rapid detection of Ascosphaera apis from suspected samples, a type of specific real-time PCR was developed in this study. In this PCR-identificaion, the microchip-based special tube and apparatus, GenSpector™, were used with two A. apis specific primers named QP-A. apis F and QP-A. apis R, were designed on the basis of the 18S rRNA gene. Real-Time PCR for the identification of pathogen was completed within 17 minutes including PCR-amplication and analysis of melting temperature. This type of extremely rapid PCR was designated as Quick Real-Time PCR in this study. For the detection and control of Chalkbrood in apiculture, this type of quick detection would be expected to use especially in field.
꿀벌 진균성 질병의 신속 확인을 위한 Ascosphera apis, Aspergillus flavus의 PCR 검출법
이혜민(Hye-Min Lee),하정순(Jung-Soon Ha),조용호(Yong-Ho Jo),남성희(Sung-Hee Nam),윤병수(Byoung-Su Yoon) 한국양봉학회 2004 韓國養蜂學會誌 Vol.19 No.2
Chalkbrood and Stonebrood are well known fungal diseases of honeybee, those are caused by the fungus Ascosphaera apis and by several species of Aspergillus, respectively. Because there is no available control method for these disease, and because co-infection with other pathogens should be escaped, the rapid detection system will be important in hive management. In this study, specific PCR method for the rapid detection of chalkbrood and stonebrood was developed. Specific primer-pairs were designed based on 18S rRNA gene of Ascosphaera apts and Aspergillus flavus. This PCR system could easy detect the existence of these pathogens from larvae samples. In addition, new universal primers was designed for amplification of 18S rRNA gene from various fungi. Using this universal PCR, un-identified strains of fungi will be determined its characterization based on 18S rRNA gene.
Multiplex-PCR을 이용한 꿀벌 노제마병, 백묵병, 미국부저병의 동시 검출
최지영(Ji-young Choi),이명렬(Myeong-lyeol Lee),남성희(Sung-hee Nam),김종길(Jong-gill Kim),최영철(Young-cheol Choi),김원태(Won-tae Kim),심하식(Ha-Sik Sim),김근영(Keun-young Kim) 한국양봉학회 2006 韓國養蜂學會誌 Vol.21 No.2
A multiplex polymerase chain reaction (multiplex PCR) was developed for the simultaneous detection and differentiation among Nosema ceranae, Ascosphaera apis and Paenibacillus larvae in honeybee. Nosema disease(N. ceranae), Chalkbrood disease(A. apis) and American Foulbrood disease(P. larvae) are highly contagious diseases in Apis mellifera and are causing considerable economic loss to beekeepers in Korean. Nosema disease is widespread and can cause extensive losses of adult bees and may also be responsible for some supersedure of queens. Chalkbrood disease is caused by Ascosphaera apis, entomopathonic fungi occurring only honeybee larval stage. American foulbrood disease, caused by the spore-forming bacterium Paenibacillus larvae, is most serious and fatal bacterial disease. Three sets of primers were selected from different genomic sequences to specifically amplify a 230 bp amplicon, specific for Nosema ceranae (NS primer); a 500 bp amplicon, specific for Ascosphaera apis (CBP primer); and a 971 bp amplicon, specific for Paenibacillus larvae (PL primer). Using the primers in conjunction (multiplex PCR) we were able to detect N. ceranae, A. apis and P. larvae among them. We proposed that the multiplex PCR was sensitive, specific and rapid tool that can serve as a useful differential diagnostic tool for detecting N. ceranae, A. apis and P. larvae in honeybee.
꿀벌의 석고병, 백묵병 현장 진단을 위한 신속 Ascosphaera apis 및 Aspergillus flavus 검출법 개발
왕지희(Ji-hee Wang),민상현(Sang-hyun Min),임수진(Su-jin Lim),윤병수(Byoung-Su Yoon) 한국양봉학회 2016 韓國養蜂學會誌 Vol.31 No.1
Rapid detection methods against Ascosphaera apis and Aspergillus flavus were developed for Chalkbrood and Stonebrood in honeybee. Specific primer sets were selected which detect A. flavus or A. apis based on 18s ribosomal RNA independently and were evaluated using real-time PCR and Ultra-fast PCR (UF-PCR). As the result, Minimal detection time was reduced until 8 min 38 seconds for A. flavus and 12 min 57 seconds for A. apis under 30 cycles UF-PCR. The detections of these methods show that even 101 (100 atto gram) molecules of DNA template could be detected for Aspergillus flavus and that 103 (10 femto gram) molecules of DNA template could be detected for Ascosphaera apis. Specific UF-PCRs are not only useful to confirm pathogendetection with naked eye, but be easy to handle a mobile PCR instrument in field. To monitoring of Chalkbrood or Stonebrood in apiary field directly, applications of on-site pathogen detection would be expected.