Genotyping of Chlamydia trachomatis from endocervical specimens in Brazil

BACKGROUND: There is no data concerning genotyping of Chlamydia trachomatis from Brazilian samples. GOAL: To characterize the genotype of C. trachomatis detected in women assisted at a STD public clinic and establish the prevalence of this infection in that population. STUDY DESIGN: Endocervical samples of a group of 100 women were tested for chlamydial infection with PCR directed to C. trachomatis cryptic plasmid. Genotyping of positive samples were done after omp1 amplification and sequencing. RESULTS: The overall prevalence of C. trachomatis infection was 19%, with the highest prevalence in women between 15 and 25 years old (68.4%). Four genotypes were found associated with endocervical infections: D, E, F, and K. Sequence analysis revealed a coinfection of genotypes D and E in 1 woman. CONCLUSIONS: To our knowledge this is the first study to characterize Brazilian C. trachomatis endocervical samples and Brazilian C. trachomatis genotype coinfection. Our results also emphasize the importance of routine diagnosis of C. trachomatis for the control of this STD.     

Confirming the Chlamydia trachomatis status of referred rectal specimens

OBJECTIVES: To assess the reliability of different laboratory methods for the detection of Chlamydia trachomatis in rectal specimens METHODS: 1782 rectal specimens confirmed as C trachomatis positive using a standard laboratory method, were forwarded to the Sexually Transmitted Bacteria Reference Laboratory (STBRL). All specimens were retested using a C trachomatis specific independent in-house real time polymerase chain reaction (PCR). If this test was negative, a second test (Artus Real-Art PCR Kit) was employed as a confirmation. A correlation between real time PCR results obtained at the reference centre (STBRL), and the method of C trachomatis detection used in the primary laboratory was undertaken. RESULTS: The percentage of specimens that could be confirmed as positive, compared with primary method of detection was as follows: C trachomatis culture 87.5%, strand displacement assay (SDA: Becton Dickinson) 93.4%, Cobas Amplicor (Roche) 89.2%, Aptima Combo Two assay (Genprobe) 83.3%, and enzyme immunoassays (EIA) 35.4%. CONCLUSIONS: High rates of confirmation can be achieved using an independent real time PCR assay to examine rectal specimens which had initially tested C trachomatis positive using nucleic acid amplification tests and chlamydia tissue culture. This is not possible for specimens that had been screened using EIA tests, which reflects the low specificity of this test when used for rectal specimens. Laboratories currently using EIA based assays to test rectal specimens should review this approach.     

Genotyping of Chlamydia trachomatis would improve contact tracing

BACKGROUND: The reported number of Chlamydia trachomatis genital infections has increased 15% annually since 1997 in Sweden. Inaccurate partner notification might be one reason. GOAL: The goals were to determine if genotyping of C trachomatis would improve partner notification and to study the duration of infection. STUDY DESIGN: Sexual networks were constructed. C trachomatis isolates from 231 individuals attending the Orebro STD clinic during 1 year were typed by sequencing of the omp1 gene. RESULTS: All individuals were traced and diagnoses were established in 30 of 161 networks. More than one genotype was seen in seven networks. The mean duration of C trachomatis infection in each network was calculated to be 23 weeks. CONCLUSION: Genotyping could be a useful tool in partner notification when there are discrepant or uncommon genotypes. Limited clinic catchment areas create information difficulties that obstruct accurate contact tracing.     

沙眼衣原体的基因分型及其临床意义

目的　探讨对沙眼衣原体更为简便的分型方法及基因分型的临床意义。方法　利用聚合酶链反应 (PCR)扩增沙眼衣原体外膜蛋白基因 (omp1 )片段 ,以核酸内切酶作酶切 ,1 0 %聚丙烯酰胺凝胶电泳 ,银染色后观察带型分布。结果　扩增 1 5种血清型omp1基因 ,产物为 871bp的DNA片段。用AluⅠ酶切此片段 ,观察到 1 5种血清型呈现具有特征性的带型分布 ,但C组带型较难区别。再用HpaⅡ、HinfⅠ及EcoRⅠ三重酶切 ,则可将C组各型完全区分开来。对 74株沙眼衣原体临床株进行分型 ,发现E ,F ,G ,D在本观察人群中占优势 ,偶尔 ,也见到B ,H ,J等型别。还观察到 1例F/D混合型。沙眼衣原体的型别与临床表现之间未见有明显的关联 ,但发现D型感染者常为抗沙眼衣原体抗体高滴度者。结论　omp1基因分型技术可以作为流行病学调查中的有用的研究工具     

Genotyping of Portuguese Chlamydia trachomatis urogenital isolates.

OBJECTIVE: To determine the prevalence of the different Chlamydia trachomatis genotypes in Portuguese patients. METHODS: Urogenital isolates (n = 240) derived from attenders of various clinics in the Lisbon area were differentiated into genovars by genotyping with restriction fragment length polymorphism (RFLP) analysis of the PCR amplified omp1 gene. RESULTS: Genotype E was the most common for both men (47.9%) and women (43.8%). Genotypes D and F were the second most prevalent for men (11.3%) and genotype H was the second most prevalent for women (19.5%). Genotypes F, G, D, in women and H, G, I, in men, were found in a lower percentage of cases. Genotypes B, Ba, J, K, L1 and L2 were very rarely detected. CONCLUSIONS: With one exception, the overall distribution of Chlamydia trachomatis genotypes in our study is similar to what has been observed in other western countries. The only exception is the unusual prevalence of genotype H among women. The clinical manifestations associated with this and other genotypes were similar.     

多重聚合酶链反应用于D－K型沙眼衣原体基因分型的研究

目的摸索一套泌尿生殖道沙眼衣原体基因分型的多重聚合酶链反应法。方法选择某一型与其它各型均不同的可变区部位作为下游引物,以恒定区 1区的高度保守区作为各型的公用上游引物,优化反应条件,进行标准株和临床野生株的多重聚合酶链反应扩增,并与聚合酶链反应－限制性内切酶片段多态性方法比较。结果用摸索出的反应系统和条件,对 8个标准株进行扩增,扩增出与理论值一致的片段,并可直接在普通水平琼脂糖凝胶进行电泳。特异性检测证明：①衣原体各型之间没有非特异性交叉扩增,②对泌尿生殖道病原和寄生微生物进行扩增,未扩出任何 DNA带。对临床野生株分型结果证明：复合聚合酶链反应阳性率为 100％,而聚合酶链反应－限制性内切酶片段多态性为 94.44％。结论本套衣原体基因分型的多重聚合酶链反应法特异、敏感、快速、实用,最终能用于临床分型。     

Sequential cervical specimens and the isolation of Chlamydia trachomatis: factors affecting detection

For 260 consecutive patient visits by women to a clinic for sexually transmitted diseases, four cervical specimens were cultured in duplicate for detection of Chlamydia trachomatis. Sixty-one positive results were detected by at least one of the four specimens; the first two specimens detected 67-69% and the last two 80-82% of the 61 positives. The difference in these isolation rates is statistically significant (P = 0.003). Duplicate cultures of the same specimen did not significantly increase detection rates. A combination of two specimens could increase the number detected by 44.7% beyond the results of a single-specimen culture. Contamination rates were higher for the first two specimens. Routine cleaning of the cervical canal with a swab before the taking of specimens should reduce contamination and increase the probability of obtaining infected cells when they are present.     

Evaluation of patient-administered tampon specimens for Chlamydia trachomatis and Neisseria gonorrhoeae

BACKGROUND: The patient-administered tampon specimen has proven to be an easy and sensitive method for the diagnosis of genital Chlamydia trachomatis and Neisseria gonorrhoeae infections in women by polymerase chain reaction (PCR). This method avoids the need for endocervical sampling and stringent criteria for transport. GOAL: To evaluate two commercial amplification systems for the detection of C trachomatis and N gonorrhoeae from tampon specimens. STUDY DESIGN: A group of 400 positive and negative tampon specimens tested by an in-house PCR method were selected from a pool of more than 2,000 previously collected tampons. Overall, 93 C trachomatis-positive and 77 N gonorrhoeae-positive specimens were evaluated. Each specimen was tested by Roche Cobas Amplicor and Abbott LCx (LCR), and results were compared to the in-house PCR method. RESULTS: Detection of C trachomatis by both assays was not significantly different from the in-house PCR assay. Fewer tampons were positive for N gonorrhoeae by LCR than either the in-house assay (P = 0.0001) or by Roche Amplicor (P = 0.01). However, tampon specimens tested by Roche Amplicor required DNA extraction to achieve comparative sensitivity. CONCLUSION: Both commercial assays can be applied to tampon-collected specimens for automated detection of sexually transmitted diseases. The detection of C trachomatis was similar to the in-house PCR test for both assays (P = 0.73, 0.68). Detection of N gonorrhoeae resulted in fewer positive tampon specimens when tested by ligase chain reaction than both Roche Amplicor and in-house PCR.     

连接酶链反应检测尿标本中沙眼衣原体

采用连接酶链反应（LCR）检测尿标本中沙眼衣原体（CT），并对阳性标本以聚合酶链反应（PCR）、Clearview法对照，结果发现LCR检出的44例均为真阳性，表明LCR是检测CT的一种非侵入性、敏感性及特异性很高的方法。     

Nationwide increase of Chlamydia trachomatis infection in Finland: highest rise among adolescent women and men

BACKGROUND: Since 1995, the incidence of Chlamydia trachomatis infection has been increasing in Finland, although there have been no major changes in public sexually transmitted disease (STD) services or screening practice. OBJECTIVES: The objective was to study whether the change in C. trachomatis incidence is significant and to identify specific risk groups. METHODS: The incidence rates for all C. trachomatis cases notified by laboratories to the National Infectious Disease Register (NIDR) in Finland in 1995-2000 were calculated by gender, age, and domicile. Data from a sentinel STD surveillance network was used to analyze changes of risk-taking behavior in age groups with the highest C. trachomatis rates. RESULTS: During the 6-year study period, laboratory surveillance data documented an increase in the incidence rate from 23.4 per 10,000 to 29.2 per 10,000. The increase was most evident among people living in nonurban densely populated areas. Highest increase, 1.37-fold (95% confidence interval [CI], 1.29-1.46) in women and 1.69-fold (CI, 1.47-1.92) in men, occurred in the youngest age group (10-19 years old). In 2000, more women, but not men, in the age group of 10-29 years reported 5 or more annual sex partners (18.8%; CI, 16.3-21.6) than in 1995 (8.3%; CI, 5.7-11.5). CONCLUSION: National surveillance of C. trachomatis infection based on laboratory notification documented increasing incidence rates, especially among adolescents and young people. This risk group should be a target for screening and educational programs to control the epidemic of C. trachomatis infections.     

直肠部位淋球菌及沙眼衣原体感染的诊治

直肠位于肛缘2 cm以上,长约12～15 cm,上接乙状结肠,下接肛管,通过直肠黏膜与肛管皮肤之间的齿状线与肛管分界,表面覆盖柱状上皮,是性传播疾病在男男同性恋人群(MSM)中的好发部位,女性也可因接受肛交在此部位发生性传播感染.事实上,直肠部位的性传播疾病并不少见[1-6],而且直肠部位的性传播疾病可以增加HIV的易感性[7],因此加强直肠部位性传播疾病的预防及治疗对控制HIV的流行有重要意义.本文就直肠部位较为常见的淋球菌及沙眼衣原体感染的临床特点、检查方法及治疗做一综述.     

Pooled specimens for Chlamydia trachomatis: new approach to increase yield and cost efficiency.

Pooled specimens from the urethra and cervix accounted for 97% of 101 positive chlamydial isolations in 332 women, and this yield compared favourably with the individual yield from either the urethra (77%) or the cervix (88%). Pooling specimens caused no apparent increase in toxicity to the cell culture system. These results indicate the advantages, in terms of higher yield and no higher cost, of combining the urethral and cervical specimens in one container.     

PCR for detection of Chlamydia trachomatis in endocervical, urethral, rectal, and pharyngeal swab samples obtained from patients attending an STD clinic.

OBJECTIVE: To investigate, by use of the Amplicor PCR in a routine setting, the recovery rate of Chlamydia trachomatis in ano-rectal and pharyngeal swab samples obtained from males and females attending an STD clinic in relation to sexual practices, symptoms, and signs. DESIGN: Data regarding sexual practices, and symptoms and signs related to the rectum and pharynx, were obtained from 196 females and 208 males, including 31 homosexuals and eight bisexuals. Swab samples were obtained from the urethra, rectum, and pharynx from all the patients. An additional endocervical swab sample was obtained from the females. METHODS: All samples were analysed by the Amplicor PCR (Roche). SETTING: Rudolph Bergh's Hospital, a clinic for sexually transmitted diseases situated in the centre of Copenhagen, Denmark. RESULTS: The overall prevalence of urogenital C trachomatis infection was 9.2% (37/404). The specificity of the Amplicor PCR was 100% for both ano-rectal and pharyngeal swab samples. In females three (13%) of the 23 infections were detected only by testing an ano-rectal or throat swab sample. In homosexual males two (67%) of three infections were detected only by the anorectal swab sample. Ano-rectal intercourse without use of condom was reported by 44% of females and by 52% of homosexual males. Fellatio without condom use was reported by 91% of females, and 80% of heterosexual males practised cunnilingus. Pharyngeal infection, however, occurred only in females, and the presence of pharyngeal symptoms or signs seemed predictive for pharyngeal C trachomatis infection, for which the time of incubation or colonisation exceeded 3 months. The presence of ano-rectal signs or symptoms was not predictive for an ano-rectal C trachomatis infection. CONCLUSION: The Amplicor PCR can be used on ano-rectal and pharyngeal swab samples. Ano-rectal swab samples should be obtained in females and homosexual males at high risk of being infected. Pharyngeal samples should be taken in females at high risk of being infected, especially when pharyngeal signs or symptoms are present.     

性病门诊男性尿道炎患者沙眼衣原体基因分型

目的 了解性病门诊就诊的尿道炎男性患者中,沙眼衣原体血清型分布情况。 方法 采集2013年1 - 12月中国医学科学院皮肤病医院性病门诊有尿道炎症状的男性患者的尿液,荧光定量PCR检测沙眼衣原体,对沙眼衣原体阳性患者的尿液提取DNA,用巢式 PCR法扩增沙眼衣原体主要外膜蛋白基因ompA的VS1-VS2片段,然后对此片段测序,测序结果用DNAStar5.0软件与每种血清型的参考菌株做比对,分析其血清型。 结果 对2013年432例男性尿道炎患者进行了沙眼衣原体筛查,阳性标本143例,阳性率33.1%。143例沙眼衣原体阳性标本,127例扩增出ompA的VS1-VS2片段,16例未扩增出。127例阳性标本经测序分析获得9种血清型。血清型分布情况如下：E型29（22.83%）、F型28株（22.05%）、D型19（14.96%）、G型16株（12.60%）、J型16株（12.60%）、K型8株（6.30%）、H株5株（3.94%）、I型3株（2.36%）、B型3株（2.36%）,E、F、D、J、G型占85.02%。与标准菌种比对,发现127例菌株中有14株存在碱基突变,为同义突变。 结论 性病门诊男性尿道炎沙眼衣原体血清型主要是E型、F型、D型和G型,与20年前相比,E型菌株比例有所下降,J型菌株比例增高。     

南京地区性病患者沙眼衣原体感染的基因分型

目的了解南京地区性病患者泌尿生殖道沙眼衣原体感染分离株的基因型（血清型）分布情况。方法应用聚合酶链反应和限制性片段长度多态性分析,对来自南京地区性病门诊病人和性乱妇女的９４份沙眼衣原体阳性标本进行了基因分型研究。结果Ｅ型为南京地区性病高危人群最流行的沙眼衣原体基因型,占３６．２％,其次为Ｆ、Ｇ和Ｄ型,分别占１９．２％、１７．０％和１２．８％,而Ｋ、Ｊ、Ｉ、Ｈ型比较少见。发现了一个Ｂ基因型,一个Ｆ／Ｇ混合型和４个非典型基因型。结论我国南京泌尿生殖道沙眼衣原体感染流行株的基因型分布情况与国外基本相同,且存在某些基因型的变异。     

反向斑点杂交技术检测泌尿生殖道沙眼衣原体及基因分型

目的 建立反向斑点杂交技术,对泌尿生殖道沙眼衣原体感染进行检测和基因分型。方法 用Oligo软件设计寡核苷酸探针,采用巢式聚合酶链反应(PCR)扩增omp1基因的VS1-VS2序列,反向斑点杂交技术对沙眼衣原体进行检测和基因分型。结果 巢式PCR扩增omp1基因的VS1-VS2序列的敏感性与质粒PCR符合率为98.2%(56/57)。优选出11条型特异性(A、B+Ba、C、D、E、F、G、H、I、J和K)和3条群特异性寡核苷酸探针:B群(B、Ba、D和E型),C群(A、C、H、I、J和K型)和中间群(F和G型)。特异性经过基因库中的BLAST程序比较和试验优化,结果显示各探针均与标准株呈特异性杂交。56例VS1-VS2 PER阳性的临床标本杂交法检测均阳性,共检出59株沙眼衣原体,包括8个基因型,其中以E、F、D和H型为主,占77.9%,分别为25.4%,22.0%,16.9%和13.6%。发现3例混合感染占5.4%,分别为D/E、D/F和F/K。结论 反向斑点杂交技术简便且快速,可直接对临床标本沙眼衣原体进行检测和基因分型。