Verkooyen R P, Luijendijk A, Huisman W M, Goessens W H, Kluytmans J A, van Rijsoort-Vos J H, Verbrugh H A
Department of Clinical Microbiology, Academic Hospital Dijkzigt, Rotterdam, The Netherlands.
J Clin Microbiol. 1996 Dec;34(12):3072-4. doi: 10.1128/jcm.34.12.3072-3074.1996.
To determine that susceptibility of AMPLICOR Chlamydia trachomatis PCR to inhibitory factors possibly present in cervical specimens, we obtained cervical specimens from 200 gynecology patients attending our outpatient clinic. The prevalence of C. trachomatis infection was 4.1%, as determined by cell culture. All AMPLICOR specimens were tested in one procedure as described by the manufacturer, and after the specimen was spiked with C. trachomatis, several other pretreatment protocols were used. Complete inhibition of the PCR was observed in 38 (19%) cervical specimens. Heat treatment at 95 degrees C, freeze-thawing, or 10-fold dilution of the samples reduced the initial inhibition to 9, 16, or 9%, respectively. A combination of heat treatment and 10-fold dilution reduced the inhibition to 4% of the samples. A second specimen type (swabs inoculated in 0.2 M sucrose phosphate buffer [2SP]) was also evaluated. A 10-fold dilution of the spiked 2SP specimen resulted in an inhibition rate of 6%, which was comparable to that obtained by centrifugation of the 2SP specimen prior to processing. Furthermore, it was shown that the inhibition was not correlated with blood contamination. Processing the specimens on the day of collection or the day after resulted in a higher inhibition rate than did delayed processing (27.6 versus 15.5%, respectively). An inverse correlation was found between the concentration of C. trachomatis added to the sample and the rate of inhibition observed. The inhibition was partly correlated with the pH of the cervical mucosa. Decreased inhibition was found at pH values of > or = 7.5. The effects of blood, pH, and delay in processing were all evaluated by using the AMPLICOR specimen. We conclude that the susceptibility of AMPLICOR C. trachomatis PCR to inhibiting factors in cervical specimens can be significantly reduced if the pretreatment procedure includes heat treatment or the use of 2SP transport medium. Also, a 10-fold dilution of the clinical specimen followed by heat treatment will largely prevent the inhibition of this PCR.
为确定AMPLICOR沙眼衣原体聚合酶链反应（PCR）对宫颈标本中可能存在的抑制因子的敏感性，我们从200名到我院门诊就诊的妇科患者中获取了宫颈标本。通过细胞培养确定沙眼衣原体感染率为4.1%。所有AMPLICOR标本均按照制造商所述的方法在一个步骤中进行检测，并且在标本中加入沙眼衣原体后，采用了其他几种预处理方案。在38份（19%）宫颈标本中观察到PCR完全抑制。95℃热处理、冻融或样本10倍稀释分别将初始抑制率降至9%、16%或9%。热处理和10倍稀释相结合将抑制率降至样本的4%。还对第二种标本类型（接种于0.2M蔗糖磷酸盐缓冲液[2SP]中的拭子）进行了评估。加样的2SP标本10倍稀释后的抑制率为6%，这与处理前对2SP标本进行离心获得的抑制率相当。此外，结果表明抑制作用与血液污染无关。在采集当天或采集后一天处理标本的抑制率高于延迟处理（分别为27.6%和15.5%）。发现添加到样本中的沙眼衣原体浓度与观察到的抑制率呈负相关。抑制作用部分与宫颈黏膜的pH值相关。在pH值≥7.5时抑制作用降低。血液、pH值和处理延迟的影响均通过使用AMPLICOR标本进行评估。我们得出结论，如果预处理程序包括热处理或使用2SP转运培养基，AMPLICOR沙眼衣原体PCR对宫颈标本中抑制因子的敏感性可显著降低。此外，临床标本10倍稀释后再进行热处理将在很大程度上防止该PCR受到抑制。
