A nematode, which showed entomopathogenicity, was isolated from Exomala orientalis in Korea, and then identified as Rhabditis blumi. Pathogenicity of R. blumi was evaluated against major cruciferous insect pests, including Artogeia rapae, Mamestra brassicae, and Plutella xylostella. In Petri-dish tests, insect mortality was dose and time dependent, which increased with dose (0–80 dauer juveniles/larva) and time increments. In greenhouse tests, P. xylostella larvae were most susceptible to nematodes, with insect reduction rate of 88.0 %. The rate varied with vegetable species and persistence time of live nematodes on vegetable leaves after spraying. The aeration rate was an important parameter for cultivation of the nematode R. blumi. A sufficient DOT level in the culture using an air-lift bioreactor should be maintained. The nematode growth rate increased with an increasing DOT level and/or bacteria uptake rate. The maximum nematode yield of 1.75 x 105 per ml was obtained under an aeration rate of 6 vvm.

수환경에서 유해성 미생물을 검출하는 것은 환경미생물위해성 관리에서 가장 중요한 연구 분야 중 하나이다. 유해성 미생물 검출은 인체 병원성 세균과 항생제 내성을 지닌 세균의 검출을 포함한다. 기존의 배양방법은 배양이 잘 되지 않는 유해성 세균의 검출이 불가하므로, 최근에는 환경 유전체를 전체로 염기서열을 분석하는 de nove metagenomic sequencing 방법으로 병원성에 대한 진단을 하려는 방법에 대한 연구가 시작되고 있다. 본 연구에서는 수환경에서 유해 미생물 검출을 위해 Next Generation Sequencing 방법인 Illumina Hi-Seq으로 미생물군집유전체를 shot-gun sequencing해서 병원성과 항생제 내성에 대한 정량적인 정도 도출하는 신규 방법에 대해서 결과와 함께 토의할 것이다.

표고버섯좀나방(Morophagoides moriutii)은 최근 버섯 에 심각한 피해를 주는 해충으로 유충이 골목이나 종균, 자실체를 직접 가해하여 생산량을 감소시키고 상품성을 저하 시킨다. 표고버섯좀나방의 환경 친화적 방제에 이용 할 수 있는 가능성을 알아보기 위하여 국내 토양에서 탐 색된 토착 곤충병원성 선충 Steinernema carpocapsae GSN1 계통의 접종 농도별 병원성, 침입수, 증식수를 표고 버섯좀나방의 령기에 따라 조사하였다. 선충의 표고버섯 좀나방에 대한 병원성은 접종농도가 높을수록 증가하였다. 표고버섯좀나방 2령충, 3령충에서 유충 한 마리당 선충 감염태 유충(3령충) 80마리 접종부터 100%의 높은 병원 성을 나타내었고, 표고버섯좀나방 4령충은 한 마리당 선 충 감염태 유충 40마리 접종부터 100%의 치사율을 나타 내었다. 선충의 표고버섯좀나방 유충 체내로의 침입수는 접종 농도와 령기가 높을수록 선충의 침입수도 증가 하였 다. 선충 감염태 유충 5마리 농도 처리에서는 2령충 1.3마 리, 3령충 1.7마리, 4령충 1.8마리 였으나, 160마리 농도 처리에서 2령충 12.3마리, 3령충 21.1마리, 4령충 22.5마 리로 조사 되었다. 끝으로 표고버섯좀나방 유충 한 마리 당 침입한 선충의 증식수는 같은 령기에서는 선충의 접종 농도가 많아질수록 증식수가 증가하였다. 또한 표고버섯 좀나방의 령기에 따라 선충 증식수의 차이는 많았는데, 선충 감염태 유충 160마리 처리농도에서는 각각 표고버 섯좀나방 2령충이 6,335마리, 3령충이 21,660마리, 4령충 이 88,700마리로 조사되었다. 따라서 토착 곤충병원성 선 충 S. carpocapsae GSN1 계통은 표고버섯좀나방의 생물 적 방제를 위한 방제제로 활용할 가능성이 높음을 알 수 있었다.

본 연구에서는 파프리카 재배포장과 저온저장 후 미생물상을 조사하였으며, 동정된 미생물의 병원성을 검정하여 전염원 유입과 저장병 발생의 관련성을 조사하였다. 재배포장에서는 세균의 경우 과일표면에서 Bacillus spp., Clavibacter sp.와 Pseudomonas spp. 등 7종류의 세균이 분리되었으며 저온저장 후에는 인체에 유해한 균을 포함하여 Erwinia sp., Salmonella spp., Streptococcus spp. 등 8종류의 세균이 분리되었다. 재배포장과 저온저장 후 분리된 주요 진균은 Alternaria sp,, Botrytis sp,, Cladosporium sp,, Fusarium sp,와 Penicillium sp.가 분리되었으며, Geotrichum sp.은 저온저장 후 과일에서 분리되었다. 진균의 경우 분리된 균 모두 파프리카의 부패에 관여하는 것으로 나타났으나 분리된 세균을 단독으로 접종하였을 때 파프리카의 부패에 관여하지 않는 것으로 조사되었다. 이 결과로 진균에 의해 1차적으로 저장병이 발생한 후 세균에 의해 2차적 부패가 발생하는 것으로 판단된다.

토마토 재배에 사용되는 농자재를 이용하여 곤충 병원성 곰팡이인 Beauveria bassiana의 병원성에 끼치는 영향을 조사하였다. Chlorothalonil 등 13종의 살균제 첨가 배지에서는 B. bassiana 균사가 전혀 생장하지 못하였고 chlorothalonil 등 12종의 살균제 첨가 배지에서균총이 형성되지 않았다. 살충제와 친환경자재를 첨가한 모든 배지에서 균사가 생장하고 포자가 형성되었지만 대부분 배지에서 무처리와 비교했을 때 유의차를 보였다. 온실가루이에 대한 살충력은 B. bassiana 단독처리일 때보다 화학살균제(polyoxin B, mandipropamid)와 친환경농자재(sulfur) 혼합시 떨어졌으며 화학살충제(pyridaben, dinotefuran)와 식물추출물 유래 친환경농자재(pyrethrum) 혼합시에는 유의차가 없었으며 온실에서 살균제인 polyoxin B의 혼합으로 온실가루이에 대한 B. bassiana의 감염력이 떨어지는 것을 확인하였다.

 ,  , The entomopathogenic bacterium, Xenorhabdus sp., was isolated from an entomopathogenic nematode, Steinernema monticolum. When these bacteria were injected into the hemocoel of the diamondback moth, Plutella xylostella, they caused significant mortality. However, the bacterium was not pathogenic when it was administered orally. This study showed that Xenorhabdus sp. significantly enhanced oral pathogenicity of Bacillus thuringiensis (Bt) against the last instar larvae of P. xylostella. Different ratios of culture broth of Xenorhabdus sp. and Bt showed significantly different pathogenicities against P. xylostella. In field tests, the optimal bacterial mixture significantly enhanced control efficacy against P.xylostella compared to Bt treatment alone. These results demonstrated that Xenorhabdus sp. culture broth can be developed as a potent biopesticide by enhancing the insecticidal efficacy of Bt.

 ,  , The size of infective Steinernema arenarium juveniles is variable and ranges from 724 to 1408 ㎛. Effects of harvest time and infective juvenile size on pathogenicity, development, and reproduction were examined in the last instar of the great wax moth, Galleria mellonella. Harvest time of infective juveniles (IJs) of S. arenarium affected pathogenicity. IJs harvested at the 10th day from trapping were more pathogenic than those harvested the 3rd day from trapping. Mortality of G. mellonella also depending on harvest time, i.e, 100% died within 48h when IJs were harvested at the 10th day, without relation to size. However, mortality was 40% in the small size group (SSG) compared with 18% in the large size group (LSG) within 48h when IJs were harvested at the 3rd day. Establishment of S. arenarium within the host was different depending on IJ size. The number of established IJs was 1.8 in the SSG, 3.3 in the LSG, and 3.2 in the mixed size group (MSG) when IJs were harvested at the 3rd day, and 5.3 in the SSG, 7.4 in the LSG, and 7.6 in the MSG when IJs were harvested at the 10th day. The length of the female adult was 7,070.5 ㎛ in the SSG and 7,893.9 ㎛ in the LSG and that of the male was 1,460.5 ㎛ in the SSG and 1,688.2 ㎛ in the LSG when IJs were harvested at the 3rd day. The length of the female adult was 7,573.6 ㎛ in the SSG and 8,305.4 ㎛ in the LSG and that of the male adult was 1,733.4 ㎛ in the SSG and 1,794.4 ㎛ in the LSG when IJs were harvested at the 10th day. Harvest time and size of IJs did not influence numbers of progeny or size of IJS.

본 연구는 누에의 식엽성 곤충을 대체할 실험곤충으로서의 유용성과 자외선이 한국산 곤충병원성선충 (Heterorhabditis sp. 202 strain, Heterorhabditis sp. Gyeongsan strain, Steinernema sp. 223 strain, S. carpocapsae Pocheon strain, S. glaseri Dongrae strain, S. longicaudum Nonsan strain) 의 병원성과 생존에 미치는 영향을 알아보기 위하여 수행하였다. 누에에 대한 곤충병원성선충의 병원성을 조사한 결과 누에의 령기나 곤충병원성선충의 계통이나 종에 따라 상이한 병원성을 나타내었는데 5령충에 대해서는 Steinernematidae선충의 병원성이 높았고, 1령충에 대해서는 Heterorhabditidae선충의 병원성이 높았다. UV-C는 곤충병원성선충에 유해하여 노출 60분후에는 모든 선충이 치사되었다. UV-C 조사는 곤충병원성선충의 병원성에도 영향을 미쳐 노출 10분 후에도 누에나방 3령충에 대하여 6.7% 이하의 보정사충율을 나타내었다. 야외 뽕나무밭(UV량: 2.3mW/㎠)에서 곤충병원성선충, S. carpocapsae Pocheon strain을 엽면 살포 4시간 후 채취한 뽕나무 잎을 급상한 누에는 치사되지 않았다. 뽕잎에서 4.0mW/㎠의 UV 1시간 노출은 곤충병원성선충, S. carpocapsae Pocheon strain의 누에에 대한 병원성에 차이를 보이지 않았다.

Cobweb disease symptoms were observed in a mushroom farm in Buye, Korea during a disease survey in 2008-2011. Five isolates of Cladobotryum sp. were obtained from the infected caps and stipes. These isolates of Cladobotryum sp. were identified as C. mycophilum based on their morphological, cultural characteristics and analysis of the ITS sequences. Early symptoms were noticed as round, fleshy, yellowish brown lesions on mushroom caps. Late symptoms progressed when the parasitic fungus formed white cobweb circular colonies on dead or damaged pinheads, spread on the surface of the casing, and covered entirely fruiting bodies. Optimal temperature and pH for mycelial growth on MEA is 23℃ and 6.0. Microscopically the spores of the fungus are large and most 2~3 celled produced on vertically branched conidiophores. Mushroom caps turned dark brown and shrunk due to soft rot. Testing of sensitivity to selected fungicides showed that isolate was highly resistance to Mancozeb and Thiophanate-methyl, moderately sensitivity to Iprodione, and highly sensitivity to Benomyl, Prochloraz-Mn and Carbendazim.

An entomopathogenic bacterium, Xenorhabdus sp., symbiotic to Steinernema monticolum was investigated in its insecticidal activity. The bacteria induced septicemia of two lepidopteran insects (Plutella xylostella, Spodoptera exigua), a coleopteran insect (Tribolium castaneum), and a hemipteran insect (Riptortus clavatus) when they were injected into hemocoel. The bacterial culture broth contained immunosuppressive factor(s) that inhibited hemocyte nodulation in response to heat-killed bacteria. The immunosuppressive activity appeared to be caused by inhibition of two main immune-associated enzymes, phospholipase A2 (PLA2) and phenoloxidase (PO). HPLC analysis of the bacterial culture broth contained several PLA2 inhibitors. The bacterial culture broth significantly enhanced Bt pathogenicity. There results support a novel insect pest control strategy using eicosanoid-biosynthesis inhibitors.

Autographa californica nucleopolyhedrovirus (AcMNPV) has a large doublestrand DNA genome of approximately 134 kbp and harbors 156 open reading frames (ORFs). To elucidate DNA replication cascade of AcMNPV, we developed a novel baculovirus genome that can be maintained in Escherichia coli as a plasmid and can infect susceptible lepidopteran insect cells. This genome, named bAc-MK, contains a mini-F replicon and a kanamycin resistance marker. Using a convenient Tn7 transposon-based system, pPCS-S, 55 single ORF-truncated mutants were generated by random insertion into bAc-MK genome. These single ORF-truncated mutants were independently transfected into Sf9 cells, 16 of them were found affecting viral replication since they defected in producing polyhedra. Furthermore, to verify the pathogenicity of the single ORF-truncated mutants, the remaining 39 mutants were subjected to bioassay to Spodoptera exigua 3rd instar larvae. Among them, ac9-, ac49-, ac103- and ac105-knockout mutants showed higher mortality compared to that of bAc-MK. These results suggested that these ORFs could be related to pathogenicity of AcMNPV.

Effect of inoculation level on pathogenicity, development, and propagation of entomopathogenic nematode, Steinernema arenarium was investigated using the last instar of great wax moth, Galleria mellonella. Pathogenicity of S. arenarium was higher with increasing inoculation level representing 82% at the rate of 5 infective juveniles (IJs) while >98% at the rate of >10 IJs. The number of IJs penetrated into the host was 2.7, 5.0, 7.4, and 12.2 at the rate of 5, 10, 20, and 40 IJs, respectively while 24.3 at the rate of 80 IJs and 40.2 at the rate of 160 IJs. Inoculation level did not affect female adult size (4,616 to 6,444 ㎛) while affected male adult size (1,600 to 1,934 ㎛). The rate of stunted female adults was 70.2% at the inoculation level of 80 IJs and 63.7% at the inoculation level of 160 IJs. The number of progenies was 20,431, 26,696, 47,943, 50,516, 58,701, and 74,235 at the rate of 5, 10, 20, 40, 80, and 160 IJs, respectively. The body lengths of IJs were different depending on inoculation level ranging from 636 to 1,496 ㎛.

Two entomopathogenic bacteria, Xenorhabdus nematophila and Photorhabdus temperata subsp. temperata, are known to be potent against the diamondback moth, Plutella xylostella, when the bacteria are injected into the hemocoel. This study investigated any pathogenic effect of their culture broth on P. xylostella by oral administration. Only culture broth of both bacterial species did not give enough pathogenic effects by the oral administration. However, when the culture broth was orally treated together with Bacillus thuringiensis (Bt), both cell-free culture broth significantly enhanced Bt pathogenicity against the 3rd instar larvae of P. xylostella. The culture broth was then fractionated into hexane, ethyl acetate, and aqueous extracts. Most synergistic effect on Bt pathogenicity was found in ethyl acetate extracts of both bacterial species. Thin layer chromatography of these extracts clearly showed that ethyl acetate extracts of both bacterial culture broths possessed metabolites that were different to those of hexane and aqueous extracts. These results suggest that the both entomopathogenic bacteria produce and secrete different factors to give significant synergistic effect on Bt pathogenicity.

Entomopathogenic nematode, Steinernema carpocapsae GSN1 strain was evaluated for the environmentally sound control of Tebenna issikii (Lepidoptera: Choreutidae) in the laboratory. The corrected mortality of Tebenna issikii larvae was 100% at the 40 infective juveniles (Ijs)/larva 3 days after treatment with S. carpocapsae GSN1 strain in Petri dish. LC₅₀ value of S. carpocapsae GSN1 strain against Tebenna issikii was 5.7 Ijs. The mean penetration numbers of Ijs of S. carpocapsae GSN1 strain at the 5, 10, 20, 40 and 80 Ijs/larva in a Tebenna issikii larva were 1.4, 1.4, 3.2, 5.6 and 11.9 Ijs/larva, respectively. However penetration rate of Ijs of S. carpocapsae GSN1 strain at 5 Ijs/larva was the highest among other nematode concentrations. Progeny of S. carpocapsae GSN1 strain in a Tebenna issikii larva was higher with increasing nematode concentration.

Korean entomopathogenic nematodes were introduced into cultural management of turfgrasses to enhance white grub control in golf courses for saving labour and expenses and contribution to giving safe playing and working places for golfers and superintendents by environmentally friendly control strategy. The turfgrass height influenced efficacy of entomopathogenic nematodes. Efficacy was higher at short turfgrass both in pot using Galleria mellonella larvae and in golf courses using 2nd instar of Exomala orientalis and Ectinohoplia rufipes as baits. Aeration increased the efficacy of entomopathogenic nematodes which were lower virulent. Pre-aeration was more effective than post-aeration. Exomala orientalis larval population was reduced 80.4 and 66.0% in the pre-aeration and post-aeration with entomopathogenic nematode, Steinernema carpocapsae Pocheon strain compared to 35.4% in the no aeration treatment.

골프장의 문제해충들 중 녹색콩풍뎅이(Popillia quadriguttata) 성충의 방제를 위하여 세 종의 한국산 곤충병원성선충 Steinernema carpocapsae KCTC 0981BP 계통 (ScK), S. glaseri Dongrae 계통(SgD), Heterorhabditis sp. KCTC 0991BP 계통 (HsK)을 이용하여 실험을 수행하였다. 3일 후, 녹색콩풍뎅이 성충 한 마리당 900마리의 선충을 처리하였을 때, 치사율은 ScK가 , HsK가 , SgD가 를 각각 나타냄으로써 이용한 선충 모두가 높은 병원력을 나타내었다. 성충 한 마리 당 900마리의 선충을 처리한 3일 후의 치사율은 ScK는 , HsK는 , SgD는 였다. 선충의 녹색콩풍뎅이 성충 표면부착율과 침입율은 접종 선충의 종과 밀도에 따라 차이가 있었다. 즉, 성충한 마리당 900마리의 선충을 처리하였을 때 녹색콩풍뎅이 성충의 표면에서 검출된 HsK의 비율은 였으며, SgD는 , ScK는 였다. 녹색콩풍뎅이 성충 내에 침입한 선충의 검출여부에 따른 선충의 침입율은 성충 한 마리 당 900마리의 선충을 처리한 3일 후 ScK는 , HsK와 SgD는 각각 를 보였다. 반면, 성충 한 마리 당 90마리의 선충을 처리한 3일 후의 ScK 검출율은 , SgD는 였으나 HsK는 발견되지 않았다.