作者想感谢 Curetis 有限公司支持这项研究。

本研究获当地道德委员会批准 (046/09, Rev. 3)。所有患者均获得知情同意和数据保密声明。
1. 共同愿望
注: 手术应在手术室或具有类似无菌条件的干预室进行。护士或医生的帮助是有帮助的, 但不是强制性的。
<ol><li>将病人放在仰卧位的检查床上。确保没有衣物的利益。使用电动剪刀缩短浓密或长体毛。不要用剃刀除去身体的毛发。如果髋关节被刺穿, 在前后方向放置一个荧光。如果膝盖被刺穿, 在病人的膝盖下放置一个膝部, 这样膝盖就会在略微弯曲的位置13。</li>
	<li>用下列无菌设备准备仪器表: 无菌拭子、内皮无菌覆盖褶皱、一次性无菌手术刀、尖头外科刀片 (型 #11) 和无菌套管的无菌10毫升注射器 (20 口径, 80 毫米)。分别设置酒精性皮肤消毒剂、采血管和儿科血培养瓶。</li>
	<li>穿上手术服、头巾和口罩, 戴上无菌手套。</li>
	<li>在穿刺部位彻底冲洗病人的皮肤 (如高级窝, 用于膝部和前外侧门区) 与皮肤消毒剂. 注: 请注意所使用的消毒剂所需的曝光时间 (通常是九十年代在膝部, 120s 在臀部为酒精消毒剂)。</li>
	<li>
用内皮不育的覆盖褶皱覆盖穿刺部位。识别穿刺部位。 注意: 对于膝盖, 正确的位置是2厘米以上的上髌骨杆和2厘米侧向侧面。对于髋关节, 找到前上棘和移动尾直到与上部联合。确保该网站是侧向股动脉 (约4厘米), 这总是可以触15。<ol>
<li>通过皮肤 (3-4 毫米宽和深), 用尖头手术刀刀片在穿刺现场做一个小刺切口。不要使用局部麻醉药, 因为它们可能在微生物培养中造成假阴性, 因为它们具有抑菌作用。</li>
	</ol>
</li>
	<li>
将套管连接到注射器的口滑。将套管插入穿刺切口。在膝关节的上凹处, 瞄准45°角的背和内侧, 朝向位于髌骨顶部下方的凹槽, 并将针插入大约5厘米的深度. 通过抽出注射器的活塞来吸取关节液。
	<ol>
<li>使用荧光指导时, 穿刺髋关节。将针插入穿刺部位, 瞄准内侧°的角度。在荧光, 确保针头尖端指向假肢颈部。在吸气时, 将套管推进到大约8厘米 (更深的脂肪患者) 的深度, 直至达到关节液。吸气时, 将针放在位置, 以避免刮去假体表面。 注: 针尖端与假体接触可确保关节内的定位。通常, 不超过5毫升的液体可以吸气形成髋关节, 膝关节将产生超过10毫升。</li>
		<li>取出套管后, 将无菌伤口敷料涂抹在刺伤切口上。为防止血肿的形成, 用绷带包扎腿部, 并轻微按压。</li>
	</ol>
</li>
	<li>
将收集到的联合吸气转移到样品容器中。确保全无菌的工作技术。
	<ol>
<li>用于小儿血培养烧瓶与吸气关节液的接种, 用酒精消毒剂对儿科血培养瓶的膜进行消毒。把新鲜的套管放到注射器上, 然后将1到3毫升的吸气液注入血液培养瓶。混合由温和的鼓动或旋转16。</li>
		<li>为准备一个本机关节抽吸标本, 取出注射器的套管。打开一个没有添加剂的采血管, 并注入1毫升的吸气到这个收集管。更换并固定盖子进行 PCR 分析。 注: 将样品及时运送到微生物学实验室进行 PCR 分析。从同一样品中, 传统的微生物有氧和厌氧培养也可以设置为17。</li>
		<li>将1至4毫升的关节抽吸物转化为含有 EDTA 的血液收集管, 用于细胞计数和分化2。将样品运送到生物化学实验室, 无任何延迟。</li>
	</ol>
</li>
</ol>2. PCR 诊断
<ol><li>确保所有三设备 (lysator、分析仪和驾驶舱; 查看材料表) 是否正常运行。列出在工作台上执行测试所需的所有用品 (PCR 盒、主混合管、取样管和取样管帽、0-200 µL 微和无菌吸管提示)。</li>
	<li>在启动程序前先解冻主混合管, 在室温下设置。所有的消耗品 (主混合管, 墨盒, 和样品管) 已经 prelabeled 与条形码。</li>
	<li>取下样品管的盖子。取180µL 收集的标本 (联合抽吸, 见1节; 或超声液, 见第4节) 与吸管和转移到样品管。将样品管与含有蛋白激酶 K 的样品管帽和内部控制基因合在一起, 用于整个工作流程的质量控制。</li>
	<li>在驾驶舱, 通过触摸左下角的 "新测试" 按钮打开操作软件。通过触摸屏输入患者的数据或样本 ID。选择标本时, 首先从下拉菜单中选择 "骨盒", 然后以 "骨科" 为应用模式, 最后选择 "滑液" 或 "超声液" 作为样本类型。按 "开始" 按钮。</li>
	<li>扫描样品管底部的条形码。将样品管插入 lysator。 注: 裂解过程将自动启动, 大约需要30分钟才能完成。在 lysator 屏幕上计时器的倒计时将显示溶解结束的时间。</li>
	<li>选择驾驶舱屏幕上的样本, 从 lysator 中解锁样品管。取出样品管从 lysator 和关闭 lysator 的顶盖。将样品管转移到 PCR 盒的取样管槽中。将解冻 mastermix 管插入 PCR 盒的 mastermix 插槽中。</li>
	<li>按照驾驶舱显示器上的说明, 用 mastermix 和取样管扫描 PCR 盒。</li>
	<li>将 PCR 墨盒插入分析器的指定墨盒插槽中。完成后, 分析器将释放墨盒。 注: PCR 条件由制造商预设, 用户不能更改。有关详细信息, 请参阅制造商的应用程序指南。PCR 过程将自动启动, 大约需要4ĥ10分钟。</li>
	<li>卸下并丢弃墨盒。在驾驶舱触摸屏上, 选择左下角的 "当前测试", 然后选取正确的样本 ID 来显示测试结果。将结果保存在计算机的内存中, 并打印或导出 (通过 USB 驱动器) 以供进一步参考。 注意: 将 PCR 的结果, 包括病原体的鉴定和抗性标志物的检测, 及时报告给外科医生。一个具有114脱氧核糖核酸 (DNA) 目标的面板, 细菌和真菌病原体通常发现在种植体感染和软组织感染, 以及最常见的抗生素抗性标志物的分析。</li>
</ol>3. 手术过程: 术中标本采集
注: 翻修手术需要专家水平的技能和专业知识;有关外科手术程序的全面概述, 请参阅外科医生的教科书18。完整和详细的外科手术的描述将远远超出了本文的范围。这里的重点是关于样品收集和 pre-analytics 的细节。在翻修手术中, 避免单次注射抗生素预防, 不损害手术过程中获得的微生物样本的结果。建议遵守此规则, 但此做法有争议19,20。只要有可能, 使用现有的手术方法到关节。额外的手术方法将损害软组织和增加疤痕的形成。
<ol><li>解剖组织直到关节囊暴露。在准备好的时候用注射器进行关节炎, 这样进一步的关节液就可以被收集到无菌注射器中, 以便进一步分析, 如上文所述 (步骤 1.7. 1-1. 7.3)。 注: 在取出植入物和组织样本之前, 不得使用防腐剂冲洗液。</li>
	<li>取出关节植入物取出后立即将植入物部分转移到一个无菌的塑料容器中, 和防水盖。 注: 在教科书文献 (膝部,如: 乔森·威尔兹et al., 16.4 章21;臀部,例如: 克拉斯et al., 章 14.5.122)。如制造商的指示所述, 可能需要具体的工具。种植体去除技术在不同种植体类型和固定方法上有很大差异。一组凿子和钻头, 滑动锤和万能髓内钉清除装置有助于去除骨性综合植入物23,24。</li>
	<li>使用无菌尖锐刮、镊子和钳, 从骨植入界面中取出膜组织。将组织的代表性样本转移到无菌的塑料容器中, 螺旋盖子, 作为微生物学和组织学标本。 注: 铰刀可用于清除所有软组织的髓质管。同时, 取滑膜组织和关节囊样本进行检查, 并将其储存在无菌容器中。总共必须收集至少四标本。</li>
	<li>立即将所有样品和说明部件送往微生物学实验室。 注: 然后可以给出抗生素。选择正确的抗生素是必不可少的, 必须是一个个性化的决定25,26。治疗概念必须由外科医生和微生物学的跨学科小组事先决定。</li>
	<li>完成前植入部位的清创清除任何组织, 显示碎片的迹象 (如聚乙烯磨损, 金属或水泥颗粒) 或感染和坏死 (化脓, 缺血性, 纤维蛋白涂层, 膜或 "泥" 组织)。取出任何已死或 osteitic 的骨头。卸下所有异物, 如螺钉、导线、cerclages、骨水泥碎片或 non-resorbable 缝合线 (参见克拉斯et al., 14.5.322)。</li>
	<li>使用50毫升的注射器来冲洗手术部位, 用伤口消毒剂进行选择。用脉冲灌洗系统充分 (至少3升) 的林格溶液冲洗。可能需要一个间隔来稳定联合27。</li>
	<li>关闭伤口使用抗菌涂层可深缝合胶囊或筋膜 (编织 polyglactin 缝合, 三氯生涂层, usp 2), 和皮下组织 (编织 polyglactin 缝合, 三氯生涂层, usp 0)。使用 non-resorbing 中断的缝合线 (monofilic 聚丙烯, usp 0 或 usp 2-0) 关闭皮肤。</li>
</ol>4. 超声
注意: 要小心工作, 严格菌。使用第二类生物安全工作台与层流气流进一步处理说明材料。
<ol><li>打开一个塑料容器, 将说明假肢从手术室 (见步骤 3.2) 置于层流气流工作台下, 并添加无菌0.9% 氯化钠溶液, 直到说明异物覆盖至少 80% (约500-800 毫升)。</li>
	<li>关闭塑料容器。在高强度的三十年代, 用涡流搅拌机彻底摇动或搅动塑料容器. 将塑料容器转到超声浴缸。检查超声浴中的液位。 注: 超声浴液水平必须与塑料容器内的相同。确保塑料容器不能被淹没, 如有必要, 在超声浴缸中添加或除去液体。</li>
	<li>几种的样本为5分钟, 频率为40±2赫, 功率密度0.22 ±0.04 瓦特/cm2。在三十年代彻底摇动或漩涡标本。 注: 超声1-5 分钟之间的时间是最好的溶解生物膜, 没有任何影响细菌的生存能力28。</li>
	<li>在层流工作台内, 收集50毫升的超声流体, 并将其转移到无菌、紧密密封的管中。</li>
	<li>要创建一个浓缩的超声流体, 离心管10分钟, 在 4200 x g. 留下10毫升的残余体积, 用吸管丢弃剩余的上清液。重移和涡流。</li>
	<li>接种合适的培养基 (如: 血琼脂, 巧克力琼脂, Sabouraud 琼脂, Schaedler 琼脂, 卡那霉素-万古霉素琼脂, 或乙酸汤), 每0.5 毫升浓缩超声液。接种的儿科血培养烧瓶2毫升如上所述 (见步骤 1.7.1)。将1毫升浓缩超声液转移到无添加剂的采血管中进行 PCR 分析。</li>
	<li>将接种的培养基转移到孵化器 (35 ° c, 5% CO2)。培养14天的文化, 每天检查生长。</li>
	<li>14天后, 丢弃没有增长的区域性并报告为负。如果检测到生长, 使用标准鉴定技术和药敏试验进行病原体鉴定。及时向外科医生报告结果。 注: 在这里, 使用基质辅助激光 desorbtion/电离-飞行时间 (MALDI) 光谱对病原体进行鉴定。使用半自动分析仪29、30、31进行抗菌药敏试验。</li>
</ol>

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J., Patel, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prosthetic+joint+infection.">Prosthetic joint infection.</a> Clin Microbiol Rev. 27 (2), 302-345 (2014).</li><li>Portillo, M. 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作者没有什么可透露的。

异物感染是骨科和创伤外科中的一个新兴问题, 其治疗费用昂贵, 住院时间长, 受累关节功能缺损。鉴别诊断具有挑战性。许多研究人员和临床医生正在关注这个课题, 努力寻找更精确和可靠的方法来诊断异物相关感染。到目前为止, 在诊断路径32中正在评估和实现许多不同的诊断工具。
超声程序是一种无价的方法, 它比传统的组织标本6具有更好的灵敏度。在我们的研究中, 我们发现超声流体培养物的敏感性为 88.9%, 特异性为61.5%。从组织标本和关节物的传统文化都具有66.7% 的敏感性, 其特异性分别为82.3% 和84.6%。其他研究人员显示了类似的结果, 这些常规的微生物学程序6,33,34,35。经常有人批评超声程序容易受到污染, 因此在处理样品时必须特别注意。
在组织标本或体液中检测致病病原体的 DNA 的可能性是耐人寻味的。NAT 是一个快速的过程, 可以提供几个小时内的结果, 与耗时的微生物培养方法相比。毫无疑问, NAT 在检测病原体方面具有很好的灵敏度, 但却持有检测污染物的风险, 因此缺乏特异性36。这种 PCR 技术在诊断 PJI 的巨大好处, 特别是在接受抗生素治疗接近手术或长期7,8的患者中得到了记录。研究表明, 超声液的 NAT 可能进一步增加灵敏度和特异性37。不幸的是, 由于耗时的工作流程, 这项技术在实验室中并不经常使用。
PCR 技术在一般情况下有一定的局限性: NAT 检测到的 DNA 在生存和不细菌之间没有区别, 使得解释结果变得困难。广范围 PCR 将只检测核糖体16S 核糖核酸 (16S rRNA), 而不是区分病原体37。更具体的系统并不例行检测基因编码的抗生素耐药性标记。因此, 有针对性的抗生素治疗可能会受到限制, 没有进一步的敏感性测试。
本研究的系统克服了这些局限性, 区分特定细菌和识别基因编码的抵抗标记。总的来说, NAT 检测可以被认为是一个快速和有用的互补, 以确认 PJI7,8,9,38。
有必要在联合吸入和手术中预先计划: 样品容器必须是无菌的和准备好的, 并且迅速样品运输必须是可利用的。外科医生应该对计划中的程序和期望的样品材料微生物学。在进行联合吸入时, 必须严格消毒条件, 以防止医源性感染关节。在脂肪患者中, 联合穿刺可能是有挑战性的, 透视指导可以是有益的。翻修手术需要非常高水平的专业技能, 而且应该由熟练的外科医生来完成。说明材料不应留在手术室任何时间超过必要, 但被送往微生物学尽快。本议定书中的一个非常关键的部分是潜在的污染风险。不仅在采集样品的同时, 在处理微生物实验室的样品时, 所有涉及的人员 (整形外科医生、护士、技术员、微生物学) 必须快速准确地工作, 并在程序中进行适当的培训。
超声和 NAT 是诊断种植体相关感染的重要工具。然而, 这一结果应始终受到认真的质疑。我们建议在圆桌讨论中讨论骨科医生、微生物学和传染性疾病专家以及病理学家对个体化治疗策略的看法。
在 PJI 的诊断路径中实现这一技术可以有几个优点。这是一个快速诊断, 结果在几个小时内。由于分析了几种基因编码的抗性标记, 有针对性的抗生素治疗可以发生在很早的阶段, 在临床病程。作为一个副作用, 广泛的抗生素可以保存在他们真正需要的迹象。其他样本类型 (例如组织标本、拭子、血肿、等) 也可以根据我们的协议进行调查。由于 PCR 磁带是一个闭合的系统, 因此在用户端不需要或可能进行故障排除。该协议的任何修改都必须由制造商来执行。制造商不断地对软件和硬件 (墨盒) 进行更改, 以增强系统, 但是在更新版本的确切更改中没有公开任何细节。

痛苦的置换是骨科手术中的一个诊断挑战。无菌性松动后, PJI 是植入失败的第二大原因, 并在关节置换术后出现了严重的并发症。PJI 难以治疗, 难以诊断。漏诊的 PJI 可能会导致复发性感染, 主要的发病率, 功能丧失和生活质量的丧失。因此, 在开始治疗措施之前, 所有在疼痛的关节置换术中都应排除感染。
患者病史、临床检查、CRP 血检和白细胞计数, 以及受影响关节的造影或 szintigraphy 构成基本诊断1。术前常规也应包括在无菌条件下的联合吸入。获得的吸出物是进一步诊断的非常有价值的材料。
除了细胞计数和细胞分化的联合抽吸作为行之有效的化验2, 检测蛋白质生物标志物可能有助于鉴别诊断3,4,5。传统的微生物培养仍然是病原体检测的金标准。生物膜, 由 gram-positive 和 gram-negative 细菌, 并附着于种植体表面, 是一个主要的致病因素, 在 PJI。因此, 在 2007年, 在骨科手术中对异物相关感染的诊断中实施了超声程序, 扰乱了去除植入物的生物膜, 从而允许病原体检测。细菌从而从他们的休息的形式被采取到一个活跃形式, 再做检测在文化可能。超声去除的植入物显示出比组织标本 (78.5% 与 60.8%)6更高的灵敏度。
核酸扩增试验 (NAT), 如 PCR, 最近已进入范围的临床医生和微生物学诊断 PJI。特别是在手术前接受抗生素治疗的患者中, 发现 PCR 诊断有利于识别致病生物7,8,9。最近, 一个新的多重 PCR 系统, 专门设计的植入和组织感染 (), 介绍了。该 cartridge-based 系统提供了多种基因标记, 用于鉴定病原体和抗生素耐药性标记。在它的应用范围是许多征兆 (假肢联合传染, 外科站点传染, 心脏病相关的传染, 导管伴生的传染, 糖尿病脚传染, 深皮肤和组织传染, 植入物传染,和烧伤伤口感染), 和一个广泛的样本材料面板可用于此技术 (超声液, 滑液, 拭子, 组织, 脓, 吸/渗出物, 和生物膜)10,11,12。
与常规微生物学相比, 该程序的最大优点是速度: 致病病原体可在数小时内确定。此外, 这个 PCR 系统检测到一个广泛的基因编码的抗性标记的面板, 允许启动靶向抗生素治疗早期。在 PCR 的协助下, 外科医生可以在诊断程序11的早期阶段区分感染和感染患者。在这里, 我们提出的协议来执行这个多重 PCR, 快速诊断 PJI 从联合吸气和超声液。

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			<td height="42" style="height:42px;width:216px;">sterile plastic container&#160;</td>
			<td style="width:333px;">Lock &#38; Lock, Distributor ISI GmbH, Solingen, Germany</td>
			<td style="width:119px;">EAN 8803733184403</td>
			<td style="width:232px;">Transport container for implants</td>
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			<td height="21" style="height:21px;">Bactosonic 14.2&#160;</td>
			<td>Bactosonic, Bandelin, Berlin, Germany</td>
			<td align="right" style="width:119px;">3290</td>
			<td>&#34;Sonication machine&#34;</td>
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			<td height="21" style="height:21px;width:216px;">50ml Falcon tubes</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">352070</td>
			<td>Centrifugation</td>
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			<td height="21" style="height:21px;">PEDS medium blood culture flasks&#160;</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">442194</td>
			<td>culture medium</td>
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			<td height="21" style="height:21px;">Columbia agar with 5% sheep blood</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">254071</td>
			<td>culture medium</td>
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			<td height="21" style="height:21px;">Mac Conkey agar</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">254078</td>
			<td>culture medium</td>
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			<td height="21" style="height:21px;">chocolate agar&#160;</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">254089</td>
			<td>culture medium</td>
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			<td height="21" style="height:21px;">sabouraud agar&#160;</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">254096</td>
			<td>culture medium</td>
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			<td height="21" style="height:21px;">thioglycolate bouillon&#160;</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">221788</td>
			<td>culture medium</td>
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			<td height="21" style="height:21px;">Schaedler agar</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">254084</td>
			<td>culture medium</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">kanamycin/vancomycin agar</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td align="right" style="width:119px;">254077</td>
			<td>culture medium</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bactec FX blood culture system&#160;</td>
			<td>Becton &#38; Dickinson, Heidelberg, Germany</td>
			<td style="width:119px;">441386/441385</td>
			<td>culture medium</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Unyvero A50 Analyzer</td>
			<td style="width:333px;">Curetis, Holzgerlingen, Germany</td>
			<td align="right" style="width:119px;">60001</td>
			<td>PCR machine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Unyvero L4 Lysator</td>
			<td style="width:333px;">Curetis, Holzgerlingen, Germany</td>
			<td align="right" style="width:119px;">60002</td>
			<td>PCR machine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Unyvero C8 Cockpit</td>
			<td style="width:333px;">Curetis, Holzgerlingen, Germany</td>
			<td align="right" style="width:119px;">60003</td>
			<td>PCR machine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Unyvero M1 Masterix Tube</td>
			<td style="width:333px;">Curetis, Holzgerlingen, Germany</td>
			<td align="right" style="width:119px;">10002</td>
			<td>consumables PCR</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Unyvero i60 ITI Cartridge</td>
			<td style="width:333px;">Curetis, Holzgerlingen, Germany</td>
			<td align="right" style="width:119px;">10040</td>
			<td>consumables PCR</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Unyvero Sample Tube Cap</td>
			<td style="width:333px;">Curetis, Holzgerlingen, Germany</td>
			<td align="right" style="width:119px;">10004</td>
			<td>consumables PCR</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Unyvero Sample Tube</td>
			<td style="width:333px;">Curetis, Holzgerlingen, Germany</td>
			<td align="right" style="width:119px;">10003</td>
			<td>consumables PCR</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">S-Monovette 2.7ml</td>
			<td style="width:333px;">Sarstedt AG, N&#252;mbrecht, Germany</td>
			<td style="width:119px;">05.1729.001&#160;</td>
			<td>Transport container (aspirate)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">scalpel typ 11</td>
			<td style="width:333px;">pfm medical, Cologne, Germany</td>
			<td align="right">200130011</td>
			<td>scalpel for stab incision</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">PP Container 70mL 55x45mm</td>
			<td style="width:333px;">Sarstedt AG, N&#252;mbrecht, Germany</td>
			<td align="right">759,922,721</td>
			<td>Transport container (tissue specimen)</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Vicryl Plus</td>
			<td style="width:333px;">Johnson&#38;Johnson Medical GmbH, Ethicon Germany, Norderstedt, Germany</td>
			<td>VCP247H</td>
			<td>surgical suture material</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Prolene 0</td>
			<td style="width:333px;">Johnson&#38;Johnson Medical GmbH, Ethicon Germany, Norderstedt, Germany</td>
			<td>EH7920H</td>
			<td>surgical suture material</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Prolene 2/0</td>
			<td style="width:333px;">Johnson&#38;Johnson Medical GmbH, Ethicon Germany, Norderstedt, Germany</td>
			<td>EH7697H</td>
			<td>surgical suture material</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Kodan&#160; Tinktur forte farblos</td>
			<td style="width:333px;">Sch&#252;lke &#38; Mayr GmbH, Norderstedt, Germany</td>
			<td align="right" style="width:119px;">104005</td>
			<td>alcoholic skin disinfectant</td>
		</tr>
	</tbody>
</table>

novel diagnostics in revision arthroplasty: implant sonication and multiplex polymerase chain reaction

在骨科患者, 异物相关感染, 特别是周围关节感染 (PJIs), 是一个破坏性的并发症的关节置换术。感染需要复杂的治疗, 可能导致长期住院, 并造成相当大的成本。多项外科手术的修订是必要的, 在这些病人, 在功能和生活质量的损失。
常规术前诊断包括 C 反应蛋白 (CRP) 和其他生物标志物的血液检查, 以及细胞计数、分化和培养的联合吸气分析。术中标本的组织学和微生物学也是标准的程序。超声的微生物检查, 结合分子生物学技术在微生物学中的实施, 是目前用于增强 PJI 鉴别诊断的两种新技术。
本文采用 cartridge-based 多重聚合酶链反应 (PCR) 系统对关节吸气和超声液进行了逐步过程分析。结果与传统文化和 PJI 的共识标准相匹配。从组织活检、联合吸气和超声液中得到的传统微生物培养物分别表现出66.7%、66.7% 和88.9% 的敏感性, 分别为82.3%、54.6% 和61.5% 的特异性。超声液和关节液的 PCR 诊断分别表现为50.0% 和55.6% 的敏感性, 特异性为100.0%。两种 PCR 诊断结合的灵敏度为 66.7%, 特异性为100.0%。因此, 多重 PCR 技术在诊断 PJI 中具有灵敏度高、特异性强的快速诊断工具。

PJI 的存在, 由肌肉骨骼感染协会 (msi) 的共识会议确定, 当一个主要标准或至少三五次要标准存在时, 被认为被证明。主要的标准包括在两个单独的微生物样本中存在瘘管或检测病原体。小的标准包括阳性细胞计数 (> 3000 白细胞/µL) 或阳性细胞分化 (> 85% 中性粒细胞粒) 在联合抽吸, 阳性 CRP 和沉淀率的血液样本, 阳性组织学在组织样品或只在一个微生物样本中检测到一个病原体19。与 msi 金标准相比, 核酸扩增试验 (NAT) 的灵敏度和特异性得到了计算。患者详细信息 (包括检测到的病原体) 在表 1中给出。
我们自己的结果显示, 对超声液和关节抽吸 (50.0% 和 55.6%) 的 PCR 诊断有适度的敏感性, 但非常强的特异性 100%11。每当 PCR 诊断 (总氮 = 62 测试) 显示在联合吸气 (n = 31) 或超声液 (n = 31) 阳性发现, 同样的病原体被发现后, 在一个传统的微生物文化。感染病例的不同样本的 PCR 显示没有假阳性结果 (参见图 1)。
PCR 诊断未能检测到传统微生物培养方法证实的一些病原体。6出 16 (37.0%) 真病原体中的超声液样品和5出 13 (38.0%) 病原体从联合液培养中被错过了 PCR 检测。大多数情况下, PCR 诊断漏诊凝固酶阴性葡萄球菌 (8 出 11)。两种材料联合 pcr 诊断 (联合抽吸和超声液, "汇集 pcr") 正确地辨认了12在18传染案件中 (敏感性 66.7%, 95% CI: 41.0% 到 86.7%, 参见表 2)。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/55147/55147fig1.jpg"> 图 1:NAT 结果摘要图中描述了集体病人样本的总结结果。右侧是组患者匹配的 PJI 标准, 左侧是组没有。这些棒代表了核酸扩增的方法。假阳性和假阴性以红色表示, 占总数的百分比。<a href="http://cloudflare.jove.com/files/ftp_upload/55147/55147fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<table fo:keep-together.within-page="1"><tbody><tr><td>联合</td>
			<td>修订原因</td>
			<td>是否符合 msi 标准？</td>
			<td>以前的抗生素治疗？</td>
			<td>超声流体文化</td>
			<td>联合吸气培养</td>
			<td>超声文化</td>
			<td>联合吸气</td>
		</tr><tr><td>髋关节</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td>Dermabacter 人</td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td>宿根 aquatica</td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>髋关节</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>不</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>髋关节</td>
			<td>慢性脱位</td>
			<td>不</td>
			<td>不</td>
			<td>葡萄球菌 溶血</td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>髋关节</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>髋关节</td>
			<td>无菌松动</td>
			<td>不</td>
			<td>不</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>不</td>
			<td>不</td>
			<td>不</td>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>急性感染</td>
			<td>是的</td>
			<td>是的</td>
			<td>铜绿假单胞菌</td>
			<td>铜绿假单胞菌</td>
			<td></td>
			<td>胞 铜绿</td>
		</tr><tr><td>膝盖</td>
			<td>急性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>大肠杆菌 大肠杆菌</td>
			<td>大肠杆菌 大肠杆菌</td>
			<td>大肠杆菌 大肠杆菌</td>
			<td></td>
		</tr><tr><td>髋关节</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>Corynebakterium 属.</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>急性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色葡萄球菌</td>
			<td>金黄色葡萄球菌</td>
			<td>金黄色葡萄球菌</td>
			<td>葡萄球菌 葡萄球菌</td>
		</tr><tr><td>膝盖</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>链球菌 agalacticae</td>
			<td>链球菌 agalacticae</td>
			<td>链球菌 agalacticae</td>
			<td>链球菌 agalacticae</td>
		</tr><tr><td>髋关节</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>是的</td>
			<td></td>
			<td>金黄色葡萄球菌</td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>是的</td>
			<td>Staphlococcu 葡萄球菌</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td></td>
		</tr><tr><td>髋关节</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色 葡萄球菌</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td>凝固酶阴性 葡萄球菌</td>
			<td>凝固酶阴性 葡萄球菌</td>
		</tr><tr><td>髋关节</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色葡萄球菌</td>
			<td></td>
			<td>金黄色葡萄球菌</td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>急性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色 葡萄球菌</td>
			<td>金黄色 葡萄球菌</td>
			<td>凝固酶阴性 葡萄球菌</td>
			<td>凝固酶阴性 葡萄球菌</td>
		</tr><tr><td>髋关节</td>
			<td>急性感染</td>
			<td>是的</td>
			<td>不</td>
			<td></td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td>凝固酶阴性 葡萄球菌</td>
		</tr><tr><td>髋关节</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色 葡萄球菌</td>
			<td>金黄色 葡萄球菌</td>
			<td></td>
			<td></td>
		</tr><tr><td>髋关节</td>
			<td>急性感染</td>
			<td>是的</td>
			<td>不</td>
			<td></td>
			<td>金黄色 葡萄球菌</td>
			<td>凝固酶阴性 葡萄球菌</td>
			<td></td>
		</tr><tr><td>膝盖</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>金黄色葡萄球菌</td>
			<td>金黄色葡萄球菌</td>
			<td></td>
			<td>葡萄球菌 葡萄球菌</td>
		</tr><tr><td>髋关节</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>不</td>
			<td>球菌 faecialis</td>
			<td>球菌 faecialis</td>
			<td>球菌 属.</td>
			<td>球菌 属.</td>
		</tr><tr><td>膝盖</td>
			<td>慢性感染</td>
			<td>是的</td>
			<td>是的</td>
			<td>Enterocuccus 球菌</td>
			<td>Enterocuccus 球菌</td>
			<td>球菌 属.</td>
			<td>球菌 属.</td>
		</tr></tbody></table>表 1: 患者详细信息和检测到的病原体.患者细节的表格结果, 包括修订手术的原因、msi 标准的匹配以及常规微生物学和 NAAT 分析中检测到的病原体。
<table fo:keep-together.within-page="1"><tbody>
<tr>
<td>标准</td>
			<td>PCR 几种</td>
			<td>PCR 吸气</td>
			<td>聚合 PCR</td>
		</tr>
<tr>
<td>P 值</td>
			<td>0.0036</td>
			<td>0.0013</td>
			<td>0.0001</td>
		</tr>
<tr>
<td>敏感性</td>
			<td>50.0%</td>
			<td>55.6%</td>
			<td>66.7%</td>
		</tr>
<tr>
<td>特异</td>
			<td>100.0%</td>
			<td>100.0%</td>
			<td>100.0%</td>
		</tr>
<tr>
<td>正向预测值</td>
			<td>100.0%</td>
			<td>100.0%</td>
			<td>100.0%</td>
		</tr>
<tr>
<td>负预测值</td>
			<td>59.1%</td>
			<td>61.9%</td>
			<td>68.4%</td>
		</tr>
</tbody></table>表 2: 微生物方法的结果.对超声液、联合抽吸和聚合 pcr 的 pcr 诊断进行了评价的表格结果。n = 31 个样品为抽吸和超声, 分别, n = 62 为汇集的 PCR 数据。

Watch this Scientific Journal Video about Novel Diagnostics in Revision Arthroplasty: Implant Sonication and Multiplex Polymerase Chain Reaction at JoVE.com

Novel Diagnostics in Revision Arthroplasty: Implant Sonication and Multiplex Polymerase Chain Reaction

疼痛关节置换术的鉴别诊断是治疗成功的关键。术后常规进行关节穿刺。本文介绍了联合抽吸和超声液的多重聚合酶链反应, 为这些样品的快速病原检测提供了一种可行的工具。

翻修术中的新诊断: 种植超声和多重聚合酶链反应

In orthopedic patients, foreign body-associated infections, especially periprosthetic joint infections (PJIs), are a devastating complication of ...

novel-diagnostics-revision-arthroplasty-implant-sonication-multiplex

Research

JoVE Journal

Medicine

11.0K 次观看.  University Hospital Bonn. 疼痛关节置换术的鉴别诊断是治疗成功的关键。术后常规进行关节穿刺。本文介绍了联合抽吸和超声液的多重聚合酶链反应, 为这些样品的快速病原检测提供了一种可行的工具。

视频: Novel Diagnostics in Revision Arthroplasty: Implant Sonication and Multiplex Polymerase Chain Reaction

Reverse Total Shoulder Arthroplasty

Reverse total shoulder arthroplasty was initially approved for use in rotator cuff arthropathy and well as chronic pseudoparalysis without arthritis in patients who were not appropriate for tendon transfer reconstructions. Traditional surgical options for these patients were limited and functional results were sub-optimal and at times catastrophic. The use of reverse shoulder arthroplasty has been found to effectively restore these patients function and relieve symptoms associated with their disease. The procedure can be done through two approaches, the deltopectoral or the superolateral. Complication rates associated with the use of the prosthesis have ranged from 8-60% with more recent reports trending lower as experienced is gained. Salvage options for a failed reverse shoulder prosthesis are limited and often have significant associated disability. Indications for the use of this prosthesis continue to be evaluated including its use for revision arthroplasty, proximal humeral fracture and tumor. Careful patient selection is essential because of the significant risks associated with the procedure.

Reverse total shoulder arthroplasty is indicated for the treatment of conditions that cannot be treated with conventional arthroplasty or other procedures. These primarily include degenerative and incapacitating conditions with irreparable rotator cuff and loss of the normal biomechanical coupling of the shoulder.

反向共肩关节置换术

Reverse total shoulder arthroplasty was initially approved for use in rotator cuff arthropathy and well as chronic pseudoparalysis without arthritis in ...

科研

医学

RNAscope for In situ Detection of Transcriptionally Active Human Papillomavirus in Head and Neck Squamous Cell Carcinoma

The &#39;gold standard&#39; for oncogenic HPV detection is the demonstration of transcriptionally active high-risk HPV in tumor tissue. However, detection of E6/E7 mRNA by quantitative reverse transcription polymerase chain reaction (qRT-PCR) requires RNA extraction which destroys the tumor tissue context critical for morphological correlation and has been difficult to be adopted in routine clinical practice. Our recently developed RNA in situ hybridization technology, RNAscope, permits direct visualization of RNA in formalin-fixed, paraffin-embedded (FFPE) tissue with single molecule sensitivity and single cell resolution, which enables highly sensitive and specific in situ analysis of any RNA biomarker in routine clinical specimens. The RNAscope HPV assay was designed to detect the E6/E7 mRNA of seven high-risk HPV genotypes (HPV16, 18, 31, 33, 35, 52, and 58) using a pool of genotype-specific probes. It has demonstrated excellent sensitivity and specificity against the current &#39;gold standard&#39; method of detecting E6/E7 mRNA by qRT-PCR. HPV status determined by RNAscope is strongly prognostic of clinical outcome in oropharyngeal cancer patients.

RNAscope for In situ Detection of Transcriptionally Active Human Papillomavirus in Head and Neck Squamous Cell Carcinoma

The presence of high-risk HPV in head and neck tumor tissue is associated with favorable outcomes. The recently developed RNA in situ hybridization technique called RNAscope allows direct visualization of HPV E6/E7 mRNA in FFPE tissue sections.

RNAscope的<em&gt;原位</em&gt;检测转录活性的人类乳头状瘤病毒在头颈部鳞状细胞癌

The &#39;gold standard&#39; for oncogenic HPV detection is the demonstration of transcriptionally active high-risk HPV in tumor tissue. However, detection ...

Combined In vivo Optical and &#181;CT Imaging to Monitor Infection, Inflammation, and Bone Anatomy in an Orthopaedic Implant Infection in Mice

Multimodality imaging has emerged as a common technological approach used in both preclinical and clinical research. Advanced techniques that combine in vivo optical and &#956;CT imaging allow the visualization of biological phenomena in an anatomical context. These imaging modalities may be especially useful to study conditions that impact bone. In particular, orthopaedic implant infections are an important problem in clinical orthopaedic surgery. These infections are difficult to treat because bacterial biofilms form on the foreign surgically implanted materials, leading to persistent inflammation, osteomyelitis and eventual osteolysis of the bone surrounding the implant, which ultimately results in implant loosening and failure. Here, a mouse model of an infected orthopaedic prosthetic implant was used that involved the surgical placement of a Kirschner-wire implant into an intramedullary canal in the femur in such a way that the end of the implant extended into the knee joint. In this model, LysEGFP mice, a mouse strain that has EGFP-fluorescent neutrophils, were employed in conjunction with a bioluminescent Staphylococcus aureus strain, which naturally emits light. The bacteria were inoculated into the knee joints of the mice prior to closing the surgical site. In vivo bioluminescent and fluorescent imaging was used to quantify the bacterial burden and neutrophil inflammatory response, respectively. In addition, &#956;CT imaging was performed on the same mice so that the 3D location of the bioluminescent and fluorescent optical signals could be co-registered with the anatomical &#956;CT images. To quantify the changes in the bone over time, the outer bone volume of the distal femurs were measured at specific time points using a semi-automated contour based segmentation process. Taken together, the combination of in vivo bioluminescent/fluorescent imaging with &#956;CT imaging may be especially useful for the noninvasive monitoring of the infection, inflammatory response and anatomical changes in bone over time.

Combined In vivo Optical and  &#181;CT Imaging to Monitor Infection, Inflammation, and Bone Anatomy in an Orthopaedic Implant Infection in Mice

Combined optical and &#956;CT imaging in a mouse model of orthopaedic implant infection, utilizing a bioluminescent engineered strain of Staphylococcus aureus, provided the capability to noninvasively and longitudinally monitor the dynamics of the bacterial infection, as well as the corresponding inflammatory response and anatomical changes in the bone.

结合<em&gt;在体内</em&gt;光学及μCT成像监视感染，炎症和骨解剖在骨科植入物感染的小鼠

Multimodality imaging has emerged as a common technological approach used in both preclinical and clinical research. Advanced techniques that combine in ...

Individualized Stem-positioning in Calcar-guided Short-stem Total Hip Arthroplasty

Bone- and soft-tissue sparing short stems are increasingly used in total hip arthroplasty (THA). However, there are a large variety of models of short stems, differing in design and function. Calcar-guided short stems provide an anatomical curvature in the medial calcar region, thus, positioning is done individually alongside the calcar in the "round-the-corner" technique. Depending on the level of the neck's osteotomy, stems can be aligned individually in a large bandwidth of varus- and valgus anatomies. This differs from conventional total hip arthroplasty and potentially includes a severe learning curve. Given that a great variety of caput-collum-diaphyseal (CCD)-angles can be retained, the reconstruction of femoro-acetabular offsets can be achieved precisely. However, particularly extensive varus- and valgus positioning has raised concerns in regard to stability and bone remodeling. The purpose of the present manuscript is to showcase the implantation technique in calcar-guided short-stem THA and to summarize short-term clinical and radiological results.

This protocol describes the round-the-corner technique and the individualized stem-positioning of calcar-guided short stems alongside the medial calcar, depending on the level of the osteotomy. This differs from conventional total hip arthroplasty and includes a learning curve.

股骨引导下短茎全髋关节置换术的个体化茎定位

Bone- and soft-tissue sparing short stems are increasingly used in total hip arthroplasty (THA). However, there are a large variety of models of short ...

A Blood-based Test for the Detection of ROS1 and RET Fusion Transcripts from Circulating Ribonucleic Acid Using Digital Polymerase Chain Reaction

We have developed novel methods for the isolation and characterization of tumor-derived circulating ribonucleic acid (cRNA) for blood-based liquid biopsy. Robust detection of cRNA recovered from blood represents a solution to a critical unmet need in clinical diagnostics. The test begins with the collection of whole blood into blood collection tubes containing preservatives that stabilize cRNA. Cell-free, exosomal, and platelet-associated RNA is isolated from plasma in this test system. The cRNA is reverse transcribed to complementary DNA (cDNA) and amplified using digital polymerase chain reaction (dPCR). Samples are evaluated for both the target biomarker as well as a control gene. Test validation included limit of detection, accuracy, and robustness studies with analytic samples. The method developed as a result of these studies reproducibly detect multiple fusion variants for ROS1 (C-Ros proto-oncogene 1; 8 variants) and RET (rearranged during transfection proto-oncogene; 8 variants). The sample processing workflow has been optimized so that test results can consistently be generated within 72 hours of sample receipt.

Detection of circulating ribonucleic acid (cRNA) from blood is an unmet need in clinical diagnostics. Here we describe methods that characterize cRNA from non-small cell lung cancer patients using sensitive and specific digital polymerase chain reaction. The tests meet design requirements to detect fusion variants within 72 hours.

用数字聚合酶链反应检测循环核糖核酸 ROS1 和 RET 融合记录的血液检测方法

We have developed novel methods for the isolation and characterization of tumor-derived circulating ribonucleic acid (cRNA) for blood-based liquid biopsy. ...

Remote Laboratory Management: Respiratory Virus Diagnostics

An uptick in recent pandemics (Ebola, Zika, MERS, influenza, etc.) underlines the need for a more 'nimble,' coordinated response that addresses a multitude of issues ranging from transportation, access, facilities, equipment, and communication to provider training. To address this need, we have developed an innovative, scalable, logistics-enhanced, mobile, laboratory facility for emergencies and epidemics in resource-constrained global settings. Utilizing a background in clinical operations as an academic medical center, we designed a rapidly-deployable, modular BSL-2 and BSL-3 facility with user-friendly software for tracking and management of drugs and supplies in remote regions during epidemics and outbreaks. Here, we present our intermodal, mobile, expandable shipping-container laboratory units. The design of the laboratory facilitates off-grid usage by minimizing power consumption and allowing alternate water sources. The unit's information communication technology (ICT) platform provides (i) user-friendly tablet-based documentation, (ii) enhanced tracking of patients and supplies, and (iii) integrated communication onsite with built-in telehealth capabilities. To ensure quality in remote environments, we have developed a checklist for a basic laboratory workflow and a protocol for respiratory viral diagnosis using reverse-transcription polymerase chain reaction (RT-PCR). As described, this innovative and comprehensive approach allows for the provision of laboratory capability in resource-limited global environments.

A rapidly-deployable, off-grid laboratory has been designed and built for remote, resource-constrained global settings. The features and critical aspects of the logistically-enhanced, expandable, multifunctional laboratory modules are explored. A checklist for a basic laboratory workflow and a protocol for a respiratory viral diagnostic test are developed and presented.

远程实验室管理: 呼吸道病毒诊断

An uptick in recent pandemics (Ebola, Zika, MERS, influenza, etc.) underlines the need for a more 'nimble,' coordinated response that addresses a ...

In Vivo Mouse Model of Spinal Implant Infection

Spine implant infections portend poor outcomes as diagnosis is challenging and surgical eradication is at odds with mechanical spinal stability. The purpose of this method is to describe a novel mouse model of spinal implant infection (SII) that was created to provide an inexpensive, rapid, and accurate in vivo tool to test potential therapeutics and treatment strategies for spinal implant infections.
In this method, we present a model of posterior-approach spinal surgery in which a stainless-steel k-wire is transfixed into the L4 spinous process of 12-week old C57BL/6J wild-type mice and inoculated with 1 x 103 CFU of a bioluminescent strain of Staphylococcus aureus Xen36 bacteria. Mice are then longitudinally imaged for bioluminescence in vivo on post-operative days 0, 1, 3, 5, 7, 10, 14, 18, 21, 25, 28, and 35. Bioluminescence imaging (BLI) signals from a standardized field of view are quantified to measure in vivo bacterial burden.
To quantify bacteria adhering to implants and peri-implant tissue, mice are euthanized and the implant and surrounding soft tissue are harvested. Bacteria are detached from the implant by sonication, cultured overnight and then colony forming units (CFUs) are counted. The results acquired from this method include longitudinal bacterial counts as measured by in vivo S. aureus bioluminescence (mean maximum flux) and CFU counts following euthanasia.
While prior animal models of instrumented spine infection have involved invasive, ex vivo tissue analysis, the mouse model of SII presented in this paper leverages noninvasive, real time in vivo optical imaging of bioluminescent bacteria to replace static tissue study. Applications of the model are broad and may include utilizing alternative bioluminescent bacterial strains, incorporating other types of genetically engineered mice to contemporaneously study host immune response, and evaluating current or investigating new diagnostic and therapeutic modalities such as antibiotics or implant coatings.

The protocol describes a novel in vivo mouse model of spinal implant infection where a stainless-steel k-wire implant is infected with bioluminescent Staphylococcus aureus Xen36. Bacterial burden is monitored longitudinally with bioluminescent imaging and confirmed with colony forming unit counts after euthanasia.

脊柱植入物感染的体内小鼠模型

Spine implant infections portend poor outcomes as diagnosis is challenging and surgical eradication is at odds with mechanical spinal stability. The ...

Dynamic Navigation for Dental Implant Placement

In modern implantology, the application of surgical navigation systems is becoming increasingly important. In addition to static surgical navigation methods, a guide-independent dynamic navigation implant placement procedure is becoming more widespread. The procedure is based on computer-guided dental implant placement utilizing optical control. This work aims to demonstrate the technical steps of a new dynamic computer-aided implant surgery (DCAIS)&#160;system (design, calibration, surgery) and check the accuracy of the results. Based on cone-beam computed tomography (CBCT) scans, the exact positions of implants are determined with dedicated software. The first step of the operation is the calibration of the navigation system, which can be performed in two ways: 1) based on CBCT images taken with a marker or 2) based on CBCT images without markers. Implants are inserted with the aid of real-time navigation according to the preoperative plans. The accuracy of the interventions can be evaluated based on postoperative CBCT images. The preoperative images containing the planned positions of the implants and postoperative CBCT images were compared based on the angulation (degree), platform, and apical deviation (mm) of the implants. To evaluate the data, we calculated the standard deviation (SD), mean, and standard error of the mean (SEM) of deviations within planned and performed implant positions. Differences between the two calibration methods were compared based on this data. Based on the interventions performed so far, the use of DCAIS allows for high-precision implant placement. A calibration system that does not require labeled CBCT recording allows for surgical intervention with similar accuracy as a system that uses labeling. The accuracy of the intervention can be improved by training.

Dynamic computer-aided implant surgery (DCAIS) is a controlled implant surgical placement method performed without a surgical template using optical control. The real-time intraoperative control of movement and position of the surgical device simplifies the procedure and gives more freedom to the surgeon, providing similar precision as static navigation methods.

种植牙的动态导航

In modern implantology, the application of surgical navigation systems is becoming increasingly important. In addition to static surgical navigation ...

The Use of Mixed Reality in Custom-Made Revision Hip Arthroplasty: A First Case Report

The technology of 3D printing and visualization of anatomical structures is rapidly growing in various fields of medicine. A custom-made implant and mixed reality were used to perform complex revision hip arthroplasty in January 2019. The use of mixed reality allowed for a very good visualization of the structures and resulted in precise implant fixation. According to the authors' knowledge, this is the first described case report of the combined use of these two innovations. The diagnosis preceding the qualification for the procedure was the loosening of the left hip's acetabular component. Mixed reality headset and holograms prepared by engineers were used during the surgery. The operation was successful, and it was followed by early verticalization and patient rehabilitation. The team sees opportunities for technology development in joint arthroplasty, trauma, and orthopedic oncology.

A complex revision hip arthroplasty was performed using a custom-made implant and mixed reality technology. According to the authors&#39; knowledge, this is the first report of such procedure described in the literature.

混合现实在定制翻修髋关节置换术中的应用:第一份病例报告

The technology of 3D printing and visualization of anatomical structures is rapidly growing in various fields of medicine. A custom-made implant and mixed ...

Measuring Single-Cell Mitochondrial DNA Copy Number and Heteroplasmy Using Digital Droplet Polymerase Chain Reaction

The mammalian mitochondrial (mt)DNA is a small, circular, double-stranded, intra-mitochondrial DNA molecule, encoding 13 subunits of the electron transport chain. Unlike the diploid nuclear genome, most cells contain many more copies of mtDNA, ranging from less than 100 to over 200,000 copies depending on cell type. MtDNA copy number is increasingly used as a biomarker for a number of age-related degenerative conditions and diseases, and thus, accurate measurement of the mtDNA copy number is becoming a key tool in both research and diagnostic settings. Mutations in the mtDNA, often occurring as single nucleotide polymorphisms (SNPs) or deletions, can either exist in all copies of the mtDNA within the cell (termed homoplasmy) or as a mixture of mutated and WT mtDNA copies (termed heteroplasmy). Heteroplasmic mtDNA mutations are a major cause of clinical mitochondrial pathology, either in rare diseases or in a growing number of common late-onset diseases such as Parkinson's disease. Determining the level of heteroplasmy present in cells is a critical step in the diagnosis of rare mitochondrial diseases and in research aimed at understanding common late-onset disorders where mitochondria may play a role. MtDNA copy number and heteroplasmy have traditionally been measured by quantitative (q)PCR-based assays or deep sequencing. However, the recent introduction of ddPCR technology has provided an alternative method for measuring both parameters. It offers several advantages over existing methods, including the ability to measure absolute mtDNA copy number and sufficient sensitivity to make accurate measurements from single cells even at low copy numbers. Presented here is a detailed protocol describing the measurement of mtDNA copy number in single cells using ddPCR, referred to as droplet generation PCR henceforth, with the option for simultaneous measurement of heteroplasmy in cells with mtDNA deletions. The possibility of expanding this method to measure heteroplasmy in cells with mtDNA SNPs is also discussed.

Here we present a protocol for measuring absolute mitochondrial (mt)DNA copy number and mtDNA deletion heteroplasmy levels in single cells.