Mycoplasmas

Hugh B Fackrell

2/14/2018 1
 Presentation Outline
 Structure
 Classification
 Multiplication
 Clinical manifestations
 Epidemiology
 Diagnosis
 Control

2/14/2018 2
 Pleuropneumonia organism
 The mycoplasmas are essentially
bacteria lacking a rigid cell wall during
their entire life cycle, although they are
also much smaller than bacteria. The
first organism of this type was
associated with pleuropneumonia of
cattle, and was originally called the
pleuropneumonia organism (PPO).

2/14/2018 3
 General Characteristics
 smallest known free-living organisms.
 Because of the absence of cell walls, they do
not stain with the Gram stain, and they are
more pleomorphic and plastic than eubacteria.
 Giemsa stain
– they appear as tiny pleomorphic cocci, short rods,
short spirals, and sometimes as hollow ring forms.
Their diameter ranges from 0.15 u to 0.30 u.

2/14/2018 4
 Mycoplasma, Ureaplasma
 very small (0.2 x 0.8 um)
– pass through a 0.45 um filter
 No Cell wall: plasma membrane only
– resistant to antibiotics that interfere
with the integrity of cell wall;
penicillins, cephalosporins, vancomycin,
bacitracin
 susceptible to tetracycline, erythromycin

2/14/2018 5
 Structure
 The cell is enclosed by a limiting membrane
which is more similar to that of animal cells
than that of bacterial cells because of sterols
present in the membrane.
 The cytoplasm contains ribosomes,but lacks
mesosomes. There is no nuclear membrane.
 In some strains, amorphous material on the
outer surface of the membrane suggests the
existence of a capsule.

2/14/2018 6
2/14/2018 7
 Mycoplasma
 requires sterols for growth, can be
grown on laboratory media
 most are facultatively anaerobic
– Exception M. pneumoniae
 replication controversial
– replication time 1-6 hours

2/14/2018 8
 Mycoplasma pneumoniae
 AKA Eaton’s agent
– aerobic but very slow growing
 extracellular pathogen: attaches to
respiratory epithelium by an attachment
factor called P1
 interacts with a glycoprotein receptor on the
epithelial cell surface
 ciliostasis is followed by epithelial cell
destruction

2/14/2018 9
 Clinical Syndrome
 Pneumonia
– walking pneumonia frequently
confused with virus infection
– primary atypical
– clinical
 Tracheobronchitis
 Pharyngitis
– differential diagnosis from
Strep throat
2/14/2018 10
 Children most susceptible

2/14/2018 11
 No Seasonal Incidence

2/14/2018 12
 Urethritis
 1/2 of urethral infections not caused by
Chlamydia or N. gonorrhoeae.
 Caused by
– Mycoplasma hominus
– Ureaplasma

2/14/2018 13
 Infection of Tracheal ring
Organ culture

2/14/2018 14
 Destruction of host
 due to release of hydrogen peroxide
and superoxide anion.

2/14/2018 15
 Laboratory diagnosis
 Culture:
– fried egg colonies on medium containing
sterols
– Most mycoplasmas require a rich medium
containing a sterol and serum proteins for
growth.
 Serology:
– Complement Fixation test,
Hemagglutination
2/14/2018 16
 Laboratory Diagnosis
 Culture Mycoplasma from sputum,
mucous membrane swabbings or other
specimens
 direct inoculation into liquid or solid
media containing serum, yeast extract
and penicillin to inhibit contaminating
bacteria.

2/14/2018 17
 Cultural Characteristics
 Despite the lack of a cell wall, they do not
require a medium of very high osmotic
pressure.
 On solid media, they form minute,
transparent colonies.
– looks like a fried egg. The different strains vary in
their growth rate
 may take from two days to several weeks to
form a colony.
2/14/2018 18
 Fried Egg Colonies

2/14/2018 19
 Fried Egg Colonies
 Stain intensely with neutral
red or tetrazolium or
methylene blue.

2/14/2018 20
 serology: complement fixation
 on acute and convalescent serum.
 patient’s serum heated to 56C to eliminate
complement
 combine patient’s serum and known
Mycoplasma antigen in presence of added
complement. Mix.
 Incubate - add indicator system
– Red cells and anti-red cell antibody
– hemolysis occurs if complement is unused.

2/14/2018 21
 Hemagglutination
 Cold agglutinins to human O
erythrocytes.
 hemabsorption & B-hemolysis of
guinea pig red blood cells.

2/14/2018 22
 Identification
 conclusively identified by staining its
colonies with fluorescein-labelled
antibody.

2/14/2018 23
 M. pneumoniae Nucleic Acid
Probes
 specific recombinants to
oligonucleotide sequences that are only
found in Mycoplasma pneumoniae.

2/14/2018 24
2/14/2018 25
 L Forms
 Some bacteria readily give rise
spontaneously to variants that can replicate
in the form of small filterable protoplasmic
elements with defective or absent cell walls.
 These organisms, called L-forms, can also be
formed by many species when cell wall
synthesis is impaired by antibiotic treatment
or high salt concentration.

2/14/2018 26
 L Forms vs Mycoplasma
 contain a rigid cell wall, at least at one
stage of their life cycle
 no sterols in their cytoplasmic
membrane.

2/14/2018 27
 Pleuropneumonia-like organisms
 Several organisms with similar
morphological characteristics and cultural
properties have been isolated. These are
commonly referred to as pleuropneumonia-
like organisms or PPLO. A certain group of
mycoplasmas produce extremely tiny
colonies on agar plates, and are called the T-
strains.

2/14/2018 28
 Metabolism
 The parasitic mycoplasmas have truncated
respiratory systems, lacking quinones and
cytochromes.
 Another indication for the simplicity of the
electron transport chain is the finding that
the reduced nicotinamide adenine
dinucleotide (NADH) oxidase activity is
cytoplasmic.

2/14/2018 29
 Arginine dihydrolase Pathway
 pathway Complex electron transport chains
are usually membrane bound, since they
depend on the spatial organization of their
components. Ruling out oxidative
phosphorylation as an ATP-generating
system leaves only two proven ways of ATP
generation, both based on substrate level
phosphorylation. The major source for ATP
is the arginine dihydrolase pathway.

2/14/2018 30
 Metabolism
 A few species derive their energy from the
degradation of glucose or the hydrolysis of
urea.
 All species synthesize DNA, RNA, lipids and
proteins.
 Not known if they can synthesize amino
acids.
 Those species that require sterols incorporate
these sterols (mainly cholesterol) into the cell
membrane up to concentrations of 65%.
2/14/2018 31
 Multiplication
 In the absence of a rigid cell wall, the
pattern of replication is quite different from
that of typical bacteria, whose division
starts with the formation of a well-defined
septum.

2/14/2018 32
 Life Cycle of PPLO
Elementary
body

2/14/2018 33
 Fragmentation of filaments
 mechanism of division in mycoplasmas
is controversial, sequential microscopic
observation suggests that new
elementary particles arise by
fragmentation of filamentous cells
containing several discrete DNA
components.

2/14/2018 34
 DONE!!!

2/14/2018 35