Name: the hawaii protocol for scientific monitoring of coffee berry borer: a model for coffee agroecosystems worldwide
Uploaded: 2018-03-19T12:30:00.000+00:00
Duration: 14 min 29 s
Description: Watch this Scientific Journal Video about The Hawaii Protocol for Scientific Monitoring of Coffee Berry Borer: a Model for Coffee Agroecosystems Worldwide at JoVE.com

我们感谢森林布雷默提供咖啡场的无人机图像, 以及协助使用地理信息系统方法。我们感谢托马斯 Mangine, 马修·米勒, 琳赛汉密尔顿, 香农威尔逊, 布瑞娜麦卡锡和 Mehana Sabado Halpern 协助电影制作, 和两名匿名评论者对先前的草稿发表意见。这项工作是由美国农业部资助的。在本出版物中发表的意见、发现、结论或建议是作者的观点, 并不一定反映美国农业部的观点。美国农业部是一个平等机会提供者和雇主。

注意: 该协议的西班牙文翻译作为<a href="/files/ftp_upload/57204/Supplementary_file_1-Spanish_Translation_of_the_protocol.docx" target="_blank">补充文件 1</a>提供。
1. 在咖啡场内定义取样区
<ol><li>通过全球定位系统 (GPS) 仪器对咖啡场的周长进行监测。将字段坐标导入到全球信息系统 (GIS) 中, 并生成咖啡场的地图。</li>
	<li>将字段划分为 "区域" (即、多边形), 每335米2。这些将用于确保整个领域的系统随机抽样设计。</li>
</ol>2. 在电子系统中创建数据收集应用程序
<ol><li>使用电子数据收集平台, 构建由以下相互关联的数据库组成的数据收集应用程序:陷阱、区域、站点服务、气象站、和管理。 注意: 这些数据库将在协议的所有后续步骤中用于数据的收集和组织。</li>
	<li>对于补漏白数据库, 创建 "站点名称"、"陷阱编号"、"部署日期"、"字段技术人员名称"、"部署照片" 等字段, 以及指向每个陷阱的 GPS 坐标的链接。</li>
	<li>对于区域数据库, 创建 "站点名称" 和 "区域编号" 字段, 并链接到显示每个区域的地理站点地图。</li>
	<li>对于站点服务数据库, 创建 "站点名称"、"日期"、"字段技术人员名称" 和 "站点备注" 字段。在站点服务数据库中, 创建包含以下内容的嵌套数据库。<ol>
<li>包括补漏白服务以记录陷印号码 (在父陷阱数据库中有指向相关陷阱部署记录的链接)、捕获陷阱捕获的照片和陷阱计数。</li>
		<li>包括区域服务记录物候照片、浆果出没评估 (绿色浆果的总数量、被感染的绿色浆果、带有B. 白僵菌的绿色浆果和葡萄干), 以及指向父级相关区域记录的链接。区域数据库。该记录还包括每个取样树的 GPS 坐标。</li>
		<li>包括气象站服务以记录站点名称、日期、数据下载和电池检查。</li>
		<li>包括浆果解剖记录实验室技术员姓名、日期和 CBB 位置 (AB 或 CD) 和死亡率类别 (活着, 死于其他原因, 或死于白僵菌) 为每个解剖浆果。</li>
	</ol>
</li>
	<li>对于气象站数据库, 创建 "站点名称"、"站号"、"部署日期"、"字段技术员名称"、"部署照片" 等字段, 以及指向每个气象站的 GPS 坐标的链接。</li>
	<li>对于管理数据库, 创建 "站点名称"、"日期" 和 "管理实践类型" 的字段。</li>
</ol>3. 准备和部署用于监测 CBB 运动的陷阱
<ol><li>确定监视每个字段中 CBB 移动所需的陷阱数。 注: 每个字段的陷印密度应近似为大字段 (~ 1 公顷)15的小字段 (~ 0.5 公顷) 和10个陷阱的5个陷阱。</li>
	<li>使用图钉, 在每个陷阱收集杯的填充线上方做一系列排水孔, 以避免雨水稀释杀死溶液。根据制造商的指示装配漏斗圈。</li>
	<li>制备 1 L 的杀伤溶液, 由200毫升丙烯乙二醇和800毫升水组成。接下来, 准备一个由3:1 甲醇溶液组成的引诱剂混合物: 乙醇。将40毫升的引诱剂倒入塑料半透水袋 (2 毫米, 3 英寸 x 4 英寸), 并放置在集装箱中运输。 注: 在吸引 CBB 时, 半透水袋的表现比开瓶效果好, 而且由于洗脱率降低16, 还需要较少频繁地对补强诱饵进行访问。 注意: 甲醇和乙醇是高度易燃的液体, 如果吸入或摄取, 是有毒的, 并且是皮肤和眼睛刺激物。这些化学品应在通风良好的房间里处理, 同时戴上手套、护眼和防护服。</li>
	<li>通过在字段中随机分布它们来部署陷阱。放置陷阱 0.5-1.5 米以上的地面, 并清除过道。木桩可以用来有效地保护树木之间的陷阱。将站点名称和陷阱编号与每个陷阱上的永久性标记一起写入将来的标识。</li>
	<li>填充陷阱收集杯与100毫升的乙二醇杀死解决方案, 并拧紧的杯子到位。将回形针贴在每个引诱袋上, 并用回形针将袋子钩到陷阱的中心。</li>
	<li>使用配备了电子数据收集平台的移动设备, 导航到陷阱数据库, 并创建由站点、日期、陷阱号和陷阱照片组成的新部署记录。 注意: 每个字段中陷阱的位置是通过移动设备上的 GPS 自动记录的。</li>
</ol>4. 服务陷阱
<ol><li>到达该字段后, 导航到电子系统中的站点服务数据库, 并创建一个由站点名称、日期和字段技术人员名称组成的新的站点服务记录。 注: 初始陷阱服务在陷阱部署后两周进行, 此后每两周进行一次。如果需要更高分辨率的陷阱捕获数据, 则可以进行每周的陷阱服务, 尽管我们注意到双周取样足以捕获整个季节的一般运动趋势 (图 1)。</li>
	<li>在字段中找到陷阱。将细网手筛 (网格尺寸 0.8-1.0 毫米) 放在塑料容器上, 然后从收集杯中将杀死溶液倒入筛中。把杀死的解决方案转移回收集杯和周围的液体, 以确保所有的 CBB 从收集杯中删除。</li>
	<li>在新的站点服务记录中, 导航到补漏白服务数据库并创建新的陷阱服务记录。输入相关的陷阱编号, 并在背景中用站点名称和陷阱编号拍摄筛网。将照片保存到陷阱服务记录。</li>
	<li>用勺子或金属铲, 把所有的昆虫舀进装满70% 乙醇的瓶子里。用站点、日期和陷阱编号标记瓶子。</li>
	<li>用新鲜的杀死溶液重新填充收集杯, 然后拧回陷阱上。每月一次用肥皂水清洗收集杯, 冲洗, 并用新鲜的杀液代替。每月或根据需要更换引诱剂和包袋。</li>
</ol>5. 植物物候服务区
<ol><li>在站点服务记录中, 导航到区域服务数据库并创建一个新的区域服务记录。从链接的区域数据库的站点地图中选择取样区域。</li>
	<li>为了避免取样偏差, 从区域内随机选取一棵树, 将眼睛向下投射, 这样只有树的底座可见。站在选定的树的前面, 随机选择一个侧面分支围绕胸部高度。将标尺夹在所选分支上, 确保标尺不会从照相机的视野中阻挡任何生殖部分 (节点、芽、花、果)。</li>
	<li>拍摄一张照片, 确保目标分支的标尺和整体可见。拍下整棵树的第二张照片;尽量在照片中获得尽可能多的中层树冠。将这两种物候照片保存到区域服务记录中。</li>
</ol>6. 绿色浆果损害评估服务区
<ol><li>如果用于物候的分支似乎有 > 30 绿色浆果, 请计算树枝上至少豌豆大小 (~ 0.6 厘米) 和较大的浆果的数量, 并以绿色的颜色 (BBCH 鳞片 77-8517) 来表示。在区域服务记录中输入此数字。 注: 如果用于物候的分支似乎有 < 30 绿色浆果, 随意选择一个侧面分支在约胸部高度从一个树在目标区域与 > 30 绿色浆果可见。从远处做这一点, 以避免选择中的偏差。</li>
	<li>此外, 在区域服务记录中, 输入分支上 CBB 所感染的绿色浆果的数量。被感染的浆果会有一个小孔, 通常位于中央圆盘;CBB 在洞中可能或可能不可见。</li>
	<li>输入具有可见白色白僵菌真菌的绿色被感染浆果的数量。真菌可以看到在 CBB 和/或周围的入口孔。 注意: 如果有特殊的兴趣, 可能需要进一步的化验来鉴定真菌种类。</li>
	<li>输入树枝上的葡萄干 (干果) 的数量。此信息可用于了解管理实践 (例如、剥离拾取) 和 CBB 侵扰之间的关系。</li>
	<li>从树枝上收集三受感染的绿色浆果;这些将被带回实验室和解剖, 以评估 CBB 在浆果内的位置。 注: 如果用于损害评估的分支有 < 3 个受感染的绿色浆果, 则可以从区域内的其他分支获取绿色的浆果。</li>
	<li>放置在塑料容器中的绿色浆果和标签与网站和日期。把容器放在冰上的冷却器里, 直到它们可以运回实验室。 注: 理想情况下, 浆果应在收集 1-3 天内进行解剖, 以确保 CBB 的最大存活率。<ol>
<li>储存浆果 (根据需要) 在实验室14°c 三天, 几乎没有死亡 (s Fortna & r 霍宁沃思, 个人沟通)。</li>
	</ol>
</li>
	<li>对每个取样区的物候和浆果损伤评估重复步骤。 注: 应为大型养殖场 (~ 1 公顷) 取样约25个分支机构, 并向小农场取样15个分支机构 (~ 0.5 公顷)。对于解剖, 应从大型养殖场收集75种绿色的浆果, 并在每个取样日期从小农场中抽取45个。在今年的某些地方, 可能无法收集到这些浆果的数量。在这种情况下, 尽量收集50绿色浆果为大农场 (~ 1 公顷) 和30绿色浆果为小农场 (~ 0.5 公顷)。</li>
</ol>7. 计算每个陷阱中的 CBB 数
<ol><li>将粗网手筛 (网格尺寸 ~ 1.5 毫米) 放在塑料容器上, 将甲虫从收集瓶中清空到筛子中。用一个装满水的洗瓶把瓶子里的所有东西都拿出来。</li>
	<li>使用洗瓶喷雾的内容在筛, 迫使尽可能多的小昆虫通过筛尽可能。这允许更大的昆虫和碎片从样本中的小甲虫中分离出来, 并限制 CBB 的体积估计不准确。扔掉大的昆虫和碎屑, 冲洗筛子。</li>
	<li>在第二个塑料容器上放置细网手筛 (网格尺寸 ~ 1.0 毫米), 然后将第一个容器的内容倒入细网手筛。</li>
	<li>如果有超过几个百 CBB, 跳到步骤7.6。如果有少于几个百 CBB, 把细网筛放在纸巾上, 以除去多余的水。把筛子倒过来, 把所有的东西都放到一个透明的塑料盖子上。如果它们聚集在一起, 用画笔把甲虫围起来, 让它们坐到干。</li>
	<li>如果有少于几个百 CBB, 使用一个细尖的画笔或类似的实施, 以线甲虫成行, 是几个甲虫宽, 并开始计数在光显微镜下。计算甲虫的总数, 并分成 "CBB" 和 "其他" 类别。</li>
	<li>如果有超过几个百 CBB, 将 CBB 从细筛筛到一个10毫升的注射器使用金属铲。将喷射器柱放进注射器, 轻轻向下按压, 直到感觉到轻微的阻力, 同时小心不要粉碎甲虫。记录注射器上的容积值。</li>
	<li>使用上面描述的协议从容积样本中计数200甲虫。使用下面的等式确定 CBB 与样品中其他甲虫的数量。<ol>
<li>使用以下方法估计 CBB 计数: 样本的总 CBB 估计数 = (# CBB ÷ 200) x ∂ x (用注射器测量的 mL)。 注: 这里∂ = 昆虫/毫升的数量。建议对每个区域进行∂估计;在夏威夷海岛价值1033年被测量了。</li>
		<li>使用以下方法估计其他计数: 总 "其他" 甲虫估计为样品 = (# 其他÷ 200) x ∂ x (毫升测量在注射器)。</li>
	</ol>
</li>
	<li>当陷阱计数完成后, 导航到相关的补漏白服务记录并输入 CBB 和其他甲虫的数量。</li>
</ol>8. 物候照片的评分
<ol><li>从数据收集应用程序中导出咖啡物候照片。打开照片, 用附加的尺子定位树枝。为这个分支得分以下。<ol>
<li>将节点数 (叶子的附着点) 分到分支。</li>
		<li>不成熟的芽, 成熟的芽, 蜡烛, 开花, 和别针头的存在或缺席。</li>
		<li>分数的豌豆大小的绿色浆果, 未成熟的绿色浆果, 成熟的绿色浆果, 浆果显示的颜色打破, 完全成熟的浆果, 和葡萄干。</li>
	</ol>
</li>
</ol>9. 解剖浆果以确定 CBB 位置
<ol><li>将被感染的绿色浆果从冷藏库中取出, 让它们在进行浆果解剖之前, 在室温下加热10-15 分钟。这个恢复时间是重要的, 以便 CBB 可以准确地评估为活着或死亡。 注: 对被感染的浆果进行解剖, 可以确定成人 CBB 的位置。位置 AB 表明, 雌性已开始渗透到浆果, 但尚未达到胚乳;位置 CD 指示女性已进入胚乳13。</li>
	<li>使用手术刀或类似的实施, 使切片通过浆果平行的中央圆盘作为一个初步评估甲虫的位置。接下来, 制作一系列垂直于中央圆盘和入口孔周围的浅片, 以确定 CBB 是否在 AB 或 CD 位置。</li>
	<li>将 AB 和 CD 类别细分为 "活的", "死于白僵菌", "死于其他原因", 和 "甲虫失踪"。如果不清楚成年人是活着还是死了, 那就放大显微镜, 观察腿部的运动。</li>
	<li>地方计数的人入菜与水或酒精。这有助于跟踪已计算的内容, 并防止成年甲虫逃到实验室。</li>
	<li>完成站点的解剖后, 导航到相关站点服务记录中的浆果解剖数据库, 并输入每个类别中的 CBB 总数。</li>
	<li>将解剖样品放置在容器中, 在处理前冻结72小时。</li>
</ol>10. 服务手册气象站
注: 需要手动数据下载的气象站可以每周或每月提供服务, 以下载数据并确保所有传感器工作正常。天气变量, 重要的是要考虑了解 CBB 生物学可能包括降雨, 湿度, 空气和土壤温度, 太阳辐射, 光合主动辐射 (PAR), 土壤湿度, 风速/方向。
<ol><li>在现场找到手动气象站。在电子系统中, 打开相关的站点服务记录, 然后导航到气象站服务数据库。创建连接到相关气象站部署记录的新气象站服务记录。</li>
	<li>使用防水穿梭机直接连接数据记录器和笔记本电脑进行数据下载。在气象站服务记录中记下数据的下载情况。</li>
	<li>一旦数据被关闭, 手动重新启动太阳和温度/湿度记录器, 以确保他们有正确的设置 (降雨记录器不需要重新启动)。检查电池电量并根据需要更换。在电子系统中记下这一点。</li>
	<li>返回实验室后, 将最新的数据添加到完整的天气缝中, 并更新元数据记录。</li>
</ol>11. 记录管理做法
注: 有关管理做法的信息可用于了解 CBB 活动和人口规模的模式。相关的管理做法可能包括 (但不限于): 喷洒白僵菌真菌, 喷洒 pyrethins 或其他杀虫剂, 修剪, 杂草管理, 带采摘, 樱桃采摘, 从地面去除葡萄干,等。
<ol><li>在管理数据库中, 创建一个新的管理记录, 其中提供了网站名称、日期和管理实践的类型。</li>
</ol>

<ol>
	<li>Jaramillo, J., Borgemeister, C., Baker, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coffee+berry+borer+Hypothenemus+hampei+(Coleoptera:+Curculionidae):+searching+for+sustainable+control+strategies.">Coffee berry borer Hypothenemus hampei (Coleoptera: Curculionidae): searching for sustainable control strategies.</a> Bull. Entomol. Res. 96, 223-233 (2006).</li><li>Vega, F. E., Infante, F., Johnson, A. J., Vega, F. E., Hofstetter, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genus+Hypothenemus,+with+emphasis+on+H.+hampei,+coffee+berry+borer.">The genus Hypothenemus, with emphasis on H. hampei, coffee berry borer.</a> Bark beetles, biology and ecology of native and invasive species . 11, 427-494 (2015).</li><li>ISO. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Green+coffee+-Defect+reference+chart.">Green coffee -Defect reference chart.</a> International Standard ISO 10470. , 15 (2004).</li><li>Wegbe, K., Cilias, C., Decazy, B., Alauzet, C., Dufour, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estimation+of+production+losses+caused+by+the+coffee+berry+borer+(Coleoptera:+Scolytidae)+and+calculation+of+an+economic+damage+threshold+in+Togolese+coffee+plots.">Estimation of production losses caused by the coffee berry borer (Coleoptera: Scolytidae) and calculation of an economic damage threshold in Togolese coffee plots.</a> J. Econ. Entomol. 96, 1473-1478 (2003).</li><li>Burbano, E., Wright, M., Bright, D. E., Vega, F. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+record+for+the+coffee+berry+borer,+Hypothenemus+hampei,+in+Hawaii.">New record for the coffee berry borer, Hypothenemus hampei, in Hawaii.</a> J. Insect Sci. 11 (1), 117 (2011).</li><li>Kinro, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> A Cup of Aloha: The Kona Coffee Epic. , (2003).</li><li>Teuber, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Geographical+indications+of+origin+as+a+tool+of+product+differentiation:+The+case+of+coffee.">Geographical indications of origin as a tool of product differentiation: The case of coffee.</a> J. Int. Food Agribus. Mark. 22 (3-4), 277-298 (2010).</li><li>Leung, P. S., Kawabata, A. M., Nakamoto, S. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Estimated economy-wide impact of CBB for the crop years 2011/12 and 2012/13. Brief report at request of Hawaii Congressional Delegation. , 2 (2014).</li><li>Baker, P. S., Jackson, J. A. F., Murphy, S. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Natural Enemies, natural allies. Project completion report of the integrated management of coffee berry borer project, CFC/ICO/02 (1998-2002). , (2002).</li><li>Brun, L. O., Marcillaud, C., Gaudichon, V., Suckling, D. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endosulfan+resistance+in+Hypothenemus+hampei+(Coleoptera:+Scolytidae)+in+New+Caledonia.">Endosulfan resistance in Hypothenemus hampei (Coleoptera: Scolytidae) in New Caledonia.</a> J. Econ. Entomol. 82, 1311-1316 (1989).</li><li>Vega, F. E., Infante, F., Castillo, A., Jaramillo, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+coffee+berry+borer,+Hypothenemus+hampei+(Ferrari)+(Coleoptera:+Curculionidae):+a+short+review,+with+recent+findings+and+future+research+directions.">The coffee berry borer, Hypothenemus hampei (Ferrari) (Coleoptera: Curculionidae): a short review, with recent findings and future research directions.</a> Terr. Arthropod Rev. 2, 129-147 (2009).</li><li>Aristiz&#225;bal, L. F., Johnson, M., Shriner, S., Hollingsworth, R., Manoukis, N. C., Myers, R., Bayman, P., Arthurs, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Integrated+pest+management+of+coffee+berry+borer+in+Hawaii+and+Puerto+Rico:+Current+status+and+prospects.">Integrated pest management of coffee berry borer in Hawaii and Puerto Rico: Current status and prospects.</a> Insects. 8, 123 (2017).</li><li>Bustillo, A. E., Cardenas, M. R., Villalba, D., Orozco, J., Benavides, M. P., Posada, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Manejo integrado de la broca del caf&#233; Hypothenemus hampei(Ferrari) en Colombia. , 134 (1998).</li><li>Kawabata, A. M., Nakamoto, S. T., Curtiss, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recommendations+for+Coffee+Berry+Borer+Integrated+Pest+Management+in+Hawai'i+2016.">Recommendations for Coffee Berry Borer Integrated Pest Management in Hawai'i 2016.</a> Insect Pests. IP-41, (2017).</li><li>Aristiz&#225;bal, L. F., Jim&#233;nez, M., Bustillo, A. E., Trujillo, H. I., Arthurs, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monitoring+coffee+berry+borer,+Hypothenemus+hampei+(Coleoptera:+Curculionidae),+populations+with+alcohol+baited+funnel+traps+in+coffee+farms+in+Colombia.">Monitoring coffee berry borer, Hypothenemus hampei (Coleoptera: Curculionidae), populations with alcohol baited funnel traps in coffee farms in Colombia.</a> Fla. Entomol. 98 (1), 381-383 (2015).</li><li>Messing, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+coffee+berry+borer+(Hypothenemus+hampei)+invades+Hawaii:+Preliminary+investigations+on+trap+responses+and+alternate+hosts.">The coffee berry borer (Hypothenemus hampei) invades Hawaii: Preliminary investigations on trap responses and alternate hosts.</a> Insects. 3 (1), 640-652 (2012).</li><li>Arcila-Pulgar&#237;n, J., Buhr, L., Bleiholder, H., Hack, H., Meier, U., Wicke, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Application+of+the+extended+BBCH+scale+for+the+description+of+the+growth+stages+of+coffee+(Coffea+spp).">Application of the extended BBCH scale for the description of the growth stages of coffee (Coffea spp).</a> Ann. Appl. Biol. 141 (1), 19-27 (2002).</li><li>Mathieu, F., Brun, L. O., Fr&#233;rot, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+related+with+native+host+abandonment+by+the+Coffee+Berry+Borer+Hypothenemus+hampei+(Ferrari)+(Coleoptera:+Scolytidae).">Factors related with native host abandonment by the Coffee Berry Borer Hypothenemus hampei (Ferrari) (Coleoptera: Scolytidae).</a> J. Appl. Entomol. 121, 175-180 (1997).</li><li>Pereira, A. E., Vilela, E. F., Tinoco, R. S., de Lima, J. O. G., Fantine, A. K., Morais, E. G. F., Franca, C. F. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Correlation+between+numbers+captured+and+infestation+levels+of+the+Coffee+Berry-borer,+Hypothenemus+hampei:+A+preliminary+basis+for+an+action+threshold+using+baited+traps.">Correlation between numbers captured and infestation levels of the Coffee Berry-borer, Hypothenemus hampei: A preliminary basis for an action threshold using baited traps.</a> Int. J. Pest. Manage. 58 (2), 183-190 (2012).</li><li>Baker, P. S., Ley, C., Balbuena, R., Barrera, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+affecting+the+emergence+of+Hypothenemus+hampei+(Coleoptera:+Scolytidae)+from+coffee+berries.">Factors affecting the emergence of Hypothenemus hampei (Coleoptera: Scolytidae) from coffee berries.</a> Bull. Entomol. Res. 82, 145-150 (1992).</li><li>Dufour, B. P., Fr&#233;rot, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimization+of+coffee+berry+borer,+Hypothenemus+hampei+Ferrari+(Col.,+Scolytidae),+mass+trapping+with+an+attractant+mixture.">Optimization of coffee berry borer, Hypothenemus hampei Ferrari (Col., Scolytidae), mass trapping with an attractant mixture.</a> J. Appl. Entomol. 132, 591-600 (2008).</li><li>Aristiz&#225;bal, L. F., Shriner, S., Hollingsworth, R., Arthurs, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flight+activity+and+field+infestation+relationships+for+coffee+berry+borer+in+commercial+coffee+plantations+in+Kona+and+Ka'u+districts,+Hawaii.">Flight activity and field infestation relationships for coffee berry borer in commercial coffee plantations in Kona and Ka'u districts, Hawaii.</a> J. Econ. Entomol. 110 (6), 2421-2427 (2017).</li><li>Gaertner, J., Genovese, V. B., Potter, C., Sewake, K., Manoukis, N. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vegetation+classification+of+Coffea+on+Hawaii+Island+using+Worldview-2+satellite+imagery.">Vegetation classification of Coffea on Hawaii Island using Worldview-2 satellite imagery.</a> J. App. Remote Sensing. 11, 046005 (2017).</li></ol>

我们没有要报告的利益冲突。

这里描述的监测协议可以作为研究 CBB 和控制策略的重要组成部分, 对这种侵袭性咖啡害虫。我们已经在夏威夷岛2016和2017的咖啡种植季节实施了这一监测协议, 旨在优化本文所概述的过程的每一步以及随附的视频。通过这样做, 我们确保对 CBB 人口动态的重要方面进行了监测和量化, 为议定书的每一步确定了最有效的低成本材料, 并收集了关于 CBB 运动的数据,虫害, 死亡率, 咖啡植物物候, 天气和农场管理可以用来通知和改善目前的控制策略。
该协议中有许多关键步骤必须遵循以确保最佳结果。首先, 漏斗陷阱必须设置在一个统一的高度和位置之间的树木。这将确保引诱剂充分扩散到空气中, 甲虫可以从四面八方进入陷阱。第二, 在整个监测期间, 必须使用相同网格尺寸 (粗网筛≈1.5 毫米和细网筛≈1.0 毫米) 的筛, 以确保 CBB 容积估计的一致结果。第三, CBB 与其他甲虫在每个陷阱中的比例在陷阱和生长季节之间会有很大差异, 因此有必要估计这些比例, 以尽量减少陷阱计数数据中的噪声。第四, 被感染的浆果必须储存在冰上的冷却器上, 直到它们运到实验室, 之后浆果应该保存在14摄氏度, 直到解剖。在潮湿环境中的存储将导致从浆果20CBB 出现。最后, 解剖必须在收集 1-3 天内进行, 以确保 CBB 的最大存活率。如果浆果在寒冷的温度下长时间储存, CBB 的死亡率可能会发生。
在此处不包括的研究倡议可能需要其他步骤 (例如, 监视 CBB 捕食者丰度)。如果时间、资源和/或设备是限制因素, 也可以对该协议进行修改。陷阱引诱剂由3:1 甲醇组成: 乙醇可改为1:1 甲醇: 乙醇溶液, 可比较结果为21。在陷阱22中, 肥皂水也可以代替丙烯乙二醇作为杀死溶液。对于大量的 CBB (例如) 的估计数, 超过每个陷阱的数量, 可以用基于质量的 CBB 估计代替容积估计。例如, 单个 CBB 的平均干重可以用高分辨率的刻度来确定。CBB 收集的70% 乙醇可以在烤箱中烘干, 并权衡估计每陷阱 CBB 的数量。还可以通过将所有的 CBB 从一个陷阱放入一个已毕业的缸中, 连同杀死解决方案, 并允许内容结算到底部的22来进行修改的容量估计。一旦解决, CBB 填充的钢瓶的体积可能会注意到, 1 毫升的换算系数可以确定估计每陷阱捕获的 CBB 总数。最后, 咖啡种植者对他们的农场有着密切的了解, 并使用这个监测协议来估计 CBB 的出没和移动, 可能希望省略一些步骤, 包括记录物候和计算树枝上葡萄干的数量。
这里值得一提的是本协议的两个潜在限制。首先, 在胸高处取样的树枝不会捕捉早期开花的作物, 在树冠上可能开始更高的害虫。然而, 这些观察表明, 这种早开花的作物占夏威夷咖啡场总产量的很小比例。第二, 我们的协议只记述了绿色浆果的出没, 因此当颜色断裂和成熟浆果的数量高 (9月December日在夏威夷) 时, 可能无法准确地捕捉到浆果损伤的估计。
此处介绍的 CBB 监视协议与当前使用的其他监视协议相比有几个明显的优势。首先, 系统的随机抽样设计允许更均匀的取样相对于在一个曲折的模式进行取样。这种抽样设计可以更好地估计整个给定领域的浆果损伤, 并增加探测热点的潜力。第二, 在监测议定书中列入对咖啡农田至关重要的元素 (例如、物候、天气变量和管理做法) 将有助于我们了解侵入性害虫之间的动态,他们的寄主植物, 以及各种环境因素。第三, 在实地调查中使用移动电子数据收集应用程序, 可使实时数据快速有效地输入和组织到数据库中, 也可以与其他自动咖啡监测方法 (如检测) 相关。通过遥感23。这种数据收集方法的另一个重要好处是, 可以轻松地生成详细的侵扰报告, 以便及时将管理建议转发给种植者。最后, 在 CBB 生物学、咖啡植物物候、天气和管理方面收集的实时数据可纳入预测模型的开发, 可用于自定义特定咖啡种植地点的管理计划。

咖啡莓果 (Hypothenemus hampei法拉利) 是一种侵袭性害虫, 在世界主要咖啡种植地区的1,2发现。这只小甲虫的生命周期大部分是在咖啡浆果的种子中度过的, 这使得用杀虫剂喷洒的药剂难以控制。成年女性通过中央盘孔进入咖啡莓果洞, 并进入它的种子, 它建造画廊繁殖。随着幼虫的发育, 它们以胚乳为食, 对咖啡豆造成直接损害, 并导致产量和质量的随后损失3。真菌和病原体进入豆类可能会造成间接损害, 这可能导致咖啡风味的发酵和改变4。
CBB 第一次在夏威夷 ' i 海岛在2010年8月被发现了5并且迅速地传播到几乎所有800个咖啡场在科纳和 Ka ' u 地区, 两个区域是世界知名的为他们的咖啡产品的优质质量6,7.非托管和管理不善的农场可能会有超过90% 的侵扰程度, 造成巨大的经济损失。在夏威夷, 由于 CBB 所造成的经济影响估计每年约为2100万美元,8。CBB 自最初引进夏威夷岛以来一直在传播, 最近在邻近的夏威夷群岛瓦胡岛 (2014) 和毛伊岛 (2016) 被发现。艾考岛是夏威夷唯一不受 CBB 影响的咖啡馆, 但该岛3000英亩的咖啡极易受到这种高度分散的害虫的伤害。
历史上, 丹和毒死蜱等合成杀虫剂在许多国家被用来控制 CBB。然而, 对这些杀虫剂对人类和环境的毒性的关切9, 以及杀虫剂抗药性的证据10, 导致这些物质被禁止在许多国家使用。目前, 大多数咖啡种植区都依赖 IPM 方法来控制 CBB。IPMs 通常涉及环境卫生做法 (例如、修剪和剥带)、生物控制 (例如、掠夺性甲虫或寄生昆虫的释放) 以及生物农药的应用 (例如,杀虫真菌B. 白僵菌)11,12。目前在夏威夷 CBB 管理的建议还建议使用酒精诱饵陷阱和 "三十树取样方法" 开发的 Cenicafé13,14, 定期现场监测。这种抽样方法涉及随机选择一个分支, 从中冠至少有45绿色浆果, 并计数的数量, 感染和不受感染的浆果。这个过程是重复的曲折模式在整个领域, 总共30棵树每公顷 (2.5 英亩), 并用于估计数量的侵扰。
尽管夏威夷的咖啡种植者采取了许多这种 ipm 做法, 但群岛的气候、地形和文化习俗的极端异质性要求每个地点都要对 ipm 进行定制。定制 IPM 的开发将取决于一个监测程序, 其中包括咖啡农田, 咖啡害虫生物学和环境的基本要素。我们已实施全面监测 CBB 和主机的动态, 作为一个区域范围内项目的一部分, 在夏威夷, 聚集景观水平的数据, 以通知管理做法。该协议可用于世界各地的其他咖啡农田, 并将特别有用的那些经历高度多变的环境和社会经济因素需要定制 IPM 管理 CBB 人口。

<table><tbody><tr><td>funnel trap</td><td>CIRAD</td><td></td><td>Brocap trap</td></tr><tr><td>propylene glycol</td><td>Better World Manufacturing, Inc.</td><td></td><td></td></tr><tr><td>methanol</td><td>Fisher Scientific or similar supplier</td><td></td><td>CAUTION: Methanol is highly flammable, is toxic if inhaled or ingested, and is a skin and eye irritant. Wear gloves, eye protection, and protective clothing, and only use in well-ventilated rooms.</td></tr><tr><td>ethanol</td><td>Fisher Scientific or similar supplier</td><td></td><td>CAUTION: Ethanol is highly flammable, is toxic if inhaled or ingested, and is a skin and eye irritant. Wear gloves, eye protection, and protective clothing, and only use in well-ventilated rooms.</td></tr><tr><td>polypropylene resealable bags (2 Mil 3 x 4&quot;)</td><td>Uline or similar supplier</td><td>S-1292</td><td></td></tr><tr><td>thumbtack</td><td>Widely available</td><td></td><td>For making drainage holes in funnel trap</td></tr><tr><td>paperclips</td><td>Widely available</td><td></td><td>For attaching lure bag to traps</td></tr><tr><td>galvanized wire (12 gauge)</td><td>Widely available</td><td></td><td>For attaching funnel trap to stakes</td></tr><tr><td>wire cutter</td><td>Widely available</td><td></td><td></td></tr><tr><td>tomato stakes</td><td>Widely available</td><td></td><td></td></tr><tr><td>permanent marker</td><td>Widely available</td><td></td><td></td></tr><tr><td>mobile device</td><td>Apple or other supplier</td><td></td><td>iPad or smartphone equipped with camera</td></tr><tr><td>waterproof case</td><td>Widely available</td><td></td><td>For mobile device</td></tr><tr><td>data collection application</td><td>Fulcrum or similar software</td><td></td><td></td></tr><tr><td>GNSS Surveyor</td><td>Bad Elf</td><td></td><td>~1-meter positioning accuracy</td></tr><tr><td>1 mm mesh hand sieve</td><td>Widely available</td><td></td><td></td></tr><tr><td>1.5 mm mesh hand sieve</td><td>Widely available</td><td></td><td></td></tr><tr><td>20 mL glass scintillation vials</td><td>Widely available</td><td></td><td></td></tr><tr><td>label maker</td><td>Widely available</td><td></td><td></td></tr><tr><td>label tape</td><td>Widely available</td><td></td><td></td></tr><tr><td>metal lab spatula</td><td>Widely available</td><td></td><td></td></tr><tr><td>scrub brush</td><td>Widely available</td><td></td><td></td></tr><tr><td>dish soap</td><td>Widely available</td><td></td><td></td></tr><tr><td>binder clip</td><td>Widely available</td><td></td><td></td></tr><tr><td>ruler</td><td>Widely available</td><td></td><td></td></tr><tr><td>plastic tupperware</td><td>Widely available</td><td></td><td></td></tr><tr><td>cooler</td><td>Widely available</td><td></td><td></td></tr><tr><td>ice pack</td><td>Widely available</td><td></td><td></td></tr><tr><td>wash bottle</td><td>Widely available</td><td></td><td></td></tr><tr><td>papertowels</td><td>Widely available</td><td></td><td></td></tr><tr><td>fine-tipped paintbrush</td><td>Widely available</td><td></td><td></td></tr><tr><td>light microscope</td><td>Leica or similar supplier</td><td></td><td></td></tr><tr><td>clear plastic lid</td><td>Widely available</td><td></td><td></td></tr><tr><td>tally counter</td><td>Widely available</td><td></td><td></td></tr><tr><td>10 mL syringe</td><td>Widely available</td><td></td><td></td></tr><tr><td>fine-tipped forceps</td><td>Widely available</td><td></td><td></td></tr><tr><td>scalpel or razor blade</td><td>Widely available</td><td></td><td></td></tr><tr><td>freezer</td><td>Widely available</td><td></td><td></td></tr><tr><td>waterproof data shuttle</td><td>HOBO by Onset Computer Corp.</td><td>U-DTW-1</td><td></td></tr><tr><td>PAR Sensor with 3m Cable</td><td>HOBO by Onset Computer Corp.</td><td>S-LIA-M003</td><td></td></tr><tr><td>Temp/RH Sensor (12-bit) w/ 2m Cable</td><td>HOBO by Onset Computer Corp.</td><td>S-THB-M002</td><td></td></tr><tr><td>Solar Radiation Shield</td><td>HOBO by Onset Computer Corp.</td><td>RS3</td><td></td></tr><tr><td>Extra-Large Solar Panel 6 Watts</td><td>HOBO by Onset Computer Corp.</td><td>SOLAR-6W</td><td></td></tr><tr><td>Rain Gauge (0.2mm) with 2m Cable</td><td>HOBO by Onset Computer Corp.</td><td>S-RGB-M002</td><td></td></tr><tr><td>Smart Temp Sensor 12-bit w/ 2m Cable</td><td>HOBO by Onset Computer Corp.</td><td>S-TMB-M002</td><td></td></tr><tr><td>Soil Moisture - 10HS</td><td>HOBO by Onset Computer Corp.</td><td>S-SMD-M005</td><td></td></tr><tr><td>Silicon Pyranometer Sensor w/3m Cable</td><td>HOBO by Onset Computer Corp.</td><td>S-LIB-M003</td><td></td></tr><tr><td>Light Sensor Bracket</td><td>HOBO by Onset Computer Corp.</td><td>M-LBB</td><td></td></tr><tr><td>NDVI Light Sensor Bracket</td><td>HOBO by Onset Computer Corp.</td><td>M-NDVI</td><td></td></tr><tr><td>Complete 3M Tripod kit</td><td>HOBO by Onset Computer Corp.</td><td>M-TPA-KIT</td><td></td></tr><tr><td>RX3000 3G Remote Monitoring Station</td><td>HOBO by Onset Computer Corp.</td><td>RX3003-00-01</td><td></td></tr><tr><td>Global Limited Plan - RX3000 T2 4-hr</td><td>HOBO by Onset Computer Corp.</td><td>SP-806</td><td></td></tr></tbody></table>

the hawaii protocol for scientific monitoring of coffee berry borer: a model for coffee agroecosystems worldwide

咖啡浆果 (CBB) 是世界上最具破坏性的咖啡作物害虫。我们制定了一项科学的监测协议, 目的是捕获和量化这种侵入性害虫的动态和影响, 以及它在异质景观中的寄主植物的开发。这一全面监测系统的基石是及时地理数据收集 CBB 运动, 咖啡莓虫出没, 死亡率由真菌白僵菌, 和咖啡植物物候通过移动电子数据记录应用.这个电子数据收集系统允许现场记录通过内置的全球定位系统进行地理, 并得到一个气象站网络和农场管理实践记录的支持。全面监测 CBB 和寄主植物动态是夏威夷地区范围内项目的一个重要组成部分, 目的是收集景观水平数据, 以改进管理实践。咖啡农田在世界其他地区经历高度多变的环境和社会经济因素也将受益于实施本议定书, 因为它将推动发展定制的综合虫害管理 (IPM), 以管理 CBB 人口。

我们报告几个咖啡养殖场的例子, 这些实例代表了从上文所述监测协议中可以获得的结果类型。为了确定字段内部和之间的 CBB 移动模式, 给定陷阱的总捕获量可以除以部署后的天数来估计每天捕获的 CBB 数。然后, 每天捕获的 CBB 数可以平均分布在所有的陷阱中, 以确定每个陷阱每天捕获的 CBB 的平均数量 (均为 SEM;图 2)。陷阱捕捉数据可用于推断峰值飞行活动的期间18, 也可用于指导管理活动, 如修剪和B. 白僵菌喷雾。在田间浆果损伤评估中获得的百分比可能与陷阱捕捉数据进行比较, 以确定高感染期是否与峰值飞行活动19相吻合。这些信息对于决定是否仅通过陷阱监测 CBB 活动是否足以通知控制措施至关重要。浆果解剖在实验室确定 CBB 位置可以用来通知种植者何时喷洒B. 白僵菌的应用 (> 5% 的 CBB 是在 AB 位置14)。CBB 位置信息还可以与现场的损坏评估所产生的热点图一起使用, 以便向种植者通报应喷洒B. 白僵菌的区域内的近似位置 (图 3)。
通过编制关于 CBB 位置、死亡率由 CBB、植物物候学和管理实践的数据, 可以获得有关造成感染的各种因素的全面看法. 在图 4中显示的示例场中, 在生长季节早期解剖的大多数浆果都是在 AB 位置 CBB 的, 而在本季晚些时候解剖的大部分浆果则在 CD 位置上 CBB。在浆果生产的高峰期之后, 从7月下旬到12月记录了七轮樱桃收获 (图 4)。最后, 本赛季在大约一个月的时间内进行了七次B. 白僵菌的应用, 观察到的 CBB 死亡率从 0-23% (图 4) 不等。最后, 虽然这里没有天气预报, 但温度、湿度和降雨信息的增加可能会进一步深入了解驱动 CBB 的因素, 以及白僵菌对咖啡场的有效性。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/57204/57204fig1.jpg"> 图 1.平均 (SEM) CBB 捕获每一个陷阱每天进行抽样和双周间隔.这个平均捕获每天是为五漏斗陷阱散布在农场随机。在每周取样中捕捉到更多的极端峰和低谷, 而这些峰值在两周抽样中略有增加, 尽管这两个区间的总体趋势是可比的。<a href="/files/ftp_upload/57204/57204fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/57204/57204fig2.jpg"> 图 2.平均每天捕获的 CBB.这个平均捕获每天是为九漏斗陷阱散布在农场随机。在这个农场 (3月和 12月), 在2016-2017 的生长季节, CBB 飞行活动的两个主要山峰可以看到。<a href="/files/ftp_upload/57204/57204fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/57204/57204fig3v2.jpg"> 图 3.CBB 侵扰热点.这张咖啡养殖场的地图显示了在2017年6月14日的一项监测调查中观察到的 CBB 出没的热点。每个红色圆圈的大小与取样分支上的绿色的浆果数量成正比。在这个样本农场里, 总共取样了25个分支, 每根树枝上都有 0-36 种被感染的绿色浆果。<a href="/files/ftp_upload/57204/57204fig3v2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/57204/57204fig4.jpg"> 图 4.一个全面的看法 CBB 在一个样本咖啡场出没.CBB 在解剖的绿色浆果中的位置被定义为 AB (雌性已经开始进入浆果, 但尚未达到胚乳) 或 CD (雌性进入胚乳)。CBB 的死亡率 (通过白僵菌真菌)、咖啡植物物候 (每个分支的平均浆果数量) 和农场管理做法 (B. 白僵菌喷雾和樱桃采摘) 也显示在2016咖啡种植季节。<a href="/files/ftp_upload/57204/57204fig4large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>

Watch this Scientific Journal Video about The Hawaii Protocol for Scientific Monitoring of Coffee Berry Borer: a Model for Coffee Agroecosystems Worldwide at JoVE.com

The Hawaii Protocol for Scientific Monitoring of Coffee Berry Borer: a Model for Coffee Agroecosystems Worldwide

全面监测咖啡浆果和寄主植物的动态是收集景观水平的数据, 以改善管理这一入侵害虫的关键。在这里, 我们提出了一个科学监测咖啡浆果的运动, 出没, 死亡, 咖啡植物物候, 天气和农场管理通过移动电子数据记录应用的协议。

夏威夷咖啡莓科学监测协议: 全球咖啡农田模型

Coffee berry borer (CBB) is the most devastating insect pest for coffee crops worldwide. We developed a scientific monitoring protocol that is aimed at ...

the-hawaii-protocol-for-scientific-monitoring-coffee-berry-borer

Research

JoVE Journal

Environment

8.0K 次观看.  Oak Ridge Institute for Science and Education. 全面监测咖啡浆果和寄主植物的动态是收集景观水平的数据, 以改善管理这一入侵害虫的关键。在这里, 我们提出了一个科学监测咖啡浆果的运动, 出没, 死亡, 咖啡植物物候, 天气和农场管理通过移动电子数据记录应用的协议。

视频: The Hawaii Protocol for Scientific Monitoring of Coffee Berry Borer: a Model for Coffee Agroecosystems Worldwide

A Technique to Screen American Beech for Resistance to the Beech Scale Insect (Cryptococcus fagisuga Lind.)

Beech bark disease (BBD) results in high levels of initial mortality, leaving behind survivor trees that are greatly weakened and deformed.&#160;The disease is initiated by feeding activities of the invasive beech scale insect, Cryptococcus fagisuga, which creates entry points for infection by one of the Neonectria species of fungus.&#160;Without scale infestation, there is little opportunity for fungal infection. Using scale eggs to artificially infest healthy trees in heavily BBD impacted stands demonstrated that these trees were resistant to the scale insect portion of the disease complex1.&#160;Here we present a protocol that we have developed, based on the artificial infestation technique by Houston2, which can be used to screen for scale-resistant trees in the field and in smaller potted seedlings and grafts.&#160;The identification of scale-resistant trees is an important component of management of BBD through tree improvement programs and silvicultural manipulation.

A Technique to Screen American Beech for Resistance to the Beech Scale Insect Cryptococcus fagisuga Lind.

Beech bark disease is initiated by feeding activities of the beech scale insect that create fungal entry points in the bark.&#160;Trees that are resistant to the scale insect are also disease resistant.&#160;Here we present the protocol we have developed to screen individual beech trees for beech scale resistance.

一种技术，屏幕美国榉木为抵抗山毛榉介壳虫（<em&gt;隐球菌fagisuga</em&gt;林德。）

Beech bark disease (BBD) results in high levels of initial mortality, leaving behind survivor trees that are greatly weakened and deformed.&#160;The ...

科研

环境科学

Video Tracking Protocol to Screen Deterrent Chemistries for Honey Bees

The European honey bee, Apis mellifera L., is an economically and agriculturally important pollinator that generates billions of dollars annually. Honey bee colony numbers have been declining in the United States and many European countries since 1947. A number of factors play a role in this decline, including the unintentional exposure of honey bees to pesticides. The development of new methods and regulations are warranted to reduce pesticide exposures to these pollinators. One approach is the use of repellent chemistries that deter honey bees from a recently pesticide-treated crop. Here, we describe a protocol to discern the deterrence of honey bees exposed to select repellent chemistries. Honey bee foragers are collected and starved overnight in an incubator 15 h prior to testing. Individual honey bees are placed into Petri dishes that have either a sugar-agarose cube (control treatment) or sugar-agarose-compound cube (repellent treatment) placed into the middle of the dish. The Petri dish serves as the arena that is placed under a camera in a light box to record the honey bee locomotor activities using video tracking software. A total of 8 control and 8 repellent treatments were analyzed for a 10 min period with each treatment was duplicated with new honey bees. Here, we demonstrate that honey bees are deterred from the sugar-agarose cubes with a compound treatment whereas honey bees are attracted to the sugar-agarose cubes without an added compound.

The loss of honey bee colonies presents a challenge to crop pollination services. Current pollinator protection practices warrant an alternative approach to minimize the contact of honey bees to harmful pesticides using repellent chemistries. Here, we provide detailed methods for a visual tracking protocol to screen deterrents for bees.

用于蜂蜜蜜蜂筛选化学成分的视频跟踪协议

The European honey bee, Apis mellifera L., is an economically and agriculturally important pollinator that generates billions of dollars annually. Honey ...

Using Capillary Electrophoresis to Quantify Organic Acids from Plant Tissue: A Test Case Examining Coffea arabica Seeds

Carboxylic acids are organic acids containing one or more terminal carboxyl (COOH) functional groups. Short chain carboxylic acids (SCCAs; carboxylic acids containing three to six carbons), such as malate and citrate, are critical to the proper functioning of many biological systems, where they function in cellular respiration and can serve as indicators of cell health. In foods, organic acid content can have significant impact on taste, with increased SCCA levels resulting in a sour or &#34;acid&#34; taste. Because of this, methods for the rapid analysis of organic acid levels are of particular interest to the food and beverage industries. Unfortunately, however, most methods used for SCCA quantification are dependent on time-consuming protocols requiring the derivatization of samples with hazardous reagents, followed by costly chromatographic and/or mass spectrometric analyses. This method details an alternate method for the detection and quantification of organic acids from plant material and food samples using free zonal capillary electrophoresis (CZE), sometimes simply referred to as capillary electrophoresis (CE). CZE provides a cost-effective method for measuring SCCAs with a low limit of detection (0.005 mg/ml). This article details the extraction and quantification of SCCAs from plant samples. While the method provided focuses on measurement of SCCAs from coffee beans, the method provided can be applied to multiple plant-based food materials.

Using Capillary Electrophoresis to Quantify Organic Acids from Plant Tissue: A Test Case Examining Coffea arabica Seeds

This article presents a method for the detection and quantification of organic acids from plant material using free zonal capillary electrophoresis. An example of the potential application of this method, determining the effects of a secondary fermentation on organic acid levels in coffee seeds, is provided.

利用毛细管电泳从植物组织量化有机酸:一个测试用例检查<em&gt;小粒种咖啡</em&gt;种子

Carboxylic acids are organic acids containing one or more terminal carboxyl (COOH) functional groups. Short chain carboxylic acids (SCCAs; carboxylic ...

化学

Key Elements of Photo Attraction Bioassay for Insect Studies or Monitoring Programs

Optimized visual attractants will increase insect trapping efficiency by using the target insect&#39;s innate behaviors (positive photo-taxis) as a means to lure the insect into a population control or monitoring trap. Light emitting diodes (LEDs) have created customizable lighting options with specific wavelengths (colors), intensities, and bandwidths, all of which can be customized to the target insects. Photo-attraction behavioral bioassays can use LEDs to optimize the attractive color(s) for an insect species down to specific life history stages or behaviors (mating, feeding, or seeking shelter). Researchers must then confirm the bioassay results in the field and understand the limited attractive distance of the visual attractants.
The cloverleaf bioassay arena is a flexible method to assess photo attraction while also assessing a range of natural insect behaviors such as escape and feeding responses. The arena can be used for terrestrial or aerial insect experiments, as well as diurnal, and nocturnal insects. Data collection techniques with the arena are videotaping, counting contact with the lights, or physically collecting the insects as they are attracted towards the lights. The assay accounts for insects that make no-choice and the arenas can be single (noncompetitive) color or multiple (competitive) colors. The cloverleaf design causes insects with strong thigmotaxis to return to the center of the arena where they can view all the options in a competitive LED tests. The cloverleaf arena presented here has been used with mosquitoes, bed bugs, Hessian fly, house flies, biting midges, red flour beetles, and psocids. Bioassays are used to develop accurate and effective insect traps to guide the development and optimization of insect traps used to monitor pest population fluctuations for disease vector risk assessments, the introduction of invasive species, and/or be used for population suppression.

Photo-attraction bioassay arenas are used to determine the optimal light color(s) to maximize insect attraction; however bioassays and methods are specific to target insect behaviors and habitats. Customizable equipment and modifications are explained for nocturnal or diurnal and terrestrial or aerial insects.

昆虫研究或监测项目照片吸引力生物测定的关键要素

Optimized visual attractants will increase insect trapping efficiency by using the target insect&#39;s innate behaviors (positive photo-taxis) as a means ...

行为学

A Standardized Procedure for Monitoring Harmful Algal Blooms in Chile by Metabarcoding Analysis

Harmful algae blooms (HABs) monitoring has been implemented worldwide, and Chile, a country famous for its fisheries and aquaculture, has intensively used microscopic and toxin analyses for decades for this purpose. Molecular biological methods, such as high-throughput DNA sequencing and bacterial assemblage-based approaches, are just beginning to be introduced in Chilean HAB monitoring, and the procedures have not yet been standardized. Here, 16S rRNA and 18S rRNA metabarcoding analyses for monitoring Chilean HABs are introduced stepwise. According to a recent hypothesis, algal-bacterial mutualistic association plays a critical synergetic or antagonistic relationship accounting for bloom initiation, maintenance, and regression. Thus, monitoring HAB from algal-bacterial perspectives may provide a broader understanding of HAB mechanisms and the basis for early warning. Metabarcoding analysis is one of the best suited molecular-based tools for this purpose because it can detect massive algal-bacterial taxonomic information in a sample. The visual procedures of sampling to metabarcoding analysis herein provide specific instructions, aiming to reduce errors and collection of reliable data.

This protocol introduces steps of metabarcoding analysis, targeting 16S rRNA and 18S rRNA genes, for monitoring harmful algal blooms and their associated microbiome in seawater samples. It is a powerful molecular-based tool but requires several procedures, which are visually explained here step-by-step.

通过元条形码分析监测智利有害藻华的标准化程序

Harmful algae blooms (HABs) monitoring has been implemented worldwide, and Chile, a country famous for its fisheries and aquaculture, has intensively used ...

Methodology to Test Control Agents and Insecticides Against the Coffee Berry Borer Hypothenemus hampei

Prior to recommending insecticides to treat the coffee berry borer (CBB) Hypothenemus hampei, it is valuable to know the mortality and repellency of these insecticides against adult insects or their impact on reproductive output. However, currently available methods assess adult mortality only, limiting the selection of novel insecticides with a different mode of action. In this work, different experimental methods were examined to identify the diverse effects on the CBB under laboratory conditions. For this, green coffee fruits (GFs) were collected and disinfected by immersion in sodium hypochlorite solution followed by UV light irradiation. In parallel, CBB adults from a colony were disinfected by immersion in sodium hypochlorite solution. To assess fruit protection (preinfestation), the fruits were placed in plastic boxes, and the insecticides were applied. Then, the CBB adults were released at a rate of two CBBs per GF. The GFs were left under controlled conditions to evaluate CBB infestation and survival after 1, 7, 15, and 21 days. To evaluate insecticide efficacy after CBB infestation (postinfestation), CBB adults were released to the GFs in a 2:1 ratio for 3 h at 21 &#176;C. Infested fruits showing CBB adults with their abdomens partially exposed were selected and placed in 96-well racks, and the CBBs boring into the fruits were treated directly. After 20 days, the fruits were dissected, and the CBB biological stages inside each fruit were recorded. The GFs served as substrates that mimic natural conditions to evaluate toxic, chemical, and biological insecticides against the CBB.

Methodology to Test Control Agents and Insecticides Against the Coffee Berry Borer Hypothenemus hampei

A method using green coffee fruits (GFs) was developed to test the toxicity of insecticides against the coffee berry borer (CBB). Insecticides or toxic substances were applied to disinfected GFs before or after CBB infestation. Insect mortality, repellency, and reproductive capacity, in addition to other parameters, were evaluated.

针对咖啡浆果螟子下皮测试控制剂和杀虫剂 的方法

Prior to recommending insecticides to treat the coffee berry borer (CBB) Hypothenemus hampei, it is valuable to know the mortality and repellency of these ...

An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

Fopius arisanus (Sonan) is an important parasitoid of Tephritid fruit flies for at least two reasons. First, it is the one of only three opiine parasitoids known to infect the host during the egg stage1. Second, it has a wide range of potential fruit fly hosts. Perhaps due to its life history, F. arisanus has been a successfully used for biological control of fruit flies in multiple tropical regions2-4. One impediment to the wide use of F. arisanus for fruit fly control is that it is difficult to establish a stable laboratory colony5-9. Despite this difficulty, in the 1990s USDA researchers developed a reliable method to maintain laboratory populations of F. arisanus10-12. There is significant interest in F. arisanus biology13,14, especially regarding its ability to colonize a wide variety of Tephritid hosts14-17; interest is especially driven by the alarming spread of Bactrocera fruit fly pests to new continents in the last decade18. Further research on F. arisanus and additional deployments of this species as a biological control agent will benefit from optimizations and improvements of rearing methods. In this protocol and associated video article we describe an optimized method for rearing F. arisanus based on a previously described approach12. The method we describe here allows rearing of F. arisanus in a small scale without the use of fruit, using materials available in tropical regions around the world and with relatively low manual labor requirements.

An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

Fopius arisanus is an egg-larval parasitoid of Tephritid fruit flies that is successfully used in biological control of these important tropical pests. We describe here an optimized protocol for rearing F. arisanus on a small scale using readily available materials.

一个优化的协议饲养 Fopius arisanus Tephritid果寄生蝇

Fopius arisanus (Sonan) is an important parasitoid of Tephritid fruit flies for at least two reasons. First, it is the one of only three opiine ...

生物学

Monitoring Colony-level Effects of Sublethal Pesticide Exposure on Honey Bees

The effects of sublethal pesticide exposure to honey bee colonies may be significant but difficult to detect in the field using standard visual assessment methods. Here we describe methods to measure the quantities of adult bees, brood, and food resources by weighing hives and hive parts, by photographing frames, and by installing hives on scales and with internal sensors. Data from these periodic evaluations are then combined with running average and daily detrended data on hive weight and internal hive temperature. The resulting datasets have been used to detect colony-level effects of imidacloprid applied in a sugar syrup as low as 5 parts per billion. The methods are objective, require little training, and provide permanent records in the form of sensor output and photographs.

Sublethal doses of pesticides may affect colonies in ways that are difficult to detect using only periodic or visual methods. Methods to measure total adult bee mass, brood, and food resources by weighing hives and hive parts, photographing frames, and installing sensors, are provided. Data analysis is also addressed.

蜜蜂致死农药暴露对蜂群水平的影响

The effects of sublethal pesticide exposure to honey bee colonies may be significant but difficult to detect in the field using standard visual assessment ...

Tobacco Hornworm as an Insect Model System for Cannabinoid Pre-clinical Studies

With increased attention on cannabinoids in medicine, several mammalian model organisms have been used to elucidate their unknown pharmaceutical functions. However, many difficulties remain in mammalian research, which necessitates the development of non-mammalian model organisms for cannabinoid research. The authors suggest the tobacco hornworm Manduca sexta as a novel insect model system. This protocol provides information on preparing the artificial diet with varying amounts of cannabidiol (CBD), setting up a cultivation environment, and monitoring their physiological and behavioral changes in response to CBD treatment. Briefly, upon receiving hornworm eggs, the eggs were allowed 1-3 days at 25 &#176;C on a 12:12 light-dark cycle to hatch before being randomly distributed into control (wheat germ-based artificial diet; AD), vehicle (AD + 0.1% medium-chain triglyceride oil; MCT oil) and treatment groups (AD + 0.1% MCT + 1 mM or 2 mM of CBD). Once the media was prepared, 1st instar larvae were individually placed in a 50 mL test tube with a wooden skewer stick, and then the test tube was covered with a cheesecloth. Measurements were taken in 2-day intervals for physiological and behavioral responses to the CBD administration. This simple cultivation procedure allows researchers to test large specimens in a given experiment. Additionally, the relatively short life cycles enable researchers to study the impact of cannabinoid treatments over multiple generations of a homogenous population, allowing for data to support an experimental design in higher mammalian model organisms.

The present protocol provides instructional information for using tobacco hornworm Manduca sexta in cannabinoid research. The method described here includes all necessary supplies and protocols to monitor physiological and behavioral changes of the insect model in response to cannabidiol (CBD) treatment.

烟草角虫作为大麻素临床前研究的昆虫模型系统

With increased attention on cannabinoids in medicine, several mammalian model organisms have been used to elucidate their unknown pharmaceutical ...

Double-Staining Method to Detect Pectin in Plant-Fungus Interaction

Plant cells use different structural mechanisms, either constitutive or inducible, to defend themselves from fungal infection. Encapsulation is an efficient inducible mechanism to isolate the fungal haustoria from the plant cell protoplast. Conversely, pectin, one of the polymeric components of the cell wall, is a target of several pectolytic enzymes in necrotrophic interactions. Here, a protocol to detect pectin and fungal hyphae through optical microscopy is presented. The pectin-rich encapsulation in the cells of coffee leaves infected by the rust fungus Hemileia vastatrix and the mesophyll cell wall modification induced by Cercospora coffeicola are investigated. Lesioned leaf samples were fixed with the Karnovsky solution, dehydrated, and embedded in glycol methacrylate for 2-4 days. All steps were followed by vacuum-pumping to remove air in the intercellular spaces and improve the embedding process. The embedded blocks were sectioned into 5-7 &#181;m thick sections, which were deposited on a glass slide covered with water and subsequently heated at 40 &#176;C for 30 min. Next, the slides were double-stained with 5% cotton blue in lactophenol to detect the fungus and 0.05% ruthenium red in water to detect pectin (acidic groups of polyuronic acids of pectin). Fungal haustoria of Hemileia vastatrix were found to be encapsulated by pectin. In coffee cercosporiosis, mesophyll cells exhibited dissolution of cell walls, and intercellular hyphae and conidiophores were observed. The method presented here is effective to detect a pectin-associated response in the plant-fungi interaction.

This protocol describes a microscopical method to detect pectin in coffee-fungus interaction.

夏威夷咖啡莓科学监测协议: 全球咖啡农田模型

摘要

探索更多视频

此视频中的章节

一种技术，屏幕美国榉木为抵抗山毛榉介壳虫（

用于蜂蜜蜜蜂筛选化学成分的视频跟踪协议

利用毛细管电泳从植物组织量化有机酸:一个测试用例检查

昆虫研究或监测项目照片吸引力生物测定的关键要素

通过元条形码分析监测智利有害藻华的标准化程序

针对咖啡浆果螟子下皮测试控制剂和杀虫剂的方法

一个优化的协议饲养 Fopius arisanus Tephritid果寄生蝇

蜜蜂致死农药暴露对蜂群水平的影响

烟草角虫作为大麻素临床前研究的昆虫模型系统

检测植物-真菌相互作用中果胶的双重染色方法

夏威夷咖啡莓科学监测协议: 全球咖啡农田模型

摘要

探索更多视频

此视频中的章节

一种技术，屏幕美国榉木为抵抗山毛榉介壳虫（

用于蜂蜜蜜蜂筛选化学成分的视频跟踪协议

利用毛细管电泳从植物组织量化有机酸:一个测试用例检查

昆虫研究或监测项目照片吸引力生物测定的关键要素

通过元条形码分析监测智利有害藻华的标准化程序

针对咖啡浆果螟子下皮测试控制剂和杀虫剂 的方法

一个优化的协议饲养 Fopius arisanus Tephritid果寄生蝇

蜜蜂致死农药暴露对蜂群水平的影响

烟草角虫作为大麻素临床前研究的昆虫模型系统

检测植物-真菌相互作用中果胶的双重染色方法

针对咖啡浆果螟子下皮测试控制剂和杀虫剂的方法