Unit 2: Bacteriology
85
Lecture 3 - Pathogenic Neisseria,
Moraxella and Acinetobacter
The family Neisseriaceae consists of Gram-negative
aerobic bacteria from 14 genera, including Neisseria,
Chromobacterium, Kingella,and Aquaspirillum. The
genus Neisseria contains two important human
pathogens, N. gonorrhoeae and N. meningitidis.
Neisseria gonorrheae
Morphology and culture
Gonococci are Gram-negative, coffee-bean-shaped cocci
that are usually paired and have a diameter of
approximately 1 µm. Attachment pili on the bacterial cell
surface are responsible for their adhesion to mucosal cells.
Gonococci can be grown on moist culture mediums
enriched with protein (blood). The atmosphere for
primary culturing must contain 5–10% CO2.
Neisseria gonorrhoeae
Pathogenesis and clinical picture.
Gonorrhea is a sexually transmitted disease. The
pathogens penetrate into the urogenital mucosa, causing a
local purulent infection. In men, the prostate and
epididymis can also become infected.
Uncomplicated
gonorrhea in the adult male is an inflammatory and
pyogenic infection of the mucous membranes of the
anterior urethra. The most common symptom is a
discharge that may range from a scanty, clear or cloudy
fluid to one that is copious and purulent. Dysuria
(difficulty in urination)is often present. Inflammation of
the urethral tissues results in the characteristic redness,
swelling, heat, and pain in the region. There is intense
burning and pain upon urination. In women, the
gonococci can also cause salpingitis, oophoritis, or even
peritonitis.
Endocervical infection is the most common form of
uncomplicated gonorrhea in women. Such infections are
usually characterized by vaginal discharge and sometimes
by dysuria. About 50% of women with cervical infections
are asymptomatic. Asymptomatic infections occur in
males, as well. Males with asymptomatic urethritis are an
important reservoir for transmission and are at increased
risk for developing complications. Asymptomatic males
and females are a major problem as unrecognized carriers
of the disease, which occurs in the U.S. at an estimated
rate of over 700,000 cases per year. Gonococci reaching
the conjunctival membrane may cause a purulent
conjunctivitis, seen mainly in newborn children.
Gonococci can also infect the rectal or pharyngeal
mucosa. Hematogenously disseminated gonococci may
also cause arthritis or even endocarditis.
Determinants of the Pathogenicity of Gonococci.
Attachment pili on the surface and the outer
membrane protein Opa are responsible for adhesion to
cells of the urogenital tract. Opa also directs the invasion
process by means of endocytosis. Immune defenses
against granulocytes are based on the outer membrane
porin Por that prevents the phagosome from fusing with
lysosomes, resulting in the survival—and
proliferation—of phagocytosed gonococci in
granulocytes. The lipo-oligosaccharide (LOS) in the
outer membrane is responsible for resistance to
complement (serum resistance) as well as for the
inflammatory tissue reaction in a manner analogous to
the more complexly structured LPS of enterobacteria.
Gonococci can capture iron from the siderophilic
proteins lactoferrin and transferrin, accumulating it inside
the bacterial cells to facilitate their rapid proliferation.
An IgA1 protease produced by the gonococci hydrolyzes
secretory antibodies in the mucosal secretions. The
pronounced antigen variability of the attachment pili
and the Opa protein make it possible for gonococci to
thwart specific immune defense mechanisms repeatedly.
Diagnosis
The method of choice is detection of the pathogens by
means of methylene blue and gram staining and
culturing. Gonococci are sensitive in cultures and the
material must be used immediately after they are obtained
to inoculate. Thayer-Martin blood agar with antibiotics
added to eliminate accompanying flora, on which medium
the cultures are then transported to the laboratory. The
identification procedure involves both morphology and
biochemical characteristics. Techniques developed
recently utilize immunofluorescence or coagglutination
methods utilizing monoclonal antibodies to the main
protein of the outer membrane, Por.
Unit 2: Bacteriology
86
Direct detection in pus and secretion samples is possible
using an enzymatic immunosorbence test or detection of
gonococcus-specific DNA sequences coding for rRNA
using a gene probe.
Therapy
The agent of choice used to be penicillin G. In recent
years, however, the percentage of penicillinase-producing
strains has increased considerably all over the world. For
this reason, third-generation cephalosporins are now
used to treat uncomplicated cases of gonorrhea. They are
applied in a single dose (e.g., ceftriaxone, 250–500mg
i.m.). Good results have also been reported with single-
dose oral application of fluorinated 4-quinolones (e.g.,
0.5 g ciprofloxacin or 0.4 g ofloxacin).
Penicillin Resistance in Gonococci : The determinants of
high-level penicillin resistance in gonococci are small,
nonconjugative plasmids, which are mobilized by a
conjugative helper plasmid for transmission from one
gonococcal cell to another. The penicillin resistance
plasmids code for the TEM betalactamase that occurs
frequently in Enterobacteriaceae. It is therefore assumed
that the penicillinase gene in gonococci derived from the
Enterobacteriaceae gene pool. Low-level, inherent
resistance to penicillin is based on chromosomal genes (
penA, penB) that code for penicillin-binding proteins
with reduced affinity to penicillin. These genes are
products of mutations.
Epidemiology and prevention
Gonorrhea is a worldwide sexually transmitted disease
that occurs only in humans. Its level of annual incidence
in developed countries is estimated at 12 cases per 1000
inhabitants. The actual figures are likely to be much
higher due to large numbers of unreported cases. A
reduction in incidence seen in recent years may be due
to AIDS prophylaxis. Protective immunization for
high-risk persons is not feasible due to the antigen
variability of the organism as described above. Stopping
the spread of gonorrhea involves mainly rapid recognition
of infections and treatment accordingly. One hundred
percent prevention of ophthalmia neonatorum is possible
with a single parenteral dose of 125mg ceftriaxone. Local
prophylaxis is also practiced using a 1% solution of
silver nitrate or eye ointments containing 1%
tetracycline or 0.5% erythromycin.
Neisseria meningitidis
Morphology and culture
Meningococci are Gram-negative, coffee-bean shaped
cocci that are frequently pleomorphic and have a
diameter of 1 µm. They are nonmotile and feature a
polysaccharide capsule.
Growing meningococci in cultures requires mediums
containing blood. A concentration of 5–10% CO2
encourages proliferation.
Neisseria meningitidis scanning EM
Antigen structure. Serogroups A, B, C, D, etc. (a total
of 12) are differentiated based on the capsule chemistry.
Epidemics are caused mainly by strains of serogroup A,
sometimes by B strains as well and, more rarely, by group
C strains. Serogroups are divided into serovars based on
differences in the outer membrane protein antigens.
Pathogenesis and clinical picture
Meningococci are parasites of the nasopharynx. These
microorganisms are carried by 5–10% of the population.
If virulent meningococci colonize the nasopharyngeal
mucosa of a host lacking the antibodies, pathogen
invasion of the mucosa by means of “parasite directed
endocytosis” becomes possible. The CNS is doubtless the
preferred compartment for secondary infections,
although hematogenously disseminated pathogens can
also infect the lungs, the endocardium, or major joints.
Onset of the meningitis is usually sudden, after an
incubation period of two to three days, with severe
headache, fever, neck stiffness, and severe malaise.
Severe hemorrhagic sepsis sometimes develops
(Waterhouse-Friedrichsen syndrome).
Diagnosis
Diagnosis requires detection of the pathogen in
cerebrospinal fluid or blood by means of microscopy
and culturing techniques. For success in culturing, the
material must be used to inoculate blood agar without
delay. Identification of the pathogen is based on
identification of metabolic properties. The slide
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87
agglutination test is used to determine the serogroup.
Latex agglutination or coagglutination can be used for
direct antigen detection in cerebrospinal fluid.
Therapy
The antibiotic of choice is penicillin G. Very good results
have also been obtained with third-generation
cephalosporins, e.g., cefotaxime or ceftriaxone. It is
important to start treatment as quickly as possible to
prevent delayed damage. The advantage of
cephalosporins is that they are also effective against other
meningitis pathogens due to their broad spectrum of
action (with the exception of Listeria monocytogenes).
Epidemiology and prevention
Meningococcal infections are more frequent in the winter
and spring months. Transmission of meningococci is by
droplet infection. Humans are the only pathogen
reservoir. Sources of infection include both carriers and
infected persons with manifest disease. In developed
countries, meningitis occurs sporadically or in the form
of minor epidemics in more or less isolated collectives
(work camps, recruiting camps, school camping
facilities). The incidence level is approximately 12 cases
per 100 000 inhabitants per year. In parts of the
developing world (African meningitis belt) the level is
higher. Lethality runs to 85% if the disease is left
untreated, but is reduced to less than 1% if treatment is
begun early enough. Prophylactic antibiosis is indicated
for those in close contact with diseased persons (e.g., in
the same family). Prophylactic measures also include
treatment of carriers to eliminate this reservoir, whereby
minocylin or rifampicin must be used instead of
penicillin G. Prophylactic immunization can be
achieved with a vaccine made from the purified capsule
polysaccharides A, C, Y, and W-135. There is no
serogroup B vaccine, since the capsule in serogroup B
consists of polyneuraminic acid, which the immune
system does not recognize as a foreign substance.
Moraxella and Acinetobacter
The taxonomic definitions of these genera are still
inconclusive. Bergey’s Manual of Systematic
Bacteriology groups both under the family
Moraxellaceae.These bacteria are short, rounded rods,
often coccoid, sometimes also diplococcoid. Their
natural habitat is either human mucosa (Moraxella) or
the natural environment (Acinetobacter).
Moraxella.
The genus comprises two medically important species:
Moraxella catarrhalis
. Component of the normal flora of
the upper respiratory tract. May be responsible for:
pneumonia, acute exacerbation of chronic bronchitis,
otitis media (up to 20% in children), and sinusitis. About
90% of all strains produce one of the so-called BRO
penicillinases, so that therapy with a penicillinase-stable
betalactam antibiotic is indicated.
Moraxella lacunata
. Formerly Diplobacterium Morax-
Axenfeld. Can cause conjunctivitis and keratitis. The
reason why this organism is now rarely found as a
pathogen in these eye infections is unknown.
Acinetobacter
In immunodeficient persons, A. baumannii, A.
calcoaceticus, and other species can cause nosocomial
infections (urinary tract infections, pneumonias,
wound infections, sepsis). Clinical strains of these
species often show multiresistance to antibiotics, so that
treatment of these infections may prove difficult.
Summary:
Neisseria are Gram-negative, aerobic cocci that are often
arranged in pairs.They are typical mucosal parasites that die
rapidly outside the human organism. Culturing on enriched
nutrient mediums is readily feasible.
Neisseria gonorrheae is the pathogen responsible for
gonorrhea (“clap”).
Infection results from sexual intercourse. The organisms
adhere to cells of the urogenital tract by means of attachment
pili and the protein Opa, penetrate into the organism using
parasite-directed endocytosis and cause a pyogenic infection,
mainly of the urogenital epithelium. An infection is
diagnosed mainly by means of microscopy and culturing of
purulent secretions. The therapeutic of choice is penicillin G.
Alternatives for use against penicillinase-positive gonococci
include third-generation cephalosporins and 4-quinolones. N.
meningitidis is a parasite of the nasopharyngeal mucosa.
These meningococci cause meningitis and sepsis. Diagnosis
involves detection of the pathogens in cerebrospinal fluid
and blood. The disease occurs sporadically or in the form of
minor epidemics in children, youths, and young adults. The
antibiotics of choice are penicillin G and third-generation
cephalosporins.
The family Neisseriaceae includes aerobic, Gram-
negative cocci and rods, the most important of which are the
human pathogens N. gonorrheae and N. meningitidis. Other
species in the genus Neisseria are elements of the normal
mucosal flora.