Pirro F, Wieler L H, Failing K, Bauerfeind R, Baljer G
Institut für Hygiene und Infektionskrankheiten der Tiere, Giessen, Germany.
Vet Microbiol. 1995 Feb;43(2-3):131-41. doi: 10.1016/0378-1135(94)00089-f.
Previous or present infection with Shiga-like toxin producing E. coli (SLTEC) was detected by an indirect neutralization assay of antibody titer. Bovine colostra and sera blocked the cytotoxic effects of Shiga-like toxin on Vero cell monolayers. SLT neutralizing antibodies were present in 84.0% (189/225) of the colostrum samples from randomly chosen cows in Bavaria, Germany. While all of the colostra with neutralizing activity reacted with SLT-I, only 14.7% neutralized both SLT-I and -II. Approximately 93.0% (37/40) of sera from heifers had SLT neutralizing activity. To quantify the neutralizing antibodies, colostra were tested in the Vero cell assay for their capability to reduce the 50% cytotoxic dose (CD50) of SLT standards, where the neutralizing units/ml (nu/ml) equal the log10 of CD50 reduction. Almost half of reactive colostra (48.7%) reduced the CD50 of the SLT-I standard by 10(4) to 10(5) (4-5 nu/ml). Higher reactivity (5-7 nu/ml) was found in 46.5% of positive colostra. The remaining colostra samples had over 7 nu/ml. To determine if the colostra were blocking receptors for SLT on Vero cells, cells were preincubated with colostra, and SLT was later added. No neutralizing activity was detected, indicating the reactivity of colostra was directed against SLT. When the colostra were subjected to ammonium sulphate precipitation and DEAE anion exchange chromatography, high levels of neutralizing activity were found in the IgG1 containing fractions. Colostrum fractions were tested for SLT-I binding antibodies in a capture ELISA, based on the binding of SLT-I to the toxin receptor analogue P1-glycoprotein. Only fractions from colostra with over 5 nu/ml were reactive in this assay, indicating the ELISA was less sensitive than the Vero cell assay. The results support the theory that SLTEC exposure of cows in Germany is more widespread than expected from epidemiological studies based on bacterial isolation. This possibly indicates a higher risk of human SLTEC infection via beef and milk products.
通过抗体滴度的间接中和试验检测先前或当前是否感染产志贺样毒素大肠杆菌（SLTEC）。牛初乳和血清可阻断志贺样毒素对Vero细胞单层的细胞毒性作用。在德国巴伐利亚随机选取的奶牛的初乳样本中，84.0%（189/225）含有SLT中和抗体。虽然所有具有中和活性的初乳都与SLT-I发生反应，但只有14.7%的初乳能同时中和SLT-I和-II。约93.0%（37/40）的小母牛血清具有SLT中和活性。为了量化中和抗体，在Vero细胞试验中检测初乳降低SLT标准品50%细胞毒性剂量（CD50）的能力，其中中和单位/毫升（nu/ml）等于CD50降低值的log10。几乎一半有反应的初乳（48.7%）将SLT-I标准品的CD50降低了10⁴至10⁵（4 - 5 nu/ml）。46.5%的阳性初乳具有更高的反应性（5 - 7 nu/ml）。其余初乳样本的反应性超过7 nu/ml。为了确定初乳是否阻断Vero细胞上SLT的受体，先将细胞与初乳预孵育，随后加入SLT。未检测到中和活性，这表明初乳的反应性是针对SLT的。当对初乳进行硫酸铵沉淀和DEAE阴离子交换层析时，在含有IgG1的组分中发现了高水平的中和活性。基于SLT-I与毒素受体类似物P1-糖蛋白的结合，在捕获ELISA中检测初乳组分中的SLT-I结合抗体。只有反应性超过5 nu/ml的初乳组分在该试验中有反应，这表明ELISA的敏感性低于Vero细胞试验。结果支持这样一种理论，即德国奶牛接触SLTEC的情况比基于细菌分离的流行病学研究所预期的更为普遍。这可能表明通过牛肉和奶制品感染人类SLTEC的风险更高。
