Ligase Chain Reaction Research Articles

Screening for sexually transmitted infections in substance abuse treatment programs

Objectives: We evaluated the prevalence of the sexually transmitted infections (STIs) chlamydia and gonorrhea in clients at a methadone maintenance program and a residential detoxification program. Methods: We collected urine specimens for chlamydia and gonorrhea ligase chain reaction testing and assessed sexual, substance abuse and STI histories. Results: Of 700 subject assessments, 490 occurred among detoxification clients and 210 in methadone maintenance. Chlamydia trachomatis was detected in 5/700 (0.9, 95% CI=0.1–1.8%) and Neisseria gonorrhoeae in none. All chlamydia infected subjects were recruited from the detoxification program. Subjects reported high risk sexual behavior: 17% reported commercial sex exchange, and 22% reported inconsistent condom use with multiple sexual partners during the prior 2 months. Conclusion: Based on prevalence in Boston, MA, universal screening for STI in substance abuse treatments programs is not warranted. However, routine screening for younger substance abusers and in communities with high prevalence should be considered.

The clinical and economic consequences of screening young men for genital chlamydial infection.

Wide-scale application of urine-based screening of asymptomatic men for chlamydial infection has not been thoroughly assessed. The goal was to compare clinical and economic consequences of three strategies: (1). no screening, (2). screening with ligase chain reaction (LCR) assay of urine, and (3). prescreening urine with a leukocyte esterase test (LE) and confirming positives with LCR. We used a decision analytic model. At a chlamydia prevalence of 5%, the no screening cost was US dollars 7.44 per man screened, resulting in 522 cases of pelvic inflammatory disease (PID) per 100000 men. LE-LCR was most cost-effective, preventing 242 cases of PID over no screening at an additional cost of US dollars 29.14 per male screened. The LCR strategy prevented 104 more cases of PID than LE-LCR but cost US dollars 22.62 more per male screened. For this to be more efficient than LE-LCR, the LCR assay cost needed to decline to <or=US dollars 18. At a chlamydia prevalence of 5%, LE-LCR is the most efficient use of resources. If LCR cost decreases or chlamydia prevalence increases, the LCR strategy is favored.

Opportunistic screening for genital chlamydial infection. II: Prevalence among healthcare attenders, outcome, and evaluation of positive cases

Objectives: To determine the prevalence and treatment outcomes among young women screened opportunistically for genital Chlamydia trachomatis and to evaluate the impact of screening in those participating. Design: An opportunistic...

Opportunistic screening for genital chlamydial infection. I: Acceptability of urine testing in primary and secondary healthcare settings

Objectives: To determine the acceptability of opportunistic screening for Chlamydia trachomatis in young people in a range of healthcare settings. Design: An opportunistic screening programme (1 September 1999 to 31...

Ligase chain reaction.

AbstractThe ligase chain reaction (LCR) is one of many techniques developed in recent years to detect specific nucleic acid sequences by amplification of nucleic acid targets. The LCR has been used for genotyping studies to detect tumors and identify the presence of specific genetic disorders such as sickle cell disease caused by known nucleotide changes that occur as a result of point mutations and has now become widely used in infectious disease detection, both in the diagnostic and research settings, primarily focusing on infections caused by microbes that have proven difficult to detect by traditional culture techniques. The LCR is now recognized as the method of choice for detection of urogenital infections due to Chlamydia trachomatis because of its greater sensitivity as compared to traditional cell culture or nonamplifed DNA probes or antigen-detection assays. Other uses of the LCR have also been reported (1–8). When used for detection of infectious diseases, amplification tests such as the LCR have the additional advantages in that they do not require viable organisms in a specimen, a single specimen can be used to detect multiple different pathogens, provided suitable primers are available, and easily obtained specimens such as urine can be used for diagnostic purposes, making screening of large numbers of persons practical, as well as facilitating research to better understand the epidemiology of specific diseases.KeywordsChlamydia TrachomatisUrine SpecimenAbbott DiagnosticsLigase Chain ReactionLigase Detection ReactionThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Prevalence of Chlamydia trachomatis and acceptability of screening in asymptomatic women attending antenatal clinics

Antenatal genital tract infection with Chlamydia trachomatis (CT) is associated with a two to threefold risk of preterm birth. The prevalence of CT in women attending antenatal clinics is unknown. The aim of this study was to determine the prevalence of infection and acceptability of screening for CT in an asymptomatic antenatal population. Women attending antenatal clinics in Glasgow (Princess Royal Maternity Hospital) and Aberdeen (Aberdeen Royal Infirmary) were asked to provide a first void urine sample for screening for CT using the ligase chain reaction. Women were also sent a postal questionnaire which asked for their views of screening. The prevalence of CT was highest in the under-20 age group, with a percentage prevalence (95% CI) of 12.1% (8.6 – 16.7) (see Table). Neither the presence of symptoms or signs of infection nor parity were correlated significantly with the risk of CT carriage. 77% of women thought that antenatal clinics were appropriate settings for screening for CT.

Pregnant adolescents at risk: Sexual behaviors and sexually transmitted disease prevalence

Objective: The purpose of this study was to determine the level of high-risk sexual behaviors and the prevalence of chlamydia and gonorrhea among pregnant adolescents. Study Design: These analyses used data from 203 pregnant and 209 nonpregnant adolescents who were recruited from public health clinics. Data sources included interviewer-administered questionnaires, ligase chain reaction tests for chlamydia/gonorrhea in the third trimester of pregnancy, and state health department reports of chlamydia/gonorrhea. Statistical analyses included logistic regression. Results: Pregnant adolescents were significantly more likely to have not used condoms during sexual intercourse in the past 30 days compared with nonpregnant adolescents, although other sexual risk behaviors were reduced. Nineteen percent of pregnant adolescents had chlamydia or gonorrhea diagnosed during the pregnancy. Conclusion: Pregnant adolescents have high levels of sexually transmitted diseases during pregnancy, and many adolescents use condoms inconsistently. Prenatal care providers may be in a unique position to decrease sexually transmitted diseases among pregnant adolescents by encouraging the reduction of risk behaviors and aggressively screening for sexually transmitted diseases as part of routine prenatal care. (Am J Obstet Gynecol 2003;188:63-70.)

Screening of mutations in the ras family of oncogenes by polymerase chain reaction-based ligase chain reaction.

AbstractLung cancer is the most common fatal type of cancer in the developed world. The overwhelming majority of cases of lung cancer are caused by tobacco products (1), and even with the best therapeutic approaches, less than 15% of diagnosed cases survive 5 years (2). It has been noted that even after smoking cessation, the risk of lung cancer remains elevated for over 15 years (3). This observation clearly indicates that lung cancer has a protracted course developing over a 10-20-yr period, from the moment in which an epithelial cell becomes initiated by chronic exposure to chemical insults until a clinically evident cancer is detected. The long latency period generated by this biology provides an important window of opportunity to find transformed precancerous cells in high-risk populations and intervene in a timely manner.KeywordsMagnesium AcetateLigase Chain ReactionInvariant PrimerTaqMan ReactionPositive Control Cell LineThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

The accuracy and efficacy of screening tests for Chlamydia trachomatis: a systematic review.

Screening women for lower genital tract infection with Chlamydia trachomatis is important in the prevention of pelvic inflammatory disease, ectopic pregnancy and infertility. This systematic review aims to state clearly which of the available diagnostic tests for the detection of C. trachomatis would be most effective in terms of clinical effectiveness. The review included all studies published from 1990 onward that evaluated diagnostic tests in asymptomatic, young, sexually active populations. Medline and Embase were searched electronically and key journals were hand-searched. Further studies were identified through the Internet and contact with experts in the field. All studies were reviewed by two reviewers and were scored by Irwig's assessment criteria. Additional quality assessment criteria included a documented sexual history and recording of previous chlamydial infection. The reviews were subjected to meta-analysis and meta-regression. The 30 studies that were included examined three types of DNA-based test--ligase chain reaction (LCR), PCR and gene probe--as well as enzyme immuno-assay (EIA). The results showed that while specificities were high, sensitivities varied widely across the tests and were also dependent on the specimen tested. Pooled sensitivities for LCR, PCR, gene probe and EIA on urine were 96.5%, 85.6%, 92% and 38%, respectively, while on cervical swabs the corresponding sensitivities of PCR, gene probe and EIA were 88.6%, 84% and 65%. Meta-analysis demonstrated that DNA amplification techniques performed best for both urine and swabs in low prevalence populations. We conclude that nucleic acid amplification tests used on non-invasive samples such as urine are more effective at detecting asymptomatic chlamydial infection than conventional tests, but there are few data to relate a positive result with clinical outcome.

Acceptability of sexually transmitted infection screening among women in short-term substance abuse treatment.

Substance abuse treatment centers provide an opportunity to offer sexually transmitted infection (STI) screening to a high-risk and hard-to-reach population. The goal was to assess STI prevalence, risk factors, and acceptability of STI screening among females at substance abuse treatment centers with use of urine testing by ligase chain reaction and self-collected swab specimens. Adult, female inpatients were offered free testing and treatment for chlamydia infection, gonorrhea, and trichomonas infection. Interviews were conducted to collect risk behavior data. Eighty-six percent of inpatients (180/209) accepted testing. Twenty-three percent (41/177) had an STI. Of those with an STI, 90% (37/41) had trichomonas infection. All 41 infected patients received treatment. Drug use before sex, exchange of sex for money/drugs, and any gynecological complaint were significantly associated with infection. Most women were uninsured (76%). Only 45% had undergone a medical examination in the past year. STI screening is highly acceptable among women in substance abuse treatment centers. Substance users are at high risk for STIs and may not otherwise receive medical care.

Optimal resource allocation for curing Chlamydia trachomatis infection among asymptomatic women at clinics operating on a fixed budget.

The goal was to determine the optimal strategy for screening coverage, test selection, and treatment for infection in asymptomatic women for a given family-planning-program budget. We developed a resource allocation model to determine the optimal strategy using data from 5078 visits by women universally screened for infection in a publicly funded family planning clinic system in Philadelphia. We maximized the number of infected women cured from the clinic perspective and maximized the cost-savings from the healthcare system perspective. The model incorporated the following age distributions: <20 years (27%), 20 to 24 years (30%), and >24 years (43%), with prevalences of 10.6%, 6.9%, and 2.3%, respectively. We modeled two screening test assays (DNA probe and ligase chain reaction [LCR] for cervical specimens) and two treatments (doxycycline and azithromycin). The model allowed for different test and treatment choices by age group. At the baseline annual budget of $6 per visit, the strategy that maximized both the number of infected women cured and cost savings would be to screen all women with DNA probe and to treat all women with positive tests with azithromycin. This strategy would result in 183 women cured at a cost-savings of $140,176. Sensitivity analysis showed that the total budget had a great impact on the optimal strategy, incorporating screening coverage, test selection, and treatment. Using resource allocation models enables clinic managers operating with a fixed budget to identify a strategy that maximizes the number of asymptomatic women cured and cost savings when the clinic age distribution and age-specific prevalences are known.

Duration of persistence of gonococcal DNA detected by ligase chain reaction in men and women following recommended therapy for uncomplicated gonorrhea.

Neisseria gonorrhoeae infection remains relatively common in the United States, representing a public health challenge. Ligase chain reaction (LCR) is both highly sensitive and specific for the detection of N. gonorrhoeae in urine and patient-obtained vaginal swab specimens. Because of the LCR test's exquisite sensitivity, it may potentially detect DNA from nonviable organisms following effective therapy, leading to false-positive test results and unnecessary additional treatment. The purpose of the present study was to determine the duration that gonococcal DNA is detectable by LCR following therapy for uncomplicated gonococcal infection. One hundred thirty men and women between the ages of 16 and 50 years presenting to a sexually transmitted disease clinic with urogenital gonorrhea were enrolled. After the standard history was taken and a genital examination was done, the patients were asked to submit either a urine specimen (men) or a urine specimen plus a self-obtained vaginal swab specimen (women) for N. gonorrhoeae testing by LCR at the initial visit and each day during the study period. At enrollment, patients were treated with single doses of ofloxacin, cefixime, or ceftriaxone. The median time to a negative urine LCR test result was 1 day for the men (mean, 1.6 +/- 0.14 days) and 2 days for the women (mean, 1.7 +/- 0.19 days). Among the women the clearance time was significantly longer for vaginal specimens (mean, 2.8 +/- 0.30 days) than for urine specimens (mean, 1.7 +/- 0.11 days). Irrespective of patient gender and specimen type, gonococcal DNA can be expected to be absent from urogenital specimens within 2 weeks following successful therapy.

Quantification of Chlamydia trachomatis elementary bodies in urine by ligase chain reaction.

Modification of the standard Chlamydia trachomatis ligase chain reaction (LCR) detection assay resulted in a quantitative test. Sample rates from C. trachomatis standards ranging from 32 to 1,048,576 elementary bodies (EB)/ml of urine exhibited a linear correlation with concentration. Quantitative LCR (qLCR) was used to measure the number of EB per milliliter in 158 urine samples from women in Madagascar that tested positive for C. trachomatis by the standard LCR detection assay. C. trachomatis concentrations displayed an apparent bimodal distribution, with approximately one-third of samples (37%) in a peak ranging from 32 to 1,015 EB/ml (median = 297 EB/ml) and the remainder (63%) in a grouping with relatively higher concentrations, ranging from 1,086 to 218,670 EB/ml (median = 7,389 EB/ml). qLCR will be useful for studies of chlamydial infection aimed at understanding the associations of organism burden with clinical manifestations and transmission.

Failure of Commercial Ligase Chain Reaction To Detect Mycobacterium tuberculosis DNA in Sputum Samples from a Patient with Smear-Positive Pulmonary Tuberculosis Due to a Deletion of the Target Region

Failure of Commercial Ligase Chain Reaction To Detect <i>Mycobacterium tuberculosis</i> DNA in Sputum Samples from a Patient with Smear-Positive Pulmonary Tuberculosis Due to a Deletion of the Target Region

Screening for Chlamydia trachomatis infection is indicated for women under 30 using emergency contraception

A total of 838 women attending a large family planning clinic in Scotland for emergency contraception were offered screening for Chlamydia trachomatis infection. 569 were screened using ligase chain reaction test in first void urine at the time of presenting for emergency contraception and were retested 1 or 2 weeks later. Women aged under 20 and over 30 years were significantly more likely to decline to be tested than women aged 20 to 30. The prevalence of chlamydia was 7.6% in woman aged 24 or less, 5.3% in women aged 25 to 29, and 1.2% in women aged 30 or more. Only two women (< 1%) who tested negative at the time of using EC were positive 1 or 2 weeks later. Women under age 30 who use EC should be offered screening for chlamydia infection and testing at the time they attend for EC is adequate to detect the great majority of infected women.

Use of ligase chain reaction for enhanced detection of Phytophthora infestans

Sequences of the internal transcribed spacer region 2 (ITS 2) of ribosomal DNA from 10 different species of Phytophthora were aligned, and regions of dissimilarity were used to construct ligase chain reaction (LCR) primers for detecting the late blight pathogen, Phytophthora infestans. Ligase chain reaction primers allow differentiation of species down to a single base pair difference. The primers have greater specificity than previously described polymerase chain reaction (PCR) primers and allow differentiation of P. infestans, Phytophthora mirabilis, and Phytophthora phaseoli, which contain identical ITS sequences, from many other Phytophthora species. Ligase chain reaction was coupled with PCR to increase sensitivity of P. infestans detection and can serve as an enhancement to existing PCR protocols. By labeling LCR primers with biotin and digoxigenin, the LCR was combined with an ELISA-based detection system to improve sensitivity and ease of application.

Parallel observation of four methods for screening women under 25 years of age for genital infection with Chlamydia trachomatis

Objectives: To compare four methods of screening women for Chlamydia trachomatis in an obstetrics and gynaecology department. Study design: A total of 303 healthy women under 25 years were recruited from antenatal, induced abortion, and family planning clinics. Each underwent parallel testing of endocervical specimens by enzyme immunoassay (EIA) and ligase chain reaction (LCR), vulval swabs by LCR, and urine by LCR. Outcome measures included sensitivity, specificity, acceptability of each method, and the influence of pregnancy. Results: Overall prevalence (95% CI) was 9.9% (6.8–14%). All methods had a high rate of detection (75–100%), not affected by pregnancy. Urine was most acceptable, followed by vulval swabbing. Conclusions: Opportunistic screening of women under 25 years attending obstetric and gynaecology affiliated clinics found high rates of C. trachomatis infection. Both urine and vulval swab methods were highly sensitive, acceptable, and not affected by pregnancy status. Due to pragmatic issues surrounding the urine method, screening by vulval swabs deserves wider recognition.

Early identification of Mycobacterium tuberculosis complex in BACTEC cultures by ligase chain reaction.

A total of 1431 acid-fast bacilli (AFB) in Bactec culture vials from 1427 patients was differentiated by the Bactec NAP method and tested by the LCx Mycobacterium tuberculosis ligase chain reaction system. In all, 1321 of 1325 M. tuberculosis complex (MTBC) isolates were correctly detected by the LCx assay. All the 106 non-tuberculous mycobacteria (NTM) isolates were negative by the LCx assay. No false MTBC-positive result was obtained from NTM isolates. However, the LCx assay failed to detect four MTBC isolates from one patient. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were 99.7, 100%, 100% and 96.4%, respectively. These data suggest that the LCx system can be used to identify MTBC in AFB-positive Bactec broth cultures when the growth index is > or = 100. The method gives a 100% PPV and allows a faster turnaround time for MTBC than the NAP test.

Coexistence of urethritis with genital ulcer disease in South Africa: influence on provision of syndromic management

Objective: To assess whether syndromic management of genital ulcer disease was sound, if based on the premise that men with genital ulcers rarely have a concomitant urethral infection. Methods: Specimens...

Accuracy of results obtained by performing a second ligase chain reaction assay and PCR analysis on urine samples with positive or near-cutoff results in the LCx test for Chlamydia trachomatis.

Nucleic acid amplification assays such as the ligase chain reaction and PCR have encountered reproducibility problems. The initial extract and a newly extracted aliquot of urine specimens (n = 120) which had signal-to-cutoff (S/CO) ratios above 0.80 by the LCx Chlamydia assay were retested. Nucleic acid was extracted from an additional urine sample for testing by the AMPLICOR PCR Chlamydia assay. Fifteen percent (18 of 120) of the urine specimens were negative by all repeat tests (initial mean S/CO ratio by the LCx Chlamydia assay, 0.93; S/CO ratio range, 0.80 to 3.30). Repeat testing of the 102 specimens with possible positive results by the LCx Chlamydia assay by use of the initially extracted aliquot confirmed the results for 95 (93.1%) of the specimens; repeat testing of a newly extracted aliquot confirmed the results for 87 (85.3%) of the specimens. Twenty specimens had discordant results by the two repeat LCx Chlamydia assays. A total of 78 of 102 (76.5%) of the specimens were positive by the AMPLICOR PCR, and the AMPLICOR PCR confirmed the results for 82.1% (78 of 95) and 89.6% (78 of 87) of the specimens positive by the two repeat LCx Chlamydia assays, respectively. Some of the discrepancies observed by multiple repeat tests may have been due to specimen mislabeling or contamination during performance of the procedure rather than to the LCx Chlamydia assay. Both assays suffered from a lack of reproducibility on repeat testing with a small proportion of specimens, probably due to the presence of low levels of DNA, the presence of variable amounts of amplification inhibitors, and the loss of DNA during extraction.