1. Lab Testing for STIs and UTIs
Treponema pallidum -
Syphilis
Corkscrew-shaped,motile G(-)
spiral bacteria
Presentation: 3rd
most common
bacterial STI;onset
2-3 weeks after
exposure
Sx: primary syphilis
with non-tender
chancrewith
smooth margins;
secondary syphilis
with flu-like
symptoms and skin
rash (specifically on
palms and soles);
tertiary with CNS
involvement
Primary and
secondary stages
most contagious
Females: possible
transiently
bacteremic without
symptoms; can
pass infection to
fetus up to 8 yrs
followinginitial
infection
Dx based on clinical
presentation and
(+) serology
Specimens:
serous exudates
from primary
chancre,
condyloma latum,
mucous patch
lesion,lymph
node aspirate;
also blood or CSF
Identification: not routinely cultured; can be visualized on dark field or immunofluorescence
microscopy.Non-treponemal tests measure IgG/IgM to lipoidal material released from
damaged host cell or to pathogenic lipids/lipoproteins.Nucleic Acid Amplification Testing
(NAAT) used with cervical swab,vaginal swab,or urinespecimen.
Non-specific serological tests:
 Rapid Plasma Reagin Card (RPR): most commonly used test but ~ 30% of patients will not
have + test until later in courseof disease; carbon beads coated w/ synthetic Ag (cardiolipin
lecithin,and cholesterol) to detect Ab in serum; (+) result= black clumps againstwhite
background.
Biologic falsepositives:maternal Ab in neonatal syphilis
 Venereal Disease Research Laboratory (VDRL) test: slidemicroflocculation testthat detects
Ab usingsynthetic Ag; primarily for CSF and not useful for very early or advanced stages of
disease
Ab-specific serological tests:
 TP-PA: (Treponema pallidumparticleagglutination):gelatin particles coated with T. pallidum
pallidum Ag; serial dilutions of ptserum evaluated via microtiter plate
 Enzyme immunoassays (EIAs): pt serum added to microtiter plates containingwells coated
with T. pallidum pallidum Ag, then labeled anti-human IgG added to measure specific IgGin
the patient’s serum
 FTA-ABS (fluorescentAb assay) and MHA-TP (microhemagglutination assay) detect Ab
againstsyphilis;notfor screening but used to confirm infection,except in first3-4 weeks
post-exposure, usingblood or CSF (FTA-ABS only)
Western blot with T. pallidumAg used for independent verification of dx

2. Neisseria Gonorrhoeae-
Gonorrhea
“Bean-shaped” G(-) diplococci
with flattened sides adjacent
sides,cell wallsappear outlined
Presentation: 2nd
most common
bacterial STI and
causeof urethral
discharge;onset 2 -
3 d after exposure
Sx: intense dysuria
and purulent
discharge;may be
asymptomatic
Uncomplicated
Gonorrhea:
infection of
columnar epithelial
cells in mucosal
membranes of
lower genital tract
Other
presentations:
Oropharyngeal,
anorectal,ocular
Specimens:
Genital tract,
urine(firstvoid),
anal area,
oropharynx,
conjunctiva,
Bartholin’s glands,
fallopian tubes,
endometrium,
blood,jointfluid,
skin lesions,
neonate gastric
contents
30 – 60% males
have frequent co-
infection w/ C.
trachomatis;
recommend test
for syphilisand
HIV
Collection Method:
 Mucosal surface
sampled with
Dacron swabs -
fatty acids in
cotton swabs may
kill Gonorrhoeae
 Urethral samples >
1 hr after urination
 Discard anorectal
swabs
contaminated w/
fecal material and
repeat
Culture: should always beperformed to
test for β-lactamaseproduction
Media: inoculation directly onto media
and incubated at 35 – 37° C in CO2
enriched atmosphere
 Chocolateagar supplemented w/
IsoVitaleX (supplies factors Vand X) –
nonselectivemedia for cultivation
 Modified Thayer-Martin (MTM), Martin-
Lewis (ML) and New York City (NYC)
media - selectivemedia for use with
samples containingnormal flora
 Non-culture methods: ELIZA and nucleic
acid test (NAAT)
Transport: rapidly loses viability in
buffered, nonnutritive transportmedia
Storage: CO2 enriched environment when
incubator not available
Identification: Patient
samples may have large#
PMNs, some having
intracellular G(-)
diplococci;numbers of
intracellular pathogen low
in both early and late
phases of infection
Male: presumptive Dx
based on patient hx,
clinical signs & symptoms,
and G(-) diplococci within
PMNs
Female: urogenital tract
contains flora closely
resemblingN.
Gonorrhoeae; G(-) results
only presumptive until
confirmed by (+) culture
or (+) PCR

3. Chlamydiatrachomatis
G(-)
Nonmotile G(-) coccobacillary
organisms lined up likea
“school of fish”
Presentation:
Most common bacterial STI and most common causeof urethral
discharge;most frequently reported STI in U.S.
Sx: most asymptomatic but if symptoms arepresent, onset 1-3
weeks after exposure
Females: may have abnormal vaginal dischargeor burningduring
urination;if spread to fallopian tubes then may have lowback or
abdominal pain;cervical infection can spread to rectum
Males:may have urethral itching/burning,discharge,or burning
duringurination
Two forms:
 Elementary bodies (EB): small,dense spherical bodies with
diameter of 0.6 – 0.4 µm; extracellular formand is infectious
 Reticulate bodies (RB): larger cocci with diameter of 0.6 – 1.0
µm; intracellular formthatis metabolically activewithin host
cell
Culture and Identification:
 Difficultto culture; culture usingHeLa or McCoy host cells in tissue
culture(second most sensitivetest)
Presence of inclusion bodies differential for C.trachomatis vs.C.
psittici or C.pneumoniae
 PCR based tests most sensitive
 DFA staining3rd mostsensitivefor detection
 Non-culture methods: ELIZA and nucleic acid test(NAAT)
 Rapid tests areavailableto detect Ab against C.trachomatis strains
L1, L2, and L3 that causelymphogranuloma venereum; may yield false
positiveresults dueto cross-reactivity with other C. trachomatis
strains,so resultmustbe confirmed with more sensitivetesting
standards

4. Non-gonococcal urethritis
(NGU):
Onset 10 -14 d, less severe Sx
than Gonorrhea (often
asymptomatic)
 > 100,000 (105) = UTI/ bacteruria
 104 – 105 = possiblebacteruria
 103 – 104 = suspect, retest and use clean catch
 < 103 = normal levels and any bacteria likely normal flora
Urine sedimentation rate: ~ 75% of patients with UTIs will havemore
than 100 WBCs/mL of urine
Gram stain:1-2 bacteria per 1000x microscopic field yieldspresumptive
dx of bacteruria
Escherichia coli
G(-)
Rare causeof
urethral discharge
Group 1: produces acid by fermentation of
lactosein Bacturcultmedium – resultis
yellow
E. coli, Citrobacter, Enterobacter,
Streptococci, Enterococci
Urine specimen must be transported to lab within 30 minutes to
prevent falsely elevated counts – E. coli generation time ~ 20 min.
Bacturcult test:
 Culture medium supplyingnutrients for growth of various species of
aerobic bacteria
 Proteus swarminginhibited by para-nitropheyl glycerol
 Contains phenol red indicator,which turns yellowin presence of acid
and red in alkalineconditions
Pure cultures: discretecolonies with detectable characteristics
produced by medium; presumptive differentiation into 3 groups
 Group 1: ferments lactoseand produces acid  yellow medium
 Group 2: produce rose to orange medium
 Group 3: alkalineconditions  magenta medium
Mixed cultures should be checked for divergent characteristicsand
may indicatecontamination;especially common in females when urine
stream contacts external genitalia
 Mixed cultures with less than 1000 bacteria /mL of urine=
contamination (1 species normally dominant)
Limits of Bacturcult test: presumptive results,color changes can be
masked by drug effects (dyes, antibacterials),results may vary based on
condition of patient and viability of bacteria in specimen,the results
must be evaluated in lightof total clinical information obtained,and
mixed cultures may produce inconclusivecolor changes thatdo not
directly correlatewith relativeproportion of bacteria in the sample
Enterobacter
G(-)
Citrobacter
G(-)
Proteus
G(-)
Group 3: produces alkalineconditions in
Bacturcultmedium – resultis magenta
(purple-red)
Proteus and Pseudomonas
Pseudomonas
G(-)
Klebsiella pneumoniae
G(-)
Group 2: conditions produced by these
bacteria (includingsomePseudomonas
species and some Enterococci species)
resultin rose to orange coloration of
Bacturcultmedium
Klebsiella pneumoniae, Staphylococci,
Streptococci and Enterococci,
Pseudomonas
Enterococci
G(+)
Streptococcus
G(+)
Staphylococcus
G(+)
Uristix 4 Reagent Strips: strips contain 4 separatereagent zones to test for presence of glucose,
protein, nitrite and leukocytes in urine
 Glucose: glucoseoxidaseand peroxidaseenzymes present catalyzereaction of color change
rangingfrom green to brown
 Protein: at constant pH, yellow is protein (-) and any color rangingfromyellow-green to green to
blue-green is protein (+)
 Nitrite: at acidicpH,dietary nitrate is converted to nitriteby G(-) bacteria in urine,producinga
pink color
 Leukocytes: production of a purplecolor indicates thepresence of granulocytic leukocytes
containingesterases thatcatalyzehydrolysis of chromogenic ester substrates on test strip;
presumptive test indicatingpresenceof PMNs – PMN content must be verified and pathogen
must be identified