In this article we will discuss about the laboratory diagnosis of various sexually transmitted diseases (STDs). The various sexually transmitted diseases whose diagnosis has been discussed are: 1. Urethritis 2. Genital Ulcers 3. Vaginal Discharge.
Sexually Transmitted Disease # 1. Urethritis:
Causative agents of urethritis are listed in Table 16.1:
Urethral, vaginal, cervical, and anal exudates are examined and cultured.
Gram-stained smear of discharge reveal abundant pus cells (polymorphonuclear leucocytes). About one in twenty of the pus cells, contains plenty of bean-shaped Gram-negative intracytoplasmic diplococci.
(i) Male patients:
A positive smear is virtually diagnostic of gonococci.
It is sometimes difficult to interpret the smear finding in mixed normal flora in females. Veillonella parvula is also a Gram-negative diplo-coccus normally present in vaginal flora.
Moreover, asymptomatic carriage is high in females, especially with endocervical lesion. Hence positive identification of the organism by culture or immuno-fluorescent study using genetic probes (e.g. DNA probe) is necessary to confirm the diagnosis.
A direct wet film of freshly collected specimen shows motile trichomonad with polymorphonuclear leucocytes.
Microscopical examination of vaginal secretions in 10% potassium hydroxide show yeast cells.
4. Non-gonococcal urethritis:
Smear stained by Giemsa stain shows intracytoplasmic inclusion bodies, suggestive of C. trachomatis. When diagnosis of C. trachomatis cannot be made by light microscopy, a definitive diagnosis is to be made by either culture or by antigen detection.
Specimen is inoculated into McCoy or HeLa cell tissue cultures. Tissue culture is not routinely available.
(iii) Detection of antigen:
Smear made from exudate is examined by immunofluorescence test with a monoclonal antibody or by ELISA to detect LPS antigens (either soluble or in the elementary body cell wall) of C. trachomatis.
Sexually Transmitted Disease # 2. Genital Ulcers:
1. Hard chancre (Syphilis) — by T. pallidum
2. Soft chancre (Chancroid) – by H. ducreyi
3. Herpes progenitalis – by Herpes simplex virus types 1 and 2, primarily type 2.
4. Donovanosis (granuloma inguinale) – by Calymmatobacterium granulomatis.
5. Lymphogranuloma venereum – by C. trachomatis types L1, L2, L3.
Laboratory diagnosis — See Table 16.2.
(i) Dark-field microscopy of specimen collected from a syphilitic primary chancre as well as secondary rash may reveal motile Treponema pallidum.
(ii) Gram stain of exudate from cases of chancroid may show Gram-negative ovoid bacilli. The bacteria en mass have configuration of “shoals of fish”.
(iii) Giemsa/Wright staining of exudate of genital herpes often show multinucleated giant cells or intra-nuclear inclusions.
(iv) Castaneda’s method:
Chlamydiae though Gram-negative, but they stain well by Castaneda’s method and Giemsa stain. The inclusions in cell culture are stained by both the stains well.
In donovanosis, Donovan bodies (round cocco- bacilli, 1 x 2 μm) may be found in smear from ulcer stained by Giemsa stain. The Donovan bodies lie within the cystic spaces in the cytoplasm of large mononuclear cells. Capsule appears as a dense acidophilic zone around the bacterium, resembling a closed safety pin or “telephone handle”.
2. Serological test:
Syphilis is diagnosed in most patients on the basis of serological tests. Positive result is obtained from about two weeks after the appearance of primary sore.
Two types of tests: nonspecific (non-treponemal) and specific (treponemal) tests are used (Table 16.3):
(a) Non-specific tests:
Non-treponemal tests measure IgG and IgM antibodies (known as reagin antibodies) produced against lipids released from damaged cells during the early stage of the disease which are also present on cell surface of treponemes.
(b) Specific or treponemal tests:
These are used to confirm positive VDRL or RPR tests. The most commonly used treponemal tests are FTA-ABS test and MHA-TP test. These tests can become positive before the non-treponemal tests in early syphilis, or remain positive when the non-treponemal tests revert to negative in some patients with late syphilis.
Western blot test using whole cell T. pallidum as antigen has recently been used successfully to confirm non-treponemal tests. VDRL and TPHA are more useful tests for diagnosis of syphilis in most laboratories. When these tests are done together, they provide an effective screen for early and late syphilis.
(ii) Herpes genitalis:
Serological tests (e.g. CFT) with patient’s HSV infection. The antibody titre is usually within 32, especially in genital herpes.
1. Syphilis is best treated with large doses of penicillin. Long-acting benzathine penicillin is recommended in early syphilis, and penicillin G for congenital and late syphilis. Tetracycline and doxycycline’s are alternative drugs.
2. Herpes simplex can be treated with acyclovir.
3. Chancroid is treated with sulfonamides or aminoglycosides.
Sexually Transmitted Disease # 3. Vaginal Discharge:
Causes include gonorrhoea, infection due to C. trachomatis, bacterial vaginosis, trichomonas vaginalis and Vulvovaginal candidiasis.
The vaginal discharge may be due to more than one of the above mentioned microorganisms.
1. pH — greater than 4.5.
2. Amine test:
When vaginal fluid is mixed with a 10% solution of KOH, fishy odor is immediately liberated. The odor is due to volatile amines present in vaginal fluid.
3. Saline microscopy:
Vaginal discharge contains clue cells (squamous epithelial cells coated with coccobacillary organisms). Leucocytes are virtually absent.
The normal vaginal flora of lacto- bacilli in vagina is largely replaced by G. vaginalis and other anaerobes.