Family Neisseria

Neisseria: General Features

• Neisseria are Gram-negative cocci that typically appear in pairs with the opposing sides flattened,
imparting a "kidney bean" appearance.
• They are nonmotile, non–spore-forming, and non–acid-fast.
• Their cell walls are typical of Gram-negative bacteria, with a peptidoglycan layer and an outer
membrane containing endotoxic glycolipid complexed with protein.
• Two species cause disease in humans: Neisseria meningitidis and Neisseria gonorrhoeae.
• The structural elements of N meningitidis and N gonorrhoeae are the same, except that the
meningococcus has a polysaccharide capsule external to the cell wall.
• Gonococci and meningococci require an aerobic atmosphere with added carbon dioxide and enriched
medium for optimal growth.
• Gonococci grow more slowly and are more fastidious than meningococci, which can grow on routine
blood agar.
• All Neisseria are oxidase-positive.
• Species are defined by growth characteristics and patterns of carbohydrate fermentation.
• Both pathogenic species possess pili and outer membrane proteins (OMPs), which vary in their
function and antigenic composition.
• The outer membrane of pathogenic Neisseria contains a variant of lipopolysaccharide (LPS) in which
the side chains are shorter and lack the repeating polysaccharide units found in the LPS of most other
Gram-negative bacteria. This short-chain neisserial LPS is called lipooligosaccharide (LOS).
• The lipid A and core oligosaccharide are structurally and functionally similar to other Gram-negative
LPS.
• The pili, OMPs, and LOS are antigenic and have been used in typing schemes.
Neisseria Meningitidis
 Bacteriology

• Meningococci produce medium-sized smooth colonies on

blood agar plates after overnight incubation.
• Carbon dioxide enhances growth, but is not required.
• Twelve serogroups have been defined on the basis of the
antigenic specificity of their polysaccharide capsule.
• The most important disease-producing serogroups are A, B, C,
W-135, and Y.
• In addition to the group polysaccharides, individual N
meningitidis strains may contain two distinct classes of pili and
multiple classes of OMPs including porins and adherence
proteins, some of which have structural and functional
similarities to those found in gonococci.
• The function of other OMPs is unknown.
Virulence factors of the meningococcus include:
1) Capsule: A polysaccharide capsule surrounds the bacterium and is
antiphagocytic, as long as there are no specific antibodies to coat (opsonize)
the bacterium.
• Neisseria meningitidis is classified into serogroups based on different
capsular polysaccharides, which are antigenic (stimulate a human antibody
response).
• There are 9 serotypes of meningococcus (designated A,B, C, D, X, Y, Z,
W135, and 29E).
• Meningitis is caused by groups A, B, and C.
2) Endotoxin (LPS): The meningococci can release blebs of endotoxin, which
causes blood vessel destruction (hemorrhage) and sepsis. The blood vessel
hemorrhage is seen on the skin as tiny, round, red dots of hemorrhage called
petechiae (a petechial rash). This same hemorrhaging process can damage
the adrenal glands.
3) IgA1 protease: This is only found in pathogenic species of Neisseria. This
enzyme cleaves IgA (a type of antibody) in half.
4) Neisseria meningitidis can extract iron from human transferrin via a non-
energy requiring mechanism.
 Meningococcal Disease

• Meningococci are usually quiescent members of the nasopharyngeal flora

but may produce fulminant infection of the bloodstream and/or central
nervous system (CNS).
• There is little warning; localized infections that precede systemic spread are
rarely recognized.
• The major disease is an acute, purulent meningitis with fever, headache,
seizures, and mental signs secondary to inflammation and increased
intracranial pressure.
• Even when the CNS is not involved, N meningitidis infections have a
marked tendency to be accompanied by rash, purpura, thrombocytopenia,
and other manifestations associated with endotoxemia.
• This bacterium causes one of the few infections in which patients may
progress from normal health to death in less than a day.
• It can also spread quickly in family, school, and even national outbreaks.
• High-risk groups are:
• 1) Infants aged 6 months to 2 years
• 2) Army recruits
 Transmission and Epidemiology

• Because meningococci do not survive long in the environment, these bacteria are usually
acquired through close contact with secretions or droplets.
Upon reaching their portal of entry in the nasopharynx, the meningococci attach there using pili.
• In many people, this can result in simple asymptomatic colonization.
• In the more vulnerable individual, however, the meningococci are engulfed by epithelial cells
of the mucosa and penetrate into the nearby blood vessels, along the way damaging the
epithelium and causing pharyngitis.
• Meningococcal meningitis has a sporadic or epidemic incidence in late winter or early
spring.
• The continuing reservoir of infection is humans who harbor the pathogen in the
nasopharynx.
• The scene is set for transmission when carriers live in close quarters with nonimmune
individuals, as might be expected in families, day care facilities, college dormitories, and
military barracks.
• The highest risk groups are young children (6 to 36 months old) and older children and
young adults (10 to 20 years old).
• Every year, in what is called “the meningitis belt” in sub-Saharan Africa, a meningococcal
epidemic sweeps through, coinciding with the dry season which runs from approximately
December to May.
• In 2009, a particularly large outbreak killed more than 2,100 people in Niger and Nigeria and
infected tens of thousands.
• Many more would have been affected except for a massive mobilization of vaccine.
• In the space of 4 months, 7.5 million people were vaccinated.
 Manifestations

• The most common form of meningococcal infection is acute purulent

meningitis, with clinical and laboratory features similar to those of
meningitis from other causes.
• A prominent feature of meningococcal meningitis is the appearance of
scattered skin petechiae, which may evolve into ecchymoses or a diffuse
petechial rash.
• These cutaneous manifestations are signs of the disseminated intravascular
coagulation (DIC) syndrome, which is part of the endotoxic shock brought on
by meningococcal bacteremia (meningococcemia).
• Meningococcemia sometimes occurs without meningitis and may progress
to fulminant DIC and shock with bilateral hemorrhagic destruction of the
adrenal glands (Waterhouse–Friderichsen syndrome).
• However, the disease is not always fulminant, and some patients have only
low-grade fever, arthritis, and skin lesions that develop slowly over a period
of days to weeks.
• Meningococci are a rare cause of other infections such as pneumonia, but it
is striking that localized infections are almost never recognized in advance of
systemic disease.
 Immunity
• Immunity to meningococcal infections is related to group-specific
antipolysaccharide antibody, which is bactericidal and facilitates
phagocytosis.
• The bactericidal activity is due to complement-mediated cell lysis via the
classical complement pathway.
• The peak incidence of serious infection occurs between 6 months and 2
years of age. This corresponds to the nadir in the prevalence of antibody in
the general population, which is the time between loss of transplacental
antibody and the appearance of naturally acquired antibody.
• By adult life, serum antibody to one or more meningococcal serogroups is
usually present, but an immune deficit to the other serogroups remains.
• Infections appear when populations carrying virulent strains mix (college,
summer camp, military barracks) and susceptible individuals acquire a new
strain of a serogroup to which they lack group-specific antibody.
• Protective antibody is stimulated by infection and through the carrier state,
which produces immunity within a few weeks.
 Diagnosis
• Diagnosis involves Gram stain and culture of the meningococcus from blood,
cerebrospinal fluid, or petechial scrapings.
• Neisseria grow best on blood agar that has been heated so that the agar turns brown
(called chocolate agar).
• The classic medium for culturing Neisseria is called the Thayer-Martin VCN media.
This is chocolate agar with antibiotics, which are included to kill competing bacteria.
• V stands for vancomycin, which kills gram-positive organisms.
• C stands for colistin (polymyxin) which kills all gram-negative organisms (except
Neisseria).
• N stands for nystatin, which eliminates fungi.
• Therefore, only Neisseria (both Neisseria meningitidis and Neisseria gonorrhoeae)
are able to grow on this culture medium.
• The addition of a high concentration of CO2 further promotes the growth of
Neisseria.
• Presumptive identification of the genus is obtained by a Gram stain and oxidase
testing on isolated colonies
• In the laboratory, the differentiation between the Neisseria species is based on
Neisseria meningitidis‘ ability to produce acid from maltose metabolism, while
Neisseria gonorrhoeae cannot!
 Treatment
• Prompt treatment with penicillin G or ceftriaxone is
required at the first indication of disseminated
meningococcemia.
• Close contacts of an infected patient are treated with
rifampin.
• Immunization with purified capsular polysaccharides
from certain strains (groups A, C, Y, and W135) is
currently available and used for epidemics and in
high-risk groups.
• The group B polysaccharide does not induce
immunity, so a vaccine is not available at present.
Neisseria Gonorrhoeae
 Bacteriology

• N gonorrhoeae grows well only on chocolate agar and on specialized

medium enriched to ensure its growth.
• It requires carbon dioxide supplementation.
• Small, smooth, nonpigmented colonies appear after 18 to 24 hours and
are well developed (2 to 4 mm) after 48 hours.
• Gonococci possess numerous pili that extend through and beyond the
outer membrane, which are structurally similar to those of meningococci.
• The gonococcal outer membrane is composed of phospholipids, LPS, LOS,
and several distinct OMPs.
• The OMPs include porins (PorA and PorB) and adherence proteins known
as Opa.
• Opa proteins are a set of at least 12 proteins that get their name from the
opaque appearance they give to colonies as a result of adhesion between
gonococcal cells.
• A variable number of the Opa proteins may be expressed at any one time.
 Virulence factors of the gonococcus include:
• 1) Pili: Neisseria gonorrhoeae has complex genes coding for
their pili.
• These genes undergo multiple recombinations, resulting in
the production of pili with hypervariable amino acid
sequences. These changing antigens in the pili protect the
bacteria from our antibodies, as well as from vaccines aimed
at producing antibodies directed against the pili.
• The pili adhere to host cells, allowing the gonococcus to
cause disease. They also serve to prevent phagocytosis,
probably by holding the bacteria so close to host cells that
macrophages or neutrophils are unable to attack.
• 2) Protein II: This outer membrane protein is also involved in
adherence to host cells.
 Pathogenesis and Virulence Factors

• Successful attachment is key to the organism’s ability to cause disease.

• Gonococci use specific chemicals on the tips of fimbriae to anchor themselves to
mucosal epithelial cells.
• They only attach to nonciliated cells of the urethra and the cervix, for example.
• Once the bacterium attaches, it invades the cells and multiplies on the basement
membrane.
• The fimbriae may also play a role in slowing down effective immunity.
• The fimbrial proteins are controlled by genes that can be turned on or off, depending
on the bacterium’s situation.
• This phenotypic change is called phase variation. In addition, the genes can rearrange
themselves to put together fimbriae of different configurations.
• This antigenic variation confuses the body’s immune system.
• Antibodies that previously recognized fimbrial proteins may not recognize them once
they are rearranged.
• The gonococcus also possesses an enzyme called IgA protease, which can cleave IgA
molecules stationed on mucosal surfaces.
• In addition, it pinches off pieces of its outer membrane. These “blebs,” containing
endotoxin, probably play a role in pathogenesis because they can stimulate portions of
the nonspecific defense response, resulting in localized damage.
 Transmission and Epidemiology

• N. gonorrhoeae does not survive more than 1 or 2

hours on fomites and is most infectious when
transferred to a suitable mucous membrane.
• Except for neonatal infections, the gonococcus
spreads through some form of sexual contact.
• The pathogen requires an appropriate portal of entry
that is genital or extragenital (rectum, eye, or throat).
• Gonorrhea is a strictly human infection that occurs
worldwide and ranks among the most common
sexually transmitted diseases.
• Approximately 10% of infected males and 50% of
infected females experience no symptoms, it is often
spread unknowingly.
 Gonococcal Disease in Men
• A man who has unprotected sex with an infected person
can acquire a Neisseria gonorrhoeae infection.
• This organism penetrates the mucous membranes of the
urethra, causing inflammation of the urethra (urethritis).
• Although some men will remain asymptomatic, most will
complain of painful urination along with a purulent urethral
discharge (pus can be expressed from the tip of the penis).
• Both asymptomatic and symptomatic men can pass this
infection to another sexual partner.
• Possible complications of this infection include
epididymitis, prostatitis, and urethral strictures.
• Fortunately, this disease is easily cured by a small dose of
ceftriaxone.
Gonorrhea in men and women. The majority of cases in women are asymptomatic. Local
extension up the fallopian tubes causes salpingitis. The majority of men have acute urethritis,
and only a small percentage have local extension the epididimus. A very small part of either
spectrum results in bacteremia and disseminated gonococcal infection.
 Gonococcal Disease in Women

• Like men, women can also develop a gonococcal urethritis, with painful burning on
urination and purulent discharge from the urethra. However, urethritis in women is
more likely to be asymptomatic with minimal urethral discharge.
• Neisseria gonorrhoeae also infects the columnar epithelium of the cervix, which
becomes reddened and friable, with a purulent exudate.
• A large percentage of women are asymptomatic.
• If symptoms do develop, the woman may complain of lower abdominal discomfort, pain
with sexual intercourse (dyspareunia), and a purulent vaginal discharge.
• Both asymptomatic and symptomatic women can transmit this infection.
• A gonococcal infection of the cervix can progress to pelvic inflammatory disease (PID).
• PID is an infection of the uterus (endometritis), fallopian tubes ( salpingitis), and/or
ovaries ( oophoritis).
• Clinically, patients can present with fever, lower abdominal pain, abnormal menstrual
bleeding, and cervical motion tenderness (pain when the cervix is moved by the
doctor's examining finger). Menstruation allows the bacteria to spread from the cervix
to the upper genital tract.
• It is therefore not surprising that over 50% of cases of PID occur within one week of the
onset of menstruation.
• The presence of an intrauterine device (IUD) increases the risk of a cervical gonococcal
infection progressing to PID.
 Complications of PID include:

1) Sterility: The risk of sterility appears to increase with each gonorrhea

infection. Sterility is most commonly caused by scarring of the fallopian
tubes, which occludes the lumen and prevents sperm from reaching the
ovulated egg.
2) Ectopic pregnancy: The risk of a fetus developing at a site other than
the uterus is significantly increased with previous fallopian tube
inflammation (salpingitis). The fallopian tubes are the most common site
for an ectopic pregnancy. Again, with scarring down of the fallopian
tubes, there is resistance to normal egg transit down the tubes.
3) Abscesses may develop in the fallopian tubes, ovaries, or peritoneum.
4) Peritonitis: Bacteria may spread from ovaries and fallopian tubes to
infect the peritoneal fluid .
• 5) Peri-hepatitis (Fitz-Hugh-Curtis syndrome): This is an infection by
Neisseria gonorrhoeae of the capsule that surrounds the liver. A patient
will complain of right upper quadrant pain and tenderness. This
syndrome may also follow chlamydial pelvic inflammatory disease.
 Gonococcal Disease in Both Men and Women

1) Gonococcal bacteremia: Rarely, Neisseria gonorrhoeae can

invade the bloodstream. Manifestations include fever, joint pains,
and skin lesions (which usually erupt on the extremities).
Pericarditis, endocarditis, and meningitis are rare but serious
complications of a disseminated infection.
2) Septic arthritis: Acute onset of fever occurs along with pain and
swelling of 1 or 2 joints.
Without prompt antibiotic therapy, progressive destruction of the
joint will occur.
Examination of synovial fluid usually reveals increased white blood
cells.
Gram stain and culture of the synovial fluid confirms the diagnosis,
revealing gram-negative diplococci within the white blood cells.
Gonococcal arthritis is the most common kind of septic arthritis in
young, sexually active individuals.
 Gonococcal Disease in Infants
• Neisseria gonorrhoeae can be transmitted from
a pregnant woman to her child during delivery,
resulting in ophthalmia neonatorum.
• This eye infection usually occurs on the first or
second day of life and can damage the cornea,
causing blindness.
• Erythromycin eye drops, which are effective
against both Nisseria gonorrhoeae and
Chlamydia, are given to all newborns.
• Gonococcal conjunctivitis can also occur in
adults.
 Immunity
• The apparent lack of immunity to gonococcal infection
has long been a mystery.
• Among sexually active persons with multiple partners,
repeated infections are the rule rather than the exception.
• Both serum and secretory antibodies are generated
during natural infection, but the levels are generally low,
even after repeated infections.
• Another aspect is that even when antibodies are formed,
antigenic variation defeats their effectiveness and allows
the gonococcus to escape immune surveillance.
• Antigenic variation of pili, Opa proteins, and LOS is
particularly likely to be important.
 Diagnosis

• Gram Smear
• The presence of multiple pairs of bean-shaped, Gram-negative
diplococci within a neutrophil is highly characteristic of
gonorrhea when the smear is from a genital site.
• The direct Gram smear is more than 95% sensitive and specific
in symptomatic men.
• Unfortunately, it is only 50% to 70% sensitive in women, and its
specificity is complicated by the presence of other bacteria in
the female genital flora that may have a similar morphology.
• A positive Gram smear is generally accepted as diagnostic in
men. It should not be used as the sole source for diagnosis in
women or when the findings have social (divorce) or legal
(rape, child abuse) implications.
 Diagnosis
• Culture
• In men, the best specimen is urethral exudate or urethral scrapings (obtained with a loop or
special swab).
• In women, cervical swabs are preferred over urethral or vaginal specimens.
• The highest diagnostic yield in women is with the combination of a cervical and an anal canal
culture; this is because some patients with rectal gonorrhea have negative cervical cultures.
• Throat or rectal cultures in men are needed only when indicated by sexual practices.
• Swabs may be streaked directly onto culture medium or promptly transmitted (in less than 4
hours) to the laboratory in a suitable transport medium.
• The selective medium (eg, Martin–Lewis agar) is an enriched selective chocolate agar with
antibiotics. The exact formulation has changed over the years, but includes agents active
against Gram-positive bacteria (vancomycin), Gram-negative bacteria (colistin, trimethoprim),
and fungi (nystatin, anisomycin) at concentrations that do not inhibit N gonorrhoeae.
• Colonies appear after 1 to 2 days of incubation in carbon dioxide at 35°C.
• They may be identified as Neisseria by demonstration of typical Gram stain morphology and a
positive oxidase test.
• Classically, speciation is by carbohydrate degradation pattern, but this approach has been
replaced by immunologic procedures (immunofluorescence, coagglutination, enzyme
immunoassay) using monoclonal antibodies to unique antigens.
• Neisseria species other than N gonorrhoeae are unusual in genital specimens, but speciation
is the only way to be certain of the diagnosis.
 Diagnosis
• Direct Detection
• Much effort has been directed at developing immunoassay and nucleic acid
hybridization methods that detect gonococci in genital and urine
specimens without culture.
• Such methods have particular importance for screening populations in
which culture is impractical.
• Of these, only the nucleic acid amplification methods have the sensitivity
to substitute for culture and are now widely used in public health
laboratories. The cost/benefit ratio of these tests has been improved by
combining them with Chlamydia detection, which targets the same clinical
population.
• DNA amplification methods are combined with Chlamydia detection
• Serology
• Attempts to develop a serologic test for gonorrhea have not yet achieved
the needed sensitivity and specificity.
• A test that would detect the disease in asymptomatic patients would be
very useful in control of this disease.
 Treatment

• Penicillin was traditionally the drug of choice, but a large

percentage of isolates now are able to produce
penicillinase.
• Others are tetracycline resistant.
• As alternatives, practitioners have been using quinolones
(like ciprofloxacin) or cephalosporins.
• But there is constantly rising resistance to quinolones, as
well.
• β-Lactamase–producing strains are highly resistant
• Ceftriaxone, quinolones, and azithromycin are
recommended therapy
• Quinolone and azithromycin resistance is still uncommon
 Prevention

• Condoms provide a high degree of protection against both

infection with N gonorrhoeae and transmission to a sexual
partner.
• Spermicides and other vaginal foams and douches are not
reliable protection.
• The classic public health methods of case–contact tracing
and treatment are important, but difficult because of the
size of the infected population.
• The availability of a good serologic test would greatly aid
control, as it has for syphilis.
• The development of a vaccine is a high but distant goal.
• Achieving it awaits further understanding of immunity and
its relationship to the shifting target provided by the
gonococcus.