Many adjunctive tests are available to help evaluate male-factor infertility if the initial evaluation fails to lead to a diagnosis. One guiding principle in this era of cost-containment is to order tests only if they will change patient management.

Semen Leukocyte Analysis

White blood cells (leukocytes) are present in all ejaculates and may play important roles in immune surveillance and clearance of abnormal sperm. Leukocytospermia or pyospermia, an increase in leukocytes in the ejaculate, is defined as > 1 × 10⁶ leukocytes/mL semen and is a significant cause of male subfertility. The prevalence of pyospermia ranges from 2.8% to 23% of infertile men. In general, neutrophils predominate among inflammatory cells (

Table 42-9). This condition is detected by a variety of diagnostic assays, including differential stains (eg, Papanicolaou), peroxidase stain that detects the peroxidase enzyme in neutrophils, and immunocytology.

The reason for the immune cell infiltrate in pyospermia is poorly understood, although some proposed causes include an inflammatory response associated with infection, sensitization of the immune system to sperm antigens, or a reaction to low-grade toxins like cigarette smoke or alcohol.

Antisperm Antibody Test

The testis is a curious organ in that it is an immunologically privileged site, probably owing to the blood-testis barrier. Autoimmune infertility may occur when the blood-testis barrier is broken and the body is exposed to sperm antigens. Trauma to the testis and vasectomy are 2 common ways in which this occurs, giving rise to antisperm antibodies (ASA). ASA may be associated with impaired sperm transport through the reproductive tract or impairment in egg fertilization. An assay for ASA should be obtained when:

1. The semen analysis shows sperm agglutination or clumping.
2. Low sperm motility exists with history of testis injury or surgery.
3. There is confirmation that increased round cells are leukocytes.
4. There is unexplained infertility.

Antisperm antibodies can be found in 3 locations: serum, seminal plasma, and sperm-bound. Among these, sperm-bound antibodies are the most relevant. The antibody classes that appear to be clinically relevant include IgG and IgA. IgG antibody is derived from local production and from transudation from the bloodstream (1%). IgA is thought to be purely locally derived.

Hypoosmotic Swelling Test

The most clinically useful measure of sperm viability is cell motility. However, a lack of motility does not necessarily signify absent viability. Indeed, there are clinical conditions, such as immotile-cilia syndrome and extracted testicular sperm, in which there may be immotile but otherwise presumably healthy sperm. Such sperm can now be used clinically for micromanipulation and in vitro fertilization (IVF). Cell viability can be evaluated noninvasively by using the physiologic principle of hypoosmotic swelling. Conceptually, viable cells with functional membranes should swell when placed in a hypoosmotic environment (25 mM citrate and 75 mM fructose). Since sperm have tails, the swelling response is very obvious in that tail coiling accompanies head swelling. This sperm test is indicated in cases of complete absence of sperm motility.

Sperm Penetration Assay (SPA)

It is possible to measure the ability of human sperm to penetrate a specially prepared hamster egg in a laboratory setting. The hamster egg allows interspecies fertilization but no further development. This form of bioassay can give important information about the ability of sperm to undergo the capacitation process as well as penetrate and fertilize the egg. Infertile sperm would be expected to penetrate and fertilize a lower fraction of eggs than normal sperm. The indications for the diagnostic SPA are limited to situations in which functional information about sperm are needed, that is, to further evaluate couples with unexplained infertility and to help couples decide whether intrauterine insemination (IUI) (good SPA result) or IVF and micromanipulation (poor SPA result) is the appropriate next treatment.

Sperm-Cervical Mucus Interaction

Infertility can be caused by impaired sperm transport in the female reproductive tract. An evaluation of interaction between sperm and cervical mucus is one way to assess the quality of the transport process. One way to examine sperm-mucus interaction is by measuring the rate of sperm movement through a pool of cervical mucus on a microscope slide or within a capillary tube. This assay involves controls, in which sperm are placed in seminal fluid instead of cervical mucus. An abnormal cervical mucus-sperm interaction may suggest infertility treatment in which sperm are placed beyond the cervix into the uterus (IUI).

Chromosomal Studies

Subtle genetic abnormalities can present as male infertility. It is estimated that between 2% and 15% of infertile men with azoospermia (no sperm count) or severe oligospermia (low sperm counts) will harbor a chromosomal abnormality on either the sex chromosomes or autosomes. A blood test for cytogenetic analysis (karyotype) can determine if such a genetic anomaly is present. Patients at risk for abnormal cytogenetic findings include men with small, atrophic testes, elevated FSH values, and azoospermia. Klinefelter syndrome (XXY) is the most frequently detected sex chromosomal abnormality among infertile men (

Figure 42-7).

Cystic Fibrosis Mutation Testing

A blood test is indicated for infertile men who present with cystic fibrosis (CF) or the much more subtle condition, CAVD. Similar genetic mutations are found in both patients, although the former group is generally considered to have an atypical form of CF, in which the scrotal vas deferens is nonpalpable. Approximately 80% of men without palpable vasa will harbor a CF gene mutation. Recent data also indicate that azoospermic men with idiopathic obstruction and men with a clinical triad of chronic sinusitis, bronchiectasis, and obstructive azoospermia (Young syndrome) may be at higher risk for CF gene mutations.

Y Chromosome Microdeletion Analysis

As many as 7% of men with oligospermia and 15% of azoospermic men have small, underlying deletions in one or more gene regions on the long arm of the Y chromosome (Yq). Several regions of the Y chromosome have been implicated in spermatogenic failure, identified as AZFa, b, and c (

Figure 42-8). Deletion of the DAZ (deleted in azoospermia) gene in the AZFc region is the most commonly observed microdeletion in infertile men. Fertility is possible in men with these deletions with IVF and micromanipulation of sperm. A polymerase chain reaction-based blood test can examine the Y chromosome from peripheral leukocytes for these gene deletions and is recommended for men with low or no sperm counts and small, atrophic testes.

Radiologic Testing

A. Scrotal Ultrasound
High-frequency (7.5-10 mHz) ultrasound of the scrotum has become a mainstay in the evaluation of testicular and scrotal lesions. In men who have a hydrocele within the tunica vaginalis space, the testis may be nonpalpable and should undergo ultrasound to confirm that it is normal. Any abnormality of the peritesticular region should also undergo a scrotal ultrasound to determine its characteristics or origin.

Recently, scrotal color Doppler ultrasonography has been used to investigate varicoceles (

Figure 42-9). By combining measurements of blood-flow patterns and vein size, both physiologic and anatomic information can be obtained for more accurate assessment. The diagnostic criteria that define a varicocele vary from study to study, but in general, a pampiniform venous diameter of > 2-3 mm is considered abnormal. Retrograde blood flow through the veins with a Valsalva maneuver is also an important radiologic feature of a varicocele.

B. Venography
Venography is generally accepted as the most accurate way to diagnose varicoceles. Although found by palpation in approximately 30-40% of subfertile men, varicoceles can be detected by venography in 70% of patients. Renal and spermatic venography is fairly invasive and is usually performed through percutaneous cannulization of the internal jugular vein or common femoral vein. Venographically, a varicocele is defined by a Valsalva-induced, retrograde flow of contrast material from the renal vein into the scrotal pampiniform plexus. This test is expensive and technician-dependent; at present its main indications are to guide simultaneous percutaneous treatment of varicoceles or for the diagnosis of varicocele recurrence after prior treatment.

C. Transrectal Ultrasound (TRUS)
High-frequency (5-7 mHz) TRUS offers superb imaging of the prostate, seminal vesicles, and ejaculatory ducts. Transrectal ultrasound has virtually replaced surgical vasography in the diagnosis of obstructive lesions that cause infertility. Demonstration by TRUS of dilated seminal vesicles (> 1.5 cm in width) or dilated ejaculatory ducts (> 2.3 mm) in association with a cyst, calcification, or stones along the duct is highly suggestive of ejaculatory duct obstruction (

Figure 42-10). In addition, prostatic abnormalities such as tumors and congenital anomalies of the vas, seminal vesicle, or ejaculatory ducts are easily defined. The indications for TRUS in infertility include low ejaculate volumes in association with azoospermia or severe oligospermia and decreased motility.

D. Computed Tomography Scan or Magnetic Resonance Imaging of the Pelvis
The imaging techniques of CT and MRI can help define reproductive tract anatomy. However, since the advent of TRUS, these studies have relatively few indications. They include evaluation of a patient with a solitary right varicocele, a condition often associated with retroperitoneal pathology, and evaluation of the nonpalpable testis.