Microbes

Microbes are studied under

Bacteriology: study of bacteria
Mycology: study of fungi
Virology: study of virus
Serology: study of serum; in vitro antigen antibody reactions
GENERAL CHARACTERISTICS OF BACTERIA
Structure & classifcation of bacteria
Bacteria are microorganisms which have a primitive nucleus & are called
prokaryotes. t do a like actinomycetales. not contain chlorophyll. !hey are
unicellular & donot show true branching e"cept in higher bacter !hey di#er
from eukaryotes in the following ways.
$rokaryotes %bacteria& 'ukaryotes%(ungi)
$roto*oa&
+. ,ell wall $eptidoglycan ,hitin
-. ,ytoplasmic
membrane
Sterols absent $resent
.. /uclear
membrane
0bsent $resent
Classifcation
,lassi1cation involves the arrangement of organisms into a ta"onomic group
on the basis of similarities. !he ma2or characteristics of microorganisms fall
into the following categories:
+. Morphological characteristics:
,ell shape) si*e & structure; cell arrangement; occurrence of special
structures & developmental forms; staining reactions & motility &
3agellar arrangement.
-. ,hemical composition:
!he various chemical constituents of the cells.
.. ,ultural characteristics
/utritional re4uirements & physical conditions re4uired for growth) &
the manner in which growth occurs.
5. Metabolic characteristics
!he way in which microorganism obtains) utili*es & stores energy)
carries out metabolic reactions & regulates them.
6. 0ntigenic characteristics
Special large chemical compounds %antigens& of the cell distinctive for
certain kind of microorganisms.
7. 8enetic characteristics
,haracteristics of hereditary material of cell) 9/0 & occurences &
function of other kinds of 9/0 that may be present. '". $lasmid.
:. $athogenicity
!he ability of the organism to cause disease in various plants or
animals or other organisms.
;. 'cological characteristics
<abitat & the distribution of the organism in nature & the interactions
between & among species in natural environment.
=. ,lassi1cation based on relationship between host & microorganism:
Microorganisms are classi1ed on the basis of relationship between
host & microorganism as Saprophytes) parasites.
Bacteria are classi1ed basing on their shape.
a. Cocci %>okkos:berry& spherical or oval bacteria.
b. Bacilli rod shaped bacteria.
c. !ibrios ,omma shaped) curved rods
d. S"irilla Spiral bacteria) which are rigid & cannot bend.
e. S"iroc#etes Spiral bacteria which are 3e"ible.
f. Actino$%cetes Bacteria which are long & 1lamentous) & show
branching similar to fungi.
g. M%co"las$a are bacteria without a cell wall. Since cell wall gives
shape to a bacterium) mycoplasma lack de1nite shape) & so can
assume any shape.
Arran&e$ent
Some bacteria show a typical arrangement or grouping. !hus) there are:
a' Sta"#%lococci ,occi arranged in irregular groups resembling a bunch
of grapes.
b' Stre"tococci(stre"to bacilli ,occi or bacilli arranged in chains.
c' )i"lococci ,occi arranged in pairs.
*' Cuneifor$ arran&e$ent 0rranged at angles to each other) as in
,orynebacterium diphtheria.
)e$onstration of bacteria $icrosco"% & stainin&
Bacteria are microscopic & invisible to the naked eye.
Most of the pathogenic bacteria have a si*e range of ?.6@ to +6 @. !he
resolution %si*e of smallest ob2ect that can be seen& of unaided eye is about
-?? @. <ence the study of bacteria re4uires a microscope. !he resolution of a
compound microscope is usually ?.-6 @ %-6?nm&.
'lectron microscope gives a resolution of +nm.
,ompound
microscope
'lectron microscope
+ Smallest ob2ect visuali*ed ?.- microns +nm
- Ma"imum magni1cation +???" +)??)???"
. Source of light Sunlight or electronic
light
Beam of electrons
5 Medium in which rays travel 0ir Vacuum
!he commonest) widely used microscope in a microbiology laboratory is a
compound microscope. t consists of the following parts:
+. Stand: !he stand consists of a A shaped foot. Modern microscopes
have a 3at base.
-. Body: !he body is a , shaped structure attached to the stand. t holds
various functional parts of the microscope like optical tube) stage)
condenser & mirror. !his has two types of ad2ustments) the coarse &
the 1ne ad2ustment.
.. !he optical tube: !he optical system is mounted in a tube called optical
tube. 0t the upper end) the optical tube carries the eye piece %lens
nearest to the eye&. 0t its lower end) it has a revolving disc shaped
nose piece to which are attached ob2ective lenses %lenses nearest to
ob2ect&. !he ob2ective lenses are of various powers: low power B6")
+?"C high power %5?"& & oil immersion %+??"&.
5. Stage: !his is a hori*ontal platform situated below the nose piece.
!he stage accommodates the glass slide on which the microorganism
is mounted. t is called mechanical stage) as the slide can be moved in
various directions smoothly & accurately. t has a hole in the centre to
allow light rays to pass through the ob2ect on the slide.
6. Substage condenser: !his is a system of lenses which can condense &
focus the rays of light on to the ob2ect on the slide.
7. ris diaphragm: !his is present 2ust below the condenser. !his controls
the amount of light.
:. Mirror: !his is situated below the iris diaphragm. !his serves to focus
the light into the optical system) from the source. !he mirror is plane
on one side & concave on the other modern microscopes have a bulb
in the place of the mirror) which can act as source of light if connected
to electricity.
E+a$ination of fres#, unstaine*, li-in& "re"arations of bacteria
!hese are also called wet preparations. 0s the bacteria are live) they are
potentially infectious & care should be taken during handling.
!hese are two types of wet preparations.
a. <anging drop method
!his is mainly used to test for the motility of bacteria. !rue motility can
be identi1ed by change in place & direction of bacteria; where as false
motility %Brownian movement& is identi1ed as only a change in
direction but not place.
b. Det 1lm method
Det 1lms are used for bacteria like treponema palladium in chancre
3uid) vibrio cholera in stool sample) ova) cysts & larvae of intestinal
parasites) tropo*oites of trichomonas vaginalis in vaginal 3uid &
micro1laria in blood.
E+a$ination of staine* "re"arations
!he clinical specimens or cultures of bacteria can be smeared on a slide)
dried) 1"ed) stained & e"amined under oil immersion. Bacterial protoplasm is
translucent & cannot be visuali*ed if not stained. !he p< of bacterial
protoplasm is acidic) & so basic stains are used usually in a ?.6E
concentration. !he various staining methods are:
a. Simple staining: 9ilute a4ueous solutions of basic dyes such as
methylene blue) methyl violet) safranine or basic fuchsin are used.
Ases:
F Si*e) shape & arrangement of bacteria can be observed.
F Spores of bacteria appear as circular rings attached to bacteria.
b. /egative staining: n this procedure) the organism remains unstained
but the background gets the stain. !he techni4ue can be wet or dry.
ndia ink or +?E /igrosin is used.
Ases:
F !o study the capsules of bacteria like pneumococci) klebsiella)
bacillus anthracis or capsule of fungi like ,ryptococcus neoformans.
c. 8ramGs Staining: !his is the most widely used stain in bacteriology. t is
named after <ans ,hristian 8ram %+;;5& who describes the techni4ue.
!his is di#erential stain.
$rinciple: !o the 1"ed smear of bacteria on the slide) a primary stain is
added. !he stain is then removed %decolourisation& with a decolourising
agent %usually acetone or alcohol&. !hereafter) another stain of a
contrasting colour %counterstain& is added) to visuali*e the
decolourised cells.
Ases:
!he bacterial population can be divided by 8ramGs staining into two
groups.
8ram positive: !hose bacteria that retain the primary stain & resist
decolourisation %eg: Staphylococci) Streptococci&
8ram negative: !hose bacteria that get decolourised & take the
counter stain. %eg: '.coli) Salmonella&
8ramGs reactivity related to the composition of cell wall & p< of
protoplasm.
d. 0cid fast staining: !his is another di#erential staining techni4ue)
mainly for bacteria which are diHcult to be stained by gramGs staining
%Mycobacteria) /ocardia) Spores of bacteria&.
$rinciple:
!o the 1"ed smear on the slide) primary stain %IiehlF/eelsenGs carbol
fuchsin& is added. 8entle intermittent heat is applied for better
penetration of the stain into the cell. !he smear is then decolourised
with a dilute acid. ,ounterstain is then applied.
Ases:
!he organisms which take up the primary stain & resist acid
decolourisation are called Jacid fastG bacteria. Mycobacterium
tuberculosis is strongly acid fast %-?E sulphuric acid& & mycobacterium
leprae less %6E sulphuric acid&.
Anlike the gram positive character which is relative & is lost by
e"cessive decolourisation) acid fastness is an absolute character. So)
addition of acid & duration of its contact with the smear must be
liberal. 0cid fastness is ascribed to the presence of Jmycolic acidG in the
bacterial cell wall.
e. mpregnation stains: !hese are used for thin) slender bacteria like
spirochetes. Silver stains are used) which get deposited on the
organism & increase its width. !hus bacteria are made visible under
the microscope. %e.g: (ontanaGs Stain&.
f. 0lbertGs stain: !his is a special stain used to demonstrate
intracytoplasmic granules in corynebacterium diphtheria. !he bacteria
are stained pale green) & the granules bluish purple.
BACTERIAL CELL SI.E & FORM, /ARTS OF BACTERIAL CELL
!he various components of a bacterial cell are:
+. ,ell wall: gives shape to bacterium
-. ,ytoplasmic membrane: for active transport of materials into & outside
the cell.
.. ,ytoplasm: contains ribosomes) mesosomes & granules
5. /ucleus: determines the characters of the bacterium.
6. (lagella: useful for locomotion
7. (imbriae %pili&: help in adhesion to surfaces
:. Spore: helps to survive adverse conditions
;. Mesosomes: these are points from which the cell starts dividing by
binary 1ssion
=. Kibosomes: used for protein synthesis
+?. nclusion granules: act as store houses of energy.
+. ,ell wall
!his is a rigid outer structure which provides shape to the bacterium &
protect it from the e"ternal osmotic pressure which is 6F-? times lower
than the internal osmatic pressure. 0 bacterium which which loses its
cell wall undergoes bursting. !he main constituent cell wall is
peptidoglycan which is also called murein or muramic acid.
!he cell wall of gram positive bacteria di#ers from that of gram negative
bacteria
S'N
o
Gra$ /ositi-e Cell 0all Gra$ Ne&ati-e Cell 0all
+. !hicker !hinner
-. !eichoic acid present Lipopolysaccharide %L$S& present
.. $eptidoglycan +7F;? nm thick -nm thick
1ses of Cell 0all
%i& 8ives shape to bacterium
%ii& Maintains integrity & protects the bacterium from bursting
%iii& Kesponsible for gramGs reactivity
%iv& n gram negative bacteria) L$S is responsible for endoto"ic activity
%v& 0ntigenic & induces antibody formation
-. ,ytoplasmic membrane
t is an elastic semipermeable membrane) beneath the cell wall.
%i& !his encloses the internal structures of bacteria like cytoplasm)
ribosomes & the nuclear ring
%ii& t gives origin to certain bacterial appendages like 3agella &
1mbriae
%iii& !he septal & lateral mesosomes also take origin from the
cytoplasmic membrane.
.. /ucleus
Bacteria are prokaryotic %$roMprimitive) >aryonMnucleus& & hence
nuclear membrane is absent. !he 9/0 is suspended in the cytoplasm
in the form of a ring) with attachment toa septal mesosome. !he
bacterial 9/0 contains +??? genes) which determine the characters of
bacteria. Bacterial cell division starts with the nuclear division.
5. $lasmid
0part from the nuclear 9/0) bacteria also possess e"trachromosomal
9/0 in some occasions. !he e"trachromosomal 9/0 element e"isting
freely in the cytoplasm) unattached to the nucleus is called JplasmidG.
$resence of plasmid confers certain additional characters to the
organism like drug resistance or to"in production. $lasma replicates
independently of the bacterial replication which means that) when the
bacterium replicates only one daughter cell gets the plasmid while the
other does not. $lasmids propagate themselves between bacteria
either by transduction %through which plasmid is transferred&.
mportance of plasmids
%i& ,onfer on bacteria certain additional characters like drug
resistance or to"igenicity.
%ii& Bacteria containing JKG plasmid %resistance to antibiotics& are well
known to cause hospital infections
%iii& $lasmids are widely used in transport of important genetic
characters between bacteria %genetic engineering&.
6. ,apsule
Some bacteria possess a viscid layer outer to the cell wall. !his is
called capsule & provides a slimy outer envelope to the bacterium.
7.
8rowth & nutrition of bacteria
,ulture media
Laboratory methods of identi1cation of bacteria
0ntibiotic sensitivity testing: di#usion & dilution tests in brief
Bacterial coloni*ation on various parts of the body
,ollection transport of clinical samples to laboratory