NEISSERIACEAE (PATHOGENIC)
VIRULENCE FACTORS
I. GENERAL CHARACTERISTICS
- Obligate Aerobe
- Non-motile
- Non-spore forming
- Capnophilic: Prefer increased CO2 (3-
10%)
- Optimal growth: humid atmosphere
- Gram negative diplococci
- NOTE: All Neisseria species are gram
negative diplococci except: Neisseria
elongata, Neisseria
weaveri, and Neisseria bacilliformis - rod
shaped
- NOTE: All species are cytochrome
oxidase- and catalase-positive except
for N. elongata and N.
bacilliformis, which are catalase-
negative.
- Neisseria gonorrhoeae (often called
gonococci) and Neisseria meningitidis
(meningococci) are the
primary human pathogens of the genus.
• N. gonorrhoeae is not considered to be
part of the normal biota and is always
pathogenic.
• N. meningitidis may be found as a
commensal inhabitant of the upper
respiratory tract of carriers,
but it can also become an invasive
pathogen.
II.
- Pathogenic Neisseria spp. have
several characteristics that contribute to
their virulence, including the
following:
a. Receptors for human transferrin
b. Capsule (N. meningitidis): has many
strains (9 serogroups) based on
capsular polysaccharide
• A, B, C, Y and W-135: most often
associated with epidemics
c. Pili (fimbriae)
d. Cell membrane proteins
e. Lipooligosaccharide (LOS) or
endotoxin;
f. Immunoglobulin A (IgA) protease
III. Neisseria gonorrhoeae
- Humans are the only natural host for
N. gonorrhoeae, the agent of gonorrhea-
an acute pyogenic
infection of non-ciliated columnar and
transitional epithelium; infection can be
established at any site
where these cells are found.
- Gonococcal infections are primarily
acquired by sexual contact and occur
primarily in the urethra,
endocervix, anal canal, pharynx, and
conjunctiva.
- The first use of the term gonorrhea,
meaning a “flow of seed,” was in the
second century when the
urethral discharge was mistaken for
semen.
- Clinical Infections:
• Gonorrhea has a short incubation
period of approximately 2 to 7 days
• In men, acute urethritis, usually
resulting in:
✓purulent discharge
✓dysuria (painful urination)
• N. gonorrhoeae strains with a
nutritional requirement for arginine,
hypoxanthine, and uracil (AHU
strains) are often isolated from
asymptomatic men (3-5%)
• The endocervix is the most common
site of infection in women.
• 50% of cases in women may be
asymptomatic leading to complications
such as pelvic
inflammatory disease, which may cause
sterility, ectopic pregnancy, or
perihepatitis (Fitz-Hugh–
Curtis syndrome).
• Other conditions associated with N.
gonorrhoeae include anorectal and
oropharyngeal infections.
Infections in these sites are more
common in men who have sex with men
but can also occur in
women.
• Newborns can acquire ophthalmia
neonatorum- a gonococcal eye infection,
during vaginal delivery
through an infected birth canal.
• Ocular infections can occur in adults
because of inoculation of the eye with
infected genital secretions or, rarely, as
a result of a laboratory accident.
- Laboratory Diagnosis:
a. Specimen collection & transport
• The specimen of choice for genital
infections in men is the urethra and in
women is the endocervix. In men,
purulent discharge can be collected
directly onto a swab for culture.
• N. gonorrhoeae is inhibited by SPS.
✓Gelatin can be added to the media to
neutralize the effects of SPS.
• When no apparent discharge is
present, the swab is inserted up to 2 cm
into the anterior urethra and slowly
rotated to collect material. Swabs for
rectal culture should be inserted 4 to 5
cm into the anal canal. Disinfectants
should be avoided in preparing the
patient for collection of the specimen.
• Calcium alginate and cotton swabs are
inhibitory to N. gonorrhoeae, so Dacron
or rayon swabs are preferred.
• Direct plating of the specimen to
gonococcal selective media gives
optimal results.
• Several commercial transport systems,
such as James E. Martin Biological
Environmental
Chamber (JEMBEC) plates, Gono-Pak
and Transgrow allow direct plating in the
clinical setting.
b. Direct microscopic examination
• Smears for direct Gram stains should
be prepared from urogenital specimens.
• The demonstration of gram-negative
intracellular diplococci from the urethral
discharge with
culture is evidence of gonococcal
infection.
c. Culture
• N. gonorrhoeae typically does not grow
on SBA; the medium of choice for
cultivation is CHOC agar. Because
CHOC agar supports the growth of
many other organisms found as
commensals in specimens collected for
the recovery of gonococci, a selective
medium is necessary.
d. Incubation
• Inoculated plates should be incubated
at 35° C in a 3% to 5% CO2
atmosphere. Scented or
colored candles may be inhibitory to the
gonococci, so only white wax candles
are used in the candle extinction jar.
Cultures are examined daily for growth
and held for 72 hours.
e. Colony morphology
• Colonies of N. gonorrhoeae on CHOC
agar or selective agars are small, gray
to tan, translucent, and raised after 24 to
48 hours of incubation.
• T1 and T2 – piliated (virulent):
Colonies are smaller, raised and appear
bright in reflected light
• T3 – T5 – non-piliated (avirulent):
Larger, flatter colonies
f. Oxidase test
• The oxidase test must be done on all
suspected isolates of N. gonorrhoeae
• Rgt.:1% tetramethyl-p-
phenylenediamine dihydrochloride/ 1%
dimethyl – p – phenylenediamine
dihydrochloride
g. Superoxol test
• (+) production of gas bubbles from
30% H2O2
• Performed using the procedure in
Catalase test
✓Vigorous bubble: N. gonorrhoeae
✓Weak, delayed bubbling:
N.meningitidis
h. Carbohydrate utilization
• Cystine trypticase agar (CTA),
containing 1% of the individual
carbohydrate and phenol red as
a pH indicator.
• If the organism uses the particular
carbohydrate, acid, characterized by a
yellow color, is produced in 24 to 72
hours.
• N. gonorrhoeae is positive for glucose
only.
i. Almost all strains of N. gonorrhoeae:
susceptible to penicillin
• (PPNG): penicillinase-producing N.
gonorrhoeae
• (CMRNG): chromosomally mediated
resistant N. gonorrhoeae to both
penicillin and tetracycline
• Cephalosphorins: recommended for
treatment
IV. Neisseria meningitidis
- Similar to N. gonorrhoeae, N.
meningitidis is also found only in
humans. However, N. meningitidis can
be found as a commensal as well as an
invasive pathogen.
- Can be found on the mucosal surfaces
of the nasopharynx and oropharynx in
30% of the population.
- Clinical infections:
a. Meningococcemia, or sepsis, may
occur with or without meningitis and
carries a 25% mortality
rate, even if treated.
• The disease becomes fulminant and
spreads rapidly, causing DIC, septic
shock, or hemorrhage in the adrenal
glands (Waterhouse-Friderichsen
syndrome).
b. Waterhouse-Friderichsen Syndrome
(W-135)
• Adrenal hemorrhage observed during
sepsis, as in the Waterhouse-
Friderichsen syndrome, may be
attributable to endotoxemia occurring
during or shortly after stimulation of the
adrenal cortex by infection.
- Laboratory diagnosis
a. Specimen collection & transport
• Cerebrospinal fluid (CSF), blood,
nasopharyngeal swabs and aspirates,
joint fluids, and less commonly sputum
and urogenital sites.
• N. meningitidis is also inhibited by
SPS.
✓Gelatin can be added to the media to
neutralize the effects of SPS.
b. Direct microscopic examination
• Intracellular and extracellular gram-
negative diplococci.
• At least 1 mL of CSF should be
centrifuged (increases the detection) at
1000 x g for 10
minutes and the sediment used for slide
and culture.
c. Culture & incubation
• Specimens for isolation of N.
meningitidis should be cultured on SBA
and CHOC agar.
• Cultures should be incubated in the
same atmospheric conditions described
for N.
gonorrhoeae and examined daily for 72
hours.
d. Carbohydrate utilization
• Definitive identification of N.
meningitidis can be made by the use of
carbohydrate methods, chromogenic
enzyme tests, or multi-test assays.
• In carbohydrate methods, N.
meningitidis uses glucose and maltose.
- N. lactamica (delayed lactose
fermenter), generally a nonpathogenic
species that may mimic N.
meningitidis, can also grow on selective
media. A rapid o-nitrophenyl-β-D-
galactopyranoside (ONPG)
test, which detects lactose utilization, is
usually positive in 30 minutes for N.
lactamica.
NEISSERIACEAE (NON-
PATHOGENIC)
I. GENERAL CHARACTERISTICS
- Normal flora
- Rare cause of infection
- Sporadically implicated with:
• Meningitis
• Endocarditis
• Prosthetic valve infections
• Bacteremia
- May share some characteristics
(morphology and biochemical) with the
pathogenic Neisseria
- Must be identified and separated from
the pathogenic ones
II. Neisseria cinerea
- Resembles with N. gonorrhoeae in
these aspects:
• Biochemical properties (ferments
glucose)
• Colony characteristic (T3 type in CA)
- Methods of Differentiation:
• COLISTIN SUSCEPTIBILITY Test
• INABILITY of N. cinerea to grow on
MTM
• INABILITY of N. gonorrhoeae to grow
on MHA
• NEGATIVE RESULTS in SUGAR
FERMENTATION TEST using CTA
(N.cinerea)
III. Neisseria flavescens
- Yellow pigmented specie
- ASACCHAROLYTIC (Cannot ferment
sugars)
- Differentiated from N.cinerea by its
ability to grow on SBA at 22oC and
yellow colonies
IV. Neisseria lactamica
- Mistaken with N.meningitidis
- Can grow in MTM, ferments Glucose
and Maltose in the CHO utilization test
- flora of the pharynx of infants and
children
- The only specie that ferments lactose
in the CHO utilization (Definitive tool for
differentiation)
V. Neisseria mucosa
- Has very mucoid colonies
- Isolated from nasopharynx of children
- Rare cause of pneumonia in children
- May be confused with N.sicca and
N.subflava biovar perflava (based on
CHO fermentation)
- Reduces nitrate to nitrogen gas
(Definitive tool for differentiation)
VI. Neisseria polysaccharea
- Isolated from throats of healthy
children
- Produces large amounts of
extracellular polysaccharide in media
with 1% sucrose
- Misidentified as N.meningitidis (due to
its positive glucose and maltose
utilization)
- Ability to grow on NA and
polysaccharide production with 1% or
5% sucrose (Tool for differentiation)
VII. Neisseria sicca
- Dry, wrinkled, adherent, breadcrumb
like colonies
- Found in RT of adults
VIII.Neisseria subflava
- Flora of the upper RT
- Rare cause of serious bacteremia,
meningitis and septicemia
- Manifestation – resembles
meningococcal infection
IX. Moraxella catarrhalis
- Resembles Neisseria in morphology
and biochemical properties
- OXIDASE POSITIVE; CATALASE
POSITIVE
- Flora of the upper RT
- Opportunistic pathogen
- Grows on NA (N.gonorrhea and
N.meningitidis can’t)
- DNAse positive (N.gonorrhea and
N.meningitidis negative)
- Fails to utilize CHO in the CTA medium
- Beta lactamase +
- “Hockey puck” colonies on SBA &
CHOC agar
- produce Blue color (+) on Butyrate
Disk/ Tributyrin HOH Test (N.gonorrhea
= no color change -)
X. Moraxella bovis
- Agent of pink eye conjunctivitis
- DNAse + like M.catarrhalis
- Oxidase -
- Catalase +
- Resembles Neisseria especially in
specimens collected from vaginal and
genital cultures
- OXIDASE TEST (Definitive tool for
differentiation)