Medical Microbiology

Chapter 1 Introduction
[Requirement]
Master the definition of microorganism
Master the classification of microorganism
[Class hour: 1 hours ]
[Outline]
I. Microbes
1. Definition (microorganism)
2. Classification
3. categories according to structure
1) Non—cellular type
Virus: no cellular structure
Parasite in living cell
Only contain one kind of nucleic acid molecule , DNA/RNA
2)Prokaryotic type
No nuclear membrane or mitotic apparatus nuclear region can be seen ,
composed of DNA.
No separate internal membrane bound organelles .

Bacterium Mycoplusma
Chlamydia Riclcettsia
Spirochetes Actinomycetes
3) Eulcaryotic type
intracellular membrane —enclosed organelles
Fungi : hyphae
yeast
nucleus : two membrane layers
Endoplasmic Reticulum (ER) rough ER
 Smooth ER
Mitochondria
4.Distribution of Microorganisms: 1) In environments.
2) In human organisms
II Microbiology
1 .research objective : Pathogenic Microbes
1)biological properties
2)pathogenesis and immune response
3) Diagnosis and protection
2. History
Leeuwenhock : invent Microscope in 1674 .
Pasteur : pasteurization , Vaccine
Koch : solid medium purify bacteria
Pathogenic microbe criterion
Lister : disinfection aseptic technique
Iwanovsky virus.
3. Modern M
1) M o s t b a c t e r i a w e r e c o n t r o l e d b y a n t i b i o t i
—> resistence plasmid.
Normal flora—opportunistic pathogens
(flora disequilibrium super-infection )
Hospital acquired infections.
New bacteria : Helicobacter pylori—chronic gastritis.
2) Viral research made progress.
AIDS
3) New diagnostic techniques .
ELISA
PCR :polymerase chain reaction.
4) New type vaccine .
5) Microbial genomic program, MGP .
Chapter 2 Bacterial shape and structure
[Requirement]
Master the structure of bacteria and their functions
Master the following concepts :
1. plasmid 2. flagellum 3. pillus 4. capsule
5.L-form bacterium 6. mesosome 7. lipopolysaccharide(LPS)
Understand the size and shape of bacteria.
[Class hour: 3 hours ]
[Outline]
Section I . Bacterial Morphology
Single cellular prokaryotic microbe

I . Size of bacteria
Measure unit um(micrometer)
Coccus 1 um bacillus 2—3um
II Shape of bacteria
1. coccus (cocci)
1) diplococcus : in pairs
2) streptococcus: long chain
3) staphylococcus: irregular cluster
2. bacillus (bacilli): rod
3. Spirilla bacterium
1)vihrio v. cholera
2)Spirillum
4. Campylobacter
C. jejuni
Helicobacter H. pylori
 Section II Bacterial structure
Superficial layer cell wall
(cell envelope) cell membrane
mesosome
inner -nucletid
plasmid
specific structure -external 1 flagellum 2 pillus
supper-facial :capsule
inner: endodpor .
I Basic stucture
（I）cell wall
1. f u n c t i o n : 1 ) p r o t e c t i o n 2 ) k e e p the constant shape . 3)antigeni
material
2. structure and chemical composition peptidoglycan（mucopeptide）
1)polysaccharide backbone . N-acetyl glucosamine link N-acetyl muramic
acid with 1.4 –glucosidic bond
2)t etrape ptide si de chai n link m urami c aci d . ala glu lys ala 3)pent
bridge G+ 【L—glycine】
G－ diaminopimelic acid
3. Special components of Gram-positive cell wall
1. teichoic acids wall teichoic acid
membrane teichoic acid

function :
1) bind Mg2+(magnesium) upply of this ion to the cell
2) provide the cell with its consistency
3) adhesion ------pathogenicity
4) antigenicity
2. polysaccharides
may contain a variety of sugars.
4. Special components of Gram-Negative cell wall .
1) lipopolysaccharide, LPS
a complex and unique glycolipid consisting of there distinct but covalent
linked regions:
（1）lipid A glucosamine disaccharide units connected by pyropho
bridge . endotoxin , non-genus specific
（2）core polysaccharide
（3）specific polysaccharide “o”Ag
2) Outer membrane : exchange .receptor(sex pili phaqe)
3) Lipoprotein
Are firmly but non covalently attached to the peptido-glycan
membrane
4) periplasmic space .
between out membrane and inner membrane binding protein
hydrolytic enzymes

function （1）transport .nonspecific diffusion

（2）barrier: resist-many antibiotic
comparision of cell wall G + G－
（II）Cell membrane
mesosome : invagination vesicular membrane increase membrane area
chondriosome
septal mesosome : cell division
（III）Cytoplasm
plasmid
extra-chromosomal genetic material circle , double streands DNA, replicated
independently , carry genetic information, control a wide rang
bacteria.
（Fertility）factor F ---------control sex pili
factor R----------control drug resistance
 col faction ------control E col: to produce bacteriocin.
II. Special structure of bacteria
（I）capsule
secret a slime layer outside the cell wall
composed of polysacchride or polypeptid
pneumococcus anthracis
>0.2um capsule
<0.2um microcapsule
slime layer Washed off

does not appear to be associated with the cell.

Function condition , enviroment host body
Function
1. anti—phagocytosis surface phagocytosis .opsonic phagocytosis
2. rnti-dry
3. antigenicity
4. typing
（II）flagella
long filamentous appendage.
Originate in the protoplasmic membrance
Function :1. motility 2. antigenicity “H”Ag 3. pathogenicity somebacterin
（III）pili （pilus）
filamentous appendage on the surface of bacteria, shorter .straighter ,finer than
flagella.
1. common pili 100—200/cell
adherente o o b rgan a f t s acteria,o m m
dhere ,a o uface
with pathogenicity.
2. sex pili 1—4/cell
male bacteria with sex pili
transfer of genetic material（DNA）during bacteria conjugation
 F+--------àF－
Section 3. Special living form of bacteria.
1. spore
round or elliptic minute body formed inside the bacteria —endospore
dormant form （resting forms ）
vegetative form
1) forming: inadequate nutrition
2) structure （1）core DNA（chromosome）synthesis of proteins. Energy
（2）inner membrane
(3) cortex （4）coat （5）exosporium
3) germination
spore vegetative form
activation initiation outgrowth
4) function of spore
highly resistant to heat . chemical .dry.
reason:（1）many layers .thick coat
（2）little water. 40% free water
80% -----------
（3）large amount of calcium dipicolinate
（4）heat —stable enzyme （DPA,吡啶二羧酸钙）
destroy spore: autoclave.121°C 15—30′
（II）Bacteria L—Forms cell wall deficient form
1) Morphology: friend egg —spherical body.
Colonies large body
Filamentous colony —filamentous forms.
2) Media . high osmotic .reversion
3) Pathogenicity
Similar to the infection of virus or m
（yocr og pa lnai ss m
m as without
wall）
Infiltration of mononuclear cells and lymphocytes and different from that o
 the bacterial infections , with mainly infiltration of neutrophils.
Chapter3. Bacterial multiplication and metabolism
[Requirement]
Master the mode and spead of B. reproduction.
Master the reason that obligate anaerobes can’t grow in free oxygen condition.
Master the metabolic products of bacteria.
Master the following concepts:
1.pyrogen 2. antibiotics 3. bacteriocin
Understand bacterial nutrition.
Understand the growth and reproduction of bacteria
[Class hour: 2 hours ]
[Outline]
I .Bacterial nutrition
1. nutrient material
(1) w a t e r （ 2 ） c a r b o n s o u r c e : e n（
e r3g）
y Nitrogen source constituen
protein（4）inorganic ions (5)growth
2. nutrient type 1)autotrophy 2)heterotroph saprophyte

parasite
II. Bacterial growth and reproduction
1. growth condition (1)enough nutrients (2) Suitable PH (3) suitable temperature
(4)suitable air
obligate aerobe
obligate anaerobe
facultative anaerobe
microaerophilic bacterium
Reason that obligate anaerobes con’t grow in free uxygen condition:(1) Lack of
cytochrome and cytochromase----àEh 300mv/120mv
( 2 ) l a c k o f s u p e r o x i d e d i s m u t a s e ( S
[O2－] H2O.
III. mode and spead of B. reproduction
1)Mode. binary flssion . chromosome replication . synthesize cell membrane an
wall G+B chromosome bind mesosome G－B. chromosome bind membrane.
2)speed generation time 2 0 - - - - 3 0 m i 2n 1 E. 5. - c- -o -l -i 2 10 0HhR 1 M
0 Ty
Tuberculosis
(1) lag phase adapeatation
(2) logarithmic phase most rapid reproduction
(3) stationary phase .rate of reproduction rate of deed
(4) decline phase rate of dead > rate of reproduction.
IV. Bacterial Metabolism
1. Bacterial enzyme
Exoenzyme hydrolase
Some pathogenicity —coagulase
Endoenzyme metabolic enzyme
Respiratory enzyme
Metabolic products
1. Catabolic products and biochemical reaction
2. Anabolic products and clinical significance
1) pyrogen
p o l y s a c c h a －r i cd ee l lo f w a Gl l ( L P S ) a n t
121°C, 30’. Fluid infusion reaction
2) toxin and invasive enzymes
exotoxin
endotoxin
enzyme
3) pigment
water—soluble P. aerogenosa ---green
fat ---soluble S. aureus —golden
4) antibiotics
Killing or inhibiting substances produced
 microorganisms
Actinomycete
5) bacteriocin
protein by certain bacteria , which can kill or inhibit the growth of related
strains.

Chapter 4. Bacteriophage
[Requirement]
Master the interrelation between phage and bacteria
Master the following concepts :

1. lysogenic state 2. lysogenic phage (temperate phage)

3.lysogenec bacteria 4.prophage

Understand the.size, shape and structure of phage.
[Class hour: 1 hours ]
[Outline]
Infecting bacterial virus .
(1) viral common properties
smallest
simple structure DNA/RNA
parasite in living cell
(2) widespread existence
(3) high host –specific parasitism
I. Biological properties
1. Shape and structure
Seen by EM

Tadpol microsphere slim rod collar

head core DNA/RNA tail pipe-like
capsid protein coat base plate
2. antigenicity tail fib
 3. resistive
>Bacteria 75°C 30′
sensitive to rays
II. Interrelation between phage and bacteria
1. ph. Reproduction and lyzing B. virulent ph . which can replicated
cell and released in lysis of B .
replicatic cycle.
1) absorption and tail pins and fibers absorb teichoic receptor of B,
call wall G+
G－
Pr. LPS
Surface structure . sex pili
2) p enetration
:l — sysosome l S like
c ubstance
D
into bacteria.
3) Biosynthesis structure
Phage DNA transcribe mRNA protein
enzyme

phage DNA replication

4) lysis
encode lysosome lysis B. release
2. lysogenic state
infected bacterial phage doesn’t replicate it’s gene ,integrated with bacteri
DNA, its replication is associated with bacterial DNA
lysogenic phage (temperate phage)
lysogenec bacteria
prophage : The DNA of temperate phage integrated into bacterial DNA.

Chapter 5 . Bacterial inheritance and variation.

[Requirement]
Master the characteisticrs of plasmid.
Master the mechanism of mutation and gene transfer of bacteria.
Master the following concepts :
1. heredity(inberitance) 2. variation 3. Plasmid
4. Mutation 5. genetic transfer 6. Reassortment 7. Prion

[Class hour: 2 hours ]

[Outline]
Concept:
(1) h e r e d i t y ( i n b e r i t a n c e ) g e n e r a l s t a b i l i t y o n “ l i k e n e s s ” i n t h e c h a r a c
progeny and parent.
(2) variation . the difference between progeny and parent He
genetic substance changes .nonberitable variation enviroment changes
I. Genetic substance
1. chromosome
consists of a circular double strand DNA molecular, control bacter
life without introne.
2. Plasmid
Extrochromosomal genetic substance.
Circle double strand DNA
（1） autonomous replication
binding to chromosome —episome
（2） tranfer from a bacteria to another
conjugative plasmid : F+ serpili

nonconjugative plasmid: phage

（3）contral most of auxiliary functions of bacterial cell
eg. Antibiotic resistance —R plasmid .
the production of fimbriae—F
bacteriocin —E. coli con.
（4）bacterium can carry one or several plasmid
 （5）dispensable, it can be lost .
（6）different size : small—a few genes .large—hundreds of genes.
3. phage.
III. variation examples.
1. shape and structure variation .
eg . L form
Y . pestic 3—6%. Nacl pleomorphism
Protenus media—H colony

Media(1%carbolic acid) O colony(no flagella)

2. colony variation
smooth colony rough colony
3. virulence variation vaccine
BCG : Bacilla of calmetter-Gueria 230 passages, 13 years.
4. Resistance variation
III. Mechanism of bacterial variation mutation. Transfer and combination .
1.Mutation
（1） concept: a stable heritable change of bacterial gene ,

spontaneous 10－6～10－9

inducing inutagen
（2） type of mutation
molecular foundation of mutation .
Bacterial nucleotide sequence change
i. base replacements
transation (转换) A—G, C—T
transversion（转换）A—T C—G
ii. base deletion . one base or fragment lost
iii. base insertion . one base or fragment
 type （according to code ）
missense mutation: replacement in one code one amino acid
changes

nonsense mutation: replacement in one code change into stop code

frame shift mutation: deletion or insertion one base or fragment

type (according to biological properties)
resistant mutation
nutrient deficient mutation
iethal mutation
conditional lethal mutation
2. mutation and selection
fluctuation test
replica test(Ames test)
3.genetic transfer.
Genetic substance of B.is tranfered to another bacteria.
Donor bacteria : provide genetic substand

Recipient bacteria : accept genetic substand

1) transformation
recipient B. takes up exogenous DNA of donor B.
eg : Griffith’s experiment
competence : bacterial state. In which bacteria can able to take up DNA
environment in logarithmic phase
2) transduction .
donor bacterial DNA is transfered to recipient bacterial by phage .
（1） general transduction: any fragment of donor bacteria is transfered.
（2） Specific transduction: the fragment near attachment is transfered.
（3） Lysogenic conversion
 Bacteria acquired new properties by phage lysogenization.
Eg C, diphtheria
3. conjuqation
bacterial DNA is transferred from donor bacteria to recipient bacteria by F p
Hfr(high frequent recombinant bacterium )
（1）F plasmid F’ plasmid intergrated into chromosome.
F plasmid of high freguent recombinant bacteria separated from
chromosome, carrying a neighbour DNA.
Sexduction.
（2）R plasmid
conjugative R plasmid resistance transfer factor RTF
resistance factor (RF)
the function of RTF is similar to F plasmid

nonconjugative R plasmid.
The result of gene tranfer
i. exogenous DNA was degraded
ii exogenous DNA from circular ds DNA
autonomous replication
abortive infection
iii. exogenous DNA recombine with endogenous DNA
hemologeous recombination
site-spcific recombination
eg phage
transposable elements insertion sequence
transposon

IV. Medical application

1, Bacterial indentification
2. prevention treatment of diseases. Eg . vaccine
 3. screening potential carcinogen . Ames test
4. genetic engineering
（1）prepare objective gene endonuclease E COR I
（2） objective gene recombines with vector
vector: plasmid .phage .
（ 3 ） r e c o m b i n e d D N A t r a n s f e r t o
（4）Screen positive cell.resistance—antibiotics.
(5) Amplify objective gene and express.
Chapter 6 Disinfection and sterilization
[Requirement]
Master the condition of physical methods of disinfection.
Master the mechanism of the antiseptic effect of chemical agents.
Master the following concepts :
1.disinfection 2. sterilization 3. asepsis 4. antisepsis
[Class hour: 1 hours ]
[Outline]:
Concept
1. disinfection : To pathogenic microorganisms (vegetative form)
2. sterilization : To kill all microorganisms . pathogen
vegetative /spore.
3. asepsis : a state of sterility (no living bacteria) asepeic technique
4. antisepsis:: To inhibit growth and reproduction of bacteria.
I. Physical methods
1. heat lethal
make bacterial protein denature . bacterial DNA degrade, inju
membrane .
dry heat: vegatatine 80---100°C 1hr
spore 160°C 2hrs
flame
hot air oven 160°C 2―3hrs
 moist heat
advantages over dry heat
(1) Bacteria absorb H2O , proteinis easy to solidify and denature .
(2) Strong penetration
Eg. 100 th layer: dry heat 130-140°C 4hrs。72.5°C/100 th
Moist heat 105°C 3hrs. 101°C/100 th
(3) lalent heat gas –liquid .
pasteurization 62°C 30′ 72°C ， 30″
Boiling 100°C 5′
Steaming and intermittent sterilization Arnold 3times.
Autoclaving ： 1.05 kg/ cm2， 121.3°C 15－30′－spore
2. radiation
1) ultraviolet light 200---300nm
interfere DNA replication dimmer of Thymine .
strongest spectrum 260nm DNA absort specfurm
weak penetration : definite time and intensity
stimulate skin, eyes,.
Application : air . operation room
2) iDnizing radiation : eg ,x-ray . r-ray.
3. filtration
fiter heat－labile solution
eg. Serum . toxin .antibiotics. etc.
seitz filter: K>EK>EK-S
glassical filter: G1 G2 G3……6
thin membrane filter
4. other: low temperature( preserve bacteria)
II. Chemical disinfection
Disinfectant
Antiseptic
1. chemical agents and mechanism
 1) heavy –metal salts
(1) bind to –SH and destroy it, inhibit activity of enzymes.
(2) Protein denature. Merbromin .thimerosal 0.1%disinfectant
0.01%antiseptic
2) Oxidizer
H2 O 2 KMnO 4 CHCOOH(过氧乙酸)
Halogen（iodine ，chlorinated lime）
（1）SH S—S— enzyme
(2) destroy amino group, indol group.
(3) denature protein
3) surfactant
combine with phospholipid
increast membrane permeability
bromogeramine(新洁尔灭)
domiphen（杜灭芬）
4） aldehyde（醛类）
farmaldehyde、
enzyme deactivity
5） alcohol 70％ alcohal protein denature。
6）phenol （酚类）protein denature ， destroy cell membrane。
2． Application
skin： 2.5％碘酒 70％酒精
mucosa 2％ 红汞 0.1％新洁尔灭
drinking water 漂白粉
patients excreta 5％ phenol 2％lyson
air: formalin spray
3． Affecting factors（1）concentration and time (2) properties and quantity
(3)emperature and PH (4) antagonist: organs protein , excreta
Chapter 7. Normal flora
[Requirement]
Master the initiate infection of opportunistic pathogenic B.
Master the functions of normal flora
Master the following concepts :
1. normal flora 2. opportunistic pathogenic B
[Class hour: 1 hours ]
[Outline]
Concept
1. normal flora: The microorgomison that parasitize on the body surface or trac
connecting with external, don’t harm the host in ordinary condition.
2. opportunistic pathogenic B
some bacteria are unable to cause in ordinary condition , they initiate infection.
If(1) they leave their ordinary habitant and gain access to other part.
(2) host immunity reduce.
(3) Owing to the wide use of antibiotic, hormone and anti-concer druges.
I. Distribution of normal flora
Skin
Mouth carity anaerobes
Intestinal tract: anaerobes: aerobes =1000:1
Vagina: Lactobacillus.
II. Role of normal flora.
1. Biological barrier: antagonism
2. nutrient synthesize : aa. VB. K. enhance absorb.
3. immue: enhance and develop and maturation of immue system
III. Flore disequilibrium (dysbacteriosis)
Balance of normal flora was broken. Flor
(superinfection) widely use antibiotics, sensitive bacteria were killed , t
resistant bacteria reproduce rapidly.
Chapter 8. Bacterial pathogencity
[Requirement]
Master the mechanism of bacterial virulence
[Class hour: 3 hours ]
[Outline]

Section 1. Introduction
I. pathogenicity
1. Definition
Denotes the ability of B. to cause disease.
S. typhi typhoid
M. tuberculosis tuberculosis
N . gonococcus gonorrhea
staphylococci straom
compare: S. aureus, S, epider, S, Saprophy
Major properties of three species of staphylococci
2. Virulence
It refers to extent of pathogencity
Medium lethal dose, LD50
Medium infective dose , ID50
II. Pathogenic B, and non-pathogenic B.
III. Infection and disease
1. Immunity: The host can protect itself from and com
disease.
2. Bacteria: (1)Virulence invasiveness B . component
B. enzymes
Toxins endotoxin
exotoxin
(2) the amount of invading bacteria
(3) the portal of entry
All p eople
a “ ref infected”
b u d w rom n irthf i a ntil a eath,
infections. Normal flora pathogenic B. Translocation .Dysbacteriosis
IV. Mode of modern infection
Pathogenic B. healthy host
 Non-pathogenic B compromised
Normal flora host
(conditional pathogen)
Reason: (1) broad spectrum antibiotic
(2) method of modern .treatment and diagnosis.
Section 2. Bacterial virulence
I . Invasiveness
The ability of B . to resist host defence, colonize, multiply and spread.
1. Surface structure
1).adhesive factor —adhesin
（1）pili
V.cholera
S. desentery specific epithelial cells
N. gonococci
Small intestine rice –water stools
Segmented intestine abdominal cramp (colon)
Urethra and vagina urethritis
Vaccine
(2). lipoteichoic acid, LTA
group A strept . LTA fibrillae adhere to
M-protein LTA receptors
2) antiphagocytic factors
(1) capsule >0.2um
surface phagocytosis: H,C3b
C3b Bb
(2) micro-capsule<0.2um
S.typhi Vi-Ag
E, coli K –Ag
(3) flagella: V . choleva
 H. pylori
2. Exoenzyme
(1) coagulase: S. aureus
fibrinogen fibrin
surround bacteria
(2) hyaluronidase (spreading factor)
hydrolyze hyaluronic acid tissue loose B. spreads
(3) streptokinase. SK. Lyse fibrin
(4) streptodornase, SD, resolve DNA.
II. Toxin
1. Exotoxin
(1) excreted by living cells , mainly G+ B.
(2) Polypeptide
(3) Heat-unstable, 60°C, 1-2hr destroy
(4) Strong antigenicity，exotoxin toxiod
(5) highly toxic
(6) high selection for tissues
exotoxin subunit A： toxicity
subunitB: non-toxicity
bind receptor of sensitive
cell
a. neurotoxin： tetanospasmin spinal cord
b. cytotoxin : diphtherotoxin inhibit: elongation factor 2
cell protein synthesis
c. enterotoxin
cholera toxin
2.Endotoxin
(7) i n t e g r a l p a r t －o fB . GC e l l wall. .Liberated upo
disintegration.
(8) LPS, main toxic part: Lipid A
 (9) Heat-stable: 160°C 2－4hr
(10) Can’t converted into toxiod
(11) Weakly toxic
(12) Non-specific
Function: a, fever
endotoxin stimulate reticuloendothelial cell release
endothelial cell
endogenous pyrogen(IL-1) thermo-regulatory centers
hypothalamus.
Repeated injection of endotoxin gives less and less fever response.
Reason: “tolerance” reticuloendothelia blockade
IgM antibody to LPS.
(2) WBC reaction
LPS induce neutrophi releasing factor WBC except S. typhi
(3) infectious shoclc
LPS blood platelet ,WBC,complement, kinin. Vasoactive ,substances,eg,
Serotonin, kallikrein. Kinins micro- circulation failure
(4) disseminated intravascular coagulation
DIC
Section 3. Occurance and development of infection
I. Origin of infection
1. Exogenous infection
(1) patient
(2) carrier
2. Endogenous infection
II.The mode of infection
1. resp. tract
2. digestive tract
3. damaged skin or mucous membranes
4. arthropod vector
 5. contact: sex contact
III. The type and outcome of infection
1. inapparent infection
2. apparent infection
toxemia
bacteremia
septicemia
pyemia
3. Carrier state