Wu F M, Beuchat L R, Wells J G, Slutsker L, Doyle M P, Swaminathan B
Center for Food Safety, University of Georgia, Griffin 30223-1797, USA.
Int J Food Microbiol. 2001 Dec 4;71(1):93-9. doi: 10.1016/s0168-1605(01)00559-1.
Isolating Escherichia coli O157:H7 from batches of alfalfa seeds used to produce sprouts implicated in human illness has been difficult, perhaps due to nonhomogenous and very low-level contamination and inaccessibility of the pathogen entrapped in protected areas of the seed coat. We evaluated the effectiveness of various treatments in releasing E. coli O157:H7 from seeds. The influence of homogenization (blending or stomaching for 1 or 2 min), rinsing method (shaking for 5 min), soaking time (0. 1, 3, 6, or 15 h), soaking temperature (4 or 21 degrees C), and the addition of surfactants (0.1%, 0.5%, or 1.0% Tween 80 or Span 20) to rinse water was determined. Blending or stomaching for 1 or 2 min, and soaking for 1 h or longer at 4 or 21 degrees C, respectively, resulted in maximum release of E. coli O157:H7 from seeds. Soaking seeds at 37 degrees C for 15 h increased cell populations of E. coli O157:H7 by approximately 3.6 log10 CFU/g, likely due to bacterial growth. The maximum number of cells released from seeds by rinse water containing 1.0% Span 20 was at 21 degrees C, whereas at 37 degrees C, 0.1% or 0.5% Tween 80 was more effective. Detection of E. coli O157:H7 on seeds stored at 37 degrees C for up to 13 weeks and on sprouts derived from these seeds was compared. E. coli O157:H7 inoculated on seeds at 2.0 log10 CFU/g was detected after storage of seeds for up to 8 weeks at 37 degrees C and in sprouts produced from the seeds. The pathogen was not detected on seeds stored for 13 weeks at 37 degrees C and was not isolated from sprouts produced from these seeds. Identifying seed treatment methods that enhance removal of E. coli O157:H7 from alfalfa seeds can aid the isolation and enumeration of the pathogen on seeds. With a combination of optimal conditions for detecting the pathogen, i.e. soaking seeds for 1 h and pummeling seeds for 1 min, an enrichment step in modified tryptic soy broth (TSB), and the use of immunomagnetic beads for separation of E. coli O157:H7 cells, E. coli O157:H7 was detected in alfalfa seeds incubated at 37 degrees C for up to 8 weeks as effectively as in sprouts produced from the seeds.
从用于生产与人类疾病相关豆芽的苜蓿种子批次中分离出大肠杆菌O157:H7一直很困难，这可能是由于污染不均匀且水平极低，以及病原体被困在种皮的保护区域内难以接触到。我们评估了各种处理方法从种子中释放大肠杆菌O157:H7的效果。确定了匀浆（搅拌或均质1或2分钟）、冲洗方法（振荡5分钟）、浸泡时间（0.1、1、3、6或15小时）、浸泡温度（4或21摄氏度）以及在冲洗水中添加表面活性剂（0.1%、0.5%或1.0%吐温80或司盘20）的影响。分别搅拌或均质1或2分钟，以及在4或21摄氏度下浸泡1小时或更长时间，可使大肠杆菌O157:H7从种子中最大程度地释放出来。在37摄氏度下将种子浸泡15小时会使大肠杆菌O157:H7的细胞数量增加约3.6个对数10 CFU/g，这可能是由于细菌生长。含有1.0%司盘20的冲洗水在21摄氏度下从种子中释放的细胞数量最多，而在37摄氏度下，0.1%或0.5%吐温80更有效。比较了在37摄氏度下储存长达13周的种子以及由这些种子长出的豆芽上大肠杆菌O157:H7的检测情况。接种在种子上的大肠杆菌O157:H7浓度为2.0个对数10 CFU/g，在37摄氏度下储存长达8周的种子以及由这些种子长出的豆芽中都能检测到。在37摄氏度下储存13周的种子上未检测到该病原体，并且从这些种子长出的豆芽中也未分离出该病原体。确定能够增强从苜蓿种子中去除大肠杆菌O157:H7的种子处理方法，有助于在种子上分离和计数该病原体。通过结合检测该病原体的最佳条件，即浸泡种子1小时并研磨种子1分钟，在改良胰蛋白胨大豆肉汤（TSB）中进行富集步骤，并使用免疫磁珠分离大肠杆菌O157:H7细胞，在37摄氏度下培养长达8周的苜蓿种子中检测大肠杆菌O157:H7的效果与从这些种子长出的豆芽中一样有效。
