Neisseria genus

Gram negative cocci

TAXONOMY

Neisseria species and

Branhamella (Moraxella)
catarrhalis are classified with
the genera Moraxella, Kingella
and Acinetobacter in the family
Neisseriaceae.
General description:

coccae, diameter 0.6- 1 m;

bean shaped and paired, with
their flat sides touching;
Gram negative, but can show
resistance to decolouration;
none develop flagella or spores,
but capsules can be found on
the pathogens;
non- motile, strictly aerobic.
Neisseria
Classification:
- according to growth requirements are:
- fastidious: N. meningitidis , N.
gonorrhoeae (menongococci, and
gonococci respectively)(complex growth
requirements: grow best on choco-late agar
supplemented with solubilized starch, under
increased CO2 tension, at 37C);
- non fastidious: present in normal
flora of colonised mucosa
Ex.: N.lactamica, N.mucosa, N. sicca, N.
subflava, N.cinerea etc;
Biochemical identification:

catalase and oxidase positive;

show weak metabolic activity;
sugar fermentation tests;
kit contains: glucose, fructose,
sucrose;
Neisseria

16 species, 10 in humans
Neisseria
CSF Gram smear
Classification

Fastidious Neisseria:
- N. meningitidis;
- N. gonorrhoeae.
Non-fastidious Neisseria:
- N. sicca;
- N. lactamica;
- N. flava;
- N. subflava;
- N. perflava, etc.
N. meningitidis

Microscopical characteristics: intra- and

extracellular disposition in smears from
CSF.
Growth characteristics:

less fastidious than gonococci:

- blood agar, chocolate agar, in wet atmos-

phere, CO2, 3- 5% stimulate growth;
- 18- 24 hours: S type colonies, diameter:1- 4
mm; non hemolytic, non pigmented.
N. meningitidis- classification

Serogroups: meningococci can be

grouped into 13 serogroups according to
antigenic differences in capsular polysa-
ccharides (A, B, C 90%, J, and W-135
the most frequent; others: X, Y, Z, 29e, L
etc.).
- Serogroups A, B, C and Y are wide-
spread and are most frequently involved
as human pathogens.
Serotypes: serogroups B and C are
subdivided into serotypes according to
outer membrane proteins (Europe).
Immunity

these antigenic structures stimulate

synthesis of protective antibodies,
except serogroup B (similar with
E. coli K1).
Capsular B antigen is not recog-
nized as non self structure and
dont stimulate antibody produc-
tion.
 Determinants of pathogenicity

Adhesion factors: pili mediate adhe-

sion to the oropharyngeal mucosa and
to the meningeal tissue.
Capsule: antiphagocytic properties.
Particular LPS LOS
(lipooligosaccharides)
IgA proteases: protect bacteria against
the effect of the secretory IgA.
Pathogenicity factors:

Meningococcus can colonize the

mucosa of the superior respiratory
tract (oropharynx and nasopharynx);
pilli are involved in adhesion on cell
receptors;
IgA are involved and IgA proteases
that destroy IgA molecule, present
on the surface of mucosa.
Invasiveness
Pathogenicity factors:

Antiphagocytic factors:
polysaccharidic capsule;
proteic structure of external membrane (their
involvement is sustained especially for group
B meningococci).
Can cause severe infections- toxic shock,
because of endotoxin production in high
amount (this endotoxin is different compared to
Enterobacteriaceae).
Distal chains have low number of saccha-
rides: lipooligosaccharides- LOS)
Pathogenesis

N. meningitidis attaches to the cells of

the nasopharynx following the inhalation
of contaminated droplets.
In a non-immune, susceptible host, it
invades locally, disseminates
hematogenously, and reaches the
meninges, its primary target tissue.
Once the bacteria reach the meninges,
they proliferate, causing inflammation.
 Immunity

Complement-fixing IgM and IgG

antibodies can promote bacterial elimina-
tion from circulation and tissues.
Complement-deficient individuals
(particularly those deficient in C5, C6, C8
or other components of the terminal
sequence) are approximately 8000
times more susceptible to infections
caused by N. meningitidis than normal
individuals.
Clinical disease
Clinical disease

A febrile illness, usually self-limiting,

is the mildest form of meningo-
coccal infection.
A blood culture may reveal Neisseria.
Meningitis: severe headache, fever,
stiff neck, vomiting, neurologic signs
(e.g., cranial nerve palsies corresponding to
inflammation of the basal meninges), myalgias,
arthralgias, altered mental status and
coma.
 Clinical disease

Acute meningococcemia is systemic

infection with N. meningitidis leading to
septic shock, widespread petechial or
hemorrhagic skin lesions, disseminated
intravascular coagulation (DIC), and
death within 8-12 hours of the initial
appearance of symptoms.
Meningococcemia
Clinical disease

The Waterhouse-Friderichsen
syndrome (fulminant meningo-
coccemia of short evolution) is
characterized by myocarditis
and bilateral hemorrhagic
destruction of the adrenals.
Waterhouse-Friderichsen syndrome
(fulminant meningococcemia)
Waterhouse-Friderichsen syndrome
(fulminant meningococcemia)
Epidemiology

N. meningitidis is transmitted by
respiratory secretions and is highly
contagious.
Carriers harbor Neisseria in the
nasopharynx but may be asympto-
matic.
The carrier rate varies from 3-20%
in nonepidemic areas to as high as
50-95% during epidemics.
High transmission rates are found in
day-care centers and military
barracks.
Diagnosis

Examination of CSF is useful for

the establishment of the
diagnosis of meningitis.
A gram-staining will not precisely
identify the organism, but it will
provide information about mor-
phology and staining characte-
ristics.
CSF
CSF
Blue methylen staining
Neisseria Gram smear
14.12
Diagnosis

Culture and antibiotic susceptibility

tests should be performed in order to
determine the treatment protocol for
the patient.
Rapid diagnostic tests (latex aggluti-
nation) fail to yield information
concerning antibiotic susceptibility.
Blood cultures.
Cultivation (chocolate agar)
Cultivation

S type colonies, diameter: 1- 4

mm;
non hemolytic, non pigmented.
Oxidase (+)
Identification
Species identification is based on
sugar degradation.
Is done by inoculation on special
medium - CTA (cystin tripsin agar+ sugar
phenol red or rapid tests - 44 tests).
Maningococci: glucose + maltose +;
Supplementary identification:
capsular polysaccharidic antigen
with specific antiserum (latex
agglutination) A, B, C, J, W 135
Biochemical identification
Glucose Sucrose Lactose Maltose
G (+) M(+) G (-) M(-) G (-) M (-) G (-) M (+)
N meningitidis versus
N. gonorrhoeae
N meningitidis
Mini API
Diagnosis

Other cultures
Cultures from nasopharyngeal
secretions are indicated;
Treatment

Penicillin G is effective when large doses

are administered intravenously.
In the last years were isolated strains with
modified Penicillin sensitivity (Spain,
England, USA, South Africa).
MIC: 0.1- 1 g/ ml- low level resistance,
which, in most cases dont affect the
treatment for MIC < 0.5 g/ml.
If MIC > CSF concentration, is used
- Cefotaxime,
- Ceftriaxone or
- Chloramphenicol.
Treatment

Changes in Penicillin sensitivity are

caused by PBP 2, and gene that
codify this PBP are resembling with
those of pneumococcal resistance
(mosaic genes) acquired by
transformation.
The mosaic genes are from
commensal Neisseria: N. flavescens,
N. lactamica, N. cinerea.
Treatment

This resistance is a cause of major

concern- for surveillance of antibiotic
sensitivity for meningococcal strains.
Testing: similar to pneumococci
(Oxacilin 1g/ml).
For resistant strains, MIC by
quantitative method: E test.
Other resistance mechanisms: beta-
lactamase production with increased
MIC (few strains in the literature).
Treatment

Ceftriaxone and other third

generation cephalosporins
distribute to the CSF and, because
of their broad spectrum, are effective
against other bacteria often isolated
from patients with meningitis.
Treatment

Chloramphenicol, which has

excellent meningeal penetration, is
an adequate choice for patients
allergic to penicillin.
It has been suggested that the administration of
corticosteroids to reduce the inflammatory response
may improve patient outcome.
Control and prevention

During epidemics, both chemopro-

phylaxis (for short-term protection) and
immunization (for long-term protection)
are indicated.
Chemoprophylaxis.
Prevention with rifampin or ciprofloxacin
is recommended for individuals with close
patient contact (e.g., family members)
because of their increased risk of infection.
Day-care and medical personnel may also
benefit from chemoprophylaxis in special
cases.
Epidemiology

Transmission: respiratory route;

Pflugge droplets. Low resistance in
environment.
Transmission after long contact with
source.
There is a risk for meningitis in family
contacts or those coming from a
children or teenager community
(recruits).
Appear in first 10 days after contact.
Epidemiology

Risk for infection occurrence is 500-

800 times increased compared to
infection in general population. More
frequent in cold and wet season.
In Europe, USA - majority of menin-
gococcal infections - group B, then
C- endemic or sporadic infections.
Meningitis A- epidemic.
Control and prevention

Immunoprophylaxis.
A vaccine constituted by the capsular
polysaccharide of serotypes A and C is
available.
A quadrivalent vaccine, containing the
polysaccharides of serotypes A, C, Y,
and W-135, has recently been
introduced.
Conjugate vaccines made with A and
C polysaccharides and a non-toxic
mutant of diphtheria toxin are currently
being evaluated.
Neisseria gonorrhoeae

Morphology: in pathological
products as coffee grains, inside
PMN cytoplasm, that allow
etiological diagnosis only after
direct microscopic examination.
Gram staining
Urethral pus
Samples prelevation

If the patient develops skin

lesions, those can also be
cultured;
Joint fluid, tracheal aspirate, or
urethral exsudate may be
examined by Gram stain and
cultured, depending on the
patients symptoms.
9.12
Growth characteristics

More fastidious than meningococci;

Grow well on special culture media
(HYL Horse blood, Yeast extract,
Liver extract), Thayer Martin,
NYC (New York City)- transparent culture
media that detects also Mycoplasma hominis and
Ureaplasma urealyticum.
CO2 atmosphere (carboxiphil bacteria).
Cultivation from contaminated
samples
Antibiotics for commensals
inhibition
Culture (+) in 24-48 hours
Cultivation

Colonies: small, transparent,

corresponding to virulent strains
(Type 1 and 2 colonies); Type 3
and 4 non-virulent strains.
Colonies
Cultivation
Identification

sugar fermentation- glucose

degradation by oxidative
pathway;
auxotyping- dependence of
strain on some amino acids;
there are > 30 auxotypes;
AHU auxotype: these strains
need next for their growth:
arginine, hypoxanthine, uracil.
Specificity antigens

Fimbrial antigens that are different,

and their existence make vaccine
preparation difficult.
Explain chronic evolution of gonoco-
ccal infection.
Pilli have 2 regions:
- antigenically constant, involved in
receptor attachment;
- hypervariable region- antigens are
immunogenic.
Specificity antigens

Antibodies appear when the

concentration is critical- determine
elimination of that strain, with
selection of new antigenic variants,
for which the non specific defense
mechanisms are not prepared.
Protein 1 (major) gonococcus
serotyping with epidemic importance.
These serotypes are more frequently
encountered in systemic infections.
Pathogenicity factors

pilli proteins;
proteins from external membrane are
involved in adhesion;
produce IgA protease spleet IgA, and
colonization is favored;
antiphagocytic capsule effect- proteins from
external membrane that inhibit bacterial
phagocytosis or ensure bacterial survival
inside phagocyte; dont have capsular
polysaccharide.
Endotoxin - LOS, systemic toxic potential,
compared to meningococci and also have
cytotoxic effect.
Pathogenic factors
 A major porin protein, P. I (Por), in the outer
membrane of the bacterium is thought to be the
invasin that mediates penetration of a host cells
Neisseria gonorrhoeae can produce one or several
outer membrane proteins called Opa (P.II)
proteins. Increase attachment that become ferme.
These proteins are subject to phase variation.
 Rmp - reduction-modifiable protein (P.III) is an
outer membrane protein found in all strains of N.
gonorrhoeae. It does not undergo antigenic variation
and is found in a complex with Por and LOS.
It shares partial homology with the OmpA protein of Escherichia coli.
Antibodies to Rmp, induced either by a neisserial
infection or by colonization with E. coli, tend to
block bactericidal antibodies directed against Por
and LOS.
In fact, anti-Rmp antibodies may increase
susceptibility to infection by N. gonorrhoeae.
Infections
Bacteria is adjusted to sexual transmission; with
different locations in male and female.
Male:

- urethral mucosa
- evolution as acute urethritis with urethral purulent
secretion, in 90% cases.

Diagnosis: laboratory diagnosis, treatment.

In incorrectly treated infections or in those with

asymptomatic infections - infection become chronic
with high risk for complications - ascendant
infections (epididimitis, prostatitis, elderly patients-
urethral strictures);
homosexual- rectal mucosa infection: rare, clinically
manifest as acute proctitis.
Infections
Female infection evolves in endocol- endocervicitis.
In the vicinity: urethritis, proctitis (contamination of
rectal mucosa during defecation, with genital
secretions or sexual intercourse).

50% cases- asymptomatic infection with two

consequences:

- female: infection reservoir;

- high risk for chronic infection- pelvic inflammatory
disease;
- acute evolution acute salpingitis, pelviperitonitis, or
late complications leading to pelvic inflammatory
disease (PID) sterility or ectyopic pregnancy.
Clinical disease

Pharyngitis, pneumonia, and presen-

tations usually associated with N.
gonorrhoeae (e.g., urethritis, chronic
meningococcemia low-grade fever,
joint pain, and skin lesions).
Infections

Girls: gonococcus infection is a

sign of promiscuity.
Localization: vulvo- vaginal
mucosa.
Complications
 Complications

Both in male and female (more frequent

in female), infection can evolve as
complication (1-3%):
systemic infection localized in
joints: septic arthritis; skin manifes -
tations: macular, papular, necrosis
lesions.
Rare: endocard or meningeal localization.
Other receptive mucosa: pharyngeal-
minimal or asymptomatic expression;
gonococcus conjunctivitis, purulent in
new born.
Invasive infections

serotype AHU, highly Penicillin

sensitive
hosts with reduced levels of C5,
C6, C8.
Diagnosis - microscopy
Diagnosis

Acute infections direct

microscopically examination.
Chronic infections

small number of bacteria; cant

be differentiated from commen-
sal Neisseria that colonize
vaginal mucosa in women.
In this condition- culture.
Microscopically examination
Bacteriological examination

Pathological product:
prelevation with dacron swab.
Cotton is toxic.
Amies transport media, in less
than 6 hours should arrive in the
lab.
Inoculation on culture media.
Oxidase (+)
Oxidase (+)
Catalase (+)
Biochemical identification tests
(glucose +, maltose -, sucrose -)
Mini API galeries (identification)
Colistin sensitivity (resistant)
Acridine orange staining (UV
light examination)
Treatment

Initially was with Penicillin.

1976- increase of penicillin MIC-
dosage increase (from 100.000
UI to 4.8 mil IU). Unique dose is
preferred for treatment.
1976: penicillinase producing
strain.
Treatment

MIC values couldnt meet the

treatment.
New antibiotics, initially
Spectimonycin- efficient in genital
locations, but not strains from
pharynx.
There are Spectinomycin resistant
strains.
Tetracycline was also used, but also
tetracycline resistant strains were
isolated.
Treatment

Today- Ceftriaxone in unique

dose- infection is stopped; other
genital infections are resolved.
Chromosomal resistance by
PBP and changes in permea-
bility ( MIC 1-2 g/ml and those beta- lactamase
positive: > 32 g/ml).
Treatment

Second place: Chlamydia

trachomatis - ceftriaxone in unique
dose + unique dose of Azithromycin/
7 day.
Also the sexual partners should be
treated.
Detection, by serologic diagnosis, of
other possible sexually transmitted
infections (syphilis, HIV/ AIDS).
Epidemiology:

encountered only in humans

always pathogenic.
Transmission: sexually; exception:
toilet objects, recently contaminated.
Vertical transmission - mother with
genital infection; risks for infection
transmission.
Females are more exposed to
gonococcus infection.
Number of contacts increase the risk
for transmission.
Epidemiology:

Prophylaxis
Non specific measures: detection
of sources, educational strate -
gies, avoiding the sexual contact
with occasional partners.
In new born, instillation of Ag
NO3 1% solution.
Non- fastidious Neisseria

N. lactamica, N. mucosa, N.
sicca, N. flavescens, N. subflava
with 3 biotypes: subflava, flava,
flavescens,
N. elongata, N. cinerea.
*N elongata : catalase negative, elongate,
bacillary shape.
Others, have the same
characteristics.
Growth characteristics:

on ordinary media, at 35- 37o C.

Majority can grow on enriched
media, at 22 oC.
According to colony morphology
and sugar fermentation is
differentiated the species.
Infections:

Rare: lower RTI in immuno-

supressed patients;
bacteriemia with localization in
endocard and meninges.
Infections:

Meningitis: N. lactamica, N. mucosa,

N. flava
Bacteriemic severe evolution: N.
lactamica, N. subflava
Endocarditis: N. mucosa, N. sicca,
N. subflava, N. cinerea, N. elongata
Others infections: ostheomielytis,
arthritis, cellulites, sinusitis, otitis,
lung severe infections, conjunctivitis.
Treatment

Natural antibiotic sensitivity; the

same sensitivity spectrum as Gram
positive cocci except Penicillin M
and lyncosamides, to which are
naturally resistant.
There is the possibility of genetic
transfer increase in frequency of
Penicillin resistant strains for N.
flavescens, N. cinerea, N. lactamica,
the source for mosaic genes.
N. lactamica
N. lactamica
N. cinerea
Moraxella genus (former Brahamella)
subgenus Branhamella
cocci shape
B. catarrhalis
subgenus Moraxella rod
shape species
M. lacunata
M. nonliquefaciens
M. osloensis
Branhamella catarrhalis

the only medically important

species; appears only in
humans.
microscopical characteristics:
similar to Neisseria.
in pathological product- there is
a tendency for discoloration,
more difficult in Gram staining,
intra and extrecellular.
B. catarrhalis

Growth characteristics:- non

fastidious bacteria; grow on ordinary
media.
Better development on blood agar/
chocolate agar.
Appear in 24 hours, aerobic
conditions.
Non pigmented, non hemolytic
colonies, white- grey appearance.
After 24 hours- center is acuminated;
slippery on the media.
B. catarrhalis

Colonization of RT and there is a

difference with age: more frequent
children are colonized with this
species.
Involved in LRTI, URTI, in cases with
obstructive bronchopneumopathy.
Can be isolated alone or in
association with: H. influenzae, S.
pneumoniae.
Infections

Can cause pneumonia,

empiema.
In children- the third etiologic
agent of acute otitis media.
Infections

Acute infections: otitis, sinusitis,

bronchopulmonar infections,
keratitis, supurative arthitis.
Severe infections: endocarditis,
meningitis.
Old persons: respiratory diseases,
chronic bronchitis, acute lobar
pneumonia
Unknown pathogenic mechanism.
Diagnosis:

microscopically examination
Cultivation: non- fastidious
Identification: catalase, oxidase
positive and non fermenter
Antibiogram.
B. catarrhalis
B. catarrhalis
Non-fastidious bacteria
B. catarrhalis
B. catarrhalis
The lack of sugars
fermentations
DNA-ase (+)
Colistine sensitivity test (rare
strains are resistant)
B. catarrhalis: treatment

Sensitive to: second, third

generation cephalosporins,
macrolides, Tetracycline,
Chloramphenicol, Rifampicine,
Fluoroquinolones.
Cothrimoxazole (SXT): variable
sensitivity.
B. catarrhalis: treatment

>90% strains are resistant to

penicillin (beta-lactamases
production).
Treatment: amoxicilline +
clavulanic acid or orale
cephalosporins, resistant to
beta-lactamases.
 subgenus Moraxella
Microscopy: rods very short, in pairs or short
chains
non- fastidious, except M. osloensis
Infections:
M. lacunata (Moraxs bacillus):
conjunctivitis, keratitis
M. nonliquefaciens: bronchopneumonia.
M. osloensis: meningitis, septicemia,
ostheomielytis, septic arthritis (is
commensal of genital tract; others, commensals on
respiratory tract).
Treatment

Moraxella subgenus: sensitive

to penicilline, cephalosporines,
amynoglicosides, tetracycline,
rifampicine, polymixines.