T11: medical bacteriology

5 features of bacteria

• unicellular (single cell)

• prokaryotic organism

• microscopic organism

• multiplies by binary fission → genetically identical

• free living or parasite

1 bacterial colony is generated from

1 cell

colony count unit

CFU/ml

bacterial cell morphology

coccus (sphere)

bacillus (rod)

spirillum

spirochete

bacterial cell arrangements

pairs & singles

chains

clusters

gram (+) bacteria colour in gram stain

purple

gram (-) bacteria colour in gram stain

red

name the 3 internal structures of bacteria

cell wall, cytoplasm, nucleic acid

name the 3 external structures of bacteria

capsule, flagella, pili (fimbriae) & sex pili

special structure of bacteria

endospore

difference between cell wall in gram-negative bacteria & gram-positive bacteria

gram (-) bacteria: thin peptidoglycan cell wall, have outer membrane, lipopolysaccharide

gram (+): thick peptidoglycan cell wall, presence of teichoic & lipoteichoic acid

which bacteria cannot be identified using gram stain

acid-fast bacteria

example of acid-fast bacterium

microbacterium tuberculosis

endotoxin

• what kind of toxin

• present in which type of bacteria

• part of what portion of bacteria

• heat-stable toxin

• gram-negative only

• lipopolysaccharide

internal structures of bacteria

• does bacteria have nucleus

bacteria have no nucleus, hv nucleoid only

chromosome of bacteria

single chromosome with circular dsDNA

plasmid function

carries antibiotic resistance gene

ribosome subunit in bacteria

70S (30S+50S)

organelles in bacteria

bacteria has no organelles

horizontal gene transfer is

Movement of genetic material between organisms (mainly bacteria) that is not parent-to-offspring (not vertical inheritance)

name the 4 horizontal gene transfer mechanisms in bacteria & describe them

1. conjugation - plasmid transferred from donor to recipient

2. transformation - uptake of free DNA from environment

3. transduction - bacterial DNA transferred from 1 bacterium to another via bacteriophage (virus that infects bacteria)

4. vesiduction - gene transfer through extracellular vesicles

external structure that is a major virulence factor of bacteria

capsule

name for capsule or slime layer than surrounds bacterial cell wall

glycocalyx

function of glycocalyx/ bacterial capsule & give example

immune evasion - eg. antiphagocytosis (inhibits phagosome & lysosome fusion, blocks phagocytosis)

cell surface appendages in bacteria

flagella & pilli

flagella function

motility (chemotaxis = movement of organism towards or away from chemical stimulus)

pili function

adhesion of bacteria to mucous membrane (esp. urinary tract) & DNA transfer btwn bacteria (sex pili)

endospore is & function

dormant bodies produced by bacteria during starved conditions

allows bacteria to endure harsh conditions - is highly heat & chemical resistant

endospore present in what types of bacteria

only gram (+)

capsule, flagella, pilli found where

some gram(+) & gram (-) bacteria

5 factors affecting growth of bacteria

1. pH

2. temperature

3. salt condition

4. O2 requirement

5. nutrients: carbon & nitrogen sources

what to do if want to see bacterial colony

grow on agar plate

name the 3 different types of culture media

selective media

differential media

enrichment media

selective media

suppresses unwanted microbes & encourages growth of desired microbes

differential media

distinguish between different bacteria growing on same plate by visible changes

enrichment media

encourages growth of desired microbe (without necessarily inhibiting others)

blood agar composition & use

• contains mammalian blood

• used to isolate fastidious organisms & detect hemolysis

(fastidious organism = microbe that needs special nutrients or conditions to grow)

chocolate agar composition & use

• contains RBCs that have been lysed by slow heating

• grows fastidious bacteria

MacConkey agar function

• selective & differential media

• inhibits growth of gram-positive bacteria & promotes growth of gram-negative bacteria

method of growing anaerobic bacteria

cultivation of anaerobes using anaerobic jar (GasPak system)

anaerobic indicator is

methylene blue

hemoculture use

culturing a patient’s blood sample to detect the presence of microorganisms

for suspected bacteremia

hemoculture - grey label for

aerobic bacteria in adults

hemoculture - gold label for

anaerobic bacteria

hemoculture - pink label for

aerobic bacteria in children

hemoculture - red label for

TB/fungus

what is hemolysis in blood agar

how bacteria breaks down RBCs in blood agar plates

beta hemolysis is what & state observation around bacterial colony

RBCs fully lysed → clear zone around colonies

alpha hemolysis is what & state observation around bacterial colony

RBCs partially broken down → greenish/brown discolouration around colonies

gamma hemolysis is what & state observation around bacterial colony

no RBC destruction, RBCs not lysed at all → no change in blood agar

bacterial metabolism is

all biochemical reactions that occur in cell/organism

state 2 biochemical - single enzyme tests

catalase & coagulase test

catalase test purpose

distinguishes btwn Staphylococcus & streptococcus

coagulase test purpose

distinguishes btwn Staphylococcus aureus (coagulase positive) & other coagulase-negative staphylococci

state the 2 bacterial classification methods

bergey’s manual

modern classification

how does bergey’s manual classify bacteria

morphology, biochemical tests, immunological tests

how does modern classification classify bacteria

according to genetic sequence: 16s rRNA

state the 4 phenotypic classifications of bacteria

• morphology - colony morpho, cell morpho, cell structure

• physiology - growth characteristics, nutrition, metabolism

• antimicrobial pattern - antibiotic susceptible or resistance

• immunogenicity pattern - antigenic characteristic or serotyping

what test used to measure bacterial morphology

staining, culture, microscopic

what test used to measure bacterial physiology

culture media, culture condition, biochemical test - bacterial metabolism

what test used to measure antimicrobial pattern

antimicrobial susceptibility test

antimicrobial susceptibility test is

determine which antibiotics can effectively treat an infection & to detect resistance patterns

antimicrobial susceptibility test methods are

disc diffusion methods (zone of inhibition, clear areas around bacteria colony), MIC & MBC (minimum inhibitory concentration & minimum bactericidal concentration)

what test used to measure immunogenicity pattern

(describes how the immune system responds to a substance, often a drug or vaccine)

serology test & immunoassay

genotypic classification measures what & specify

genetics - DNA or RNA, gene mutation

state the tests used in genotypic classification

PCR, RT-PCR, DNA sequencing, DNA hybridization

state the 2 types of staining techniques

simple & differential staining

simple staining is & use

use of a single stain for visualisation of morphological shape & arrangement

differential staining is & use

use of 2 contrasting stains separated by a decolourizing agent for identification & visualization of structure

name the differential staining used for identification

gram stain, acid-fast stain

name the differential staining used for visualization of structure

spore stain & capsule stain

gram stain distinguishes bacteria (gram-positive & gram-negative) based on

difference in cell wall compartment

cell wall characteristics of gram-positive bacteria

• thick peptidoglycan layer

• lipoteichoic acid at cell wall

cell wall characteristics of gram-negative bacteria

• thin peptidoglycan cell wall

• presence of lipopolysaccharides at surface

• presence of outer membrane layer

name the 4 dyes used in gram staining

1. crystal violet (purple) (basic dye)

2. gram’s iodine

3. alcohol

4. safranin (red) (counter stain)

state the use of each dye in gram staining

1. crystal violet causes both cell walls to affix the dye

2. gram’s iodine causes the dye to be trapped in the cell wall of the gram (+) bacteria, has no effect on gram (-)

3. alcohol = decolorisation, gram (+) remains purple but gram (-) becomes colourless

4. safranin (counterstain): stains the gram (-) bacteria, turns the colourless cell to pink-red

state the colour of gram (+) & gram (-) bacteria at the end of gram staining test

gram (+) = violet

gram (-) = red

after gram-stain, what procedure next

view slides under microscope, objective lens 100x

state the gram-reaction, cell morpho & cell arrangement for Staph aureus

gram-positive cocci in cluster

state the gram-reaction, cell morpho & cell arrangement for Streptococcus pyogenes

gram-positive cocci in chain

state the gram-reaction, cell morpho & cell arrangement for S.pneumoniae

gram-positive diplococci (lancent shaped)

state the gram-reaction, cell morpho & cell arrangement for neisseria gonorrhoeae & neisseria meningitidis

gram-negative diplococci

state the gram-reaction, cell morpho & cell arrangement for bacillus cereus

gram-positive bacilli in chain w/ central spore

distinctive characteristic of acid-fast bacteria

mycolic acid/ high lipid content in cell wall

name the 2 methods used to stain acid-fast bacilli

1. ziehl-neelsen (heat method)

2. kinyoun’s method (cold method)

name for staining method used to detect acid-fast bacilli through fluorescence microscope

auramine-rhodamine staining

AFB staining used to detect acid-fast bacilli through which type of microscope

light microscope

positive test for AFB staining thru light microscope

red long rods → presence of acid-fast bacilli

positive test for auramine-rhodamine staining

red-orange/yellow/reddish-yellow fluorescence → presence of acid-fast bacilli

AFB grading

• 1-2 AFBs seen per whole smear

doubtful positive → scanty (inadequate)

AFB grading

• 1-9 AFBs seen per 100 fields

(tung 100 fields, hen AFB yuu 1-9 un)

AFB 1+, positive

AFB grading

• 1-9 AFBs seen per 10 fields

AFB 2+

AFB grading

• 1-9 AFBs seen per single field

AFB 3+, positive

AFB grading

• >9 AFBs seen per single field

AFB 4+, positive

modified acid-fast stain use

detects partially acid-fast organisms that have low amount of mycolic acids in their cell wall

characteristic of bacteria detected by modified acid-fast stain

branching, filamentous bacteria

give examples for partially acid-fast bacteria

nocardia, actinomyces

what is non-gram staining bacteria

bacteria that is not or poorly detected by gram stain

examples of non-gram staining bactera

1. obligated intracellular bacteria (bacteria that cannot survive or multiply outside host cell) → chlamydiae, rickettsiae

2. mycoplasma spp.

3. legionella spp.

4. treponema pallidum

treponema pallidum cause what

syphillis