成像FlowCytobot(IFCB)是一种可现场部署的流式细胞仪,越来越多地用于监测有害藻类。IFCB基于它们的叶绿素-a荧光和/或它们散射(侧向散射)的光量来获取悬浮颗粒的图像。本研究假设基于荧光的图像采集会低估Dinphysisspp。,非组成型混合营养的一个属,当猎物有限时。这是因为Dinphysisspp。通过摄取纤毛虫猎物Mesodiniumspp获得质体。,在没有猎物的情况下失去光合能力和自发荧光。甚至小花的Dinphysisspp。可能是剧毒的,导致腹泻性贝类中毒(DSP),强调准确检测低丰度的重要性。为了探索这个,进行了实验室实验,以确定用于Dinphysisacuminata饲喂培养的最佳IFCB设置,以及在普吉特海湾收集的IFCB观测值的现有时间序列(华盛顿,U.S.A)用于比较Dinphysisspp。通过侧向散射与荧光触发的样品相对于Mesodiniumspp的丰度估计。丰度。这项研究引入了一种定量方法来优化目标有害藻类的检测,该方法可以在多个IFCB中重复进行,并证明了IFCB校准对Dinphysisspp的影响。检测。实验室实验表明,用于基于荧光的图像采集的IFCB设置需要相当灵敏,才能准确检测D.acuminata细胞。无论使用何种方法触发图像采集,校准不良的IFCB都可能会错过相当大比例的尖锐湿疣丰度。田间试验结果表明,Dinphysisspp的生理状态。当触发荧光时,可以影响IFCB的检测。在IFCB未能检测到Dinphysisspp的7天期间观察到了这一点。当细胞在荧光上触发时,细胞仍然使用侧向散射触发方法进行检测以及通过显微镜观察。在此期间,Mesodiniumspp.没有检测到,Finphysisspp的单个细胞的IFCB衍生的自发荧光水平。很低,和不到50%的Dinphysisspp。细胞在显微镜下表现出自发荧光。一起,这表明Dinphysisspp具有独特的摄食生态。当细胞饥饿时,可能会影响IFCB对它们的检测。
The Imaging FlowCytobot (IFCB) is a field-deployable imaging-in-flow cytometer that is increasingly being used to monitor harmful algae. The IFCB acquires images of suspended particles based on their chlorophyll-a fluorescence and/or the amount of light they scatter (side scattering). The present study hypothesized that fluorescence-based image acquisition would undercount Dinophysis spp., a genus of non-constitutive mixotrophs, when prey is limited. This is because Dinophysis spp. acquire plastids via ingestion of their ciliate prey
Mesodinium spp., and lose photosynthetic capacity and autofluorescence in the absence of prey. Even small blooms of Dinophysis spp. can be highly toxic and result in diarrhetic shellfish poisoning (DSP), highlighting the importance of accurately detecting low abundances. To explore this, laboratory experiments were conducted to determine optimal IFCB settings for a fed culture of Dinophysis acuminata, and an existing time series of IFCB observations collected in Puget Sound (Washington, U.S.A) was used to compare Dinophysis spp. abundance estimates from samples triggered via side scattering versus fluorescence in relation to
Mesodinium spp. abundance. This study introduces a quantitative approach for optimizing the detection of target harmful algae which can be repeated across multiple IFCBs and demonstrates the effects of IFCB calibration on Dinophysis spp. detection. The laboratory experiments showed that IFCB settings for fluorescence-based image acquisition need to be fairly sensitive to accurately detect D. acuminata cells. A poorly calibrated IFCB can miss a significant proportion of D. acuminata abundance whatever the method used to trigger the image acquisition. Field results demonstrated that the physiological status of Dinophysis spp. can influence their detection by the IFCB when triggering on fluorescence. This was observed during a 7-day period when the IFCB failed to detect Dinophysis spp. cells when triggering on fluorescence while cells were still detected using the side scattering triggering method as well as observed by microscopy. During this period,
Mesodinium spp. was not detected, IFCB-derived autofluorescence level of individual cells of Dinophysis spp. was low, and less than 50 % of Dinophysis spp. cells exhibited autofluorescence under the microscope. Together, this indicates that the unique feeding ecology of Dinophysis spp. may affect their detection by the IFCB when cells are starved.