CN1196084A - 检测非嗜链烷烃微生物 - Google Patents

检测非嗜链烷烃微生物 Download PDF

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CN1196084A
CN1196084A CN96196939A CN96196939A CN1196084A CN 1196084 A CN1196084 A CN 1196084A CN 96196939 A CN96196939 A CN 96196939A CN 96196939 A CN96196939 A CN 96196939A CN 1196084 A CN1196084 A CN 1196084A
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R·A·奥拉
M·S·菲尔德
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Abstract

一种检测采自病人的样本中非嗜链烷烃微生物存在与否的方法,包括提供一个包含水溶液的容器,并用样本接种该水溶液。将碳源覆盖的载玻片放入容器中。在暴露于样本后分析载玻片,可检测样本中非嗜链烷烃微生物存在与否。还公开了相关的装置。

Description

检测非嗜链烷烃微生物
本发明涉及一种检测样本中非嗜链烷烃微生物存在与否的方法和相关装置。
临床样品中的非嗜链烷烃微生物的鉴定是病人药物治疗的一个重要部分。通常对涉及的微生物的特性进行有目的的推测，因此用一种简单有效的方法和装置来改进鉴定这些微生物的方法将是有利的。
本文所采用的术语“非嗜链烷烃微生物”指依赖于非链烷烃碳源的任何微生物。这些非嗜链烷烃微生物的实例包括，但不局限于下面：结核分枝杆菌，副结核分枝杆菌；麻风分枝杆菌；葡萄球菌属；链球菌属；大肠杆菌；李斯特氏菌属；布鲁氏菌属：Humemophilus；密螺旋体属；肺炎球菌属；梭菌属；隐球菌属；球孢子菌属；和组织胞浆菌属。而且，本文所采用的术语“病人”指动物界的成员，包括人类，它们的身体样本用本发明的方法和装置处理。
美国专利号5,153,119和5,316,918公开了用于鉴定和检测鸟细胞内结核分枝杆菌(“MAI”)(一种嗜链烷烃微生物)的抗生素敏感性的方法和设备。这些专利的发明人为Robert-A.Ollar，他是本文公开的本发明的一个共同发明人。该方法涉及提供一种包含一种水溶液的容器并将样品接种到溶液中。此后，将链烷烃覆盖的载玻片放进该容器中。接着观察载玻片中的MAI生长的存在与否。
尽管在Dr.Ollar的专利中公开了有效、可行和经济的方法，但仍然需要一种简单和有效的方法来检测非嗜链烷烃微生物的存在与否。发明概述
本发明满足或超过了上述以及其它需要。一种检测来自病人的样本中的非嗜链烷烃微生物存在与否的方法包括：提供一种包含一种水溶液的容器并用样本接种水溶液。将用碳源覆盖的载玻片放进该容器中。暴露于样本后分析载玻片，可检测样本中非嗜链烷烃微生物的存在与否。
还公开了一种相关装置。该装置包括用于容纳水溶液的容器和被碳源覆盖、适宜于被放置在容器中的载玻片。碳源引诱非嗜链烷烃微生物。碳源可被包括在与载玻片粘合的明胶基质中或可为与本身粘附于载玻片的许多凝胶颗粒离子性或亲和性结合的碳源。附图简要说明
当与所附单一的附图结合起来阅读时，可从下面的本发明的详细描述中获得对本发明的完整的理解，其中附图显示在接种了样品的水溶液中支撑被碳源覆盖的载玻片的试管的前正面观。发明详述
本发明的方法和装置提供了一种可行、有效且经济的鉴定非嗜链烷烃微生物的方法，现参考单一的附图，描述非嗜链烷烃微生物鉴定装置10的具体实例。装置10包括包含水溶液13(例如Czapek培养液)的标准试管12和密封试管12的棉塞16。根据本发明，用于检测非嗜链烷烃微生物存在与否的样本被接种到水溶液13中。接着将具有包括或含有碳源20的覆盖层的载玻片18放入试管12中，应理解到水溶液13不应含有任何碳源，因为希望在载玻片18中提供唯一的碳源20，以使非嗜链烷烃微生物有效地生长，从而在载玻片18中而不在水溶液13中被检出。可分析通过人的肉眼可以观察到或不能观察到的载玻片18中的生长以检测非嗜链烷烃微生物的存在与否。优选地，需要24小时的最少保温时间用于发生生长。为了在保温期后分析载玻片18，可以使用火焰灭菌匙突擦刮载玻片18并在类似琼脂胰蛋白酶大豆琼脂(TSA)上再培养。如果擦出物包含生长，可使用传统微生物学方法或使用DNA提取方法来分析TSA中的生长，其中DNA提取方法包括有机溶剂提取或柱层析提取。
接种到试管12中的样本可为血液样品；任何活体解剖或组织样本；胃液；尿；脑脊液；鼻咽粘液或唾液。这些样本可从医院医生办公室或急诊室中的病人，例如通过已知技术用已知的标准方法而获得。
载玻片18中的碳源20可包括含有碳源的明胶基质。碳源可为选自下面的一种或多种：葡萄糖，果糖，甘油，甘露醇，天冬酰胺和酪蛋白以及其它。另一实施方案可包括提供载玻片，用粘附剂覆盖载玻片并确保多个凝胶颗粒与粘附剂结合，接着可将碳源与凝胶颗粒离子性或亲和性地结合。
可通过下面方法制备具有包含碳源的明胶基质的载玻片18。首先将容器如实验室烧杯装100ml蒸馏水。向烧杯中加入二(2)克琼脂(明胶基质)和三(3)克碳源(如葡萄糖)。随后煮沸混合物并蒸气灭菌，将含有碳源的熔化的明胶基质倒入置于热平板上的平皿中。在此方法中，明胶基质/碳源保持熔化状态。此后，将灭菌的载玻片插入熔化的明胶基质/碳源中，并被其覆盖。将被覆盖的载玻片从平皿中移出，并让其静置1分钟或两分钟以使其上的覆盖层20固化。接着准备将具有含碳源的明胶基质的覆盖层的载玻片放入含有水溶液13和样本的试管12中。
制备载玻片的另一种方法包括：首先用粘附剂如火棉胶覆盖载玻片，接着将大量凝胶颗粒(购自Pharmacia of Parsippany，New Jersey)加入粘附剂中。凝胶颗粒直径约为1微米。现将具有凝胶颗粒覆盖层的载玻片浸在含有碳源(如葡萄糖)的缓冲试剂中来将碳源以离子或亲和方式与凝胶颗粒相连。
可使用本发明方法鉴定的非嗜链烷烃微生物包括任何依赖于非链烷烃碳源的微生物，非嗜链烷烃微生物包括，但不局限于结核分枝杆菌；副结核分枝杆菌；麻风分枝杆菌；葡萄球菌属；链球菌属，大肠杆菌；李斯特氏菌属；布鲁氏菌属；Humemophilus；密螺旋体属；肺炎球菌属；梭菌属；隐球菌属；球孢子菌属和组织胞浆菌属。
应该理解本发明已经公开了一种检测样本中非嗜链烷微生物存在与否的方法和相关装置。该方法有效、可行，并且不涉及使用昂贵和复杂的装置。还公开了相关装置。
虽然已公开了本发明具体的实施方案，但本领域技术人员应该理解根据所公开的所有教导，可对细节进行各种修改和改变。因此，所公开的具体方案意在仅仅说明，而不限制被赋予所附权利要求的全部范围和其所有和全部等同物的本发明的范围。

Claims (17)

1.一种检测采自病人的样本中非嗜链烷烃微生物存在与否的方法，所述方法包括：
提供一包含水溶液的容器；
用所述样本接种所述水溶液；
将用碳源覆盖的载玻片放入所述容器中；和
在暴露于所述样本后，分析所述载玻片以检确所述非嗜链烷烃微生物存在与否。
2.权利要求1的方法，包括采用以含有所述碳源的明胶基质覆盖的载玻片作为所述载玻片。
3.权利要求1的方法，包括通过首先将许多凝胶颗粒粘附到所述载玻片上，并接着将所述碳源结合到所述凝胶颗粒上来提供所述载玻片。
4.权利要求3的方法，其中所述碳源与所述凝胶颗粒离子性结合。
5.权利要求3的方法，其中所述碳源与所述凝胶颗粒亲和性结合。
6.权利要求3的方法，包括将所述凝胶颗粒通过粘附剂粘附到所述载玻片上。
7.权利要求6的方法，包括采用火棉胶作为所述粘附剂。
8.权利要求1的方法，包括采用葡萄糖、果糖、甘油、甘露醇、天冬酰胺和酪蛋白中的一种或多种作为所述碳源。
9.权利要求1的方法，包括采用选自血液、胃、尿、脑脊液、鼻咽粘液和唾液的一种作为所述样本。
10.一种便于检测采自病人的样本中非嗜链烷烃微生物存在与否的装置，所述装置包括：
一种用于容纳水溶液的容器；和
一个用碳源覆盖的载玻片，所述载玻片适宜于被放在所述容器中，并且所述碳源引诱所述非链烷烃微生物。
11.权利要求10的装置，其中所述载玻片被含有所述碳源的明胶基质覆盖。
12.权利要求10的装置，其中所述载玻片被其上已结合所述碳源的许多凝胶颗粒所覆盖。
13.权利要求12的装置，其中所述碳源与所述凝胶颗粒离子性结合。
14.权利要求12的装置，其中所述碳源与所述凝胶颗粒亲和性结合。
15.权利要求12的装置，其中所述凝胶颗粒通过粘附剂粘附于所述载玻片。
16.权利要求15的装置，其中所述粘附剂为火棉胶。
17.权利要求10的装置，其中所述碳源为葡萄糖、果糖、甘油、甘露醇、天冬酰胺和酪蛋白中的一种或多种。
CN96196939A 1995-09-14 1996-08-23 检测非嗜链烷烃微生物 Pending CN1196084A (zh)

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US5854013A (en) * 1995-09-14 1998-12-29 Infectech, Inc. Method of determining the presence or absence of a nonparaffinophilic microorganism in a specimen
US5832529A (en) * 1996-10-11 1998-11-03 Sun Microsystems, Inc. Methods, apparatus, and product for distributed garbage collection

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US3826717A (en) * 1973-02-26 1974-07-30 V Gilbert Quantitative antibiotic test container
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US4683201A (en) * 1984-10-09 1987-07-28 Eli Lilly And Company Antibiotic A80190-producing Actinomadura oligospora and process
US4692407A (en) * 1985-01-31 1987-09-08 Forsyth Dental Infirmary For Children Method for the determination of Streptococcus mutans
US4683195A (en) * 1986-01-30 1987-07-28 Cetus Corporation Process for amplifying, detecting, and/or-cloning nucleic acid sequences
US4683202A (en) * 1985-03-28 1987-07-28 Cetus Corporation Process for amplifying nucleic acid sequences
US5316918A (en) * 1989-10-24 1994-05-31 Infectech, Inc. Method and apparatus for testing MAI (mycobacterium avium-intracellulare) for antimicrobial agent sensitivity
US5153119A (en) * 1989-10-24 1992-10-06 Infectech Inc. Method for speciating and identifying mai (mycobacterium avium-intracellulare)

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