미포장충류(Nosema spp. (NS))는 양봉꿀벌에 심각한 문제를 야기시키는 기생충으로 효과적인 방제물질의 선발이 무엇보다 중요하다. 본 연구는 노제마병과 기타 꿀벌의 발생유행시기의 구명과 더불어 3가지 노제마병 방제물질(M1 = 벌꿀희석의 레몬쥬스; M2 = 설탕시럽 혼합의 카모마일 추출물; M3 = 설탕시럽 혼합의 항생물질 스트리베트)을 평가하고자 수행하였다. 꿀벌 성충과 유충집단의 질병 유행시기를 년간 조사하였으며, 야외 및 실험실 조건에서 노제마병에 대한 M1, M2, M3의 효과를 평가하였다. 조사결과 극소수의 꿀벌 성충과 유충 질병이 발견되었다. 노제마 병은 겨울과 봄 기간 저온과 고습조건에서 검출되었다. 포장실험에서 M2는 36.66%까지 발병억제 능력을 보였으며, 반면M3는 23.33%, M1는 13.33%의 억제효과를 보였다. 실내실험에서 M2가 방제효과가 가장 좋았고, 그 다음 M1와 M3 이었다. 3가지 방제물질은 병에 감염된 꿀벌성충의 생존력을 크게 높이는 것으로 나타났다. 본 연구는 노제마병 방제를 위한 천연물질로 카모마일의 잠재적 방제효과를 제시하고 있다.

Nosema disease caused by the microsporidia Nosema apis and Nosema ceranae are a honey bee pathogen parasitizing. Nosema disease symptoms include digestive and absorption disorders because the spores damage epithelial tissue and potentially causing colony death. Recently, N. ceranae has been reported as an important threat to honey bee health. Turmeric (Curcuma longa L.) Curcuma tonga L. belongs to the family Zingiberaceae and is a perennial, tropical herb. Turmeric, the powdered rhizome, is used for medicinal purposes. The aim of this study was to evaluate the potential of Turmeric (Curcuma longa L.) for the control of N. ceranae in honeybees. For the study, we infected with N. ceranae spore through dosed and fed with the turmeric extraction at difference concentration. The data show that the turmeric extraction was not toxic for bee at least at 1% and the bees fed with 0.5 % turmeric extraction had significantly lower infection rates. This data suggest that turmeric could be useful in alternative strategies for the control of N. ceranae.

The population of managed honeybees has been dramatically declining the recent past in worldwide. N. ceranae causessignificant detriment to honey production and results in economic losses critically. In our knowledge, Fumagillin is theonly antibiotic approved for control of nosemosis in honeybees. In this study, to select isolate with anti-Nosema activityagainst N. ceranae. Entomopathogenic fungi cultural filtrates were screened using in vitro polar tube germination assay.These fungi cultural filtrates were used to evaluate the safety of honeybees and their inhibition of nosemosis. As a result,P. marquandii 364 and Pochonia sp. 60 showed inhibitory activity on the growth of N. ceranae in honeybees and didnot significantly affect the survival rate of honeybees. These may be employed as antibiotic agents and a good featureto be used in the development of new biocontrol agents of nosemosis.

Honeybees have faced many diseases which treaten bee colony including a serious population decline phenomenon called Colony Collapse Disorder (CCD). Nosema ceranae is a pathogen cause nosemosis, which is now wide spread around the world. According to the genome sequencing for N. ceranae, it has been identified that the presence of machinery for RNA silencing. Microsporidia N. ceranae that are obligated intracellular parasites depend on their host for energetic and metabolic needs. Here we selected the several genes from mitosome of N. ceranae to develop RNAi for the control of Nosema. Especially, TOM40, FNR1, FNR2 and Nar1 were chosen. After infection of N. ceranae, the Honeybee were treated with RNAi either by using only one or combining two or more. The infection rate and specific gene silecing in Nosema were analyzed.

The population of managed honey bees has been dramatically declining the recent past in worldwide. The one of most common disease of bees is nosemosis, the nosemosis is caused by microsporidia in the genus Nosema. Nosema apis and N. ceranae have been described as honeybee pathogens. These microsporidia are highly evoloved fungi with an obligately intracellular parasitic lifstyle. The disease causes significant detriment to honey production and results in economic losses. In our knowledge, Fumagillin is the only antibiotic approved for control of nosemosis in honey bees, however this antibiotic may have unintended effects on the honey bee host, ultimately contributing to increased prevalence and pathogenicity of Nosema. Therefore, we screened anti-Nosema substances from entomopathogenic fungal culture filtrates using in vitro polar tube germination assay. These fungal metabolites are employed as antibiotic agents. As results, Total 3 samples (23% of 13 total samples) showing the germinating inhibition against N. ceranae. This screening method may be useful for the detection of anti-Nosema substances from various samples and selected samples in this study may be a good feature to be used in the development of a new biocontrol method of nosemosis.

A Duplex polymerase chain reaction (PCR) was developed for the simultaneous detection and differentiation among Nosema spp. and Vairimorpha spp. from Lepidoptera insects. Two sets of primers were selected from different genomic sequences to specifically amplify an 831 bp amplicon within the SSU rRNA gene, specific for both Nosema spp. and Vairimorpha spp. (MSSR primer); a 542 bp amplicon within the SSU rRNA gene, specific for Vairimorpha spp. (VSSU primer). Using the primers in conjunction (duplex PCR) it was possible to detect Nosema spp. and Vairimorpha spp. to differentiate between them. The sensitivity of this PCR assay was approximately 10 spores per milliliter. It is proposed that the duplex PCR is a sensitive, specific and rapid tool that can serve as a useful differential diagnostic tool for detecting Nosema spp. and Vairimorpha spp. in Lepidoptera insect.

Nosema disease caused by microsporidia which belongs to fungi is a major cause of honey production loss and is worldwide in distribution. The most common method of nosema disease controlling is the application of the fumagillin. It is an antibiotic derived from the fungus Aspergillus fumigates. It has been used for many years to treat microsporidiosis caused by Nosema in honeybees. However, Fumagillin does not kill Nosema spores, and has rapidly deteriorating potency after application, resulting in only partial and temporary anti-Nosema effect, since new bees emerge constantly in a colony, and re-application is required several times a year. Also it is no longer practised for the associated risk of honey contamination. In this study we compare the effect in spore reduction of Cordyceps militaris extract(CMX) and of fumagillin. Three weekly administrations of CMX showed a noticeable decrease in the number of spores infecting the honey bee intestines, without negative effects for adults and comb health. Therefore this product could play an important role in prevention and reduction of nosema disease without risks of honey with antibiotics.

The purpose of this study was to identify fast and effectively the Nosema disease of bumble bees from Gangwon province in Korea. Bumble bees are crucial pollinators of various crops and microsporidia are the critical infections of these hosts. The origin of the bumble bees is probably being known from the Britain. Also, the following species of bumble bees have been used around the world: Bombus terrestris, Bombus lucorum, Bombus occidentalis, Bombus ignites and Bombus impatiens. When bumble bees are infected with N. bombi, their abdomens can become distended, paralyzed and infected workers and they often become sluggish and die early. We have identified the morphology of the microsporidium by light and electron microscopy and have found that the morphology of the microsporidium is rounded spore morph, with fairly small spores as described before in many other articles. For the specific and sensitive diagnosis of the microsporidian parasite N. bombi in bumble bees, we have developed the improved method of polymerase chain reaction (PCR) conditions for expeditious diagnosis. Two pairs of primers were tested on N. bombi and the related microsporidia Nosema apis and Nosema sp., both of which infect bumble bees and honey bees and further we have verified and analyzed DNA sequence data of N. bombi in bumble bees by using the BLAST server at the National Center for Biotechnology Information.

The bumblebee, Bombus terrestris, has played an important role as one of the alternative pollinators since the outbreak of honeybee collapse disorder. Recently, pathogens and parasites such as viruses, bacteria and mites, which affect the life span and fecundity of their host, have been discovered in B. terristris. In order to detect the microsporidian pathogen, Nosema Spp. in the field populations of B. terristris, we collected adults and isolated their genomic DNA for diagnostic PCR. The PCR primers specific for Nosema Spp. were newly designed and applied to gene amplification for cloning. Only small subunit ribosomal RNA(SSU rRNA) gene of N. ceranae was successfully amplified and sequenced among examined genes, which indicates that N. ceranae mainly infects the examined field population of B. terristris. To detect of SSU rRNA gene, two regions of SSU rRNA gene were selected by primary PCR analysis and further analyzed in quantitative real-time PCR(qRT-PCR). The qRT-PCR analysis demonstrated that SSU rRNA of N. ceranae was detected at concentrations as low as 0.85 ng/μl genomic DNA. This result suggests that the detection via qRT-PCR can be applied for the rapid and sensitive diagnosis of N. ceranae infection in the field population as well as risk assessment of B. terristris.

Bombus terrestris has played an important role in the pollination in agricultural fields for the alternatives in colony collapsing in the honeybee. Recently, some pathogens or parasites such as viruses, bacteria, mites have been discovered in B. terristris, which affects its life span and fecundity. In order to detect a microsporidian, Nosema apis. in the field population, we collected honeybees and isolated genomic DNA. PCR primers specific for 16S ribosomal RNA (16S rRNA) were synthesized and applied to gene amplification for cloning and quantitative real-time PCR (qRT-PCR). The amplified gene was cloned and sequenced to confirm the 16S rRNA gene. qRT-PCR analysis showed the detection limit of 16S rRNA of Nosema apis was approximately 0.5 ng/μl genomic DNA. This result suggests that detection via qRT-PCR can be applied for the diagnosis of pathogen infection.

Nosema disease caused by Nosema apis which belongs to fungi is a major cause of honey production loss and is worldwide in distribution. N. apis infects the epithelial cells of the digestive system of adult honeybees. Nosema causes significant losses in population size of honeybee. There are about 25 thousand beekeepers caring for approximately 1,697,000 colonies in Korea. Honey production totaled almost 38,505 metric tons in 2010. This production was estimated to be worth about 274 billion Korean won. To determine infection level of nosema disease during the season, adult worker bees were collected from two colonies of experiment apiary from January to October. Our results indicate that the infection level of nosema disease was increased in spring and autumn. Also we initiated a survey of honeybee colonies on the blooming period of Acacia to determine the prevalence of N. apis. Twenty two hives owned by 18 beekeepers were sampled for this study. Bees were collected on 24th and 25th May of 2012. Nosema spore counts ranged from zero to 5,266,000 spores per bee. The average number of nosema spores per bee was calculated to be 1,375,000. Approximately 86% of the apiaries examined were infected with nosema, based on the presence of spores in the flowering period of Acacia. This indicates that nosema is the predominant species affecting honeybee colonies.

Taxonomic resolution of the Nosema/Vairimorpah clade has been augmented with DNA sequences of the small subunit (SSU) and large subunit (LSU) ribosomal RNA (rRNA) and the arrangement of SSU and LSU. Based on the two characteristics, the clade is largely divided into two, i.e. 'true' Nosema sub-group and 'non-true' Nosema sub-group within the clade. Our study shows that a novel Nosema species isolated from Pieris rapae has mixed characteristics of the 'true' and non 'true' Nosema sub-group based on the topology of SSU and LSU sequences, and rRNA of the isolate is normally organized. Additionally, the length of ITS can be a diagnostic tool to distinguish 'true' Nosema from non 'true' Nosema in the Nosema/Vairimorpha clade based on its nucleotide length as reported before. To our knowledge, this is the first report of recombination event in the Nosema/Vairimorpha clade.

A multiplex polymerase chain reaction (PCR) was developed for the simultaneous detection and differentiation among Nosema apis and Nosema ceranae in honeybee. Three sets of primers were selected from different genomic sequences to specifically amplify a 831 bp amplicon within the SSU rRNA gene, specific for both N. apis and N. ceranae (MSSR primer); a 375 bp amplicon within the SSU rRNA gene, specific for N. apis (NA primer); and a 1,131 bp amplicon within SSU rRNA gene, specific for N. ceranae (NC primer). Using the primers in conjunction (multiplex PCR) we were able to N. apis and N. ceranae and to differentiate between them. The sensitivity of this PCR assay was approximately 102 spores per milliliter. We proposed that the multiplex PCR was sensitive, specific and rapid tool that can serve as a useful differential diagnostic tool for detecting N. apis and N. ceranae in honeybee.

Microsporidia are obligate fungal intracellular parasites of all animal taxa. Among them the genus Nosema (Nosematidae) is known as the most common entomopathogen. Of these parasites, the ribosomal organization is one of the most pronounced molecular characteristics. One type is the normalarrangement of small subunit (SSU)-internal transcribed spacer (ITS)-large subunit (LSU) in the DNA sequence order. The other is the reverse arrangement of LSU-ITS-SSU. The latter is assigned to be the ‘true’ Nosema in the Nosema/Vairimorpha clade. However, we found that the SSU sequence of a strain of Nosema species having the normal arrangement of its rRNA sequence seemed to be more closely related to the ‘true’ Nosemagroup. Consequently we have further analyzed the complete sequence of rRNA. The results imply that there might be arecombination event in its rRNA evolution and/or the strain may form a novel group near the ‘true’ Nosema group. Interestingly both SSU and LSU of the ‘true’ Nosema and others may be under different selection pressure. We have also found that the size of ITS is distinct between the ‘true’ Nosema and other microsporidian species within the Nosema/Vairimorpha clade. This feature should be a useful diagnostic tool to distinguish the ‘true’ Nosema from others in the clade.

A multiplex polymerase chain reaction (PCR) was developed for the simultaneous detection and differentiation among Nosema apis and Nosema ceranae in honeybee. Three sets of primers were selected from different genomic sequences to specifically amplify a 831 bp amplicon within the SSU rRNA gene, specific for both N. apis and N. ceranae (MSSR primer); a 375 bp amplicon within the SSU rRNA gene, specific for N. apis (NA primer); and a 1,131 bp amplicon with in SSU rRNA gene, specific for N. ceranae (NC primer). Using the primers in conjunction (multiplex PCR) we were able to N. apis and N. ceranae and to differentiate between them. The sensitivity of this PCR assay was approximately 102spores per milliliter. We proposed that the multiplex PCR was sensitive, specific and rapid tool that can serve as a useful differential diagnostic tool for detecting N. apis and N. ceranae in honeybee.