Cytopathologic detection of Chlamydia trachomatis in vaginopancervical (Fast) smears

The value of the Papanicolaou-stained vaginopancervical (Fast) smear in the detection of chlamydial infection has been disputed. We examined 116 satisfactory Fast smears from 203 women enrolled in the Johns Hopkins Fertility Control Clinic and compared tissue-culture results with cytopathologic detection using various published morphologic criteria. All Chlamydia culture-positive cases were reviewed, and certain cytologic features considered helpful in the detection of chlamydial infection in cervical smears obtained from this selected high-risk population were identified. The changes that had the highest correlation with tissue culture included fine vacuolation of metaplastic endocervical cells, giving their cytoplasm a rarefied "moth-eaten" appearance. Using these criteria, cytopathologic changes of chlamydial infection were observed in 24 of 28 cases of tissue-culture-positive cases and in 8 of 88 tissue-culture-negative cases. The sensitivity and specificity of the Fast-smear cytodiagnosis of Chlamydia infection utilizing these morphologic changes and compared with tissue culture were 86% and 91%, respectively. Other cytologic features, including inflammatory background and intracytoplasmic structures consistent with initial and intermediate chlamydial bodies within the metaplastic cells, were found to be useful although less specific and less sensitive. The implications of these diagnostic features, the conditions to be considered in their differential diagnosis, and the pitfalls of chlamydial cytodiagnosis and the chlamydia culture studies have been critically reviewed. Study design and the high unsatisfactory cervical smear rate are discussed.     

沙眼衣原体诊断技术研究进展

沙眼衣原体（Ct）是一种特殊的病原体,具有与革兰氏阴性细菌相似的细胞壁,含有DNA和RNA两种类型的核酸,严格寄生于宿主细胞内,沙眼衣原体是一种能够通过滤器,以二分裂方式繁殖的原核细胞型微生物.它具有两种形态：在细胞外具有高度传染性的为原体 在细胞内进行复制、无传染性的为始体.它可以引起非淋球菌尿道炎等许多泌尿生殖道相关疾病,近年来其感染率和危害性已超过淋病奈瑟菌而居性传播疾病之首,眼部衣原体侵入人体眼结膜和角膜引起沙眼和包涵体结膜炎,是世界范围致盲的首要病因.约80%的被感染女性无临床症状,感染反复迁移,造成病理改变,可导致复杂的并发症.因此,早期、简便、快速、特异地发现Ct,对临床的诊断,疾病的早期治疗和预防其流行等具有重要的意义.目前,对沙眼衣原体的诊断方法主要有培养法,免疫学法和分子生物学法.     

Value of papanicolaou smear in detection of Chlamydia trachomatis infection

This study was undertaken to assess the value of Papanicolaou smear for the diagnosis of Chlamydia trachomatis infection. The study was both retrospective (groups I and II) and prospective (group III). Group I consisted of 41 smears with cytomorphological changes proposed by Gupta, Kiviat, or Shiina. Group II was a control group, consisting of 30 cytologically normal smears. All these smears were subjected to specific immunofluorescent (IF) staining under identical conditions to confirm the diagnosis. In group III, 40 consecutive duplicate cervical smears were collected from patients attending the Sexually Transmitted Disease Clinic. One smear was routinely examined, and the specific IF staining was done on the other smear. The results in all the three groups were analysed. It was concluded that Papanicolaou smear is not useful in the detection of Chlamydia trachomatis infection.     

Characterization of Chlamydia trachomatis omp1 gene among sexually transmitted disease patients in south China

Chlamydia trachomatisinfections are the leadingcause of bacterial sexually transmitted diseases(STD) [1] .Fifteen prototypic serovarslabelled Ato Kand L1,L2,and L3were initiallyrecognisedby polyclonal antibodies ,and additional immuno-variants (Ba ,Da ,Ia ,L2a ,etc) ,whichin somepublications are referred to as distinct serovars ,have been identified by monoclonal antibodies .Most serovars can cause urogenital infections andare associated with a spectrumof clinical diseases[2] ,including ur…     

实时荧光定量PCR检测沙眼衣原体的研究

目的采用TaqMan探针建立检测沙眼衣原体的实时荧光定量PCR（real-time PCR）方法。方法根据沙眼衣原体外膜蛋白A的基因（ompA）序列设计引物和探针,以克隆的ompA部分基因片段作DNA模板,建立实时荧光定量检测方法。结果建立的荧光定量PCR检测方法的最低检出限为5 copies/反应,检测线性范围100～107线性关系良好（r2=0.997）,比巢式PCR敏感100倍;且与鹦鹉热衣原体、淋球菌、解脲脲原体、大肠杆菌等病原菌DNA以及人基因组DNA均无交叉反应,表明该方法具有良好的特异性。以巢式PCR作参比,建立的荧光定量PCR法检测沙眼衣原体的阳性符合率为100.00%,阴性符合率为95.09%,总符合率为96.81%。结论建立的检测沙眼衣原体实时荧光定量PCR具有特异性强和敏感性高的特点,可快速检测样本中微量沙眼衣原体DNA,适用于对沙眼衣原体进行大规模筛选。     

沙眼衣原体套式（Nested）PCR检测研究

本文报告沙眼衣原体（ＣＴ）的套式（Ｎｅｓｔｅｄ）ＰＣＲ检测方法，本方法ＣＴ隐匿性质粒为靶基因，外套引物采用国外学者所报告灵敏度和特异性较高的序列，内套引物自行设计，经方法学考核表明本法灵敏度和特异性极高。１４６例临检标本套式ＣＴ，ＰＣＲ阳性检出率为３６．３％而市售ＰＣＲ试剂盒（其引物序列与本文外套引物相同）阳性检出率仅为４．１％，前者明显高于后者（Ｐ〈０．０１）。     

Recovery of cytomegalovirus and Chlamydia trachomatis from vaginal tampons

Herpes simplex virus (HSV), cytomegalovirus (CMV), and Chlamydia trachomatis are important agents in venereal and neonatal disease. Vaginal tampon culture for HSV has previously been demonstrated to be a simple and effective technique for quantitative culture of cervical secretions. We have evaluated the tampon culture as a means of performing quantitative cultures for CMV and C trachomatis. Cell-free and cell-associated CMV were quantitatively recovered from vaginal tampons when extraction was performed within one hour of tampon inoculation. However, when tampons were stored, there was a rapid loss of infectivity over time at all storage temperatures except -70 degrees C. C trachomatis was quantitatively recovered from tampons stored at less than or equal to 4 degrees C for four days. When stored at -70 degrees C, C trachomatis was stable on tampons for more than one week. Because HSV, CMV, and C trachomatis are stable in a single transport medium, a tampon stored at 4 degrees C briefly or at -70 degrees C for one week could be utilized for the detection of all three agents.     

Genotyping of Chlamydia trachomatis strains from cultured isolates and nucleic acid amplification test-positive specimens

Urogenital strains of Chlamydia trachomatis are divided into several serogroups (D-K). Since these serovars are represented with differing prevalence in the population a serotyping of strains is necessary, when characterising the epidemiological situation. The aim of this study was the genotyping of C. trachomatis strains, the comparison of the results with those of serotyping, and the genotyping of positive specimens using commercial nucleic acid amplification tests (NAAT). The Chlamydia trachomatis major outer membrane protein gene (omp1) from 55 isolated strains and 36 NAAT-positive specimens was amplified by polymerase chain reaction (PCR). The restriction fragment length polymorphism (RFLP) patterns of these amplicons were compared with those of reference strains. The genotypes E and F were found to be most prevalent. The results are discussed considering other studies, genovariants and epidemiology.     

Chlamydia trachomatis causing neonatal conjunctivitis in a tertiary care center

Chlamydia trachomatis is considered a major aetiological agent of conjunctivitis in newborns. The objective of the present study was to determine the aetiology of neonatal conjunctivitis and clinico-epidemiological correlates of chlamydial ophthalmia neonatorum. Fifty-eight newborns with signs and symptoms of conjunctivitis were studied. Conjunctival specimens were subjected to Gram staining, routine bacteriological culture, culture for Neisseria gonorrhoeae and direct fluorescent antibody (DFA) staining for diagnosis of C. trachomatis infection. C. trachomatis was detected in 18 (31%) neonates. Findings suggest that since C. trachomatis is the most common cause of neonatal conjunctivitis, routine screening and treatment of genital C. trachomatis infection in pregnant women and early diagnosis and treatment of neonatal Chlamydial conjunctivitis may be considered for its prevention and control.     

E-选择素在生殖道沙眼衣原体感染中的作用研究

目的 研究E-选择素在生殖道沙眼衣原体感染中的作用.方法 将64只小鼠生殖道沙眼衣原体感染模型随机分成2组,实验组中的小鼠用人工合成E-选择素干预,对照组中的小鼠用生理盐水进行干预,统计两组在1w,2w,3w,4w不同时间点上的感染小鼠阴道脱落菌量以及沙眼衣原体感染率.结果 成功建立生殖道沙眼衣原体感染的小鼠模型:与对照组相比较,人工合成E-选择素干预后1w和2w的生殖道沙眼衣原体感染率(90.2％vs 77.4％,85.3％vs 70.1％)显著增高,而阴道脱落菌量显著增高(4.758 ±0.225 vs 3.210 ±0.315,2.698±0.177 vs 1.809±0.203),两组间差异具有统计学.在干预后3w两组的沙眼衣原体感染率分别为34％和24％,阴道脱落菌量分别为1.412±0.134和1.201±0.189,两组间差异无统计学意义;两组在干预后4w的沙眼衣原体感染率均为0且无阴道脱落菌量.结论 E-选择素能促进生殖道沙眼衣原体感染的发生率以及增加沙眼衣原体的毒力,本文为进一步阐明生殖道沙眼衣原体感染的发病机制提供实验依据.     

Clearance of Chlamydia trachomatis– induced polyserositis in SCID mice requires both CD4+ and CD8+ cells

To characterize the role of specific lymphocyte subsets in Chlamydia trachomatis infection, we established a murine model using the mouse pneumonitis agent (MoPn) of C. trachomatis and C.B-17 scid/scid (SCID) mice which lack functional B and T cells. After intraperitoneal inoculation with the bacteria, SCID mice developed polyserositis with pleuritis, pericarditis, and perihepatitis. Within 8 weeks post infection, SCID mice succumbed to the disease, whereas immunocompetent congenic C.B-17+/+ mice resolved the infection. Adoptive transfer of immune spleen cells into MoPn-infected SCID mice resulted in a complete elimination of the agent and prevention of polyserositis as measured by quantitative chlamydial culture, direct immunofluorescence and histopathological analysis. Selective reconstitution of MoPn-infected SCID mice with immune B lymphocytes, CD4⁺ T cells or CD8⁺ T cells alone did not influence the chlamydial load in the lung and liver of infected SCID animals, resulting in a polyserositis as observed in untreated MoPn-infected SCID mice. However, co-transfer of both CD4⁺ T cells and CD8⁺ T cells led to a significant reduction of chlamydiae in quantitative organ culture coupled with unremarkable histopathology. These data confirm that T cell-mediated immune responses are essential for immune protection in chlamydial infection, although total eradication of the agent could not be achieved. Further experiments are needed to stress the importance of a concerted action of B and T lymphocytes, as indicated by the complete protective efficacy of transferred splenocytes. Received: 9 April 1998     

Unclassified serovars of Chlamydia trachomatis isolated from Japanese infants

Objective: To analyze antigenic and genetic variations of Chlamydia trachomatis among the serovars obtained from Japanese infants.
Methods: The polymerase chain reaction (PCR) was used to amplify a large part of the major outer-membrane protein gene, and restriction fragment length polymorphism (RFLP) was used to identify the serovars of C. trachomatis from nasopharyngeal and conjunctival swabs from Japanese infants and neonates.
Results: The typing of 10 nasopharyngeal isolates gave the following results: seven E, one H, and two unclassified serovars. The typing of seven conjunctival isolates gave the following results: five D, one F, and one unclassified serovar. Reactive patterns of these unclassified strains, determined by PCR-RFLP, to monoclonal antibodies were different from those of 15 reference serovars.
Conclusions: Characterization of unclassified variants will allow more detailed epidemiologic studies of perinatal C. trachomatis infections in Japan.     

Relationships between Chlamydia trachomatis antibody titers and tubal pathology assessed using transvaginal hydrolaparoscopy in infertile women

PROBLEM: Since transvaginal hydrolaparoscopy (THL) was introduced as the first-line procedure in the early stages of the exploration of the adnexal structures in infertile women, it has been shown that THL is a less traumatic and a more suitable outpatient procedure than diagnostic laparoscopy. This study was performed to investigate the relationships between Chlamydia trachomatis antibody titers and tubal pathology assessed using THL in infertile women. METHODS: The C. trachomatis antibody titers (IgG and IgA) were evaluated by ELISA. The posterior of the uterus and the tubo-ovarian structures were carefully observed, and tubal passage using indigocarmine was confirmed using THL. THL was carried out in 32 infertile women having C. trachomatis antibody in their sera between May 1999 and October 2001. Unilateral salpingectomy had been performed on two of the 32 patients. RESULTS: Tubal occlusion was confirmed in 20 (32.3%) of the 62 tubes, while peritubal adhesion was diagnosed in 37 (59.7%) of the 62 tubes. Using receiver operating characteristics curves, the cut-off value of C. trachomatis IgG antibody titer to predict tubal occlusion was determined to be 3.55. Tubal occlusion was observed in 16 (51.6%) of the 31 tubes in patients with the C. trachomatis IgG antibody titer of more than 3.55, which was significantly higher in four (12.9%) of the 31 tubes having the antibody titer less than 3.55 (P = 0.004). However, there was no correlation between C. trachomatis IgG antibody titer and peritubal adhesion. As for C. trachomatis IgA antibody titer, there was no correlation between antibody titer and tubal occlusion or peritubal adhesion. CONCLUSIONS: These results suggest that C. trachomatis infection is significantly associated with tubal pathology. Although the cut-off value of C. trachomatis IgG antibody titer to predict the existence of tubal occlusion was shown to be 3.55, we would suggest that THL or standard laparoscopy is performed to consider appropriate treatments in patients with past C. trachomatis infection because of the high prevalence of peritubal adhesion.     

100例不育症患者解脲支原体和沙眼衣原体定量分析

目的为了分析不育症和非淋茵性尿道炎患者感染解脲支原体和沙眼衣原体定量检测结果的差异.方法荧光定量多聚酶链式反应(FQ-PCR)技术连续对100例(有97例作了UU检测,84例作了CT检测)本院生殖中心不育症患者及同时时662例(有470例作了UU检测,540例作了CT检测)普通性病和妇科门诊非淋患者CTDNA和UUDNA进行了定量检测.结果不育组UU阳性率=57/97(58.76%),UU拷贝对数值6.016±1.044;CT阳性率=7/84(8.33%),CT拷贝对数值4.968±2.190.普通非淋组UU阳性率=209/470(46.60%),UU拷贝对数值6.131±1.385;CT阳性率=62/540(11.48%),CT拷贝对数值5.894±1.943.结论UU是引起非淋和不育症的主要病原体.UU相对于CT对人类生育的危害更为严重.不育组和普通非淋组UU的定量结果没有统计学差异,而普通非淋组的CT的拷贝数则高于不育组,P＜0.05.     

Chlamydia trachomatis antigen detection in pregnancy and its verification by antibody blocking assay

Purpose: To detect the prevalence of genital infection caused by Chlamydia trachomatis in pregnant women and also to confirm the positive results using blocking antibody assay. Methods: Endocervical specimens were collected from 200 symptomatic and asymptomatic pregnant women attending the ANC OPD at M P Shah Medical College, Jamnagar. The samples were tested for presence of Chlamydia trachomatis antigen using the monoclonal antibody. Blocking antibody assay was used to further verify the positive results. Results: Out of 200 pregnant women, 38 (19%) were found positive for Chlamydia trachomatis antigen. Out of the 68 symptomatic patients, C. trachomatis antigen was detected in 26.4%. After verification of the positive samples 13.6% of the asymptomatic pregnant women were found to be harbouring the infection in their genital tract. Two (5.2%) out of the 38 positive samples, on verification with the blocking antibody assay, were found to be false positive by IDEIA,^TM thus the specificity of the IDEIA^TM being 94.8%. In patients with previous history of abortions, 27.7% were tested positive for C. trachomatis infection. Conclusions: Significant number of pregnant women shad C. trachomatis antigen in their endocervical canal, which can be easily diagnosed by this simple enzyme immuno assay having a specificity of 94.8%. Verification of positive results by antibody blocking assay can further improve the specificity of this non-culture test. Asymptomatic patients should also be screened for the infection. History of previous abortions places the patient at a higher risk for C. trachomatis infection thus such patients should be definitely tested for chlamydia infection.     

Usefulness of transvaginal hydrolaparoscopy in investigating infertile women with Chlamydia trachomatis infection

BACKGROUND: A new technique called transvaginal hydrolaparoscopy (THL) was recently developed for the exploration of the tubo-ovarian structures in infertile patients without obvious pelvic pathology. This study was performed to investigate the usefulness of THL to evaluate Chlamydia trachomatis tubal infertility. METHODS: Forty-one women with primary and secondary infertility participated in this study. Fourteen had past C. trachomatis infection. In 38 (92.7%) of the 41, access to the pouch of Douglas was obtained. In total, 71 (93.4%) out of 76 adnexa were clearly visualized. Thirty-seven patients were analysed and compared their tubal passages and peritubal adhesions using both hysterosalpingography (HSG) and THL. Twenty-four tubes from 14 patients with past C. trachomatis infection and 44 tubes from 23 patients without a history of C. trachomatis infection were compared. RESULTS: For the diagnosis of the tubal passage, there were no significant differences in the discrepancy rates between HSG and THL, in patients with and without past C. trachomatis infection. In 14 (58.3%) of the 24 tubes from patients with past C. trachomatis infection and in eight (18.2%) of the 44 tubes from patients without infection, peritubal adhesion was diagnosed only by THL. There was a significant difference in the discrepancy rates of the diagnosis of peritubal adhesion between HSG and THL in the two groups (P = 0.0007 ). CONCLUSIONS: These results suggest that C. trachomatis infection is highly associated with peritubal adhesion which is difficult to diagnose by HSG. Therefore, in C. trachomatis antibody-positive patients, exclusion of tubal pathology by THL or standard laparoscopy should be carried out to consider appropriate treatments. Although THL is not a substitute for laparoscopy, it can be proposed as a first line procedure in the early stages of the infertility investigation.     

荧光定量PCR检测沙眼衣原体及临床应用

目的：用荧光定量聚合酶链反应（FQ－PCR）检测沙眼衣原体的含量,探讨FQ－PCR在衣原体性尿道炎诊断中的价值。方法：应用荧光探针标记引物的荧光定量聚合酶链反应对172例疑为沙眼衣原体感染患者标本进行检测,并与常规PCR（电泳－EB染色）进行比较。结果：FQ－PCR阳性率为19．8％,常规PCR法为20．9％,两法符合率为63．9％。女性宫颈分泌物标本FQ－PCR阳性为29．2％,常规PCR法为16．7％;男性分泌物FQ－PCR阳性率为16．9％,常规PCR法为13．8％。FQ－PCR特异性较常规PCR法高。结论;FQ－PCR在扩增中实时在线检测,并选择理想的标准曲线做出较精确的临值分析,具有较高的特异性和敏感性,对病原体的诊断有一定的临床意义。