이 연구는 부여 105AS-13.2.3-BQ-B1 교육부에서 동물의 국과 식물 건강 검사 및 검역, 농업위원회, 집행 위안 그랜트 103-2313-B-197-002-MY3에서에 의해 지원되었다 과학 기술 (MOST)의.

1. 준비 <ol><li> 50 %의 설탕 시럽 1 L합니다. 1 L의 DDH 2 O.에 1kg 자당 녹이고 </li><li> BLD에서 pyriproxyfen (PPN) 솔루션을 준비합니다. 10,000 ppm의 PPN 원액 1.1 L을 4 ° C에서 2 DDH O. 스토어 멸균 1 L에 PPN 100 ㎖ 용액을 희석. </li><li> 다음 실험을위한 BLD 0.1, 1, 10 및 100 ㎎ / ㎏ (PPM)의 최종 농도로 PPN 원액을 희석. </li><li> (식민지 레벨) PPN-시럽을합니다. 다음 실험에 대한 50 %의 설탕 시럽 10 및 100 ppm으로 최종 농도로 PPN 주식을 희석. </li><li> 꿀 꿀벌 사육. 참고 : 여기, 실험 위치는 국립 일란 대학 (NIU) 양봉장, 이순신 - 란시, 대만은; (GPS 좌표 : N24.747278, E121.746200). <ol><li> 꿀벌 (아피스 mellifera이 L.)는 식품 수량 매주 식민지 확인하고 (꿀 저장 영역이 비어) 필요한 경우 1리터 50 %의 설탕 시럽을 공급. 건강한 정의일반적 산란 퀸 각 콜로니의 꿀벌 빗 9 프레임으로서 콜로니. </li></ol></li><li> 기본 애벌레 다이어트 (BLD)의 100 mL로 준비합니다. (DDH 2 O) 멸균 증류수, 탈 이온수에 6 % D- 글루코오스, 프 룩토 오스 6 %, 1 % 효모 추출물을 용해시키고, 50 % 로얄 젤리 보충. 4 ° C에서 보관하지만 3 일 이상합니다. 실험 전에 35 ° C로 사전 따뜻하고 3 일 이내에 사용합니다. </li></ol> 생체 수유 방법 2. 노트 : 생체 내에서 공급 법은 헨리 등의 알로부터 변형되었다. (20) <ol><li> 꿀 꿀벌 유충 선택 및 라벨링. <ol><li> 4 프레임 부에 퀸을 제한하면서 4 프레임 5 개 프레임으로 건강한 콜로니 9 프레임 분할 퀸 excluder입니까. 산란을 위해 4 프레임 섹션에서 적어도 하나 개의 빈 프레임을 남겨주세요. </li><li> 여왕이 1 일 산란 후계란의 외관을 검사 프레임 1 일령 유충이 부화까지 72 시간 (4 일째)에 벌통 내부의 알을 유지한다. 손으로 테스트 하이브 밖으로 (24 시간 내에 부화) 1 일 된 노동자 유충을 포함하는 프레임 중 하나를 가지고 꿀벌 브러시 프레임에서 꿀벌 노동자를 제거합니다. </li><li> 투명 슬라이드 (크기 = 길이 X 너비 X 두께 29.7 mm X 21mm X 0.1 mm)를 가진 프레임을 덮고 압정 (도 1a)와 프레임의 가장자리에 상기 투명 슬라이드 손톱. </li><li> 각각의 치료를위한, 임의로 하나 50 일령 유충 (4 일째)을 선택하고, 상기 투명 슬라이드 영구 마커 펜을 사용하여 각 브루 셀 마크 (도 1a 및도 1b). 참고 : 다른 치료 사이의 혼동을 피하기 위해 영구 마커 펜을 사용하여뿐만 아니라 프레임과 투명 슬라이드의 각 치료의 정보를 작성합니다. 표시된 슬라이드를 제거하고 생체에 대한 유지 &lt;/ EM&gt; 급전 관찰. </li></ol></li><li> 생체 먹이. <ol><li> 의 흡기 양에 따라 각각 10 μL, 10 μL, 20 μL로 일 1, 2, 3에 피펫으로 각 표지 브루 셀 PPN-BLD (0.1, 1, 10, 100 PPM)의 상이한 농도를 추가 유충의 나이. 대조군에 BLD (NO PPN)의 동일한 금액을 추가합니다. 이에 따라, 각 분류 브루 셀 PPN-BLD의 총 투여 량은 4, 40, 400, 4000 페이지에 축적한다. 참고 : 표시된 투명 슬라이드하여 표시 무리 세포를 인식하고 공급 한 후 표시된 투명 슬라이드를 제거합니다. 일벌에 의해 무리 세포의 청소를 방지하기 위해 공급하는 신선한 BLD를 사용합니다. </li><li> 더 관찰의 원래 식민지로 PPN 처리 된 프레임을 돌려줍니다. 참고 : 각각의 치료는 네 개의 식민지에서 네 생물 반복 있습니다. </li></ol></li><li> 7 일에서 처리 유충의 관측. <ol><li> 일을 관찰하려면전자 표지 무리 세포의 사망률과 상한 율을 기록하기 위해 프레임에 표시된 투명 슬라이드를 반환 유충을 PPN은 처리. </li></ol></li><li> 하루에 13에서 처리 번데기와 우화 요금의 관측. <ol><li> 덮인 무리 세포의 꿀벌 왁스를 제거합니다. </li><li> 브루 셀에 소프트 팁 핀셋을 넣고 클램프 번데기를 아주 약간 부드럽게 번데기를 꺼내. </li><li> 각 웰 아래 기관 조직의 이중층으로 24 웰 조직 배양 플레이트에 번데기로 이동. 번데기 전송 중에 손상과 사망률을 기록합니다. </li><li> (약 팔일) 출현 될 때까지 34 ° C, 70 % 상대 습도 인큐베이터에서 24 웰 조직 배양 플레이트 유지. </li><li> 관찰과 번데기와 등장 꿀벌을 기록합니다. </li></ol></li><li> 통계 <ol><li> ± SD 평균값으로 기록 된 데이터와 현재 계산 </li><li> SA에 의해 변동 (ANOVA)을 이용하여 분석 데이터를 분석S 및 다른 처리의 두 방법의 차이를 분석하기 위해 최하위 차 (LSD)를 사용하여 시험. P - 값 &lt;0.05로 통계적으로 유의 한 정의합니다. 테이블의 동일한 열에서 서로 다른 글자는 통계적 분석에 의해 현저한 효과를 나타냈다. </li></ol></li></ol> 벌집에서 식민지 수준에서 PPN 3. 독성 <ol><li> 꿀벌 그룹을 설정합니다. <ol><li> 프레임 4 (파트 A) 및도 5의 프레임 (파트 B) 4 프레임 부에 여왕으로 제한하면서 건강한 콜로니 9 개 프레임을 분할하는 수직 퀸 excluder입니까. 산란을위한 4 프레임 부분에 적어도 하나 개의 빈 프레임을 남겨주세요. 참고 : 다른 여왕 부품 사이에 이동 여왕을 방지하기 위해 여왕 부분의 상단에있는을 excluder 넣습니다. </li><li> 여왕이 1 일 산란 한 후, 달걀 모양의 프레임을 선택합니다. 손으로 A 부분에서 계란을 포함하는 적절한 프레임을 가지고 꿀벌 근로자를 제거꿀벌 브러시로 프레임입니다. </li><li> 투명 슬라이드 프레임을 덮고 압정와 프레임의 가장자리에 상기 투명 슬라이드 손톱. </li><li> 임의로 함유 달걀 100 개 브루 셀을 선택하여 상기 투명 슬라이드 영구 마커를 사용하여 각 브루 셀을 표시한다. 그룹 1. 다른 치료 사이의 혼동을 피하기 위해 영구 마커를 사용하여 프레임과 투명 슬라이드의 각 치료의 정보를 기록 이러한 100 개 무리 세포를 할당합니다. </li><li> 3 일 A 부분에 표시된 프레임을 반환 한 후 알을 낳기 위해 파트 B에 여왕을 전송합니다. </li><li> 일일 후, 파트 B의 프레임을 검사 한 적절한 프레임을 선택하고, 단계 3.1.4에 기술 된 바와 같이 계란 함유 100 브루 셀 라벨. 그룹이 이러한 100 개 무리 세포를 할당합니다. </li><li> 3 일 파트 B로 표시된 프레임을 반환 한 후 알을 낳기에이 부분에 여왕을 전송합니다. </li><li> 6 번 파트 A와 파트 B의 여왕 교환을 반복하고 수치 그룹을 할당,각각 (도 2). 9 개 그룹의 총이 있어야합니다. </li></ol></li><li> 일 13 PPN 설탕 시럽 T reat 꿀벌 콜로니 (도 2). <ol><li> 1 L 플라스틱 꿀벌 급전 박스 (10) 또는 100 PPM (W X L x 높이 = 20 cm X 30cm X 3.5 cm)를 함유하는 50 % PPN 설탕 시럽을 추가 실험 콜로니의 프레임의 상부에 박스를 넣어. 참고 : 무리 세포가 봉인 된 것처럼 그룹 1 13 일째에 PPN을받지 않습니다. </li><li> 단계 2.2에 기재된 바와 같이 1 L 50 % 설탕 시럽 (NO PPN)와 대조군 사료. </li></ol></li><li> 5 일 (그림 2)에서 각 그룹에 대해 100 개 라벨 무리 세포의 계란 부화 속도로 일일 된 유충 기록을 계산합니다. 꿀 꿀벌 계란은 일반적으로 0일 유충으로 부화 3 일이 걸릴; 따라서 확인하고 100 계란 함유 하루 1에 무리 세포를 라벨 및 부화 속도의 비율을 얻기 위해 하루 5에서 각 그룹에 대해 1 일 된 유충의 수를 계산합니다. </li><li> 기음날 (11) (도 2)에서 각 그룹 (100 개) 표지 브루 셀의 애벌레 캐핑 레이트 캡핑 브루 셀 기록 ount. 6 ~ 7 일 된 애벌레 무리 세포는 애벌레 pupating 꿀 꿀벌 노동자 꿀벌 왁스로 제한됩니다. </li><li> 번데기 성숙을 관찰하고 17 일 (도 2)에서 각 그룹 (100 개) 표지 브루 셀의 우화 율을 기록한다. <ol><li> 덮인 무리 세포의 꿀벌 왁스를 제거하고 부드럽게 부드러운 밀고 족집게와 번데기를 꺼내. 각 웰 아래 기관 조직의 이중층으로 24 웰 조직 배양 플레이트에 번데기로 이동. </li><li> 출현 될 때까지 34 ° C, 70 % RH에서 항온 24 웰 조직 배양 판을 유지한다. </li><li> 관찰과 번데기를 기록하고 그룹 9까지 각 그룹 (49 실험 일)를 위해 꿀벌을 등장. 참고 : 각 치료는 네 생물 반복 있습니다. </li></ol></li><li> 통계 <ol><li> 기록 데이터를 계산하고, 평균으로서 본± SD. </li><li> 치료의 쌍 사이의 유의 한 차이를 분석하여 (예를 들어 0 PPM / 10 PPM 10 PPM / 100 ppm을 0 PPM / 100 PPM) 학생의 양측의 t -test를 사용하여 각 그룹이다. P - 값 &lt;0.05처럼 통계적으로 유의 한 정의합니다. </li></ol></li></ol>

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Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+impact+of+pyriproxyfen+on+the+development+of+honey+bee+(Apis+mellifera+L.)+colony+in+field.">The impact of pyriproxyfen on the development of honey bee (Apis mellifera L.) colony in field.</a> J. Asia Pac. Entomol. 19 (3), 589-594 (2016).</li><li>Dennehy, T. J., DrGain, B. A., Harpold, V. S., Brink, S. A., Nichols, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Whitefly Resistance to Insecticides in Arizona: 2002 and 2003 Results. , 1926-1938 (2004).</li><li>Yang, E. C., Wu, P. S., Chang, H. C., Chen, Y. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+sub-lethal+dosages+of+insecticides+on+honey+bee+behavior+and+physiology.">Effect of sub-lethal dosages of insecticides on honey bee behavior and physiology.</a> , (2010).</li><li>Fourrier, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Larval+exposure+to+the+juvenile+hormone+analog+pyriproxyfen+disrupts+acceptance+of+and+social+behavior+performance+in+adult+honey+bees.">Larval exposure to the juvenile hormone analog pyriproxyfen disrupts acceptance of and social behavior performance in adult honey bees.</a> PLoS One. 10, e0132985 (2015).</li><li>Hanley, A. V., Huang, Z. Y., Pett, W. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+dietary+transgenic+Bt+corn+pollen+on+larvae+of+Apis+mellifera+and+Galleria+mellonella.">Effects of dietary transgenic Bt corn pollen on larvae of Apis mellifera and Galleria mellonella.</a> J. Apicult.Res. 42 (4), 77-81 (2003).</li><li>Kaftanoglu, O., Linksvayer, T. A., Page, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rearing+honey+bees,+apis+mellifera,+in+vitro+1,+effects+of+sugar+concentrations+on+survival+and+development.">Rearing honey bees, apis mellifera, in vitro 1, effects of sugar concentrations on survival and development.</a> J. Insect Sci. 11 (96), 1-10 (2011).</li><li>Vandenberg, J. D., Shimanuki, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Technique+for+rearing+worker+honey+bees+in+the+laboratory.">Technique for rearing worker honey bees in the laboratory.</a> J. Apicult. Res. 26 (2), 90-97 (1987).</li><li>Peng, Y. S. C., Mussen, E., Fong, A., Montague, M. A., Tyler, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+chlortetracycline+on+honey+bee+worker+larvae+reared+in+vitro.">Effects of chlortetracycline on honey bee worker larvae reared in vitro.</a> J. Invertebr.Pathol. 60 (2), 127-133 (1992).</li><li>Bitondi, M. M., Mora, I. M., Simoes, Z. L., Figueiredo, V. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Apis+mellifera+pupal+melanization+program+is+affected+by+treatment+with+a+juvenile+hormone+analogue.">The Apis mellifera pupal melanization program is affected by treatment with a juvenile hormone analogue.</a> J. Insect Physiol. 44 (5-6), 499-507 (1998).</li><li>Zufelato, M. S., Bitondi, M. M., Simoes, Z. L., Hartfelder, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+juvenile+hormone+analog+pyriproxyfen+affects+ecdysteroid-dependent+cuticle+melanization+and+shifts+the+pupal+ecdysteroid+peak+in+the+honey+bee+(Apis+mellifera).">The juvenile hormone analog pyriproxyfen affects ecdysteroid-dependent cuticle melanization and shifts the pupal ecdysteroid peak in the honey bee (Apis mellifera).</a> Arthropod Struct. Dev. 29 (2), 111-119 (2000).</li><li>Santos, A. E., Bitondi, M. M., Simoes, Z. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hormone-dependent+protein+patterns+in+integument+and+cuticular+pigmentation+in+Apis+mellifera+during+pharate+adult+development.">Hormone-dependent protein patterns in integument and cuticular pigmentation in Apis mellifera during pharate adult development.</a> J. Insect Physiol. 47 (11), 1275-1282 (2001).</li><li>Brouwers, E. V. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucose/Fructose+ratio+in+the+food+of+honeybee+larvae+during+caste+differentiation.">Glucose/Fructose ratio in the food of honeybee larvae during caste differentiation.</a> J. Apicult.Res. 23 (2), 94-101 (1984).</li><li>Howe, S. R., Dimick, P. S., Benton, A. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Composition+of+freshly+harvested+and+commercial+royal+jelly.">Composition of freshly harvested and commercial royal jelly.</a> J. Apicult. Res. 24 (1), 52-61 (1985).</li></ol>

저자는 더 경쟁 재정적 이해 관계가 없음을 선언합니다.

여왕 제한 알을 낳는 방법과 퀸 교환 방법은이 프로토콜 내에서 필드 테스트를 위해 꿀벌 그룹을 설정하기위한 중요한 단계이다. 여왕 제한 알을 낳는 방법은 꿀벌의 수명주기의 동기화를 가능하게한다. 그 결과, 연구진은 살충제의 서로 다른 용량으로 치료를 위해 같은 나이 1 일 된 유충을 선택할 수 있습니다. 퀸 교환 방법, 여왕이 부 A (4 개 프레임) 및 살충제 살충제 잔류 물의 영향을 평가하?...

환경에서 농약의 존재는 가장 심각한 문제 중 하나 인 영향을 미치는 꿀 꿀벌 1, 2, 3의 생명. 몇몇 연구는 꿀벌 식민지 꿀벌 제품에서 살충제 잔류의 일반적인 존재를 증명하고있다. 대만, 농약의 평균 응용 프로그램 11-12kg / (2005 년부터 2013 년) 매년 하였다. 대만에서 사용되는 농약의 양이 5 EU 국가보다 높은, 그리고 라틴 아메리카 국가 4. 즉, apicultural 환경은 특히 대만과 가능성이 다른 국가에서 심각한 농약 스트레스를 겪고있다. 꿀벌 API를 mellifera이 농업 시스템 (6)의 주요 매개자 중 하나이며 또한 꿀 가치있는 제품을 생산하고 있습니다. 그러나 꿀벌은 박람회는다양한 농약이 농약에 대한 ED 꿀과 꽃가루 7, 8을 수집 할 때 농약을 살포 한 꽃에서 먹이 찾아 돌아다 후 벌통에 다시 가져올 수 있습니다. 또한 자체가 두드러기 9, 10, 11 내 해충 문제를 제어하기 위해 목표로 양봉에 의해 농약에 노출 될 수 있습니다. 꿀벌 유충이 자신의 개발, 유충, 무인 항공기에 대한 간호사 꿀벌에 의해 공급되고 있기 때문에, 심지어 여왕은이 농약으로 오염 된 감로 및 꽃가루 (12)에 노출 될 수 있습니다. 꿀벌에 대한 다양한 농약의 독성 (13)를 해결해야합니다. 많은 노력은 환경 잔류 농약의 문제를 평가하기 위해 이루어졌다. 양 외. 에서 꿀벌 유충의 개발에 신경 독성 살충제 이미다클로 프리드의 영향을 테스트이 벌집과 이미다클로 프리드의 서브 치사량은 성인 꿀벌 후각 (14)의 결합 동작의 결과를보고했다. 또한 Urlacher 등. 실험실 조건 (15)에서 꿀 꿀벌 노동자의 학습 성능에 유기 인계 살충제, 클로르 피리 포스의 하위 치명적인 영향을 조사했다. 이전 연구에서 우리는 애벌레 꿀벌 16 (PPN) pyriproxyfen 곤충 생장 조절제의 영향 (IGR)을 평가. 본 논문에서는 꿀벌의 발전에 화학적 영향을 평가하기위한 방법을 제시한다. 꿀벌 공급 방법을 설명하고 하나의 개별 꿀벌 또는 콜로니에 적용 하였다. 첫째, 우리는 생체 내에서 개별 꿀벌에 대한 농약의 영향을 평가하기 위해 식민지에서 유충에 농약에 오염 된 기본 애벌레 다이어트 (BLD)의 서로 다른 농도를 시험했다. 우리는 자연 CONDIT을 시뮬레이션 진행벌통 내에 농약 오염 시럽을 사용하여 이온 살충제. 이 방법에서, 널리 해충 (17)에 대해 사용되며, 꿀벌 유충 및 번데기 (16), (18, 19)의 발전에 유해하다 PPN은 현장에서 농약의 부정적인 효과를 나타내는 지표가 될 것이다.

<table border="0" cellpadding="0" cellspacing="0" width="887">
	<tbody>
		<tr height="22">
			<td height="22" style="height:22px;width:193px;">Honey bee box</td>
			<td style="width:243px;">SAN-YI Honey Factory</td>
			<td style="width:155px;">W1266</td>
			<td style="width:297px;">Honeybees rearing</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:193px;">Queen excluder (between frames)</td>
			<td style="width:243px;">SAN-YI Honey Factory</td>
			<td style="width:155px;">I1575</td>
			<td style="width:297px;">Queen limitation&#160;</td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:193px;">Queen excluder (on top )</td>
			<td style="width:243px;">SAN-YI Honey Factory</td>
			<td style="width:155px;">I1566</td>
			<td style="width:297px;">Queen limitation on top&#160;</td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:193px;">Bee brush</td>
			<td style="width:243px;">SAN-YI Honey Factory, Taiwan</td>
			<td>W1414</td>
			<td style="width:297px;">clean the bees on frame gently</td>
		</tr>
		<tr height="18">
			<td height="18" style="height:18px;width:193px;">Bee feeder</td>
			<td style="width:243px;">SAN-YI Honey Factory, Taiwan</td>
			<td>P0219</td>
			<td style="width:297px;">feed sugar syrup to colony</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;">Transparent slide</td>
			<td style="width:243px;">Wan-Shih-Chei, Taiwan (http://www.mbsc.com.tw/a01goods.asp?s_id=40)</td>
			<td>1139</td>
			<td style="width:297px;">Mark the larval area on the frames (Material: Polyethylene Terephthalate, PET) (Size= Length*Width*thick= 29.7mm*21mm*0.1mm)</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:193px;">24 well tissu culture plate</td>
			<td style="width:243px;">Guangzhou Jet Bio-Filtration Co., Ltd</td>
			<td style="width:155px;">TCP011024</td>
			<td style="width:297px;">Rearing pupae from extraction</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:193px;">Autoclave</td>
			<td style="width:243px;">Tomin medical equipmenco., LTD.</td>
			<td style="width:155px;">TM-321</td>
			<td style="width:297px;">Make sterilized distilled deionized water (ddH2O)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:193px;">P20 pipetman</td>
			<td style="width:243px;">Gilson</td>
			<td style="width:155px;">F123600</td>
			<td style="width:297px;">Add PPN into bee larval food pool</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:193px;">Incubator&#160;</td>
			<td style="width:243px;">Yihder Co., Ltd.</td>
			<td style="width:155px;">LE-550RD</td>
			<td style="width:297px;">Rearing pupae from extraction</td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:193px;">Kimwipes</td>
			<td style="width:243px;">COW LUNG INSTRUMENT CO., LTD</td>
			<td style="width:155px;">KCS34155</td>
			<td style="width:297px;">Rearing pupae from extraction</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:193px;">Royal jelly</td>
			<td style="width:243px;">National Ilan University (NIU)</td>
			<td style="width:155px;">NIU</td>
			<td style="width:297px;">Make basic larval diet (BLD)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:193px;">D-(+)-Glucose</td>
			<td style="width:243px;">Sigma</td>
			<td style="width:155px;">G8270</td>
			<td style="width:297px;">Make basic larval diet (BLD)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:193px;">D-(-)-Fructose</td>
			<td style="width:243px;">Sigma</td>
			<td style="width:155px;">F0127</td>
			<td style="width:297px;">Make basic larval diet (BLD)</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:193px;">Yeast extract</td>
			<td style="width:243px;">CONDA, pronadisa</td>
			<td style="width:155px;">1702</td>
			<td style="width:297px;">Make basic larval diet (BLD)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:193px;">Sucrose</td>
			<td style="width:243px;">Taiwan sugar coporation</td>
			<td style="width:155px;">E01071010</td>
			<td style="width:297px;">Make sugar syrup for bee food</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:193px;">Pyriproxyfen (11%)</td>
			<td style="width:243px;">LIH-NUNG CHEMICAL CO.. LTD.</td>
			<td style="width:155px;">Registration No. 1937</td>
			<td style="width:297px;">Insect growth regulator (IGR) used in the experiment</td>
		</tr>
	</tbody>
</table>

evaluating the effect of environmental chemicals on honey bee development from the individual to colony level

The presence of pesticides in the beekeeping environment is one of the most serious problems that impacts the life of a honey bee. Pesticides can be brought back to the beehive after the bees have foraged on flowers that have been sprayed with pesticides. Pesticide contaminated food can be exchanged between workers which then feed larvae and therefore can potentially affect the development of honey bees. Thus, residual pesticides in the environment can become a chronic damaging factor to honey bee populations and gradually lead to colony collapse. In the presented protocol, honey bee feeding methods are described and applied to either an individual honey bee or to a colony. Here, the insect growth regulator (IGR) pyriproxyfen (PPN), which is widely used to control pest insects and is harmful to the development of honey bee larvae and pupae, is used as the pesticide. The presenting procedure can be applied to other potentially harmful chemicals or honeybee pathogens for further studies.

꿀벌 필드 시험, 여왕이 산란을위한 4 프레임 부에 한정되었다. 이 단계는 한 프레임에 무리 밀도를 증가시키고 이후의 관찰을 용이하게 할 수있다. 각각의 치료는 표시되었고, 꿀벌 &#39;개발은 분명 투명 슬라이드를 통해 관찰되었다. 벌집 꿀벌 유충에 PPN-BLD의 생체 먹이 정확하게 콜로니에 꿀벌의 개발 PPN의 영향을 평가하기 위해 수행 하였다. 생체 공급 법?...

Watch this Scientific Journal Video about Evaluating the Effect of Environmental Chemicals on Honey Bee Development from the Individual to Colony Level at JoVE.com

Evaluating the Effect of Environmental Chemicals on Honey Bee Development from the Individual to Colony Level

여기에서 우리는 제시 방법은 개별 꿀벌과 벌집 콜로니 모두 농약 오염 된 식품을 공급한다. 절차는 기본 유생 다이어트 생체 공급함으로써, 또한 벌집 콜로니의 자연 상태에서 개별 꿀벌의 살충제 효과를 평가한다.

식민지 수준에 개인에서 꿀 꿀벌 개발에 대한 환경 화학 물질의 효과를 평가

The presence of pesticides in the beekeeping environment is one of the most serious problems that impacts the life of a honey bee. Pesticides can be ...

evaluating-effect-environmental-chemicals-on-honey-bee-development

Research

JoVE Journal

Environment

9.0K 조회수.  National Ilan University.  여기에서 우리는 제시 방법은 개별 꿀벌과 벌집 콜로니 모두 농약 오염 된 식품을 공급한다. 절차는 기본 유생 다이어트 생체 공급함으로써, 또한 벌집 콜로니의 자연 상태에서 개별 꿀벌의 살충제 효과를 평가한다. 

비디오: Evaluating the Effect of Environmental Chemicals on Honey Bee Development from the Individual to Colony Level

Automated Gel Size Selection to Improve the Quality of Next-generation Sequencing Libraries Prepared from Environmental Water Samples

Next-generation sequencing of environmental samples can be challenging because of the variable DNA quantity and quality in these samples. High quality DNA libraries are needed for optimal results from next-generation sequencing. Environmental samples such as water may have low quality and quantities of DNA as well as contaminants that co-precipitate with DNA. The mechanical and enzymatic processes involved in extraction and library preparation may further damage the DNA. Gel size selection enables purification and recovery of DNA fragments of a defined size for sequencing applications. Nevertheless, this task is one of the most time-consuming steps in the DNA library preparation workflow. The protocol described here enables complete automation of agarose gel loading, electrophoretic analysis, and recovery of targeted DNA fragments. 
In this study, we describe a high-throughput approach to prepare high quality DNA libraries from freshwater samples that can be applied also to other environmental samples. We used an indirect approach to concentrate bacterial cells from environmental freshwater samples; DNA was extracted using a commercially available DNA extraction kit, and DNA libraries were prepared using a commercial transposon-based protocol. DNA fragments of 500 to 800 bp were gel size selected using Ranger Technology, an automated electrophoresis workstation. Sequencing of the size-selected DNA libraries demonstrated significant improvements to read length and quality of the sequencing reads.

This manuscript describes an automated gel size selection approach for purifying DNA fragments for next-generation sequencing. The Ranger Technology provides complete automation of the entire process of agarose gel loading, electrophoretic analysis, and recovery of targeted DNA fragments allowing for high-throughput and high quality next-generation sequencing libraries.

자동 젤 사이즈 선택은 환경 물 샘플에서 준비 차세대 시퀀싱 라이브러리의 질을 향상시키기 위해

Next-generation sequencing of environmental samples can be challenging because of the variable DNA quantity and quality in these samples. High quality DNA ...

연구

환경과학

Exploring the Effects of Atmospheric Forcings on Evaporation: Experimental Integration of the Atmospheric Boundary Layer and Shallow Subsurface

Evaporation is directly influenced by the interactions between the atmosphere, land surface and soil subsurface. This work aims to experimentally study evaporation under various surface boundary conditions to improve our current understanding and characterization of this multiphase phenomenon as well as to validate numerical heat and mass transfer theories that couple Navier-Stokes flow in the atmosphere and Darcian flow in the porous media. Experimental data were collected using a unique soil tank apparatus interfaced with a small climate controlled wind tunnel. The experimental apparatus was instrumented with a suite of state of the art sensor technologies for the continuous and autonomous collection of soil moisture, soil thermal properties, soil and air temperature, relative humidity, and wind speed. This experimental apparatus can be used to generate data under well controlled boundary conditions, allowing for better control and gathering of accurate data at scales of interest not feasible in the field. Induced airflow at several distinct wind speeds over the soil surface resulted in unique behavior of heat and mass transfer during the different evaporative stages.

A protocol for the design and construction of a soil tank interfaced to a small climate controlled wind tunnel to study the effects of atmospheric forcings on evaporation is presented. Both the soil tank and wind tunnel are instrumented with sensor technologies for the continuous in situ measurement of environmental conditions.

증발에 대기 강제력의 효과를 탐구 : 대기 경계층과 얕은 표면 하의의 실험 통합

Evaporation is directly influenced by the interactions between the atmosphere, land surface and soil subsurface. This work aims to experimentally study ...

Removal of Trace Elements by Cupric Oxide Nanoparticles from Uranium In Situ Recovery Bleed Water and Its Effect on Cell Viability

In situ recovery (ISR) is the predominant method of uranium extraction in the United States. During ISR, uranium is leached from an ore body and extracted through ion exchange. The resultant production bleed water (PBW) contains contaminants such as arsenic and other heavy metals. Samples of PBW from an active ISR uranium facility were treated with cupric oxide nanoparticles (CuO-NPs). CuO-NP treatment of PBW reduced priority contaminants, including arsenic, selenium, uranium, and vanadium. Untreated and CuO-NP treated PBW was used as the liquid component of the cell growth media and changes in viability were determined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay in human embryonic kidney (HEK 293) and human hepatocellular carcinoma (Hep G2) cells. CuO-NP treatment was associated with improved HEK and HEP cell viability. Limitations of this method include dilution of the PBW by growth media components and during osmolality adjustment as well as necessary pH adjustment. This method is limited in its wider context due to dilution effects and changes in the pH of the PBW which is traditionally slightly acidic&#160;however; this method could have a broader use assessing CuO-NP treatment in more neutral waters.

Removal of Trace Elements by Cupric Oxide Nanoparticles from Uranium In Situ Recovery Bleed Water and Its Effect on Cell Viability

Production bleed water (PBW) was treated with cupric oxide nanoparticles (CuO-NPs) and cellular toxicity was assessed in cultured human cells. The goal of this protocol was to integrate the native environmental sample into a cell culture format assessing the changes in toxicity due to CuO-NP treatment.

우라늄에서 구리 산화물 나노 입자에 의한 미량 원소의 제거<em&gt; 현장에서</em&gt; 복구 블리드 물과 세포 생존에 미치는 영향

In situ recovery (ISR) is the predominant method of uranium extraction in the United States. During ISR, uranium is leached from an ore body and extracted ...

A Flow-through Exposure System for Evaluating Suspended Sediments Effects on Aquatic Life

This paper describes the Fish Larvae and Egg Exposure System (FLEES). The flow-through exposure system is used to investigate the effects of suspended sediment on various aquatic species and life stages in the laboratory by using pumps and automating delivery of sediment and water to simulate suspension of sediment. FLEES data are used to develop exposure-response curves between the effects on aquatic organisms and suspended sediment concentrations at the desired exposure duration. The effects data are used to evaluate management practices used to reduce the interactions between aquatic organisms and anthropogenic causes of suspended sediments. The FLEES is capable of generating total suspended solids (TSS) concentrations as low as 30 to as high as 800 mg/L, making this system an ideal choice for evaluating the effects of TSS resulting from many activities including simulating low ambient levels of TSS to evaluating sources of suspended sediments from dredging operations, vessel traffic, freshets, and storms.

A robust and flexible flow-through exposure system designed to maintain sediment in suspension is presented. The system is used to investigate the effects of suspended sediment on various aquatic species and life stages in the laboratory.

수산 생명에 일시 중지를 평가하면 퇴적물 효과에 대한 플로우를 통해 노출 시스템

This paper describes the Fish Larvae and Egg Exposure System (FLEES). The flow-through exposure system is used to investigate the effects of suspended ...

Extraction of Organochlorine Pesticides from Plastic Pellets and Plastic Type Analysis

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment during manufacturing and transport. Because of their environmental persistence, they are widely distributed in the oceans and on beaches all over the world. They can act as a vector of potentially toxic organic compounds (e.g., polychlorinated biphenyls) and might consequently negatively affect marine organisms. Their possible impacts along the food chain are not yet well understood. In order to assess the hazards associated with the occurrence of plastic pellets in the marine environment, it is necessary to develop methodologies that allow for rapid determination of associated organic contaminant levels. The present protocol describes the different steps required for sampling resin pellets, analyzing adsorbed organochlorine pesticides (OCPs) and identifying the plastic type. The focus is on the extraction of OCPs from plastic pellets by means of a pressurized fluid extractor (PFE) and on the polymer chemical analysis applying Fourier Transform-InfraRed (FT-IR) spectroscopy. The developed methodology focuses on 11 OCPs and related compounds, including dichlorodiphenyltrichloroethane (DDT) and its two main metabolites, lindane and two production isomers, as well as the two biologically active isomers of technical endosulfan. This protocol constitutes a simple and rapid alternative to existing methodology for evaluating the concentration of organic contaminants adsorbed on plastic pieces.

Microplastics act as vector of potentially toxic organic contaminants with unpredictable effects. This protocol describes an alternative methodology for assessing the levels of organochlorine pesticides adsorbed on plastic pellets and identifying the polymer chemical structure. The focus is on pressurized fluid extraction and attenuated total reflectance Fourier transform infrared spectroscopy.

플라스틱 펠렛 및 플라스틱 유형 분석에서 유기 염소 농약의 추출

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment ...

Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation

Dimethylated thioarsenicals such as dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV), which are produced by the metabolic pathway of dimethylarsinic acid (DMAV) thiolation, have been recently found in the environment as well as human organs. DMMTAV and DMDTAV can be quantified to determine the ecological effects of dimethylated thioarsenicals and their stability in environmental media. The synthesis method for these compounds is unstandardized, making replicating previous studies challenging. Furthermore, there is a lack of information about storage techniques, including storage of compounds without species transformation. Moreover, because only limited information about synthesis methods is available, there may be experimental difficulties in synthesizing standard chemicals and performing quantitative analysis. The protocol presented herein provides a practically modified synthesis method for the dimethylated thioarsenicals, DMMTAV and DMDTAV, and will help in the quantification of species separation analysis using high performance liquid chromatography in conjunction with inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The experimental steps of this procedure were modified by focusing on the preparation of chemical reagents, filtration methods, and storage.

Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation

This article presents modified experimental protocols for dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV) synthesis, inducing dimethylarsinic acid (DMAV) thiolation through mixing of DMAV, Na2S, and H2SO4. The modified protocol provides an experimental guideline, thereby overcoming limitations of the synthesis steps that could have caused experimental failures in quantitative analysis.

DMMTAV 및 환경 응용 프로그램에 대 한 DMAV 를 사용 하 여 DMDTAV 의 준비: 합성, 정화, 및 확인

Dimethylated thioarsenicals such as dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV), which are produced by the metabolic ...

The Effect of the Application of Thyme Essential Oil on Microbial Load During Meat Drying

Meat is a high protein meal that is used in the preparation of jerky, a popular food snack, where preservation and safety are important. To assure food safety and to extend the shelf life of meat and meat products, the use of either synthetic or natural preservatives have been applied to control and eliminate foodborne bacteria. A growing interest in the application of natural food additives for meat has increased. Microorganisms, such as Escherichia coli, contaminate meat and meat products, causing foodborne illnesses. Therefore, it is necessary to improve the meat conservation process. However, the use of essential oils when the meat is being dried has not been deeply studied. In this regard, there is an opportunity to increase the value of dried meat and reduce the risk of foodborne illnesses by applying essential oils during the drying process. In this protocol, we present a novel method of applying thyme essential oil (TEO) during meat drying, specifically in vapor form directly in a drying chamber. For evaluation, we use Minimal Inhibitory Concentration (MIC) to detect the number of harmful bacteria in the treated samples compared to raw samples. The preliminary results show that this method is a viable and alternative option to synthetic preservatives and that it significantly reduces microbial load in dried meat.

Microorganisms such as Escherichia coli that contaminate meat products cause foodborne illnesses. The use of essential oils in the meat drying process has not been deeply studied. Here, we present a novel method of applying thyme essential oil to meat during drying to reduce the microbial load in dried meat.

고기 건조 하는 동안 미생물 부하에 백 리 향 에센셜 오일의 응용 프로그램의 효과

Meat is a high protein meal that is used in the preparation of jerky, a popular food snack, where preservation and safety are important. To assure food ...

Protocol for Acute and Chronic Ecotoxicity Testing of the Turquoise Killifish Nothobranchius furzeri

The killifish Nothobranchius furzeri is an emerging model organism in the field of ecotoxicology and its applicability in acute and chronic ecotoxicity testing has been demonstrated. Overall, the sensitivity of the species to toxic compounds is in the range with, or higher than, that of other model species.
This work describes protocols for acute, chronic, and multigenerational bioassays of single and combined stressor effects on N. furzeri. Due to its short maturation time and life-cycle, this vertebrate model allows the study of endpoints such as maturation time and fecundity within four months. Transgenerational full life-cycle exposure trials can be performed in as little as 8 months. Since this species produces eggs that are drought-resistant and remain viable for years, the on-site culture of the species is not needed but individuals can be recruited when required. The protocols are designed to measure life-history traits (mortality, growth, fecundity, weight) and critical thermal maximum.

Protocol for Acute and Chronic Ecotoxicity Testing of the Turquoise Killifish Nothobranchius furzeri

In this work, we describe an acute, chronic and multigenerational bioassay to study the effects of single and combined stressors on the Turquoise killifish Nothobranchius furzeri. This protocol is designed to study life-history traits (mortality, growth, fecundity, weight) and critical thermal maximum.

급성 및 만성 Ecotoxicity 청록색 송사리 Nothobranchius furzeri 의 테스트에 대 한 프로토콜

The killifish Nothobranchius furzeri is an emerging model organism in the field of ecotoxicology and its applicability in acute and chronic ecotoxicity ...

Measuring Hypopharyngeal Gland Acinus Size in Honey Bee (Apis mellifera) Workers

The nurse hypopharyngeal glands produce the protein fraction of the worker and royal jelly that is fed to developing larvae and queens. These paired glands that are located in the head of the bee are highly sensitive to the quantity and quality of pollen and pollen substitutes that the nurse bee consumes. The glands get smaller when nurses are fed deficient diets and are large when they are fed complete diets. Because nurse hypopharyngeal gland size is a robust indicator of nurse nutrition, it is essential that those studying honey bee nutrition know how to measure these glands. Here, we provide detailed methods for dissecting, staining, imaging, and measuring nurse bee hypopharyngeal glands. We present comparisons of unstained and stained tissue and data that were used to study the impact of pollen on gland size. This method has been used to test how diet impacts hypopharyngeal gland size but has further use for understanding the role of these glands in hive health.

Measuring Hypopharyngeal Gland Acinus Size in Honey Bee Apis mellifera Workers

Hypopharyngeal gland acinus size is a robust measure of nurse honey bee nutrition. Here, we provide a detailed protocol for dissecting, staining, imaging, and measuring nurse bee hypopharyngeal gland acini.

꿀 꿀벌 (Apis mellifera) 노동자의 Hypopharyngeal 동맥 Acinus 크기 측정

The nurse hypopharyngeal glands produce the protein fraction of the worker and royal jelly that is fed to developing larvae and queens. These paired ...

Composition and Distribution Analysis of Bioaerosols Under Different Environmental Conditions

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, we described a protocol for multiple analyses of the biological compositions in environmental PM. Five experiments are presented: (1) PM number monitoring by using a laser particle counter; (2) PM collection by using a cyclonic aerosol sampler; (3) PM collection by using a high-volume air sampler with filters; (4) culturable microbes collection by the Andersen six-stage sampler; and (5) detection of biological composition of environmental PM by bacterial 16SrDNA and fungal ITS region sequencing. We selected hazy days and a livestock farm as two typical examples of the application in this protocol. In this study, these two sampling methods, cyclonic aerosol sampler and filter sampler, showed different sampling efficiency. The cyclonic aerosol sampler performed much better in terms of collecting bacteria, while these two methods showed the same efficiency in collecting fungi. Filter samplers can work under low temperature conditions while cyclonic aerosol samplers have a sampling limitation for temperature. A solid impacting sampler, such as an Andersen six-stage sampler, can be used to sample bioaerosols directly into the culture medium, which increases the survival rate of culturable microorganisms. However, this method mainly relies on culture while more than 99% of microbes cannot be cultured. DNA extracted from the culturable bacteria collected by the Andersen six-stage sampler and samples collected by cyclonic aerosol sampler and filter sampler were detected by bacterial 16S rDNA and fungal ITS region sequencing.All the methods above may have wide application in many fields of study, such as environmental monitoring and airborne pathogen detection. From these results, we can conclude that these methods can be used under different conditions and may help other researchers further explore the health impacts of environmental bioaerosols.

Understanding the biological composition of environmental particulate matter is important for the study of its significant impacts on human health and disease spread. Here, we used three types of bioaerosol sampling methods and a biological analysis of airborne microbes to better explore airborne microbial communities under different environmental conditions.

구성 및 다른 환경 조건에서 Bioaerosols의 유통 분석

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, ...