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>
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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 border="0" cellpadding="0" cellspacing="0" width="729">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">funnel trap</td>
			<td style="width:213px;">CIRAD</td>
			<td style="width:119px;"></td>
			<td style="width:181px;">Brocap trap</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">propylene glycol</td>
			<td style="width:213px;">Better World Manufacturing, Inc.</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">methanol</td>
			<td style="width:213px;">Fisher Scientific or similar supplier</td>
			<td style="width:119px;"></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 height="42">
			<td height="42" style="height:42px;width:216px;">ethanol</td>
			<td style="width:213px;">Fisher Scientific or similar supplier</td>
			<td style="width:119px;"></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 height="42">
			<td height="42" style="height:42px;width:216px;">polypropylene resealable bags (2 Mil 3 x 4&#34;)</td>
			<td style="width:213px;">Uline or similar supplier</td>
			<td style="width:119px;">S-1292</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">thumbtack</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td>For making drainage holes in funnel trap</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">paperclips</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td>For attaching lure bag to traps</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">galvanized wire (12 gauge)</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td>For attaching funnel trap to stakes</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">wire cutter</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">tomato stakes</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">permanent marker</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">mobile device</td>
			<td style="width:213px;">Apple or other supplier</td>
			<td style="width:119px;"></td>
			<td>iPad or smartphone equipped with camera</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">waterproof case</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td>For mobile device</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">data collection application</td>
			<td style="width:213px;">Fulcrum or similar software</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">GNSS Surveyor</td>
			<td style="width:213px;">Bad Elf</td>
			<td style="width:119px;"></td>
			<td>~1-meter positioning accuracy</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">1 mm mesh hand sieve</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">1.5 mm mesh hand sieve</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">20 mL glass scintillation vials</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">label maker</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">label tape</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">metal lab spatula</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">scrub brush</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">dish soap</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">binder clip</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">ruler</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">plastic tupperware</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">cooler</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">ice pack</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">wash bottle</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">papertowels</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">fine-tipped paintbrush</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">light microscope</td>
			<td style="width:213px;">Leica or similar supplier</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">clear plastic lid</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">tally counter</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">10 mL syringe</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">fine-tipped forceps</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">scalpel or razor blade</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">freezer</td>
			<td style="width:213px;">Widely available</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">waterproof data shuttle</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td style="width:119px;">U-DTW-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">PAR Sensor with 3m Cable</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>S-LIA-M003</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">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 height="21">
			<td height="21" style="height:21px;width:216px;">Solar Radiation Shield</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>RS3</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Extra-Large Solar Panel 6 Watts</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>SOLAR-6W</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Rain Gauge (0.2mm) with 2m Cable</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>S-RGB-M002</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">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 height="21">
			<td height="21" style="height:21px;">Soil Moisture - 10HS</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>S-SMD-M005</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Silicon Pyranometer Sensor w/3m Cable</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>S-LIB-M003</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Light Sensor Bracket</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>M-LBB</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NDVI Light Sensor Bracket</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>M-NDVI</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Complete 3M Tripod kit</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>M-TPA-KIT</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">RX3000 3G Remote Monitoring Station</td>
			<td>HOBO by Onset Computer Corp.</td>
			<td>RX3003-00-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">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

7.9K 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

Optimized Negative Staining: a High-throughput Protocol for Examining Small and Asymmetric Protein Structure by Electron Microscopy

Structural determination of proteins is rather challenging for proteins with molecular masses between 40 - 200 kDa. Considering that more than half of natural proteins have a molecular mass between 40 - 200 kDa1,2, a robust and high-throughput method with a nanometer resolution capability is needed. Negative staining (NS) electron microscopy (EM) is an easy, rapid, and qualitative approach which has frequently been used in research laboratories to examine protein structure and protein-protein interactions. Unfortunately, conventional NS protocols often generate structural artifacts on proteins, especially with lipoproteins that usually form presenting rouleaux artifacts. By using images of lipoproteins from cryo-electron microscopy (cryo-EM) as a standard, the key parameters in NS specimen preparation conditions were recently screened and reported as the optimized NS protocol (OpNS), a modified conventional NS protocol 3 . Artifacts like rouleaux can be greatly limited by OpNS, additionally providing high contrast along with reasonably high&#8208;resolution (near 1 nm) images of small and asymmetric proteins. These high-resolution and high contrast images are even favorable for an individual protein (a single object, no average) 3D reconstruction, such as a 160 kDa antibody, through the method of electron tomography4,5. Moreover, OpNS can be a high&#8208;throughput tool to examine hundreds of samples of small proteins. For example, the previously published mechanism of 53 kDa cholesteryl ester transfer protein (CETP) involved the screening and imaging of hundreds of samples 6. Considering cryo-EM rarely successfully images proteins less than 200 kDa has yet to publish any study involving screening over one hundred sample conditions, it is fair to call OpNS a high-throughput method for studying small proteins. Hopefully the OpNS protocol presented here can be a useful tool to push the boundaries of EM and accelerate EM studies into small protein structure, dynamics and mechanisms.

More than half of proteins are small proteins (molecular mass &#60;200 kDa) that are challenging for both electron microscope imaging and three-dimensional reconstructions. Optimized negative staining is a robust and high-throughput protocol to obtain high contrast and relatively high resolution (~1 nm) images of small asymmetric proteins or complexes under different physiological conditions.

优化负染:通过电子显微镜检查小和不对称蛋白结构的高通量协议

Structural determination of proteins is rather challenging for proteins with molecular masses between 40 - 200 kDa. Considering that more than half of ...

科研

环境科学

Experimental Protocol for Manipulating Plant-induced Soil Heterogeneity

Coexistence theory has often treated environmental heterogeneity as being independent of the community composition; however biotic feedbacks such as plant-soil feedbacks (PSF) have large effects on plant performance, and create environmental heterogeneity that depends on the community composition. Understanding the importance of PSF for plant community assembly necessitates understanding of the role of heterogeneity in PSF, in addition to mean PSF effects. Here, we describe a protocol for manipulating plant-induced soil heterogeneity. Two example experiments are presented: (1) a field experiment with a 6-patch grid of soils to measure plant population responses and (2) a greenhouse experiment with 2-patch soils to measure individual plant responses. Soils can be collected from the zone of root influence (soils from the rhizosphere and directly adjacent to the rhizosphere) of plants in the field from conspecific and heterospecific plant species. Replicate collections are used to avoid pseudoreplicating soil samples. These soils are then placed into separate patches for heterogeneous treatments or mixed for a homogenized treatment. Care should be taken to ensure that heterogeneous and homogenized treatments experience the same degree of soil disturbance. Plants can then be placed in these soil treatments to determine the effect of plant-induced soil heterogeneity on plant performance. We demonstrate that plant-induced heterogeneity results in different outcomes than predicted by traditional coexistence models, perhaps because of the dynamic nature of these feedbacks. Theory that incorporates environmental heterogeneity influenced by the assembling community and additional empirical work is needed to determine when heterogeneity intrinsic to the assembling community will result in different assembly outcomes compared with heterogeneity extrinsic to the community composition.

Understanding the role of environmental heterogeneity in species coexistence has typically focused on types of heterogeneity that are extrinsic to the community&#8217;s species composition. We provide novel detailed methods for creating soil heterogeneity treatments using soils subject to plant-soil feedback conditioning, or heterogeneity intrinsic to the community composition.

用于操纵植物诱导土壤异质性的实验协议

Coexistence theory has often treated environmental heterogeneity as being independent of the community composition; however biotic feedbacks such as ...

A Protocol for Conducting Rainfall Simulation to Study Soil Runoff

Rainfall is a driving force for the transport of environmental contaminants from agricultural soils to surficial water bodies via surface runoff. The objective of this study was to characterize the effects of antecedent soil moisture content on the fate and transport of surface applied commercial urea, a common form of nitrogen (N) fertilizer, following a rainfall event that occurs within 24&#160;hr&#160;after fertilizer application. Although urea is assumed to be readily hydrolyzed to ammonium and therefore not often available for transport, recent studies suggest that urea can be transported from agricultural soils to coastal waters where it is implicated in harmful algal blooms. A rainfall simulator was used to apply a consistent rate of uniform rainfall across packed soil boxes that had been prewetted to different soil moisture contents. By controlling rainfall and soil physical characteristics, the effects of antecedent soil moisture on urea loss were isolated. Wetter soils exhibited shorter time from rainfall initiation to runoff initiation, greater total volume of runoff, higher urea concentrations in runoff, and greater mass loadings of urea in runoff. These results also demonstrate the importance of controlling for antecedent soil moisture content in studies designed to isolate other variables, such as soil physical or chemical characteristics, slope, soil cover, management, or rainfall characteristics. Because rainfall simulators are designed to deliver raindrops of similar size and velocity as natural rainfall, studies conducted under a standardized protocol can yield valuable data that, in turn, can be used to develop models for predicting the fate and transport of pollutants in runoff.

A rainfall simulator was used to apply a consistent rate of uniform rainfall to packed soil boxes in a study of the fate and transport of urea, a nonpoint source environmental contaminant. Under uniform soil and rainfall conditions, antecedent soil moisture content exerted strong control over urea loss in surface runoff.

一个协议，用于开展降雨模拟研究土壤径流

Rainfall is a driving force for the transport of environmental contaminants from agricultural soils to surficial water bodies via surface runoff. The ...

A Protocol for Collecting and Constructing Soil Core Lysimeters

Leaching of nutrients from land applied fertilizers and manure used in agriculture can lead to accelerated eutrophication of surface water. Because the landscape has complex and varied soil morphology, an accompanying disparity in flow paths for leachate through the soil macropore and matrix structure is present. The rate of flow through these paths is further affected by antecedent soil moisture. Lysimeters are used to quantify flow rate, volume of water and concentration of nutrients leaching downward through soils. While many lysimeter designs exist, accurately determining the volume of water and mass balance of nutrients is best accomplished with bounded lysimeters that leave the natural soil structure intact. 
Here we present a detailed method for the extraction and construction of soil core lysimeters equipped with soil moisture sensors at 5 cm and 25 cm depths. Lysimeters from four different Coastal Plain soils (Bojac, Evesboro, Quindocqua and Sassafras) were collected on the Delmarva Peninsula and moved to an indoor climate controlled facility. Soils were irrigated once weekly with the equivalent of 2 cm of rainfall to draw down soil nitrate-N concentrations. At the end of the draw down period, poultry litter was applied (162 kg TN ha-1) and leaching was resumed for an additional five weeks. Total recovery of applied irrigation water varied from 71% to 85%. Nitrate-N concentration varied over the course of the study from an average of 27.1 mg L-1 before litter application to 40.3 mg L-1 following litter application. While greatest flux of nutrients was measured in soils dominated by coarse sand (Sassafras) the greatest immediate flux occurred from the finest textured soil with pronounced macropore development (Quindocqua).

A detailed method for extraction and assembly of intact soil core lysimeters and their use for study of leachate and associated loss of nutrients from surface applied poultry litter is demonstrated.

一种用于收集和构建土芯协议测渗计

Leaching of nutrients from land applied fertilizers and manure used in agriculture can lead to accelerated eutrophication of surface water. Because the ...

Protocol for Microplastics Sampling on the Sea Surface and Sample Analysis

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of scientific publications address microplastic pollution of the sea surface. The protocol below describes the methodology for sampling, sample preparation, separation and chemical identification of microplastic particles. A manta net fixed on an &#187;A frame&#171; attached to the side of the vessel was used for sampling. Microplastic particles caught in the cod end of the net were separated from samples by visual identification and use of stereomicroscopes. Particles were analyzed for their size using an image analysis program and for their chemical structure using ATR-FTIR and micro FTIR spectroscopy. The described protocol is in line with recommendations for microplastics monitoring published by the Marine Strategy Framework Directive (MSFD) Technical Subgroup on Marine Litter. This written protocol with video guide will support the work of researchers that deal with microplastics monitoring all over the world.

The protocol below describes the methodology for: microplastics sampling on the sea surface, separation of microplastic and chemical identification of particles. This protocol is in line with the recommendations for microplastics monitoring published by the MSFD Technical Subgroup on Marine Litter.

协议塑料微粒采样的海面和样品分析

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of ...

Building Double-decker Traps for Early Detection of Emerald Ash Borer

Emerald ash borer (EAB) (Agrilus planipennis Fairmaire), the most destructive forest insect to have invaded North America, has killed hundreds of millions of forest and landscape ash (Fraxinus spp.) trees. Several artificial trap designs to attract and capture EAB beetles have been developed to detect, delineate, and monitor infestations. Double-decker (DD) traps consist of two corrugated plastic prisms, one green and one purple, attached to a 3 m tall polyvinyl chloride (PVC) pipe supported by a t-post. The green prism at the top of the PVC pipe is baited with cis-3-hexenol, a compound produced by ash foliage. Surfaces of both prisms are coated with sticky insect glue to capture adult EAB beetles. Double-decker traps should be placed near ash trees but in open areas, exposed to sun. Double-decker trap construction and placement are presented here, along with a summary of field experiments demonstrating the efficacy of DD traps in capturing EAB beetles. In a recent study in sites with relatively low EAB densities, double-decker traps captured significantly more EAB than green or purple prism traps or green funnel traps, all of which are designed to be suspended from a branch in the canopy of ash trees. A greater percentage of double decker traps were positive, i.e., captured at least one EAB, than the prism traps or funnel traps that were hung in ash tree canopies.

Effective traps to attract and capture the emerald ash borer (EAB) are a key element of detecting and managing this invasive pest. Double-decker traps, placed in full sun near ash trees, incorporate visual and olfactory cues and were more likely to capture EAB than other trap designs in field trials.

建筑双层疏水阀早期检测翡翠灰螟

Emerald ash borer (EAB) (Agrilus planipennis Fairmaire), the most destructive forest insect to have invaded North America, has killed hundreds of millions ...

Protocol for Assessing the Relative Effects of Environment and Genetics on Antler and Body Growth for a Long-lived Cervid

Cervid phenotype can be placed into one of two categories: efficiency, which promotes survival over extravagant morphometric growth, and luxury, which promotes growth of large weaponry and body size. Populations of the same species display each phenotype depending on environmental conditions. Although antler and body size of male white-tailed deer (Odocoileus virginianus) varies by physiographic region in Mississippi, USA and is strongly correlated with regional variation in nutritional quality, the effects of population-level genetics from native stocks and previous re-stocking efforts cannot be disregarded. This protocol describes how we designed a controlled study, where other factors that influence phenotype, such as age and nutrition, are controlled. We brought wild-caught pregnant females and six-month-old fawns from three distinct physiographic regions in Mississippi, USA to the Mississippi State University Rusty Dawkins Memorial Deer Unit. Deer from the same region were bred to produce a second generation of offspring, allowing us to assess generational responses and maternal effects. All deer ate the same high-quality (20% crude protein deer pellet) diet ad libitum. We uniquely marked each neonate and recorded body mass, hind foot, and total body length. Each subsequent fall, we sedated individuals via remote injection and sampled the same morphometrics plus antlers of adults. We found that all morphometrics increased in size from first to second generation, with full compensation of antler size (regional variation no longer present) and partial compensation of body mass (some evidence of regional variation) evident in the second generation. Second generation males that originated from our poorest quality soil region displayed about a 40% increase in antler size and about a 25% increase in body mass when compared to their wild harvested counterparts. Our results suggest phenotypic variation of wild male white-tailed deer in Mississippi are more related to differences in nutritional quality than population-level genetics.

Phenotypic differences among cervid populations may be related to population-level genetics or nutrition; discerning which is difficult in the wild. This protocol describes how we designed a controlled study where nutritional variation was eliminated. We found that phenotypic variation of male white-tailed deer was more limited by nutrition than genetics.

评估环境和遗传因素对鹿茸和长寿命的鹿的身体生长的相对影响的议定书 》

Cervid phenotype can be placed into one of two categories: efficiency, which promotes survival over extravagant morphometric growth, and luxury, which ...

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.

绿松石鳉鱼的急性和慢性毒性测试协议Nothobranchius furzeri

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

The Plant Infection Test: Spray and Wound-Mediated Inoculation with the Plant Pathogen Magnaporthe Grisea

Plants possess a powerful system to defend themselves against potential threats by pathogenic fungi. For agriculturally important plants, however, current measures to combat such pathogens have proved too conservative and, thus, not sufficiently effective, and they can potentially pose environmental risks. Therefore, it is extremely necessary to identify host-resistance factors to assist in controlling plant diseases naturally through the identification of resistant germplasm, the isolation and characterization of resistance genes, and the molecular breeding of resistant cultivars. In this regard, there is need to establish an accurate, rapid, and large-scale inoculation method to breed and develop plant resistance genes. The rice blast fungal pathogen Magnaporthe grisea causes severe disease symptoms and yield losses. Recently, M. grisea has emerged as a model organism for studying the mechanisms of plant-fungal pathogen interactions. Hence, we report the development of a plant virulence test method that is specific for M. grisea. This method provides for both spray inoculation with a conidial suspension and wounding inoculation with mycelium cubes or droplets of conidial suspension. The key step of the wounding inoculation method for detached rice leaves is to make wounds on plant leaves, which avoids any interference caused by host penetration resistance. This spray/wounding protocol contributes to the rapid, accurate, and large-scale screening of the pathotypes of M. grisea isolates. This integrated and systematic plant infection method will serve as an excellent starting point for gaining a broad perspective of issues in plant pathology.

The Plant Infection Test: Spray and Wound-Mediated Inoculation with the Plant Pathogen Magnaporthe Grisea

Here, we present a protocol to test plant virulence with the plant pathogen Magnaporthe grisea. This report will contribute to the large-scale screening of the pathotypes of fungi isolates and serve as an excellent starting point for understanding the resistant mechanisms of plants during molecular breeding.

植物感染试验: 以植物病原菌为载体的喷雾和伤口介导接种

Plants possess a powerful system to defend themselves against potential threats by pathogenic fungi. For agriculturally important plants, however, current ...

Harvesting Venom Toxins from Assassin Bugs and Other Heteropteran Insects

Heteropteran insects such as assassin bugs (Reduviidae) and giant water bugs (Belostomatidae) descended from a common predaceous and venomous ancestor, and the majority of extant heteropterans retain this trophic strategy. Some heteropterans have transitioned to feeding on vertebrate blood (such as the kissing bugs, Triatominae; and bed bugs, Cimicidae) while others have reverted to feeding on plants (most Pentatomomorpha). However, with the exception of saliva used by kissing bugs to facilitate blood-feeding, little is known about heteropteran venoms compared to the venoms of spiders, scorpions and snakes.
One obstacle to the characterization of heteropteran venom toxins is the structure and function of the venom/labial glands, which are both morphologically complex and perform multiple biological roles (defense, prey capture, and extra-oral digestion). In this article, we describe three methods we have successfully used to collect heteropteran venoms. First, we present electrostimulation as a convenient way to collect venom that is often lethal when injected into prey animals, and which obviates contamination by glandular tissue. Second, we show that gentle harassment of animals is sufficient to produce venom extrusion from the proboscis and/or venom spitting in some groups of heteropterans. Third, we describe methods to harvest venom toxins by dissection of anaesthetized animals to obtain the venom glands. This method is complementary to other methods, as it may allow harvesting of toxins from taxa in which electrostimulation and harassment are ineffective. These protocols will enable researchers to harvest toxins from heteropteran insects for structure-function characterization and possible applications in medicine and agriculture.

Although many insects in the suborder Heteroptera (Insecta: Hemiptera) are venomous, their venom composition and the functions of their venom toxins are mostly unknown. This protocol describes methods to harvest heteropteran venoms for further characterization, using electrostimulation, harassment, and gland dissection.

从刺客臭虫和其他 Heteropteran 昆虫身上采集毒液毒素

Heteropteran insects such as assassin bugs (Reduviidae) and giant water bugs (Belostomatidae) descended from a common predaceous and venomous ancestor, ...

夏威夷咖啡莓科学监测协议: 全球咖啡农田模型

摘要

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此视频中的章节

优化负染:通过电子显微镜检查小和不对称蛋白结构的高通量协议

用于操纵植物诱导土壤异质性的实验协议

一个协议，用于开展降雨模拟研究土壤径流

一种用于收集和构建土芯协议测渗计

协议塑料微粒采样的海面和样品分析

建筑双层疏水阀早期检测翡翠灰螟

评估环境和遗传因素对鹿茸和长寿命的鹿的身体生长的相对影响的议定书》

绿松石鳉鱼的急性和慢性毒性测试协议Nothobranchius furzeri

植物感染试验: 以植物病原菌为载体的喷雾和伤口介导接种

从刺客臭虫和其他 Heteropteran 昆虫身上采集毒液毒素

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

优化负染:通过电子显微镜检查小和不对称蛋白结构的高通量协议

用于操纵植物诱导土壤异质性的实验协议

一个协议，用于开展降雨模拟研究土壤径流

一种用于收集和构建土芯协议测渗计

协议塑料微粒采样的海面和样品分析

建筑双层疏水阀早期检测翡翠灰螟

评估环境和遗传因素对鹿茸和长寿命的鹿的身体生长的相对影响的议定书 》

绿松石鳉鱼的急性和慢性毒性测试协议Nothobranchius furzeri

植物感染试验: 以植物病原菌为载体的喷雾和伤口介导接种

从刺客臭虫和其他 Heteropteran 昆虫身上采集毒液毒素

评估环境和遗传因素对鹿茸和长寿命的鹿的身体生长的相对影响的议定书》