Module 8 Lecture
Neisseria Classification
domain: bacteria
phylum: proteobacteria (gram negative with different shape cells)
class: B-proteobacteria (classification dependent on rRNA gene)
order: Neisseriales
family: Neisseriaceae
genus: Neisseria
species (14 total): Neisseria gonorrhoaea & Neisseria meningitidis
genome — 1 chromosome; 2.1 megabases; 2,000 genes total; GC content 52%
GC content is not important for placement in phylum
Neisseria general characteristics
gram negative with diplococcus arrangement
look like coffee bean with scanning electron microscope
very small; ~0.6-1 um
nonmotile
non-spore forming
can be found in mucus membranes such as respiratory tract & urogenital tract
many normal flora; those aside from the two discussed in leture (e.g. can be found in throat)
catalase positive (convert H2O2 to H2O and O2)
oxidase positive
has cytochrome C oxidase & electron transport chain that is required for transfer of electrons to the final acceptor O2
aerobic — like 21% oxygen
capnophile — like 3-5% CO2
facultative intracellular pathogens
can get inside host cells & grow; considered fastidious (picky)
need lots of growth factors which limits media they can grow on
sensitive to temperature
Neisseria gonorrhoeae general characteristics
very fastidious
primary pathogen in humans
grown on lysed RBC which provide additional growth factors that would otherwise not be available
any antibiotics present in the agar media do not impact the growth of Neisseria but instead help the media select which colonies grow
no growth on blood agar plate; only grows on chocolate agar
referred to as gonococcus
colony morphology
small, tan, translucent
morphology is dependent on pili presence/absence
(+) pili — typically smaller, raised and tend to reflect light
(-) pili — larger, dull, flat
Neisseria gonorrhoeae virulence factors
pili/fimbriae function to help with adhesion to host cells
target specific receptor on epithelial cells
pili show antigenic variability — can change how they look overall
can look different to host to become undetectable; host cannot recognize if infected twice
evade immune system
lipo-oligosaccharide (LOS)
gram negative have LPS cellular component
N. gonorrhoeae has a modified lipopolysaccharide called lipooligosaccharide
causes endotoxic shock (stimulate nonspecific immune response in host)
immune response ineffective against invading microbe
modification — absent O side chain; replaced with a branched component
LPS is released as blebs during rapid growth
atypical of other gram negatives
IgA protease
sIgA antibody prevalent in mucous to kill bacteria that is under the mucus
Neiserria prefers to live in the mucus & the IgA protease chews up sIgA; lives and survive in mucus
some outer membrane proteins that promote adhesion
example: Opa
antigenic variable
no capsule present!!
survival in blood is loss; no dissemination
transferrin receptor
transferrin is the molecule humans use to bind iron
bind up free iron to prevent bacterial infection bc they require it for growth
Neisseria have a receptor that bind our iron and steal supply
humans as only host target because Neisseria require an iron supply and they possess the same transferrin receptor
Neisseria gonorrhoeae disease
ophthalmia neonatorum
an eye infection that is acquired during vaginal birth; often to asymptomatic mother
newborn babies are required to receive antibiotic eyedrops
gonorrhea - sexually transmitted disease
there are cases where the bacteria can infect the throat & one can acquire anal rectal infection
can by asymptomatic / nonspecific symptoms
incidence of N. gono - 350,000 cases
heavily underreported because females are asymptomatic and because it is a STD
mortality is very low; rare dissemination into bloodstream because it does not have a capsule
if capsule present then it manifests as arthritis in joints
in females the bacteria are found in the cervix and urethra
symptoms include cervicitis and urethritis
can experience painful urination & pain during sexual intercourse
purulent discharge may happen
~50% are symptomatic
problematic because pathogen can travel to fallopian tubes & cause damage and PID (pelvic inflammation disease)
can lead to infertility and ectopic pregnancy
in males the bacteria are found in the urethral epithelium
symptoms include urethritis & purulent discharge
~95% are symptomatic
infrequently (rare) the pathogen will ascend into the testes and cause inflammation in the epididymis
N. gonorrhoeae lab identification
gram negative
fastidious pathogen; must be notified that handling this pathogen because it has certain mediums it will grow on and it will be susceptible to drying and temperature changes
cannot store in the fridge because they would die & must be transported in special media
when swabs are taken, use special swabs because the pathogen is sensitive to certain types
calcium alginate will kill the bacteria
specimen differs in the genders
males — urethral swab (do not need to consider normal flora)
perform direct gram stain; gram negative diplococcus
females — endocervix swab
will not perform direct smear because normal flora present
non-urogenital specimen (in throat or rectum)
swab the area & notify of the pathogen
gonococcal selective media to culture (because the pathogen is often referred to gonococcus)
must be quick due to temperature fluctuations and to prevent desiccation
will not grow on just blood agar
modified thayer martin medium — chocolate agar + lots of antibiotics to reject growth of fungi and other Neisseria species
high have CO2 content (~3-5%)
culture will look small, raised and will reflect light
oxidase positive
catalase positive
Cysteine trypticase agar + carbohydrates under to distinguish gram negative diplococci, oxidase positive colonies recovered from gonococcal media
tubes with different carbon sources - glucose, maltose, lactose, sucrose
if the bacteria can use that source, it turns red → yellow
N. gonorrhoeae is glucose positive (can only use glucose as carbon source)
more ID methods include chromogenic substrate, coagglutination, modified conventional/chromogenic enzyme, and fluorescence associate monoclonal antibodies
non culture test methods include nucleic acid amplification test (like PCR) and nucleic acid hybridization
many tests for identification suggest sexual abuse in children (not ophthalmic neonatal)
N. gonorrhoeae treatment & prevention
high antibiotic resistance
developed resistance in several ways
plasmid-mediated penicillin resistance
chromosome-mediated resistance
plasmid-mediated high-level tetracycline resistance
chromosome-mediated spectromyocin resistance
chromosome-mediated fluoroquinolone resistance
treatment with antibiotics
cephalosporins (B-lactamase resistant B lactams)
often treated for chlamydia as well
infection does not confer immunity
antigenic variability with the pili and LPS so host cell cannot recognize invasion
if treated and infection goes away, the person can get infected once more
no vaccine but there is work on one
N. meningitidis general characteristics
meningococcus
colony morphology
a bit bigger than N. gonorrhoeae colonies
small, smooth, glistening
tan appearance on chocolate agar
blue tint on BAP
encapsulated
distinguishes from N. gonorrhoeae
~13 serotypes
regulated process; dependent on gene expression
fastidious pathogen but not as much as N. gonorrhoeae because it can grow on BAP
can be found naso/oropharynx carriers (asymptomatic)
3-30% carriers; pathogen is usually unencapsulated
N. meningitidis virulence factors
pili — used for adhesion
exhibit antigenic variability & there are multiple genes that encode these pili
bacteria can change which is presented on surface to evade host immune system
transferrin receptor
binds host transferrin and siphon host iron source away
LOS
can cause endotoxic shock (by lipid A component)
modified in that it is missing outer O site chain and the core is branched
blebs secreted when bacteria are growing during exponential phase
IgA protease chews up sIgA in mucus membranes of human hosts
capsule helps block phagocytosis & promotes survival in the blood
can cause more systemic infections
N. meningitidis disease
acquired by close contact & respiratory secretion from asymptomatic carriers in the nasal or oropharynx
dorms / barracks
infects the pharynx of the infected and gains access to the blood
carrier → infected symptomatic host → pharynx → cross epithelial barrier → access to bloodstream → travels to meninges → coma → death
meningitis
inflammation of the meningeal layers of the brain and spinal cord
symptoms experienced include frontal headache, very stiff neck, vomiting, nausea, and fast progression to coma
fatality rate is 10-15%
onset of symptoms on the order of 12 hours
touch chin to chest & if pain associated then likely have meningitis
meningococcemia
bacteremia (bacteria in blood)
gram negative & LOS with lipid A component that induces endotoxic shock
drop in blood pressure and intravascular coagulation (hemorrhaging into the skin)
symptoms purpuric rash from the hemorrhaging - no blanching if spots are pressed
fatality rate 25-50%
critical that patients are treated immediately with antibiotics
N. meningitidis lab identification
specimen usually from sterile sites like blood & cerebral spinal fluid
perform direct gram stain
CSF smear yields tiny gram negative diplococcus
in culture, it can grow on chocolate agar & BAP
to find source of infection perform nasopharynx or oropharynx swab which would need to be grown on selective media (gonococcal selective agar)
colony morphology should appear small, raised, tan on chocolate agar & blue on SBA
oxidase positive
catalase positive
cysteine trypticase agar + carbohydrates test with four carbon sources
glucose positive & maltose positive
to distinguish serotypes of the capsule, use slide agglutination test
N. meningitidis treatment and prevention
low antibiotic resistance
for meningitis antibiotic of choice in intravenous penicillin
for meningococemia treat with intravenous cephalosporin
vaccine is available
Menactra
protects against a few serological groups A, C & Y
Moraxella
classification
domain: bacteria
phylum: proteobacteria
class: gamma-proteobacteria
order: Pseudomonadales
family: Moraxellaceae
genus: Moraxella
species: Moraxella catarrhalis
genome — 1.86 megabases 42% GC content, about 2,000 genes total
Moraxella catarrhalis general characteristics
direct smear on otitis media (inner ear infection)
gram negative diplococcus
growth on chocolate agar plate & SBA
colony morphology
“wagon-wheel” — tan ring surrounding a yellow center, irregular edges
hockey pucks — take inoculating loop & press on colony, tightly aggregated and scoot across media surface
opaque and tan/yellow coloration
reservior: commensal of upper respiratory tract (in children more than adults)
can lead to opportunistic infections
Moraxella catarrhalis virulence factor and disease
LOS that can cause endotoxic shock
formation of bioflims
pathogen does not disseminate; localized infection
3rd most common cause of middle ear infections
Moraxella catarrhalis lab identification
Moraxella catarrhalis is a secondary infection; a cold or viral infection first hits and then there is secondary infection with ear pain (specimen is ear exudate)
culture - “wagon-wheel,” hockey puck
catalase positive
oxidase positive
cysteine trypticase agar + carbohydrates test with four carbon sources
asaccharolytic - does not give positive result in any tubes
Moraxella catarrhalis treatment
most produce B-lactamase so B-lactams are ineffective (no penicillin!!)
choice of B-lactamase resistant B-lactams antibiotics
upper generation cephalosporins
for prevention - no vaccine