**4. Evaluation**

In patients whose history and physical examination suggest urethritis, Gram staining of urethral discharge is the first-line laboratory test, and detection of >5 white blood cells (WBC)/per oil immersion field allows a rapid diagnosis of urethritis. In some publications, >2 WBC/high power field (HPF) was used as the threshold value based on the argument that this would provide a more sensitive diagnosis. However, this has not been supported by other studies. The cut-off value accepted by the European Association of Urology (EAU) is >5 polymorphonuclear lymphocytes (PMNL)/HPF. This method has high specificity and sensitivity both for the diagnosis of urethritis and determining the presence or absence of gonococcal infection [24].

A positive leukocyte esterase test of first-void urine or >10 WBC/HPF in the sediment of first-void urine is also diagnostic criterion for urethritis [25].

*Chlamydia* is not detectable by Gram staining because it is a small obligate cellborne parasitic bacteria. In a patient with pyuria and suspected urethritis based on history and physical examination, detecting no bacteria on Gram staining raises a strong suspicion of nongonococcal urethritis pathogens, most of which are *Chlamydia* [26].

All male patients with suspected urethritis should undergo NAATs, which are the gold standard for the diagnosis of *N. gonorrhoeae* and *C. trachomatis*, the most common urethritis pathogens. *N. gonorrhoeae* should be included in the NAAT panel even if it was not detected in Gram staining [25]. Even if gonococci were detected in Gram staining, there may be a concurrent chlamydial infection, so evaluation with NAATs should still be done [7].

Methods used in the diagnosis of *C. trachomatis* infection include cytological examination, cell culture, antigen quantification, direct fluorescent antibody tests, enzyme

#### *Chlamydia Infection from Androgical Perspective DOI: http://dx.doi.org/10.5772/intechopen.110045*

immunoassays, and NAATs if nucleic acid from the pathogen is detected. Among these, NAATs have the highest sensitivity [27]. This method has found widespread use worldwide [16]. Whereas, only urethral swab samples can be used for cultures and hybridization tests, NAATs can also be done using a first-void urine sample, with similar efficacy. NAATs work by amplifying and detecting chlamydial DNA from a very small number of organisms in clinical samples using specific primers and enzymes [10].

*C. trachomatis* culture is mostly used in treatment failure and to assess resistance to administered treatment [2]. As obligate intracellular parasites, *Chlamydia* cannot be grown in culture media. The living cell environment is necessary for their reproduction [28]. Therefore, as cell culturing requires an experienced team and a well-equipped laboratory, is difficult, and takes time, the use of this technique in the diagnosis of *C. trachomatis* has been replaced in recent years by nucleic acid screening tests, which are molecular techniques that provide faster results and have high specificity and sensitivity. NAAT is the gold standard diagnostic method for urogenital chlamydial infection and can be performed using urethral swab samples collected with a Dacron- or rayon-tipped plastic swab or cytobrush, or using firstvoid urine. Other swabs containing cotton may inhibit *C. trachomatis* [29]. Sampling is done by inserting a dry swab 3−4 cm into the anterior urethra and rotating it within the urethra before withdrawing. However, the patient should not have urinated in the last 1−2 h [20]. Likewise, for NAATs of first-void urine, the patient should not have urinated within the last 20−60 min. A sample of 10−20 mL is collected at the start of urination without cleaning the urethra. Some publications indicate that urethral swabs are less sensitive than urine in men but have the same specificity [10].

In men who have sex with men, samples for *Chlamydia* and *N. gonorrhoeae* testing should be obtained from the sites of possible sexual contact [2]. Although a normal urine sample is negative in these patients, it should not be forgotten that 70% of extragenital (oral and/or anal) sites may yield positive NAAT results [3].

If the urethral smear is normal and symptoms are inconclusive, repeating the smear in the morning with first-void urine is recommended. The patient should be advised to avoid excessive fluid intake the day before to ensure that urination is not urgent in the morning and they can give a first-void urine sample in the laboratory. If a symptomatic man has a negative smear, a positive leukocyte esterase test of firstvoid urine aids in the diagnosis of urethritis [2].

The EAU guideline also strongly recommends NAATs for chlamydia and gonorrhea before empirical treatment, if possible. However, treatment should be initiated immediately upon diagnosing urethritis in men with severe symptoms, without waiting for the results of chlamydia, gonorrhea, and *M. genitalium* tests. Patients with mild symptoms and microscopically low leukocyte counts (5–15 PMNL/HPF) are reevaluated after 3−7 days. A urethral smear is obtained early in the morning. NAAT and gonorrhea culture results are also examined when available. Urethritis can sometimes resolve spontaneously without treatment. If laboratory tests are positive and the urethritis persists according to microscopic findings, appropriate antibiotic treatment targeting the microorganism isolated at this second visit should be initiated, bearing local resistance patterns in mind [2]. If symptoms do not resolve in 3−4 weeks, urethritis is classified as persistent. In this case, evaluation with NAATs (including for *T. vaginalis*) should be repeated 4 weeks after the end of treatment [25]. Because men with chlamydial, gonorrheal, or trichomonal infections are at high risk of reinfection, they should be reevaluated by repeating the tests 3 months later. Although it is not an FDA-approved test for *Trichomonas* and *Mycoplasma*, NAAT is performed in many reference and commercial laboratories [7].

The immune response can affect the development of nongonococcal urethritis. A high microbial load (>1000/copies/mL in first-void urine) is a strong predictor of nongonococcal urethritis [2].
