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Sexually Transmitted Diseases

ChlamydiaScreening Tests To Detect Chlamydia trachomatis and Neisseria gonorrhoeae Infections - 2002

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Prepared by
Robert E. Johnson, M.D.1
Wilbert J. Newhall, Ph.D.1
John R. Papp, Ph.D.2
Joan S. Knapp, Ph.D.2
Carolyn M. Black, Ph.D.2
Thomas L. Gift, Ph.D.1
Richard Steece, Ph.D.3
Lauri E. Markowitz, M.D.1
Owen J. Devine, Ph.D.1
Cathleen M. Walsh, Dr.P.H.1
Susan Wang, M.D.1
Dorothy C. Gunter, M.P.H.1
Kathleen L. Irwin, M.D.1
Susan DeLisle, M.P.H.1
Stuart M. Berman, M.D.1
Division of Sexually Transmitted Diseases Prevention
National Center for HIV, STD, and TB Prevention, CDC
Division of AIDS, STD, and TB Laboratory Research
National Center for Infectious Diseases, CDC
Association of Public Health Laboratories
Washington, D.C.


Since publication of CDC's 1993 guidelines (CDC. Recommendations for the prevention and management of Chlamydia trachomatis infections, 1993. MMWR 1993;42[No. RR-12]:1--39), nucleic acid amplification tests (NAATs) have been introduced as critical new tools to diagnose and treat C. trachomatis and Neisseria gonorrhoeae infections. NAATs for C. trachomatis are substantially more sensitive than previous tests. When using a NAAT, any sacrifice in performance when urine is substituted for a traditional swab specimen is limited, thus reducing dependence on invasive procedures and expanding the venues where specimens can be obtained. NAATs can also detect both C. trachomatis and N. gonorrhoeae organisms in the same specimen. However, NAATs are usually more expensive than previous tests, making test performance from an economic perspective a key consideration.

This report updates the 1993 guidelines for selecting laboratory tests for C. trachomatis with an emphasis on screening men and women in the United States. (In this report, screening refers to testing persons in the absence of symptoms or signs indicating C. trachomatis or N. gonorrhoeae infection.) In addition, these guidelines consider tests from an economic perspective and expand the previous guidelines to address detection of N. gonorrhoeae as well as C. trachomatis infections. Because of the increased cost of NAATs, certain laboratories are modifying manufacturers' procedures to improve test sensitivity without incurring the full cost associated with screening with a NAAT. Such approaches addressed in these guidelines are pooling of specimens before testing with a NAAT and additional testing of specimens whose non-NAAT test result is within a gray zone. This report also addresses the need for additional testing after a positive screening test to improve the specificity of a final diagnosis.

To prepare these guidelines, CDC staff identified pertinent concerns, compiled the related literature published during 1990 or later, prepared tables of evidence, and drafted recommendations. Consultants, selected for their expertise or disciplinary and organizational affiliations, reviewed the draft recommendations. These final guidelines are the recommendations of CDC staff who considered contributions from scientific consultants. These guidelines are intended for laboratorians, clinicians, and managers who must choose among the multiple available tests, establish standard operating procedures for collecting and processing specimens, interpret test results for laboratory reporting, and counsel and treat patients.

The material in this report originated in the National Center for HIV, STD, and TB Prevention, Harold W. Jaffe, M.D., Acting Director, and the Division of Sexually Transmitted Diseases Prevention, Harold W. Jaffe, M.D., Acting Director; and the National Center for Infectious Diseases, James M. Hughes, M.D., Director, and the Division of AIDS, STD, and TB Laboratory Research, Jonathan E. Kaplan, M.D., Acting Director.


An estimated 3 million Chlamydia trachomatis infections occur annually among sexually active adolescents and young adults in the United States (1). The majority of persons with C. trachomatis infection are not aware of their infection because they do not have symptoms that would prompt them to seek medical care ( 2). Consequently, screening is necessary to identify and treat this infection.

Untreated, C. trachomatis infections can lead to serious complications. In certain studies, ≤40% of women with untreated C. trachomatis infections experience pelvic inflammatory disease (PID) (3,4). Of these, the majority have symptoms that are too mild or nonspecific for them to seek medical treatment. Regardless of symptom severity, the consequences of PID are severe. Of those with PID, 20% will become infertile; 18% will experience debilitating, chronic pelvic pain; and 9% will have a life-threatening tubal pregnancy (5). C. trachomatis infection during pregnancy leads to infant conjunctivitis and pneumonia and maternal postpartum endometritis.

Among men, urethritis is the most common illness resulting from C. trachomatis infection. Complications (e.g., epididymitis) affect a minority of infected men and rarely result in sequelae. Among men who engage in receptive anal intercourse, the rectum is a common site of C. trachomatis infection. Rectal infections are usually asymptomatic, but can cause symptoms of proctitis or proctocolitis. C. trachomatis can cause conjunctivitis among adults and is a cause of sexually acquired reactive arthritis.*

Estimated tangible costs of C. trachomatis illness in the United States exceed $2.4 billion annually (6). Also critical are the intangible costs, including the psychological and emotional injury caused by infertility and ectopic pregnancy.

As of December 2000, all 50 states and the District of Columbia had enacted laws requiring the reporting of C. trachomatis cases. In 2001, C. trachomatis infections were the most commonly reported communicable infections, with a total of 783,242 reports to CDC (7). Rates of C. trachomatis infection for women are highest for adolescents (2,536/100,000 among women aged 15--19 years) and young adults (2,447/100,000 among women aged 20--24 years). These age groups had the highest rates of infection for men as well, although the peak rate occurred among men aged 20--24 years (605/100,000).

Reported rates of C. trachomatis infections have risen significantly during 1987--2001 (51--278 cases/100,000 persons) ( 7). This increase is probably caused by a combination of factors, including an increased awareness of the need to screen women for C. trachomatis infection, resulting in the initiation of screening programs in both public and private health-care settings, improvement in the sensitivity of diagnostic tests, improved surveillance and reporting systems, and continued high infection rates.

Introduction of large-scale screening programs (e.g., one initiated in the Department of Health and Human Services Region X [Alaska, Idaho, Oregon, and Washington] family planning clinics in 1988) have been followed by a reduction in C. trachomatis positivity rates by ≤60% (7--9). C. trachomatis screening programs have been initiated throughout the United States that are based on such demonstration projects.

In 2001, Neisseria gonorrhoeae was second in frequency only to C. trachomatis among reported communicable infections in the United States, with 361,705 reported cases (7). The age distribution of N. gonorrhoeae infections is similar to that for C. trachomatis infections. Also similar to C. trachomatis, uncomplicated N. gonorrhoeae infection is usually confined to the mucosa of the cervix, urethra, rectum, and throat; N. gonorrhoeae infection is often asymptomatic among females; and, if untreated, N. gonorrhoeae infection can lead to PID, tubal infertility, ectopic pregnancy, and chronic pelvic pain (10). N. gonorrhoeae usually causes symptomatic urethritis among males, and occasionally results in epididymitis. Rarely, local infection disseminates to cause an acute dermatitis tenosynovitis syndrome, which can be complicated by arthritis, meningitis, or endocarditis (10). Also, similar to C. trachomatis, N. gonorrhoeae can be acquired at birth. N. gonorrhoeae neonatal infection can cause severe conjunctivitis, which can result in blindness if untreated and, rarely, sepsis with associated meningitis, endocarditis, or arthritis. After the introduction of a national control program in the mid-1970s, the overall rate of reported N. gonorrhoeae infection had declined by 74% during 1975--1997 (7, 11). However, the rate increased by 8.1% during 1997--1999, followed by a limited decline in 2000--2001.

Culture testing for C. trachomatis and N. gonorrhoeae has been the reference standard against which all other tests have been compared. However, other tests have been needed because culture methods for C. trachomatis are difficult to standardize, technically demanding, and expensive. Culture for either agent is associated with problems in maintaining the viability of organisms during transport and storage in the diverse settings in which testing is indicated. Thus, diagnostic test manufacturers have developed nonculture tests that do not require viable organisms, including tests that can be automated. The first nonculture screening tests for C. trachomatis and N. gonorrhoeae included enzyme immunoassays (EIAs), which detect specific chlamydial or gonococcal antigens, and direct fluorescent antibody (DFA) tests for C. trachomatis, which use fluorescein-conjugated monoclonal antibodies that bind specifically to bacterial antigen in smears. These antigen-detection tests were followed by nucleic acid hybridization tests, which detect C. trachomatis-specific or N. gonorrhoeae-specific deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) sequences. With the availability of these nonculture tests, screening programs for C. trachomatis were initiated, and screening programs for N. gonorrhoeae began to change from culture to using the more convenient and, in remote settings, more reliable nonculture methods. The primary drawback of these tests, chiefly for C. trachomatis, is that they fail to detect a substantial proportion of infections (12--23). Consequently, a new generation of nonculture tests, called nucleic acid amplification tests (NAATs), were developed that amplify and detect C. trachomatis-specific or N. gonorrhoeae-specific DNA or RNA sequences. These tests are substantially more sensitive than the first generation nonculture tests (12--23).

These guidelines are intended to assist laboratorians, clinicians, and managers 1) select screening tests for C. trachomatis or N. gonorrhoeae from the complex array of tests available; 2) establish standard operating procedures for collecting, processing, and analyzing specimens; and 3) interpret test results for laboratory reporting, counseling, and treating patients. The guidelines were developed through literature reviews and extended consultation with non-CDC sexually transmitted disease (STD) specialists.

Testing Technologies

The following discussion is a review of the complex array of technologies now available for laboratory diagnosis of C. trachomatis and N. gonorrhoeae infections. For this report, technologies are subdivided into those that are designed for 1) batch testing in a laboratory or 2) point-of-care testing as single tests or a limited number of tests performed while patients await results. Laboratory-based tests include culture, NAATs, nucleic acid hybridization and transformation tests, EIAs, and DFA tests. Point-of-care tests have long included the Gram-stained smear for N. gonorrhoeae. Point-of-care tests for C. trachomatis include solid-phase EIAs and a solid-phase optical immunoassay. The leukocyte esterase test (LET) is a dipstick test that is applied to urine specimens to screen for urinary tract inflammation (see Methods To Enhance Performance or Reduce Costs ). Personnel, quality assurance, and quality control requirements relating to the use of all tests for medical care are published in the Clinical Laboratory Improvement Amendments of 1988 (CLIA) regulations (24) and are linked to testing complexity.

Laboratory-Based Tests

Culture Tests

C. trachomatis Culture. Cell culture for C. trachomatis involves inoculating a confluent monolayer of susceptible cells with an appropriately collected and transported specimen. After 48--72 hours of growth, infected cells develop characteristic intracytoplasmic inclusions that contain substantial numbers of C. trachomatis elementary and reticulate bodies. These unique inclusions are detected by staining with a fluorescein-conjugated monoclonal antibody that is specific for the major outer membrane protein (MOMP) of C. trachomatis.

Cell culture methods vary among laboratories, leading to probable substantial interlaboratory variation in performance (25). For example, because of a larger inoculum and reduced risk of cross-contamination, the shell vial method of culture is more sensitive and specific than the( 96-well microtiter plate method (26,27). In certain laboratories, higher sensitivities are obtained by performing a blind pass in which an inoculated cell monolayer is allowed to incubate for 48--72 hours, after which the monolayer is disrupted and used to inoculate a fresh monolayer that is stained after another cycle of growth (28).

Tissue culture detection of C. trachomatis is highly specific if a C. trachomatis-MOMP-specific stain is used, because stained C. trachomatis inclusions have a unique appearance. Less specific inclusion-detection methods using EIA, iodine, and Giemsa are not( recommended (29,30). Certain CDC consultants believe that commercial stains employing monoclonal antibodies directed against lipopolysaccharide (LPS), which are genus-specific rather than species-specific, are more sensitive and more economical than species-specific monoclonal antibody stains directed against MOMP. Such stains might be suitable for routine use, but a species-specific stain would be preferable in situations requiring increased specificity.

The high specificity of cell culture and ability to retain the isolate make cell culture the first choice when the results will be used as evidence in legal investigations. In addition, cell culture is the only method by which a clinical isolate can be obtained for antimicrobial susceptibility testing. The relatively low sensitivity, long turnaround time, difficulties in standardization, labor intensity, technical complexity, stringent transport requirements, and relatively high cost are the primary disadvantages of cell culture isolation of C. trachomatis. Additional information regarding cell culture for C. trachomatis is available elsewhere (14,28,31).

N. gonorrhoeae Culture. Methods of gonococcal culture have been well-described elsewhere (12,32). Specimens are streaked on a selective (e.g.,Thayer-Martin or Martin-Lewis) or nonselective (e.g., chocolate agar) medium if specimens are from nonsterile or sterile sites, respectively. Inoculated media are incubated at 35ºC--36.5ºC in an atmosphere supplemented with 5% CO2 and examined at 24-hour intervals for ≤72 hours. Culture media for N. gonorrhoeae isolation include a base medium supplemented with chocolatized equine or bovine blood to support the growth of the gonococcus; selective media differ from routine culture media in that they contain antimicrobial agents (i.e., vancomycin, colistin, and an antifungal agent), which inhibit the growth of other bacteria and fungi. Supplemental CO2 can be supplied by a CO2 incubator, candle-extinction jar, or CO2-generating tablets.

A presumptive identification of N. gonorrhoeae isolates recovered from a genital specimen on selective medium can be made with a Gram stain and oxidase test. A presumptive identification indicates only that a Gram-negative, oxidase-positive diplococcus (e.g., a Neisseria species or Branhamella [Moraxella] catarrhalis) has been isolated from a specimen. Certain coccobacilli, including Kingella denitrificans, might appear to be Gram-negative diplococci in Gram-stained smears. A confirmed laboratory diagnosis of N. gonorrhoeae cannot be made on the basis of these tests alone. A presumptive test result is sufficient to initiate antimicrobial therapy, but additional tests must be performed to confirm the identity of an isolate as N. gonorrhoeae. Culture isolation is also suitable for nongenital tract specimens. Using selective media is necessary if the anatomic source of the specimen normally contains other bacterial species.

The sensitivity of culture can be monitored (i.e., quality-controlled) by evaluating results among males with urethral discharge. Culture results are compared with results obtained by using Gram-stained smear or by using nonselective medium. Vancomycin-sensitive strains, which were relatively common in certain areas in the past, now appear to be an uncommon cause of false-negative cultures (10 ,12 ,33 ,34).

The advantages of culture are high sensitivity and specificity, low cost, suitability for use with different types of specimens, and the ability to retain the isolate for additional testing. Retention of the isolate for additional testing might be indicated for medicolegal purposes, antimicrobial susceptibility determination, and subtyping of isolates. The major disadvantage of culture for N. gonorrhoeae is that specimens must be transported under conditions adequate to maintain the viability of organisms. Another disadvantage is that a minimum of 24--72 hours is required from specimen collection to the report of a presumptive culture result.


The common characteristic among NAATs is that they are designed to amplify nucleic acid sequences that are specific for the organism being detected. Similar to other nonculture tests, NAATs do not require viable organisms. The increased sensitivity of NAATs is attributable to their ability to produce a positive signal from as little as a single copy of the target DNA or RNA. Commercial tests differ in their amplification methods and their target nucleic acid sequences. The Roche Amplicor® (manufactured by Roche Diagnostics Corporation, Basel, Switzerland) test uses polymerase chain reaction (PCR); the Abbott LCx® (Abbott Laboratories, Abbott Park, Illinois) test uses ligase chain reaction (LCR); and the Becton Dickinson BDProbeTec ET (Becton, Dickinson and Company, Franklin Lakes, New Jersey) test uses strand displacement amplification to amplify C. trachomatis DNA sequences in the cryptic plasmid that is found in >99% of strains of C. trachomatis. The Gen-Probe APTIMA (Gen-Probe, Incorporated, San Diego, California) assay for C. trachomatis uses transcription-mediated amplification (TMA) to detect a specific 23S ribosomal RNA target. These nucleic acid amplification methods are also used to detect N. gonorrhoeae. The target for the Roche Amplicor test for N. gonorrhoeae is a 201 base pair sequence within the cytosine methyltransferase gene M:Ngo P11. The Abbott LCx test for N. gonorrhoeae detects a 48 base-pair sequence in the Opa genes, ≤11 copies of which occur per cell, whereas the BDProbeTec ET for N. gonorrhoeae detects a DNA sequence that is within the multicopy pilin gene-inverting protein homologue. The Gen-Probe APTIMA Combo 2 version of TMA detects the 16S ribosomal RNA (rRNA) of N. gonorrhoeae. Additional overviews of the various amplification methods can be found in other published reports ( 16 ,35 ,36) and in materials available from the manufacturers.

The majority of commercial NAATs have been cleared by the Food and Drug Administration (FDA) to detect C. trachomatis and N. gonorrhoeae in endocervical swabs from women, urethral swabs from men, and urine from both men and women. In addition, other specimens (e.g., those from the vagina [37--50] and eye [51--53]) have been used with satisfactory performance, although these applications have not been cleared by FDA. Testing of rectal and oropharyngeal specimens with NAATs has had limited evaluation and is not recommended.

The ability of NAATs to detect C. trachomatis and N. gonorrhoeae without a pelvic examination or intraurethral swab specimen (for males) (e.g., by testing urine) is a key advantage of NAATs, and this ability facilitates screening males and females in other than traditional screening venues (e.g., STD and family planning clinics). A disadvantage of NAATs is that specimens can contain amplification inhibitors that result in false-negative results. Certain manufacturers provide amplification controls to detect inhibition. Selected NAATs might be substantially less sensitive than non-NAATs when performed on urine than when performed on endocervical specimens ( 54--56) or male urethral swabs (54,56). Although not documented by published head- to-head studies, the majority of CDC consultants believe that non-NAATs are substantially less sensitive than NAATs when used on urine specimens. The nucleic acid primers employed by commercial NAATs for C. trachomatis are not known to cross-react with DNA from other bacteria found in humans. However, the primers employed by certain NAATs for N. gonorrhoeae might cross-react with nongonococcal Neisseria species (54 ,56 ,57). NAATs are also more susceptible than non-NAATs to false-positive results because of contamination if strict quality control procedures are not applied.

Nucleic Acid Hybridization (Nucleic Acid Probe) Tests

Two nucleic acid hybridization assays are FDA-cleared to detect C. trachomatis or N. gonorrhoeae: the Gen-Probe PACE® 2 and the Digene Hybrid Capture® II assays. Both the PACE and Hybrid Capture assays can detect C. trachomatis or N. gonorrhoeae in a single specimen. The PACE 2C test and the Hybrid Capture II CT/GC versions of these tests do not differentiate between the two organisms and, when positive, should be followed by tests for each organism to obtain an organism-specific result. In the Gen-Probe hybridization assays, a DNA probe that is complementary to a specific sequence of C. trachomatis or N. gonorrhoeae rRNA hybridizes with any complementary rRNA that is present in the specimen (58). A competitive probe version of the PACE 2 assay is commercially available to augment specificity. In this version, the test is repeated on initially positive specimens with and without adding an unlabeled probe. The unlabeled probe competitively inhibits binding of the labeled probe; a reduction in signal when the assay is performed with the unlabeled probe is interpreted as verification of the initial positive test result.

RNA hybridization probes in the Digene assay are specific for DNA sequences of C. trachomatis and N. gonorrhoeae, including both genomic DNA and cryptic plasmid DNA (59). Technical requirements and expertise necessary for performing nucleic acid hybridization tests are similar to those for the EIAs, which are described in this report. One of the advantages of the nucleic acid hybridization tests is the ability to store and transport specimens for ≤7 days without refrigeration before receipt and testing by the laboratory.

Nucleic Acid Genetic Transformation Tests

The Gonostat® test (Sierra Diagnostics, Incorporated, Sonora, California) uses a gonococcal mutant that grows when transformed by DNA extracted from a swab specimen containing N. gonorrhoeae. N. meningitidis causes false-positive results (60). The test has received limited evaluation in published studies (61--64), which include an evaluation of its use with mailed specimens (62). A genetic transformation test is not available for detection of C. trachomatis infection.

EIA Tests

A substantial number of EIA tests have been marketed for detecting C. trachomatis infection. By contrast, the performance and cost characteristics of EIA tests for N. gonorrhoeae infection have not made them competitive with culture (32). C. trachomatis EIA tests detect chlamydial LPS with a monoclonal or polyclonal antibody that has been labeled with an enzyme. The enzyme converts a colorless substrate into a colored product, which is detected by a spectrophotometer. Specimens can be stored and transported without refrigeration and should be processed within the time indicated by the manufacturer. One disadvantage of the EIA methods that detect LPS is the potential for false-positive results caused by cross-reaction with LPS of other microorganisms, including other Chlamydia species (28, 30, 65, 66). Manufacturers have developed blocking assays that verify positive EIA test results. The test is repeated on positive specimens with the addition of a monoclonal antibody specific for chlamydia LPS. The monoclonal antibody competitively inhibits chlamydia-specific binding by the enzyme-labeled antibody; a negative test result when using the blocking antibody is interpreted as verification of the initial positive test result. EIA tests should not be used with rectal specimens because of cross-reactions with fecal bacteria.

DFA Tests

Depending on the commercial product used, the antigen that is detected by the antibody in the C. trachomatis DFA procedure is either the MOMP or LPS molecule. Specimen material is obtained with a swab or endocervical brush, which is then rolled over the specimen well of a slide. After the slide has dried and the fixative applied, the slide can be stored or shipped at ambient temperature. The slide should be processed by the laboratory in ≤7 days after the specimen has been obtained. Staining consists of covering the smear with fluorescein-labeled monoclonal antibody that binds to C. trachomatis elementary bodies. Stained elementary bodies are then identified by fluorescence microscopy. Only C. trachomatis organisms will stain with the anti-MOMP antibodies used in commercial kits. The anti-LPS monoclonal antibodies used in certain commercial kits can cross-react with nonchlamydial bacterial species, as well as with LPS of C. pneumoniae and C. psittaci. DFA with a C. trachomatis-specific anti-MOMP monoclonal antibody is considered to be highly specific, when performed by an experienced microscopist. An additional advantage of DFA is that the quality of endocervical smears can be assessed by checking for the presence of columnar cells (see Collecting and Transporting Specimens for Screening). DFA requires that the laboratorian be competent in fluorescent microscopy and adequately trained in identifying fluorescein-stained C. trachomatis elementary bodies. DFA is best suited for laboratories that test a limited number of specimens, because the procedure is fatiguing and time-consuming. DFA tests have not been established as an initial test for the direct detection of N. gonorrhoeae in clinical specimens.

Serology Tests

Serology has limited value in testing for uncomplicated genital C. trachomatis infection and should not be used for screening because previous chlamydial infection frequently elicits long-lasting antibodies that cannot be easily distinguished from the antibodies produced in a current infection. More specific information regarding serologic assays for C. trachomatis antibody has been reported elsewhere (14). A serologic screening or diagnostic assay is not available for N. gonorrhoeae.

Point-of-Care Tests

C. trachomatis Point-of-Care Tests

Tests for C. trachomatis have been developed that can be performed within 30 minutes, do not require expensive or sophisticated equipment, and are packaged as single units. The results are read qualitatively. These so-called rapid or stat tests can offer advantages in physicians' offices, small clinics and hospitals, detention centers, and other settings where results are needed immediately (e.g., when decisions need to be made regarding additional testing or treatment while the patient is still present). These tests are classified under CLIA as tests of moderate complexity (24). Accordingly, personnel standards, quality control, quality assurance, and proficiency testing requirements apply when performing FDA-cleared C. trachomatis tests that are rapid enough to qualify as point-of-care tests. In addition, these tests are usually less sensitive and more expensive than laboratory-based C. trachomatis tests that require longer to perform. Similar to EIAs, these tests use antibodies against LPS that detect all three Chlamydia species that infect humans and are subject to the same potential for false-positive results caused by cross-reactions with other microorganisms. A point-of-care test should not be selected if it is performed in a laboratory after the patient's visit. Such use of point-of-care tests is inappropriate because sensitivity and specificity are typically less, controls less rigorous, and costs higher than for tests designed for laboratory use (67).

Gram Stain for N. gonorrhoeae

Gram stain is a key tool for the diagnosis of gonococcal urethritis in men, but its application to screening is limited because of the requirement for an intraurethral swab specimen if discharge is not present at the urethral meatus. A Gram stain for the presumptive diagnosis of N. gonorrhoeae infection is performed on thin smears of urethral exudate from men and is presumptively positive if the smear contains typical Gram-negative diplococci within polymorphonuclear (PMN) leukocytes. Unfortunately, other Neisseria species have a similar appearance. Although commensal Neisseria species are not normal flora of anogenital sites, isolates of Neisseria meningitidis and nonpathogenic Neisseria species have been reported occasionally from anogenital sites among both men and women (12).

As a point-of-care test, Gram stain is most reliable for the presumptive identification of N. gonorrhoeae in urethral exudates from men. The sensitivity and specificity of a Gram stain for males with symptomatic urethritis are comparable to culture isolation followed by oxidase testing and Gram staining of the isolate (32 ,68 ,69). Gram-negative extracellular diplococci without any intracellular diplococci might be observed in smears from men with early symptomatic infections. The sensitivity of Gram stain for males with asymptomatic urethral infection has not been determined. Gram-negative intracellular diplococci might be observed with certain infections caused by nongonococcal Neisseria species (e.g., N. cinerea).

The sensitivity of Gram stains of endocervical specimens is lower than for urethral specimens from men with symptomatic gonorrhea (10 ,32 ,69), and adequate specificity requires a skilled microscopist. For these reasons, Gram stain is not recommended for testing for N. gonorrhoeae infection among women. Gram stains of pharyngeal specimens are not recommended because N. meningitidis and commensal Neisseria species colonize the pharynx (12). As with point-of-care tests for C. trachomatis, the Gram-stained smear is classified under CLIA as a moderate complexity test for urethral and endocervical specimens (24). Gram stains from all other body sites are categorized as high complexity. A trained laboratorian is required for preparing and reading high-complexity Gram-stained smears.

* These guidelines do not address trachoma and lymphogranuloma venereum, which rarely occur in the United States.

The term cleared is used by the Food and Drug Administration (FDA) to describe the process they use to review applications to market the class of diagnostic tests that includes C. trachomatis and N. gonorrhoeae tests discussed in these guidelines. The term approved is used by FDA to describe a more rigorous process they use to review applications to market classes of diagnostic tests that involve, for example, higher levels of risk if the test result is erroneous than is the case for C. trachomatis or N. gonorrhoeae.