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Sexually

Transmitted Diseases
Common Diseases
Dr.T.V.Rao MD

Dr.T.V.Rao MD

What are Sexually Transmitted Diseases


STDs are infections that are spread from person to person through intimate sexual contact. STDs are dangerous because they are easily spread and it is hard to tell just by looking who has an STD. 1 in 4 teenagers has an STD.(Western Statistics)
Dr.T.V.Rao MD 2

STDs
STDs are diseases and infections which are capable of being spread from person to person through:
sexual intercourse oral-genital contact or in non-sexual ways. IV drug Congenitally transmitted
Dr.T.V.Rao MD 3

Common STIs
Chlamydia Gonorrhea Genital Herpes (HSV-2) Genital Warts (HPV) Hepatitis B

HIV and AIDS Pubic Lice Syphilis Trichomoniasis

Dr.T.V.Rao MD

STDs of Concern
Actually, all of them Sores (ulcers)
Syphilis Genital herpes (HSV-2, HSV-1)

Others uncommon in the U.S.


Lymphogranuloma venereum Chancroid Granuloma inguinale
Dr.T.V.Rao MD 5

Do Patients Want to Know? About STDs


92.4% wanted to know if they were infected 90.8% wanted to know if their partners were infected 65% expected the test as part of STD screening
Dr.T.V.Rao MD

STDs of Concern (continued)


Drips (discharges) Gonorrhea Chlamydia Nongonococcal urethritis / mucopurulent cervicitis Trichomonas vaginitis / urethritis Candidiasis (vulvovaginal, less problems in men)

Other major concerns


Genital HPV (especially type 16, 18) and Cervical Cancer
Dr.T.V.Rao MD 7

Genital Ulcer Diseases Does It Hurt?


Painful
Chancroid Genital herpes simplex

Sores

Painless
Syphilis Lymphogranuloma venereum Granuloma inguinale
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Treponema pallidum The Agent of Syphilis


Spirochete Obligate human parasite

Transmission Sexual Trans-placental Percutaneous following contact with infectious lesions Blood Transfusion No reported cases of transmission since 1964
Dr.T.V.Rao 5 MD 9

Syphilis The Great Imitator


Infectious Dose: ~57 organisms1

Incubation Period 21 days (median) 3 clinical stages of syphilis

Primary:
Painless sore (chancre) at inoculation site Secondary: Rash, Fever, Lymphadenopathy, Malaise Tertiary/Latent: CNS invasion, organ damage The physician that knows syphilis knows medicine. Sir William Osler
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Primary Syphilis - Clinical Manifestations


Incubation: 10-90 days (average 3 weeks) Chancre
Early: macule/papule erodes Late: clean based, painless, indurated ulcer with smooth firm borders Unnoticed in 15-30% of patients Resolves in 1-5 weeks HIGHLY INFECTIOUS
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Primary Syphilis

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Primary Syphilis - Chancre

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Primary Syphilis - Chancre

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Primary Syphilis Chancre

Source: Florida STD/HIV Prevention Training Center

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Primary Syphilis Chancre


Represents haematogenous dissemination of spirochetes Usually 2-8 weeks after chancre appears Findings:
rash - whole body (includes palms/soles) mucous patches Condylomata lata - HIGHLY INFECTIOUS constitutional symptoms

Sn/Sx resolve in 2-10 weeks


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S o r e s

Sores

Secondary Syphilis Rash

Source: Florida STD/HIV Prevention Training Center

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Secondary Syphilis: Generalized Body Rash

Sores

Source: CDC/NCHSTP/Division of STD Prevention, STD Clinical Slides

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Secondary Syphilis Rash

Source: Florida STD/HIV Prevention Training Center

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Secondary Syphilis Rash

Source: Cincinnati STD/HIV Prevention Training Center

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Secondary Syphilis

Source: Diepgen TL, Yihune G et al. Dermatology Online Atlas

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Secondary Syphilis Condylomata Lata

Sores

Source: Florida STD/HIV Prevention Training Center

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Congenital Syphilis
Congenital syphilis usually occurs following vertical transmission of T. pallidum from the infected mother to the fetus in utero, but neonates may also be infected during passage through the infected birth canal at delivery.
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DIAGNOSIS OF SYPHILIS
1. History and clinical examination. 2. Dark-field microscopy: special technique use to demonstrate the spirochete as shiny motile spiral structures with a dark background. The specimen includes oozing from the lesion or sometimes L.N. aspirate. It is usually positive in the primary and secondary stages and it is most useful in the primary stage when the serological tests are still negative.
Dr.T.V.Rao MD 24

Laboratory Testing
Direct examination of clinical specimen by dark-field microscopy or fluorescent antibody testing of sample. Non-specific or non-treponemal serological test to detect reagin, utilized as screening test only. Specific Treponemal antibody tests are used as a confirmatory test for a positive reagin test.
Dr.T.V.Rao MD 25

Diagnosis of Syphilis
Evaluation based on three factors: Clinical findings.
Demonstration of spirochetes in clinical specimen. Present of antibodies in blood or cerebrospinal fluid.

More than one test should be performed. No serological test can distinguish between other Treponemal infections.
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Provisional Diagnosis of Syphilis


A presumptive diagnosis is possible with sequential serologic tests (e.g. VDRL, RPR), using the same testing method each time. A fourfold change in titer (e.g. from 1:8 to 1:32) is usually considered to be clinically significant. Confirmatory tests should be performed
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Other Diagnostic Tests


Serological tests of syphilis. Biopsy rarely needed. It shows endarteritis obliterans with inflammatory reaction containing a plenty of plasma cells. Granuloma may found in tertiary syphilis.
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Laboratory Diagnosis of Syphilis


The Uncommon Methods
Rabbit Infectivity Test (RIT)
High Sensitivity and Specificity Long turn-around-time Limited to research settings
http://www.els.net

Dark Field Microscopy


Useful only during primary infection Technician expertise required

Immunostaining
Direct fluorescent antibody or silver stain

Polymerase Chain Reaction (PCR)


Not commercial available
textbookofbacteriology.net

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Non-specific or lipoidal tests A. VDRL (venereal disease research laboratories) - It is useful for the screening, diagnosis and follow up. - The results can be qualitative or qualitative

SERLOGICAL TESTS OF SYPHILIS: STS

False positive results

1. Acute type: usually low titre and dont persist for more than 6 months 2. Chronic type: usually last for more than 6 months B. Rapid plasma reagin test. C. Wasserman test not used more
Dr.T.V.Rao MD 30

Laboratory Diagnosis of Syphilis

Serology

The Common Methods

Mainstay for syphilis testing Two classes of serologic tests


Non-treponemal Treponemal
Dr.T.V.Rao 8 MD 31

A. Reiter protein complement fixation test. B. Fluorescent Treponemal antibody/absorption test, FTA/ABS. the most specific and most sensitive . C. Treponema pallidum

Specific serological tests of Syphilis

haemagglutination test- TPHA- D.


Treponema pallidum immobilization testTPI
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Serologic Tests for Syphilis:


Non-Treponemal Assays
Principle:
T. pallidum infection leads to the production of reagin Reagin Antibodies to substances released from cells damaged by T. pallidum Reagin reacts with cardiolipin Cardiolipin a phospholipid component of certain eukaryotic and prokaryotic membranes

Examples of non-treponemal tests:


Rapid Plasma Reagin (RPR) Venereal Disease Research Laboratory (VDRL)
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Serologic Tests for Syphilis:


Non-Treponemal Assays
RPR and VDRL are agglutination assays
Charcoal Cardiolipin

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Serologic Tests for Syphilis:


Non-Treponemal Assays
RPR and VDRL are agglutination assays

Charcoal Cardiolipin Reagin Serum or CSF

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VDRL

Each preparation of antigen suspension should first be examined by testing with known positive or negative serum controls. The antigen particles appear as short rod forms at magnification of about 100x. Aggregation of these particles into large or small clumps is interpreted as degrees of positivity Reactive on left, non-reactive on right
Dr.T.V.Rao MD 36

Rapid Plasma Reagin Test - RPR


General screening test, can be adapted to automation. CANNOT be performed on CSF. Antigen
VDRL cardiolipin antigen is modified with choline chloride to make it more stable attached to charcoal particles to allow macroscopic reading antigen comes prepared and is very stable.

Serum or plasma may be used for testing, serum is not heated.


Dr.T.V.Rao MD 37

RPR
Test Procedure:
Serum or plasma added to circle on card and spread. One drop of antigen from a needle capable of delivering 60 drops/mL is added. Rotate at 100 rpms/minute for 8 minutes. Results are read macroscopically.

Daily quality control:


20 gauge needle checked for delivery of 60 drops/mL Rotator checked for 100 rpms/minute Room temperature must be 23-29 C. Three levels of control must be run and give appropriate results.

RPR appears to be more sensitive than the VDRL.


Dr.T.V.Rao MD 38

Treponema pallidum haemagglutination (TPHA)


Adapted to micro techniques (MHA-TP) Tanned sheep RBCs are coated with T. pallidum antigen from Nichols strain. Agglutination of the RBCs is a positive result.
Dr.T.V.Rao MD 39

Non-Treponemal Tests:
Advantages

Rapid turnaround time Minutes Inexpensive No specialized instrumentation required Usually revert to negative following therapy Can be used to monitor response to therapy
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Non-Treponemal Tests:
Limitations
Results are subjective Intra- and Inter-laboratory variability Non-specific False positive results can result from other infectious or non-infectious conditions EBV, Lupus, etc. Limited sensitivity in early/primary syphilis and in late/latent syphilis Low throughput Problematic for high volume laboratories
Dr.T.V.Rao 13 MD 41

Non-Treponemal Tests:
Limitations, continued
Possibility for prozone effect High levels of antibody may inhibit the agglutination reaction To identify prozone, labs must serially dilute samples

Undilute

1:2

1:4

1:8

1:16

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Serologic Tests for Syphilis: Treponemal Assays

Principle: Infection leads to production of specific antibodies directed against T. pallidum Treponemal tests detect IgG or total IgM/IgG antibodies directed against T. pallidum
Dr.T.V.Rao 15 MD 43

Serologic Tests for Syphilis:


Treponemal Assays
Microhemagglutination assay (MHA) Fluorescent treponemal antibody (FTA-ABS) Treponema pallidum particle agglutination (TP-PA) Enzyme Immunoassay (EIA) Multiplex Flow Immunoassay (MFI)
FTA-ABS TP-PA Conventional EIA

www.mastgrp.biz Yellow wells = positive

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Treponemal Assays:
Advantages
High Specificity Possibly higher sensitivity during early and late syphilis stages compared to non-treponemal tests Newer Methods Objective result interpretation Automation option High throughput High reproducibility/precision
Dr.T.V.Rao 20 MD 45

Fluorescent Treponemal Antibody Absorption Test (FTA-ABS) Diluted, heat inactivated serum added to Reiters strain of T. pallidum to remove cross reactivity due to other Treponemes. Slides are coated with Nichols strain of T. pallidum and add absorbed patient serum.
Dr.T.V.Rao MD 46

Fluorescent Treponemal Antibody Absorption Test (FTA-ABS)


Slides are washed, and incubated with antibody bound to a fluorescent tag. After washing the slides are examined for fluorescence. Requires experienced personnel to read. Highly sensitive and specific, but time consuming to perform.
Dr.T.V.Rao MD 47

Treponemal Assays:
Limitations
Remain positive despite treatment Cannot be used to monitor response to therapy Conventional Methods Subjective interpretation requiring technician expertise to read Newer Methods Expensive instrumentation Higher cost/test
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Traditional Algorithm
Non-treponemal test (e.g., RPR) Reactive Treponemal test (e.g., FTA) Non-reactive Negative for syphilis

Reactive Syphilis

Non-reactive

Negative for syphilis

Advantages: Results show good correlation with disease status Rapid, inexpensive screening method Excellent option for laboratory with small throughput Recommended by the CDC

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Traditional Algorithm
Non-treponemal test (e.g., RPR) Reactive Treponemal test (e.g., FTA) Non-reactive Negative for syphilis

Reactive Syphilis

Non-reactive

Negative for syphilis

Disadvantages: Manual (RPR) and subjective interpretation Screening method is non-specific and may lead to false-positive results Not suitable for high throughput laboratories Potentially lower sensitivity for detecting early syphilis and late/latent disease
Dr.T.V.Rao 25 MD 50

Reverse Algorithm
Treponemal test (eg, EIA) Reactive Non-Treponemal test (eg, RPR) Non-reactive Negative for syphilis

Reactive Syphilis

Non-reactive Second Treponemal Test (e.g., TP-PA) Reactive Non-reactive

Evaluation Required*

Negative for syphilis

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Every Pregnant women Needs Screening

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Genital Herpes Simplex

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Genital Herpes Simplex - Clinical Manifestations


Direct contact may be with asymptomatic shedding Primary infection commonly asymptomatic; symptomatic cases sometimes severe, prolonged, systemic manifestations Vesicles painful ulcerations crusting Recurrence a potential Diagnosis:
Culture Serology (Western blot) PCR
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Epidemiology of Genital Herpes


One of the 3 most common STDs, increased 30% from late 70s to early 90s 25% of US population by age 35 HSV-2: 80-90%, HSV-1: 10-20% (majority of infections in some regions) Most cases subclinical Transmission primarily from subclinical infection Complications: neonatal transmission, 55 enhanced HIV transmission, psychosocial issues
Dr.T.V.Rao MD

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Underdiagnoses of Genital Herpes


779 women attending STD clinic 372 genital herpes diagnosis:
363 HSV-2 antibody positive 9 HSV-1 culture positive lesions

Of the 372 diagnosed with genital herpes


82 (22%) symptomatic 14 (4%) viral shedding without symptoms 60 (14%) history of symptoms 216 (58%) HSV-2 antibody without viral shedding or history of symptoms
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Genital Herpes Simplex

Source: Diepgen TL, Yihune G et al. Dermatology Online Atlas

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Genital Herpes Simplex

Source: CDC/NCHSTP/Division of STD, STD Clinical Slides

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Drips
Gonorrhea Nongonococcal urethritis Chlamydia Mucopurulent cervicitis Trichomonas vaginitis and urethritis Candidiasis
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Gonorrhea Clinical Manifestations


Urethritis - male
Incubation: 1-14 d (usually 2-5 d) Sx: Dysuria and urethral discharge (5% asymptomatic) Dx: Gram stain urethral smear (+) > 98% culture Complications

Urogenital infection - female


Endocervical canal primary site 70-90% also colonize urethra Incubation: unclear; sx usually in l0 d Sx: majority asymptomatic; may have vaginal discharge, dysuria, urination, labial pain/swelling, abd. pain Dx: Gram stain smear (+) 50-70% culture Complications
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Gonorrhea

Source: Florida STD/HIV Prevention Training Center

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Gonorrhea Gram Stain

Source: Cincinnati STD/HIV Prevention Training Center

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Diagnosis not Easy


Three levels of diagnosis are defined on the basis of clinical findings or the results of laboratory diagnostic tests. A definitive diagnosis of gonorrhoea must be obtained for medico legal purposes.
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Diagnosis of Gonorrhoea
Suggestive diagnosis is defined by the presence of: A mucopurulent endocervical or urethral exudate on physical examination and sexual exposure to a person infected with N. gonorrhoea.
Dr.T.V.Rao MD 64

Presumptive diagnosis of gonorrhoea is made on the basis of one of the following three criteria:
Typical gram-negative intracellular diplococci on microscopic examination of a smear of urethral exudate from men or endocervical secretions from women*; Growth of a gram-negative, oxidase-positive diplococcus, from the urethra (men) or endocervix (women), on a selective culture medium, and demonstration of typical colonial morphology, positive oxidase reaction, and typical gram- negative morphology;
Dr.T.V.Rao MD 65

Presumptive Diagnosis of Gonorrhoea


The observation of gram-negative, intracellular diplococci on microscopic examination of endocervical secretions from women must be supported by a positive result from a test from either 2 or 3 to make a laboratory diagnosis of presumptive N. gonorrhoea.
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Definitive diagnosis of gonorrhoea requires:


Isolation of N. gonorrhoea from sites of exposure (e.g., urethra, endocervix, throat, rectum) by culture (usually a selective medium) and demonstrating typical colonial morphology, positive oxidase reaction, and typical gramnegative morphology

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Definitive diagnosis of gonorrhoea requires:


Confirmation of isolates by biochemical, enzymatic, serologic, or nucleic acid testing e.g., carbohydrate utilization, rapid enzyme substrate tests, serologic methods such as coagglutination or fluorescent antibody tests supplemented with additional tests that will ensure accurate identification of isolates, or a DNA probe culture confirmation technique.
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Newer Methods in Diagnosis of

Gonorrhoea
Detection of N. gonorrhoea by a nonculture laboratory test (Antigen detection test (e.g., Gonozyme [Abbott]), direct specimen nucleic acid probe test (e.g., Pace II [GenProbe]), nucleic acid amplification test (e.g., LCR [Abbott]).
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Definitive diagnosis of gonorrhoea requires:


Isolation of N. gonorrhoea from sites of exposure (e.g., urethra, endocervix, throat, rectum) by culture (usually a selective medium) and demonstrating typical colonial morphology, positive oxidase reaction, and typical gram-negative morphology and

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Definitive diagnosis of gonorrhoea requires:


Confirmation of isolates by biochemical, enzymatic, serologic, or nucleic acid testing e.g., carbohydrate utilization, rapid enzyme substrate tests, serologic methods such as coagglutination or fluorescent antibody tests supplemented with additional tests that will ensure accurate identification of isolates, or a DNA probe culture confirmation technique.
Dr.T.V.Rao MD 71

Drips

Nongonococcal Urethritis
Etiology:
20-40% C. trachomatis 20-30% genital mycoplasmas (Ureaplasma urealyticum, Mycoplasma genitalium) Occasional Trichomonas vaginalis, HSV Unknown in ~50% cases

Sx: Mild dysuria, mucoid discharge Dx: Urethral smear 5 PMNs (usually 15)/OI field Urine microscopic 10 PMNs/HPF Leukocyte esterase (+) 72
Dr.T.V.Rao MD

Chlamydia

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Chlamydia
Chlamydia is an infection of the penis, vagina, throat, or tube that carries urine. Chlamydia is caused by bacteria (a kind of germ). You get it by having sex with someone who has Chlamydia. Chlamydia can be spread by the vagina, penis, mouth, or anus.
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Chlamydia
Too many people are continuing to have unsafe sex, put themselves at risk of STIs and the serious consequences associated with infection, including infertility. "On-going investment in programmes to increase sexual health awareness, condom use and testing, particularly for groups at most risk, is vital.
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Chlamydia trachomatis
More than three million new cases annually Responsible for causing cervicitis, urethritis, Proctitis, lymphogranuloma venereum, and pelvic inflammatory disease Direct and indirect cost of chlamydial infections run into billions of dollars Potential to transmit to newborn during delivery
Conjunctivitis, pneumonia
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Normal Cervix

Dr.T.V.Rao MD Source: Claire E. Stevens, Seattle STD/HIV Prevention Training Center

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Chlamydia Cervicitis

Dr.T.V.Rao MD Source: St. Louis STD/HIV Prevention Training Center

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Mucopurulent Cervicitis

Dr.T.V.Rao MD Source: Seattle STD/HIV Prevention Training Center

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Drips

Laboratory Tests for Chlamydia


Tissue culture has been the standard
Specificity approaching 100% Sensitivity ranges from 60% to 90%

Non-amplified tests
Enzyme Immunoassay (EIA), e.g. Chlamydiazyme
sensitivity and specificity of 85% and 97% respectively useful for high volume screening false positives

Nucleic Acid Hybridization (NA Probe), e.g. Gen-Probe Pace2


sensitivities ranging from 75% to 100%; specificities greater than 95% detects chlamydial ribosomal RNA able to detect gonorrhea and chlamydia from one swab need for large amounts of sample DNA
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Laboratory Tests for Chlamydia


(continued)

DNA amplification assays


polymerase chain reaction (PCR) ligase chain reaction (LCR)

Sensitivities with PCR and LCR 95% and 85-98% respectively; specificity approaches 100% LCR ability to detect chlamydia in first void urine
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Chlamydia Direct Fluorescent Antibody (DFA)

Source: Centers for Disease Control and Prevention

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Drips

Pelvic Inflammatory Disease (PID)


l0%-20% women with GC develop PID In Europe and North America, higher proportion of C. trachomatis than N. gonorrhoeae in women with symptoms of PID CDC minimal criteria
uterine adnexal tenderness, cervical motion tenderness

Other symptoms include


endocervical discharge, fever, lower abd. pain

Complications:
Infertility: 15%-24% with 1 episode PID secondary to GC or chlamydia 7X risk of ectopic pregnancy with 1 episode PID chronic pelvic pain in 18%
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Chancroid
The combination of a painful genital ulcer and tender Suppurative inguinal adenopathy suggests the diagnosis of Chancroid A probable diagnosis of Chancroid, for both clinical and surveillance purposes, can be made if all of the following criteria are met: 1) the patient has one or more painful genital ulcers; 2) the patient has no evidence of T. pallidum infection by dark field examination of ulcer exudate or by a serologic test for syphilis performed at least 7 days after onset of ulcers;
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Chancroid
3) the clinical presentation, appearance of genital ulcers and, if present, regional lymphadenopathy are typical for Chancroid; and 4) a test for HSV performed on the ulcer exudate is negative.
Dr.T.V.Rao MD 85

Chancroid
A definitive diagnosis of Chancroid requires the identification of H. ducreyi on special culture media that is not widely available from commercial sources; even when these media are used, sensitivity is <80% (145).
Dr.T.V.Rao MD 86

Granuloma Inguinale (Donovanosis)


Granuloma inguinale is a genital ulcerative disease caused by the intracellular gramnegative bacterium Klebsiella granulomatis (formerly known as Calymmatobacterium granulomatis). The disease occurs rarely in the United States, although it is endemic in some tropical and developing areas, including India; Papua, New Guinea; the Caribbean; central Australia; and southern Africa (192,193).
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Granuloma Inguinale (Donovanosis)


Clinically, the disease is commonly characterized as painless, slowly progressive ulcerative lesions on the genitals or perineum without regional lymphadenopathy; subcutaneous granulomas (pseudoboboes) might also occur. The lesions are highly vascular (i.e., beefy red appearance) and bleed easily on contact.
Dr.T.V.Rao MD 88

Granuloma Inguinale (Donovanosis)


The causative organism is difficult to culture, and diagnosis requires visualization of darkstaining Donovan bodies on tissue crush preparation or biopsy. No FDA-cleared molecular tests for the detection of K. granulomatis DNA exist, but such an assay might be useful when undertaken by laboratories that have conducted a CLIA verification study.
Dr.T.V.Rao MD 89

Lymph granuloma Venereum


Lymph granuloma venereum (LGV) is caused by C. trachomatis serovars L1, L2, or L3 . The most common clinical manifestation of LGV among heterosexuals is tender inguinal and/or femoral lymphadenopathy that is typically unilateral.
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Lymph granuloma Venereum


Diagnosis is based on clinical suspicion, epidemiologic information, and the exclusion of other Aetiologies for proctocolitis, inguinal lymphadenopathy, or genital or rectal ulcers. C. trachomatis testing also should be conducted, if available.
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Lymph granuloma Venereum


Genital and lymph node specimens (i.e., lesion swab or bubo aspirate) can be tested for C. trachomatis by culture, direct immunofluorescence, or nucleic acid detection. NAATs for C. trachomatis are not FDA-cleared for testing rectal specimens, although some laboratories have performed the CLIA validation studies that are needed to provide results for clinical management. Additional molecular procedures (e.g., PCR-based genotyping) can be used to differentiate LGV from non-LGV C. trachomatis, but these are not widely available.
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Lymph granuloma Venereum


chlamydia serology (complement fixation titers >1:64) can support the diagnosis of LGV in the appropriate clinical context. Comparative data between types of serologic tests are lacking, and the diagnostic utility of serologic methods other than complement fixation and some micro immunofluorescence procedures has not been established.
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An estimated 5 million new cases occur each year in women and men. Occurs in vagina of women so may be sexually transmitted to men using infected washcloths and towels. It is transmitted to the baby during delivery. It also can occur in the urethra (carries urine to penis) in men, doesnt have symptoms usually. SYMPTOMS: Appear within 5 to 28 days of exposure Women usually have a vaginal discharge that FEMALE SYMPTOMS: Itching and burning at the outside of the opening of the vagina and vulva. Painful and frequent urination Heavy, unpleasant smelling greenish, yellow discharge MALE SYMPTOMS: Usually nothing, or discomfort in urethra, inflamed head of the penis.
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Trichomoniasis

Bacterial Vaginitis
Controversy: STD - yes or no Need for treatment
1980: only if patient complains 2002: increased risk of:
Preterm birth / premature rupture of membranes Amniotic fluid infection Chorioamnionitis / Postpartum endometritis Pelvic inflammatory disease Postsurgical infection Cervical intraepithelial neoplasia Mucopurulent cervicitis Acquisition of HIV Dr.T.V.Rao infection MD

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Life at Risk with Sexually Transmitted Infections Best Choice Play safe

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Programme Created by Dr.T.V.Rao MD for Medical and Paramedical Students in the Developing World

Email
doctortvrao@gmail.com

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