中国科技网10月9日报道（张微 编译）以色列本-古里安大学和美国麻省理工学院的研究人员开发出一种高敏感度，节约成本的技术，只需在24小时之内，就能快速完成空气、土壤、水和农产品中的细菌性病原体检测。

根据这项研究的领导者，正在本-古里安大学扎克伯格水研究所攻读博士学位的以斯拉奥洛夫斯基博士介绍，“现场样本中快速可靠的病原体检测对于公共卫生、安全和环境监测是至关重要的。目前在食品、水或临床应用上使用的方法需要依赖实验室以及花费大量的时间培养技术，比如在乳品业、废水和径流处理这样的活动中，需要在环境样本中实时监测病原体。

这项发表在《水、空气&土壤污染》期刊网络版上的研究，定义了一个准确的、价格低廉的、高通量的以及快速的替代方法来检测不同环境样本中的病原。“这是首次在种类广泛的环境样本类型中，全面评估病原体浓度的研究，通过标准(或传统)方法，利用完整的并行测试，实现多病原检测。

“我们能够在24小时内准确地确定土壤样品中的沙门氏菌，而传统的方法需要四到五天，而且需要分类，”以斯拉奥洛夫斯基博士说。“我们还成功地确定了一种致命的感染，铜绿假单胞菌，存在于气溶胶中，是国内污水处理系统中的产生的。结果表明，我们开发的技术提供了一种广泛的方法，在具有挑战性的环境样品中的快速，高效，可靠地检测出相对低浓度的致病微生物。

为了评估这项技术，各种类型的环境样本，包括气溶胶，各种土壤类型，废水和蔬菜表面(番茄)，被同时侵入了沙门氏菌或铜绿假单胞菌。研究人员选择了这些病原是因为他们是主要的致病病原，在环境中有着较高的生存潜能，而且在浓度低的时候很难被准确地检测出来。

“当这个方法应用于没有侵入病原的现场样本时，我们的方法显著优于标准方法，能够在收到样本一天之内检测出病原，” 以斯拉奥洛夫斯基博士说。“由于这个集中而且经济的检测程序告诉我们在一天之内重点检测哪里，因此我们不需要花费几天时间来检测数以百计的样本和子样本。”

这个快速检测方法同时使用了两种技术，一个是用于微生物检测的进化的“MPN-type enrichment”技术，再加上广泛用于分子生物学用于实时监测DNA扩增的“qPCR”技术。

“我们大大缩短了之前的方案，也不再使用任何名牌昂贵的试剂对DNA进行提纯和净化，也不用增加工作流程，从原始样品很容易地进行到qPCR实验，” 以斯拉奥洛夫斯基博士说。

土壤、水和蔬菜样本的检测是高度敏感的(低至每一个细胞的测试)，研究人员认为需要额外的步骤来进一步提高检测水平，使得这项技术能够反映出气溶胶中的低病原浓度(特别是低浓度感染剂量)。

研究人员推荐今后用这种方法来检测其他病原，如军团菌，金黄色葡萄球菌和空肠弯曲杆菌，其中第二个病菌是食源性疾病的最常见原因。

Researchers develop rapid method for water, soil pathogen screening

Researchers at Ben-Gurion University of the Negev (BGU) and the Massachusetts Institute of Technology (MIT) have developed a highly sensitive, cost-effective technology for rapid bacterial pathogen screening of air, soil, water, and agricultural produce in as little as 24 hours.

According to Ezra Orlofsky Ph.D, who led the research while working on his doctorate at the BGU Zuckerberg Institute for Water Research, "Rapid and reliable pathogen detection in field samples is critical for public health, security and environmental monitoring. Current methods used in food, water or clinical applications rely on labor and time-intensive culturing techniques while activities such as dairy farming, wastewater and runoff treatment necessitates real-time monitoring of pathogens in environment samples."

The study, published online in the Water, Air & Soil Pollution journal (Springer) defines an accurate, inexpensive, high-throughput, and rapid alternative for screening of pathogens from various environmental samples. "This is the first study to comprehensively assess pathogen concentrations in such a broad variety of environmental sample types while achieving multiple pathogen detection with complete parallel testing by standard (or traditional) methods," Orlofsky explains.

"We accurately identified Salmonella (S. enterica) in environmental soil samples within 24 hours, while traditional methods take four to five days and require sorting," Orlofsky says. "We also successfully identified a sometimes-fatal infection, Pseudomonas aeruginosa, in aerosols generated by a domestic wastewater treatment system. The results suggest that the developed method presents a broad approach for the rapid, efficient and reliable detection of relatively low densities of pathogenic organisms in challenging environmental samples."

To evaluate the technology, a variety of environmental samples, including aerosols, various soil types, wastewater and vegetable surface (tomato), was concurrently spiked with Salmonella enterica and/or Pseudomonas aeruginosa. The researchers chose these pathogens because they are leading causes of illness, have high survival potential in the environment and are considered difficult to detect accurately at low concentration.

"When applied to non-spiked field samples, our method outperformed the standard methods substantially, while detecting pathogens within a day of receiving the samples," says Orlofsky. "Since this focused and economical screening procedure tells us exactly where to look within a day, we don't need to monitor hundreds of samples and sub-samples over several days."

The two techniques used concomitantly are an evolved "MPN-type enrichment" ("Most Probable Number") used in microbiology testing, coupled with "qPCR," (quantitative polymerase chain reaction) widely used in molecular biology to monitor the amplification of DNA in real time.

"We considerably shortened previous protocols, do not use any name-brand expensive re-agents for DNA extraction and purification, and increased the procedure and workflow to segue easily from raw sample to qPCR assays," says Orlofsky.

While detection in soil, water and vegetable samples was highly sensitive (as low as one cell per test), the researchers believe additional steps are required to further improve the detection levels such that they reflect low pathogen concentrations (especially ones with low infective doses) in aerosols.

The researchers recommend applying this method in the future to other pathogens such as Legionella pneumophilia, (Legionnaire's Disease), Staphylococcus aureus (Staph infection) and Campylobacter jejuni, the second most common cause of foodborne illness.

来源：中国科技网—科技日报