Definitions for the level of evidence (I-IV) and grade of recommendation (A-C) are provided at the end of the "Major Recommendations" field.
Summary of Recommended Tests for Chlamydia trachomatis
Use of Tests with Appropriate Specimens
|First catch urine
||3 (No longer recommended)
NAAT: nucleic acid amplification test
- Test of choice
- Test of choice, not licensed
- Not recommended
*Men and women with persistent urethral symptoms where tests from other sites are negative or women who have undergone a hysterectomy
#Men who have sex with men (MSM) and commercial sex workers (CSW)
Appropriate Specimens for Men and Women
||First catch urine (specimen of choice) or urethral swab
|Men who have sex with men (MSM)
||First catch urine (specimen of choice) or urethral swab
Pharyngeal and rectal swab
|Women undergoing speculum examination
|Women not requiring speculum examination
||Self taken lower vaginal swab (specimen of choice) or first catch urine*
*Please note first catch urine samples may be less sensitive than endocervical or self taken lower vaginal swabs for the detection of C. trachomatis.
A variety of different tests are available to detect C. trachomatis in the genital tract. Their appropriate use depends on the characteristics of the test itself, the correct choice of sample and the clinical presentation of the patient. Currently there are no enzyme immuno-assays (EIAs), point of care tests (POCT) or deoxyribonucleic acid (DNA) probe technology that can be recommended for use in the diagnosis of C. trachomatis as they show inferior sensitivity and specificity to that of the recommended tests, the nucleic acid amplification tests (NAATs).
Nucleic Acid Amplification Tests
The role of nucleic acid amplification technology in the routine diagnosis of C. trachomatis infections has evolved over the last decade. There are a number of commercial assays currently available for routine use. The four (listed) below are commonly used in clinical practice:
- Abbott RealTime PCR assay (Abbott m2000, Abbott Diagnostics)
- BD ProbeTec ET, Strand displacement amplification (SDA, Becton Dickinson)
- COBAS Taqman, Polymerase chain reaction assay (Real-time PCR, Roche Diagnostics)
- GenProbe Aptima assay, Transcription mediated amplification assay (TMA, GenProbe)
These commercial assays all detect both viable and non-viable organisms but differ in their target sequence and their method of amplification. These assays also offer dual detection of C. trachomatis and Neisseria gonorrhoeae from a single specimen. For further information regarding the detection of N. gonorrhoeae using NAATs please see the National Guideline Clearinghouse (NGC) summary of the British Association of Sexual Health and HIV (BASHH) guideline UK national guideline for the management of gonorrhoea in adults, 2011 and BASHH/Health Protection Agency (HPA) guidance at www.hpa.org.uk .
NAATs are the tests of choice for urethral, cervical, vaginal (self-taken and clinician-obtained) and first catch urine specimens because of their superior sensitivity and high specificity (Ib, Grade A). All of the above commercial NAATs show adequate sensitivity and specificity. The testing platform selected must have a positive predictive value (PPV) over 90% and detect all known variants (IV, Grade C).
It is beyond the remit of these guidelines to recommend any one NAAT above another. The choice of testing platform will depend on a variety of factors including:
- The volume of samples to be processed
- Cost of reagents/equipment
- The relative sensitivity and specificity of the individual tests for different clinical specimens
- Whether the test is used to detect C. trachomatis alone or as a combined test for C. trachomatis and N. gonorrhoeae
No single test provides 100% sensitivity and specificity. Test problems include inhibitors, contamination, reproducibility and hormonal factors, which can lower sensitivity.
Although not licensed for these sites, NAATs may be used and potentially give valid results from pharyngeal and rectal specimens (IIa, Grade B). This should be validated locally for the individual platform used.
Confirming Positive NAATs by Another Technique
Only another NAAT is sensitive enough to confirm a positive result. (IIa, Grade B). Currently the HPA guidelines recommend that every positive chlamydia result should be confirmed using a NAAT, preferably with an assay of equal sensitivity but with a different target.
However recent data suggests that confirmatory testing may be unnecessary given that >90% of positive NAAT results will be confirmed. (III, Grade B) The data are available for genital specimens and for some platforms only. Further work is required to validate this strategy for extra-genital specimens. Regardless of the site tested, clinicians need to be aware of the potential for false positive results, particularly when using the test in a low prevalence population.
When the test result is equivocal (unconfirmed reactive), arrangements should be made to re-test the original sample and request a further sample. Where possible this sample should be tested using a NAAT assay of equal sensitivity but with a different target. (IIa, Grade B)
Inhibitors can be identified in specimens from all sites, in particular first-void urine. An internal amplification control to identify inhibition should be used and is available using some commercial kits. Not all NAATs include an internal control (see individual manufacturer's instructions).
Transport, Storage and Handling of Samples
Requirements for the transport, storage and handling of samples vary between commercial assays and the manufacturer's instructions should be followed. It is recommended that results should be available within seven working days of the specimen being taken. If supplementary testing of a sample is required then the results should be available within 14 working days of the specimen being taken.
All efforts need to be made to ensure all staff adhere to the correct test procedures, avoid sampling errors and environmental contamination. Participation in an External Quality Assurance (EQA) programme needs to be encouraged (essential in an accredited laboratory) to minimise common errors and ensure that reproducibility of testing is maintained.
New Variant Chlamydia trachomatis
In November 2006 a C. trachomatis strain with a deletion in the cryptic plasmid was discovered in Sweden (new-variant – nvCT). The deletion, 377bp in length, affected the target sequence of some commercial tests resulting in false negative results. Isolated cases were found in Norway, Ireland, Denmark, France and Scotland. It is uncertain why this strain appears here and only enhanced surveillance will show whether it will be found elsewhere.
The target sequence of some commercially available kits has since been modified, but not all kits are capable of detecting the nvCT. Current NAATs are being modified and withdrawn so it is important that clinicians and microbiologists are aware of the status of the test they use. Further variants could occur and may not be detected by current commercial assays.
Lymphogranuloma venereum (LGV) is caused by the more invasive L serovars (L1, L2, L2a, and L3) of C. trachomatis. Since the end of 2003, an ongoing outbreak of LGV proctitis has been reported in Europe and North America among men who have sex with men (MSM), which has been strongly associated with human immunodeficiency virus (HIV) infection. It is recommended that all MSM with a positive rectal chlamydia NAAT who report rectal symptoms or who are a contact of someone with LGV should have a sample sent to the Sexually Transmitted Bacteria Reference Laboratory (STBRL), Health Protection Agency Centre for Infections, London, United Kingdom [UK] or the Scottish Bacterial Sexually Transmitted Infections Reference Laboratory (SBSTIRL). A real-time polymerase chain reaction (PCR) assay is now available at STBRL and SBSTIRL which performs well for the detection of LGV, non-LGV or dual infections from rectal specimens (IV, Grade C). Further information regarding the LGV enhanced surveillance protocol is available at http://www.hpa.org.uk/Topics/InfectiousDiseases/InfectionsAZ/LGV/EnhancedSurveillanceSystem/ and the LGV guideline is available at http://www.bashh.org/documents/92/92.pdf .
The traditional method of diagnosing C. trachomatis was by cell culture. Although chlamydiae are bacteria, they cannot be cultivated in non-living or cell free media. However, few laboratories in the UK still offer this service. Cell culture procedures are expensive, labour intensive and time consuming. Cell culture allows viable isolates to be obtained from cases where therapeutic failure is suspected, to test for antimicrobial resistance. At best the sensitivity of cell culture is probably no more than 75%, although specificity should be 100% if a C. trachomatis major outer membrane protein (MOMP) specific stain is used. Cell culture is no longer required for medico-legal purposes (IV, Grade C).
Appropriate Specimens for Testing for Chlamydia trachomatis
The performance of different tests for C. trachomatis can be influenced by the test specimen used.
First Catch Urine (FCU)
First catch urine comprises the first 15-50 mls of urine passed at any time of the day (see individual pack inserts). The patient must not have urinated for at least one hour (or 2 hours for some kits). FCU is licensed in both men and women for most NAATs, FCU in women is less sensitive than using endocervical or self taken lower vaginal specimens. FCUs are the preferred specimens for men (IIa, Grade B).
Cervical samples are suitable for all tests. Specimens should be taken during a speculum examination with the swab inserted into the cervical os using the manufacturers swab collection packs and rotated for a few seconds (IIa, Grade B).
Both male and female urethral samples are suitable for all tests. For men either a urethral specimen or first catch urine are ideal specimens although a urethral specimen may cause discomfort. For a male urethral specimen the swab is inserted into the urethra 2-4 cm and rotated one or more times (Grade C). A urethral sample may be required in women with persistent urethral symptoms in whom specimens for C. trachomatis at other sites are negative or in those who have undergone a hysterectomy (IV, Grade C). For a female urethral specimen introduce the swab 1 cm into the urethra and rotate one or more times (IV, Grade C).
A cotton tipped swab should be rubbed over the posterior pharynx and tonsillar crypts.
Pharyngeal samples are licensed for use with the tissue culture technique (IIa, Grade A).
NAATs are not licensed for use with pharyngeal specimens but accumulating evidence suggests they perform well (IIa, Grade B). Pharyngeal specimens should be taken and tested for C. trachomatis in MSM (IIa, Grade B) and commercial sex workers (CSW) reporting sexual behaviours which may result in pharyngeal infection (IV, Grade C). There is insufficient evidence to recommend testing for pharyngeal C. trachomatis using NAATs in heterosexual men and women (IV, Grade C).
There is limited data regarding self taken pharyngeal specimens among MSM but what there is suggests similar sensitivity and specificity to samples obtained by healthcare workers.
A cotton tipped swab should be rubbed against the rectal wall. This should ideally be taken at proctoscopy but data suggests that rectal swabs taken without proctoscopy have similar sensitivity.
Tissue culture is validated for detecting C. trachomatis from rectal specimens (IIa, Grade B). There are no licensed NAATs for the detection of C. trachomatis in rectal specimens but data is available supporting the validity of these tests for use here. Routinely available NAATs for C. trachomatis will detect all serovars including LGV (IIa, Grade B). Rectal specimens should be taken and tested for C. trachomatis in MSM (IIa, Grade B) and CSWs (IV, Grade C) reporting sexual behaviours which may result in rectal infection. There is insufficient evidence to recommend testing for rectal C. trachomatis using NAATs in heterosexual men and women (IV, Grade C).
There is limited data regarding self taken rectal specimens among MSM but this suggests similar sensitivity and specificity compared to samples obtained by healthcare workers via proctoscopy.
Vaginal Specimens (VS)
Insert the swab into the vagina, about two inches and gently rotate the swab for 10 to 30 seconds.
Some commercially available NAATs are licensed for use with vaginal samples, either clinician obtained or self-taken (further information is available from each manufacturer's kit insert). Vaginal specimens have been demonstrated by a number of workers to produce similar sensitivity to cervical testing (IIa, Grade B). Vulval specimens are not recommended for testing for C. trachomatis.
A cotton tipped swab should be used to for C. trachomatis from the conjunctiva. NAATs have a higher sensitivity to detect infection from the conjunctiva than other methodologies.
Specimens from the Glans Penis
NAATs have poor sensitivity to detect C. trachomatis from clinician and self-taken specimens from the glans penis and cannot be recommended (IIa, Grade B).
Factors Which May Alter Recommended Tests or Test Sites
Recommendations for testing are unaltered for:
- Sexual contacts of known chlamydia infection
- Sex workers – pharyngeal and rectal specimens should be taken and tested for C. trachomatis in CSWs (IV, Grade C) reporting sexual behaviours which may result in infection at these sites.
- Pregnant women
- Presence or absence of symptoms
However testing in women who have undergone a hysterectomy should be undertaken using either a FCU, vulval-vaginal (VV) specimen or a urethral swab (IV, Grade C).
Sexual Assault Victims
Culture is no longer recommended for detecting C. trachomatis at all exposed sites following sexual assault in adults because of its low sensitivity. It is recommended that a NAAT be taken from all exposed sites. (IIa, Grade C). Confirmation of a positive NAAT for C. trachomatis should be undertaken using a NAAT with a different target in medico-legal cases. This is available in some laboratories and the Sexually Transmitted Bacterial Reference Laboratory or the SBSTIRL.
Frequency of Repeat Testing in an Asymptomatic Patient
Re-exposure to a possible source of chlamydia should lead to re-screening if the patient re-presents. However there is no evidence currently to guide the frequency of repeat testing in those without a clear history of re-exposure to chlamydia. The Department of Health (DoH) Chlamydia Screening Programme for under 25's recommends repeat testing annually or every time someone has a new sexual partner.
When to Test Following Potential Exposure to Infection
Individuals should be advised to have a test for chlamydia with a NAAT when they first present and, if potential exposure occurred within the last two weeks, they should also be asked to return for a repeat NAAT two weeks after the exposure (IV, Grade C). (http://www.bashh.org/documents/1743/1743.pdf )
Test of Cure (TOC) Following C. trachomatis treatment
TOC (repeat testing to confirm clearance of infection) is not routinely recommended if:
- Standard treatment has been given
- There is confirmation that the patient has adhered to therapy
- There is no risk of re-infection
However, if these criteria cannot be met or if the patient is pregnant a TOC is advised. This should be taken using the same technique and sample type as used for the initial testing. Few data are however available regarding the optimal time to undertake a TOC. It is recognised that NAATs will detect residual DNA/ribonucleic acid (RNA) even after successful treatment of the organism four to six weeks after treatment (IIb, Grade B).
Levels of Evidence
||Type of Evidence
||Evidence obtained from meta-analysis of randomised controlled trials
||Evidence obtained from at least one randomised controlled trial
||Evidence obtained from at least one well-designed controlled study without randomisation
||Evidence obtained from at least one type of well-designed quasi-experimental study
||Evidence obtained from well-designed, non-experimental descriptive studies, such as comparative studies, correlation studies and case control studies
||Evidence obtained from expert committee reports or opinions and/or clinical experience of respected authorities
Grades of Recommendation
|A (Evidence levels Ia, Ib)
||Requires at least one randomised controlled trial as part of the body of literature of overall good quality and consistency addressing the specific recommendation
|B (Evidence levels IIa, IIb, III)
||Requires availability of well-conducted clinical studies but no randomised clinical trials on the topic of recommendation
|C (Evidence level IV)
||Requires evidence from expert committee reports or opinions and/or clinical experience of respected authorities. Indicates absence of directly applicable studies of good quality